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39057403_p44
|
39057403
|
sec[3]/p[1]
|
4. Conclusions
| 4.21875 |
biomedical
|
Study
|
[
0.99951171875,
0.00028514862060546875,
0.00017130374908447266
] |
[
0.9990234375,
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Spiculiferosides A, B, and C exhibited moderate cytotoxic activities against human embryonic kidney HEK293, melanoma SK-MEL-28, breast cancer MDA-MB-231, and colorectal carcinoma HCT 116 cell lines but significantly inhibited proliferation and colony formation in HCT 116 cells. Spiculiferoside C demonstrated the highest anti-cancer activity among the investigated compounds. The molecular mechanism of anti-cancer action of this compound was associated with the induction of cell cycle arrest at the G2/M phase through regulation of CDK2, CDK4, cyclin D1, and p21WAF1/CIP1 proteins’ expression and inhibition of mitogen-activated protein kinases of MAPK/ERK1/2 signaling cascade.
|
[
"Alla A. Kicha",
"Dmitriy K. Tolkanov",
"Timofey V. Malyarenko",
"Olesya S. Malyarenko",
"Alexandra S. Kuzmich",
"Anatoly I. Kalinovsky",
"Roman S. Popov",
"Valentin A. Stonik",
"Natalia V. Ivanchina",
"Pavel S. Dmitrenok"
] |
https://doi.org/10.3390/md22070294
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278278_p0
|
PMC11278278
|
sec[0]/p[0]
|
1. Introduction
| 4.152344 |
biomedical
|
Study
|
[
0.9990234375,
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[
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High-risk uterine sarcomas (HRUS) are generally aggressive tumors with an unfavorable outcome . The preoperative diagnosis of uterine sarcoma is difficult but important, especially to determine the choice of surgical approach. Patients with uterine leiomyosarcoma (LMS), high-grade endometrial stromal sarcoma (HGESS), and undifferentiated uterine sarcoma (UUS) have similar symptoms, including abnormal uterine bleeding (AUB), rapid uterine growth, and lower abdominal discomfort or pain . Despite efforts, there is still no reliable diagnostic method to differentiate between uterine leiomyoma and sarcoma . Numerous previous studies have described the challenges associated with sonographic diagnosis of uterine sarcoma . In one of the largest multicenter studies regarding sonographic findings in uterine sarcoma involving 195 patients, uterine sarcomas were observed sonographically as solid masses with heterogeneous echogenicity, occasionally featuring irregular cystic areas, and were predominantly moderately or very well vascularized . However, Köhler et al. recently published a preoperative risk score that can significantly facilitate the diagnosis of uterine sarcoma based on medical history, clinical, and sonographic findings, and made recommendations for further diagnostic procedures.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p1
|
PMC11278278
|
sec[0]/p[1]
|
1. Introduction
| 3.943359 |
biomedical
|
Review
|
[
0.99609375,
0.002826690673828125,
0.0008754730224609375
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[
0.04840087890625,
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Uterine LMS arise from the uterine muscle and are usually surrounded by the myometrium . Therefore, performing endometrial biopsy in patients with LMS is considered less sensitive than in patients with other uterine malignancies such as endometrial carcinoma, carcinosarcoma, and adenosarcoma. However, HGESS and USS regularly develop in the endometrium and are therefore more amenable to endometrial diagnostic procedures. Curettage, endometrial sampling, and endometrial biopsy remain the most used diagnostic procedures in patients with abnormal uterine bleeding. These are different endometrial diagnostic procedures but are often used synonymously in the literature. Additional hysteroscopy improves sensitivity and should be performed when possible .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p2
|
PMC11278278
|
sec[0]/p[2]
|
1. Introduction
| 3.810547 |
biomedical
|
Study
|
[
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[
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Even after endometrial biopsy, many uterine sarcomas are inadequately treated, mostly when histologically misdiagnosed as leiomyomas or other benign lesions. The evidence for the predictive value of endometrial biopsy in patients with endometrial cancer is well defined, while that for uterine sarcoma is still poorly defined . However, the sensitivity of endometrial biopsy for uterine sarcoma, regardless of its indication, is unknown. In a meta-analysis of the results of endometrial biopsy for postmenopausal bleeding, one of the main symptoms of HRUS, uterine sarcoma is not mentioned at all .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p3
|
PMC11278278
|
sec[0]/p[3]
|
1. Introduction
| 4.089844 |
biomedical
|
Study
|
[
0.9990234375,
0.0006780624389648438,
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[
0.77978515625,
0.0013866424560546875,
0.2186279296875,
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As the efficacy of radiotherapy and chemotherapy is generally poor in uterine sarcoma, surgical therapy is considered the gold standard. The choice of surgical approach, whether minimally invasive or open with or without tissue morcellation, is a crucial point in the treatment, with controversial results regarding its influence on outcomes . Raspagliesi et al. have investigated the impact of morcellation on survival outcomes in 125 patients with uterine sarcoma and reported that patients undergoing morcellation experienced a 3-fold increased risk of death . Reichert et al. recently published a retrospective analysis of 301 LMS and showed that morcellation doubles the risk of locoregional recurrence (HR = 2.11, 95%-CI 1.41–3.16, p < 0.001) and surprisingly prolongs the time to distant metastases (HR = 0.52, 95%-CI 0.32–0.84, p = 0.008), while no influence on overall survival was observed . Therefore, adequate surgery in terms of total hysterectomy without tumor injury remains the treatment of choice, while inadequate surgery must be strongly avoided .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278278_p4
|
PMC11278278
|
sec[0]/p[4]
|
1. Introduction
| 4.011719 |
biomedical
|
Study
|
[
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[
0.998046875,
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0.0002416372299194336
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Due to the very high number of endometrial biopsies performed for uterine bleeding and the extremely low incidence of HRUSs of approximately 0.4 per 100,000 women, its sensitivity cannot be determined. Therefore, the primary outcome of this study is to determine the detection rate of endometrial biopsy in patients with HRUSs and its most common indications. Secondary outcomes include evaluating the impact of preoperative biopsy on surgical approach and the rate of inadequate surgery, as well as survival data.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p5
|
PMC11278278
|
sec[1]/p[0]
|
2. Material and Methods
| 4.105469 |
biomedical
|
Study
|
[
0.990234375,
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[
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Materials: For this study, data on 467 patients with histologically proven high-risk uterine sarcomas (HRUS) by hysterectomy were retrospectively retrieved from the consultation database of the German Clinical Center of Excellence for Genital Sarcomas and Mixed Tumors (DKSM, University Medicine Greifswald, Germany). Inclusion criteria were patients with early or locally advanced stage leiomyosarcoma (LMS), high-grade endometrial stromal sarcoma (HGESS), or undifferentiated uterine sarcoma (UUS), who underwent hysterectomy between 2011 and 2021 with or without preoperative endometrial biopsy. Exclusion criteria were patients with metastatic uterine sarcoma, due to the influence of distant metastases on surgical procedure, postoperative management, and prognosis. Heterologous sarcomas and malignant mixed tumors were also excluded. Diagnoses and treatment were performed in different German in- and outpatient hospitals or clinics. Histologic specimens were also reviewed by various German pathology institutes.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278278_p6
|
PMC11278278
|
sec[1]/p[1]
|
2. Material and Methods
| 4.011719 |
biomedical
|
Study
|
[
0.99365234375,
0.00592041015625,
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[
0.99853515625,
0.0006103515625,
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0.0003483295440673828
] |
Methods: Using available medical records, patient’s characteristics, symptoms, diagnostic procedures including sonographic findings, indications for endometrial biopsy, histopathological findings, following surgical approach, and survival data were collected and analyzed. Preoperative endometrial curettage was performed with or without hysteroscopy. Hysteroscopic findings were not analyzed and the role of hysteroscopy in the diagnosis was not evaluated in this study. A pipelle biopsy was not performed in any case. We used the term endometrial biopsy in the following. All histological findings of endometrial biopsy were categorized into clinically relevant benign and malignant groups, which could have an impact on the operative planning and were compared with the definitive final diagnosis of sarcoma.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p7
|
PMC11278278
|
sec[1]/p[2]
|
2. Material and Methods
| 3.054688 |
biomedical
|
Study
|
[
0.99658203125,
0.0025768280029296875,
0.0006103515625
] |
[
0.97509765625,
0.0234832763671875,
0.0006542205810546875,
0.0008993148803710938
] |
Surgical interventions with high-power morcellation, tumor perforation (spontaneous or iatrogenic), or other tumor injury were considered as inadequate surgical procedures. In this study, the term AUB refers exclusively to intermenstrual bleeding and postmenopausal bleeding. Heavy uterine bleeding was not added to AUB here.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278278_p8
|
PMC11278278
|
sec[1]/p[3]
|
2. Material and Methods
| 2.732422 |
biomedical
|
Study
|
[
0.994140625,
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] |
[
0.9931640625,
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0.0006957054138183594,
0.000530242919921875
] |
Because of the potential influence on prognosis and survival, data of menopausal status, tumor diameter, morcellation, adjuvant therapy including chemotherapy, and/or radiotherapy were determined in the two groups with and without endometrial biopsy.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p9
|
PMC11278278
|
sec[1]/p[4]
|
2. Material and Methods
| 3.890625 |
biomedical
|
Study
|
[
0.99951171875,
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[
0.99853515625,
0.0009469985961914062,
0.0004119873046875,
0.00007802248001098633
] |
Using SPSS version 27, OS rates of LMS and HGESS/UUS were determined by Kaplan–Meier analysis concomitant with the log-rank test. Differences in categorial variables were compared using the chi-square test or Fisher’s exact test and in continuous variables by means of the t -test. A p -value of <0.05 was considered statistically significant.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p10
|
PMC11278278
|
sec[1]/p[5]
|
2. Material and Methods
| 1.415039 |
biomedical
|
Other
|
[
0.97607421875,
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0.01043701171875
] |
[
0.2203369140625,
0.7705078125,
0.0015230178833007812,
0.007396697998046875
] |
Missing clinical data were requested by the attending facility. All patients included gave their written consent to data collection and the use of anonymized personal records for research purposes. All procedures performed in this study were in accordance with the ethical standards of the local Institutional Research Committee of University Medicine (identifier: B 034/19) and with the 1964 Helsinki declaration and its later amendments.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p11
|
PMC11278278
|
sec[2]/p[0]
|
3. Results
| 4.011719 |
biomedical
|
Study
|
[
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[
0.99853515625,
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From all 467 patients, a total of 52 (11.1%) patients showed distant metastasis (58.1% with and 41.9% without endometrial biopsy) after further clinical investigations by preoperative or mainly postoperative imaging (CT, MRI). Of the remaining 415 HRUS, 303 (73.0%) were LMS and 112 (27.0%) HGESS/UUS. Because of the highly comparable clinical characteristics as well as prognosis and outcome, HGESS and UUS were combined into one group.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
PMC11278278_p12
|
PMC11278278
|
sec[2]/p[1]
|
3. Results
| 4.105469 |
biomedical
|
Study
|
[
0.98193359375,
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] |
[
0.99658203125,
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All patients underwent preoperative vaginal and partially abdominal sonography. An endometrial biopsy was performed in 42.9% (n = 178) of patients, 33% (n = 100) of LMS, and 69.6% (n = 78) of HGESS/UUS ( Table 1 ). The frequency of AUB was significantly higher in HGESS/UUS (67.0%) than in LMS (49.8%) ( p < 0.01). Solitary AUB was the main indication for endometrial biopsy in 48% of LMS and 50% of HGESS/UUS. A combination of bleeding events with other symptoms, abnormal intracavitary uterine sonographic findings, macroscopically visible tumors in the cervical canal such as polyps, or myoma in the statu nascendi were the indications in 41.0% of LMS and 35.9% of HGESS/UUS cases. Indications due to symptoms other than bleeding events, including heavy menstrual bleeding, were recorded in 11.0% of LMS and 14.1% of HGESS/UUS.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p13
|
PMC11278278
|
sec[2]/p[2]
|
3. Results
| 4.046875 |
biomedical
|
Study
|
[
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[
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Sonographic uterine findings suspicious for sarcoma were described by Köhler et al. . Only intracavitary sonographic suspicious findings were considered indications for biopsy. Intramural suspicious findings without AUB were not an indication for biopsy. Imaging procedures mostly after endometrial biopsy including the cohort of M1 were performed by CT (abdomen and/or chest) in 40.8%, MRI (abdomen) in 8.2%, CT plus MRI in 7.8%, chest-X-ray in 2%, and PET-CT in 1.2%. In only five cases, the indications of endometrial biopsy were the results of imaging procedures together with a suspicious sonographic finding.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p14
|
PMC11278278
|
sec[2]/p[3]
|
3. Results
| 4.085938 |
biomedical
|
Study
|
[
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[
0.9990234375,
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A summary of the patients’ and tumors’ characteristics is shown in Table 1 . The mean age (years) of women who underwent endometrial biopsy (56.3 in LMS and 58.9 in HGESS/UUS) was higher than that of women who did not undergo endometrial biopsy (52.1 in LMS and 55.7 in HGESS/UUS), reaching statistical significance only in the LMS group ( p < 0.001). Postmenopausal status was observed in 49.3% (n = 100) of patients with LMS who did not receive endometrial biopsy and 66% (n = 66) of patients who did and in 64.7% (n = 22) of patients with HGESS/UUS without biopsy and 71.8% (n = 56) with biopsy. In summary, 51.5% (122) of patients with no biopsy and 68.5% (122) of patients with biopsy were postmenopausal.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p15
|
PMC11278278
|
sec[2]/p[4]
|
3. Results
| 4.136719 |
biomedical
|
Study
|
[
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] |
[
0.9990234375,
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] |
The tumor diameter of LMS and HGESS/UUS was significantly lower in the biopsy group (8.86 and 8.68 cm) compared to the non-biopsy group (11.47 and 12.69 cm) ( Table 1 ). Patients who underwent preoperative endometrial biopsy were significantly less likely to undergo inadequate surgery (28.0% (n = 28) of LMS and 38.5% (n = 30) of HGESS/UUS) than those who underwent surgery without biopsy (42.4% (n = 86) of LMS and 64.7% (n = 22) of HGESS/UUS). Morcellation of the tumor, the most common type of inadequate surgery, was significantly less common with biopsy at 21% (n = 21) of LMS and 11.5% (n = 9) of HGESS/UUS ( Table 1 ) than without biopsy at 33% (n = 67) of LMS and 32.4% (n = 11) of HGESS/UUS; in summary, 16.9% (n = 30) with biopsy versus 32.9% (n = 78) without (chi-square test p < 0.001).
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278278_p16
|
PMC11278278
|
sec[2]/p[5]
|
3. Results
| 3.955078 |
biomedical
|
Study
|
[
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] |
[
0.998046875,
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] |
The specific preoperative histological diagnosis by endometrial biopsy was correctly made in 30.0% of LMS and in 33.3% of HGESS/UUS ( Table 2 ).
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p17
|
PMC11278278
|
sec[2]/p[6]
|
3. Results
| 4.074219 |
biomedical
|
Study
|
[
0.99853515625,
0.00125885009765625,
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] |
[
0.9990234375,
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The abovementioned histologic findings were categorized into clinically relevant groups ( Table 3 ). Thus, the diagnosis of uterine sarcoma or unspecified malignant mesenchymal tumor was correct in 45.0% of LMS and 73.1% of HGESS/UUS. If misdiagnosed carcinosarcomas and unspecified carcinomas by endometrial biopsy were considered and added, the number of positive malignant findings increased to 78.2% of HGESS/UUS ( Table 3 ).
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999999 |
PMC11278278_p18
|
PMC11278278
|
sec[2]/p[7]
|
3. Results
| 4.070313 |
biomedical
|
Study
|
[
0.9990234375,
0.0006461143493652344,
0.00021386146545410156
] |
[
0.9990234375,
0.00028061866760253906,
0.0005359649658203125,
0.00008660554885864258
] |
Excluding the diagnosed variants of LM or STUMP, benign lesions (e.g., endometrial polyp adenomyosis, glandular hyperplasia, or endometrial stromal nodules) were diagnosed in 30.0% of LMS and 14.1% of HGESS/UUS. Normal histologic findings were described in 22.0% of LMS and only 7.7% of HGESS/UUS. Benign lesions and normal findings were significantly less frequent in HGESS/UUS (21.8%) than in LMS (52%) ( p < 0.01).
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278278_p19
|
PMC11278278
|
sec[2]/p[8]
|
3. Results
| 4.066406 |
biomedical
|
Study
|
[
0.99853515625,
0.0013380050659179688,
0.0002815723419189453
] |
[
0.9990234375,
0.00039958953857421875,
0.0004475116729736328,
0.0001195073127746582
] |
There was no significant difference in the 5-year and 10-year overall survival rates in patients suffering from high-risk uterine sarcomas (LMS and HGESS/UUS) with (46.9% and 30.9%, respectively) or without (53.8% and 36.8%, respectively) preoperative endometrial biopsy (log rank 0.09) .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p20
|
PMC11278278
|
sec[2]/p[9]
|
3. Results
| 3.300781 |
biomedical
|
Study
|
[
0.99072265625,
0.007843017578125,
0.0012044906616210938
] |
[
0.99658203125,
0.0021953582763671875,
0.0010423660278320312,
0.00041604042053222656
] |
There was also no significant difference in the 5-year and 10-year locoregional recurrence-free interval (log rank = 0.81) .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p21
|
PMC11278278
|
sec[2]/p[10]
|
3. Results
| 4.097656 |
biomedical
|
Study
|
[
0.9990234375,
0.0007777214050292969,
0.00025200843811035156
] |
[
0.9990234375,
0.0002689361572265625,
0.00038433074951171875,
0.00009185075759887695
] |
In contrast, the 5-year and 10-year distant recurrence-free interval was significantly higher in the non-biopsy group (58.3% and 46.4%, respectively) compared with the biopsy group (46.1% and 37.9%, respectively) (log rank = 0.046) , with a higher rate of distant metastases. In multivariable analysis, the distant metastasis-free interval was only dependent on intraoperative morcellation (HR 0.68, 95%-CI 0.48–0.96, p = 0.031) and postmenopausal status (HR 1.53, 95%-CI 1.11–2.1, p = 0.08), but not on the performed preoperative biopsy (HR 1.25, 95%-CI 0.92–1.69, p = 0.14).
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999994 |
PMC11278278_p22
|
PMC11278278
|
sec[2]/p[11]
|
3. Results
| 4.0625 |
biomedical
|
Study
|
[
0.9990234375,
0.0007305145263671875,
0.0003044605255126953
] |
[
0.99951171875,
0.00026726722717285156,
0.000324249267578125,
0.0000776052474975586
] |
In addition, we separately analyzed the impact of endometrial biopsy on survival data for LMS and HGESS/UUS (plots not shown). Regarding OS and locoregional recurrence-free interval, the data were consistent with those of all HRUS. An effect on 5- and 10-year distant metastasis-free interval was observed (56.4% and 37.6% without and 39.5% and 19.8% with biopsy, respectively) but was not statistically significant (log rank = 0.19).
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278278_p23
|
PMC11278278
|
sec[2]/p[12]
|
3. Results
| 4.105469 |
biomedical
|
Study
|
[
0.9990234375,
0.0007867813110351562,
0.0003185272216796875
] |
[
0.9990234375,
0.0002448558807373047,
0.00043582916259765625,
0.00007522106170654297
] |
Biopsy per se does not affect the local recurrence-free survival in morcellated and non-morcellated women (log rank p = 0.84). This finding is supported by the fact that in morcellated patients, biopsy also has no effect on local recurrence-free survival (log rank 0.361). In the non-biopsy group, morcellation is associated with significantly worse local recurrence-free survival as shown in univariate analysis (log rank < 0.001). However, in multivariate analysis, only morcellation (HR 1.76, CI 1.23–2.52, p = 0.002) and postmenopause (HR 1.48, CI 1.03–2.12, p = 0.03) had a significant influence on local recurrence-free survival, but not biopsy (HR 1.11, CI 0.77–1.58, p = 0.566).
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p24
|
PMC11278278
|
sec[2]/p[13]
|
3. Results
| 2.445313 |
biomedical
|
Study
|
[
0.8564453125,
0.138916015625,
0.004512786865234375
] |
[
0.92236328125,
0.0665283203125,
0.0015201568603515625,
0.00955963134765625
] |
A total of 28 patients received chemotherapy, 4 in the biopsy group and 24 in the non-biopsy group. In addition, 12 patients in each group received radiotherapy (brachytherapy with or without external beam radiation). The rate of chemotherapy and radiotherapy was too low to have an impact on the survival data.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p25
|
PMC11278278
|
sec[3]/p[0]
|
4. Discussion
| 3.910156 |
biomedical
|
Study
|
[
0.99951171875,
0.0003533363342285156,
0.0002818107604980469
] |
[
0.78662109375,
0.0038547515869140625,
0.208984375,
0.0005488395690917969
] |
The detection rate of high-risk uterine sarcomas (HRUSs) by preoperative endometrial biopsy is relatively low. The correct specific histologic diagnosis could be achieved in only 30.0% of leiomyosarcoma (LMS) and 33.3% of high-grade endometrial stromal sarcoma and undifferentiated uterine sarcoma (HGESS/UUS). In fact, there are no prospective or large retrospective studies on the impact of endometrial biopsy in uterine sarcoma in general and in HRUSs in particular, regardless of the indication. There are limited retrospective data with relatively small numbers of cases on the detection rate of uterine sarcomas by endometrial biopsy. Most publications only refer to leiomyosarcoma.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p26
|
PMC11278278
|
sec[3]/p[1]
|
4. Discussion
| 3.759766 |
biomedical
|
Study
|
[
0.9970703125,
0.0023975372314453125,
0.0004527568817138672
] |
[
0.9990234375,
0.0006771087646484375,
0.00019741058349609375,
0.00014472007751464844
] |
A total of 178 women (42.9% of all patients) underwent endometrial biopsy prior to surgery. The numbers were significantly higher in patients with HGESS and UUS (69.6%) than in LMS (33.0%), confirming the fact that AUB occurs significantly more often when HGESS and UUS are diagnosed.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999999 |
PMC11278278_p27
|
PMC11278278
|
sec[3]/p[2]
|
4. Discussion
| 4.046875 |
biomedical
|
Study
|
[
0.998046875,
0.0016193389892578125,
0.0002046823501586914
] |
[
0.97216796875,
0.002471923828125,
0.0247344970703125,
0.000530242919921875
] |
AUB as the main indication for endometrial biopsy is considered the main symptom in patients with uterine sarcoma. It indicates penetration of the sarcoma into the uterine cavity or primary development within the endometrium . AUB has been described in 46% to 57% of LMS and even more frequently in 56.4% to 65% of HGESS/UUS . AUB was the main indication for endometrial biopsy in 48% of LMS and 50% of HGESS/UUS, while a combination of AUB and abnormal intracavitary sonographic findings or a macroscopically visible tumor in the cervical canal was the indication in 41.0% of LMS and 35.9% of HGESS/UUS. Other symptoms and findings, as well as heavy menstrual bleeding, were the indication for endometrial biopsy in 11.0% of LMS and 14.1% of HGESS/UUS.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999999 |
PMC11278278_p28
|
PMC11278278
|
sec[3]/p[3]
|
4. Discussion
| 4.082031 |
biomedical
|
Study
|
[
0.9990234375,
0.0006055831909179688,
0.00024378299713134766
] |
[
0.99951171875,
0.00035119056701660156,
0.00029015541076660156,
0.00007992982864379883
] |
According to our data, AUB occurred mostly as intermenstrual or postmenopausal bleeding in about half of the patients with LMS (49.8%) and significantly more frequently in patients with HGESS/UUS (67.0%) ( p < 0.01). This difference is because HGESS and UUS usually arise in the endometrium and spread very quickly into the uterine cavity. In contrast, the majority of LMS are located primarily in the myometrium and invade the cavity at a later stage .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p29
|
PMC11278278
|
sec[3]/p[4]
|
4. Discussion
| 4.082031 |
biomedical
|
Study
|
[
0.9990234375,
0.0008592605590820312,
0.0001913309097290039
] |
[
0.9990234375,
0.00037026405334472656,
0.0006432533264160156,
0.00011903047561645508
] |
Our analysis showed that the correct specific histologic diagnosis by endometrial biopsy was unsatisfactorily low in 30.0% and 33.3% of LMS and HGESS/UUS, respectively. From the pathologists’ point of view, a correct diagnosis based on endometrial biopsy could be achieved in only 30% of cases . However, if the results refer to all malignant uterine tumors (including carcinosarcomas and carcinomas) detected by endometrial biopsy, the detection rate is significantly higher with up to 45.0% in LMS and 78.2% in HGESS/UUS. Relevant preoperative misdiagnosis (benign lesions and normal findings) was observed in 52.0% of LMS and 21.8% of HGESS/UUS.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
PMC11278278_p30
|
PMC11278278
|
sec[3]/p[5]
|
4. Discussion
| 4.03125 |
biomedical
|
Study
|
[
0.9990234375,
0.0006551742553710938,
0.00014591217041015625
] |
[
0.99853515625,
0.0005731582641601562,
0.0007996559143066406,
0.0001418590545654297
] |
Kho et al. recently published a similar analysis of 79 patients with uterine LMS who underwent endometrial sampling with or without diagnostic hysteroscopy followed by hysterectomy; 31.6% of LMS could be diagnosed preoperatively by endometrial sampling alone and 66.7% when sampling was combined with hysteroscopy . The impact of diagnostic hysteroscopy on the detection rate of endometrial biopsy was not evaluated in this study.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278278_p31
|
PMC11278278
|
sec[3]/p[6]
|
4. Discussion
| 3.976563 |
biomedical
|
Study
|
[
0.99951171875,
0.00035119056701660156,
0.0003666877746582031
] |
[
0.9677734375,
0.0004360675811767578,
0.031646728515625,
0.00017714500427246094
] |
Hinchcliff et al. evaluated the role of endometrial biopsy in the preoperative detection of 68 uterine LMS and showed that 51.5% of cases were diagnosed preoperatively as LMS or spindle cell proliferation. Only 35% of cases were correctly diagnosed as uterine LMS . In the study of Skorstad et al., 142 of 209 women (67.9%) with LMS underwent preoperative endometrial biopsy. A malignant finding without a specific diagnosis was described in 38.7% of cases . In the study of Bansal et al., preoperative endometrial sampling suggested an invasive tumor in 86% and could predict the general histologic diagnosis of sarcoma in only 64%, but not the specific entity . Therefore, an exact comparison with our data was not possible.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p32
|
PMC11278278
|
sec[3]/p[7]
|
4. Discussion
| 3.771484 |
biomedical
|
Study
|
[
0.99951171875,
0.0005578994750976562,
0.0001742839813232422
] |
[
0.99560546875,
0.0018177032470703125,
0.0021991729736328125,
0.000324249267578125
] |
Studies and case reports of endometrial biopsy for stromal sarcoma are even rarer than for LMS. Jin et al. reported only 23 cases of UUS and 47 cases of LGESS; endometrial biopsy was performed in only 65% and 29.8% of cases, respectively. Overall, the correct diagnosis was achieved in 89.6% of this small group .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999999 |
PMC11278278_p33
|
PMC11278278
|
sec[3]/p[8]
|
4. Discussion
| 3.556641 |
biomedical
|
Study
|
[
0.99853515625,
0.0013246536254882812,
0.00018715858459472656
] |
[
0.9892578125,
0.00772857666015625,
0.0018301010131835938,
0.0009984970092773438
] |
Li et al. described 114 cases of uterine sarcoma. Preoperative endometrial biopsy was performed in 48 patients. A diagnosis of uterine malignancy was made in 36 patients (75%) .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p34
|
PMC11278278
|
sec[3]/p[9]
|
4. Discussion
| 2.933594 |
biomedical
|
Study
|
[
0.998046875,
0.0012493133544921875,
0.0005435943603515625
] |
[
0.9951171875,
0.004058837890625,
0.0002624988555908203,
0.00034928321838378906
] |
In a previous study of the French Sarcoma Group, data from 39 patients with HGESS and UUS were analyzed. Only 15 patients (38.5%) underwent preoperative endometrial biopsy, and 7 patients (46.6%) were found to have uterine sarcoma .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p35
|
PMC11278278
|
sec[3]/p[10]
|
4. Discussion
| 3.416016 |
biomedical
|
Study
|
[
0.9990234375,
0.0007596015930175781,
0.0002956390380859375
] |
[
0.99658203125,
0.0025348663330078125,
0.0005383491516113281,
0.0002772808074951172
] |
Tanner et al. studied 21 uterine sarcomas (HGESS and UUS); 11 patients (50%) received endometrial biopsy. The pathological diagnosis was correct in seven (64%) of the analyzed tumors .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p36
|
PMC11278278
|
sec[3]/p[11]
|
4. Discussion
| 3.146484 |
biomedical
|
Study
|
[
0.998046875,
0.0016803741455078125,
0.00044345855712890625
] |
[
0.97705078125,
0.0206146240234375,
0.001369476318359375,
0.0008721351623535156
] |
Wais et al. identified 302 patients with various uterine sarcomas, and 114 (51%) underwent endometrial sampling. There was evidence of sarcoma in 65% of cases. The main indications for this procedure were AUB including PMB .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p37
|
PMC11278278
|
sec[3]/p[12]
|
4. Discussion
| 4.070313 |
biomedical
|
Study
|
[
0.99951171875,
0.0004131793975830078,
0.0001881122589111328
] |
[
0.97705078125,
0.0009016990661621094,
0.0217132568359375,
0.00023245811462402344
] |
Despite efforts, there are still no reliable diagnostic procedures to differentiate between uterine leiomyoma and sarcoma . Transcervical core needle biopsycan only be applied in order to achieve clarification in cases in which the diagnosis remains uncertain despite having performed all of the known diagnostic measures . However, this is definitely not a diagnostic method for intermenstrual and postmenopausal bleeding. Nevertheless, experiences with this approach have been surprisingly positive in selected cases. In one study, transcervical core needle biopsies were taken from 453 women whose tumors were LMS-suspicious (high SI in T1/2 W MRI and/or high LDH-values and/or rapidly growing tumor with 3-fold gain in volume within one year) . Among 372 eligible cases, only seven sarcomas were found. Nonetheless, false-negative results must be reconsidered, in particular for spindle-cell sarcomas . One reason for this is because mitoses, atypia, and necroses can in fact be concentrated in so-called “hot spots”. There is a real risk of missing these hot spots using core biopsy, so that, essentially, only positive findings can be truly relied on.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p38
|
PMC11278278
|
sec[3]/p[13]
|
4. Discussion
| 4.082031 |
biomedical
|
Study
|
[
0.9990234375,
0.0006651878356933594,
0.0001583099365234375
] |
[
0.9990234375,
0.0003161430358886719,
0.0005927085876464844,
0.00011444091796875
] |
According to our data, the preoperative histologic diagnosis of sarcoma has an important impact on the mode of sarcoma surgery, whereas misdiagnosis could lead to incorrect surgical treatment, resulting in suboptimal therapeutic approaches, prolonging the operative time, or in some cases leading to a second surgical procedure. As expected after endometrial biopsy, the indication of sarcoma for surgery increased from 26.4% to 57% in LMS and from 38.3% to 84.6% in HGESS/UUS. Consistently, the number of inadequately treated patients decreased from 42.4% to 28% in LMS and from 64.7% to 38.5% in HGESS/UUS ( p < 0.05 for both groups). The number of morcellated sarcomas decreased significantly from 33.0% to 21% in LMS ( p = 0.016) and from 32.4% to 11.5% in HGESS/UUS ( p = 0.017). To the best of our knowledge, there are no studies in the literature on the impact of preoperative endometrial biopsy on surgical approach.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p39
|
PMC11278278
|
sec[3]/p[14]
|
4. Discussion
| 4.070313 |
biomedical
|
Study
|
[
0.9990234375,
0.00072479248046875,
0.0002446174621582031
] |
[
0.99951171875,
0.0002372264862060547,
0.00036025047302246094,
0.00008541345596313477
] |
However, based on our data, preoperative endometrial biopsy does not affect overall survival in patients with HRUS. Even when the histologic result of endometrial biopsy was considered whether malignant or non-malignant, no significant difference in overall survival was seen in both groups. This is consistent with the results published by Kho et al. with a median overall survival of 50 months in LMS and no significant difference between patients diagnosed before or after hysterectomy . The biopsy group in our study had more prognostically favorable variables, such as a smaller tumor diameter and lower morcellation rate, than unfavorable variables, such as postmenopausal status (the latter only for LMS) ( Table 1 ).
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p40
|
PMC11278278
|
sec[3]/p[15]
|
4. Discussion
| 3.921875 |
biomedical
|
Review
|
[
0.99853515625,
0.0008997917175292969,
0.0005655288696289062
] |
[
0.309814453125,
0.004589080810546875,
0.68505859375,
0.0006508827209472656
] |
The correlation between intrabdominal morcellation of uterine sarcoma and outcome were reported previously . However, only a few studies have evaluated the long-term outcomes in these patients, with a clear impact on disease-free interval and very heterogenous results regarding overall survival . Similar results regarding morcellation and outcomes in uterine smooth muscle tumor of uncertain malignant potential were demonstrated . Morcellation intraabdominally should be avoided, if the diagnosis of sarcoma could not be excluded, otherwise specimen retrieval via a mini-laparotomic incision seems to be a suitable alternative .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
PMC11278278_p41
|
PMC11278278
|
sec[3]/p[16]
|
4. Discussion
| 4.195313 |
biomedical
|
Study
|
[
0.9990234375,
0.0006928443908691406,
0.0003504753112792969
] |
[
0.9990234375,
0.0002524852752685547,
0.0005726814270019531,
0.00009679794311523438
] |
The effect of endometrial biopsy on recurrence-free interval has not been investigated in the literature. As shown in a study from our group, tumor morcellation doubles the risk of locoregional recurrence rate . As tumor morcellation was performed significantly less often with biopsy (32.5% with vs. 55.4% without biopsy, p < 0.001), the locoregional recurrence-free interval remained unaffected between the two groups (log rank = 0.81). Regarding morcellation, biopsy per se does not affect the local recurrence-free survival in morcellated and non-morcellated women (log rank p = 0.84). In the non-biopsy group, morcellation is associated with significantly worse local recurrence-free survival as shown in univariate analysis (log rank < 0.001) (graphs not shown). However, in multivariate analysis, only morcellation (HR 1.76, CI 1.23–2.52, p = 0.002) and postmenopause (HR 1.48, CI 1.03–2.12, p = 0.03) but not the biopsy significantly influenced the local recurrence-free survival (HR 1.11, CI 0.77–1.58, p = 0.566). The reduction in the morcellation rate is obviously an advantage of biopsy, although it does not influence the local recurrence rate of morcellated patients. As was also shown in a previous study , tumor morcellation significantly halves the risk of distant metastases (HR = 0.52). This may explain why the distant metastasis-free interval is longer without biopsy, because the proportion of morcellated tumors is significantly higher in this group. But, in the multivariable analysis, distant metastasis was only dependent on morcellation (HR 0.68, 95%-CI 0.48–0.96, p = 0.031) and postmenopausal status (HR 1.53, 95%-CI 1.11–2.1, p = 0.08) but not on biopsy performed (HR 1.25, 95%-CI 0.92–1.69, p = 0.14). As the proportion of postmenopausal women in both groups is not significantly different, only the effect of morcellation on the recurrence-free interval of distant metastases remains. The phenomenon of morcellation on distant metastases cannot be explained at present and has already been discussed in the work of Reichert et al. . Similar results were reported by Nobre .
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p42
|
PMC11278278
|
sec[3]/p[17]
|
4. Discussion
| 1.90332 |
biomedical
|
Study
|
[
0.98828125,
0.005619049072265625,
0.006206512451171875
] |
[
0.94091796875,
0.055908203125,
0.0016794204711914062,
0.0017261505126953125
] |
Due to the small number of cases who underwent radiotherapy or chemotherapy, and their unproven efficacy, chemotherapy and radiotherapy were not included or considered in this analysis.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278278_p43
|
PMC11278278
|
sec[3]/p[18]
|
4. Discussion
| 4.003906 |
biomedical
|
Study
|
[
0.99951171875,
0.0003943443298339844,
0.0002033710479736328
] |
[
0.9990234375,
0.00029587745666503906,
0.0005130767822265625,
0.00008863210678100586
] |
Strengths and limitations of this study: Our study used centralized data and was limited by its observational retrospective nature, which may introduce selection bias. The data were collected from several centers in Germany and the histological examination was performed in different institutions, which limits our ability to identify factors that may affect the quality of the results. As mentioned in the Section 2 , the comparison with the current literature was limited by the fact that our study only examined complete endometrial curettage, whereas other groups used different methods of analysis, such as endometrial sampling or endometrial biopsy, according to the literature.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
PMC11278278_p44
|
PMC11278278
|
sec[4]/p[0]
|
5. Conclusions
| 4.085938 |
biomedical
|
Study
|
[
0.99951171875,
0.00035691261291503906,
0.0001844167709350586
] |
[
0.99853515625,
0.0006356239318847656,
0.0007433891296386719,
0.00010055303573608398
] |
The detection rate of histologically proven high-risk uterine sarcoma (HRUS) by endometrial biopsy is up to 33%; meanwhile, its sensitivity is unknown when endometrial biopsy was performed due to abnormal uterine bleeding or abnormal intracavitary sonographic finding. Uterine malignancies, including misdiagnosed carcinomas and carcinosarcomas, were detected in 45.0% of LMS and 78.2% of HGESS/UUS by endometrial biopsy.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p45
|
PMC11278278
|
sec[4]/p[1]
|
5. Conclusions
| 3.978516 |
biomedical
|
Study
|
[
0.99853515625,
0.0010395050048828125,
0.0005025863647460938
] |
[
0.86328125,
0.00705718994140625,
0.1290283203125,
0.00051116943359375
] |
The preoperative diagnosis of HRUSs by endometrial biopsy has a significant impact on the surgical procedure. Therefore, inappropriate surgery, mainly in the form of tumor morcellation, is performed significantly less. Preoperative endometrial biopsy does not affect OS. By reducing the rate of tumor morcellation, endometrial biopsy reduces the risk of the locoregional recurrences induced by this inadequate procedure. This appears to be a benefit of biopsy. According to the available data, the distant recurrence-free interval is only indirectly prolonged in the non-biopsy group.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278278_p46
|
PMC11278278
|
sec[4]/p[2]
|
5. Conclusions
| 3.875 |
biomedical
|
Other
|
[
0.994140625,
0.005252838134765625,
0.000705718994140625
] |
[
0.1539306640625,
0.70068359375,
0.1419677734375,
0.0031890869140625
] |
Endometrial biopsy is a cornerstone in the management of AUB, no matter whether sarcoma or other malignant uterine tumors are expected. However, benign or normal findings, observed in up to 52% of LMS and 21.8% of HGESS/UUS, can also provide a false sense of security. To minimize the rate of false negative findings, only clearly indicated endometrial biopsies should be performed.
|
[
"Zaher Alwafai",
"Verena M. C. Reichert",
"Paula Spring",
"Marek Zygmunt",
"Günter Köhler"
] |
https://doi.org/10.3390/jcm13144048
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278294_p0
|
PMC11278294
|
sec[0]/p[0]
|
Introduction
| 4.253906 |
biomedical
|
Study
|
[
0.9990234375,
0.000316619873046875,
0.0004944801330566406
] |
[
0.9287109375,
0.0021343231201171875,
0.06878662109375,
0.0002167224884033203
] |
In vivo analysis provides more accurate information for indicating or predicting the processes occurring in the complex living organisms compared with the ex vivo study. Combining in vivo sampling with liquid chromatography (LC)/gas chromatography-mass spectrometry (GC-MS) analysis is the most powerful technique for obtaining comprehensive information, as it can yield both targeted and nontargeted data with quantitative capability. Microdialysis (MD) is a widely used method for in vivo sampling over the years, especially valued for its ability to collect samples continuously over time. However, MD analysis is limited by a few drawbacks that create the need for complementary and/or alternative sampling methods. First, sampling of nonpolar compounds can present a problem in MD analysis. As nonpolar compounds are typically highly bound to tissue matrix, they are only available in very low free concentration levels that limit their detection and quantitation. In some cases, such compounds can also adsorb onto the membrane and/or tubes used in MD. Secondly, collected samples require subsequent sample preparation, such as liquid extraction or solid-phase extraction, prior to chromatographic separation and MS analysis. In addition to being time-consuming and labour intensive, these additional sample preparation steps can also lead to the decomposition of some unstable compounds, and thus negatively impact the accuracy and precision of the final data.
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999994 |
PMC11278294_p1
|
PMC11278294
|
sec[0]/p[1]
|
Introduction
| 4.347656 |
biomedical
|
Study
|
[
0.9990234375,
0.00046515464782714844,
0.0006995201110839844
] |
[
0.5546875,
0.00597381591796875,
0.438720703125,
0.0006074905395507812
] |
Solid-phase microextraction (SPME), in which a small amount of extraction phase is immobilized onto a solid support such as a fiber to extract analytes from samples, combines sampling, sample preparation, and enrichment into a single step. In human applications, SPME was first successfully applied for in vivo analysis of human breath, saliva, blood in vein and skin surfaces. The subsequent development of biocompatible coating materials using biosafe coating and supporting material has expanded the use of SPME and enabled in vivo analysis of tissue through direct insertion of fibers into the tissues. The miniaturized nature of SPME enables minimally invasive sampling, while its non-exhaustive extraction mechanism causes minimal perturbation to the living system, as only a small portion of analytes is extracted via free concentration . These advantages allow researchers to acquire ‘snapshots’ of unique biological processes taking place in regions not otherwise easily sampled via traditional methods. For instance, they have permitted an in-depth metabolomics study with high temporal and spatial resolution by in vivo SPME sampling of brain tissue of awake and freely moving rats . The using of in vivo SPME instead of regular tissue biopsy can also alleviate animal suffering and reduce the consumption of animal life.
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278294_p2
|
PMC11278294
|
sec[0]/p[2]
|
Introduction
| 4.1875 |
biomedical
|
Study
|
[
0.99951171875,
0.00043392181396484375,
0.00019407272338867188
] |
[
0.93603515625,
0.00313568115234375,
0.06005859375,
0.00054168701171875
] |
After more than 10 years adventures, going through ex vivo biofluidic, tissue, in vivo mouse, rat, dog and monkey studies, in vivo SPME has been finally used for human applications in the monitoring of doxorubicin (DOX) concentration during the in vivo lung perfusion (IVLP) surgery . Pulmonary metastases are secondary cancerous cells that develop in approximately 30% of patients with malignant tumors. IVLP is used to locally deliver DOX, a powerful anti-cancer drug that can cause multi-organ toxicities, to isolated lung tissue during surgical resection in an effort to reduce its toxicity to other organs while enabling the use of higher doses in the targeted organs. However, during IVLP, the concentration levels and distribution of DOX in different parts of the lung must be precisely controlled so as to prevent insufficient application or excessive dosing. While traditional methods of monitoring that rely on tissue biopsy collected from the terminal parts of the organ are invasive and cannot provide spatial resolution, the successful implementation of in vivo SPME-LC-MS for analysis of lung tissue of two patients undergoing IVLP surgery for a period of 3 h has enabled quantitative results with both temporal and spatial resolution. This ground-breaking achievement thus opens new horizons for in vivo SPME bioanalytical applications in medicine and beyond. Within this context, we provide here a brief future perspective on in vivo SPME focusing on human applications. Fig. 1 Coupling in vivo solid-phase microextraction (SPME) with (A) liquid chromatography-mass spectrometry (LC-MS) for post-surgery analysis or (B) direct MS for real-time analysis during surgery. ESI: electrospray ionization; MOI: microfluidic open interface. Fig. 1
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278294_p3
|
PMC11278294
|
sec[1]/p[0]
|
Perspectives on the future of in vivo SPME devices
| 3.978516 |
biomedical
|
Study
|
[
0.99951171875,
0.00020694732666015625,
0.0004448890686035156
] |
[
0.98486328125,
0.01476287841796875,
0.00036644935607910156,
0.00017547607421875
] |
In the aforementioned IVLP application, a commercially available biocompatible SPME fiber (diameter, 285 μm; coating thickness, 45 μm; coating length, 15 mm) using a mixed-mode coating (C8 + benzenesulfonic acid particles embedded in polyacrylonitrile) while affixed to a nitinol support was used. The device can be further optimized to maximize its applicability for in vivo tissue analysis.
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278294_p4
|
PMC11278294
|
sec[1]/p[1]
|
Perspectives on the future of in vivo SPME devices
| 4.15625 |
biomedical
|
Study
|
[
0.9990234375,
0.00041961669921875,
0.0005326271057128906
] |
[
0.84619140625,
0.009429931640625,
0.14404296875,
0.0004353523254394531
] |
Firstly, although the nitinol support is chemically and physically stable, it can be easily bent during insertion into lung tissue, thus requiring special care during the operation. Additionally, traversing into resistant tissue such as muscle requires the use of a sheath needle or guide cannula, which adds an inconvenient step to the workflow. Given that nitinol is a metal characterized by a darker hue and the device is small, doctors observed that particular care needs to be taken to not lose fibers placed in the tissue, especially when multiple fibers are used simultaneously. Further optimization of the device should include both the use of a stronger support material such as a medical-grade stainless-steel needle, and the addition of a suitable holder on top of the devices to facilitate visual tracking and removal. Secondly, the mixed-mode coating used for this application is usually activated by organic solvents prior to sampling to ensure better extraction efficiency and reproducibility. However, sterilization of the fibers prior to insertion by autoclave deactivates the coating by removing the solvents, thus resulting in lower efficiency and reproducibility. In this regard, future directions should take into consideration both the development of new fiber sterilization methods and the development of new coating materials that do not require activation via solvents. Finally, the continued development of highly efficient coating materials for SPME is important for the analysis of different target analytes. Nowadays, most of the coating materials for SPME are primarily suitable for extraction of nonpolar compounds; the development of biocompatible coating materials with good extraction efficiency towards polar analytes would greatly increase the applicability of SPME.
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278294_p5
|
PMC11278294
|
sec[2]/p[0]
|
Direct coupling of in vivo SPME with MS: enabling real time monitoring during surgery
| 4.089844 |
biomedical
|
Study
|
[
0.99951171875,
0.0004382133483886719,
0.00016951560974121094
] |
[
0.9873046875,
0.01056671142578125,
0.001918792724609375,
0.0002760887145996094
] |
In the aforementioned application, samples extracted via in vivo SPME were submitted to LC-MS analysis after surgery for investigating the general trends of metabolic processes. However, for point-of-care analysis, particularly for individuals with advanced-stage cancer, metabolic processes can vary significantly among patients due to the disparities in their physiological conditions. In this sense, real-time monitoring of anti-cancer drugs during surgery could provide useful information to the medical team, enabling more precise adjustments to dosage in different targeted areas throughout the surgery while also helping inform doctors on other decisions related to the surgical procedure. For example, IVLP requires re-perfusion with clean blood following DOX delivery to remove any drug residue in tissue after chemotherapy; knowing the exact drug concentration left in the tissue would aid physicians in timing the re-perfusion step adequately.
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278294_p6
|
PMC11278294
|
sec[2]/p[1]
|
Direct coupling of in vivo SPME with MS: enabling real time monitoring during surgery
| 4.140625 |
biomedical
|
Study
|
[
0.99951171875,
0.00023818016052246094,
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] |
[
0.93115234375,
0.01280975341796875,
0.055694580078125,
0.0003497600555419922
] |
While ambient MS techniques developed for real-time MS analysis, such as the intelligent knife and the mass spectrometer pen, are available for diverse medical applications, these are limited to providing qualitative information on cut tissue (knife) or the surface of tissue (pen). Coupling in vivo SPME to direct/ambient MS analysis could provide quantitative data with spatial resolution via insertion of SPME needles into the targeted tissue areas followed by subsequent instrumental analysis. Taking advantage of the sample clean up function of SPME, rinsing fibers with water for a few seconds following extraction to remove nonspecific attachments on the coating would be sufficient for direct desorption and analysis by MS. For such applications, the newly developed automated microfluidic open interface (MOI) could be employed to facilitate the instrumental workflow following extraction from tissue . Using this automated interface, a medical professional would just need to place the rinsed fiber into an open chamber and then remove it after 10 s; data would load onto the computer software interface in real time to quickly aid the team in adjusting concentration levels as needed. In addition to the MOI, other direct/ambient MS techniques such as nano electrospray ionization (ESI)-MS or probe ESI (PESI)-MS can also be considered.
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278294_p7
|
PMC11278294
|
sec[2]/p[2]
|
Direct coupling of in vivo SPME with MS: enabling real time monitoring during surgery
| 2.744141 |
biomedical
|
Other
|
[
0.994140625,
0.0016574859619140625,
0.004344940185546875
] |
[
0.090576171875,
0.90771484375,
0.0009417533874511719,
0.0006284713745117188
] |
The biggest challenge posed by the proposed direct MS technique is that the MS instrument should be placed in close proximity to where the extractions are being performed, either in the surgery room or somewhere adjacent. Traditional MS instruments can be quite large and noisy due to the vacuum pumps; as such, the use of portable MS instruments might be a solution to this limitation.
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999999 |
PMC11278294_p8
|
PMC11278294
|
sec[3]/p[0]
|
Future applications: from targeted drug monitoring to untargeted metabolomics studies
| 3.960938 |
biomedical
|
Review
|
[
0.99755859375,
0.0004627704620361328,
0.00213623046875
] |
[
0.2568359375,
0.00717926025390625,
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0.00035071372985839844
] |
For in vivo applications, SPME has its unique feature when compared with other techniques. At present, only two distinct applications of in vivo SPME have been validated for human use, DOX monitoring during IVLP and, more recently, untargeted profiling of human brain tissue . The development and implementation of in vivo SPME for human applications is not a venture initiated from ground zero; SPME has already been employed in a variety of applications using large animal models for different purposes such as monitoring of folinic acid, 5-fluorouracil, and oxaliplatin during IVLP, quality assessments of graft (such as kidney and liver) transplantations, and monitoring of dynamic changes during ex vivo heart perfusion. Recent human applications provide robust evidence that translations of in vivo SPME applications using large animal models (such as pig and monkey) to human studies are feasible and bolster our confidence in expanding the use of this technology for various future applications.
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278294_p9
|
PMC11278294
|
sec[3]/p[1]
|
Future applications: from targeted drug monitoring to untargeted metabolomics studies
| 3.78125 |
biomedical
|
Other
|
[
0.9990234375,
0.0003204345703125,
0.0007290840148925781
] |
[
0.1878662109375,
0.79296875,
0.0186767578125,
0.0006699562072753906
] |
In addition to in vivo SPME sampling for various applications, integration with advanced analytical technologies, like direct MS for rapid quantification of target analytes, or LC/GC coupled with high-resolution MS and ion mobility-MS for untargeted metabolomics screening, can yield comprehensive insights into living systems.
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
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https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278294_p10
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PMC11278294
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sec[4]/p[0]
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CRediT author statement
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other
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Other
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[
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Wei Zhou : Writing - Original draft preparation, and Reviewing and Editing; Runshan W. Jiang : Writing - Reviewing and Rditing; Barbara Bojko : Writing - Reviewing and Editing; Janusz Pawliszyn : Supervision, Writing - Reviewing and Editing.
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
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PMC11278294
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Declaration of competing interest
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other
|
Other
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The authors declare that there are no conflicts of interest.
|
[
"Wei Zhou",
"Runshan Will Jiang",
"Barbara Bojko",
"Janusz Pawliszyn"
] |
https://doi.org/10.1016/j.jpha.2023.12.008
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
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39057378
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1. Introduction
| 4.277344 |
biomedical
|
Study
|
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More than 200 species of the fungal genus Colletotrichum devastatingly affect a wide array of agricultural crops including vegetables, fruits, forest trees, cereals, and legumes by causing anthracnose, fruit rot, and blights . These fungi are ranked among the top ten most significant pathogens in the world because of their omnipresence and economic impact, and they provide model pathosystems for plant–pathogen interaction studies . Unlike some Colletotrichum species with narrow host ranges , the C. acutatum species complex (CASC) and the C. gloeosporioides species complex (CGSC) exhibit broader host specificity. These complexes encompass multiple species that cause rot disease in apple and various other crops, including small fruits, citrus, and peach. Some of the dominant species causing apple bitter rot in the Mid-Atlantic are C. fioriniae , C. chrysophilum, and C. noveboracense . The enormous economic loss imposed by bitter rot on the apple industry could be managed by fungicide application or using resistant cultivars, which are currently not available. However, these strategies are unattainable due to the diverse genetic makeup of Colletotrichum species, lack of known genetic resistance within domesticated apple germplasm , and the development of fungicide-resistant strains. Therefore, a thorough investigation at the genomic level to explore and discover potential pathogenic mechanisms of Colletotrichum species is needed.
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
39057378_p1
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39057378
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sec[0]/p[1]
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1. Introduction
| 4.199219 |
biomedical
|
Study
|
[
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Hemibiotrophism is the most common lifestyle utilized by Colletotrichum species. To carry out this lifestyle, Colletotrichum spp. use various specialized structures to colonize multiple plant parts such as leaves, stems, and fruits. Upon landing on the plant tissue, fungal conidia adhere to the host surface and form specialized structures called appressoria which allow the fungus to penetrate the host cuticle via a penetration peg forming at the base of appressorium . The primary hyphae emerging from the penetration peg accelerate nutrient acquisition from colonized plant tissues. The switch from biotrophy to the necrotrophic phase is directed by the production of secondary infectious hyphae which then will invade neighboring cells and kill the host tissue .
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
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1. Introduction
| 4.878906 |
biomedical
|
Study
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Establishing a successful infection by Colletotrichum species relies on a large spectrum of expressed and secreted pathogenicity and virulence factors such as effector proteins, carbohydrate active enzymes (CAZyme), and secondary metabolites . Effectors are both proteins or non-proteinaceous molecules secreted into plant hosts to facilitate infection, compromise the plant defense system, and/or in some cases trigger the host defense responses . Given the diverse host range of Colletotrichum spp., there are specialized effectors developed for fine-tuned adjustments to specific hosts or conserved effectors mediating infection processes for multiple hosts . CAZymes directly contribute to the degradation of plant cell wall components to liberate carbohydrates upon host colonization that also serve as carbon sources for fungal growth and proliferation . CAZymes are categorized into six major groups: glycoside hydrolases (GHs), glycosyl transferases, polysaccharide lyases (PLs), carbohydrate esterases (CEs), auxiliary activity enzymes (AA), and carbohydrate-binding modules (CBMs) . CAZymes in Colletotrichum spp. outnumber those of any other ascomycete fungi sequenced so far . Fungi owe their adaptability, survival, and pathogenicity to secondary metabolites (SMs), which in a fungal context, refers to the production of molecules termed natural products or secondary metabolites that are not part of primary metabolism. SMs are small molecules, synthesized by enzymes encoded by biosynthetic gene clusters (BGCs) in their genomes . Secondary metabolites are of great research interest as these molecules include toxins and beneficial natural products such as antibiotics, and they are important for ecological interactions of fungi with other microbes and their environment. In comparison to fungi like Verticillium and Fusarium , some Colletotrichum species contain higher numbers of SM gene clusters in genomes varying from 41 in C . salicis to 73 in C. truncatum . The identification and comparison of pathogenicity gene repertoires of different Colletotrichum species is paramount to understand plant–pathogen interaction, assess the present risks of these species, and ultimately develop economic and effective disease management approaches.
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
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1. Introduction
| 4.160156 |
biomedical
|
Study
|
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Comparative genomics and genome-scale analyses of Colletotrichum spp. offer an invaluable platform for unraveling the intricate patterns underlying speciation, pathogenesis, host specificity, and evolutionary relationships within this genus . Thanks to advancements in high-throughput sequencing, over 250 fully sequenced genomes of Colletotrichum are currently available in the National Center for Biotechnology Information (NCBI) repository . However, given the growing number of species and species complexes in the genus, more studies on genome sequencing and genome-mining analyses are needed to fill the existing research gaps. Therefore, conducting comparative genomics analyses and an analysis of the pathogenicity of gene repertoires of Colletotrichum species in CASC and CGSC, particularly for newly identified species like C. noveboracense and C. chrysophilum in apple, not only will shed light on host specificity, the pathogenic adaptation of these species, and evolutionary relationships but will also help to evaluate the present and future risk of these pathogens in the agricultural industry.
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
39057378_p4
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39057378
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|
1. Introduction
| 4.171875 |
biomedical
|
Study
|
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The aim of the current study was to utilize genomics in different Colletotrichum spp. to: 1. provide genomics resources for a total of nine isolates of four Colletotrichum species ( C. fioriniae , C. chrysophilum , C. noveboracense , and C. nupharicola ) that have not yet been sequenced and/or are from isolates from a specific geographic region, 2. construct phylogenetic trees using newly sequenced genome data to compare to previous MLST findings, 3. discover and discuss CAZyme, effector, and SM gene clusters within and across Colletotrichum spp. Our immediate and long-term goals are to identify targets for functional gene studies that may mediate pathogenicity, virulence, and/or quiescence in the different fungal species that can be targeted for the development of new control strategies (e.g., dsRNA development). In addition, we would like to explore more omics-based technologies (e.g., transcriptomics, proteomics, and metabolomics) using these isolates and their genomes to ascertain the biological underpinnings of their widespread success and observed geographic dominance in specific areas throughout the country.
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
39057378_p5
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39057378
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2.1. Fungal Strains, Genomic DNA Extraction, and Whole Genome Sequencing
| 4.21875 |
biomedical
|
Study
|
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Full genome sequences of two isolates of C. fioriniae (ACFK5 and ACFK16), three isolates of C. chrysophilum (AFK154, AFK26, and PMKnsl-1), and one isolate of C. noveboracense (PMBrms-1) were obtained from distinctive lesions on bitter rot-infected apple fruit in New York, Pennsylvania, and Virginia . In addition, the genomes of C. noveboracense Coll940 collected from Juglans nigra in Oklahoma and two isolates of C. nupharicola (CBS470 and Coll922) were isolated from yellow waterlily ( Nuphar lutea ) in Washington and New Jersey, respectively . The genomic DNA was extracted from 5-day-old single-spore cultures via the protocol described by Yelton et al. with slight modification. The whole genome sequencing was performed by the Beijing Genomics Institute (BGI) in Shenzhen, China, using the DNBSEQ short read platform (MGI Tech Co., Ltd. a subsidiary of BGI Group, Shenzhen, China) for 350 bp libraries with the paired-end 150 bp sequencing strategy, as described in the DNBseq De Novo service overview manual (BGI) yielding 46× to 103× coverage, sufficient for genome assembly. Genomes were assembled using SPADES 3.15.2 (Center for Algorithmic Biotechnology, St. Petersburg State University, St. Petersburg, Russia) for isolates Coll940 and AFK154 and MEGAHIT v1.2.9 (Computational Genomics Lab, Hong Kong University of Science and Technology, Kowloon, Hong Kong, China) for isolates CBS470, ACFK5, ACFK16, PMKnsl-1, AFK26, PMBrms-1, and Coll922, and annotated using the MAKER pipeline (University of Utah, Salt Lake City, UT, USA). More detailed materials and methods are described in Khodadadi et al. .
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
39057378_p6
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39057378
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2.2. Phylogenomics
| 4.394531 |
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|
Study
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Three independent approaches were utilized to analyze the phylogeny of the Colletotrichum species selected for this study. First, high-throughput average nucleic identity (ANI) was performed through the fastANI software v1.33 . Here, genome sequences retrieved from NCBI for each of the nine subject genomes were used as input into fastANI with the assemblies for Coll940 and PMBrms-1 ( C. noveboracense ) used as the query list and the other seven assemblies as the reference list. A cluster map of the fastANI results was generated using the ANIclustermap script . The second approach was to use the OrthoFinder ( https://github.com/davidemms/OrthoFinder ) to construct a phylogenetic tree based on conserved gene sequence similarity. The protein FASTAs for each of the nine assemblies were used as input with the -m MSA option enabled for the generation of maximum likelihood trees from multiple sequence alignments (MSAs). The resulting newick tree was visualized using the ETE 3 toolkit’s newick tree viewer (Centre for Genomic Regulation, Barcelona, Spain) . Lastly, a tree was generated based on highly conserved single-copy orthologous genes. BUSCO (EMBL Bioinformatics Core Facility, Heidelberg, Germany) was implemented to identify single-copy orthologs in each genome. BUSCO was executed in the -m genome mode with the genome sequence FASTA for the ten assemblies. The Glomerellales ODB10 database of BUSCO was implemented as the lineage parameter option. The BUSCO result plot was generated using the generate_plot.py script included with BUSCO. Phylogenetic relationships between the single-copy ortholog genes present in each assembly were investigated using the BUSCO Phylogenomics python pipelines . Within this pipeline, MSA was conducted using MUSCLE (Robert C. Edgar, Harvard University, Cambridge) and alignment trimming was performed by trimAl (Salvador Capella-Gutiérrez, Toni Gabaldón, and José M. Sánchez-Pulido, Barcelona, Spain) . A consensus phylogenetic tree was determined using IQ-tree (Alexandros Stamatakis, Heidelberg Institute for Theoretical Studies and Karlsruhe Institute of Technology, Heidelberg, Germany) on the resulting supermatrix generated by the BUSCO_phylogenomics.py pipeline with the -m MFP option enabled and -B for bootstrapping set to 1000. The resulting newick tree was visualized using the ETE 3 toolkit’s newick tree viewer . To complement the phylogenetic analyses conducted using the nine subject genomes, we reperformed the BUSCO phylogeny and fastANI analyses using an additional nine genomes ( Supplemental Table S1 ). These nine genomes were previously identified as belonging to the acutatum phylogenetic subclade and offered an opportunity to assess broader species-level relationships with our subject genomes . One species listed in Liu et al. in that subclade, C. godetiae , was not included due to a BUSCO complete score of <90%, suggesting an incomplete genome assembly. Furthermore, AFKH109 has a similarly low BUSCO score and thus was removed from the BUSCO phylogenetic analysis of the 18 assemblies. However, both assemblies were included in the broader fastANI analysis.
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
39057378_p7
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39057378
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sec[1]/sec[2]/p[0]
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2.3. Mining for CAZymes, Secondary Metabolic Gene Clusters, and Effectors
| 4.246094 |
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Fungal versions of the antibiotics and Secondary Metabolites Analysis SHell (antiSMASH) 6.0 pipeline https://fungismash.secondarymetabolites.org/ (University of Göttingen, Germany) with “relaxed” detection strictness was used for mining our Colletotrichum genomes for secondary/specialized metabolite (SM) biosynthetic gene clusters. We used the dbCAN2 meta server for analyzing and predicting six major classes of carbohydrate-active enzymes in our newly sequenced genomes of Colletotrichum species (University of Alberta, Edmonton, Canada). HMMER (Sean Eddy, Howard Hughes Medical Institute, Janelia Research Campus, Ashburn, VA, USA) with E-Value < 1e-15, coverage > 0.35, eCAMI (University of Göttingen, Germany) with important_k_mer_number >= 5, k_mer size = 8, and DIAMOND (Benjamin Buchfink and collaborators, Max Planck Institute for Developmental Biology, Tübingen, Germany) with a cut-off E-value of <1e-102 for CAzymes prediction were used as the default thresholds . Candidates found by at least two predictive tools were retrieved. For each CAZy class, the number of enzyme modules and the families they belong to are reported. Finally, we applied EffectorP 3.0 (University of Copenhagen, Denmark) (fungal mode parameter) and SignalP 6.0 mode Eukarya (Technical University of Denmark, Kongens Lyngby, Denmark) with a significance threshold = 0.7 to predict known apoplastic and cytoplasmic effectors in 9 isolates of Colletotrichum species .
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
39057378_p8
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39057378
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|
3.1. Colletotrichum spp. Genomes
| 4.148438 |
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As described in Khodadadi et al. , a total of 16,621 protein coding genes were predicted in each Colletotrichum species assembly. While the genome assembly metrics varied for each of the Colletotrichum species sequenced, all genomes had over 95% coverage of complete BUSCOs. The GC content of the genomes sequenced ranged from 49.33 to 53.94%, with C. fioriniae and C. chrysophilum having the upper and lower ranges, respectively. The estimated genome sizes for the sequenced isolates ranged from 49.33 to 59.08 Mb. The estimated sequencing coverage ranged from 46× to 103×, with the lowest and highest coverages obtained for the C. nupharicola and C. fioriniae isolates, respectively ( Table 1 ).
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
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39057378_p9
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39057378
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3.2. Phylogenomic Analysis
| 4.371094 |
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Study
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A novel Colletotrichum species, designated C. noveboracense , was identified as the causal agent of apple bitter rot disease in New York and Pennsylvania . Several isolates (PMBrms-1 and AFKH109) from these affected regions, along with a single endophytic isolate from Juglans nigra in Oklahoma (Coll940) , formed a highly supported distinct clade based on multi-locus sequence analysis (MLSA). Initial phylogenetic analyses using Bayesian inference with a three-gene approach ( ITS , TUB2 , and ApMat ) revealed a well-supported clade (BI PP = 1.0) containing the isolates subsequently identified as C. noveboracense . Further Bayesian analysis employing seven loci ( ACT , TUB2 , CAL , GAPDH , GS , ITS , and ApMat ) and other locus combinations consistently placed C. noveboracense as a sister taxon to C. nupharicola (PP = 0.95) within the Colletotrichum gloeosporioides species complex (CGSC) . However, C. nupharicola is readily distinguishable within CGSC based on morphological characteristics. This host-specific species exhibits significantly slower growth on potato dextrose agar (PDA) and possesses conidia with both length and width exceeding those of other species in CGSC .
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
39057378_p10
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39057378
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sec[2]/sec[1]/p[1]
|
3.2. Phylogenomic Analysis
| 4.371094 |
biomedical
|
Study
|
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The current study was expanded to encompass a larger dataset, incorporate the informative APN2 locus known to enhance resolution within CGSC, and evaluate the novel clade using the stringent Genealogical Concordance Phylogenetic Species Recognition (GCPSR) criteria. This comprehensive approach confirmed the isolates as a strongly supported clade, distinct from all other taxa within CGSC . These substantial morphological disparities between C. noveboracense and C. nupharicola warranted a more comprehensive analysis. To understand the evolutionary relationships among species, a genome-based method was utilized. The alignment-free method via fastANI software was used and is typically implemented in the determination of species for prokaryotes. However, previous works using small Eukaryote (fungal) genomes and genes were successful . When employed for our dataset, ANI identified a group with a similar sequence identity that contained ACFK5 and ACFK16 belonging to CASC, whereas the remaining eight assemblies, belonging to CGSC, grouped together with ANI values of 96.5–99.9% similarity . For our three assemblies in question, AFKH109, Coll940, and PMBrms-1 grouped together with CBS470 into subclades branching from Coll922. The same result was observed when constructing a tree based on gene trees of orthologs shared between assemblies . This result was determined using ortholog trees based on protein similarities via the OrthoFinder software . Another independent approach used BUSCO gene sets from the Glomerellales ODB10 database . Over 6618 single-copy BUSCO sequences were identified in each genome, except for AFKH109 which had 5830 . This analysis suggests that the AFKH109 assembly is of lower quality. We used the BUSCO_phylogenomics pipeline to construct phylogenetics trees using these highly conserved gene sets . The pipeline used 6714 BUSCO sequences that were complete, single-copy, and were found in at least four assemblies. The resulting consensus tree was highly similar to the prior two approaches (phylogenetic and sequence similarity) but separated ACFK5 and ACFK16 from a single clade. Again, AFKH109, Coll940, and PMBrms-1 grouped together with CBS470 into a subclade with the same subtending branch for Coll922. Consistently across all three methods, CBS470 and Coll940 were most closely related. AFKH109 and PMBrms-1 either grouped together or were subtending the CBS470 and Coll940 clades in that order, respectively. Based on our genome analysis, CBS470 ( C. nupharicola ) exhibits a closer genetic similarity to three C. noveboracense isolates (Coll940, PMBrms-1, and AFKH109) compared to other isolates. This suggests a potentially closer evolutionary relationship between C. nupharicola and these specific C. noveboracense isolates. As a complementary approach, we also reconstructed the BUSCO phylogeny and fastANI analysis using previously identified acutatum subclade members . These broader analyses support our previously stated conclusions as the phylogenetic structure was preserved between our nine subject genomes and the broader Colletotrichum species members . Moreover, the average nucleotide identity preserved these relations in this broader comparison . This finding supports previous phylogenetic efforts using eight informative loci . The inferred phylogeny suggests a significant evolutionary distance between CASC and CGSC.
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
39057378_p11
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3.3. Bioinformatic Prediction of Effectors
| 4.136719 |
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Study
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Comparative genomics approaches lend themselves to the discovery of an array of predicted effectors, CAZymes, and SM gene clusters, which may not have otherwise been elucidated by classical genetic, pathological, and/or biochemical approaches. These studies often lead to the elucidation of both known and novel loci that can then be targeted for functional studies. Hence, we sought to utilize the Colletotrichum spp. genomes that we previously elucidated , with tools to discover a holistic view of factors that are routinely associated with fungal virulence, pathogenicity, and small molecule/toxin potential.
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
39057378_p12
|
39057378
|
sec[2]/sec[2]/p[1]
|
3.3. Bioinformatic Prediction of Effectors
| 4.402344 |
biomedical
|
Study
|
[
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Effector prediction was conducted using EffectorP 3.0 for nine different isolates consisting of four different Colletotrichum spp. . This tool has been implemented to mine many different fungal genomes and has been recently used to explore Colletotrichum truncatum . In our study, we found 390 to 581 predicted effectors with the lowest number in C. fioriniae and the highest number in C. chrysophilum AFK26 . Predicted apoplastic effectors ranged from 241 to 321 and cytoplasmically targeted effectors ranged from 77 to 120. Some effectors were categorized as both having the potential to be localized in either the cytoplasm or apoplasm. Rao and Nandeneni found between 200 and 400 predicted effectors in four different Colletotrichum spp., of which C. fioriniae showed around 300 which is similar to our findings. Effector prediction using EffectorP 3.0 in Lu et al.’s study identified a large number of candidate effectors in different Colletotrichum species (288 to 608 per genome). This analysis suggests significant variation in predicted effector numbers between Colletotrichum species. Further clustering analysis within the genus revealed that about 20% of these candidate effectors are core effectors, present in all Colletotrichum species examined. Another 70% represent conserved effectors with orthologs (similar genes) identified in some Colletotrichum species. Interestingly, each species also harbors a unique set of species-specific effectors, ranging from 4.1% to 15.6% of their total predicted effectors. These findings suggested a potential link between the conservation patterns of candidate effectors and the host range and virulence of Colletotrichum pathogens . Moreover, de Queiroz et al. conducted a comprehensive analysis of candidate effector proteins in two physiological races (83.501 and 89 A2 2-3) of the pathogen Colletotrichum lindemuthianum and found a total of 353 and 349 effectors, respectively. Interestingly, over 63% of these effectors share common features: they are rich in cysteine, contain repetitive amino acid sequences, and/or possess nuclear localization signals . Additionally, analysis revealed several conserved protein domains shared among the C. lindemuthianum effector candidates. Later, they extended the analysis to nine other Colletotrichum species. The number of predicted effectors varied across these species, ranging from 247 in Colletotrichum graminicola to 446 in Colletotrichum orbiculare . Notably, all analyzed species shared twelve conserved protein domains within their predicted effector candidates . Discrepancies in numbers determined in this study, with isolates from other studies, and between isolates of a similar species could be due to an array of factors that relate to genome coverage, annotation quality, and differences in bioinformatic programs utilized to analyze various genomes. It is of interest that overall, the total numbers of predicted effectors can be generalized as three Colletotrichum spp. ( C. nupharicola, C. noveboracense, and C. chrysophilum ) had higher numbers than C. fioriniae. Notwithstanding, the biological ramifications of these observations may not be realized at this time and require systematic functional study in the fungus via gene deletion, RNA silencing, and/or overexpression in planta to ascertain their role in fungal–host interactions.
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
39057378_p13
|
39057378
|
sec[2]/sec[3]/p[0]
|
3.4. Carbohydrate Active Enzyme (CAZyme) Classes
| 4.476563 |
biomedical
|
Study
|
[
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CAZymes are enzymes involved in complex carbohydrate synthesis or breakdown . These enzymes are enriched in many fungal taxa including those whose members engage in plant associations (e.g., phytopathogens). The study and identification of CAZymes has been of great interest due to their biotechnological potential and the possibility of using their profiles as clues to understand fungal lifestyles and their evolution which includes plant pathogenic potential . This study showed the comparison of carbohydrate-active enzyme (AA, CE, GH, and PL) profiles between species, providing insights into their diverse carbohydrate degradation capabilities . A previous study showed that plant pathogenic fungi contain the largest number of CAZymes . In this study, the total number of predicted CAzyme genes for the sequenced isolates ranged from 593 to 831. This is a high number when compared to other species in the genus like C. australisiense with 541 genes and C. siamense with 507 genes but similar when compared to Colletotrichum camelliae with 836 genes . All the assessed isolates show a similar trend in the abundance of each identified CAZyme category . The identified categories from least to most abundant CAZymes in all the analyzed species are as follows: (1) carbohydrate-binding molecules (CBMs), (2) polysaccharide lyases (PLs), (3) carbohydrate esterases (CEs), (4) glycosyl transferases (GTs), (5) auxiliary activities (AAs), and (6) glycoside hydrolases (GH)s. While glycoside hydrolases are the CAZyme category with the most predicted genes for the assessed Colletotrichum spp. in this study, pectin, a glycoside hydrolase substrate, comprises from 5.6 to 10% of the apple cell wall components and hemicellulose 2 to 4.1% . This bioinformatic prediction sheds light on the metabolic potential of these species, on understanding the basic biology of virulence, and the potential industrial application for the degradation of complex carbohydrates. In addition, gene expression analyses, for individual loci, specific CAZyme classes, and a systems-based RNAseq/functional approach of these genes in Colletotrichum spp. will leverage our genomics findings to further understand how these loci enhance host specificity and/or underpin aggressiveness in host–parasite interactions.
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999994 |
39057378_p14
|
39057378
|
sec[2]/sec[4]/p[0]
|
3.5. Biosynthetic Gene Cluster Distribution
| 4.476563 |
biomedical
|
Study
|
[
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Fungi produce unique molecules (natural products/secondary metabolites) through specialized biosynthetic gene clusters (BGCs). Unlike those for growth, these BGCs (2–20+ genes) create a chemical arsenal (toxins and antibiotics) vital for fungal interactions with other microbes and their environment . In this study, we used antiSMASH to predict the nine Colletotrichum spp. SM gene cluster repertoire. We found that the number of predicted SM gene clusters ranged from 46 in C. floriniae ACFK5 to 75 in C. noveboracense Coll940 . These numbers of clusters are typical for the genus as previous studies have reported 71 for C. camelliae , 85 SM gene clusters for Colletotricum siamense, and 55 for Colletotricum australisinense . In addition, all of the sequenced strains contained similar profiles in the type of BGC that were predicted. The assessed species contain the greatest proportion of type-1 polyketide synthases, i.e., T1PKS-type backbone genes in their genomes, followed by terpene backbone genes, and finally nonribosomal peptide-synthetase (NRPS) or NRPS-like genes. These findings are congruent with what Liu et al. found when comparing C. siamense and C. australisinense . The cercosporin gene cluster is present for all the assessed samples but its homology varies between the sequenced isolates to the representative cluster of Cercospora betiicola . The role of polyketide synthases (PKSs) in promoting host penetration by Colletotrichum species has been recognized for a considerable time . While all the SM gene cluster predictions are valuable, most of the detected clusters have not been characterized, representing a great opportunity to further mine, characterize, and understand their ecological role in Colletotrichum spp. Secondary metabolites (SMs) produced by fungal phytopathogens exhibited a robust correlation with both their pathogenicity and host range . Application of these phytotoxic SMs to host leaves induced disease symptoms mirroring those observed in anthracnose caused by Colletotrichum species. This finding underscores the critical role these metabolites play in the pathogenesis and infection mechanisms employed by these fungi .
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
39057378_p15
|
39057378
|
sec[2]/sec[4]/p[1]
|
3.5. Biosynthetic Gene Cluster Distribution
| 4.527344 |
biomedical
|
Study
|
[
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Of the reported BGCs in this study, alternapyrone is conserved throughout all the assessed Colletotrichum species. Alternapyrone is a polyketide predicted to be synthesized by five gene products named altA to altD . For this cluster, the backbone gene encodes for an iterative type I polyketide synthase (pksN), one gene encodes for a FAD/FMN-dependent oxygenase/oxidase, and three genes are annotated as a cytochrome P-450. The alternapyrone BGC was first described in Alternaria solani , the early blight disease pathogen of tomato and potato, and recently has been reported to be produced by Parastagonospora nodorum , a fungal wheat pathogen . Limited information can be found on the compound bioactivity, but data suggest that alternapyrone displays cytotoxic activity and some derivatives inhibit wheat seed germination . This finding along the complete BGC could further be explored to unravel individual species secondary metabolite production and the importance of their role in ecological and interspecies interactions. This research holds significant promise for the improved management of apple bitter rot. Our work provides a foundation for developing targeted control strategies by identifying candidate genes crucial for Colletotrichum spp. virulence and pathogenicity. These genes, including effectors, CAZyme genes, and secondary metabolite gene clusters, represent potential targets for novel control mechanisms. RNA interference or gene editing techniques could be employed to silence or disrupt these genes, hindering fungal function. Furthermore, by comparing the genomes of various Colletotrichum species that infect apples, this study sheds light on the genetic basis of host specificity. This newfound knowledge can be harnessed to develop apple cultivars exhibiting enhanced resistance to bitter rot disease.
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
39057378_p16
|
39057378
|
sec[3]/p[0]
|
4. Conclusions
| 4.464844 |
biomedical
|
Study
|
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This study utilized whole genome sequencing of nine Colletotrichum isolates encompassing four species ( C. fioriniae , C. chrysophilum , C. noveboracense, and C. nupharicola ) to gain insights into their pathogenicity, host specificity, and evolutionary relationships. Phylogenomic analyses confirmed the distinctiveness of C. noveboracense as a novel causal agent of apple bitter rot disease. Notably, C. noveboracense and C. nupharicola displayed a closer evolutionary relationship compared to other species in the CGSC and the CASC. Comparative genomics revealed a vast repertoire of potential virulence factors, including predicted effector proteins, carbohydrate-active enzymes (CAZymes), and secondary metabolite (SM) gene clusters. To solidify our findings and translate them into actionable strategies, future research should pursue several avenues. Functional validation of the identified candidate genes is crucial. Techniques like targeted gene deletion, overexpression, and RNA silencing can elucidate which genes are essential for Colletotrichum spp. virulence and pathogenicity. Additionally, omics approaches such as transcriptomics, proteomics, and metabolomics offer a deeper understanding of gene expression during infection and the production of metabolites by Colletotrichum spp. These comprehensive data can reveal further targets for control strategies. Finally, the wealth of genetic information gleaned from this study holds immense potential for developing more specific and accurate molecular diagnostic tools . These tools can effectively identify Colletotrichum spp. responsible for apple bitter rot, facilitating earlier intervention and improved disease management. Our findings provide a valuable resource for further functional studies aimed at elucidating the specific roles of these virulence factors in Colletotrichum spp. Additionally, these data lay the groundwork for the development of novel and targeted disease management strategies for crops susceptible to Colletotrichum infection.
|
[
"Fatemeh Khodadadi",
"Dianiris Luciano-Rosario",
"Christopher Gottschalk",
"Wayne M. Jurick",
"Srđan G. Aćimović"
] |
https://doi.org/10.3390/jof10070493
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
PMC11278320_p0
|
PMC11278320
|
sec[0]/p[0]
|
Introduction
| 4.484375 |
biomedical
|
Study
|
[
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Currently, a series of vaccine candidates against emerging viral pandemic COVID-19 are in development worldwide, and several of these candidate vaccines are either authorized for emergency use or approved for use in humans. 1 , 2 , 3 , 4 Among these, nucleic acid (NA)-based vaccines contain a section of genetic material derived from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that prompts a person’s own cells to generate parts of the targeted virions, and as a result, generate immunity. 1 , 2 Despite mRNA having received major interest and entering a new era in vaccinology, RNA molecules exhibit a low stability and must be protected from enzymatic digestion to achieve successful targeted delivery. 4 Recently, lipid nanoparticle (LNP) technology was successfully advanced and became more sophisticated, highlighting the gap between the advantages of mRNA carrier platforms and the availability of automated microfluidic manufacturing processes to scale up their production. 4 , 5 , 6 However, mRNA technology requires ultracold transportation, which limits the feasibility for communities lacking cold chain facilities. 4 In contrast to mRNA vaccines, DNA molecules possess high stability and do not require ultracold storage and distribution 7 ; nevertheless, the assistance of either a viral vector to deliver genetic materials to the body’s cells or an electroporation device into the muscle or skin of the host is necessary to achieve efficacious DNA vaccine delivery and protective immunity against SARS-CoV-2 infection. 8 , 9 , 10 Such systems face the obstacle that the use of viral vectors generally elicits limited transgene capacity and broad preexisting immunity in humans. However, the electroporation device often results in substantial cell death caused by high-voltage pulses. Therefore, it should be very interesting to extend the range of LNP systems to DNA vaccine delivery and immunogenicity.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278320_p1
|
PMC11278320
|
sec[0]/p[1]
|
Introduction
| 4.28125 |
biomedical
|
Study
|
[
0.99951171875,
0.0003478527069091797,
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[
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] |
The LNP commonly contains four hybrid lipid components, and each component makes a contribution to structural stability and functional activity. 4 , 11 The components include cholesterol (for cell transfection), phospholipid (a helper lipid for particle structure), ionizable cationic lipid (for cellular uptake and endosomal escape of NA), and PEGylated lipid (for LNP stability/circulation). Cholesterol and phospholipids are building block components embedded in the lipid bilayer of the cell membrane. 4 The structural function of ionizable cationic lipids is the key factor in the expression kinetics and endosomal escape activities of NA antigen, which are associated with vaccine-induced adaptive immune responses. 11 Alternatively, PEGylated lipids act as surface engineering agents (surfactants) that stand out from the surface of LNPs. 4 These lipids reduce surface free energy between the lipophilic core and aqueous phase as well as nonspecific binding to proteins by steric repulsion. The obtained LNPs entrap and protect the designed DNA from enzymatic degradation during systemic circulation and facilitate endocytosis and endosomal escape. 4 For the first time, we repurposed an amphiphilic bioresorbable copolymer (ABC) comprising hydrophilic poly(ethylene glycol) (PEG) and lipophilic poly(lactic acid) (PLA) as an alternative to PEGylated lipids together with cholesterol, a helper lipid (phospholipid), and an ionizable cationic lipid to prepare hybrid LNPs. The immunogenicity of the DNA-LNP vaccines was determined in hamsters for the induction of protective immunity against heterologous virus challenge or as a heterologous boosting tool against SARS-CoV-2 variants.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278320_p2
|
PMC11278320
|
sec[1]/sec[0]/p[0]
|
Self-assembled DNA-LNP vaccine
| 4.527344 |
biomedical
|
Study
|
[
0.99951171875,
0.0003981590270996094,
0.00022077560424804688
] |
[
0.99658203125,
0.0007066726684570312,
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0.00016808509826660156
] |
The formation of the DNA-LNP vaccine involves a self-assembling process between two fluid streams: an acidic buffer solution of DNA encoding protein antigen and a water-miscible solution comprising the lipid mixture. 5 Automated microfluidic systems have offered a robust tool for homogenizing processes and controlling the synthesis of nanoparticles in a versatile and reproducible manner . This apparatus, equipped with a series of toroidal channel mixers, induces chaotic advection, so the lipid components (dissolved in ethanol) and genetic molecules (dissolved in acidic buffer solution) can diffuse and self-assemble between ethanol-water interfaces. 5 , 6 Typically, DNA-LNP assemblies were uniform at nanoscale as measured by dynamic light scattering (DLS) technologies. The prepared DNA-LNPs showed a spherical shape with nanostructures under transmission electron microscopy (TEM) images. Notably, the process is spontaneous—in other words, no extra energy is required to produce the nanoparticles. An efficient workflow can be achieved by steady pressure-driven flow streams such that the process was operated by controlling the total flow rate and the mixing ratio under laminar flow conditions. Figure 1 Self-assembled DNA-LNP vaccine Typically, 2 miscible fluids, an aqueous solution of DNA and an organic solution comprising mixtures of lipids dissolved in ethanol, were separately pumped into the Y-shaped inlet channel of the microfluidic chip. A series of toroidal channel mixers induces chaotic advection, so the lipid components and DNA molecules can diffuse and self-assemble between ethanol-water interfaces under laminar flow conditions. Generally, cholesterol and phospholipids play a structural role in LNPs, while ionizable cationic lipid (SM-102) enables the capture of DNA in aqueous solutions and the formation of DNA-lipid complexes. In addition, the amphiphilic bioresorbable copolymer PLA-PEG serves as a surface engineering agent (surfactant) to stabilize the lipid core and aqueous phase, yielding DNA-LNPs with uniform structures at nanoscale size under DLS and TEM images.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
PMC11278320_p3
|
PMC11278320
|
sec[1]/sec[0]/p[1]
|
Self-assembled DNA-LNP vaccine
| 4.371094 |
biomedical
|
Study
|
[
0.99951171875,
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[
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Formulation efficiency and particle size are important physicochemical characteristics of DNA-LNPs, and determining these characteristics is fundamentally important for rational vaccine design. The work started by studying the effect of the content of PEGylated lipid on the prospective efficiency of LNPs . After passing through the inlet channel of the microfluidic chip, stable nanoparticles of uniform distribution (namely DNA-LNP-FM) can be prepared with a predetermined cholesterol:DSPC (1,2-distearoyl- sn -glycero-3-phosphocholine):SM-102 (8-[(2-hydroxyethyl)[6-oxo-6-(undecyloxy)hexyl]amino]-octanoic acid, 1-octylnonyl ester):DMG-PEG molar ratio of 38.5:10:50:1.5. By removing the DMG-PEG, a non-degradable lipid, the DNA-LNP (DNA-LNP-F1) conserved the encapsulation efficiency (EE) of LNPs, while the DNA recovery rate (RR) was lower (∼10%); in addition, the DNA-LNPs tended to aggregate with a broader particle size scale, polydispersity, and zeta potential. Supplementation with the appropriate ingredients of PLA-PEG helps DNA-LNPs stabilize and shrink the particles as more building block components are embedded in the LNP structure. DNA-LNP-F2 and DNA-LNP-F3 represent DNA-LNPs with 1.5% PLA-PEG and 3.0% PLA-PEG, respectively. Afterward, we investigated the biocompatibility of DNA-LNPs; this can be conducted via the treatment of HEK293 cells with DNA-LNPs, followed by assessment of cell viability. Within the 72-h incubation period, the treatment of DNA-LNP-FM induced a higher percentage of cell death in HEK293 cells compared to medium alone at each time point. Interestingly, the potency of cell death was reduced when the surface-active agents within the LNP was replaced as PLA-PEG. Collectively, we identified the cell death effects of DNA-LNPs on cells and suggested that the selection of surface-active agents may play a crucial role in DNA-LNP-induced cell death. A detailed description of the NA fundamental characterization and transfection efficiency in the cell culture study and in vivo distribution assessment of PLA-PEG-surface engineered DNA-LNPs is summarized in Figures S1–S5 , using plasmid DNA encoding EGFP and/or green click beetle luciferase protein (CBGr99) as a payload reporter. In Figure S2 , we observed that there was no or low fluorescence signal in the free DNA group, indicating low transfection. Notably, low transfection efficiency can be overcome after encapsulation into LNPs or with conventional Lipofectamine 2000 transfection reagent. However, Lipofectamine treatment induced a higher percentage of cell death in HEK293 cells compared to the groups of DNA-LNP and free DNA. Similar to the experiments with GFP-encoding DNA, quantification of mRNA transcription and protein expression was investigated in HEK293 cells during 72-h post-transfection with luciferase-encoding DNA. As shown in Figure S3 A, there was no or low luciferase mRNA transcription in the free DNA group. In contrast, there was a dramatic increase in mRNA transcription after encapsulation into LNPs at each time point (24, 48, and 72 h). Interestingly, higher luciferase activity was observed for DNA-LNP formulation compared to the conventional transfection reagent. In vivo distribution assessment in mice with plasmid DNA encoding luciferase protein showed that the luminescence signals were durable and long lasting for at least 1 month . Notably, we found that PEGylated lipid contents did not play a critical role in the transfection of DNA-LNP formulation in the cell culture system , as they act as surface modification agents (surfactants) rather than serving a structural function in the expression kinetics and endosomal escape activities of the DNA molecule. It is interesting to note that upon storage at 37°C, the diameter of PLA-PEG-surface engineered DNA-LNP formulations remained unchanged, and the luminescence signal produced by the cells decreased the least. However, the diameters of the DNA-LNP formulations increased upon storage at 4°C, and the luminescent signal produced by cells decreased the most . Figure 2 Impact of PEGylated lipid on the physicochemical characteristics and biocompatibility of DNA-LNPs TSomi DNA-LNPs were characterized in terms of EE%, RR%, z-average particle size (nm), PDI, and zeta potential (mV), respectively. The biocompatibility of DNA-LNPs was conducted by the treatment of HEK293 cells with DNA-LNPs, followed by the assessment of cell viability. DNA-LNP-F1, -F2, -F3, and -FM represent DNA-LNPs without PEGylated lipid or with 1.5% PLA-PEG, 3.0% PLA-PEG, or 1.5% DMG-PEG, respectively. The cell relative viability was calculated as the percentage viability compared to medium control group at each time point. Data are represented as the mean ± SD of 3 replicates of each sample. The results are representative of 3 independent experiments.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278320_p4
|
PMC11278320
|
sec[1]/sec[0]/p[2]
|
Self-assembled DNA-LNP vaccine
| 4.1875 |
biomedical
|
Study
|
[
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[
0.99951171875,
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] |
Other parameters, such as the contents of the DNA, the helper lipid (DSPC), the ionizable cationic lipid (SM-102), and the ratio of PEGylated lipid to helper lipid (P/H), affect the EE and structural stability of the LNPs . As can be observed, in increasing the DNA content in LNPs, the DNA-LNPs tended to aggregate with a broader particle size distribution and lower DNA EE and RR. When the DNA-LNPs are prepared with different DSPC concentrations of fixed cholesterol/SM-102/PLA-PEG ratios, higher EE and RR are obtained only when the DSPC molar ratio is higher than 10. The DNA-LNPs present a smaller particle size and zeta potential when DSPC increases, as more building block components are embedded in the LNP structure. Increasing the DNA content could enlarge the particles, while increasing the DSPC content could shrink the particles. By adjusting SM-102 content could effectively drive DNA recovery and lead to property changes in LNPs. Last but not least, although PLA-PEG (PEGylated component) and DSPC (helper component) play a crucial role in the fundamental fingerprints of DNA-LNPs, we did not find the impact of the P/H ratio on the EE and structural stability of the LNPs to be within the designated ranges. Figure 3 Parameter tuning on the physicochemical characteristics of DNA-LNPs (A) Library of LNP chemical constituents. (B) TSomi DNA-LNPs were characterized in terms of EE%, RR%, z-average particle size (nm), PDI, and zeta potential (mV), respectively. The results are represented as the mean ± SD of 3 replicates of each sample. The standard molar ratio of lipids was cholesterol:DSPC:SM-102:PLA-PEG of 38.5:10:50:1.5, with a fixed cholesterol content of 516 μg.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999999 |
PMC11278320_p5
|
PMC11278320
|
sec[1]/sec[1]/p[0]
|
COVID-19 DNA vaccine efficacy
| 4.203125 |
biomedical
|
Study
|
[
0.99951171875,
0.00031876564025878906,
0.00020575523376464844
] |
[
0.99951171875,
0.00018405914306640625,
0.00046896934509277344,
0.00007510185241699219
] |
COVID-19 vaccines based on the initial global spread of the first SARS-CoV-2 genomic sequence, Wuhan-1, are lacking in eliciting cross-protective immunity against the emergence of variants. 1 , 12 Since new pandemic waves of COVID-19 are exacerbated by emerging SARS-CoV-2 variants, a feasible strategy for developing broadly protective immunity is receiving an annual vaccination or boosting with occasional updates. 1 , 13 , 14 We used plasmid DNA encoding the SARS-CoV-2 Omicron variant trimeric-spike (TSomi) gene as a model antigen to elucidate the role of DNA-LNP in vaccine immunogenicity. Previously, we investigated the use of LNP as a promising platform for new-generation mRNA/DNA vaccine technology. 14 , 15 The results demonstrated that LNP-encapsulated DNA could elicit higher humoral responses than those by a 10-times dosage of free DNA. 15 Here, we further investigated the immunological aspects of DNA-LNP surface engineered with an ABC. This investigation was conducted by studying the immunogenicity and efficacy profiles through intramuscular injection of DNA-LNP-F2 in hamsters, which is a useful animal model for evaluating COVID-19 vaccines. 3 DNA-LNP-F2 was chosen as it is a compromise between transfection efficacy and biocompatibility. It should be noted that LNP-F2 possesses the same molar ratio of lipid mixture as an approved LNP-based mRNA vaccine (LNP-FM in the present study), with the modification that the surface-active agent (DMG-PEG) has been replaced with an equivalent amount of PLA-PEG.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278320_p6
|
PMC11278320
|
sec[1]/sec[1]/p[1]
|
COVID-19 DNA vaccine efficacy
| 4.289063 |
biomedical
|
Study
|
[
0.99951171875,
0.0004906654357910156,
0.00019788742065429688
] |
[
0.9990234375,
0.00025343894958496094,
0.0006346702575683594,
0.00011092424392700195
] |
We conducted animal experiments on the efficiency of DNA-LNP vaccines against SARS-CoV-2 and its variants. Hamsters were administered prime-boost doses of 10 and 30 μg TSomi DNA-LNP, respectively. Tris buffer was used as the negative control. Serum samples were collected at predetermined time points for the evaluation of specific immunoglobulin G (IgG) antibody responses and protective immunity against both homologous and heterologous virus strains of SARS-CoV-2. Following the sequential vaccination regimen represented in Figure 4 A, we found that the sera obtained from TSomi DNA-LNP-vaccinated hamsters could generate effective Omicron-specific neutralizing antibodies and IgG antibodies , indicating the merit of the vaccination strategy in COVID-19 protection. Interestingly, the same serum samples could neutralize Wuhan virus infection; even the neutralizing antibody titers were rather reduced . These data are similar to the previous reports in which sera obtained from the hosts received Omicron-specific mRNA/DNA or protein vaccines possessed low cross-neutralization antibody titers against the Wuhan virus. 14 , 15 , 16 We subsequently investigated whether TSomi DNA-LNP could induce protective immunity in hamsters when challenged with the Wuhan strain virus. Notably, Wuhan strain virus infection induced severe disease in human and hamster animal models. 14 , 15 It is therefore interesting to investigate whether the TSomi DNA vaccine could induce protective immunity against Wuhan strain virus challenge. After infection with Wuhan virus, we found that the hamsters vaccinated with TSomi DNA-LNP did not show a decrease in body weight during 6 days of monitoring at either 10 or 30 μg dosages; however, the hamsters that received Tris buffer alone showed a 10% reduction in body weight . As depicted in Figure 4 D, the viral load in the lung showed a mild reduction on day 3 in the 10-μg DNA-LNP group compared to the buffer control, and this reduction continued to progress through day 6. However, it is important that the loaded virus was not fully eradicated from the host at this dosage. It is worth noting that increasing the dosage to 30 μg DNA-LNP effectively generated valuable viral load attenuation characteristics. The pathogenesis in the lung at day 6 was analyzed, and based on the data, the buffer control group showed severe inflammation, while very low inflammation was found in the TSomi DNA-LNP vaccination groups . Taken together, these data revealed that the TSomi DNA-LNP vaccine is capable of inducing protective immunity against a Wuhan virus challenge. Figure 4 Vaccine efficacy of LNP-formulated DNA sequences encoding SARS-CoV-2 Omicron variant spike genes against the Wuhan original strain (A) Schematic diagram of the vaccination regimen. Hamsters ( n = 8) were injected with TSomi DNA-LNP through the i.m. route on day 0. At week 2, all hamsters were boosted through i.m. administration of the same vaccine formulations. (B) The VN antibodies in serum samples at week 6 are expressed as the individual values with the geometric mean titer (GMT). The Omicron TS-specific IgG titers are shown in Figure S6 . In (C)–(F), the hamsters were challenged with Wuhan SARS-CoV-2 at week 6 post-vaccination. (C) Body weight change (%) after virus challenge. (D) Viral load in the lungs of infected hamsters at days 3 and 6 post-infection (d.p.i.). (E) Histopathologic examination of tissue sections (4 μm) from the lungs of infected hamsters was performed by staining with H&E and examined using a microscope (original magnifications, ×40 and ×400). (F) Pathologic severity was scored, and the p value was calculated by 1-way ANOVA with Tukey’s multiple comparison test. ∗ p < 0.05.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278320_p7
|
PMC11278320
|
sec[1]/sec[1]/p[2]
|
COVID-19 DNA vaccine efficacy
| 4.105469 |
biomedical
|
Study
|
[
0.99951171875,
0.00037026405334472656,
0.0002620220184326172
] |
[
0.99951171875,
0.00019478797912597656,
0.00024008750915527344,
0.0000603795051574707
] |
We next aimed to evaluate whether Omicron-specific antibodies can be driven by a hybrid-type vaccine booster. As illustrated in Figure 5 A, hamsters were first administered two doses of Wuhan mRNA-LNP vaccine and generated Wuhan-specific IgG antibodies, as shown in Figure S7 . At week 26, the hamsters were boosted with an Omicron-specific DNA-LNP vaccine. As shown in Figure 5 B, boosting with the TSomi DNA-LNP vaccine resulted in a robust neutralizing titer against the Omicron variant, with all five hamsters exhibiting titers exceeding 80 (seroconversion). This finding implied that a boosting dose of TSomi DNA-LNP could offer protective efficacy against virus challenge. Figure 5 Boost effect of TSomi DNA-LNP on Omicron-specific VN antibodies in hamsters that were previously injected with 2 doses of Wuhan mRNA-LNP (A) Schematic diagram of the vaccination regimen. Hamsters ( n = 5) were injected with 10 μg Wuhan mRNA-LNP through the i.m. route on days 0 and 21. The Wuhan TS-specific IgG titers are shown in Figure S7 . At week 26, the hamsters were boosted through i.m. administration of 10 μg TSomi DNA-LNP. (B) The Omicron-specific VN antibodies in serum samples prior to booster vaccination and 2 weeks post-booster are expressed as the individual values with the GMT. The dotted horizontal line represents seroconversion (SCR). ∗ p < 0.05.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278320_p8
|
PMC11278320
|
sec[2]/p[0]
|
Discussion
| 4.371094 |
biomedical
|
Study
|
[
0.99951171875,
0.00036525726318359375,
0.00020813941955566406
] |
[
0.99853515625,
0.00028824806213378906,
0.0009160041809082031,
0.00010037422180175781
] |
The first generation of LNP carriers has been elaborated from proprietary lipids. Some of these components have been used as excipients in mRNA vaccines against COVID-19 and small interfering RNA (siRNA) treatment against polyneuropathy with US Food and Drug Administration (FDA) approval. 4 While allergic reactions to vaccines are rare in clinics trials, lipid components used in conventional LNP formulations are speculatively implicated in some severe cases, such as anaphylaxis. 17 , 18 In fact, the US Centers for Disease Control and Prevention and the US FDA recommend that individuals with a history of severe allergic reactions to any component of these vaccines/drugs refrain from receiving mRNA COVID-19 vaccines. 17 In regard to this, a growing interest in designing booster doses has spurred efforts to increase the number of well-defined lipids in the preparation of LNP-formulated genetic vaccines. Although recent studies have reported on a series of novel ionizable cationic lipids or helper lipids for screening nanoparticle-based DNA delivery, 19 , 20 there is a lack of in-depth analysis on the impact of the choice of PEGylated lipids and the relative proportions of lipid ingredients on the transfection efficiency and safety aspects of DNA vaccines. The present work is the first investigation on surface engineering of DNA-LNPs using diblock PLA-PEG copolymer as an alternative to PEGylated lipids. In this form, high levels of gene regulation and cellular transfection of DNA-LNPs were found in HEK293 cells; in contrast, substantial protein expression was found for free DNA. Dual-function PLA-PEG makes the obtained nanostructured lipid carrier an LNP of great versatility. Because of stability, PLA-PEG is known to be an AB-type ABC used for the design of sustained drug/vaccine delivery systems that have received great attention because they offer structure-function relationships in response to temperature stimuli. 21 This system could help further design an optimal vesicle for NA delivery that does not require cold chain storage and distribution. Regarding biocompatibility, we suggest using PEG segments of molecular weight 2,000 on the one hand, as it possesses beneficial properties, including the steric hindrance and clearance of LNPs 4 ; on the other hand, PLA was chosen as the lipophilic block, as it provides a worthwhile way to ensure the absorbability of DNA-LNPs during post-vaccination. 21 As a result, designer vaccine LNPs obtained worthwhile bioresorbable nanoparticles that help reduce the risk of side effects and adverse events.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278320_p9
|
PMC11278320
|
sec[2]/p[1]
|
Discussion
| 4.222656 |
biomedical
|
Study
|
[
0.99951171875,
0.00037407875061035156,
0.0001729726791381836
] |
[
0.99853515625,
0.000244140625,
0.0009036064147949219,
0.00009196996688842773
] |
In the present study, the results from virus infection showed that Omicron-specific DNA-LNP vaccine candidates can induce protective immunity against Wuhan strain SARS-CoV-2 challenge . We previously used angiotensin-converting enzyme 2 transgenic mice to demonstrate that such cross-protective immunity is mediated by CD8 + T cells. 14 Interestingly, we also demonstrated that immunization of DNA provided greater protection than mRNA immunization in hamsters, using the same dosage and LNP formulation. 15 It is known that the ability to remodel the tumor microenvironment and the generation of antigen-specific CD8 + T cells are crucial in successful tumor immunotherapy. 22 With this potential in mind, the anticipated research tasks could be extended for further evaluation of novel molecular therapeutic strategies into human trials, aiming to generate therapeutic immunity against cancer. Insofar as the vaccination route was concerned, the currently approved/authorized NA-based COVID-19 vaccines are administered intramuscularly (i.m.), which fail to provide protection against SARS-CoV-2 virus replication in the nasopharynx. However, the clinical trial results showed that intranasal vaccination with viral vector DNA did not produce a consistent mucosal antibody response nor a strong systemic response. 23 To effectively deliver NA genomes to mucosa-associated lymphoid tissue, it is necessary to incorporate an immunostimulatory adjuvant. This facilitates the mucosal immune system to recognize antigens, leading to the generation of protective mucosal immune responses. Further studies are warranted to find an optimal dosage of combination adjuvant to develop a nasal spray vaccine for both mucosal activity and neutralizing antibody response.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278320_p10
|
PMC11278320
|
sec[2]/p[2]
|
Discussion
| 4.082031 |
biomedical
|
Study
|
[
0.99951171875,
0.00019443035125732422,
0.00014150142669677734
] |
[
0.99365234375,
0.0007648468017578125,
0.005313873291015625,
0.00011038780212402344
] |
In summary, successful DNA vaccination can be achieved with the aid of an LNP approach that comprises a hybrid mixture of cholesterol, helper lipid, ionizable cationic lipid, and an AB-type amphiphilic bioresorbable copolymer. The rational design of DNA-LNP conciliates between efficacy and safety, which offers new insights into COVID-19 vaccine boosters. The outcomes of this study will provide critical mechanistic insights into boosting the vaccine power of lipid nanoparticles and direction for the optimization of genetic vaccine formulations against possible future emerging infectious diseases/pathogens.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278320_p11
|
PMC11278320
|
sec[3]/sec[0]/p[0]
|
Plasmid construction and characterization
| 4.117188 |
biomedical
|
Study
|
[
0.99951171875,
0.00024962425231933594,
0.00018548965454101562
] |
[
0.99951171875,
0.00027680397033691406,
0.000263214111328125,
0.00005906820297241211
] |
A plasmid DNA encoding SARS-CoV-2 TSomi was used as a vaccine antigen in immunogenicity studies, 10 , 14 , 15 which was optimized for human codon usage and synthesized by GenScript Biotech (Piscataway, NJ), as previously described. 10 In the cell culture study and in vivo distribution assessment, plasmid DNA sequences encoding EGFP or CBGr99 were used as reporter genes for monitoring the transfection/expression of the DNA-LNP formula . The TSomi and CBGr99 genes were subcloned into the clinically used vector pVAX1 with the Kozak sequence incorporated at the 5′ end of the genes, as previously described. 10 GFP was expressed by a pcDNA3.1 backbone plasmid vector. All plasmids were transformed into Escherichia coli DH5α cells (ECOS101, Yeastern Biotech, Taipei City, Taiwan) for plasmid amplification, followed by extraction and purification using an endotoxin-free Qiagen column system .
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278320_p12
|
PMC11278320
|
sec[3]/sec[1]/p[0]
|
DNA-LNP formation
| 4.132813 |
biomedical
|
Study
|
[
0.99951171875,
0.00021195411682128906,
0.00018596649169921875
] |
[
0.9990234375,
0.0006613731384277344,
0.00027942657470703125,
0.00007927417755126953
] |
Cholesterol was purchased from Sigma-Aldrich . SM-102 and DMG-PEG were purchased from Cayman Chemical . DSPC was purchased from Avanti Polar Lipids . The AB-type diblock copolymer PLA-PEG with molecular characteristics of 20 wt % PLA and 80 wt % PEG was synthesized by ring-opening polymerization of lactide on PEG monomethyl ether (number average molecular weight ∼2,000), as described previously. 24 DNA-LNP formation was elaborated using a microfluidic mixer system (NanoAssemblr Ignite; Precision Nanosystems, Vancouver, BC, Canada). Lipid mixtures (cholesterol:helper lipid:ionizable cationic lipid:PEGylated lipid) were dissolved in ethanol at predetermined molar ratios. DNA was dissolved in 6.25 mM sodium acetate aqueous solution, providing an acidic environment at pH 5.2. Typically, a volumetric flow ratio was set at 1/3 (ethanol/aqueous) and a total flow rate of 12 mL/min for the mixing of two streams flowing through microfluidic channels, where a 2-mg lipid mixture (700 μL) with cholesterol/DSPC/SM-102/PLA-PEG molar composition of 38.5/10/50/1.5 was combined with 100 μg DNA-containing buffer solution (2,100 μL). The crude formulations were diluted with a total 120 mL of 20 mM Tris-HCl buffer (pH 7.5) and concentrated with Amicon Ultra centrifugal filters . The recovered products were further sterilized by filtration using a 0.45-μm filter and stored in Tris buffer until use. mRNA-LNP formulation was synthesized in the same manner with an equivalent amount of mRNA.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278320_p13
|
PMC11278320
|
sec[3]/sec[2]/p[0]
|
Measurements
| 4.128906 |
biomedical
|
Study
|
[
0.99951171875,
0.00021398067474365234,
0.00018477439880371094
] |
[
0.99951171875,
0.0002868175506591797,
0.0003829002380371094,
0.00005805492401123047
] |
Various analytical techniques were conducted to provide a range of fundamental information of LNP, including its surface morphology using TEM techniques , particle size and electrophoretic mobility using light-scattering techniques (Malvern Zetasizer Pro-blue, Malvern Panalytical, Malvern, UK), and DNA determination using a fluorescence detection method . The particle size and electrophoretic mobility were measured by DLS and electrophoretic light scattering, respectively; and data were converted to z-average particle size (diameter, nm), polydispersity index (PDI), and zeta potential (mV). EE% of the DNA in the LNP was calculated as the following equation: EE% = (D 0 – D 1 )/D 0 × 100, where D 0 and D 1 represent total DNA in solution after and before LNP lysis, respectively. The RR% of DNA was calculated as the percentage of D 1 divided by the initial amount of DNA added.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
PMC11278320_p14
|
PMC11278320
|
sec[3]/sec[3]/p[0]
|
In vitro transfection
| 4.089844 |
biomedical
|
Study
|
[
0.99951171875,
0.00033783912658691406,
0.00022554397583007812
] |
[
0.9990234375,
0.0006856918334960938,
0.00027942657470703125,
0.0000629425048828125
] |
A group of 24-well plates were seeded with 1 × 10 5 HEK293 cells in DMEM supplemented with 10% fetal bovine serum at a total volume of 1 mL per well. At 24-h post-seeding, cells were transfected in triplicate in the presence of 1 μg DNA-LNP. Cells transfected with the commercial transfection reagent TurboFect or Lipofectamine 2000 were used as a positive control. The plates were then incubated for 3 days at 37°C and 5% CO 2 in air. The cells were stained with trypan blue solution (0.4%) following the supplier’s instructions and analyzed with the Countess 3 FL Automated Cell Counter (Thermo Fisher Scientific). The cell viability was denoted as the cells that have clear cytoplasm. The following reporter genes were selected as indicators for transfection efficiency: EGFP and CBGr99.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278320_p15
|
PMC11278320
|
sec[3]/sec[4]/p[0]
|
Hamster vaccination and challenge
| 1.37207 |
biomedical
|
Other
|
[
0.97900390625,
0.0011987686157226562,
0.0198211669921875
] |
[
0.0546875,
0.943359375,
0.001102447509765625,
0.0009708404541015625
] |
Syrian hamsters were obtained from the National Laboratory Animal Breeding and Research Center (Taipei, Taiwan), housed at the Animal Center of the National Health Research Institutes (NHRI), and maintained in accordance with institutional animal care protocols .
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
PMC11278320_p16
|
PMC11278320
|
sec[3]/sec[4]/p[1]
|
Hamster vaccination and challenge
| 3.607422 |
biomedical
|
Study
|
[
0.9990234375,
0.0003552436828613281,
0.0006561279296875
] |
[
0.99853515625,
0.0011682510375976562,
0.0002624988555908203,
0.00009083747863769531
] |
The vaccine immunogenicity study was conducted in 6- to 12-week-old hamsters. Two sets of experiments were performed: (1) sequential vaccination with TSomi DNA-LNP and (2) sequential vaccination with Wuhan-specific mRNA-LNP, followed by TSomi DNA-LNP boosting dose.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |
PMC11278320_p17
|
PMC11278320
|
sec[3]/sec[4]/p[2]
|
Hamster vaccination and challenge
| 4.132813 |
biomedical
|
Study
|
[
0.99951171875,
0.0005064010620117188,
0.00019252300262451172
] |
[
0.99951171875,
0.0002868175506591797,
0.0003418922424316406,
0.00008797645568847656
] |
For the sequential DNA vaccination experiment, hamsters ( n = 8) were vaccinated i.m. with two doses of vaccine candidates at a 2-week interval. Three vaccination groups were compared as follows: group 1 received Tris buffer only; group 2 received 10 μg DNA-LNP; and group 3 received 30 μg DNA-LNP. At weeks 2, 4, and 6 following vaccination, serum samples were obtained through retroorbital blood sampling to assess the Omicron-specific antibody titers. The presence of Omicron-specific IgG antibodies in the sera was determined by ELISA, as described previously. 14 The virus-neutralizing (VN) antibodies were determined as described below. Four weeks after the final vaccination, the hamsters under isoflurane anesthesia were challenged intranasally with 10 5 TCID 50 (50% tissue culture infectious dose) SARS-CoV-2 in a 50-μL volume. Their body weights were monitored every day after the challenge for 6 days. Subsequently, four hamsters from each group were sacrificed on days 3 and 6 after challenge for viral load quantification and pathologic examination. The viral load was determined by homogenizing left lung tissues in 2 mL PBS using a gentleMACS Dissociator (Miltenyi Biotec, Auburn, CA). After centrifugating the lung homogenate at 600 × g for 5 min, the virus-containing supernatant was collected for live virus titration by TCID 50 assay, as previously described. 15 , 19
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278320_p18
|
PMC11278320
|
sec[3]/sec[4]/p[3]
|
Hamster vaccination and challenge
| 4.105469 |
biomedical
|
Study
|
[
0.99951171875,
0.00033402442932128906,
0.0002110004425048828
] |
[
0.99951171875,
0.0003142356872558594,
0.00028061866760253906,
0.0000635981559753418
] |
For the hybrid-type vaccine booster experiment, hamsters ( n = 5) were vaccinated i.m. with two doses of 10 μg mRNA-LNP at a 3-week interval. The production process of the mRNA encoding SARS-CoV-2 Wuhan trimeric spike was performed as previously described. 15 Briefly, mRNA was synthesized by in vitro transcription reaction by T7 RNA polymerase-mediated transcription from a linearized pT7TS plasmid DNA template. The final mRNA utilizes CleanCap Reagent AG 3′ OMe as 5′ cap and full replacement of uridine with N1-methylpseudouridine-5′-triphosphate. At week 26, the hamsters were boosted i.m. with 10 μg TSomi DNA-LNP. Serum samples were collected at predetermined time points for the evaluation of systemic antibody responses.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999997 |
PMC11278320_p19
|
PMC11278320
|
sec[3]/sec[5]/p[0]
|
VN assay
| 4.15625 |
biomedical
|
Study
|
[
0.99951171875,
0.0003674030303955078,
0.00016820430755615234
] |
[
0.99853515625,
0.0007781982421875,
0.0003559589385986328,
0.00010180473327636719
] |
For the VN assay, 200 TCID 50 per well of SARS-CoV-2 virus were incubated with 2-fold-diluted hamster sera at a starting dilution of 1:40 with M199 medium . Mixtures of virus and serum were transferred to monolayers of Vero cells and incubated at 37°C and 5% CO 2 for 4 days. The neutralizing titer was defined as the reciprocal of the highest serum dilution at which the infectivity of the SARS-CoV-2 virus at 200 TCID 50 for Vero cells was completely neutralized in 50% of quadruplicate inoculations. Infectivity was identified by the presence of cytopathy after 4–5 days of incubation, and the neutralization titer was calculated using the Reed-Muench method. For calculation purposes, an undetectable level was scored as a titer equal to 20.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278320_p20
|
PMC11278320
|
sec[3]/sec[6]/p[0]
|
H&E staining and pathologic scoring
| 4.078125 |
biomedical
|
Study
|
[
0.99951171875,
0.0002300739288330078,
0.000152587890625
] |
[
0.9990234375,
0.0004131793975830078,
0.0003070831298828125,
0.00006705522537231445
] |
The lung tissue samples of infected hamsters were excised, fixed with 4% paraformaldehyde overnight, and embedded in paraffin by a routine histopathological examination method performed by the core pathology facility at the NHRI. We stained 4-μm sections with H&E and analyzed using a Leica DFC 5400 digital camera. The captured images were processed using Leica Application Suite version 4.13. To quantify the severity of lung tissue damage, the pathologic score was scored as described in a previous study. 14
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999996 |
PMC11278320_p21
|
PMC11278320
|
sec[3]/sec[7]/p[0]
|
Statistical analysis
| 4.023438 |
biomedical
|
Study
|
[
0.99951171875,
0.0002574920654296875,
0.00013899803161621094
] |
[
0.9990234375,
0.0003857612609863281,
0.0003948211669921875,
0.00006598234176635742
] |
The statistically significant difference between each vaccination group was assessed by GraphPad Prism software. The viral load in the lungs of infected hamsters and the pathologic severity scoring of lung sections after viral challenge were determined, and the p value was calculated by an ANOVA model with Tukey’s multiple comparison test. The antibody titers were compared by performing log-transformed values. The comparison of antibodies before and after the boost was performed by a two-tailed paired t test. A value of 0.05 was considered an indication of significance.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999995 |
PMC11278320_p22
|
PMC11278320
|
sec[4]/p[0]
|
Data and code availability
| 0.943359 |
other
|
Other
|
[
0.34814453125,
0.00284576416015625,
0.64892578125
] |
[
0.0199432373046875,
0.978515625,
0.0009026527404785156,
0.0006194114685058594
] |
The findings of this study are available within this paper or are available from the corresponding author upon reasonable request.
|
[
"Chung-Hsiang Yang",
"Kuan-Yin Shen",
"Hui-Min Ho",
"Chiung-Yi Huang",
"Yu-Jhen Cheng",
"Chih-Chun Pu",
"Fang-Feng Chiu",
"Wan-Chun Huang",
"Hung-Chun Liao",
"Hsin-Wei Chen",
"Ching-Len Liao",
"Shih-Jen Liu",
"Ming-Hsi Huang"
] |
https://doi.org/10.1016/j.omtn.2024.102261
|
N/A
|
https://creativecommons.org/licenses/by/4.0/
|
en
| 0.999998 |