Analysis of related factors for pathological upgrading of cervical biopsy from CIN3 to cancer after conical resection.

BACKGROUND: To investigate related factors for postoperative pathological upgrading of cervical biopsy to cervical cancer (CC) in patients with cervical intraepithelial neoplasia (CIN)3 after conical resection. METHODS: This retrospective study collected data from patients diagnosed with CIN3 by cervical biopsies at the author's Hospital between January 2012 and December 2022. The primary outcome was the pathological results of patients after conical resection. The pathological findings were categorized into the pathological upgrading group if postoperative pathology indicated CC, while those with normal, inflammatory, or cervical precancerous lesions were classified into the pathological non-upgrading group. The factors associated with upgrading were identified using multivariable logistic regression analysis. RESULTS: Among 511 patients, there were 125 patients in the pathological upgrading group (24.46%). The patients in the upgrading group were younger (47.68 ± 9.46 vs. 52.11 ± 7.02, P < 0.001), showed a lower proportion of menopausal women (38.40% vs. 53.02%, P = 0.0111), a lower proportion of HSIL (40.00% vs. 57.77%, P = 0.001), a higher rate of HPV-16/18 positive (25.60% vs. 17.36%, P = 0.011), a higher rate of contact bleeding (54.40% vs. 21.50%, P < 0.001), lower HDL levels (1.31 ± 0.29 vs. 1.37 ± 0.34 mmol/L, P = 0.002), higher neutrophil counts (median, 3.50 vs. 3.10 × 109/L, P = 0.001), higher red blood cell counts (4.01 ± 0.43 vs. 3.97 ± 0.47 × 1012/L, P = 0.002), higher platelet counts (204.84 ± 61.24 vs. 187.06 ± 73.66 × 109/L, P = 0.012), and a smaller platelet volume (median, 11.50 vs. 11.90 fL, P = 0.002).The multivariable logistic regression analysis showed that age (OR = 0.90, 95% CI: 0.86-0.94, P < 0.001), menopausal (OR = 2.68, 95% CI: 1.38-5.22, P = 0.004), contact bleeding (OR = 4.80, 95% CI: 2.91-7.91, P < 0.001), and mean platelet volume (OR = 0.83, 95% CI: 0.69-0.99, P = 0.038) were independently associated with pathological upgrading from CIN3 to CC after conical resection. CONCLUSION: Age, menopausal, contact bleeding, and mean platelet volume are risk factors of pathological upgrading from CIN3 to CC after conical resection, which could help identify high risk and susceptible patients of pathological upgrading to CC.

USP7 promotes decidualization of ESCs by STAT3/PR axis during early pregnancy.

Decidualization is an essential process for embryo implantation during early pregnancy. Defective decidualization is a critical leading cause of early pregnancy loss (EPL). Ubiquitin-specific protease 7 (USP7) is a deubiquitinating enzyme that is involved in uterine function. This study aimed to explore the underlying mechanism by which USP7 regulates decidualization in EPL. We found that USP7 was downregulated in the decidual tissue of EPL patients. Upregulation of USP7 enhanced decidualization in endometrial stromal cells (ESCs), with increased decidualized biomarkers IGFBP1 and PRL and progesterone receptor A/B (PR-A/B) expression. Moreover, we found that signal transducer and activator of transcription 3 (STAT3) is a direct target of USP7 in ESCs. USP7 bound to STAT3 by deubiquitination and increased STAT3 levels in ESCs. Suppression of STAT3 impeded the USP7-promoted cell viability, decidualization, and PR-A/B expression of ESCs. USP7 promoted the decidualization of ESCs through the STAT3/PR signaling pathway during early pregnancy, which provides new insight into the pathological mechanism of EPL and may contribute to the clinical treatment of EPL.

Primary clear cell adenocarcinoma of the rectovaginal septum: A rare case report.

RATIONALE: Primary clear cell adenocarcinoma of the rectovaginal septum is a very rare event. PATIENT CONCERNS: We reported a case of a 55-year-old woman diagnosed with a lump in the vaginal rectal septum after undergoing hysterectomy with bilateral salpingo-oophorectomy in 2017, who was admitted to our department due to vaginal bleeding. Magnetic resonance imaging of the pelvis indicated the vaginal rectal space cystic and solid mass about 110 mm × 100 mm × 140 mm in size. DIAGNOSIS: The pathological diagnosis of postoperative was clear cell adenocarcinoma. INTERVENTIONS: Abdominal laparotomy showed a solid block of the vaginal rectal septum. Surgery was performed to reduce the tumor. OUTCOMES: This patient received 8 courses of combined chemotherapy courses after surgery for the residual lesion and achieved a complete response. LESSONS: Due to the rare observation of the growth pattern, the cell morphology and immune phenotype are not specific, and clinical and pathological diagnosis is difficult. Introducing the diagnosis and treatment of this case and reviewing the literature provide a relevant reference for clinicians identification and diagnosis and treatment of this rare case.

TRIM44 Promotes Endometrial Carcinoma Progression by Activating the FRS2 Signalling Pathway.

The Tripartite Motif Containing 44 (TRIM44) is highly expressed in a variety of tumours. However, the TRIM44's role in endometrial carcinoma (EC) progression remains unknown. To investigate the TRIM44's role in the development and metastasis of EC, we detected TRIM44 expression in EC cell lines and surgical specimens from patients with EC using immunohistochemistry, real-time reverse transcription-polymerase chain reaction, and western blotting analysis. The biological functions of TRIM44 by loss-of-function analysis in RL95-2 and Ishikawa cells were studied. The effect of TRIM44 on the progression of EC in terms of cell proliferation, apoptosis, and invasion was examined and revealed its underlying mechanism in vitro using EC cell lines and in vivo using mouse xenograft models. The TRIM44's expression was positively correlated with EC progression and poor prognosis. The TRIM44 knockdown reduced the EC cell proliferation and invasion while promoting cell apoptosis. Mechanism experiments showed that the TRIM44 interacts with Fibroblast Growth Factor Receptor Substrate 2 (FRS2) and negatively regulates the expression of Bone Morphogenetic Protein 4(BMP4), ß-catenin, and Transforming Growth Factor Beta Receptor 1(TGF-ßR1). Moreover, the effect of TRIM44 overexpression on EC cell proliferation, invasion, and apoptosis is reversed by the FRS2 knockdown. Our study may provide a new perspective on targeting the TRIM44/FRS2 signaling pathway in treating EC, which deserves further investigation.

Performance characterization of PCR-free whole genome sequencing for clinical diagnosis.

ABSTRACT: To evaluate the performance of polymerase chain reaction (PCR)-free whole genome sequencing (WGS) for clinical diagnosis, and thereby revealing how experimental parameters affect variant detection.Five NA12878 samples were sequenced using MGISEQ-2000. NA12878 samples underwent WGS with differing deoxyribonucleic acid (DNA) input and library preparation protocol (PCR-based vs PCR-free protocols for library preparation). The depth of coverage and genotype quality of each sample were compared. The performance of each sample was measured for sensitivity, coverage of depth and breadth of coverage of disease-related genes, and copy number variants. We also developed a systematic WGS pipeline (PCR-free) for the analysis of 11 clinical cases.In general, NA12878-2 (PCR-free WGS) showed better depth of coverage and genotype quality distribution than NA12878-1 (PCR-based WGS). With a mean depth of ∼40×, the sensitivity of homozygous and heterozygous single nucleotide polymorphisms (SNPs) of NA12878-2 showed higher sensitivity (>99.77% and >99.82%) than NA12878-1, and positive predictive value exceeded 99.98% and 99.07%. The sensitivity and positive predictive value of homozygous and heterozygous indels for NA12878-2 (PCR-free WGS) showed great improvement than NA128878-1. The breadths of coverage for disease-related genes and copy number variants are slightly better for samples with PCR-free library preparation protocol than the sample with PCR-based library preparation protocol. DNA input also influences the performance of variant detection in samples with PCR-free WGS. All the 19 previously confirmed variants in 11 clinical cases were successfully detected by our WGS pipeline (PCR free).Different experimental parameters may affect variant detection for clinical WGS. Clinical scientists should know the range of sensitivity of variants for different methods of WGS, which would be useful when interpreting and delivering clinical reports.

Expression and potential role of CXCL5 in the pathogenesis of intrauterine adhesions.

OBJECTIVE: C-X-C motif chemokine ligand 5 (CXCL5), a member of the chemokine family, is associated with remodeling of connective tissues. However, its role in formation of intrauterine adhesions (IUA) remains unclear. We aimed to investigate the expression and mechanism underlying the role of CXCL5 in IUA. METHODS: Expression of CXCL5 in IUA was detected by immunohistochemistry in a rat model of IUA and by real-time PCR and western blotting in patients with IUA. The protein levels of matrix metalloproteinase 9 (MMP9) and transcription factor p65 in human endometrial cells were assessed by western blotting after CXCL5 overexpression. RESULTS: Protein expression of CXCL5 was significantly decreased in the endometria of IUA rats compared with that of control and sham-operated rats. Real-time PCR and western blotting in patients with IUA showed similar results to those from the rat model. After overexpression, CXCL5 significantly upregulated expression of MMP9 and slightly upregulated expression of p65 in human endometrial cells. CONCLUSIONS: CXCL5 plays an important role in IUA formation after endometrial injury. We propose a molecular mechanism to explain formation of IUA, including downregulation of MMP9 by low CXCL5 expression. These findings provide valuable information for the prevention and targeted therapy of IUA.

miR-219a suppresses human trophoblast cell invasion and proliferation by targeting vascular endothelial growth factor receptor 2 (VEGFR2).

OBJECTIVE: Vascular endothelial growth factor (VEGF) plays a critical role in regulating trophoblast cell invasion and proliferation, involved in a variety of pregnancy complications, such as spontaneous abortion and pre-eclampsia. Numerous studies have revealed that microRNAs (miRNAs) are participated in a series of molecular processes that regulate cell function, such as cell invasion, proliferation, and apoptosis. Vascular endothelial growth factor receptor 2 (VEGFR2), a receptor of VEGF, has been shown to be involved in trophoblast function. However, the relation between miRNA and VEGFR2 and their role in trophoblast function remain to be elucidated. METHODS: The effect of miR-219a on the trophoblast function has been explored using luciferase reporter, transwell, qRT-PCR, western blot, bromodeoxyuridine (BrdU), ELISA, immunofluorescent staining, and tube formation assays. RESULTS: In the current study, we observed that through targeted inhibition of VEGFR2 expression by miR-219a, the function of VEGFR2 as well as the downstream PI3K/AKT/NF-κB signaling pathway were suppressed, leading to suppression of trophoblastic proliferation and invasion. Moreover, upregulation of VEGFR2 restored the miR-219a-inhibited cell proliferation, invasion, and tube formation. CONCLUSIONS: These results revealed that miR-219a played crucial roles in negatively regulating trophoblastic proliferation and invasion by suppression of the PI3K/AKT/NF-κB signaling pathway by targeting VEGFR2, therefore serving as a potential treatment method for the complications of pregnancy caused by trophoblastic dysregulation.

Elevated miR-23a impairs trophoblast migration and invasiveness through HDAC2 inhibition and NF-κB activation.

Preeclampsia (PE) is a pregnancy-specific disorder characterized by the onset of hypertension and proteinuria with onset after the 20th week of gestation. The pathogenesis of PE is attributed to increased trophoblast cell death and poor trophoblast migration/invasiveness. This study investigates the function of microRNA-23a (miR-23a) in PE and its effects on migration and invasion of trophoblast cells HTR-8/SVneo. We found higher expression of miR-23a in placental tissue samples from PE pregnant women compared to samples from normal pregnant women. Enhancing miR-23a expression by its specific mimic reduced HTR-8/SVneo cell migration and invasion and increased HTR-8/SVneo cell apoptosis. The dual-luciferase reporter gene assay revealed miR-23a binding with HDAC2. We found that HDAC2 was poorly expressed in placental tissue samples from PE pregnant women, and its expression correlated inversely with miR-23a expression. HTR-8/SVneo cells showed diminished HDAC2 expression upon miR-23a elevation and increased HDAC2 expression upon miR-23a inhibition. Lentivirus-mediated HDAC2 knockdown mimicked the effects of miR-23a on HTR-8/SVneo cells and led to NF-κB activation. Similarly, HDAC2 overexpression and NF-κB inhibition both abrogated the effects of miR-23a on HTR-8/SVneo cells, suggesting that miR-23a reduced HTR-8/SVneo cell migration and invasion and increased HTR-8/SVneo cell apoptosis by HDAC2 inhibition and NF-κB activation. In summary, these results support a novel role of miR-23b in invasion and apoptosis of trophoblast cells, and imply that targeting miR-23b may be a new avenue for treating PE.

circ-ZUFSP regulates trophoblasts migration and invasion through sponging miR-203 to regulate STOX1 expression.

Recurrent spontaneous abortion (RSA), defined as two or more consecutive pregnancy losses before 12 weeks of gestation with or without previous live births. Circular RNAs (circRNAs) are a novel class of endogenous noncoding RNAs that play important roles in gene expression regulation and trophoblasts function during embryo development. This study aimed to evaluate the function mechanism of circRNAs regulating trophoblasts function in the occurrence and progression RSA. Through overexpression and down-regulation of circ-ZUFSP, we investigated the effect of circ-ZUFSP on the function of trophoblasts and found loss of circ-ZUFSP suppressed trophoblasts migration and invasion in vitro. Moreover, loss of circ-ZUFSP regulated trophoblasts migration and invasion via regulation of miR-203. Furthermore, STOX1 was revealed to a target of miR-203, and down-regulation of STOX1 reversed circ-ZUFSP enhanced cell invasion, suggesting that circ-ZUFSP might regulate STOX1 expression through sponging miR-203 in HTR-8/SVneo cells. In addition, low expression of circ-ZUFSP, STOX1 and high expression of miR-203 were testified in placental tissues of RSA mice. Our study demonstrated a molecular mechanism of circ-ZUFSP regulating trophoblasts migration and invasion, which might provide a novel indicator for early diagnosis and potential treatment of RSA.

Long non-coding RNA Linc00483 accelerated tumorigenesis of cervical cancer by regulating miR-508-3p/RGS17 axis.

OBJECTIVES: The aim of this study was to uncover the underlying mechanisms of cervical cancer progression and provide potential therapeutic targets for its treatment in clinic. MATERIALS AND METHODS: Real-Time qPCR was used to determine the expression levels of Linc00483, miR-508-3p and RGS17 mRNA in cervical cancer tissues and cell lines. Terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) assay was conducted to determine cell apoptosis. Western Blot was performed to detect protein expression levels. Wound healing and Transwell assay were employed to determine cell migration and invasion respectively. Online software (TargetScan, miRDB and miR TarBase) were used to predict the regulating mechanisms of Linc00483, miR-508-3p and RGS17, which were validated by dual-luciferase reporter gene system. In vivo tumor-bearing mice models were established to validate the cellular results. RESULTS: Linc00483 aberrantly overexpressed in both cervical cancer tissues and cell lines comparing to the Control groups. Knock-down of Linc00483 inhibited cervical cancer cell proliferation, invasion as well as migration, and promoted cell apoptosis. In addition, miR-508-3p was identified as the downstream target of Linc00483, and miR-508-3p inhibitor abrogated the inhibiting effects of downregulated Linc00483 on cervical cancer cell viability. Furthermore, the expression levels of Linc00483 was positively correlated with RGS17 in the clinical samples and overexpressed Linc00483 increased RGS17 expression levels in cervical cancer cells by sponging miR-508-3p. The in vivo experiments showed that knock-down of Linc00483 inhibited cervical cancer cell tumorigenesis and lung metastasis in mice models. CONCLUSIONS: Knock-down of Linc00483 inhibited the development of cervical cancer by regulating miR-508-3p/RGS17 axis.

TGF-ß1 alleviates HgCl2 induced apoptosis via P38 MAPK signaling pathway in human trophoblast cells.

It is well known that embryonic development can be perturbed by environmental factors such as heavy metals. Mercury is one of the most significant threats to the environment and human health. Mercury can damage many parts of the human body, including lungs, kidneys, nerves and fetus. However, the effect of mercury on human embryo remains unknown. Here, we showed that HgCl2 treatment resulted in a significant increase in apoptosis in HTR-8/SVneo cells. However, the effect of HgCl2 on apoptosis was partially reduced by the combination treatment with TGF-ß1 and HgCl2 in HTR-8/SVneo cells. Moreover, HgCl2 treatment gradually decreased the expression of TGF-ß1 in a dose dependent manner. Furthermore, a P38 MAPK inhibitor, SB202190, decreased the cell apoptosis and caspase activation induced by HgCl2 in trophoblast cells. In addition, TGF-ß1 alleviated HgCl2 induced apoptosis of HTR-8/SVneo cells via p38 MAPK signaling pathway, which was involved in the TAK1 expression. These results might provide a theoretical basis for mercury induced trophoblast associated embryo damage and a potential avenue of intervention.

Effect of STOX1 on recurrent spontaneous abortion by regulating trophoblast cell proliferation and migration via the PI3K/AKT signaling pathway.

STOX1 is a transcription factor that is implicated in the high prevalence of human gestational diseases. It has been studied in various types of gestational diseases using different molecular and cellular biological technologies. However, the effect and detailed mechanism of storkhead box 1 (STOX1) in recurrent spontaneous abortion (RSA) remain unknown. This study aimed to explore the effect and detailed mechanism of STOX1 in human trophoblast cells. The result showed that downregulation of STOX1 by short hairpin RNA (shRNA) led to a decrease in proliferation and migration in HTR-8/SVneo cells, while it induced the apoptosis of HTR-8/SVneo cells. Moreover, the result showed that trophoblast cells expressed lower levels of pAKT and p85 subunits after treatment with STOX1 shRNA when compared with control. However, overexpression of STOX1 obviously increased the pAKT and p85 protein expressions. Transfection of pcDNA-AKT plasmid increased cell proliferation and migration in HTR-8/SVneo cells while suppressed the apoptosis of HTR-8/SVneo cells. Furthermore, inhibition of the PI3K/Akt pathway by a specific inhibitor promoted cell apoptosis and aggravatedly suppressed cell proliferation and migration of HTR-8/SVneo cells. On the other hand, upregulation of the PI3K/Akt pathway could increase the relative expression level of Bcl-2 and decrease the relative expression levels of Bax and Bim, while inhibition of the PI3K/Akt pathway led to adverse results. Our results demonstrated that inhibition of STOX1 could suppress trophoblast cell proliferation and migration, while promote apoptosis through inhibiting the PI3K/Akt signaling pathway. These findings might provide a new fundamental mechanism for regulating RSA and could be used to prevent and treat RSA in clinic.