Vaginal reconstruction by collagen scaffolds loaded with vaginal epithelial and smooth muscle cells in pigs.

In women, a healthy and functional vagina is important for the maintenance of a good quality of life. Various factors, including congenital anomalies, cancer, trauma, infections, inflammation, or iatrogenic injuries, can lead to damage or loss of the vaginal structure, necessitating repair or replacement. Often, such reconstruction procedures involve the use of nonvaginal tissue substitutes, like segments of the large intestine or skin, which are less than ideal both anatomically and functionally. Therefore, there is an urgent need to develop new methods of vaginal reconstruction. In this study, we established a new method for isolation and expansion of vaginal epithelial and smooth muscle cells. Subsequently, collagen scaffolds designed for vaginal reconstruction were loaded with vaginal epithelial and smooth muscle cells in vitro and tested in vivo using the vaginal excision pig model. The results showed that the collagen scaffold loaded with vaginal epithelial and smooth muscle cells significantly promotes the reconstruction of the vagina compared with small intestinal submucosa (SIS) membrane or bare collagen scaffold. Notably, the reconstructed vaginal tissues exhibit remarkable similarity to their normal counterparts, encompassing not only the vaginal epithelium and smooth muscle but also the intricate networks of blood vessels and nerves. These compelling results underscore the feasibility of a tissue engineering approach in vaginal reconstruction, offering promising prospects for improving the quality of life in affected individuals.

Tumor-derived Endothelial Cell: Important Etiological Factors in Endometriosis.

BACKGROUND AND AIM: Endometriosis (EMS) is a very complex disease with high heterogeneity. Recently, single-cell RNA sequencing (scRNA-seq) has been applied to comprehensively characterize cellular heterogeneity. Here, we built a new transcriptomic profile of EMS cellular signatures. METHODS: Three women diagnosed with endometriosis were recruited. Their fresh eutopic endometrium (EM) and ectopic endometrium (EC) tissues were sampled during surgery. ScRNA-seq was performed on 10x Genomics Chromium. RESULTS: Thirty cell clusters were identified as more than ten different cell types using cell type-specific marker genes. Re-clustering analysis revealed five subtypes of endothelial cells (ECs). Compared to EM, the proportion of tumor-derived ECs (IGFBP3+) was significantly increased in EC (43.8 vs. 16.0%). 63 differentially expressed genes (DEGs) between tumor-derived ECs and normal ECs were enriched in "angiogenesis", such as EFNB2, DLL4, and THSD7A. Subsequently, 114 retrospective EMS cases were included in clinical validation studies of EFNB2. It was co-expressed with PECAM1 and IGFBP3 and significantly increased in EC. Meanwhile, the recurrence rate of women with EFNB2++ expression was significantly higher than that of EFNB2+ cases (p <0.05). CONCLUSIONS: The significant increase in tumor-derived ECs characterized by neovascularization may be an important pathological feature of EMS. In addition, EFNB2 plays an important role and is closely related to the recurrence of EMS.

The impact of alfentanil supplementation on the sedation of bronchoscopy: A meta-analysis of randomized controlled trials.

BACKGROUND: The efficacy of alfentanil supplementation for the sedation of bronchoscopy remains controversial. We conduct a systematic review and meta-analysis to explore the influence of alfentanil supplementation on the sedation during bronchoscopy. METHODS: We search PubMed, EMbase, Web of science, EBSCO, and Cochrane library databases through December 2019 for randomized controlled trials (RCTs) assessing the effect of alfentanil supplementation versus placebo for the sedation during bronchoscopy. This meta-analysis is performed using the random-effect model. RESULTS: Five RCTs are included in the meta-analysis. Overall, compared with control group for bronchoscopy, alfentanyl supplementation is associated with significantly reduced coughing scores (Std. MD = -0.55; 95% CI = -0.96 to -0.14; P = 0.009) and dose of propofol (Std. MD = -0.34; 95% CI = -0.64 to -0.04; P = 0.03), but reveals the increase in hypoxemia (RR = 1.56; 95% CI = 1.17 to 2.08; P = 0.002). CONCLUSIONS: Alfentanyl supplementation benefits to reduce coughing scores and dose of propofol for bronchoscopy, but increases the incidence of hypoxemia. The use of alfentanyl supplementation for bronchoscopy should be with caution.

Down-regulation of PBK inhibits proliferation of human endometrial stromal cells in thin endometrium.

BACKGROUND: Thin endometrium (TE) is a challenging clinical issue in the reproductive medicine characterized by inadequate endometrial thickness, poor response to estrogen and no effective treatments currently. At present, the precise pathogenesis of thin endometria remains to be elucidated. We aimed to explore the related molecular mechanism of TE by comparing the transcriptome profiles of late-proliferative phase endometria between TE and matched controls. METHODS: We performed a bulk RNA-Seq (RNA-sequencing) of endometrial tissues in the late-proliferative phase in 7 TE and 7 matched controls for the first time. Differential gene expression analysis, gene ontology enrichment analysis and protein-protein interactions (PPIs) network analysis were performed. Immunohistochemistry was used for molecular expression and localization in endometria. Human endometrial stromal cells (HESCs) were isolated and cultured for verifying the functions of hub gene. RESULTS: Integrative data mining of our RNA-seq data in endometria revealed that most genes related to cell division and cell cycle were significantly inhibited, while inflammation activation, immune response and reactive oxygen species associated genes were upregulated in TE. PBK was identified as a hub of PPIs network, and its expression level was decreased by 2.43-fold in endometria of TE patients, particularly reduced in the stromal cells, which was paralleled by the decreased expression of Ki67. In vitro experiments showed that the depletion of PBK reduced the proliferation of HESCs by 50% and increased the apoptosis of HESCs by 1 time, meanwhile PBK expression was inhibited by oxidative stress (reduced by 76.2%), hypoxia (reduced by 51.9%) and inflammatory factors (reduced by approximately 50%). These results suggested that the insufficient expression of PBK was involved in the poor endometrial thickness in TE. CONCLUSIONS: The endometrial transcriptome in late-proliferative phase showed suppressed cell proliferation in women with thin endometria and decreased expression of PBK in human endometrial stromal cells (HESCs), to which inflammation and reactive oxygen species contributed.

Diffuse Alveolar Hemorrhage With Avelumab Maintenance Therapy.

Immune checkpoint blockade is a rapidly expanding therapeutic modality in oncology. However, its adverse effects extend beyond the cytotoxicity of conventional chemotherapy. Pneumotoxicity associated with immune checkpoint therapy presents a diagnostic conundrum that has been further complicated by the COVID-19 pandemic. We report a case of a patient with metastatic urothelial carcinoma who developed diffuse alveolar hemorrhage (DAH) following treatment with avelumab.

The efficacy of amniotic membrane-mediated sequential double-barrier therapy for the treatment of postoperative intrauterine adhesions.

ABSTRACT: To study the efficacy of using amniotic membrane, balloon and intrauterine device (IUD) as barrier therapy to prevent re-adhesion after hysteroscopic adhesiolysis.A total of 45 patients diagnosed with intrauterine adhesions in Changzhou Maternal and Child Health Hospital from June 2014 to December 2017 were included in this retrospective case control study. According to different postoperative isolation barrier methods, the patients were divided into group A (Foley balloon + fresh amniotic membrane Day1 + IUD Day7) (22 cases) and group B (Foley balloon Day1 + IUD Day7) (23 cases). Three months after the surgery, the second hysteroscopy was performed to observe the condition of the uterine cavity and the improvement of menstruation, and to monitor the thickness of the endometrium.The efficacy of hysteroscopic procedure in group A was significantly higher than that of group B (P < .05). After 3 months of treatment, the improvement rate of menstruation was significantly higher in group A than in group B (P < .05). Endometrial thickness in both group A and B was significantly increased compared with that before the surgery (P < .05). The postoperative endometrium of group A was significantly thicker than that of group B (P < .05).Amniotic membrane-mediated sequential double-barrier method is clinically feasible for preventing recurrent intrauterine adhesions.

miR-129 Blocks Secondary Hyperparathyroidism-Inducing Fgf23/αKlotho Signaling in Mice with Chronic Kidney Disease.

BACKGROUND: Secondary hyperparathyroidism, a condition of excess parathyroid hormone (PTH, Pth) production, is often seen in chronic kidney disease (CKD) patients with elevated fibroblast growth factor 23 (FGF23, Fgf23). Elevated FGF23 levels stimulate secondary hyperparathyroidism-associated parathyroid αKlotho signaling. As overexpression of rationally selected microRNAs can suppress target gene activation, we hypothesized that microRNA-based suppression of parathyroid FGF23/αKlotho axis activity may be a potential strategy to combat secondary hyperparathyroidism. METHODS: In vitro luciferase assays and human parathyroid adenoma cell experiments were used to determine miR-129-1-3p's effects on αKlotho expression in vitro. We also studied the effects of parathyroid-specific miR-129-1 overexpression (miR-129Ox) in CKD and non-CKD mice and parathyroid tissue cultures derived therefrom. RESULTS: miR-129-1-3p directly targets the αKlotho mRNA strand in human parathyroid cells. miR-129Ox CKD mice and control CKD mice displayed comparable serum levels of calcium, phosphate, Fgf23, and 1,25-dihydroxyvitamin D (1,25(OH)2D). However, miR-129Ox CKD mice displayed reduced parathyroid αKlotho expression and lower circulating Pth levels. In vitro culture of miR-129Ox CKD murine parathyroid tissue showed suppressed responses to Fgf23, with decreased Pth secretion and diminished cell proliferation after four days. CONCLUSIONS: miR-129 negatively regulates pro-proliferative, Pth-inducing Fgf23/α​Klotho signaling in the parathyroid glands of CKD mice.

PPM1A suppresses the proliferation and invasiveness of RCC cells via Smad2/3 signaling inhibition.

BACKGROUND: Cytokine therapies show promise in treating renal cell carcinoma (RCC). Transforming growth factor beta 1 (TGF-ß1) is a cytokine whose downstream Smad2/3 signaling activity is inhibited by the protein phosphatase Mg2+/Mn2+-dependent 1 A (PPM1A). Here, we hypothesized that PPM1A may be involved in suppressing RCC cell aggressiveness through its negative regulation of Smad2/3. METHODS: We quantified PPM1A expression from RCC tumors and matching healthy tissue and performed a Kaplan-Meier survival analysis. In silico analysis on PPM1A was performed using Cancer Genome Atlas-Kidney Renal Clear Cell Carcinoma and Clinical Proteomic Tumor Analysis Consortium RCC cohort data. We tested four RCC cell lines and selected the ACNH and A498 cells lines as expressing the greatest PPM1A levels. We assayed the effects of RNAi-mediated PPM1A silencing on invasiveness, proliferation, colony formation, and Smad2/3 phosphorylation in untreated and TGF-ß1-stimulated ACNH and A498 cells. A nude mouse A498 xenograft tumor model was constructed to validate PPM1A's effects in vivo. RESULTS: PPM1A levels are reduced in RCC tumors and are negatively correlated with RCC grade and stage. Below-median PPM1A expression is associated with reduced overall survival in RCC patients. PPM1A silencing promoted cellular invasiveness, proliferation, colony formation, and Smad2/3 phosphorylation under TGF-ß1-stimulated conditions but not under untreated conditions. These effects of PPM1A were shown to be dependent on Smad2/3. Intratumor PPM1A overexpression inhibited A498 xenograft tumor growth. CONCLUSIONS: This study establishes a direct link between PPM1A's suppression of Smad2/3 signaling and RCC cell aggressiveness. PPM1A could potentially serve as a biomarker for RCC cell aggressiveness.

Tissue Infiltrating Immune Cells as Prognostic Biomarkers in Endometrial Cancer: A Meta-Analysis.

BACKGROUND: The association between tumour-infiltrating immune cells and the prognosis of endometrial cancer (EC) is controversial due to the smaller sample sizes and limited statistical power of the extant studies. We carried out a meta-analysis of the relationship between tumour-infiltrating immune cells and EC survival outcomes. METHODS: A literature search in multiple databases was carried out up to December 2019. Pooled hazard ratio (HRs) and 95% confidence intervals (CIs) were calculated by the Z-test to assess the association between infiltrating immune cells and overall survival (OS), progression-free survival (PFS), relapse-free survival (RFS), disease-specific survival (DSS), and disease-free survival (DFS). A subgroup analysis was performed based on the localisation of immune cells in tumour parenchyma or stroma, immune markers, and the International Federation of Gynecology and Obstetrics stage. Heterogeneity and publication bias between studies were evaluated by Cochran's Q-test and Egger regression test, respectively. RESULTS: Seventeen studies were included in the analysis. The pooled HR of OS, PFS, DSS, and DFS indicated that a high CD8+ T cell density was associated with a favorable prognosis in EC patients. A significant relationship was found between a high density of CD45RO+ T cells and a favorable OS in EC patients, but the FoxP3+ T cell density was not associated with either OS or RFS. A high TAM density was associated with a worse PFS. However, a sensitivity analysis indicated that the findings of PFS and DSS in CD8+ T cell and PFS in TAM were not robust results. CONCLUSION: This is the first meta-analysis of the relationship between tumour-infiltrating immune cells and the prognosis of EC. High CD8+ and CD45RO+ T cell densities in tumours were associated with favorable outcomes in EC patients.

A signature motif in LIM proteins mediates binding to checkpoint proteins and increases tumour radiosensitivity.

Tumour radiotherapy resistance involves the cell cycle pathway. CDC25 phosphatases are key cell cycle regulators. However, how CDC25 activity is precisely controlled remains largely unknown. Here, we show that LIM domain-containing proteins, such as FHL1, increase inhibitory CDC25 phosphorylation by forming a complex with CHK2 and CDC25, and sequester CDC25 in the cytoplasm by forming another complex with 14-3-3 and CDC25, resulting in increased radioresistance in cancer cells. FHL1 expression, induced by ionizing irradiation in a SP1- and MLL1-dependent manner, positively correlates with radioresistance in cancer patients. We identify a cell-penetrating 11 amino-acid motif within LIM domains (eLIM) that is sufficient for binding CHK2 and CDC25, reducing the CHK2-CDC25 and CDC25-14-3-3 interaction and enhancing CDC25 activity and cancer radiosensitivity accompanied by mitotic catastrophe and apoptosis. Our results provide novel insight into molecular mechanisms underlying CDC25 activity regulation. LIM protein inhibition or use of eLIM may be new strategies for improving tumour radiosensitivity.

Expression of VEGF and HIF-1α in locally advanced cervical cancer: potential biomarkers for predicting preoperative radiochemotherapy sensitivity and prognosis.

Locally advanced cervical cancer (LACC) is an early-stage cervical cancer characterized by a local tumor diameter of ≥4 cm. Patients with LACC have a relatively poor prognosis. Although preoperative radiochemotherapy (PRCT) might offer a valuable opportunity for subsequent radical surgery, surgeons should also consider the nonresponsive rate, the adverse effects of PRCT, and the surgical complications before designing a treatment plan. Therefore, biomarkers for predicting PRCT sensitivity and prognosis in patients with LACC are of high importance. We investigated the prognostic significance of vascular endothelial growth factor (VEGF) and hypoxia inducible factor-1α (HIF-1α) in patients with LACC. A total of 43 patients with LACC who underwent PRCT (one course each of intravenous chemotherapy and after-loading intracavitary brachytherapy followed by a radical hysterectomy) during the period 2009-2014 were included in this study. VEGF and HIF-1α expression levels were evaluated by immunohistochemistry in LACC lesions before and after PRCT. In addition, we analyzed the association of these proteins with the clinical response and pathological findings of pelvic lymph node metastasis (PLNM) after the subsequent surgery. The total clinical response rate was 81.39% after PRCT, including five complete responses and 30 partial responses. VEGF and HIF-1α expression before PRCT was significantly higher than after PRCT (VEGF: 85.71% vs 66.67%; HIF-1α: 83.33% vs 59.52%, P<0.05). In addition, the same trend was found in patients with PLNM compared to those without PLNM (VEGF: 100% vs 77.78%; HIF-1α: 100% vs 74.07%, P<0.05). The areas under the receiver operating characteristic curves were 0.896 and 0.835 when using pre-PRCT VEGF and HIF-1α expression levels, respectively, to diagnose PLNM in patients with LACC. Serial detection of VEGF and HIF-1α demonstrated a sensitivity of 66.67% and specificity of 88.89%. These findings suggest that VEGF and HIF-1α expressions are potential biomarkers for PRCT and have great clinical significance for the prediction of PRCT response and prognosis in patients with LACC.

miR-424(322) reverses chemoresistance via T-cell immune response activation by blocking the PD-L1 immune checkpoint.

Immune checkpoint blockade of the inhibitory immune receptors PD-L1, PD-1 and CTLA-4 has emerged as a successful treatment strategy for several advanced cancers. Here we demonstrate that miR-424(322) regulates the PD-L1/PD-1 and CD80/CTLA-4 pathways in chemoresistant ovarian cancer. miR-424(322) is inversely correlated with PD-L1, PD-1, CD80 and CTLA-4 expression. High levels of miR-424(322) in the tumours are positively correlated with the progression-free survival of ovarian cancer patients. Mechanistic investigations demonstrated that miR-424(322) inhibited PD-L1 and CD80 expression through direct binding to the 3'-untranslated region. Restoration of miR-424(322) expression reverses chemoresistance, which is accompanied by blockage of the PD-L1 immune checkpoint. The synergistic effect of chemotherapy and immunotherapy is associated with the proliferation of functional cytotoxic CD8+ T cells and the inhibition of myeloid-derived suppressive cells and regulatory T cells. Collectively, our data suggest a biological and functional interaction between PD-L1 and chemoresistance through the microRNA regulatory cascade.