Association between gut microbiota and endometriosis: a two-sample Mendelian randomization study.

Background: Recent studies have shown that an imbalance in gut microbiota (GM) may not always be associated with endometriosis (EMS). To investigate this further, we conducted a two-sample Mendelian randomization study. Methods: MR analysis was performed on genome-wide association study (GWAS) summary statistics of GM and EMS. Specifically, the MiBioGen microbiota GWAS (N = 18,340) was used as exposure. The FinnGen study GWAS (8,288 EMS cases and 68,969 controls) was used as outcome. We primarily used the inverse variance weighted (IVW) method to analyze the correlation and conducted a sensitivity analysis to verify its reliability. Results: (1) MR analysis: The results of the IVW method confirmed that a total of 8 GM taxa were related to the risk of EMS. Class-Melainabacteria (p = 0.036), family-Ruminococcaceae (p = 0.037), and genus-Eubacteriumruminantium (p = 0.015) had a protective effect on EMS, whereas order-Bacillales (p = 0.046), family-Prevotellaceae (p = 0.027), genus-Anaerotruncus (p = 0.025), genus-Olsenella (p = 0.036) and genus-RuminococcaceaeUCG002 (p = 0.035) could increase the risk of EMS. (2) Sensitivity analysis: Cochrane's Q test (p > 0.05), MR-Egger intercept method (p > 0.05), and leave-one-out method confirmed the robustness of MR results. Conclusion: This study performed a MR analysis on two large national databases and identified the association between 8 GM taxa and EMS. These taxa could potentially be utilized for indirectly diagnosing EMS and could lead to novel perspectives in research regarding the pathogenesis, diagnosis, and treatment of EMS.

IL-6 secretion of CD4+ T cells stimulated by LC3-positive extracellular vesicles in human epithelial ovarian cancer

Ovarian cancer (OC) as the most fatal gynecological malignancy worldwide, with epithelial ovarian cancer (EOC) being the predominant and most lethal form, poses a serious threat to human health. LC3-positive extracellular vesicles (LC3+ EVs) promote tumorigenesis by educating CD4+ T cells in a murine melanoma model. However, regulation of LC3+ EVs in human EOC remains largely unknown. MethodsDifferential analysis of Rab8a, Hsp90α and Il6 expression was performed using GEPIA2. The number of LC3+ EVs and the frequency of Heat shock protein 90α+ LC3+ EVs (HSP90α+ LC3+ EVs) in the ascites of EOC patients were tested by flow cytometry. IL-6, IL-10, IFN-γ, IL-4 and TGF-β were measured by ELISA. CD4+ T cells were isolated from peripheral blood of healthy human donors using MACS magnetic bead technology. ResultsHigher Rab8a, Hsp90a and Il6 expression of cancer tissues compared with normal adjacent tissues in OC were found. The level of IL-6 was positively correlated with LC3+ EVs number, HSP90α+ LC3+ EVs percentage in the ascites, and ROMA index of the patient. In addition, elevated IL-6 production by CD4+ T cells induced by LC3+ EVs was observed, which was suppressed by anti-HSP90α or anti-TLR2. ConclusionsLC3+ EVs level and HSP90α+ LC3+ EVs percentage were associated with elevated IL-6 in the ascites of EOC patients. HSP90α on LC3+ EVs from human EOC could stimulate CD4+ T cell production of IL-6 via TLR2. (AU)

IL-6 secretion of CD4+ T cells stimulated by LC3-positive extracellular vesicles in human epithelial ovarian cancer.

BACKGROUND: Ovarian cancer (OC) as the most fatal gynecological malignancy worldwide, with epithelial ovarian cancer (EOC) being the predominant and most lethal form, poses a serious threat to human health. LC3-positive extracellular vesicles (LC3+ EVs) promote tumorigenesis by educating CD4+ T cells in a murine melanoma model. However, regulation of LC3+ EVs in human EOC remains largely unknown.  METHODS: Differential analysis of Rab8a, Hsp90α and Il6 expression was performed using GEPIA2. The number of LC3+ EVs and the frequency of Heat shock protein 90α+ LC3+ EVs (HSP90α+ LC3+ EVs) in the ascites of EOC patients were tested by flow cytometry. IL-6, IL-10, IFN-γ, IL-4 and TGF-ß were measured by ELISA. CD4+ T cells were isolated from peripheral blood of healthy human donors using MACS magnetic bead technology.  RESULTS: Higher Rab8a, Hsp90a and Il6 expression of cancer tissues compared with normal adjacent tissues in OC were found. The level of IL-6 was positively correlated with LC3+ EVs number, HSP90α+ LC3+ EVs percentage in the ascites, and ROMA index of the patient. In addition, elevated IL-6 production by CD4+ T cells induced by LC3+ EVs was observed, which was suppressed by anti-HSP90α or anti-TLR2.  CONCLUSIONS: LC3+ EVs level and HSP90α+ LC3+ EVs percentage were associated with elevated IL-6 in the ascites of EOC patients. HSP90α on LC3+ EVs from human EOC could stimulate CD4+ T cell production of IL-6 via TLR2.

Tumor-released autophagosomes induces CD4+ T cell-mediated immunosuppression via a TLR2-IL-6 cascade.

BACKGROUND: CD4+ T cells are critical effectors of anti-tumor immunity, but how tumor cells influence CD4+ T cell effector function is not fully understood. Tumor cell-released autophagosomes (TRAPs) are being recognized as critical modulators of host anti-tumor immunity during tumor progression. Here, we explored the mechanistic aspects of TRAPs in the modulation of CD4+ T cells in the tumor microenvironment. METHODS: TRAPs isolated from tumor cell lines and pleural effusions or ascites of cancer patients were incubated with CD4+ T cells to examine the function and mechanism of TRAPs in CD4+ T cell differentiation and function. TRAPs-elicited CD4+ T cells were tested for their suppression of effector T cell function, induction of regulatory B cells, and promotion of tumorigenesis and metastasis in a mouse model. RESULTS: Heat shock protein 90α (HSP90α) on the surface of TRAPs from malignant effusions of cancer patients and tumor cell lines stimulated CD4+ T cell production of IL-6 via a TLR2-MyD88-NF-κB signal cascade. TRAPs-induced autocrine IL-6 further promoted CD4+ T cells secretion of IL-10 and IL-21 via STAT3. Notably, TRAPs-elicited CD4+ T cells inhibited CD4+ and CD8+ effector T cell function in an IL-6- and IL-10-dependent manner and induced IL-10-producing regulatory B cells (Bregs) via IL-6, IL-10 and IL-21, thereby promoting tumor growth and metastasis. Consistently, inhibition of tumor autophagosome formation or IL-6 secretion by CD4+ T cells markedly retarded tumor growth. Furthermore, B cell or CD4+ T cell depletion impeded tumor growth by increasing effector T cell function. CONCLUSIONS: HSP90α on the surface of TRAPs programs the immunosuppressive functions of CD4+ T cells to promote tumor growth and metastasis. TRAPs or their membrane-bound HSP90α represent important therapeutic targets to reverse cancer-associated immunosuppression and improve immunotherapy.

Octreotide reverses the resistance of A2780/Pacliaxel ovarian cancer cell line to paclitaxel chemotherapy in vitro.

OBJECTIVE: To study the anti-tumor effects of octreotide on A2780/Taxol ovarian cancer cells in vitro, and further explore its potential molecular mechanism. MATERIALS AND METHODS: Immunocytochemistry was performed to determine the expression of SSTR2 on A2780/Taxol cells. Octreotide at different concentrations (0, 1.25, 2.5, 5.0, 10.0, and 20.0 nmol/ml) were administrated to A2780/Taxol cells in vitro. CCK-8 assay was used to measure the effects on cell proliferation, and the cytometry of octreotide determined the cell apoptosis. The expressions of SSTR2 MDR1, and vascular endothelial growth factor (VEGF) messenger ribonucleic acid (mRNA) were investigated by quantitative reverse transcription polymerase chain reaction (RT-qPCR) assay and the expressions of the above protein were investigated after A2780/Taxol was treated with octreotide for 48 hours by western blot in vitro. RESULTS: Positive expression of SSTR2 was observed on the membrane of A2780/Taxol cells. The proliferation of A2780/Taxol cells was gradually inhibited with increasing octreotide concentration in a concentration-dependent and time-dependent manner. Meanwhile, flow cytometry data demonstrated the octreotide-induced cell apoptosis. The results of SSTR2 mRNA suggested that there was no significant difference between each concentration group of octreotide (P > 0.05). Compared with the control group, both the MDR1 and VEGF mRNA decreased in a dose-dependent manner following 48 hours of treatment of octreotide (P < 0.05). The results of western blot showed that octreotide decreased the expressions of SSTR2, MDR1, and VEGF protein in a dose-dependent manner (P < 0.05). CONCLUSIONS: Octreotide significantly inhibits ovarian cancer's proliferation and promotes its apoptosis via the cell surface expression of SSTR2. It could be used as a new targeted drug for treatment of ovarian cancer.

Does nonylphenol promote the growth of uterine fibroids?

OBJECTIVE: To study the effect and mechanism of action of nonylphenol (NP), an environmental oestrogen, on uterine leiomyoma (UL) cells. METHODS: Primary culture and subculture of human UL cells, identified as smooth muscle cells by immunocytochemical staining with a monoclonal anti-α-smooth muscle actin antibody, were performed. The viability of cells treated with various concentrations of NP for 24, 48 and 72h was determined by CCK-8 assay. mRNA expression of oestrogen receptor α (ERα), insulin-like growth factor 1 (IGF-1) and vascular endothelial growth factor (VEGF) was detected using real-time quantitative polymerase chain reaction, and protein expression was detected using Western blot analysis for all groups. RESULTS: NP promoted the growth of UL cells and expression of ERα, IGF-1 and VEGF; this was positively correlated with the concentration and duration of NP treatment. CONCLUSION: NP promotes the growth of UL cells. The mechanism of action appears to be over-expression of IGF-1 and VEGF, up-regulated by ERα, resulting in the growth of UL cells.

An evidence in vitro for the influence of bisphenol A on uterine leiomyoma.

OBJECTIVE: To study the effect and mechanism of environmental estrogens bisphenol A (BPA) on uterine leiomyoma (UL) cells. METHODS: Primary cultures and subcultures of human UL cells, which were identified by immunocytochemical staining with a monoclonal anti-α-smooth muscle actin antibody, were performed. The viability of cells in different concentration of bisphenol A of 24h, 48h and 72h were analyzed by CCK-8. The expressions of mRNA of ERα, IGF-1 and VEGF in all groups were detected by real-time quantitative PCR assay, and then the expressions of proteins detected by Western blot assay. RESULTS: BPA promoted the growth of UL cells and the expressions of ERα, IGF-1 and VEGF, which had positive correlation with the concentration and action time of BPA treatment. CONCLUSION: BPA promotes the growth of leiomyoma cells. The expressions of IGF-1, VEGF can be up-regulated by ERα, which may be possible mechanism of BPA promote the growth of leiomyoma cells.

A multicenter case-control study on screening of single nucleotide polymorphisms in estrogen-metabolizing enzymes and susceptibility to uterine leiomyoma in han chinese.

Uterine leiomyoma (UL) is an estrogen-responsive benign tumor in the female reproductive system and the main risk of hysterectomy for women. However, gene polymorphism of estrogen-metabolizing enzymes may lead to the different susceptibility to UL. We detected 10 single mucleotide polymorphisms in three key estrogen metabolite enzymes (COMT, CYP1A1, CYP1B1) in a Chinese Han population consisting of 800 patients and 800 healthy women from five different medical centers. The genetic polymorphism of rs3087869 (IVS1+2329C>T) (OR 3.200, 95% CI 1.614-6.345) and rs4680 (Val158Met) (OR 5.675, 95% CI 2.696-11.942) loci on COMT, rs1048943 (Ile462Val) (OR 4.629, 95% CI 2.216-9.672) and rs4646422 (Gly45Asp) (OR 3.240, 95% CI 1.624-6.461) loci on CYP1A1 and rs1065827 (Ala119Ser) (OR 5.635, 95% CI 2.990-10.619) locus on CYP1B1 were the risk factors to UL development and rs1056836 (Leu432Val) (OR 0.188, 95% CI 0.061-0.575) locus on CYB1B1 may be the protective factor to UL. The results provide a theoretical basis for genetic screening and early intervention to UL-susceptible populations.