scholarly journals si-SNHG5-FOXF2 Inhibits TGF-β1-induced Fibrosis in Human Primary Endometrial Stromal Cells by Wnt/β-catenin Signaling Pathway

2020 ◽  
Author(s):  
Limin Liu ◽  
Guobin Chen ◽  
Taoliang Chen ◽  
Wenjuan Shi ◽  
Haiyan Hu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Signaling Pathway ◽  
Stromal Cells ◽  
Target Genes ◽  
Cell Model ◽  
Ex Vivo ◽  
Endometrial Stromal Cells ◽  
Downstream Target ◽  
Related Proteins ◽  
Tgf Β1

Abstract Background: Intrauterine adhesions (IUAs) are manifestations of endometrial fibrosis characterized by inflammation and fibrinogen aggregation in the extracellular matrix (ECM). The available therapeutic interventions for IUA are insufficiently effective in the clinical setting for postoperative adhesion recurrence and infertility problems. In this study, we investigated si-SNHG5-FOXF2 can serve as a molecular mechanism for inhibition of IUA fibrosis ex vivo.Methods: FOXF2, TGF-β1 and collagens expression levels were measure by a Microarray sequencing analysis in three normal endometrium group and six IUA patients. We induced primary HESCs into MFs to develop an IUA cell model with various concentrations of TGF-β1 at various times. Downstream target genes for FOXF2 were screened by chromatin immunoprecipitation combined with whole- genome high-throughput sequencing (ChIP-seq). We investigated ECM formation, cell proliferation and Wnt/β-catenin signaling pathway-related proteins in primary HESCs with FOXF2 downregulation by quantitative reverse transcription-polymerase chain reaction (qRT-PCR), western blotting (WB), immunohistochemistry (IHC), Flow cytometry, EdU and CCK8 assays. We identified lncRNA SNHG5 as the upstream regulatory gene of FOXF2 through RNA immunoprecipitation (RIP), RNA-pulldown and FISH. Finally, we examined FOXF2 expression, ECM formation, cell proliferation and Wnt/ β-catenin signaling pathway-related proteins in primary HESCs with FOXF2 downregulation.Results: FOXF2 was highly expressed in endometrium from patients with IUA. Treatment of primary HESCs with 10ng / ml TGF-β1 for 72 h was found to be most effective for developing an IUA cell model. FOXF2 regulated multiple downstream target genes including collagen, VIM and cyclin D2/DK4 by ChIP-seq and ChIP-PCR. FOXF2 downregulation inhibited TGF-β1-mediated primary HESC fibrosis including ECM formation, cell proliferation and Wnt/β-catenin signaling pathway-related proteins expressions. We identified lncRNA SNHG5 as an upstream gene that directly regulated FOXF2 by RIP-seq, qRT-PCR, WB and FISH. SNHG5 downregulation suppressed FOXF2 expression in the IUA cell model, resulting in synergistic repression of Wnt/β-catenin signaling pathway activation and thereby altering TGF-β1-mediated ECM aggregation in endometrial stromal cells ex vivo. Conclusions: Regulation of the Wnt/β-catenin signaling pathway and ECM formation by si-SNHG5-FOXF2 effectively inhibited the profibrotic effect of TGF-β1 on primary HESCs. It can provide a molecular basis for antagonizing TGF-β1-mediated fibrosis in primary HESCs.

scholarly journals si-SNHG5-FOXF2 inhibits TGF-β1-induced fibrosis in human primary endometrial stromal cells by the Wnt/β-catenin signalling pathway

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Limin Liu ◽  
Guobin Chen ◽  
Taoliang Chen ◽  
Wenjuan Shi ◽  
Haiyan Hu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Stromal Cells ◽  
Target Genes ◽  
Cell Model ◽  
Ex Vivo ◽  
Signalling Pathway ◽  
Endometrial Stromal Cells ◽  
Downstream Target ◽  
Related Proteins ◽  
Tgf Β1

Abstract Background Intrauterine adhesions (IUAs) are manifestations of endometrial fibrosis characterized by inflammation and fibrinogen aggregation in the extracellular matrix (ECM). The available therapeutic interventions for IUA are insufficiently effective in the clinical setting for postoperative adhesion recurrence and infertility problems. In this study, we investigated whether si-SNHG5-FOXF2 can serve as a molecular mechanism for the inhibition of IUA fibrosis ex vivo. Methods FOXF2, TGF-β1 and collagen expression levels were measured by microarray sequencing analysis in three normal endometrium groups and six IUA patients. We induced primary human endometrial stromal cells (HESCs) into myofibroblasts (MFs) to develop an IUA cell model with various concentrations of TGF-β1 at various times. Downstream target genes of FOXF2 were screened by chromatin immunoprecipitation combined with whole-genome high-throughput sequencing (ChIP-seq). We investigated ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related proteins in primary HESCs with FOXF2 downregulation by quantitative reverse transcription-polymerase chain reaction (qRT-PCR), western blotting (WB), immunohistochemistry (IHC), flow cytometry, ethylenediurea (EdU) and CCK8 assays. We identified long noncoding RNAs (lncRNA) SNHG5 as the upstream regulatory gene of FOXF2 through RNA immunoprecipitation (RIP), RNA pulldown and fluorescence in situ hybridization (FISH). Finally, we examined FOXF2 expression, ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related proteins in primary HESCs upon FOXF2 downregulation. Results FOXF2 was highly expressed in the endometrium of patients with IUA. Treatment of primary HESCs with 10 ng/ml TGF-β1 for 72 h was found to be most effective for developing an IUA cell model. FOXF2 regulated multiple downstream target genes, including collagen, vimentin (VIM) and cyclin D2/DK4, by ChIP-seq and ChIP-PCR. FOXF2 downregulation inhibited TGF-β1-mediated primary HESC fibrosis, including ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related protein expression. We identified lncRNA SNHG5 as an upstream gene that directly regulates FOXF2 by RIP-seq, qRT-PCR, WB and FISH. SNHG5 downregulation suppressed FOXF2 expression in the IUA cell model, resulting in synergistic repression of the Wnt/β-catenin pathway, thereby altering TGF-β1-mediated ECM aggregation in endometrial stromal cells ex vivo. Conclusions Regulation of the Wnt/β-catenin signalling pathway and ECM formation by si-SNHG5-FOXF2 effectively inhibited the profibrotic effect of TGF-β1 on primary HESCs. This finding can provide a molecular basis for antagonizing TGF-β1-mediated fibrosis in primary HESCs.

scholarly journals si-SNHG5-FOXF2 inhibits TGF-β1-induced fibrosis in human primary endometrial stromal cells by the Wnt/β-catenin signalling pathway

2020 ◽  
Author(s):  
Limin Liu ◽  
Guobin Chen ◽  
Taoliang Chen ◽  
Wenjuan Shi ◽  
Haiyan Hu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Stromal Cells ◽  
Target Genes ◽  
Cell Model ◽  
Ex Vivo ◽  
Signalling Pathway ◽  
Endometrial Stromal Cells ◽  
Downstream Target ◽  
Related Proteins ◽  
Tgf Β1

Abstract Background: Intrauterine adhesions (IUAs) are manifestations of endometrial fibrosis characterized by inflammation and fibrinogen aggregation in the extracellular matrix (ECM). The available therapeutic interventions for IUA are insufficiently effective in the clinical setting for postoperative adhesion recurrence and infertility problems. In this study, we investigated whether si-SNHG5-FOXF2 can serve as a molecular mechanism for the inhibition of IUA fibrosis ex vivo.Methods: FOXF2, TGF-β1 and collagen expression levels were measured by microarray sequencing analysis in three normal endometrium groups and six IUA patients. We induced primary human endometrial stromal cells (HESCs) into myofibroblasts (MFs) to develop an IUA cell model with various concentrations of TGF-β1 at various times. Downstream target genes of FOXF2 were screened by chromatin immunoprecipitation combined with whole-genome high-throughput sequencing (ChIP-seq). We investigated ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related proteins in primary HESCs with FOXF2 downregulation by quantitative reverse transcription-polymerase chain reaction (qRT-PCR), western blotting (WB), immunohistochemistry (IHC), flow cytometry, ethylenediurea (EdU) and CCK8 assays. We identified lncRNA SNHG5 as the upstream regulatory gene of FOXF2 through RNA immunoprecipitation (RIP), RNA pulldown and fluorescence in situ hybridization (FISH). Finally, we examined FOXF2 expression, ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related proteins in primary HESCs upon FOXF2 downregulation.Results: FOXF2 was highly expressed in the endometrium of patients with IUA. Treatment of primary HESCs with 10 ng/ml TGF-β1 for 72 h was found to be most effective for developing an IUA cell model. FOXF2 regulated multiple downstream target genes, including collagen, vimentin (VIM) and cyclin D2/DK4, by ChIP-seq and ChIP-PCR. FOXF2 downregulation inhibited TGF-β1-mediated primary HESC fibrosis, including ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related protein expression. We identified lncRNA SNHG5 as an upstream gene that directly regulates FOXF2 by RIP-seq, qRT-PCR, WB, and FISH. SNHG5 downregulation suppressed FOXF2 expression in the IUA cell model, resulting in synergistic repression of the Wnt/β-catenin pathway, thereby altering TGF-β1-mediated ECM aggregation in endometrial stromal cells ex vivo. Conclusions: Regulation of the Wnt/β-catenin signalling pathway and ECM formation by si-SNHG5-FOXF2 effectively inhibited the profibrotic effect of TGF-β1 on primary HESCs. This finding can provide a molecular basis for antagonizing TGF-β1-mediated fibrosis in primary HESCs.

scholarly journals Activated Hippo/Yes-Associated Protein Pathway Promotes Cell Proliferation and Anti-apoptosis in Endometrial Stromal Cells of Endometriosis

2016 ◽  
Vol 101 (4) ◽  
pp. 1552-1561 ◽  
Author(s):  
Yong Song ◽  
Jing Fu ◽  
Min Zhou ◽  
Li Xiao ◽  
Xue Feng ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Signaling Pathway ◽  
Nude Mice ◽  
Stromal Cells ◽  
Target Genes ◽  
Critical Role ◽  
B Cell Lymphoma ◽  
Endometrial Stromal Cells ◽  
Proliferation And Apoptosis

Abstract Context: The imbalance in cell proliferation and apoptosis is considered an important role in the pathogenesis of endometriosis, but the exact mechanisms remains unclear. A newly established signaling pathway–Hippo/Yes-associated protein (YAP) pathway plays a critical role in the proliferation and apoptosis processes. However, studies focusing on Hippo/YAP pathway and endometriosis are lacking. Objective: The objective was to explore the function of the Hippo/YAP pathway in endometriosis. Setting and Design: The expression of YAP was first investigated in endometrium of women with or without endometriosis. The role of YAP in cell proliferation and apoptosis is identified by transfection of endometrial stromal cells (ESCs) in vitro, subsequent Verteporfin treatments in eutopic ESCs in vitro, and endometriosis animal model of nude mice in vivo. Results: Our results revealed that increased expression of YAP and decreased expression of p-YAP in ectopic and eutopic endometrium compared with normal endometrium. YAP knockdown in eutopic ESCs decreased cell proliferation and enhanced cell apoptosis companied with decreased expression of TEAD1, CTGF, and B-cell lymphoma/leukemia (BCL)-2; whereas overexpression of YAP resulted in increased proliferation and decreased apoptosis of normal ESCs with increased expression of TEAD1, CTGF, and BCL-2. By chromatin immunoprecipitation qPCR CTGF and BCL-2 were identified as directly downstream target genes of YAP-TEAD1 active complex. Eutopic ESCs treated with Verteporfin revealed decreased proliferation and enhanced apoptosis whereas in endometriosis animal models of nude mice treated with Verteporfin, the size of endometriotic lesions was significantly reduced. Conclusions: Our study suggests that the Hippo/YAP-signaling pathway plays a critical role in the pathogenesis of endometriosis and should present a novel therapeutic method against endometriosis.

scholarly journals Ping-Chong-Jiang-Ni Formula Induces Apoptosis and Inhibits Proliferation of Human Ectopic Endometrial Stromal Cells in Endometriosis via the Activation of JNK Signaling Pathway

2017 ◽  
Vol 2017 ◽  
pp. 1-10 ◽  
Author(s):  
Rui-Ning Liang ◽  
Pei-Shuang Li ◽  
Yang Zou ◽  
Yu-Ling Liu ◽  
Zhen Jiang ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Signaling Pathway ◽  
Stromal Cells ◽  
Migration And Invasion ◽  
Sprague Dawley ◽  
Childbearing Age ◽  
Endometrial Stromal Cells ◽  
Jnk Signaling ◽  
Jnk Signaling Pathway ◽  
Suppress Cell Proliferation

Endometriosis is a common gynecological condition in childbearing age women and its therapy in modern medicine achieves usually temporary cure. Ping-Chong-Jiang-Ni formula (PCJNF), a Chinese herbal medicine (CHM), was shown to be clinically effective on endometriosis. Meanwhile, c-Jun N-terminal kinase (JNK) signaling pathway was involved in the therapeutic process of CHM on endometriosis. Here, we explored the effect of PCJNF on the ectopic endometrial stromal cells (EESCs) from endometriosis and test whether JNK signaling was involved. After being treated with PCJNF-containing serum obtained from Sprague Dawley rat, cell proliferation, migration, invasion, and apoptosis were evaluated in EESCs, and the total and phosphorylated JNK, ERK, and p38 proteins were detected. Our results showed that PCJNF could suppress cell proliferation, migration, and invasion and induce apoptosis in EESCs. The suppressed proliferation and increased apoptosis were dependent on JNK activation. Additionally, PCJNF caused cell cycle arrest at G2/M phase and this effect was mediated by JNK signaling, while the decreased cell migration and invasion treated by PCJNF were independent of JNK signaling. In summary, our results provided the first evidence that PCJNF could suppress cell proliferation, migration, and invasion, while increasing apoptosis in EESCs, and the suppressed proliferation and enhanced apoptosis were mediated by JNK signaling.

scholarly journals MicroRNA-342 Promotes the Malignant-Like Phenotype of Endometrial Stromal Cells via Regulation of Annexin A2

2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Dan Sun ◽  
Yiting Wang ◽  
Li Wang ◽  
Xin Guo
Keyword(s):  
Cell Proliferation ◽  
Signaling Pathway ◽  
Stromal Cells ◽  
Annexin A2 ◽  
Mtor Signaling ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Mtor Signaling Pathway ◽  
Normal Endometrium ◽  
Endometrial Stromal Cells

The relevance of miRNA- (miR-) 342 to endometriosis has been highlighted, while its function in regulating the malignant-like phenotype of endometrial stromal cells which demonstrate epigenetic abnormalities that alter expression of transcription factors, remains unclear. Therefore, we sought to characterize the effects of miR-342 in endometrial stromal cell proliferation by regulating Annexin A2 (ANXA2). We first characterized the levels of miR-342 and ANXA2 in 31 cases of normal endometrium from patients with grade II-III cervical intraepithelial neoplasia or patients with hysterectomy versus ectopic endometrial tissues of 42 patients with endometriosis. miR-342 was upregulated, while ANXA2 was downregulated in ectopic endometrial tissues. Bioinformatics website and dual-luciferase reporter assay revealed that miR-342 negatively modulated ANXA2 expression. Following loss- and gain-of-function approaches, CCK-8, Transwell, and flow cytometry demonstrated that overexpression of miR-342 markedly increased cell proliferation, migration, and invasion but inhibited cell apoptotic ratio of endometrial stromal cells, which was reversed by ANXA2 elevation. Further, overexpressed miR-342 activated the PI3K/AKT/mTOR signaling pathway, as evidenced by upregulated levels of p-PI3K/PI3K, p-AKT/AKT, and p-mTOR/mTOR. Taken together, miR-342 targets ANXA2 to activate the PI3K/AKT/mTOR signaling pathway, thereby promoting the malignant-like phenotype of endometrial stromal cells, highlighting miR-342 inhibition as a promising approach for the treatment of endometriosis.

scholarly journals CHN1 promotes epithelial–mesenchymal transition via the Akt/GSK-3β/Snail pathway in cervical carcinoma

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Haoqi Zhao ◽  
Lan Wang ◽  
Shufang Wang ◽  
Xihua Chen ◽  
Min Liang ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Lymph Node ◽  
Signaling Pathway ◽  
Cervical Carcinoma ◽  
Epithelial Mesenchymal Transition ◽  
Migration And Invasion ◽  
Xenograft Tumor ◽  
Mesenchymal Transition ◽  
Gsk 3Β ◽  
Related Proteins

Abstract Background Metastasis and invasion are crucial in determining the mortality of cervical carcinoma (CC) patients. The epithelial–mesenchymal transition (EMT) is now a universal explanation for the mechanisms of tumor metastasis. Α-chimeric protein (α-chimaerin, CHN1) plays an important role in the regulation of signal transduction and development. However, the molecular regulatory relationships between CHN1 and CC progression in relation to EMT have not yet been identified. Methods The expression of CHN1 in CC tissues, adjacent tissues, and lymph node metastases from CC patients was detected by immunohistochemistry. Upregulation and knockdown of CHN1 were achieved by transfection of CC cells. The effect of CHN1 on cell proliferation was determined by CCK-8 and plate clone formation assays. Changes in migration and invasion capabilities were evaluated using scratch migration and transwell invasion assays. The effect of CHN1 overexpression and interference on xenograft tumor growth was determined by tumor weight and pathological analyses. The expression of EMT-related mRNAs was measured by qRT-PCR in transfected CC cells. EMT-related proteins and Akt/GSK-3β/Snail signaling pathway-related proteins were also evaluated by western blotting. Results CHN1 was overexpressed in CC tissues and was associated with lymph node metastasis and low survival in CC patients. Overexpression of CHN1 promoted cell proliferation, migration, and invasion in CC cells. In contrast, silencing of CHN1 inhibited these phenomena. Overexpression of CHN1 promoted tumor formation in an in vivo xenograft tumor mouse model, with increased tumor volumes and weights. In addition, CHN1 induced the expression of EMT-related transcription factors, accompanied by the decreased expression of epithelial markers and increased expression of mesenchymal markers. The Akt/GSK-3β/Snail signaling pathway was activated by overexpression of CHN1 in vitro, and activation of this pathway was inhibited by the signaling pathway inhibitor LY294002. Conclusion These results suggest that CHN1 promotes the development and progression of cervical carcinoma via the Akt/GSK-3β/Snail pathway by inducing EMT.

scholarly journals Mathematical analysis of the effect of portal vein cells on biliary epithelial cell differentiation through the Delta-Notch signaling pathway

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Masaharu Yoshihara ◽  
Teppei Nishino ◽  
Manoj Kumar Yadav ◽  
Akihiro Kuno ◽  
Takeshi Nagata ◽  
...  
Keyword(s):  
Portal Vein ◽  
Notch Signaling ◽  
Signaling Pathway ◽  
Target Genes ◽  
Notch Signaling Pathway ◽  
Epithelial Differentiation ◽  
Production Rates ◽  
Fine Grained ◽  
Downstream Target ◽  
Notch Receptors

Abstract Objective The Delta-Notch signaling pathway induces fine-grained patterns of differentiation from initially homogeneous progenitor cells in many biological contexts, including Drosophila bristle formation, where mathematical modeling reportedly suggests the importance of production rate of the components of this signaling pathway. In contrast, the epithelial differentiation of bile ducts in the developing liver is unique in that it occurs around the portal vein cells, which express extremely high amounts of Delta ligands and act as a disturbance for the amount of Delta ligands in the field by affecting the expression levels of downstream target genes in the cells nearby. In the present study, we mathematically examined the dynamics of the Delta-Notch signaling pathway components in disturbance-driven biliary differentiation, using the model for fine-grained patterns of differentiation. Results A portal vein cell induced a high Notch signal in its neighboring cells, which corresponded to epithelial differentiation, depending on the production rates of Delta ligands and Notch receptors. In addition, this epithelial differentiation tended to occur in conditions where fine-grained patterning was reported to be lacking. These results highlighted the potential importance of the stability towards homogeneity determined by the production rates in Delta ligands and Notch receptors, in a disturbance-dependent epithelial differentiation.

scholarly journals The Role of the FOXO1/β2-AR/p-NF-κB p65 Pathway in the Development of Endometrial Stromal Cells in Pregnant Mice under Restraint Stress

2021 ◽  
Vol 22 (3) ◽  
pp. 1478
Author(s):  
Jiayin Lu ◽  
Yaoxing Chen ◽  
Zixu Wang ◽  
Jing Cao ◽  
Yulan Dong
Keyword(s):  
Oxidative Stress ◽  
Stromal Cells ◽  
Restraint Stress ◽  
Target Genes ◽  
Mrna Levels ◽  
Endometrial Stromal Cells ◽  
Protein Levels ◽  
Pregnant Mice

Restraint stress causes various maternal diseases during pregnancy. β2-Adrenergic receptor (β2-AR) and Forkhead transcription factor class O 1 (FOXO1) are critical factors not only in stress, but also in reproduction. However, the role of FOXO1 in restraint stress, causing changes in the β2-AR pathway in pregnant mice, has been unclear. The aim of this research was to investigate the β2-AR pathway of restraint stress and its impact on the oxidative stress of the maternal uterus. In the study, maternal mice were treated with restraint stress by being restrained in a transparent and ventilated device before sacrifice on Pregnancy Day 5 (P5), Pregnancy Day 10 (P10), Pregnancy Day 15 (P15), and Pregnancy Day 20 (P20) as well as on Non-Pregnancy Day 5 (NP5). Restraint stress augmented blood corticosterone (CORT), norepinephrine (NE), and blood glucose levels, while oestradiol (E2) levels decreased. Moreover, restraint stress increased the mRNA levels of the FOXO family, β2-AR, and even the protein levels of FOXO1 and β2-AR in the uterus and ovaries. Furthermore, restraint stress increased uterine oxidative stress level. In vitro, the protein levels of FOXO1 were also obviously increased when β2-AR was activated in endometrial stromal cells (ESCs). In addition, phosphorylated-nuclear factor kappa-B p65 (p-NF-κB p65) and its target genes decreased significantly when FOXO1 was inhibited. Overall, it can be said that the β2-AR/FOXO1/p-NF-κB p65 pathway was activated when pregnant mice were under restraint stress. This study provides a scientific basis for the origin of psychological stress in pregnant women.

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