A novel fast hybrid capture sequencing method for high-efficiency common human coronavirus whole-genome acquisition.

Rapid and accurate sequencing of the entire viral genome, coupled with continuous monitoring of genetic changes, is crucial for understanding the epidemiology of coronaviruses. We designed a novel method called micro target hybrid capture system (MT-Capture) to enable whole-genome sequencing in a timely manner. The novel design of probes used in target binding exhibits a unique and synergistic "hand-in-hand" conjugation effect. The entire hybrid capture process is within 2.5 hours, overcoming the time-consuming and complex operation characteristics of the traditional liquid-phase hybrid capture (T-Capture) system. By designing specific probes for these coronaviruses, MT-Capture effectively enriched isolated strains and 112 clinical samples of coronaviruses with cycle threshold values below 37. Compared to multiplex PCR sequencing, it does not require frequent primer updates and has higher compatibility. MT-Capture is highly sensitive and capable of tracking variants.IMPORTANCEMT-Capture is meticulously designed to enable the efficient acquisition of the target genome of the common human coronavirus. Coronavirus is a kind of virus that people are generally susceptible to and is epidemic and infectious, and it is the virus with the longest genome among known RNA viruses. Therefore, common human coronavirus samples are selected to evaluate the accuracy and sensitivity of MT-Capture. This method utilizes innovative probe designs optimized through probe conjugation techniques, greatly shortening the time and simplifying the handwork compared with traditional hybridization capture processes. Our results demonstrate that MT-Capture surpasses multiplex PCR in terms of sensitivity, exhibiting a thousandfold increase. Moreover, MT-Capture excels in the identification of mutation sites. This method not only is used to target the coronaviruses but also may be used to diagnose other diseases, including various infectious diseases, genetic diseases, or tumors.

Follicular fluid progesterone downregulated HPGD and COX2 in granulosa cells via suppressing NF-ĸB in endometriosis†.

Increasing evidences showed that ovulatory dysfunction, possibly caused by luteinized unruptured follicular follicle syndrome (LUFS), is one of the reasons for endometriosis-related infertility. The present study was conducted to explore the potential effect of elevated progesterone in follicular fluid (FF) on ovulation in endometriosis. A prospective study including 50 ovarian endometriosis patients and 50 control patients with matched pairs design was conducted with alterations in FF and peritoneal fluid (PF) components identified by metabolomics analyses and differentially expressed genes in granulosa cells (GCs) identified by transcriptome analysis. Patients with endometriosis exhibited a significantly higher progesterone level in serum, FF, and PF. Granulosa cells from endometriosis patients revealed decreased expression of HPGD, COX-2, and suppressed NF-ĸB signaling. Similarly, progesterone treatment in vitro downregulated HPGD and COX2 expression and suppressed NF-ĸB signaling in granulosa tumor-like cell line KGN (Bena Culture Collection, China) and primarily cultured GCs, as manifested by decreased expressions of IL1R1, IRAK3, reduced pIĸBα/IĸBα ratio, and nucleus translocation of p65. On the contrary, TNF-α treatment increased expression of IL1R1, IRAK3, pIĸBα, p65, and HPGD in GCs. One potential p65 binding site was identified in the promoter region of HPGD by chromatin immunoprecipitation. In conclusion, we found that intrafollicular progesterone might downregulate HPGD and COX-2 in GCs via suppressing the NF-ĸB signaling pathway, shedding light on the mechanism underlying the endometriosis-related ovulatory dysfunction.

Fluorescence probe for selectively monitoring biothiols within cells and mouse depression diagnosis.

As a global mental disorder, depression is associated with oxidative stress in the brain. Cysteine, a reductive biothiols, regulates the oxidative situation in many biological events including the stress that occurs in the tissues. Exploring the pathology and physiology of depression is still a challenge and always in an urgent need. Thus, developing a new method that could track Cys level without the interferes from other competing substances is of great importance. Herein, we developed a fluorescence probe that could selectively sensing Cys over other biothiols. Besides, we have demonstrated its desirable performance in cellular applications and mouse brain. This work provides a new method for Cys imaging and understanding pathogenesis of depression. We hope the work described here could be used as a potential chemical approach for the diagnosis of Cys associated diseases in clinical applications.

Piperazine-Coumarin based fluorescence probe with enhanced brightness and solubility for bio-thiol detection and esophageal carcinoma diagnosis.

The development of novel fluorescent dyes for bio-thiol is of great importance in biological, clinical and pharmaceutical sciences. Given the importance of bio-thiol anticipating in numerous physiological processes, there is a great need to construct fluorescent biosensors with high quality to detect them. Fluorophores, especially those used in bio-system, usually require high-quality properties such as high brightness, good water solubility, bio-compatible and photostability. Herein, we reported a novel fluorescent probe based on piperazine-coumarin scaffold with enhanced brightness and solubility. To further demonstrate the potential clinical applications, we performed living cell fluorescence image and human esophageal carcinoma diagnosis. The result indicated that we were able to distinguish pathological tissue from normal tissue by applying this probe. Thus, we hope this design will be helpful to develop high-quality fluorophores for clinical diagnosis.

Icariin Inhibits Intestinal Inflammation of DSS-Induced Colitis Mice Through Modulating Intestinal Flora Abundance and Modulating p-p65/p65 Molecule.

BACKGROUND: Ulcerative colitis, as a kind of inflammatory bowel disease (IBD) is characterized by abdominal pain. This study aimed to investigate the effect of icariin (ICA) on the intestinal microflora of colitis mice. METHODS: Fifteen female C57BL/6 mice were randomly divided into the Control group, dextran sodium sulfate (DSS)-induced colitis (DSS) group, and ICA treatment (DSS+ICA) group. The severity of inflammation in DSS-induced colitis mice was evaluated using disease activity scoring (considering weight-loss percentage, stool-shape change, and stool-bleeding scoring). Pathological changes of mice intestinal tract were evaluated using hematoxylin-eosin (HE) staining. Serum levels of TNF-α and IL-6 were detected with enzyme-linked immunosorbent assay. Expressions of p65 and p-p65 (p-p65/p65 ratio) were analyzed using Western blot assay. 16S rDNA sequencing was used to analyze the abundance and composition of intestinal microflora. RESULTS: Compared with DSS group, ICA significantly improved disease activity (P < .05) and reduced inflammatory damage of colon tissues (P < .05) in DSS-induced colitis mice. Compared with the DSS group, mice in the ICA group demonstrated significant weight and colon length (P < .05). ICA significantly inhibited expressions of IL-6 and TNF-α compared to the DSS group (P < .05). p-p65/ p65 ratio in the DSS + ICA group was remarkably enhanced compared to the DSS group (P < .05). ICA significantly reduced the proportion and activity of Bacteroides, Helicobacteraceae, Turicibacter, and significantly increased that of beneficial microflora (Lactobacillus, Lachnospiraceae, Akkermansia), so as improved damages of colon tissues. CONCLUSION: ICA can improve intestinal flora abundance and composition of DSS-induced colitis mice, and inhibit tissue damage and inflammatory response through modulating the p-p65/p65 expression.

Single-cell transcriptomic analysis of endometriosis provides insights into fibroblast fates and immune cell heterogeneity.

BACKGROUND: Endometriosis is an oestrogen-dependent disease with an unclear aetiology and pathogenesis affecting 6-10% of the global female population, predominantly those of reproductive age. Herein, we profile the transcriptomes of approximately 55,000 single cells from three groups including ectopic endometrium, eutopic endometrium from women with endometriosis, and eutopic endometrium from healthy women to create a single-cell transcriptome atlas of endometriosis. RESULTS: We have identified 9 cell types and performed single-cell analysis of fibroblasts, and determined a potential developmental trajectory associated with endometriosis. We also identified fibroblast subpopulations related to endometriosis development and found that StAR played an important role in this process. Moreover, T cells in endometriosis were less activated or inflammatory with decreased effector CD8 + T cells, while the composition ratio of natural killer cells decreased and the percentage of monocytes/macrophages increased in endometriosis cysts. In addition, the effectiveness of immune cells in endometriosis lesions, eutopic endometrium from women with endometriosis, and eutopic endometrium from healthy women was distinct. Cell-cell interaction analyses highlighted the imbalanced immune environment in endometriosis lesions and immune cells in endometriosis could promote the development of the disease. CONCLUSION: Our study provided a systematic characterisation of endometriosis and insights into the aetiology and pathology of endometriosis.

Overexpressed MPS-1 contributes to endometrioma development through the NF-κB signaling pathway.

BACKGROUND: Endometriosis is a benign gynecological disease that shares some characteristics with malignant tumors and affects approximately 10% of women of reproductive age. Endometrioma refers to endometriosis that appears in the ovary. Metallopanstimulin-1 (MPS-1) is a component of the 40S subunit of ribosomes that has extra-ribosomal functions that contribute to the development of diseases. This study aimed to explore the expression pattern and role of MPS-1 in endometrioma development. METHODS: Quantitative real time polymerase chain reaction, western blotting, immunohistochemistry, and enzyme-linked immunosorbent assay were used to determine the expression of MPS-1 in patients with endometrioma. Following the successful knockdown of MPS-1 by siRNA, CCK-8 assays, flow cytometry, and transwell assays were performed to detect ectopic endometrial stromal cells (EcESCs) proliferation, the rate of apoptosis, and cell cycle, migration, and invasion, respectively. Western blotting was used to explore the effect of MPS-1 knockdown on protein levels in the NF-κB signaling pathway. RESULTS: The expression of MPS-1 was significantly higher in endometrioma and the serum of endometrioma patients than in the patients without endometriosis. In addition, the downregulation of MPS-1 expression inhibited EcESCs proliferation, migration, and invasion. This downregulation led to the arrest of the EcESCs cycle in the G0/G1 phase and apoptosis and depressed the NF-κB signaling pathway. CONCLUSION: MPS-1 can regulate EcESCs proliferation, motility, invasion, apoptosis, and cell cycle via the NF-κB signaling pathway in endometrioma. This may contribute to the formation or development of endometriotic foci. This study suggests the potential role of MPS-1 in the pathogenesis of endometriosis and enabled further research into the use of MPS-1 in the clinical diagnosis of endometrioma.

Tea Consumption and the Risk of Endometrial Cancer: An Updated Meta-Analysis.

Although the relationship between tea consumption and the risk of endometrial cancer has been previously analyzed in certain studies, the resulting information is still conflicting, and a previous meta-analysis yielded inconsistent results. Therefore, here, we aimed to perform an updated meta-analysis of studies on this subject in order to elucidate this relationship.We searched the literature on the PubMed, Web of Science, Scopus, and Chinese National Knowledge Infrastructure databases for studies that were published prior to September 25, 2019, and all the relevant references were examined. Ultimately, we included eight studies, and seven of them were on black tea. We used the overall relative risk values (RR) and 95% confidence intervals (CI) to evaluate the risk. The synthetic RR of the eight eligible studies demonstrated that tea consumption was not relevant to the incidence rate of endometrial cancer (RR 1.06, 95% CI 0.96, 1.18). No publication bias was found. We detected significant heterogeneity among the studies (Q = 15.84, p = 0.027, I2 = 55.8%). In conclusion, the results of this meta-analysis indicate that tea consumption is not relevant to the incidence of endometrial cancer. Further research and cohort studies should be conducted to validate our result.

Identification of key genes and pathways in endometriosis by integrated expression profiles analysis.

The purpose of this study was to integrate the existing expression profile data on endometriosis (EM)-related tissues in order to identify the differentially expressed genes. In this study, three series of raw expression data were downloaded from GEO database. Differentially expressed genes (DEGs) in three tissue types were screened. GO, KEGG pathway enrichment analysis, core differential genes (CDGs) protein-protein interaction (PPI) network and weighted gene co-expression network analysis (WGCNA) were performed, finally, the dysregulation of Hippo pathway in ectopic endometrium (EC) was detected by Western blotting. A total of 1,811 DEGs between eutopic (EU) and normal endometrium (NE), 5,947 DEGs between EC and EU, and 3,192 DEGs between EC and NE datasets were identified. After screening, 394 CDGs were obtained, and 5 hub genes identified in the PPI network. CDGs enrichment and WGCNA network analysis revealed cell proliferation, differentiation, migration and other biological processes, Hippo and Wnt signaling pathways, and a variety of tumor-related pathways. Western blotting results showed that YAP/TAZ was upregulated, and MOB1, pMOB1, SAV1, LATS1 and LATS2 were downregulated in EC. Moreover, CDGs, especially the hub genes, are potential biomarkers and therapeutic targets. Finally, the Hippo pathway might play a key role in the development of endometriosis.

LncRNA MEG3-210 regulates endometrial stromal cells migration, invasion and apoptosis through p38 MAPK and PKA/SERCA2 signalling via interaction with Galectin-1 in endometriosis.

BACKGROUND: Endometriosis is a benign gynaecological disease with malignant characteristics that severely affects women's quality of life. Long noncoding RNA maternally expressed gene 3 (LncRNA MEG3) is a tumour suppressor that is downregulated in various cancer cells and tissues, and regulates multiple biological processes. Emerging studies have revealed that the interactions between MEG3 and proteins are involved in disease progression. Galectin-1 affects cell motility, signal transduction and vascularization, and is overexpressed in endometriosis. Our study is the first to explore the role of MEG3-210 transcript in endometriosis and to reveal the regulatory mechanism mediated by the interaction between MEG3-210 and Galectin-1. MATERIALS AND METHODS: Endometrial tissues and sera from patients with endometriosis and controls were collected. qRT-PCR was performed to detect the expression of MEG3-210 in the endometrium and endometrial stromal cells (ESCs). The CCK-8 assay, the Transwell assay, flow cytometry and animal models were conducted to evaluate the functions of MEG3-210 in vitro and in vivo. Bioinformatic analysis, Western blot assays, RNA-pull down assays and RNA immunoprecipitation were used to explore the potential mechanism of MEG3-210 in endometriosis. RESULTS: Our results showed that MEG3-210 expression was lower in the eutopic endometrium of women with endometriosis. MEG3-210 downregulation promoted ESCs migration, invasion, anti-apoptosis in vitro and growth of endometriotic lesions in vivo. Furthermore, MEG3-210 downregulation could activate p38 mitogen-activated protein kinase (p38 MAPK) and inhibit cAMP-dependent protein kinase A/sarcoplasmic reticulum Ca2+ ATPase 2 (PKA/SERCA2) signalling, which was mediated by Galectin-1. The protein levels of Galectin-1 in patients with endometriosis were elevated, and Galectin-1 siRNA could reduce the size of lesions. CONCLUSION: MEG3-210 regulates ESCs through p38 MAPK and PKA/SERCA signalling via interaction with Galectin-1. The novel regulatory mechanism may provide new insights into drug therapy and the diagnosis of endometriosis.

Genetic deletion of gpr27 alters acylcarnitine metabolism, insulin sensitivity, and glucose homeostasis in zebrafish.

G protein-coupled receptors (GPCRs) comprise the largest group of membrane receptors in eukaryotic genomes and collectively they regulate nearly all cellular processes. Despite the widely recognized importance of this class of proteins, many GPCRs remain understudied. G protein-coupled receptor 27 (Gpr27) is an orphan GPCR that displays high conservation during vertebrate evolution. Although, GPR27 is known to be expressed in tissues that regulate metabolism including the pancreas, skeletal muscle, and adipose tissue, its functions are poorly characterized. Therefore, to investigate the potential roles of Gpr27 in energy metabolism, we generated a whole body gpr27 knockout zebrafish line. Loss of gpr27 potentiated the elevation in glucose levels induced by pharmacological or nutritional perturbations. We next leveraged a mass spectrometry metabolite profiling platform to identify other potential metabolic functions of Gpr27. Notably, genetic deletion of gpr27 elevated medium-chain acylcarnitines, in particular C6-hexanoylcarnitine, C8-octanoylcarnitine, C9-nonanoylcarnitine, and C10-decanoylcarnitine, lipid species known to be associated with insulin resistance in humans. Concordantly, gpr27 deletion in zebrafish abrogated insulin-dependent Akt phosphorylation and glucose utilization. Finally, loss of gpr27 increased the expression of key enzymes in carnitine shuttle complex, in particular the homolog to the brain-specific isoform of CPT1C which functions as a hypothalamic energy senor. In summary, our findings shed light on the biochemical functions of Gpr27 by illuminating its role in lipid metabolism, insulin signaling, and glucose homeostasis.

CTHRC1 overexpression promotes ectopic endometrial stromal cell proliferation, migration and invasion via activation of the Wnt/ß-catenin pathway.

RESEARCH QUESTION: Endometriosis is characterized by the occurrence of endometrial-like tissue outside the uterus. Collagen triple helix repeat containing-1 (CTHRC1) is known as a tumour-promoting factor in several neoplasms. This study aimed to examine the roles of CTHRC1 in the development and progression of endometriosis, and to unravel the underlying mechanisms. DESIGN: Quantitative real-time PCR, western blot analyses and enzyme-linked immunosorbent assay were performed to determine the expression levels of CTHRC1 in tissues and serum. In addition, CTHRC1 expression levels were knocked down by small-interfering RNA in ectopic endometrial stromal cells (EESC). Cell Counting Kit-8, fluorescence-activated cell sorting, Transwell and wound scratch assays were carried out to assess the underlying biological behaviours, and western blot analyses were performed to reveal the molecular mechanisms. RESULTS: mRNA and protein expression levels of CTHRC1 were markedly higher in ectopic endometrial tissues than in eutopic and control endometrial tissues. In addition, the serum concentration of CTHRC1 was apparently higher in the endometriosis group than the control group. Small interfering RNA knockdown of CTHRC1 suppressed the proliferation, migration, invasion and healing abilities of EESC. Furthermore, the protein expressions of key molecules in the Wnt/ß-catenin pathway showed an obvious down-regulated expression after siRNA transfection. CONCLUSIONS: These findings suggest that CTHRC1 may be partly responsible for the development and progression of endometriosis by increasing EESC proliferation, migration and invasion via the Wnt/ß-catenin pathway. CTHRC1 may thus serve as a diagnostic and therapeutic target for endometriosis.

Autocrine Production of Interleukin-34 Promotes the Development of Endometriosis through CSF1R/JAK3/STAT6 signaling.

Interleukin (IL)-34 plays a critical role in cell proliferation, differentiation, apoptosis, angiogenesis, inflammation and immunoregulation. Numerous diseases can be attributed to the dysregulation of IL-34 signaling. This study was performed to investigate the function of IL-34 in the pathogenesis of endometriosis. Firstly, by enzyme linked immunoabsorbent assay, we found that IL-34, VEGF, MMP-2 and MMP-9 were increased in the sera of patients with endometriosis. Secondly, exposure to IL-34 promoted the proliferation, migration and invasion of eutopic endometrial stromal cells (ESCs). Additionally, stimulation with IL-34 up-regulated colony-stimulating factor 1 receptor (CSF1R), p-JAK3, p-STAT6, VEGF, MMP-2 and MMP-9 in these eutopic ESCs. Treatment with AS1517499, an inhibitor of STAT6, remarkably abrogated the alterations induced by IL-34. A Chromatin immunoprecipitation (ChIP) assay demonstrated binding of STAT6 to the IL-34 promoter, further implicating STAT6 in IL-34 signaling. Notably, reverse results were obtained in ectopic ESCs with the application of an IL-34 neutralizing antibody. In vivo, AS1517499 suppressed the maintenance of endometriosis lesions in rats. In summary, autocrine production of IL-34, mediated by STAT6, promoted the development of endometriosis in vitro and in vivo through the CSF1R/JAK3/STAT6 pathway. Our research reveals the function of IL-34 in endometriosis, which may provide insight into novel therapeutic strategies for endometriosis.

Roles of cell migration and invasion mediated by Twist in endometriosis.

AIM: To investigate the roles of cell migration and invasion mediated by Twist in endometriosis. METHODS: The protein levels and locations of Twist, N-cadherin and E-cadherin were measured by Western blot and immunohistochemistry in ectopic endometrium and eutopic endometrium of ovarian endometriosis as well as normal endometrium of nonendometriosis patients. The messenger RNA (mRNA) expressions of Twist, N-cadherin and E-cadherin in these tissues were measured by quantitative reverse transcription polymerase chain reaction. Stable overexpression of Twist in eutopic endometrial stromal cells was transfected with a plasmid-mediated delivery system. The protein and mRNA expressions of N-cadherin and E-cadherin were detected by western blot and reverse transcription polymerase chain reaction. The changes of migration and invasion of endometrial stromal cells were explored by transwell. RESULTS: Levels of protein and mRNA of Twist and N-cadherin showed the highest expression in ectopic endometrium of ovarian endometriosis, while lowest in normal endometrium of nonendometriosis patients. On the contrary, the expression of E-cadherin showed highest in normal endometrium of nonendometriosis patients. The overexpression of Twist after transfection significantly upregulated the protein and mRNA expression of N-cadherin, while downregulated the protein and mRNA expression of E-cadherin. There is significant difference between groups. For transwell, the overexpression of Twist in eutopic endometrial stromal cell significantly promoted cell migration and invasion. CONCLUSION: Twist might be related with the increase of migration and invasion in endometrial stromal cells, mediated by epithelial-to-mesenchymal transition.

Activation of the CXCL16/CXCR6 Axis by TNF-α Contributes to Ectopic Endometrial Stromal Cells Migration and Invasion.

The activation of systemic and local inflammatory mechanisms, including elevated levels of chemokines and proinflammatory cytokines in endometriosis progression, is becoming more evident in the recent years. Here, we report the involvement of CXC chemokine 16 (CXCL16) and its sole receptor, CXC chemokine receptor 6 (CXCR6), in pathophysiology of endometriosis. Expression of CXCL16, but not CXCR6, was significantly upregulated in endometriotic lesions when compared to control endometrium. Additionally, serum CXCL16 was significantly elevated in women with endometriosis when compared to control group. Moreover, blockade of the CXCL16/CXCR6 axis by CXCR6 small-interfering RNA reduced the migration and invasion of ectopic endometrial stromal cells (EESCs) followed by decreased phosphorylation of ERK1/2. Furthermore, TNF-α treatment induced the expression of CXCL16 in EESCs. In conclusion, these results suggest that CXCL16/CXCR6 axis, whose expression was enhanced by TNF-α, may be associated with the increased motility of EESCs, through regulation of ERK1/2 signaling, thus contributing to the development of endometriosis. These findings indicate that the CXCL16/CXCR6 axis may contribute to the progression of endometriosis and could be served as a potential target for diagnosis and treatment.

Upregulation of S100A6 in patients with endometriosis and its role in ectopic endometrial stromal cells.

S100 calcium-binding protein A6 (S100A6) is up-regulated in many malignancies and overexpression of S100A6 has been identified associated with proliferation, migration and invasion phenotype in several cancer cells. In the present study, we explored whether S100A6 plays a role in the development of endometriosis. Significantly higher levels of mRNA and protein expression of S100A6 were observed in ectopic endometrial tissues compared to eutopic and normal endometrial tissues. Silencing of S100A6 in ectopic endometrial stromal cells (ESCs) significantly inhibited cell viability, migration and invasion. Moreover, knockdown of S100A6 suppressed p38/MAPK activity in ectopic ESCs, which can be partially attenuated by CacyBP/SIP phosphorylation inhibitor. In conclusion, our results suggest that the abnormal expression of S100A6 may contribute to the pathogenesis of endometriosis and the S100A6/CacyBP/p38 signaling may provide as a promising treatment target.

Proliferation and invasion of ovarian cancer cells are suppressed by knockdown of TRIM11.

Tripartite motif-containing (TRIM)11, an E3 ubiquitin ligase, is involved in the development of the nervous system. As an oncogene, it has also been identified in glioma, lung and colon cancer. However, few studies have been conducted on TRIM11 expression and functions in ovarian cancer. In the present study, we found that TRIM11 expression was obviously elevated in ovarian cancer tissues compared to adjacent non-cancerous tissues. Depletion of TRIM11 in A2780 and SK-OV-3 ovarian cancer cells by transfection of specific small interfering RNA significantly suppressed proliferation and inhibited invasion of cells, even induced apoptosis as indicated by both Cell Counting Kit-8, Annexin V/propidium iodide staining and Transwell assay. Furthermore, we explored the underlying mechanisms. Knockdown of TRIM11 not only affects the expression of cell apoptosis-related (Bcl-2 and Bax) and invasion-related proteins [matrix metalloproteinase (MMP)-2 and MMP-9], but also reduced the phosphorylation levels of ERK and AKT. In conclusion, we showed that TRIM11 was upregulated in ovarian cancer tissue samples and that TRIM11 may serve as an oncogene in ovarian cancer.

Correlation of polypeptide N-acetylgalactosamine transferases-3 and -6 to different stages of endometriosis.

PURPOSE: To investigate the expression patterns of N-acetyl galactosamine transferases (GalNAc-Ts)-3 and GalNAc-T6 in clinicopathologically characterized endometriosis (EMS), and to explore their clinical significance. METHODS: Ectopic and eutopic endometrial tissue samples were obtained and confirmed with CD-10 immunohistochemistry in patients with EMS (n = 12), whereas normal control endometrium was obtained from patients with uterine septum (n = 12). The mRNA and protein levels of GalNAc-T3 and GalNAc-T6 were detected in these samples using quantitative real-time PCR, immunohistochemistry, and western blotting. RESULTS: GalNAc-T3 and GalNAc-T6 were expressed in the endometrium of all groups, with no significant changes observed during the menstrual cycle. The expression of GalNAc-T3 and GalNAc-T6 in ectopic endometrium was significantly lower than that in eutopic (P < 0.05) or control endometrium (P < 0.05), whereas there were no significant differences (P > 0.05) between eutopic and control endometria. Furthermore, the expression of GalNAc-T3 and GalNAc-T6 was significantly lower in patients with stage III/IV EMS compared to patients with stage I/II (P < 0.05). CONCLUSIONS: Both GalNAc-T3 and GalNAc-T6 expression levels were downregulated in ectopic endometrium, which may increase the adhesion and invasion of endometrial cells and contribute to the development of EMS. Moreover, we found a strong correlation between the expression of GalNAc-T3 and GalNAc-T6 and different stages of EMS.

Silencing of SRA1 Regulates ER Expression and Attenuates the Growth of Stromal Cells in Ovarian Endometriosis.

Estradiol and its nuclear receptors, estrogen receptor (ER) α and ER-ß, have important functions in endometriosis, and the transcriptional activity of these receptors is modulated by coactivators and corepressors. The steroid receptor RNA activator 1 (SRA1) produces SRA long noncoding RNA (lncRNA) and SRA protein (SRAP), which regulate ER expression at the RNA and protein levels in some hormone-dependent tumors via an alternative splicing event. However, only a few are reported on their expressions in endometriosis. Here, we observed that low expression levels of SRA lncRNA and ER-α but relatively high expression levels of SRAP and ER-ß were detected in ovarian endometriotic tissues versus normal endometrial tissues. Steroid receptor RNA activator 1-small interfering RNA treatment significantly increased ER-α levels but reduced ER-ß levels in endometriotic stromal cells (ESCs). Furthermore, the treatment can also attenuate the proliferation and promote early apoptosis in these cells. Our results indicate that the regulation of ER via SRA in ovarian endometriosis may play a significant role in the growth of ESCs.

Elevated phosphatase of regenerating liver 3 (PRL-3) promotes cytoskeleton reorganization, cell migration and invasion in endometrial stromal cells from endometrioma.

STUDY QUESTION: Is phosphatase of regenerating liver-3 (PRL-3) associated with increased motility of endometriotic cells from endometrioma? SUMMARY ANSWER: Elevated PRL-3 promotes cytoskeleton reorganization, cell migration and invasion of endometrial stromal cells (ESCs) from endometrioma. WHAT IS KNOWN ALREADY: Overexpression of PRL-3 is associated with cancer cell migration, invasion and metastatic phenotype. STUDY DESIGN, SIZE, DURATION: Primary human ESCs were isolated from eutopic endometrium of women without endometriosis (EuCo, n = 10), with histologically proven endometrioma (EuEM, n = 19) and from the cyst wall of ovarian endometriosis (OvEM, n = 26). PARTICIPANTS/MATERIALS, SETTING, METHODS: The expression of PRL-3 in ESCs derived from EuCo, EuEM and OvEM at different phases of menstrual cycle were compared. The protein and mRNA levels of PRL-3 were examined by western blot and RT-qPCR, respectively. ESCs from OvEM were transfected with/without short hairpin RNA (shRNA) or small interfering RNA (siRNA). Additionally, a plasmid-mediated delivery system was used to achieve PRL-3 overexpression in ESCs from EuEM. The cellular distribution of F-actin and α-tubulin were examined by immunocytochemistry. Cell motility was evaluated by a transwell migration/invasion assay. MAIN RESULTS AND THE ROLE OF CHANCE: The protein and mRNA levels of PRL-3 are significantly elevated in ESCs from OvEM compared with EuCo and EuEM. The expression of PRL-3 was not altered between proliferative phase and secretory phase in ESCs from all groups. Knockdown of PRL-3 significantly modified the distribution of F-actin and α-tubulin cytoskeleton, inhibited cell migration and invasion. Endogenous inhibition of PRL-3 attenuated the expression of Ras homolog gene family members A and C (RhoA, RhoC), Rho-associated coiled-coil-containing protein kinase 1 (ROCK1) and matrix metalloproteinase (MMP) 9, but not MMP2 in ESCs from OvEM. Additionally, overexpression of PRL-3 in ESCs from EuEM up-regulates cell migration and invasion, and increases the expression of RhoA, RhoC, ROCK1 and MMP9. LIMITATIONS, REASONS FOR CAUTION: Lack of in vivo animal studies is the major limitation of our report. Our results should be further confirmed in a larger cohort of patients and extended to include eutopic and ectopic endometrium from patients with peritoneal endometriosis at different stages of the disease. WIDER IMPLICATIONS OF THE FINDINGS: Our study describes that elevated expression of PRL-3 contributes to the cell motility of ESCs from endometrioma. The results emphasize the importance of metastatic-related factor PRL-3 in the pathogenesis of endometrioma. STUDY FUNDING/COMPETING INTEREST: This work was supported by National Natural Science Foundation of China (No. 81170546) and Zhejiang Medicine Science and Technology Projects (No. Y13H040003). The authors declare no conflict of interest.