Long noncoding RNA BMPR1B-AS1 stability regulated by IGF2BP2 affects the decidualization in endometriosis patients through the SMAD1/5/9 pathway.

Endometriosis (EMs)-related infertility commonly has decreased endometrial receptivity and normal decidualization is the basis for establishing and maintaining endometrial receptivity. However, the potential molecular regulatory mechanisms of impaired endometrial decidualization in patients with EMs have not been fully clarified. We confirmed the existence of reduced endometrial receptivity in patients with EMs by scanning electron microscopy and quantitative real-time PCR. Here we identified an lncRNA, named BMPR1B-AS1, which is significantly downregulated in eutopic endometrium in EMs patients and plays an essential role in decidual formation. Furthermore, RNA pull-down, mass spectrometry, RNA immunoprecipitation, and rescue analyses revealed that BMPR1B-AS1 positively regulates decidual formation through interaction with the RNA-binding protein insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2). Downregulation of IGF2BP2 led to a decreased stability of BMPR1B-AS1 and inhibition of activation of the SMAD1/5/9 pathway, an inhibitory effect which diminished decidualization in human endometrial stromal cells (hESCs) decidualization. In conclusion, our identified a novel regulatory mechanism in which the IGF2BP2-BMPR1B-AS1-SMAD1/5/9 axis plays a key role in the regulation of decidualization, providing insights into the potential link between abnormal decidualization and infertility in patients with EMs, which will be of clinical significance for the management and treatment of infertility in patients with EMs.

Carbohydrate-DNA Conjugation Enabled by Glycosyl Radicals Generated from Glycosyl Sulfinates.

Herein, we report a method that enables the synthesis of carbohydrate-DNA conjugates by radical addition. Key to the success is the use of readily available, bench-stable, and unprotected glycosyl sulfinates as precursors to glycosyl radicals. The redox neutral reaction proceeds under mild and simple conditions and tolerates a broad substrate scope. A small library of carbohydrate-DNA conjugates was prepared.

Effect of sequential versus single-step culture medium on IVF treatments, including embryo and clinical outcomes: a prospective randomized study.

PURPOSE: Sequential media G5 series (Vitrolife) and single-step medium Continuous Single Culture Complete (CSC-C) (Irvine Scientific) are two different culture media. We want to examine difference between culturing effects of the two media. METHODS: To compare the fertilization and early embryo development, a prospective randomized controlled trial with sibling oocytes in infertile patients, aged ≤ 45 years with ≥ 8 oocytes (226 cycles) was conducted. Each half of the retrieved oocytes from the same patient were randomly allocated to two culture media separately. The remaining fresh cycles were randomly assigned to two culture media during the same period (179 cycles). We compared the clinical outcomes based on the total fresh ET cycles in this periods, in which the transferred embryos were only from one culture medium. RESULTS: Embryo outcomes: 226 cycles, included 176 IVF and 50 ICSI cycles, were analyzed, which correspond to 3518 inseminated or micro-injected oocytes. CLINICAL OUTCOMES: 71 (CSC-C) and 71 (G5 series) fresh ET cycles were compared. There were no significant differences in clinical outcomes and general fertilization rate. However, the fertilization rate was superior in the CSC-C when compared with G5 in ICSI cycles (76.51% vs. 67.25%, P = 0.008). In addition, the compacted embryo development rate was significantly higher in CSC-C on day 3. The cycles that had compacted embryos on day 3 demonstrated better outcomes both in embryos as well as clinically. CONCLUSIONS: CSC-C had higher fertilization rates than G5 series in ICSI cycles. In addition, the compaction rates of day 3 embryos were significantly higher in CSC-C.

Biallelic mutations in spermatogenesis and centriole-associated 1 like (SPATC1L) cause acephalic spermatozoa syndrome and male infertility.

Acephalic spermatozoa syndrome is a rare type of teratozoospermia that severely impairs the reproductive ability of male patients, and genetic defects have been recognized as the main cause of acephalic spermatozoa syndrome. Spermatogenesis and centriole-associated 1 like (SPATC1L) is indispensable for maintaining the integrity of sperm head-to-tail connections in mice, but its roles in human sperm and early embryonic development remain largely unknown. Herein, we conducted whole-exome sequencing (WES) of 22 infertile men with acephalic spermatozoa syndrome. An in silico analysis of the candidate variants was conducted, and WES data analysis was performed using another cohort consisting of 34 patients with acephalic spermatozoa syndrome and 25 control subjects with proven fertility. We identified biallelic mutations in SPATC1L (c.910C>T:p.Arg304Cys and c.994G>T:p.Glu332X) from a patient whose sperm displayed complete acephalia. Both SPATC1L variants are rare and deleterious. SPATC1L is mainly expressed at the head-tail junction of elongating spermatids. Plasmids containing pathogenic variants decreased the level of SPATC1L in vitro. Moreover, none of the patient's four attempts at intracytoplasmic sperm injection (ICSI) resulted in a transplantable embryo, which suggests that SPATC1L defects might affect early embryonic development. In conclusion, this study provides the first identification of SPATC1L as a novel gene for human acephalic spermatozoa syndrome. Furthermore, WES might be applied for patients with acephalic spermatozoa syndrome who exhibit reiterative ICSI failures.

Construction of Competing Endogenous RNA Networks Incorporating Transcription Factors to Reveal Differences in Granulosa Cells from Patients with Endometriosis.

Purpose: This study aimed to reveal the molecular differences in granulosa cells (GCs) from patients with endometriosis (EM). Methods: RNA sequencing was performed on GCs from patients with EM-related infertility (n = 3) and controls (n = 3). Differentially expressed long noncoding RNAs [differentially expressed lncRNAs (DELs), |log2 FC|>4, false discovery rate (FDR) <0.05] and genes [differentially expressed genes (DEGs), |log2 FC|>1.4, FDR <0.05] in patients with EM-related infertility and controls were screened. Protein-protein interaction (PPI) networks of the DEGs were constructed. Then, mRNA-miRNA-lncRNA pairs based on DEGs and DELs were constructed by comprehensive bioinformatics analyses. In addition, overlapping genes identified from both the PPI and mRNA-miRNA-lncRNA pairs were selected. Finally, a competing endogenous RNA (ceRNA) network incorporating transcription factors (TFs) was constructed. Results: A total of 25,806 lncRNAs and 19,684 mRNAs were detected, and 7 DELs and 46 DEGs were identified. Five hub genes from the PPI network were also identified. A single overlapping gene, NR4A2, from both the PPI network and mRNA-miRNA-lncRNA pairs was identified. Finally, a ceRNA network incorporating TFs, including one mRNA (NR4A2), one miRNA (hsa-miR-217), three lncRNAs (XIST, MCM3AP-AS1, and C17orf51), and five TFs (SRF, POLR2A, NRF1, MNT, and TCF7L2), was successfully constructed. Conclusions: The proposed ceRNA network and the prediction of TFs in GCs from EM-related infertility revealed differences in GCs from patients with EM. Importantly, the novel TFs, lncRNAs, miRNAs, and mRNAs involved in the ceRNA network might provide new insights into the underlying molecular mechanisms of EM-related infertility.

Functionalization of DNA-Tagged Alkenes Enabled by Visible-Light-Induced C-H Activation of N-Aryl Tertiary Amines.

A highly efficient approach to C(sp3)-C(sp3) bond construction via on-DNA photoredox catalysis between on-DNA alkenes and N-aryl tertiary amines was developed. The methodology demonstrated 55%-95% conversions without obvious DNA damage, as seen by qPCR tests. Furthermore, various functional groups, such as carboxylic acids, aldehydes, and aryl halides, that can be used to create library diversities were shown to be tolerant of the C-H activation conditions.

Establishment of an immortalized stromal cell line derived from human Endometriotic lesion.

BACKGROUND: Endometriosis is a benign gynecological disease with obviously feature of estrogen-dependence and inflammatory response. The applications of primary endometriotic stromal cells in research of endometriosis are restricted for short life span, dedifferentiation of hormone and cytokine responsiveness. The objective of this study was to establish and characterize immortalized human endometriotic stromal cells (ihESCs). METHODS: The endometriotic samples were from a patient with ovarian endometriosis and the primary endometriotic stromal cells were isolated from the endometriotic tissues. The primary cells were infected by lentivirus to establish telomerase reverse transcriptase (hTERT)-induced immortalized cells. Quantification of mRNA and proteins was examined by quantitative real-time polymerase chain reaction (qRT-PCR) and Western Blot. CCK-8 assay and EdU labeling assay were assigned to assess the growth of ihESCs. Karyotype assay was performed to detect the chromosomes of ihESCs. Colony formation assay and nude mouse tumorigenicity assay were used to evaluate colony-formation and tumorigenesis abilities. RESULTS: ihESCs continuously overexpressed hTERT via infection of lentivirus and significant extended the life span reaching 31 passages. The morphology, proliferation and karyotype of ihESCs remained unchanged. The expression of epithelial-mesenchymal transition (EMT) markers, estrogen-metabolizing proteins and estrogen/progesterone receptors (ERs and PRs) were unaltered. Furthermore, the treatment of estrogen increased the proliferation and EMT of ihESCs. Lipopolysaccharides (LPS) and IL-1ß remarkably induced inflammatory response. The clonogenesis ability of ihESCs was consistent with primary cells, which were much lower than Ishikawa cells. In addition, nude mouse tumorigenicity assay demonstrated that ihESCs were unable to trigger tumor formation. CONCLUSION: This study established and characterized an immortalized endometriotic stromal cell line that exhibited longer life span and kept the cellular morphology and physiological function as the primary cells. The immortalized cells remained normal feedback to estrogen and inflammatory response. Moreover, the immortalized cells were not available with tumorigenic ability. Therefore, ihESCs would be serviceable as in vitro cell tool to investigate the pathogenesis of endometriosis.

Circular RNA expression profiling and bioinformatic analysis of cumulus cells in endometriosis infertility patients.

Aim: To explore the circular RNA (circRNA) profile in cumulus cells from endometriosis-associated infertility patients. Methods: The expression of circRNAs was profiled by high-throughput sequencing. Sanger sequencing was performed to identify the backsplicing site. Six candidate circRNAs and their parental genes were measured in 30 samples by quantitative reverse transcription-polymerase chainreaction (qRT-PCR). Bioinformatics analysis was performed to predict the functions. Results: A total of 55 upregulated and 41 downregulated differentially expressed circRNAs were detected. Kyoto Encyclopedia of Genes and Genomes data indicated that these target genes were mainly involved in cumulus cell growth- and differentiation-related pathways. Hsa_circ_0072391, hsa_circ_0007299 and hsa_circ_0057799 were significantly increased, and hsa_circ_001533 was significantly decreased in endometriosis-associated infertility patients. Conclusion: The differentially expressed circRNAs might be potentially involved in pathophysiology of endometriosis-associated infertility.

Exploring Aldol Reactions on DNA and Applications to Produce Diverse Structures: An Example of Expanding Chemical Space of DNA-Encoded Compounds by Diversity-Oriented Synthesis.

A DNA-encoded chemical library (DECL) is built with combinatorial chemistry, which works by bringing chemical fragments together to generate diverse structures. However, chemical diversity of DNA-encoded chemical libraries is often limited by DNA compatible synthetic reactions. This report shows a conceptual strategy to expand chemical space of DNA-encoded chemical libraries by incorporation of diversity-oriented synthesis in DECL synthesis. We developed Aldol reactions on DNA in a combinatorial way. After obtaining DNA-tagged α, ß-unsaturated ketones which represent important chemical intermediates, many distinct structures with skeletal diversities are achieved by diversity-oriented synthesis.

RhoA/ROCK pathway mediates the effect of oestrogen on regulating epithelial-mesenchymal transition and proliferation in endometriosis.

Endometriosis is a benign gynaecological disease appearing with pelvic pain, rising dysmenorrhoea and infertility seriously impacting on 10% of reproductive-age females. This research attempts to demonstrate the function and molecular mechanism of RhoA/ROCK pathway on epithelial-mesenchymal transition (EMT) and proliferation in endometriosis. The expression of Rho family was abnormally changed in endometriotic lesions; in particular, RhoA and ROCK1/2 were significantly elevated. Overexpression of RhoA in human eutopic endometrial epithelial cells (eutopic EECs) enhanced the cell mobility, epithelial-mesenchymal transition (EMT) and proliferation, and RhoA knockdown exhibited the opposite function. Oestrogen up-regulated the RhoA activity and expression of RhoA and ROCK1/2. RhoA overexpression reinforced the effect of oestrogen on promoting EMT and proliferation, and RhoA knockdown impaired the effect of oestrogen. oestrogen receptor α (ERα) was involved with the regulation of oestrogen on EMT and proliferation and up-regulated RhoA activity and expression of RhoA and ROCK1/2. The function of ERα was modulated by the change in RhoA expression. Furthermore, phosphorylated ERK that was enhanced by oestrogen and ERα promoted the protein expression of RhoA/ROCK pathway. Endometriosis mouse model revealed that oestrogen enhanced the size and weight of endometriotic lesions. The expression of RhoA and phosphorylated ERK in mouse endometriotic lesions was significantly elevated by oestrogen. We conclude that abnormal activated RhoA/ROCK pathway in endometriosis is responsible for the function of oestrogen/ERα/ERK signalling, which promoted EMT and proliferation and resulted in the development of endometriosis.

Identification of a novel deletion mutation in DPY19L2 from an infertile patient with globozoospermia: a case report.

BACKGROUND: Male infertility is an increasing medical concern worldwide. In most cases, genetic factors are considered as the main cause of the disease. Globozoospermia (MIM102530) (also known as round-headed sperm) is a rare and severe malformed spermatospermia caused by acrosome deficiency or severe malformation. A subset of genetic mutations, such as DNAH6, SPATA16, DPY19L2, PICK1, and CCIN related to globozoospermia, have been reported in the past few years. The DPY19L2 mutation is commonly found in patients with globozoospermia. Herein, a 180-kbp homozygote deletion at 12q14.2 (g.63950001-64130000) was identified by copy number variation sequencing (CNVseq) in a patient with a globozoospermia, including the complete deletion of DPY19L2. CASE PRESENTATION: A 27-year-old patient at the First Affiliated Hospital of Xiamen University was diagnosed with infertility because, despite normal sexual activity for 4 years, his wife did not conceive. The patient was in good health with no obvious discomfort, no history of adverse chemical exposure, and no vices, such as smoking and drinking. The physical examination revealed normal genital development. However, semen tests showed a normal sperm count of 0% and the morphology was the round head. Sperm cytology showed that acrosomal enzyme was lower than normal. Reproductive hormones were in the normal range. B ultrasound did not show any abnormal seminal vesicle, prostate, bilateral testis, epididymis, and spermatic veins. The karyotype was normal, 46, XY, and no microdeletion of Y chromosome was detected. However, a homozygous deletion mutation was found in DPY19L2, which was further diagnosed as globozoospermia. CONCLUSIONS: The present study reported a male infertility patient who was diagnosed with globozoospermia. The analysis of gene mutations revealed that DPY19L2 had a homozygous mutation, which was the primary cause of globozoospermia.

Loss-of-function mutation in TSGA10 causes acephalic spermatozoa phenotype in human.

BACKGROUND: Acephalic spermatozoa is an extremely rare type of teratozoospermia that is associated with male infertility. Several genes have been reported to be relevant to acephalic spermatozoa. Thus, more genetic pathogenesis needs to be explored. METHODS: Whole-exome sequencing was performed in a patient with acephalic spermatozoa. Then Sanger sequencing was used for validation in the patient and his family. The patient's spermatozoa sample was observed by papanicolaou staining and transmission electron microscopy. Western blot and immunofluorescence were performed to detect the level and localization of related proteins. RESULTS: A novel homozygous frameshift insertion mutation c.545dupT;p.Ala183Serfs*10 in exon 8 of TSGA10 (NM_001349012.1) was identified. Our results showed misarranged mitochondrial sheath and abnormal flagellum in the patient's spermatozoa. TSGA10 failed to be detected in the patient's spermatozoa. However, the expression of SUN5 and PMFBP1 remained unaffected. CONCLUSION: These results suggest that the novel homozygous frameshift insertion mutation of TSGA10 is a cause of acephalic spermatozoa.

Genetic analysis of embryo in a human case of spontaneous oocyte activation: a case report.

Parthenogenesis, a unique form of reproduction, is normally inhibited in mammals and a human embryo with parthenogenetic origin is not considered capable of producing offspring. The aim of this report is to analyze a parthenogenetic oocyte retrieved from a patient so as to have a better understanding on parthenogenesis and causes of infertility. A 38-year-old woman presented at our center with a history of primary infertility for 10 years and underwent an IVF-ICSI cycle. Three MII oocytes retrieved and one of which presented with 1 pronucleus before conducting ICSI and developed into an embryo 30 h post-retrieval. Blastomere biopsy, genome amplification, copy number variation (CNV) analysis and MultiSNPs analysis was performed on the embryo. The results showed that only one blastomere contains DNA and CNV analysis indicated a genotype of 48, XX, +17, +17 and the genetic contribution of biopsied embryo was of exclusively maternal origin. Such analysis might be beneficial for patients with a history of oocyte spontaneous activation in diagnosing case-specific aberrations and providing individualized therapeutic strategies such as preimplantation genetic diagnosis to choose a genetic normal embryo to transplant.

Lipoxin A4 Suppresses IL-1ß-Induced Cyclooxygenase-2 Expression Through Inhibition of p38 MAPK Activation in Endometriosis.

Endometriosis is an inflammation-dependent gynecologic disorder. Increased cyclooxygenase-2 (COX-2) expression plays an important role in the development and progression of endometriosis. Lipoxin A4 (LXA4) is an endogenous anti-inflammation lipid and showed inhibitory effects on the development of endometriosis; however, the mechanism remains unclear. In this study, the overexpression of COX-2 was observed in ectopic endometrium of endometriosis patients compared to the normal endometrium of controls. Lipoxin A4 efficiently suppressed IL-1ß-induced COX-2 protein expression in ectopic endometriotic stromal cells (ESCs) via its receptor, formyl peptide receptor 2/lipoxin A4 receptor (FPR2/ALX). Antagonism of FPR2/ALX eliminated the inhibitory effect by LXA4. IL-1ß induced the activation of mitogen-activated protein kinases (MAPKs), which can promote the expression of COX-2. Pretreatment of ESCs with LXA4 inhibited the phosphorylation of p38 MAPK induced by IL-1ß. These findings suggest that inflammation and MAPKs pathways respond for the abnormal expression of COX-2, which can elucidate the pathophysiology of endometriosis. Moreover, LXA4 suppressed IL-1ß-induced COX-2 expression through inhibiting the p38 MAPK signaling protein. This research contributes for better understanding of the cellular and biological events of inflammation and anti-inflammation-mediated regulation in endometriosis.

Long non-coding RNA AFAP1-AS1 promoting epithelial-mesenchymal transition of endometriosis is correlated with transcription factor ZEB1.

PROBLEM: The role of the long non-coding RNA (lncRNA) actin filament associated protein 1 antisense RNA1 (AFAP1-AS1) in the etiology of endometriosis is unknown. METHOD OF STUDY: Expression of epithelial-mesenchymal transition (EMT) markers was quantified using qRT-PCR, immunohistochemistry, and Western blotting. The proliferation, migration, and invasion of ectopic endometrial epithelial cells and Ishikawa cells were evaluated by MTT, EdU, wound healing, and transwell assays. Inflammatory cytokine levels were detected by ELISA. Luciferase assays were used to measure activity of the ZEB1 promoter site pGL3-P886. RESULTS: AFAP1-AS1 levels were much higher in ectopic endometrial tissues than that in eutopic tissues. Expression of ZEB1, E-cadherin, and keratin was obviously higher in eutopic tissues than those in ectopic tissues. In contrast, expression of vimentin and N-cadherin was significantly lower in eutopic tissue than those in ectopic tissues. After knockdown of AFAP1-AS1, the morphology of endometrial epithelial cells varied from spindle fiber shaped to polygon epithelioid and proliferation, migration, and invasion were each inhibited. The knockdown of AFAP1-AS1 significantly inhibited expression from promoter site pGL3-P886 of the EMT-related transcription factor ZEB1. The size of subcutaneous tumours in nude mice was significantly reduced after down-regulation of AFAP1-AS1 expression. CONCLUSION: Higher expression of AFAP1-AS1 positively correlated with greater EMT in ectopic endometrium of patients with endometriosis. Knockdown of AFAP1-AS1 inhibited E2-induced activity of promoter site pGL3-P886 of transcription factor ZTB1, suggesting that AFAP1-AS1 knockdown inhibited growth of endometrial epithelial cells and that pathogenesis may be correlated with EMT.

High expression of ZEB1 in endometriosis and its role in 17ß-estradiol-induced epithelial-mesenchymal transition.

Endometriosis is an estrogen-dependent disease associated with pain and infertility. The objective of this study was to determine the expression of ZEB1 in endometriosis and its role in 17ß-estradiol (E2)-induced epithelial-mesenchymal transition (EMT). 25 patients with endometriosis and 16 endometriosis-free patients were recruited for the study. Tissue expression of EMT makers was investigated by immunohistochemistry, then the expression of ZEB1 was quantified by qRT-PCR, immunohistochemistry, and western blot. The proliferation, DNA replication, and migration and invasion in ZEB1 knockdown Ishikawa cells were further respectively performed by MTS, Edu, wound healing and transwell assays. Luciferase assay was used to measure the ZEB1 promoter activity. Our results show that protein levels of E-cadherin and Keratin 18 decreased in endometriotic tissues. Meanwhile the expressions of ZEB1, Vimentin, and N-cadherin were significantly increased in endometriotic tissues. Down-regulation of ZEB1 inhibited Ishikawa cells proliferation, migration, invasion and EMT. E2 promoted the expression of ZEB1 through the ER genomic pathway, which contributed to the EMT process. The -1401 bp - -1901 bp region in the ZEB1 promoter was the main target of the E2 activity. The present results suggest that a high expression of ZEB1 plays an important role in the pathogenesis of endometriosis, and it may serve as a potential therapeutic target for endometriosis.

Lipoxin A4 Suppresses Estrogen-Induced Epithelial-Mesenchymal Transition via ALXR-Dependent Manner in Endometriosis.

OBJECTIVE: Epithelial-mesenchymal transition (EMT) is essential for embryogenesis, fibrosis, and tumor metastasis. Aberrant EMT phenomenon has been reported in endometriotic tissues of patients with endometriosis (EM). In this study, we further investigated the molecular mechanism of which lipoxin A4 (LXA4) suppresses estrogen (E2)-induced EMT in EM. STUDY DESIGN: The EMT markers were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot in eutopic endometrial epithelial cells (EECs) or investigated by immunohistochemistry and qRT-PCR in endometriotic lesion of EM mice. The invasion and migration under different treatments were assessed by transwell assays with or without Matrigel. The messenger RNA (mRNA) and activities of matrix metalloproteinase 2 (MMP-2) and MMP-9 were determined by qRT-PCR and gelatin zymography, respectively. Luciferase reporter assay was used to measure the activity of zinc finger E-box binding homeobox 1(ZEB1) promoter. The level of E2 in endometriotic tissues was assessed by enzyme-linked immunosorbent assay. RESULTS: In eutopic EECs, stimulatory effects of E2 on EMT progress, migration, and invasion were all diminished by LXA4. Lipoxin A4 reduced E2-induced ZEB1 promoter activity. Lipoxin A4 also attenuated the phosphorylation of extracellular signal-regulated kinase and p38 mitogen-activated protein kinase induced by E2. Co-incubation with Boc-2 rather than DMF antagonized the influence of LXA4. Animal experiments showed that LXA4 inhibited the EMT progress, MMP expression, and proteinase activities of endometriotic lesion in an LXA4 receptor (ALXR) manner, which suppressed the progression of EM. ZEB1 mRNA expression was upregulated and well correlated with E2 level in human endometrium. CONCLUSION: Lipoxin A4 suppresses E2-induced EMT via ALXR-dependent manner in eutopic EECs, which reveals a novel biological effect of LXA4 in EM.

MicroRNA-488 and -920 regulate the production of proinflammatory cytokines in acute gouty arthritis.

BACKGROUND: Gout is considered one of the most painful acute conditions caused by deposition of monosodium urate (MSU) crystals within joints. Recent studies have shown that interleukin (IL)-1ß is a key inflammatory mediator in acute gouty arthritis (GA), and its level is regulated by microRNAs (miRNAs). However, the molecular mechanisms of the regulation remain unclear. METHODS: A miRNA microarray was used to analyze the miRNA expression profiles in peripheral white blood cells (WBCs) of patients with GA. THP-1 cells were transfected with miRNA mimics, stimulated by MSU crystals, and then subjected to quantitative real-time polymerase chain reaction or Western blot analysis. Levels of IL-1ß, IL-8, and tumor necrosis factor (TNF)-α in culture supernatants of THP-1 cells were measured by enzyme-linked immunosorbent assay. A luciferase reporter assay was conducted to confirm the interaction of miRNA and IL-1ß 3'-untranslated regions (UTRs). RESULTS: Combining bioinformatics and miRNA expression profiles, we found five miRNAs (hsa-miR-30c-1-3p, hsa-miR-488-3p, hsa-miR-550a-3p, hsa-miR-663a, and hsa-miR-920) that possibly target IL-1ß. Then, we demonstrated that miR-488 and miR-920 were significantly decreased in the WBCs of patients with GA and that MSU crystals could inhibit expression of miR-488 and miR-920. Upregulation of miR-488 and miR-920 could suppress MSU-induced IL-1ß protein expression in THP-1 cells, but no significant difference in IL-1ß messenger RNA levels was observed. Moreover, we found that miR-488 and miR-920 could directly target the 3'-UTR of IL-1ß. Overexpression of miR-488 and miR-920 could significantly inhibit the gene and protein expression of IL-8 and TNF-α in MSU-induced THP-1 cells. CONCLUSIONS: This study demonstrates the roles of miR-488 and miR-920 in regulating the production of proinflammatory cytokines in the pathogenesis of GA. These findings suggest that miR-488 and miR-920 could serve as potential therapeutic targets in the treatment of GA.

Effect of interleukin-1ß and lipoxin A4 in human endometriotic stromal cells: Proteomic analysis.

AIM: Lipoxin A4 (LXA4 ) can function as an endogenous 'breaking signal' in inflammation and plays an important role in the progression of endometriosis. The proteome responses to interleukin-1ß (IL-1ß) or LXA4 in human endometriotic stromal cells (ESC) are not well understood. METHODS: In this study, primary ESC were cultured from ovarian endometriosis tissue. Three groups were established: the control group; the IL-1ß stimulation group; and the IL-1ß and LXA4 incubation group. Proteins were assessed on 2-D polyacrylamide gel electrophoresis (2D-PAGE), and differentially expressed protein spots were further identified on matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MALDI-TOF-MS). Wound healing and transwell assays were performed to assess the migration and invasion of ESC after treatment. RESULTS: In total, 40 differentially expressed protein spots were identified successfully on MALDI-TOF-MS. The proteins identified were related to cell structure, metabolism, signal transduction, protein synthesis and membrane structure, processes that may be involved in the development of endometriosis. Vinculin and IL-4 were further analyzed on western blot and quantitative real-time polymerase chain reaction. Moreover, LXA4 could suppress the migration and invasion of ESC induced by IL-1ß. CONCLUSION: LXA4 may inhibit the progression of endometriosis partly by lowering or raising the effect of IL-1ß, mediated via some inflammation-related proteins (e.g. vinculin) and immune response-related protein (e.g. IL-4) in vitro.

ZEB1 promotes epithelial-mesenchymal transition in cervical cancer metastasis.

OBJECTIVE: To investigate role of Zinc finger E-box binding homeobox 1 (ZEB1) in cervical cancer tissue (squamous cell carcinoma, SCC). DESIGN: Exploratory study. SETTING: University hospital. PATIENT(S): Sixty patients with SCC, including stage CINIII (n = 10), IB1 (n = 10), IB2 (n = 10), IIA1 (n = 10), IIA2 (n = 10), and IIB (n = 10) were studied. INTERVENTION(S): Caski cells were transfected with recombinant shZEB1 lentivirus or shCtrl lentivirus to generate stable ZEB1-knockdown Caski cells. MAIN OUTCOME MEASURE(S): ZEB1 expression was analyzed by quantitative real-time polymerase chain reaction and immunohistochemistry in cervical cancer tissues. ZEB1 expression in Caski cells was down-regulated by short-hairpin RNA (shRNA) interference, and changes in ZEB1 expression corresponded with changes in the proliferation and migratory ability of Caski cells. RESULT(S): Quantitative real-time polymerase chain reaction and immunohistochemistry results revealed that ZEB1 expression and the ratio of Vimentin to E-cadherin were high in 27 of 50 SCC patients and correlated with advanced International Federation of Gynecology and Obstetrics stage, tumor size >4 cm, and parametrial invasion. However, the expression of ZEB1 in cervical cancer tissue was independent of age and SCC antigen level. Transfection of ZEB1 shRNA in Caski cells significantly decreased the messenger RNA and protein expression of ZEB1, parallel with increased expression of the epithelial marker E-cadherin and decreased expression of the mesenchymal marker Vimentin. Furthermore, the proliferation and migratory ability of Caski cells were significantly lower in the transfected group than in the nontransfected control group. CONCLUSION(S): Down-regulation of ZEB1 expression may protect the invasive front of the tumors from converting to a mesenchymal phenotype by reducing the proliferation and motility of cervical cancer cells, suggesting that ZEB1 might be a potential therapeutic target for SCC.