Identification of microRNAs involved in polycystic ovary syndrome based on High-throughput sequencing [miRNA-seq]
ABSTRACT: Aberration in miRNA expression or DNA methylation is a causal factor for numerous pathological conditions including polycystic ovarian syndrome PCOS, a common endocrine disorders and leading cause of infertility. The epigenetic interactions between miRNA and DNA methylation remain unexplored in PCOS. Few studies have reported that the regulation of miRNAs in polycystic ovary syndrome(PCOS). Our study helps to understand the molecular pathogenesis of PCOS in human ovarian granulosa cells from the perspective of post-transcriptional level. Overall design: In this study, we applied small RNA sequencing in ovarian granulosa cells of PCOS patients and normal people.
Project description:Aberration in miRNA expression or DNA methylation is a causal factor for numerous pathological conditions including polycystic ovarian syndrome PCOS, a common endocrine disorders and leading cause of infertility. The epigenetic interactions between miRNA and DNA methylation remain unexplored in PCOS. Our study identifies epigenetic alternation in ovarian granulosa cells from PCOS patients and helps to reveal the pathogenesis of PCOS. Overall design: In this study, we applied MBD sequencing in ovarian granulosa cells of PCOS patients and normal people.
Project description:Aberration in miRNA expression or DNA methylation is a causal factor for numerous pathological conditions including polycystic ovarian syndrome PCOS, a common endocrine disorders and leading cause of infertility. The epigenetic interactions between miRNA and DNA methylation remain unexplored in PCOS. Few studies have reported that the key genes and pathways involved in polycystic ovary syndrome(PCOS). Our study helps to understand the molecular pathogenesis of PCOS in human ovarian granulosa cells and identifiy genes and pathways that may be potential therapeutic targets for PCOS treatment.. Overall design: In this study, we applied RNA sequencing in ovarian granulosa cells of PCOS patients and normal people.
Project description:Polycystic ovary syndrome (PCOS) affects approximately 7% of the reproductive-age women. A growing body of evidence indicated that epigenetic mechanisms contributed to the development of PCOS. The role of DNA modification in human PCOS ovary granulosa cell is still unknown in PCOS progression. Global DNA methylation and hydroxymethylation were detected between PCOS' and controls' granulosa cell. Genome-wide DNA methylation was profiled to investigate the putative function of DNA methylaiton. Selected genes expressions were analyzed between PCOS' and controls' granulosa cell. Our results showed that the granulosa cell global DNA methylation of PCOS patients was significant higher than the controls'. The global DNA hydroxymethylation showed low level and no statistical difference between PCOS and control. 6936 differentially methylated CpG sites were identified between control and PCOS-obesity. 12245 differential methylated CpG sites were detected between control and PCOS-nonobesity group. 5202 methylated CpG sites were significantly differential between PCOS-obesity and PCOS-nonobesity group. Our results showed that DNA methylation not hydroxymethylation altered genome-wide in PCOS granulosa cell. The different methylation genes were enriched in development protein, transcription factor activity, alternative splicing, sequence-specific DNA binding and embryonic morphogenesis. YWHAQ, NCF2, DHRS9 and SCNA were up-regulation in PCOS-obesity patients with no significance different between control and PCOS-nonobesity patients, which may be activated by lower DNA methylaiton. Global and genome-wide DNA methylation alteration may contribute to different genes expression and PCOS clinical pathology.
Project description:Polycystic Ovary Syndrome (PCOS) is the most common endocrine disorder amongst women of reproductive age, whose aetiology remains unclear. To improve our understanding of the molecular mechanisms underlying the disease, we conducted a genome-wide DNA methylation profiling in granulosa lutein cells collected from 16 women suffering from PCOS, in comparison to 16 healthy controls. Samples were collected by follicular aspiration during routine egg collection for IVF treatment. Study groups were matched for age and BMI, did not suffer from other disease and were not taking confounding medication. Comparing women with polycystic versus normal ovarian morphology, after correcting for multiple comparisons, we identified 106 differentially methylated CpG sites with p-values <5.8?×?10<sup>-8</sup> that were associated with 88 genes, several of which are known to relate either to PCOS or to ovarian function. Replication and validation of the experiment was done using pyrosequencing to analyse six of the identified differentially methylated sites. Pathway analysis indicated potential disruption in canonical pathways and gene networks that are, amongst other, associated with cancer, cardiogenesis, Hedgehog signalling and immune response. In conclusion, these novel findings indicate that women with PCOS display epigenetic changes in ovarian granulosa cells that may be associated with the heterogeneity of the disorder.
Project description:Polycystic ovary syndrome (PCOS) is recognized as a general endocrine disease and reproductive disorder. Although evidence indicates that PCOS has a complex etiology and genetic basis, the pathogenic mechanisms and signal pathway in PCOS remain unclear. In this study, the normal structure of follicle and corpus luteum were observed, and no cyst nor hyperemia was observed under the light microscopic study with hematoxylin and eosin (H&E) staining. Eestosterone and progesterone were evaluated by radioimmunoassay in rat serum. The alterations of proliferative ability and cell cycle distribution of each group were assessed by Cell Counting Kit-8 (CCK8) assay and flow cytometry. The protein expression of p-mTOR/mTOR, p-PI3K/PI3K, p-AKT/AKT, and GAPDH were analyzed by western blotting. Both doses of PLB could benefit the ovarian morphology and polycystic property. PLBinduced a suppress effect on the proliferation of rat ovarian granulosa cells. In addition, PLB also induced concentration-dependent apoptosis in rat ovarian granulosa cells. The rat ovarian granulosa cells treated with PLB that the expression levels of p-AKT, p-mTOR, and p-PI3K were significantly decreased in a concentration-dependent manner. PLB not only plays a critical role in attenuating the pathology and polycystic property changes in the ovary but can also induce rat ovarian granulosa cell apoptosis through the PI3K/Akt/mTOR signal pathway. This study showed the innovative role of PLB in the pathogenesis of PCOS and provides a new therapeutic modality for the treatment of PCOS.
Project description:BACKGROUND:Women with polycystic ovary syndrome (PCOS) manifest a host of ovarian defects like impaired folliculogenesis, anovulation, and poor oocyte quality, which grossly affect their reproductive health. Addressing the putative epigenetic anomalies that tightly regulate these events is of foremost importance in this disorder. We therefore aimed to carry out DNA methylome profiling of cumulus granulosa cells and assess the methylation and transcript expression profiles of a few differentially methylated genes contributing to ovarian defects in PCOS. A total of 20 controls and 20 women with PCOS were selected from a larger cohort of women undergoing IVF, after carefully screening their sera and follicular fluids for hormonal and biochemical parameters. DNA extracted from cumulus granulosa cells of three women each, from control and PCOS groups was subjected to high-throughput, next generation bisulfite sequencing, using the Illumina HiSeq 2500® platform. Remaining samples were used for the validation of methylation status of some identified genes by pyrosequencing, and the transcript expression profiles of these genes were assessed by quantitative real-time PCR. RESULTS:In all, 6486 CpG sites representing 3840 genes associated with Wnt signaling, G protein receptor, endothelin/integrin signaling, angiogenesis, chemokine/cytokine-mediated inflammation, etc., showed differential methylation in PCOS. Hypomethylation was noted in 2977 CpGs representing 2063 genes while 2509 CpGs within 1777 genes showed hypermethylation. Methylation differences were also noted in noncoding RNAs regulating several ovarian functions that are dysregulated in PCOS. Few differentially methylated genes such as aldo-keto reductase family 1 member C3, calcium-sensing receptor, resistin, mastermind-like domain 1, growth hormone-releasing hormone receptor and tumor necrosis factor, which predominantly contribute to hyperandrogenism, premature luteolysis, and oocyte development defects, were explored as novel epigenetic candidates in mediating ovarian dysfunction. Methylation profiles of these genes matched with our NGS findings, and their transcript expression patterns correlated with the gene hypo- or hypermethylation status. CONCLUSION:Our findings suggest that the epigenetic dysregulation of genes involved in important processes associated with follicular development may contribute to ovarian defects observed in women with PCOS.
Project description:Polycystic ovary syndrome (PCOS), one of the most common endocrinal diseases among reproductive-aged women,is characterized by hyperandrogenemia, chronic oligo/anovulation and polycystic ovarian morphology. In this research, we presented microarrays to identify the differential expressed protein-coding genes and lncRNAs expression profile in the luteinized granulosa cells obtained from PCOS and healthy control patients. Overall design: The human luteinized granulosa cells (hLGCs) were retrieved from the follicular fluid of normal and PCOS woman undergoing IVF or ICSI. RNA was extracted and hybridized to microarray screening of non-coding RNAs and mRNAs.
Project description:Recent studies report the involvement of intra-ovarian factors, such as inflammation and oxidative stress, in the pathophysiology of polycystic ovary syndrome (PCOS), the most common endocrine disorder of reproductive age women. Endoplasmic reticulum (ER) stress is a local factor that affects various cellular events during a broad spectrum of physiological and pathological conditions. It may also be an important determinant of pro-fibrotic remodeling during tissue fibrosis. In the present study, we showed that ER stress was activated in granulosa cells of PCOS patients as well as in a well-established PCOS mouse model. Pharmacological inducers of ER stress, tunicamycin and thapsigargin, were found to increase the expression of pro-fibrotic growth factors, including transforming growth factor (TGF)-?1, in human granulosa cells, and their expression also increased in granulosa cells of PCOS patients. By contrast, treatment of PCOS mice with an ER stress inhibitor, tauroursodeoxycholic acid or BGP-15, decreased interstitial fibrosis and collagen deposition in ovaries, accompanied by a reduction in TGF-?1 expression in granulosa cells. These findings suggest that ER stress in granulosa cells of women with PCOS contributes to the induction of pro-fibrotic growth factors during ovarian fibrosis, and that ER stress may serve as a therapeutic target in PCOS.
Project description:Polycystic ovary Syndrome (PCOS) is a heterogeneous endocrine disorder that shows evidence of genetic predidposition among affected individuals. We have utilized the Microarray data from granulosa cells of normal and PCOS women for network construction. Human granulosa cells were isolated from ovarian aspirates from normal and PCOS women undergoing IVF and for each sample, RNA was extracted and hybridized to an Affymetrix GeneChip.
Project description:Polycystic ovary syndrome (PCOS) is one of the most common reproductive endocrine disorders accompanied by obvious metabolic abnormalities. Lower-quality oocytes and embryos are often found in PCOS women during assisted reproductive technology treatment. However, there is still no clarity about the mechanism of ovarian metabolic disorders and the impact on oocyte maturation in PCOS. The aim of this study was to understand the potential effect of the posttranslational modification on ovarian metabolic homeostasis and oocyte development potential in women with PCOS. A quantitative analysis of acetylated proteomics in ovarian granulosa cells of PCOS and control groups was carried out by mass spectrometry. There was widespread lysine acetylation of proteins, of which 265 proteins had increased levels of acetylation and 68 proteins had decreased levels of acetylation in the PCOS group. Most notably, differentially acetylated proteins were significantly enriched in the metabolic pathways of glycolysis, fatty acid degradation, TCA cycle, tryptophan metabolism, and branched-chain amino acid degradation. Acetyl-CoA acetyltransferase 1 (ACAT1) was an enzyme central to these metabolic pathways with increased acetylation level in the PCOS group, and there was a negative correlation of ACAT1 acetylation levels in PCOS granulosa cells with oocyte quality and embryo development efficiency in the clinic. Lysine acetylation changes of key enzymes in PCOS granulosa cells might attenuate their activities and alter metabolic homeostasis of follicular microenvironment for oocyte maturation and embryo development.