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Next Generation Sequencing Analysis of exosomes in follicle fluid of PCOS/non-PCOS patients

ABSTRACT: Purpose: the goals of this study are to identify the transcriptome profiling of exosome derived from follicle fluid of PCOS patients by next-generation sequencing (NGS). Methods:the transcriptome profiling of exosome derived from follicular fluid of five PCOS and five control patients were generated by deep sequencing, using Illumina GAIIx. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT–PCR validation was performed using TaqMan and SYBR Green assays Results: the transcriptome profiling including circRNA, lncRNA and mRNA were comparatively analyzed in the two groups of exosomes (exo-PCOS and exo-control). We focus on the circRNAs profile for further analysis. A circRNA candidate was considered to be differentially expressed if its levels between both experiment groups showed a statistically significant difference (p<0.05) of at least two-fold. Based on this criterion, we identified 16 differentially expressed circRNAs (5 up-regulated and 11 down-regulated circRNAs) between exo-PCOS and exo-control, including 3 annotated exonic circRNAs and 13 novel ones. Clustering analysis based on the differentially expressed circRNAs clustered the exo-PCOS and the exo-control groups (control). Conclusion: the circRNA expression profile of exosome derived from follicular fluid of five PCOS and five control patients were determined by RNA sequencing technology. 16 circRNAs showed significantly different expression in the exosome of PCOS FF. GO and KEGG pathway analysis indicated that the parental genes of the candidate circRNAs were involved in ovarian steroidogenesis, aldosterone synthesis and secretion, thyroid hormone signaling pathway, ubiquitin mediated proteolysis, Jak-STAT signaling pathway, and endocytosis pathway.

ORGANISM(S): Homo sapiens

PROVIDER: GSE145461 | GEO | 2020/09/09

REPOSITORIES: GEO

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Project description:Background: Circular RNA (circRNA) has been reported to be involved in the pathogenesis of cardiovascular disease (CVD), however, it is unclear whether circRNA carried by exosomes (exos) can be used as biomarkers for chronic coronary syndrome (CCS). This study aimed to investigate the expression pattern of exosomal circRNAs in the plasma of CCS patients, and to screen exo-circRNAs that can act as biomarkers for the diagnosis of CCS. Methods: High-throughput sequencing technology was carried out in the plasma exosomal RNA of 15 CCS patients and 15 non-cardiac chest pain patients (NCCP, control group) (sequencing cohort) to screen for differentially expressed circRNAs. Selected differentially expressed exo-circRNAs were further verified by real time polymerase chain reaction (RT-PCR) in a small-sample cohort (derivation cohort) and a large-sample cohort (validation cohort). Receiver operating characteristic (ROC) curve analysis was performed to evaluate the diagnostic value of exo-circRNAs for CCS patients. Results: By high-throughput sequencing of circRNAs in 15 CCS patients and 15 NCCP patients, 276 circRNAs differentially expressed in plasma exosomes of CCS patients were screened by using |log2(Fold change) |>1 and q-value<0.001, and 103 were up-regulated and 173 were down-regulated. Among the 103 up-regulated circRNAs, 5 circRNAs with high expression levels and significantly increased in plasma exosomes of CCS patients were selected for validation. Twice RT-PCR validation demonstrated that exo-hsa_circ_0075269 and exo-hsa_circ_0000284 were significantly up-regulated in patients with CCS (P<0.001). Circulating exo-hsa_circ_0075269 and exo-hsa_circ_0000284 yielded the area under the curve (AUC) values of 0.761 (P<0.001, 95%CI=0.669, 0.852) and 0.623 (P=0.015, 95%CI=0.522, 0.724) for CCS, respectively by ROC curve analysis. Moreover, AUC of hsa_circ_0075269 combined with hsa_circ_0000284 for diagnosing CCS was 0.741 (P<0.001, 95%CI=0.667, 0.850). Conclusion: The expression profile of circRNA in plasma exosomes of patients with CCS was significantly different from that of the control group. Plasma exo-hsa_circ_0075269 and exo-hsa_circ_0000284 have the potential to be new biomarkers for CCS diagnosis.

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Next Generation Sequencing Analysis of exosomes in follicle fluid of PCOS/non-PCOS patients

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