Acupuncture therapy protects PCOS patients with diabetes by regulating miR-32-3p/PLA2G4A pathway - PubMed (original) (raw)

. 2021 Aug 15;13(8):8819-8832.

eCollection 2021.

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Acupuncture therapy protects PCOS patients with diabetes by regulating miR-32-3p/PLA2G4A pathway

Jia Wu et al. Am J Transl Res. 2021.

Abstract

Objective: To investigate the potential miRNA-mRNA network co-expressed in polycystic ovary syndrome (PCOS) and diabetes, and explore the molecular mechanism of traditional acupuncture treatment of PCOS.

Methods: Patients with PCOS and diabetes who had undergone acupuncture treatment from January 2019 to June 2020 were recruited in this study. The potential miRNA-mRNA network co-expressed in PCOS and diabetes was obtained through bioinformatics analysis. The expression levels of candidate gen es were determined using quantitative qRT-PCR to study the effectiveness of acupuncture approach. Further, the mechanism of action of acupuncture method was determined using luciferase assay.

Results: A total of 44 patients were included in this study. The miRNA-mRNA network for PCOS was then constructed based on the results of the bioinformatics analysis. Acupuncture treatment could significantly down-regulate miR-32-3p levels and up-regulate expression of PLA2G4A. Luciferase experiments showed that miR-32-3p could affect glucose metabolism in PCOS patients through down-regulating PLA2G4A expression. Functional and pathway enrichment analysis further suported this finding.

Conclusions: MiR-32-3p regulates PLA2G4A protein expression, which is vital in the pathogenesis of PCOS and diabetes. Further, this research proved that the potential mechanism of traditional acupuncture treatment may be the downregulation of miR-32-3p, thus inhibiting PCOS and diabetes progression.

Keywords: Acupuncture therapy; diabetes mellitus; group IV phospholipases A2 (PLA2G4A); microRNAs; polycystic ovary syndrome.

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Conflict of interest statement

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Figures

Figure 1

Figure 1

Identification of DEMs and DEmiRNAs. Green dots indicate the down-regulated genes and red dots denote the genes that are upregulated. A: Venn diagram showing intersection of DEMs in GSE25462 and GSE34526 datasets; B: Heat map of DEMs from GSE25462 dataset; C: Volcano plot of GSE34526 (21655 RNAs); D: Volcano plot of GSE25462 (20486 RNAs); E: Volcano plot of GSE70318 (153 miRNAs).

Figure 2

Figure 2

Construction of miRNA-mRNA network and analysis of clinical correlation. A: The miRNA-mRNA network for DE-miRNAs and DEMs; B: Relative expression of PLA2G4A and miR-32-3p detected by qPCR. Significant differences were observed between control and acupuncture groups; C: Relative expression of PLA2G4A and miR-32-3p detected by qPCR. Significant differences were observed between groups with or without ovulation after acupuncture treatment. D: Correlation of miR-32-3p and PLA2G4A expression levels in clinical samples. The relative expression of miR-32-3p and PLA2G4A in clinical samples is negatively correlated.

Figure 3

Figure 3

miR-32-3p downregulates PLA2G4A expression. (A) miR-32-3p as the target miRNA of PLA2G4A was determined by RNAhybrid tool. Minimum free energy (MFE); (B) The relative expression of miR-32-3p in HEK-293 cells was detected by qPCR after transfected with miR-32-3p mimics; (C, D) The result of WB showed expression of PLA2G4A in HEK-293 cells transfected with miR-32-3p inhibitor or miR-32-3p mimic; (E) Results of PCR showed mRNAs level of PTEN, SMG1 and PLA2G4A in HEK-293 cells transfected with miR-32-3p inhibitor or miR-32-3p mimic; (F, G) The correlations shown are for ovarian tissues from GTEX cohorts. Correlation analysis of PLA2G4A with PTEN (F) and SMG1 (G) in ovarian tissue. (H) Analysis of ALP2G4A regulation by miR-32-3p by Luciferase reporter assays; (I) A model showing downregulation of PLA2G4A expression by miR-32-3p.

Figure 4

Figure 4

Expression of PLA2G4A in human tissues and cells. A, B: The immunofluorescent analysis of PLA2G4A was carried out. And PLA2G4A was mainly detected in cytosol; C, D: In the endometrial tissue, PLA2G4A was mainly detected in glandular cells; E: PLA2G4A was shown by human protein expression map (GTEX cohort, n=1884).

Figure 5

Figure 5

The KEGG pathway analysis and GO function analysis of pathways and functions associated with PLA2G4A were visualized by GSEA. And significant correlations were observed. A: GO enrichment analysis; B: KEGG enrichment analysis. Note: GSEA: Gene Set Enrichment Analysis.

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