Differentially expressed genes analysis of retinal vascular endothelial cells under hyperglycemia and hyperlipidemia induced by α-melanocyte-stimulating hormone

Authors:Wu Mianmian,  Guo Fang,  Li Yahong,  Liu Ke,  Zhang Yan
DOI: 10.3760/cma.j.issn.2095-0160.2019.09.002
Published 2019-09-10
Cite as Chin J Exp Ophthalmol, 2019,37(9): 694-700.

Abstract

Objective

To investigate the effect of α-melanocyte-stimulating hormone(α-MSH) on the expression of mRNA and long noncoding RNA (lncRNA) in retinal vascular endothelial cells stimulated by hyperglycemia and hyperlipidemia.

Methods

The simian retinal vascular endothelial cells (RF/6A)were cultured and divided into normal control group, model control group, 0.1 μmol/L α-MSH group, 0.5 μmol/L α-MSH group and 1.0 μmol/L α-MSH group.The cells were stained with CM-H2DCFDA to detect cell antioxidant capacity.The optimal concentration of α-MSH was screened.The cells from normal control group, model control group and α-MSH treatment group were collected at 24 hours after treatment, the total RNA was extracted, the cDNA library was constructed, and the high throughput RNA sequencing (RNA-seq) was carried out with bioinformatics analysis to analyze the expression profiling of mRNA and lncRNA.

Results

The fluorescence intensity of cells in 0.5 μmol/L α-MSH group was significantly lower than that in model control group (P<0.05). α-MSH of 0.5 μmol/L was chosen as the optimal concentration for subsequent experiments.Compared with the model control group, 243 mRNAs were significantly down-regulated, while 81 mRNAs were up-regulated in the α-MSH treatment group; 53 lncRNAs were markedly up-regulated and 6 lncRNAs were down-regulated in the α-MSH treatment group.Bioinformatics analysis showed that the major enrichment pathways of the down-regulated genes were transforming growth factor-β(TGF-β) signaling pathway, focal adhesion signaling pathway and extracellular matrix (ECM) receptor interactions pathways, and the main biological process involved was the regulation of small GTPase-mediated signal transduction.The co-expression gene enrichment pathways of differentially expressed lncRNA included ECM receptor interaction and hypoxia inducible factor-1(HIF-1) pathway, et al.These pathways were mainly involved in the biological processes, such as axon guidance and positive regulation of transcription from RNA polymerase Ⅱ promoter.

Conclusions

Under hyperglycemia and hyperlipidemia, the influence of α-MSH on the transcriptome of the retinal vascular endothelial cells manifests the downregulation of mRNA and upregulation of lncRNA.α-MSH may upregulate the lncRNA expression, which downregulates the downstream mRNA expression.

Key words:

α-Melanocyte-stimulating hormone; Diabetic retinopathy; RNA sequencing; Gene expression; Hyperglycemia; Hyperlipidemia; Sodium palmitate

Contributor Information

Wu Mianmian
Tianjin Key Laboratory of Retinal Functions and Diseases, Eye Institute and School of Optometry, Tianjin Medical University Eye Hospital, Tianjin 300384, China
Guo Fang
Li Yahong
Liu Ke
Zhang Yan
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Updated: March 25, 2020 — 12:04 pm