Gene Expression and MicroRNA Expression Analysis in Small Arteries of Spontaneously Hypertensive Rats. Evidence for ER Stress.

Teresa Palao,Perry D Moerland,Silvia M Arribas,Erik N T P Bakker,Gergely Groma,Aldo Jongejan,Ed Vanbavel,Angela Van Weert,Karl Swärd,Judith De Vos

doi:10.1371/journal.pone.0137027

Teresa Palao, Perry D Moerland + Show 8 more

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https://doi.org/10.1371/journal.pone.0137027

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Abstract

Small arteries are known to develop functional and structural alterations in hypertension. However, the mechanisms of this remodeling are not fully understood. We hypothesized that altered gene expression is associated with the development of hypertension in mesenteric arteries of spontaneously hypertensive rats (SHR). Three sublines of SHR and normotensive Wistar Kyoto rats (WKY) were studied at 6 weeks and 5 months of age. MiRNA and mRNA microarray experiments were performed and analyzed with bioinformatical tools, including Ingenuity Pathway Analysis (IPA). Principal component analysis showed a clear separation in both miRNA and mRNA expression levels between both ages studied, demonstrating strong age-related changes in expression. At the miRNA level, IPA identified differences between SHR and WKY related to metabolic diseases, cellular growth, and proliferation. The mRNAs differentially expressed between SHR and WKY were related to metabolism, cellular movement and proliferation. The most strongly upregulated gene (9.2-fold) was thrombospondin 4 (Thbs4), a protein involved in the endoplasmic reticulum (ER) stress response that activates transcription factor 6α (ATF6α). ATF6α downstream targets were also differentially expressed in SHR vs. WKY. Differential expression of THBS4, the cleaved form of ATF6α, and two of its targets were further confirmed at the protein level by western blot. In summary, these data revealed a number of genes (n = 202) and miRNAs (n = 3) in mesenteric arteries of SHR that had not been related to hypertension previously. The most prominent of these, Thbs4, is related to vascular ER stress that is associated with hypertension.

Highlights

Despite ample treatment options, hypertension remains one of the most important risk factors for cardiovascular disease
Wistar Kyoto rat (WKY)/NTac rats were obtained from Taconic Farms (Germantown, NY, USA).We studied male rats in order to minimize variations in gene expression related to the estrous cycle in female rats
WKY and spontaneously hypertensive rats (SHR) were less clearly separated in the principal component analysis and hierarchical clustering, indicating a weaker relationship of miRNA and mRNA expression with hypertension than with maturation

Summary

Introduction

Hypertension remains one of the most important risk factors for cardiovascular disease. Blood pressure levels are tightly related to peripheral resistance, which is mainly determined by small arteries and arterioles in the vascular bed. These vessels, referred to as resistance arteries, show structural (remodeling) and functional changes in hypertension, that may increase peripheral resistance and thereby blood pressure. In the current study we tested the hypothesis that gene expression is altered in the development of hypertension in mesenteric arteries of SHR as compared to WKY. To address this hypothesis, we used miRNA and mRNA microarrays, as well as bioinformatical analysis

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