MicroRNA-352 regulates collateral vessel growth induced by elevated fluid shear stress in the rat hind limb

Yinglu Guan,Wei-Jun Cai,Lin Xiao,Jutta Schaper,Wolfgang Schaper,Xiaoqiong Wu,Da Huang,Guangmin Liu,Bin Zhang,Baizhen Cai,Song Peng,Liaoying Gan,Junwen Liu,Liping Dong

doi:10.1038/s41598-017-06910-9

Abstract

Although collateral vessel growth is distinctly enhanced by elevated fluid shear stress (FSS), the underlying regulatory mechanism of this process remains incompletely understood. Recent studies have shown that microRNAs (miRNAs) play a pivotal role in vascular development, homeostasis and a variety of diseases. Therefore, this study was designed to identify miRNAs involved in elevated FSS-induced collateral vessel growth in rat hind limbs. A side-to-side arteriovenous (AV) shunt was created between the distal stump of one of the bilaterally occluded femoral arteries and the accompanying vein. The miRNA array profile showed 94 differentially expressed miRNAs in FSS-stressed collaterals including miRNA-352 which was down-regulated. Infusion of antagomir-352 increased the number and proliferation of collateral vessels and promoted collateral flow restoration in a model of rat hind limb ligation. In cell culture studies, the miR-352 inhibitor increased endothelial proliferation, migration and tube formation. In addition, antagomir-352 up-regulated the expression of insulin-like growth factor II receptor (IGF2R), which may play a part in the complex pathway leading to arterial growth. We conclude that enhanced collateral vessel growth is controlled by miRNAs, among which miR-352 is a novel candidate that negatively regulates arteriogenesis, meriting additional studies to unravel the pathways leading to improved collateral circulation.

Highlights

Collateral vessel growth is distinctly enhanced by elevated fluid shear stress (FSS), the underlying regulatory mechanism of this process remains incompletely understood
Among 3100 capture probes on the microarrays, 94 differentially expressed miRNAs (2-fold changes) were identified, among which 44 miRNAs were up-regulated and 50 were down-regulated; they are listed in the online-only Data Supplement (Figure S2 and Table S1). Among those differentially expressed miRNAs (2-fold changes), many have been described to be involved in the regulation of cell proliferation, migration, MMPs, apoptosis, angiogenesis and growth factors (Tables 1 and 2), which are important factors that contribute to collateral vessel growth, indicating the validity of our experimental approach
We screened potential miRNA seed sequences in the 3′ untranslated regions (3′-UTR) using 3 bioinformatic algorithms, and identified 15 genes with a miR-352 target site predicted by all three bioinformatic algorithms (Figure S4); one of them is IGF2R

Summary

Introduction

Collateral vessel growth is distinctly enhanced by elevated fluid shear stress (FSS), the underlying regulatory mechanism of this process remains incompletely understood. Increased expression of several biologically active substances, such as vascular endothelial cell adhesion molecules, inflammatory factors and growth factors was evident and several groups of differential mRNAs have been identified using this AV-shunt model[8, 9] These data strongly suggest that increased FSS induced by arterial occlusion controls collateral vessel growth via regulating gene expression at both the protein and mRNA levels. Based on the above-mentioned findings and our previous findings that high levels of FSS lead to differential mRNA expression in collateral vessels between the AV-shunt side and the ligature-only side[9], we hypothesized that there exists novel miRNAs that could play an important role in amplifying collateral vessel growth in response to elevated FSS

Methods

Results

Conclusion