Hypoxia-Inducible Factor 1α and 2α Have Beneficial Effects in Remote Ischemic Preconditioning Against Stroke by Modulating Inflammatory Responses in Aged Rats

Xiangnan Du,Heng Zhao,Chencheng Zhang,Sainan Wang,Jian Yang,Xiaokun Geng,Cuiying Liu,Huishan Du

doi:10.3389/fnagi.2020.00054

Abstract

Limb remote ischemic preconditioning (RIPC) has been proven to alleviate stroke injury in young rats, but its protective effect and its mechanism in aged rats are still unclear. Hypoxia-inducible factor (HIF) is one of the important markers of stroke, and its high expression plays an important role in the pathogenesis of stroke. In this study, we tested the hypothesis that RIPC could regulate the expression of HIF, leading to reduced inflammatory responses in aged rats. Stroke was induced by transient middle cerebral artery occlusion (MCAo) in aged rats, and RIPC was conducted in both hind limbs. The HIF-1α and HIF-2α mRNA and protein were examined by real-time RT-PCR and western blotting (WB). Inflammatory cytokines in the peripheral blood and brain were measured using AimPlex multiplex immunoassays. The protein levels of p-Akt, Akt, p-ERK, and ERK were examined by WB. We investigated that RIPC reduced the infarct size, improved neurological functions, and decreased the expression of HIF-1α and HIF-2α in the ischemic brain. RIPC reduced the levels of IL-1β, IL-6 and IFN-γ in the peripheral blood and the levels of IL-1β and IFN-γ in the ischemic brain 48 h post-stroke. Moreover, intraperitoneal injection of the HIF inhibitor, acriflavine hydrochloride (ACF), abolished the protection of RIPC with respect to infarct size and neurological functions and neutralized the downregulation of pro-inflammatory IL-1β, IL-6 and IFN-γ. ACF also reversed the activation of the Akt signaling pathway induced by RIPC following stroke. HIF may play a key role in RIPC, which was likely mediated by the Akt signaling pathway and systemic modulation of the inflammatory response in aged rats.

Highlights

Ischemic stroke is considered to be the third most lethal and disabling disease in the world
The tetrazolium chloride (TTC) staining results showed that remote ischemic preconditioning (RIPC) induced a 34.64% significantly reduction of infarct size from 38.85 ± 1.15% to 25.39 ± 1.12% compared with middle cerebral artery occlusion (MCAo) (Figure 1A and Supplementary Figure S2), attenuated neurological scores (Figure 1B), and improved behavioral performance in the tail hang test (Figure 1B)
The results showed that MCAo increased the levels of pro-inflammatory cytokines while RIPC significantly decreased the levels of pro-inflammatory IL-1β, IL-6 and IFN-γ in the peripheral blood (Figure 3A) and that of IL-1β and IFN-γ in penumbra tissue (Figure 3B) 48 h post-stroke in aged rats

Summary

Introduction

Ischemic stroke is considered to be the third most lethal and disabling disease in the world. According to the World Health Organization (WHO) statistics, twelve million people will suffer from stroke in the coming decade (Krishnamurthi et al, 2013). Given the increase of the senior citizen and the aggravating of the associated stroke burden, it is urgent to find effective treatments for this disease. Remote ischemic preconditioning (RIPC) has become a feasible therapeutic strategy for stroke and has been proven to provide a protective effect on young experimental animals (Wei et al, 2012; Liu et al, 2016; Yang et al, 2018) and in clinical trials Its protective effect and its mechanism in the elderlies are still unclear

Objectives

Methods

Conclusion