FGF21 induces autophagy-mediated cholesterol efflux to inhibit atherogenesis via RACK1 up-regulation.

Lin Xiaolong,Mihua Liu,Lin Jun,Lin Wensheng,Guo Dongmin,Tan Qiufen,Zeng Zhaolin,Hu Huijun,Pan Yuping,Hu Xuemei,Wang Zuo

doi:10.1111/jcmm.15118

Abstract

Fibroblast growth factor 21 (FGF21) acts as an anti‐atherosclerotic agent. However, the specific mechanisms governing this regulatory activity are unclear. Autophagy is a highly conserved cell stress response which regulates atherosclerosis (AS) by reducing lipid droplet degradation in foam cells. We sought to assess whether FGF21 could inhibit AS by regulating cholesterol metabolism in foam cells via autophagy and to elucidate the underlying molecular mechanisms. In this study, ApoE−/− mice were fed a high‐fat diet (HFD) with or without FGF21 and FGF21 + 3‐Methyladenine (3MA) for 12 weeks. Our results showed that FGF21 inhibited AS in HFD‐fed ApoE−/− mice, which was reversed by 3MA treatment. Moreover, FGF21 increased plaque RACK1 and autophagy‐related protein (LC3 and beclin‐1) expression in ApoE−/− mice, thus preventing AS. However, these proteins were inhibited by LV‐RACK1 shRNA injection. Foam cell development is a crucial determinant of AS, and cholesterol efflux from foam cells represents an important defensive measure of AS. In this study, foam cells were treated with FGF21 for 24 hours after a pre‐treatment with 3MA, ATG5 siRNA or RACK1 siRNA. Our results indicated that FGF21‐induced autophagy promoted cholesterol efflux to reduce cholesterol accumulation in foam cells by up‐regulating RACK1 expression. Interestingly, immunoprecipitation results showed that RACK1 was able to activate AMPK and interact with ATG5. Taken together, our results indicated that FGF21 induces autophagy to promote cholesterol efflux and reduce cholesterol accumulation in foam cells through RACK1‐mediated AMPK activation and ATG5 interaction. These results provided new insights into the molecular mechanisms of FGF21 in the treatment of AS.

Highlights

Atherosclerosis (AS) is a chronic and degenerative disease in which lipids accumulate in the walls of large arteries
Our results demonstrated that RACK1 knockdown significantly suppressed Fibroblast growth factor 21 (FGF21)-induced autophagy, decreased LC3-I to LC3-II conversion, up-regulated p62 expression (Figure 7A-C) and inhibited autophagosome formation (Figure 7D-F)
FGF21 decreased the levels of total cholesterol (TC), free cholesterol (FC) and cholesterol ester (CE), reduced cholesterol accumulation and increased cholesterol efflux in foam cells, but these effect reversed by RACK1 knockdown (Table 5; Figure 7G,H)

Summary

| INTRODUCTION

Atherosclerosis (AS) is a chronic and degenerative disease in which lipids accumulate in the walls of large arteries. Autophagy is an essential process whereby cells are able to breakdown large portions of the cytoplasm and cellular organelles and recycle them via a catabolic and a highly conserved process.[14] Lipids are degraded to free cholesterol and fatty acids through autophagy.[15] Besides, Wang et al[16] have reported that inhibiting the activation of mTOR activates autophagy and drives cholesterol efflux, thereby decreasing foam cell lipid accumulation, while another study suggested that lysosomal acid lipase mediated autophagy-mediated regulation of cholesterol efflux.[17] mTOR inhibitor everolimus selectively depletes macrophages in atherosclerotic. RACK1 induces the activation of AMPK to protect the liver from ischaemia-reperfusion (I/R) injury.[22] we suggested that FGF21 may induce autophagy via RACK1 to enhance lipid degradation, increase cholesterol efflux and reduce lipid concentration within foam cells, inhibiting AS. Our findings provide novel information regarding the mechanisms of FGF21 and their role in the prevention and treatment of AS

| MATERIALS AND METHODS

| Evaluation of foam cell formation

| DISCUSSION