All- trans retinoic acid induces lipophagy through the activation of the AMPK-Beclin1 signaling pathway and reduces Rubicon expression in adipocytes

Lipophagy is defined as a lipolysis pathway that degrades lipid droplet (LD) via autophagy. All - trans retinoic acid (atRA), a metabolite of vitamin A, stimulates lipolysis through hormone -sensitive lipase and a -oxidation. However, the regulation of lipolysis by atRA-induced autophagy in adipocytes remains unclear. In this study, we investigated the effect of atRA on autophagy in epididymal fat of mice and the molecular mechanisms of autophagy in 3T3 -L1 adipocytes. Western blotting showed that atRA decreased the expression of p62, a cargo receptor for autophagic degradation, and increased the expression of the lipidated LC3B (LC3B-II), an autophagy marker, in epididymal fat. Next, we confirmed that atRA increased autophagic flux in differentiated 3T3 -L1 cells using the GFP-LC3-RFP-LC3AG probe. Immunofluorescent staining revealed that the colocalization of LC3B with perilipin increased in differentiated 3T3 -L1 cells treated with atRA. The knockdown of Atg5, an essential gene in autophagy induction, partly suppressed the atRA-induced release of non-esterified fatty acid (NEFA) from LDs in differentiated 3T3 -L1 cells. atRA time -dependently elicited the phosphorylation of AMPK and Beclin1, autophagy-inducing factors, in mature 3T3 -L1 adipocytes. Inversely, atRA decreased the protein expression of Rubicon, an autophagy repressor, in differentiated 3T3 -L1 cells and epididymal fat. Interestingly, the expression of ALDH1A1, atRA-synthesizing enzymes, increased in epididymal fat with decreased protein expression of Rubicon in aged mice. These results suggest that atRA may partially induce lipolysis through lipophagy by activating the AMPK-Beclin1 signaling pathway in the adipocytes and increased atRA levels may contribute to decreased Rubicon expression in the epididymal fat of aged mice. (248/250 words) (c) 2024 The Author(s). Published by Elsevier Inc. This is an open access article under the CC BY -NC license ( http://creativecommons.org/licenses/by-nc/4.0/ )