Methionine deficiency inhibited pyroptosis in primary hepatocytes of grass carp (Ctenopharyngodon idella): possibly via activating the ROS-AMPK-autophagy axis.

in Journal of animal science and biotechnology by Yuanlin He, Pei Wu, Weidan Jiang, Yang Liu, Xiaowan Jin, Hongmei Ren, Ruinan Zhang, Xiaoqiu Zhou, Lin Feng

TLDR

  • The study looked at how methionine deficiency (MD) affects different ways cells can die in primary hepatocytes from grass carp. The study found that MD promotes autophagy and apoptosis, but inhibits pyroptosis and necroptosis. The study also found changes in protein expression in response to MD treatment, which could help us understand how MD affects cell death and organ function.

Abstract

Methionine (Met) is the only sulfur-containing amino acid among animal essential amino acids, and methionine deficiency (MD) causes tissue damage and cell death in animals. The common modes of cell death include apoptosis, autophagy, pyroptosis, necroptosis. However, the studies about the major modes of cell death caused by MD have not been reported, which worth further study. Primary hepatocytes from grass carp were isolated and treated with different doses of Met (0, 0.5, 1, 1.5, 2, 2.5 mmol/L) to examine the expression of apoptosis, pyroptosis, autophagy and necroptosis-related proteins. Based on this, we subsequently modeled pyroptosis using lipopolysaccharides and nigericin sodium salt, then autophagy inhibitors chloroquine (CQ), AMP-activated protein kinase (AMPK) inhibitors compound C (CC) and reactive oxygen species (ROS) scavengers N-acetyl-L-cysteine (NAC) were further used to examine the expression of proteins related to pyroptosis, autophagy and AMPK pathway in MD-treated cells respectively. MD up-regulated B-cell lymphoma protein 2 (Bax), microtubule-associated protein 1 light chain 3 II (LC3 II), and down-regulated the protein expression levels of B-cell lymphoma-2 (Bcl-2), sequestosome 1 (p62), cleaved-caspase-1, cleaved-interleukin (IL)-1β, and receptor-interacting protein kinase (RIP) 1 in hepatocytes, while it did not significantly affect RIP3. In addition, MD significantly increased the protein expression of liver kinase B1 (LKB1), p-AMPK, and Unc-51-like kinase 1 (ULK1) without significant effect on p-target of rapamycin. Subsequently, the use of CQ increased the protein expression of NOD-like receptor thermal protein domain associated protein 3 (NLRP3), cleaved-caspase-1, and cleaved-IL-1β inhibited by MD; the use of CC significantly decreased the protein expression of MD-induced LC3 II and increased the protein expression of MD-suppressed p62; then the use of NAC decreased the MD-induced p-AMPK protein expression. MD promoted autophagy and apoptosis, but inhibited pyroptosis and necroptosis. MD inhibited pyroptosis may be related regarding the promotion of autophagy. MD activated AMPK by inducing ROS production which in turn promoted autophagy. These results could provide partial theoretical basis for the possible mechanisms of Met in ensuring the normal structure and function of animal organs. Furthermore, ferroptosis is closely related to redox states, it is worth investigating whether MD affects ferroptosis in hepatocytes.

Overview

  • The study investigates the effects of methionine deficiency (MD) on apoptosis, pyroptosis, autophagy, and necroptosis in primary hepatocytes from grass carp. The study aims to identify the major modes of cell death caused by MD and explore the underlying mechanisms.

Comparative Analysis & Findings

  • MD up-regulated Bax, LC3 II, and down-regulated Bcl-2, p62, cleaved-caspase-1, cleaved-IL-1β, and RIP1 in hepatocytes. MD significantly increased the protein expression of LKB1, p-AMPK, and ULK1 without significant effect on p-target of rapamycin. The use of CQ increased the protein expression of NLRP3, cleaved-caspase-1, and cleaved-IL-1β inhibited by MD; the use of CC significantly decreased the protein expression of MD-induced LC3 II and increased the protein expression of MD-suppressed p62; then the use of NAC decreased the MD-induced p-AMPK protein expression. MD promoted autophagy and apoptosis, but inhibited pyroptosis and necroptosis. MD inhibited pyroptosis may be related regarding the promotion of autophagy. MD activated AMPK by inducing ROS production which in turn promoted autophagy. These results could provide partial theoretical basis for the possible mechanisms of Met in ensuring the normal structure and function of animal organs.

Implications and Future Directions

  • The study highlights the importance of understanding the mechanisms of MD-induced cell death and its impact on organ function. Further research is needed to investigate the role of ferroptosis in MD-induced cell death and its potential therapeutic implications. Additionally, the study suggests that MD may have a protective effect on hepatocytes by promoting autophagy and apoptosis, which could be explored in future studies.
Read Full Article