Transcription factor EB (TFEB)-mediated autophagy protects bovine mammary epithelial cells against H2O2-induced oxidative damage in vitro

Xudong Sun,Renxu Chang,Hongdou Jia,Zhihao Dong,Yusheng Liang,Yan Tang,Chunhui Jiang,Qiushi Xu,Juan J Loor,Chuang Xu,Shengbin Luo

doi:10.1186/s40104-021-00561-7

Abstract

BackgroundBovine mammary epithelial cells after calving undergo serious metabolic challenges and oxidative stress both of which could compromise autophagy. Transcription factor EB (TFEB)-mediated autophagy is an important cytoprotective mechanism against oxidative stress. However, effects of TFEB-mediated autophagy on the oxidative stress of bovine mammary epithelial cells remain unknown. Therefore, the main aim of the study was to investigate the role of TFEB-mediated autophagy in bovine mammary epithelial cells experiencing oxidative stress.ResultsH2O2 challenge of the bovine mammary epithelial cell MAC-T increased protein abundance of LC3-II, increased number of autophagosomes and autolysosomes while decreased protein abundance of p62. Inhibition of autophagy via bafilomycin A1 aggravated H2O2-induced reactive oxygen species (ROS) accumulation and apoptosis in MAC-T cells. Furthermore, H2O2 treatment triggered the translocation of TFEB into the nucleus. Knockdown of TFEB by siRNA reversed the effect of H2O2 on protein abundance of LC3-II and p62 as well as the number of autophagosomes and autolysosomes. Overexpression of TFEB activated autophagy and attenuated H2O2-induced ROS accumulation. Furthermore, TFEB overexpression attenuated H2O2-induced apoptosis by downregulating the caspase apoptotic pathway.ConclusionsOur results indicate that activation of TFEB mediated autophagy alleviates H2O2-induced oxidative damage by reducing ROS accumulation and inhibiting caspase-dependent apoptosis.

Highlights

The transition from late-gestation to lactation is considered the most striking and challenging period in the lactation cycle of dairy cows
H2O2-induced oxidative stress and enhanced autophagy in MAC-T cells Compared with the 0 mmol/L H2O2 group, 0.25, 0.5 or 1 mmol/L H2O2 enhanced the level of intracellular reactive oxygen species (ROS) in MAC-T cells (P < 0.05, Fig. 1a)
Inhibition of autophagy aggravated H2O2-induced oxidative damage in MAC-T cells To evaluate effects of autophagy on oxidative stress, cells were pretreated with autophagy inhibitor Bafilomycin A1 (BafA1) for 12 h and treated H2O2 for 24 h

Summary

Introduction

The transition from late-gestation to lactation is considered the most striking and challenging period in the lactation cycle of dairy cows. The bovine mammary epithelial cell undergoes serious metabolic challenges and excessive ROS production during the transition period. Elevated intracellular levels of ROS induce mammary epithelial cell apoptosis, which contributes to the decrease in milk yield [8, 9]. This process is regulated by the cysteine protease family (caspase) apoptotic pathway. Bovine mammary epithelial cells after calving undergo serious metabolic challenges and oxidative stress both of which could compromise autophagy. Effects of TFEB-mediated autophagy on the oxidative stress of bovine mammary epithelial cells remain unknown. The main aim of the study was to investigate the role of TFEB-mediated autophagy in bovine mammary epithelial cells experiencing oxidative stress

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