Increased autophagy mediates the adaptive mechanism of the mammary gland in dairy cows with hyperketonemia

Abstract

Hyperketonemia is a metabolic disease in dairy cows, associated with negative nutrition balance (NNB) induced by low dry matter intake (DMI) and increased nutrient requirements. Hyperketonemia could induce metabolic stress, which might indirectly affect mammary tissue. Autophagy is a highly conserved physiological process that results in the turnover of intracellular material, and is involved in maintaining cellular homeostasis under the challenge of metabolic stress induced by NNB. The aim of this study was to investigate the autophagy status and autophagy-related pathways AMPactivated kinase α (AMPKα) and mechanistic target of rapamycin (mTOR) in the mammary glands of dairy cows with hyperketonemia. Cows with hyperketonemia [CWH, n = 10, blood β-hydroxybutyrate (BHB) concentration 1.2 to 3.0 mmol/L] and cows without hyperketonemia (CWOH, n = 10, BHB < 1.2 mmol/L) from 3 to 12 DIM were randomly selected from the herd. The mammary tissue and blood samples were collected from these cows between 0630 and 0800 h, before feeding, at 3 to 12 d in milk. Serum concentrations of glucose, BHB, and fatty acids were determined using an autoanalyzer with commercial kits between 0630 and 0800 h, before feeding. Concentrations of fatty acids, BHB (median and interquartile range: CWH, 2.44 and 1.3, 2.82 mM; CWOH, 0.49 and 0.41, 0.57 mM), and milk fat were greater in CWH. The DMI, glucose concentration, milk production, and milk protein levels were lower in CWH. The mRNA abundance of autophagosome formationrelated gene, beclin 1 (BECN1), phosphatidylinositol 3-kinase catalytic subunit type 3 (PIK3C3), autophagy-related gene (ATG) 5, ATG7, ATG12, microtubuleassociated protein 1 light chain 3 (MAP1LC3, also called LC3) and sequestosome-1 (SQSTM1, also called p62) were greater in the mammary glands of CWH.The protein abundance of LC3-II and phosphorylation level of Unc-51-like kinase 1 (ULK1) were greater in CWH, but the total ubiquitinated proteins and protein abundance of p62 were lower. Transmission electron microscopy showed an increased number of autophagosomes in the mammary glands of CWH. Furthermore, the phosphorylation of AMPKα was greater, but the phosphorylation of mTOR was lower in the mammary glands of CWH. These results indicate that activity of mTOR pathways and autophagy activity, and upregulation of AMPKα, may be response mechanisms to mitigate metabolic stress induced by hyperketonemia in the mammary glands of dairy cows.

Adenosine 5′-monophosphate-activated protein kinase ameliorates bovine adipocyte oxidative stress by inducing antioxidant responses and autophagy

“…In dairy cows, autophagy is altered during metabolic disorders in the transition period. Enhanced autophagic activity in the liver and mammary gland has been associated with response mechanisms to alleviate metabolic stress induced by mild fatty liver and hyperketonemia (Chen et al, 2020;Li et al, 2020). At least in nonruminants experiencing metabolic disorders associated with adipose tissue dysfunction, autophagy is stimulated by oxidative stress (Zhang et al, 2012;Mizunoe et al, 2017).…”

NEFAs Influence the Inflammatory and Insulin Signaling Pathways Through TLR4 in Primary Calf Hepatocytes in vitro

“…Clinical data show that fatty liver in cows often manifests as insulin resistance. Cows with type I ketosis often exhibit low insulin levels, and those with type II ketosis often exhibit high insulin levels with decreased insulin sensitivity ( 26 , 34 , 35 ). Therefore, specific liver regulation of the insulin signaling pathway is crucial to explaining the pathogenesis of cow ketosis and fatty liver.…”

NEFA Promotes Autophagosome Formation through Modulating PERK Signaling Pathway in Bovine Hepatocytes

“…In line with our results, Chen et al indicated that enhanced formation and degradation of autophagosomes were found in the liver of cows with mild fatty liver during the perinatal period [ 15 ]. Moreover, Li et al previously suggested that activation of autophagy and an increase in the number of autophagosomes were found in the mammary tissues of cows with hyperketonemia [ 30 ]. Overall, autophagy can actively participate in the process of recycling and remobilizing nutrients in cells that serves as an adaptive mechanism for NEB.…”