Enhanced mitochondrial dysfunction and oxidative stress in the mammary gland of cows with clinical ketosis

Abstract

Ketosis is a common metabolic disorder in high-producing dairy cows during the peripartal period. Negative energy balance leads to increased circulating levels of nonesterified fatty acids (NEFA) and β-hydroxybutyrate (BHB), consequently increasing the risk of ketosis. It is well-known that NEFA and BHB can induce lipotoxicity and oxidative stress in bovine tissues/organs including the liver and adipose tissue. Although the mammary gland is one important site for NEFA and BHB metabolism, whether an overload in their concentrations within mammary cells causes oxidative stress during ketosis remains unclear. Thus, the present study compared oxidative stress status and mitochondrial function in mammary tissues harvested by biopsy from healthy (n = 15) and clinically ketotic (n = 15) dairy cows within 2 to 3 wk postpartum. Compared with healthy cows, ketotic cows had depressed daily milk yield (median: 28.92 vs. 21.56 kg) and dry matter intake (median: 22.36 vs. 19.92 kg/d), accompanied by elevated plasma NEFA (median: 0.32 vs. 1.26 mM), BHB (median: 0.52 vs. 3.69 mM), and lower plasma glucose (median: 4.55 vs. 2.13 mM). As detected by a commercial kit, a greater level of reactive oxygen species in mammary epithelial cells of ketotic cows, and greater oxidant indices including hydrogen peroxide and malondialdehyde coupled with lower antioxidant indices including glutathione peroxidase, catalase, and superoxide dismutase activities as detected by the respective biochemical kits in the homogenate of mammary tissue of ketotic cows indicated increased oxidative stress status. Lower citrate synthase activity and ATP production as detected by the respective commercial kits coupled with lower mRNA and protein abundance of mitochondrial respiratory chain oxidative phosphorylation complexes I-V (CO I-V) in ketotic cows suggested an impairment of mitochondrial function. This was supported by lower mRNA and protein abundance of nucleus-derived mitochondrial function regulators including peroxisome proliferator activated receptor gamma coactivator 1 α, mitofusin 2, nuclear respiratory factor 1, and mitochondrial transcription factor A. Lower mitochondrial membrane potential evaluated via the tetraethylbenzimidazolylcarbocyanine iodide (JC-1) labeling method and swollen mitochondria in mammary epithelial cells of ketotic cows suggested the existence of mitochondrial damage. Overall, the present study revealed extensive mitochondrial dysfunction and oxidative stress in the mammary gland of clinically ketotic cows. As such, data suggest that reduced milk yield in cows with ketosis is partly due to enhanced oxidative stress along with mitochondrial dysregulation in the mammary gland.