The periparturient period in dairy cows is the most critical phase of the lactation cycle during which cows are most susceptible to metabolic and inflammatory diseases [[1], [2], [3]]. The increased circulating concentrations of non-esterified fatty acids, ketone bodies, and lipopolysaccharide (LPS) (due to increased gut permeability) underscore the severity of the stress that cows experience during the periparturient period [4]. Under those conditions, there is some evidence that the mammary gland can absorb LPS from the blood, which augments metabolic stress within mammary cells [5,6]. The LPS can trigger a cascade of inflammatory signaling pathways [7], which induce local inflammatory responses [8,9].

Mitochondria are the primary organelles for the production of ATP that helps maintain cell homeostasis [[10], [11], [12], [13]]. Increased metabolic challenges result in mitochondrial damage and production of mitochondrial reactive oxygen species (Mito-ROS) [14]. Excessive Mito-ROS contributes to proinflammatory cytokine production by activating the Nucleotide-binding oligomerization domain-like receptor containing pyrin domain 3 (NLRP3) inflammasome pathway [15,16]. Thus, this proinflammatory response promotes inflammation and damage of mammary gland tissue, leading to a decline in milk production and quality [17].

Activation of the NLRP3 inflammasome pathway and mitochondrial damage in mammary gland have been reported in dairy cows undergoing metabolic stress [12,18]. Work in nonruminants also indicated that exogenous LPS led to mitochondrial damage and Mito-ROS overproduction, which further activated the NLRP3 inflammasome pathway [19]. Thus, alterations in mitochondrial status may contribute to metabolic stress-induced inflammatory responses in mammary gland of dairy cows.

Mitophagy, a specialized form of autophagy, is an important mitochondrial quality control mechanism that eliminates damaged mitochondria [20,21]. This physiological process is tightly regulated by the PTEN induced putative kinase 1 (PINK1). To initiate mitophagy, PINK1 accumulates in the mitochondrial outer membrane and recruits Parkin, which in turn ubiquitinates mitochondrial outer membrane proteins, eventually inducing degradation of damaged mitochondria [22]. Work with nonruminants demonstrated that overexpression of PINK1 suppressed Mito-ROS production and inactivated the NLRP3 inflammasome in Kupffer cells by activating mitophagy [23]. Interestingly, mitophagy was enhanced in ketotic dairy cows mammary gland in early lactation [24]. This may be a compensatory mechanism in the mammary gland to respond to the metabolic stress challenge. Thus, PINK1-mediated mitophagy might play a protective role against mitochondrial damage and inflammatory responses in the bovine mammary gland.

Increased metabolic stress as well as high metabolic priority results in mitochondrial damage and inflammatory responses in bovine mammary epithelial cells during early lactation period. Given the established role of PINK1 in the regulation of autophagy, mitochondrial damage and inflammatory responses in nonruminants, we hypothesized that PINK1-mediated mitophagy protects bovine mammary epithelial cells against mitochondrial damage and inflammatory responses. In the present study, PINK1 adenoviral vectors (Ad-PINK1) and PINK1 small interfering RNA (Si-PINK1) were used to upregulate and downregulate PINK1 abundance in bovine mammary epithelial cells (MAC-T). The aim of the present study was to elucidate the effects of PINK1-mediated autophagy on mitochondrial damage and inflammatory responses in bovine mammary epithelial cells in response to exogenous LPS.