d-lactate-induced ETosis in cattle polymorphonuclear leucocytes is dependent on the release of mitochondrial reactive oxygen species and the PI3K/Akt/HIF-1 and GSK-3β pathways.

d-lactate is a metabolite originating from bacterial metabolism that accumulates as a result of dietary disturbances in cattle, leading to ruminal acidosis. d-lactate exerts functions as a metabolic signal inducing metabolic reprogramming and extracellular trap (ET) release in polymorphonuclear leucocytes (PMNs). We previously demonstrated that d-lactate induces metabolic reprogramming via hypoxia-induced factor 1 alpha (HIF-1α) stabilization in bovine fibroblast-like synoviocytes (FLSs). In the present study, the role of HIF-1 in ET formation induced by d-lactate was assessed. HIF-1α stabilization in PMNs was controlled by mitochondrial reactive oxygen species (mtROS) release. Furthermore, inhibition of mitochondrial complex I and scavenging of mtROS decreased d-lactate-triggered ETosis. d-lactate-enhanced HIF-1α accumulation was dependent on the PI3K/Akt pathway but independent of GSK-3β activity. Pharmacological blockade of the PI3K/Akt/HIF-1 and GSK-3β axes inhibited d-lactate-triggered ETosis and downregulated PDK1 and LDHA expression. However, only GSK-3β inhibition decreased the expression of glycogen metabolism enzymes and prevented the decline in glycogen stores induced by d-lactate exposure. The results of this study suggest that mtROS, PI3K/Akt/HIF-1 and GSK-3β axes regulate carbohydrate metabolism adaptations that support d-lactate-induced ET formation in cattle.