The aim of the present study was to explore the effects and mechanisms of insulin on mitochondrial oxidative stress in septic acute kidney injury (AKI). Male Sprague Dawley rats were divided randomly into four groups: Control group, sham surgery group, cecal ligation and puncture (CLP) group, and CLP plus insulin group. Blood specimens and kidney tissues were obtained at 12 and 24 h after surgery as separate experiments. Analyses of histology and indicators of renal injury [blood urea nitrogen (BUN) and serum creatinine (CRE) and neutrophil gelatinase-associated lipocalin (NGAL)], mitochondrial function [adenosine triphosphate (ATP) and mitochondrial membrane potential (MMP)], oxidative stress [inducible nitric oxide synthase (iNOS), reactive oxygen species (ROS) and nitric oxide (NO)], endogenous antioxidant systems [superoxide dismutase (SOD) and glutathione (GSH)] as well as the expression of uncoupling protein (UCP), PINK1 protein (a major mediator of mitophagy), PGC1α protein (a major regulator of mitochondrial biogenesis) were performed. Compared with CLP group, the CLP plus insulin group had milder histological damage, higher levels of ATP and MMP as well as lower levels of BUN, serum CRE and NGAL, intrarenal iNOS, mitochondrial ROS and total NO. Moreover, the CLP plus insulin group demonstrated increased expression of SOD2 and UCP2. In contrast, insulin administration suppressed mitophagy meanwhile did not upregulate total GSH and induce mitochondrial biogenesis following CLP. These findings indicated that the upregulation of SOD2 and UCP2 may be involved in insulin protecting against mitochondrial oxidative stress in septic AKI.

Uncoupling protein 2 (UCP2) may be critical for intestinal barrier function which may play a key role in the development of sepsis, and insulin has been reported to have anti-inflammatory effects. Male Sprague-Dawley rats were randomly allocated into five groups: control group, cecal ligation and puncture (CLP) group, sham surgery group, CLP plus glucose-insulin-potassium (GIK) group, and CLP plus glucose and potassium (GK) group. Ileum tissues were collected at 24 h after surgery. Histological and cytokine analyses, intestinal permeability tests, and western blots of intestinal epithelial tight junction component proteins and UCP2 were performed. Compared with CLP group, the CLP + GIK group had milder histological damage, lower levels of cytokines in the serum and ileum tissue samples, and lower UCP2 expression, whereas the CLP + GK group had no such effects. Moreover, the CLP + GIK group exhibited decreased epithelial permeability of the ileum and increased expression of zonula occludens-1, occludin, and claudin-1 in the ileum. The findings demonstrated that the UCP2 and NLR family-pyrin domain-containing 3/caspase 1/interleukin 1β signaling pathway may be involved in intestinal barrier injury and that GIK treatment decreased intestinal barrier permeability. Thus, GIK may be a useful treatment for intestinal barrier injury during sepsis.

Sepsis-induced myocardial dysfunction is a critical cause of high mortality among patients with sepsis. Previously, insulin has been suggested to protect against lethal endotoxemia, while uncoupling protein 2 (UCP2) has been reported to exert beneficial effects against sepsis. Thus, this study aimed to investigate whether UCP2 is involved in insulin-elicited protection against myocardial dysfunction in lipopolysaccharide (LPS)-induced sepsis. Treatment of male SD rats with insulin for 30 min before LPS challenge improved the survival and cardiac function in endotoxemic rats, which was likely due to an insulin-dependent reduction of serum lactate dehydrogenase (LDH) activity and cardiac troponin T (cTn-T) levels, mitochondrial oxidative stress, and cardiomyocyte apoptosis. Insulin treatment also increased the Bcl-2/Bax ratio, prevented the release of cytochrome c into the cytosol, and reduced cleaved caspase-9 levels, which was determined by purifying the proteins using the His-tag Magnetic Bead Purification Kit (Fe3O4). Moreover, UCP2 was found to be upregulated in endotoxemic rats which were pretreated with insulin. To determine whether the apoptotic role of insulin is associated with UCP2 upregulation, we examined the effects of genipin, a UCP2 inhibitor, on insulin activity in LPS-treated H9c2 cells. Insulin strongly attenuated LPS-induced H9c2 cell apoptosis and stimulated UCP2 expression. However, genipin treatment eliminated the antiapoptotic effects of insulin. Thus, our results demonstrate that insulin-induced UCP2 upregulation plays a role in the protective effect of insulin against LPS-induced myocardial dysfunction.