Endocrine Abstracts

Diabetic nephropathy (DN) and insulin resistance (IR) in kidney cells are considered the main cause of end-stage renal failure. Additionally, hyperglycemia and increased fatty (FA) acids are observed in patients with type 2 diabetes mellitus, where lipid accumulation has been correlated to IR. Nonetheless, the impact of IR in early disease stages is still unclear. Moreover, proximal renal tubular cells have high mitochondrial content where aerobic respiration is their primary mechanism of ATP production, being particularly susceptible to metabolic and mitochondrial injury from hyperglycemia. Herein, we will explore the impact of mild (11 mM) to severe hyperglycemia (22 mM) as well as IR under normoglycemia (5 mM) in in vitro conditions in renal epithelial cells’ metabolic and mitochondrial bioenergetics profiles. To do so, we used a human epithelial kidney cell line (HK-2), which was cultured in increasing concentrations of glucose (5mM, 11 mM and 22 mM). Lipid cytotoxicity was used to induce insulin resistance (IR), through the administration of Palmitic Acid (PA), for 24 h. The effect of hyperglycemia and IR in the consumption/production of metabolites were analyzed by nuclear magnetic resonance. Additionally, in vivo mitochondria function was assessed by seahorse analytics, as well as mitochondrial complex I (CI) and II activities. We also evaluated free FA oxidation and lipid accumulation. Our results demonstrated that PA was able to induce IR and altered the metabolic fingerprint in HK-2 secretome since Glucose, Glutamine, Lactate, Pyruvate, Alanine, Isoleucine, and Glutamate concentrations increased in comparison to the same conditions without PA-induced IR, where gluconeogenic amino acids played a key role to supply energy. It was also observed that FA was not the preferential carbon source for HK-2 cells, resulting in its accumulation. Mitochondrial parameters (basal respiration, ATP production, proton leak, maximal and spare respiration capacity) of HK-2 cells are increased in mild hyperglycemia conditions but decreased when cultured in high hyperglycemia with IR. Furthermore, in this condition, there was a decrease in the CI activity of the electron transport chain. Our data demonstrated that progression from mild to severe hyperglycemia induces a metabolic shift, where gluconeogenic amino acids are crucial to supply energy requirements. Furthermore, this progression allied to IR induced mitochondrial dysfunction, suggesting that this time point between mild to severe hyperglycemia might disclose a “window” of opportunity to help to target and treat the early kidney disease associated with hyperglycemia and IR.