Leptin stimulates the epithelial‑mesenchymal transition and pro‑angiogenic capability of cholangiocarcinoma cells through the miR‑122/PKM2 axis

Abstract

Leptinisanadipokineminimallyknownforitsactivities or underlying mechanisms in cholangiocarcinoma. The present study explored the effects of leptin on the epithelial-mesenchymal transition (EMT) and pro-angiogenic capability of cholangiocarcinoma cells, and investigated the underlying mechanisms. Cholangiocarcinoma cells were treated with leptin, and their migration and invasion rates were investigated using Transwell assays. Furthermore, conditioned medium was collected from cholangiocarcinoma cells following leptin treatment and applied to human umbilical vein endothelial cells to assess tube formation. The expression of EMT and pro-angiogenic factors was examined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analyses. Mechanistically, the function of pyruvate kinase muscle isozyme M2 (PKM2) was assessed in leptin-induced phenotypes using siRNA targeting PKM2 (si-PKM2). Bioinformatics screening and luciferase reporter assays were used to reveal microRNA (miR)-122 as the potential mediator between leptin and PKM2. Finally, the associations between leptin and miR-122 or PKM2 levels in patients with cholangiocarcinoma were assessed by ELISA and RT-qPCR. Leptin significantly increased the EMT and pro-angiogenic capability of cholangiocarcinoma cells, visibly inhibited endogenous miR-122 expression, and upregulated PKM2. Furthermore, si-PKM2 inhibited leptin-induced migration, invasion, EMT-associated marker expression levels and the pro-angiogenic capability in cholangiocarcinoma cells. In addition, miR-122 negatively regulated the expression of PKM2. When applied together with leptin, miR‑122 was sufficient to reverse the multiple malignancy-promoting effects of leptin. Consistently, the serum leptin level positively correlated with that of PKM2, but negatively with that of miR-122 in patients with cholangiocarcinoma. Leptin, by downregulating miR-122 and elevating PKM2 expression, acts as a pleiotropic pro-malignancy cytokine for cholangiocarcinoma. Therefore, increasing miR-122 expression and inhibiting PKM2 may be future approaches for cholangiocarcinoma treatment.