Advanced glycation end‐products suppress autophagic flux in podocytes by activating mammalian target of rapamycin and inhibiting nuclear translocation of transcription factor EB

Abstract

Insufficient autophagy in podocytes is related to podocyte injury in diabetic nephropathy (DN). Advanced glycation end‐products (AGEs) are major factors of podocyte injury in DN. However, the role and mechanism of AGEs in autophagic dysfunction remain unknown. We investigated autophagic flux in AGE‐stimulated cultured podocytes using multiple assays: western blotting, reverse transcription–quantitative PCR, immunofluorescence staining, and electron microscopy. We also utilized chloroquine and a fluorescent probe to monitor the formation and turnover of autophagosomes. Mice of the db/db strain were used to model diabetes mellitus (DM) with high levels of AGEs. To mimic DM with normal levels of AGEs as a control, we treated db/db mice with pyridoxamine to block AGE formation. AGEs impaired autophagic flux in the cultured podocytes. Compared with db/db mice with normal AGEs but high glucose levels, db/db mice with high AGEs and high glucose levels exhibited lower autophagic activity. Aberrant autophagic flux was related to hyperactive mammalian target of rapamycin (mTOR), a major suppressor of autophagy. Pharmacologic inhibition of mTOR activity restored impaired autophagy. AGEs inhibited the nuclear translocation and activity of the pro‐autophagic transcription factor EB (TFEB) and thus suppressed transcription of its several autophagic target genes. Conversely, TFEB overexpression prevented AGE‐induced autophagy insufficiency. Attenuating mTOR activity recovered TFEB nuclear translocation under AGE stimulation. Co‐immunoprecipitation assays further demonstrated the interaction between mTOR and TFEB in AGE‐stimulated podocytes and in glomeruli from db/db mice. In conclusion, AGEs play a crucial part in suppressing podocyte autophagy under DM conditions. AGEs inhibited the formation and turnover of autophagosomes in podocytes by activating mTOR and inhibiting the nuclear translocation of TFEB. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

SGLT2 Inhibitor Empagliflozin and DPP4 Inhibitor Linagliptin Reactivate Glomerular Autophagy in db/db Mice, a Model of Type 2 Diabetes

“…In our study, glomerular autophagy was related negatively to parameters of glycemic status, fructosamine and glycated albumin. As it has been demonstrated previously, advanced glycation end-products impair autophagic flux in the podocytes via mTOR activation and inhibition of the nuclear translocation and activity of the pro-autophagic transcription factor EB (TFEB) [45]. In cultured mesangial cells, advanced glycation end-products inhibited autophagy via the RAGE/STAT5 axis [46].…”

Autophagy Function and Regulation in Kidney Disease

“…Podocyte injury due to high glucose reduced podocyte autophagy and treatment with ursolic acid improved podocyte injury through miR-21 inhibition and PTEN expression, which suppressed PI3K/Akt/mTOR pathway and restored autophagy [88]. In the db/db mouse model of diabetes mellitus, high levels of advanced glycation end-products (AGEs) and high glucose suppressed podocyte autophagy by activating mTORC1 and inhibiting the nuclear translocation of transcription factor EB (TFEB) [89]. Inhibition of AGE formation by pyridoxamine restored autophagic flux.…”

Thymol alleviates AGEs-induced podocyte injury by a pleiotropic effect via NF-κB-mediated by RhoA/ROCK signalling pathway

“…The incidence of diabetes is increasing yearly, and diabetic nephropathy (DN), the most common cause of end-stage renal disease (ESRD) worldwide [1,2], accounts for the high disability and mortality rate in diabetic patients. Approximately 40% of patients with type 1 or 2 diabetes will develop DN in 20-25 years with glomerular sclerosis, which requires much government spending [3].…”

“…Therefore, podocytes play a critical role in maintaining the filtration membrane integrity. Once these cells are injured, proteinuria associated with DN and the progression of DN occurs [1,22]. Damage to podocytes can lead to proteinuria and initiate glomerulosclerosis, which can result in the progressive loss of renal function [23].…”