PMID- 36621906 OWN - NLM STAT- MEDLINE DCOM- 20230111 LR - 20230111 IS - 2299-8306 (Electronic) IS - 0423-104X (Linking) VI - 73 IP - 5 DP - 2022 TI - The KLF4-p62 axis prevents vascular endothelial cell injury via the mTOR/S6K pathway and autophagy in diabetic kidney disease. PG - 837-845 LID - 10.5603/EP.a2022.0072 [doi] AB - INTRODUCTION: Diabetic kidney disease (DKD) is a complication of systemic diabetic microangiopathy, which has a high risk of developing into end-stage renal disease and death. This study explored the mechanism underlying autophagy in DKD vascular endothelial cell injury. MATERIAL AND METHODS: DKD and vascular endothelial cell injury models were established using Sprague Dawley rats and human umbilical vein endothelial cells (HUVECs). HUVECs overexpressing Kruppel-like factor 4 (KLF4) were constructed by transient transfection of plasmids. Biochemical determination of urinary protein and blood urea nitrogen (BUN), superoxide dismutase (SOD), and creatinine (Scr) levels was performed. Renal pathology was observed by periodic acid-Schiff (PAS) staining. Cell Counting Kit-8 (CCK8), terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL), and immunocytochemistry (ICC) were used to analyse the growth and apoptosis of HUVECs. Microtubule-associated protein light chain 3 (LC3) expression was observed by immunofluorescence (IF). The reactive oxygen species (ROS) levels were measured using flow cytometry. Monocyte chemoattractant protein-1 (MCP-1), KLF4, and tumour necrosis factor alpha (TNF-alpha) levels were detected using enzyme-linked immunosorbent assay (ELISA). The expression of KLF4, p62 protein, and LC3 was analysed using reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR). S6 kinase (S6K), p70 ribosomal S6 kinase (p-S6K), Beclin1, ATG5, LC3, p62, Caspase-3, mammalian target of rapamycine (mTOR), and phsophorylated mTOR (p-mTOR) expressions were detected by western blotting. RESULTS: PAS-positive substances (polysaccharide and glycogen) and S6K protein levels increased, and LC3 protein expression decreased in DKD rats. The levels of urinary protein, BUN, and Scr increased, and KLF4 decreased in DKD rats. High glucose (HG) levels decreased the proliferation and increased the apoptosis rate of HUVECs. The expression of ROS, TNF-alpha, MCP-1, and p62 increased, while the expression of SOD, KLF4, Beclin1, ATG5, and LC3 decreased in HG-induced HUVECs. KLF4 overexpression significantly increased Beclin1, ATG5, and LC3 protein expression and decreased p62 protein expression compared to the oe-NC group in HG-induced HUVECs. KLF4 overexpression inhibits the expression of Caspase-3, p-mTOR, and p-S6K in HG-induced HUVECs. CONCLUSIONS: KLF4-p62 axis improved vascular endothelial cell injury by regulating inflammation and the mTOR/S6K pathway in DKD. FAU - Wang, Xinxing AU - Wang X AD - Department of Hepatobiliary Surgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong, China. FAU - Su, Wei AU - Su W AD - Liver Gall Bladder and Pancreatic Surgery Ward, Qinghai Red Cross Hospital, Xining, Qinghai, China. FAU - Ma, Mingze AU - Ma M AD - Department of Infectious Diseases, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong, China. FAU - Zhu, Lichao AU - Zhu L AD - Department of Paediatric Surgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong, China. FAU - Gao, Ruxin AU - Gao R AD - Department of Hepatobiliary Surgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong, China. FAU - Qin, Chuan AU - Qin C AD - Department of Hepatobiliary Surgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong, China. FAU - Shao, Shuai AU - Shao S AD - Department of Hepatobiliary Surgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong, China. FAU - Gao, Dexuan AU - Gao D AD - Department of Urology, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong, China. FAU - Gao, Junlin AU - Gao J AD - Liver Gall Bladder and Pancreatic Surgery Ward, Qinghai Red Cross Hospital, Xining, Qinghai, China. FAU - Zhang, Zhenhai AU - Zhang Z AD - Department of Hepatobiliary Surgery, Shandong Provincial Hospital, Shandong University, Jinan, Shandong, China. zhangzhenhai@sdfmu.edu.cn. LA - eng PT - Journal Article PL - Poland TA - Endokrynol Pol JT - Endokrynologia Polska JID - 0370674 RN - 0 (Beclin-1) RN - EC 3.4.22.- (Caspase 3) RN - 0 (Kruppel-Like Factor 4) RN - EC 2.7.1.1 (MTOR protein, human) RN - EC 2.7.1.1 (mTOR protein, rat) RN - 0 (Reactive Oxygen Species) RN - EC 2.7.11.1 (Ribosomal Protein S6 Kinases, 70-kDa) RN - EC 1.15.1.1 (Superoxide Dismutase) RN - EC 2.7.11.1 (TOR Serine-Threonine Kinases) RN - 0 (Tumor Necrosis Factor-alpha) SB - IM MH - Animals MH - Humans MH - Rats MH - Autophagy/physiology MH - Beclin-1 MH - Caspase 3/metabolism MH - *Diabetes Mellitus MH - *Diabetic Nephropathies MH - *Kruppel-Like Factor 4/metabolism MH - Rats, Sprague-Dawley MH - Reactive Oxygen Species MH - Ribosomal Protein S6 Kinases, 70-kDa/metabolism MH - Signal Transduction MH - Superoxide Dismutase/metabolism MH - TOR Serine-Threonine Kinases/metabolism MH - Tumor Necrosis Factor-alpha MH - Human Umbilical Vein Endothelial Cells OTO - NOTNLM OT - Kruppel-like factor 4 OT - autophagy OT - diabetic kidney disease OT - human umbilical vein endothelial cells OT - mTOR/S6K pathway EDAT- 2023/01/10 06:00 MHDA- 2023/01/11 06:00 CRDT- 2023/01/09 06:42 PHST- 2021/11/16 00:00 [received] PHST- 2022/03/25 00:00 [accepted] PHST- 2022/02/22 00:00 [revised] PHST- 2023/01/09 06:42 [entrez] PHST- 2023/01/10 06:00 [pubmed] PHST- 2023/01/11 06:00 [medline] AID - VM/OJS/J/86971 [pii] AID - 10.5603/EP.a2022.0072 [doi] PST - ppublish SO - Endokrynol Pol. 2022;73(5):837-845. doi: 10.5603/EP.a2022.0072.