title: MiR-20a-5p mediates hypoxia-induced autophagy by targeting ATG16L1 in ischemic kidney injury.
abstract: OBJECTIVE
  Autophagy is a cellular homeostatic mechanism activated under stress conditions and might act as protective response for cell survival in ischemic kidney injury. The micro RNA (miRNA) network may be critically involved in the regulation of autophagy. The aim of this study was to evaluate whether miRNA regulates autophagy in ischemic kidney injury and renal proximal tubular cells under hypoxic conditions.
  
  
  METHODS
  Ischemic kidney injury was performed by clamping bilateral renal pedicles for 60min in male mice. Human kidney proximal tubular (HK-2) cells were exposed to in vitro hypoxic conditions. ATG16L1 is essential for autophagosome formation. Bioinformatics analyses were used to select the candidate miRNA, miR-20a-5p, which potentially targets ATG16L1. Gain-of-function and loss-of-function methods were employed to evaluate the effects of miRNA on autophagy. Chromatin immunoprecipitation analysis and promoter luciferase reporter assays were used to evaluate the interaction of transcriptional factors with miRNA.
  
  
  RESULTS
  Increased expression of punctate LC3 and ATG16L1, autophagy-related proteins, and down-expression of miR-20a-5p were detected in kidneys after ischemic injury and in HK-2 cells under hypoxic conditions. 3'-untranslated region luciferase reporter assays indicated that miR-20a-5p targeted ATG16L1 messenger RNA. Over-expression of miR-20a-5p reduced the expression of LC3-II and ATG16L1 in HK-2 cells under hypoxic conditions, whereas antagomiR-20a reversed the inhibition. Using RNAi against hypoxia-inducible factor-1α (HIF-1α) in HK-2 cells, we confirmed the inhibitory binding of HIF-1α to miR-20a-5p.
  
  
  CONCLUSIONS
  The signaling axis of HIF-1α, miR-20a-5p, and ATG16L1 in autophagic process might be a critical adapting mechanism for ischemic kidney injury.
authors.affiliation: Graduate Institute of Clinical Medical Science, China Medical University College of Medicine, Taichung, Taiwan; Division of Nephrology, China Medical University Hospital, Taichung, Taiwan; Department of Internal Medicine, China Medical University College of Medicine, Taichung, Taiwan.
authors.forename: I-Kuan
authors.initials: IK
authors.lastname: Wang
authors.affiliation: Graduate Institute of Clinical Medical Science, China Medical University College of Medicine, Taichung, Taiwan; Department of Dentistry, China Medical University Hospital, Taichung, Taiwan.
authors.forename: Kuo-Ting
authors.initials: KT
authors.lastname: Sun
authors.affiliation: Graduate Institute of Clinical Medical Science, China Medical University College of Medicine, Taichung, Taiwan; Division of Urology, Department of Surgery, Taichung Tzu Chi Hospital, Buddhist Tzu Chi Medical Foundation, Taichung, Taiwan.
authors.forename: Tsung-Hsun
authors.initials: TH
authors.lastname: Tsai
authors.affiliation: Graduate Institute of Clinical Medical Science, China Medical University College of Medicine, Taichung, Taiwan; Department of Anesthesiology, China Medical University Hospital, Taichung, Taiwan.
authors.forename: Chia-Wen
authors.initials: CW
authors.lastname: Chen
authors.affiliation: Graduate Institute of Clinical Medical Science, China Medical University College of Medicine, Taichung, Taiwan; Division of Cardiology, China Medical University Hospital, Taichung, Taiwan.
authors.forename: Shih-Sheng
authors.initials: SS
authors.lastname: Chang
authors.affiliation: Graduate Institute of Clinical Medical Science, China Medical University College of Medicine, Taichung, Taiwan; Division of Nephrology, Taichung Veterans General Hospital, Taichung, Taiwan.
authors.forename: Tung-Min
authors.initials: TM
authors.lastname: Yu
authors.affiliation: Division of Nephrology, Chang Gung Memorial Hospital, Taipei, Taiwan; Chang Gung University College of Medicine, Taoyuan, Taiwan.
authors.forename: Tzung-Hai
authors.initials: TH
authors.lastname: Yen
authors.affiliation: Division of Cardiology and Cardiovascular Research Center, Taipei Medical University Hospital, Taipei, Taiwan; Department of Internal Medicine, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Feng-Yen
authors.initials: FY
authors.lastname: Lin
authors.affiliation: Graduate Institute of Clinical Medical Science, China Medical University College of Medicine, Taichung, Taiwan; Division of Nephrology, China Medical University Hospital, Taichung, Taiwan.
authors.forename: Chiu-Ching
authors.initials: CC
authors.lastname: Huang
authors.affiliation: Graduate Institute of Clinical Medical Science, China Medical University College of Medicine, Taichung, Taiwan; Department of Anesthesiology, China Medical University Hospital, Taichung, Taiwan. Electronic address: cyli168@gmail.com.
authors.forename: Chi-Yuan
authors.initials: CY
authors.lastname: Li
mesh_terms: D020413:3' Untranslated Regions; D058186:Acute Kidney Injury / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001343:Autophagy; D000071183:Autophagy-Related Proteins; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D015687:Cell Hypoxia; D002460:Cell Line; D015870:Gene Expression; D006801:Humans; D007511:Ischemia / Q000235:genetics / Q000378:metabolism*; D007668:Kidney / Q000098:blood supply / Q000378:metabolism*; D008297:Male; D008810:Mice, Inbred C57BL; D035683:MicroRNAs / Q000502:physiology*; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D034622:RNA Interference
chemical_list: D020413:3' Untranslated Regions; C518959:ATG16L1 protein, human; D000071183:Autophagy-Related Proteins; D002352:Carrier Proteins; C477012:MAP1LC3A protein, human; C534451:MIRN20a microRNA, human; D035683:MicroRNAs; D008869:Microtubule-Associated Proteins
keywords: Acute kidney injury; Autophagy; Ischemia–reperfusion; MicroRNA