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Study on the effects of miR-155 on epithelial-mesenchymal transition in renal tubular epithelial cells and relevant mechanisms

NING Ya-xian1, WANG Jian-qin1, WANG Xiao-yuan2   

  1. 1. Department of Nephrology, 2. Department of Rheumatology, the Second Hospital of Lanzhou University, Lanzhou 730030, China
  • Online:2016-11-28 Published:2016-11-29
  • Supported by:

    Natural Science Foundation of Gansu Province,1308RJZA246

Abstract:

Objective · To investigate serum miR-155 expression in patients with chronic kidney disease (CKD) and to explore whether miR-155 influences epithelial-mesenchymal transition (EMT) in renal tubular epithelial cells via down-regulating Krüppel-like factor 4 (KLF4). Methods · Serum samples from 126 CKD Ⅰ
stage patients, 163 CKD Ⅲ stage patients, and 170 healthy individuals were collected. RT-PCR was used to test the serum miR-155 expression. In cell experiment, EMT in renal tubular epithelial cells was induced with 10 μg/L TGF-β. miR-155 mimic and miR-NC were transfected to renal tubular epithelial cells cultured in vitro. Changes in cell morphology were observed with optical microscope. Transwell assay was used to test the cell migration ability. Western blotting was used to examine the expressions of SMα-actin, Collagen Ⅰ, Collagen Ⅳ, E-cadherin, and KLF4. Bioinformatics predicted that the potential target gene for miR-155 was KLF4, which was confirmed with luciferase assay. Plasmids with over-expressed KLF4 were transfected to renal tubular epithelial cells and expressions of SMα-actin, collagen Ⅰ, collagen Ⅳ, E-cadherin, and KLF4 were detected. Results · CKD Ⅲ stage patients had lower serum miR-155 expression compared with healthy individuals and CKD Ⅰ stage patients. In cell experiment, the expression of SMα-actin, collagen Ⅰ, collagen Ⅳ and KLF4 renal tubular epithelial cells decreased and the expression of E-cadherin increased after being transfected with miR-155 mimic, as compared with the control group. Results of the luciferase assay showed that miR-155 significantly decreased the luciferase activity of KLF4-3'-UTR plasmid. Expressions of SMα-actin, collagen Ⅰ, and collagen Ⅳ in renal tubular epithelial cells were increased and the expression E-cadherin was decreased after being transfected with plasmids with over-expressed KLF4. Conclusion · CKD patients have low serum miR-155 expression. miR-155 can inhibit EMT in renal tubular epithelial cells by down-regulating KLF4.

Key words: chronic kidney disease, miR-155, Krüppel-like factor 4, epithelial-mesenchymal transition