title: Effect of anaphylatoxin C3a, C5a on the tubular epithelial-myofibroblast transdifferentiation in vitro.
abstract: BACKGROUND
  Tubulointerstitial renal fibrosis is the common end point of progressive kidney diseases, and tubular epithelial-myofibroblast transdifferentiation (TEMT) plays a key role in the progress of tubulointerstitial renal fibrosis. Anaphylatoxin C3a and C5a are identified as novel profibrotic factors in renal disease and as potential new therapeutic targets. The aim of this study was to investigate whether C3a, C5a can regulate TEMT by transforming growth factor-β1 (TGF-β)/connective tissue growth factor (CTGF) signaling pathway and the effects of C3a and C5a receptor antagonists (C3aRA and C5aRA) on C3a- and C5a-induced TEMT.
  
  
  METHODS
  HK-2 cells were divided into C3a and C5a groups which were subdivided into four subgroups: control group, 10 ng/ml TGF-β1 group, 50 nmol/L C3a group, 50 nmol/L C3a plus 1 µmol/L C3aRA group; control group, 10 ng/ml TGF-β1 group, 50 nmol/L C5a group, 50 nmol/L C5a plus 2.5 µmol/L C5aRA group. TGF-β1 receptor antagonist (TGF-β1RA) 10 µg/ml was used to investigate the mechanism of C3a- and C5a-induced TEMT. Electron microscopy was used to observe the morphological changes. Immunocytochemistry staining, real-time PCR and Western blotting were used to detect the expressions of a smooth muscle actin (α-SMA), E-cadherin, Col-I, C3a receptor (C3aR), C5aR, CTGF and TGF-β1.
  
  
  RESULTS
  HK-2 cells cultured with C3a and C5a for 72 hours exhibited strong staining of α-SMA, lost the positive staining of E-cadherin, and showed a slightly spindle-like shape and loss of microvilli on the cell surface. The expressions of α-SMA, E-cadherin, Col-I, C3aR, C5aR, TGF-β1 and CTGF in C3a- and C5a-treated groups were higher than normal control group (P < 0.05). C3aRA and C5aRA inhibited the expressions of α-SMA, Col-I, C3aR, C5aR, and up-regulated the expression of E-cadherin (P < 0.05). TGF-β1 and CTGF mRNA expressions induced by C3a and C5a were partly blocked by TGF-β1RA (P < 0.05).
  
  
  CONCLUSIONS
  C3a and C5a can induce TEMT via the up-regulations of C3aR and C5aR mRNA and the activation of TGF-β1/CTGF signaling pathway in vitro.
authors.affiliation: Division of Nephrology, West China Hospital of Sichuan University, Chengdu, Sichuan 610041, China.
authors.forename: Fang
authors.initials: F
authors.lastname: Liu
authors.forename: Rong
authors.initials: R
authors.lastname: Gou
authors.forename: Jun
authors.initials: J
authors.lastname: Huang
authors.forename: Ping
authors.initials: P
authors.lastname: Fu
authors.forename: Feng
authors.initials: F
authors.lastname: Chen
authors.forename: Wen-Xing
authors.initials: WX
authors.lastname: Fan
authors.forename: You-Qun
authors.initials: YQ
authors.lastname: Huang
authors.forename: Li
authors.initials: L
authors.lastname: Zang
authors.forename: Min
authors.initials: M
authors.lastname: Wu
authors.forename: Hong-Yu
authors.initials: HY
authors.lastname: Qiu
authors.forename: Da-Peng
authors.initials: DP
authors.lastname: Wei
mesh_terms: D015153:Blotting, Western; D015820:Cadherins / Q000235:genetics; D002460:Cell Line; D054338:Cell Transdifferentiation / Q000187:drug effects*; D015926:Complement C3a / Q000494:pharmacology*; D015936:Complement C5a / Q000494:pharmacology*; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects / Q000648:ultrastructure; D006801:Humans; D007150:Immunohistochemistry; D008855:Microscopy, Electron, Scanning; D058628:Myofibroblasts / Q000166:cytology* / Q000187:drug effects / Q000648:ultrastructure; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D015820:Cadherins; D015926:Complement C3a; D015936:Complement C5a