title: Diltiazem suppresses collagen synthesis and IL-1beta-induced TGF-beta1 production on human peritoneal mesothelial cells.
abstract: BACKGROUND
  After long-term treatment with continuous ambulatory peritoneal dialysis (CAPD), some patients may develop peritoneal fibrosis. Peritoneal mesothelial cells (PMCs) participate in the inflammatory reactions in the peritoneal cavity, and transforming growth factor-beta1 (TGF-beta1) and interleukin-1beta (IL-1beta) are involved in peritoneal fibrosis. Diltiazem is used frequently in patients with CAPD to treat hypertension. The objectives of this study were to examine the effects of diltiazem on collagen- and IL-1beta-induced TGF-beta1 production on human PMCs and the signalling pathway of diltiazem in this induction.
  
  
  METHODS
  Human PMCs were cultured from the enzymatic disaggregation of human omentum. Collagen synthesis was measured by [3H]proline incorporation into pepsin-resistant, salt-precipitated collagen. The expression of collagen I and III, and TGF-beta1 mRNA was evaluated by northern blotting. The production of TGF-beta1 by human PMCs was measured by immunoassay. The changes of intracellular calcium level after adding Fura-2-AM were measured by fluorescence spectrophotometry. Western blotting was used to assess mitogen-activated protein kinase (MAPK) signalling proteins.
  
  
  RESULTS
  We found that diltiazem (<0.2 mM) inhibited collagen I and III mRNA expression and collagen syntheses on a dose-dependent basis. Diltiazem (0.2 mM) suppressed IL-1beta- (5 ng/ml) induced TGF-beta1 production on human PMCs at both the protein and mRNA levels. Diltiazem (0.2 mM) also inhibited IL-1beta- (5 ng/ml) induced collagen I and III mRNA expression. Intracellular calcium levels did not change after the treatment with diltiazem, IL-1beta or both. The IL-1beta-treated human PMCs increased phospho-JNK (stress-activated c-Jun N-terminal kinase) and phospho-p38 MAPK expression, while diltiazem could suppress this phenomenon.
  
  
  CONCLUSIONS
  Diltiazem suppressed collagen synthesis of human PMCs and inhibited IL-1beta-induced TGF-beta1 production on human PMCs. This signalling transduction may be through p38 MAPK and JNK pathways instead of intracellular calcium. These results suggest diltiazem to be a potential therapeutic regimen in preventing peritoneal fibrosis and support further in vivo studies.
authors.affiliation: Department of Emergency Medicine, National Taiwan University Hospital and National Taiwan University College of Medicine, Taipei 100, Taiwan.
authors.forename: Cheng-Chung
authors.initials: CC
authors.lastname: Fang
authors.forename: Chung-Jen
authors.initials: CJ
authors.lastname: Yen
authors.forename: Yung-Ming
authors.initials: YM
authors.lastname: Chen
authors.forename: Tzong-Shinn
authors.initials: TS
authors.lastname: Chu
authors.forename: Ming-Tsan
authors.initials: MT
authors.lastname: Lin
authors.forename: Ju-Yeh
authors.initials: JY
authors.lastname: Yang
authors.forename: Tun-Jun
authors.initials: TJ
authors.lastname: Tsai
mesh_terms: D000704:Analysis of Variance; D015152:Blotting, Northern; D015153:Blotting, Western; D002478:Cells, Cultured; D003094:Collagen / Q000096:biosynthesis / Q000187:drug effects*; D004110:Diltiazem / Q000494:pharmacology*; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects; D005355:Fibrosis / Q000209:etiology / Q000473:pathology; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology*; D010531:Peritoneal Dialysis, Continuous Ambulatory / Q000009:adverse effects; D010537:Peritoneum / Q000166:cytology* / Q000187:drug effects; D011336:Probability; D018570:Risk Assessment; D012680:Sensitivity and Specificity; D016212:Transforming Growth Factor beta / Q000096:biosynthesis*
chemical_list: D007375:Interleukin-1; D016212:Transforming Growth Factor beta; D003094:Collagen; D004110:Diltiazem