Canstatin stimulates migration of rat cardiac fibroblasts via secretion of matrix metalloproteinase-2

Am J Physiol Cell Physiol. 2017 Mar 1;312(3):C199-C208. doi: 10.1152/ajpcell.00329.2015. Epub 2016 Dec 21.


Type IV collagen, a nonfibrillar type, is ubiquitously expressed in the basement membrane around cardiomyocytes. Canstatin, a cleaved product of α2 chain of type IV collagen, is an antiangiogenic factor. Because it has not been clarified whether canstatin exerts other biological activities in heart, we investigated the effects of canstatin on adult rat cardiac fibroblasts. Cell migration was determined by Boyden chamber assay. Western blotting was performed to detect secretion of matrix metalloproteinase-2 (MMP-2) and MMP-9 and phosphorylation of extracellular signal-regulated kinase (ERK). Localization of MMP-2 was detected by immunofluorescence staining. Canstatin (250 ng/ml) significantly increased migration, secretion, and activity of MMP-2 but not MMP-9. CTTHWGFTLC peptide, an MMP inhibitor and small interfering RNA (siRNA) against MMP-2 suppressed the canstatin-induced (250 ng/ml, 24 h) migration. Canstatin (250 ng/ml, 30 min) significantly increased phosphorylation of ERK. PD98059, a MEK inhibitor, significantly suppressed the canstatin-induced (250 ng/ml, 24 h) migration but not secretion of MMP-2. An increase in MMP-2 expression was observed in cytoplasm of the canstatin-treated (250 ng/ml) cardiac fibroblasts (within 30 min). Canstatin induced actin stress fiber formation, which was inhibited by Y-27632, a Rho-associated kinase inhibitor. Y-27632 also suppressed the canstatin-induced (250 ng/ml, 24 h) MMP-2 secretion. Canstatin (250 ng/ml, 30 min) failed to induce ERK phosphorylation in MMP-2 siRNA-treated cardiac fibroblasts. In conclusion, this study revealed a novel function of canstatin for inducing cell migration of adult rat cardiac fibroblasts at least in part by ERK phosphorylation through MMP-2 secretion, possibly via actin cytoskeletal change.

Keywords: canstatin; cardiac fibroblasts; matrix metalloproteinases; migration.

MeSH terms

  • Animals
  • Cell Movement / physiology*
  • Cells, Cultured
  • Collagen Type IV / metabolism*
  • Fibroblasts / cytology
  • Fibroblasts / physiology*
  • Heart / physiology*
  • MAP Kinase Signaling System / physiology*
  • Male
  • Matrix Metalloproteinase 2 / metabolism*
  • Myocardium / cytology
  • Rats
  • Rats, Wistar


  • COL4A2 protein, rat
  • Collagen Type IV
  • Matrix Metalloproteinase 2
  • Mmp2 protein, rat