Activation of Calpain-2 by Mediators in Pulmonary Vascular Remodeling of Pulmonary Arterial Hypertension

Am J Respir Cell Mol Biol. 2016 Mar;54(3):384-93. doi: 10.1165/rcmb.2015-0151OC.


Calpain mediates collagen synthesis and cell proliferation and plays an important role in pulmonary vascular remodeling in pulmonary arterial hypertension (PAH). In the present study, we investigated whether and how calpain is activated by PAH mediators in pulmonary artery smooth muscle cells (PASMCs). These data show that smooth muscle-specific knockout of calpain attenuated and knockout of calpastatin potentiated pulmonary vascular remodeling and pulmonary hypertension. Treatment of PASMCs with the PAH mediators platelet-derived growth factor (PDGF), serotonin, H2O2, endothelin-1, and IL-6 caused significant increases in calpain activity, cell proliferation, and collagen-I protein level without changes in protein levels of calpain-1 and -2. The calcium chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis (acetoxymethyl ester) (BAPTA/AM) did not affect calpain activation, but the extracellular signal-regulated kinase (ERK) 1/2 inhibitor PD98059 and knocking down of calpain-2 prevented calpain activation in PAH mediator-treated PASMCs. Mass spectrometry data showed that the phosphorylation of calpain-2 at serine (Ser) 50 was increased and the phosphorylation of calpain-2 at Ser369 was decreased in PDGF-treated PASMCs. The PDGF-induced increase in Ser50 phosphorylation of calpain-2 was prevented by PD98059, whereas dephosphorylation of calpain-2 at Ser369 was blocked by the protein phosphatase 2A inhibitor fostriecin. Furthermore, smooth muscle of pulmonary arteries in PAH animal models and patients with PAH showed higher levels of phospho-Ser50-calpain-2 (P-Ser50) and lower levels of phospho-Ser369-calpain-2 (P-Ser369). These data support that calpain modulates pulmonary vascular remodeling in PAH. PAH mediator-induced activation of calpain is caused by ERK1/2-dependent phosphorylation of calpain-2 at Ser50 and protein phosphatase 2A-dependent dephosphorylation of calpain-2 at Ser369 in pulmonary vascular remodeling of PAH.

Keywords: H2O2; platelet-derived growth factor; pulmonary hypertension; serotonin; vascular smooth muscle cells.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Calcium-Binding Proteins / genetics
  • Calcium-Binding Proteins / metabolism
  • Calpain / genetics
  • Calpain / metabolism*
  • Disease Models, Animal
  • Enzyme Activation
  • Enzyme Activators / pharmacology
  • Extracellular Signal-Regulated MAP Kinases / antagonists & inhibitors
  • Extracellular Signal-Regulated MAP Kinases / metabolism
  • HEK293 Cells
  • Humans
  • Hypertension, Pulmonary / enzymology*
  • Hypertension, Pulmonary / genetics
  • Hypertension, Pulmonary / pathology
  • Hypertension, Pulmonary / physiopathology
  • Hypoxia / complications
  • Mice, Knockout
  • Muscle, Smooth, Vascular / drug effects
  • Muscle, Smooth, Vascular / enzymology*
  • Muscle, Smooth, Vascular / pathology
  • Muscle, Smooth, Vascular / physiopathology
  • Myocytes, Smooth Muscle / drug effects
  • Myocytes, Smooth Muscle / enzymology*
  • Myocytes, Smooth Muscle / pathology
  • Phosphorylation
  • Protein Kinase Inhibitors / pharmacology
  • Protein Phosphatase 2 / antagonists & inhibitors
  • Protein Phosphatase 2 / metabolism
  • Pulmonary Artery / drug effects
  • Pulmonary Artery / enzymology
  • Pulmonary Artery / pathology
  • Pulmonary Artery / physiopathology
  • RNA Interference
  • Signal Transduction
  • Transfection
  • Vascular Remodeling* / drug effects


  • Calcium-Binding Proteins
  • Enzyme Activators
  • Protein Kinase Inhibitors
  • calpastatin
  • Extracellular Signal-Regulated MAP Kinases
  • Protein Phosphatase 2
  • Calpain
  • Capns1 protein, mouse
  • CAPN1 protein, human
  • CAPN2 protein, human
  • Capn2 protein, mouse