Calpain-2 activates Akt via TGF-β1-mTORC2 pathway in pulmonary artery smooth muscle cells.
Calpain is a family of calcium-dependent nonlysosomal neutral cysteine endopeptidases. Akt is a serine/threonine kinase that belongs to AGC kinases and plays important roles in cell survival, growth, proliferation, angiogenesis, and cell metabolism. Both calpain and Akt are the downstream signaling molecules of platelet-derived growth factor (PDGF) and mediate PDGF-induced collagen synthesis and proliferation of pulmonary artery smooth muscle cells (PASMCs) in pulmonary vascular remodeling. We found that inhibitions of calpain-2 by using calpain inhibitor MDL28170 and calpain-2 small interfering RNA attenuated Akt phosphorylations at serine-473 (S473) and threonine-308 (T308), as well as collagen synthesis and cell proliferation of PASMCs induced by PDGF. Overexpression of calpain-2 in PASMCs induced dramatic increases in Akt phosphorylations at S473 and T308. Moreover, knockout of calpain attenuated Akt phosphorylations at S473 and T308 in smooth muscle of pulmonary arterioles of mice with chronic hypoxic pulmonary hypertension. The cell-permeable-specific transforming growth factor (TGF)-β receptor inhibitor SB431542 attenuated Akt phosphorylations at both S473 and T308 induced by PDGF and by overexpressed calpain-2 in PASMCs. Furthermore, SB-431452 and knocking down activin receptor-like kinase-5 significantly reduced PDGF-induced collagen synthesis and cell proliferation of PASMCs. Nevertheless, neutralizing extracellular TGF-β1 using a cell-impermeable TGF-β1 neutralizing antibody did not affect PDGF-induced Akt phosphorylations at S473 and T308. Furthermore, inhibition of mammalian target of rapamycin complex 2 (mTORC2) by knocking down its component protein Rictor prevented Akt phosphorylations at S473 and T308 induced by PDGF and by overexpressed calpain-2. These data provide first evidence supporting that calpain-2 upregulates PDGF-induced Akt phosphorylation in pulmonary vascular remodeling via an intracrine TGF-β1/mTORC2 mechanism.
Department of Pharmacology & Toxicology, Medical College of Georgia, Augusta University, Augusta, Georgia;
Prasanna
P
Abeyrathna
Department of Pharmacology & Toxicology, Medical College of Georgia, Augusta University, Augusta, Georgia;
Laszlo
L
Kovacs
Department of Pharmacology & Toxicology, Medical College of Georgia, Augusta University, Augusta, Georgia;
Weihong
W
Han
Department of Pharmacology & Toxicology, Medical College of Georgia, Augusta University, Augusta, Georgia; Department of Medicine, Medical College of Georgia, Augusta University, Augusta, Georgia; Vascular Biology Center, Medical College of Georgia, Augusta University, Augusta, Georgia; and Research Service, Charlie Norwood Veterans Affairs Medical Center, Augusta, Georgia ysu@augusta.edu.
Yunchao
Y
Su
D000818:Animals; D000077214:Becaplermin; D001549:Benzamides / Q000494:pharmacology; D002154:Calpain / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004149:Dioxoles / Q000494:pharmacology; D004151:Dipeptides / Q000494:pharmacology; D004195:Disease Models, Animal; D004789:Enzyme Activation; D006801:Humans; D006976:Hypertension, Pulmonary / Q000201:enzymology / Q000209:etiology / Q000473:pathology; D000860:Hypoxia / Q000150:complications / Q000201:enzymology; D000076225:Mechanistic Target of Rapamycin Complex 2; D018345:Mice, Knockout; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism*; D020574:Proto-Oncogene Proteins c-sis / Q000494:pharmacology; D011651:Pulmonary Artery / Q000201:enzymology; D034622:RNA Interference; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000378:metabolism; D012263:Ribonucleosides / Q000494:pharmacology; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection; D053773:Transforming Growth Factor beta1 / Q000378:metabolism*; D066253:Vascular Remodeling / Q000187:drug effects
C459179:4-(5-benzo(1,3)dioxol-5-yl-4-pyridin-2-yl-1H-imidazol-2-yl)benzamide; D001549:Benzamides; D015853:Cysteine Proteinase Inhibitors; D004149:Dioxoles; D004151:Dipeptides; D046912:Multiprotein Complexes; D047428:Protein Kinase Inhibitors; D020574:Proto-Oncogene Proteins c-sis; D018125:Receptors, Transforming Growth Factor beta; D012263:Ribonucleosides; C508960:TGFB1 protein, human; D053773:Transforming Growth Factor beta1; D000077214:Becaplermin; C023764:triciribine; D003094:Collagen; D000076225:Mechanistic Target of Rapamycin Complex 2; D051057:Proto-Oncogene Proteins c-akt; D058570:TOR Serine-Threonine Kinases; D002154:Calpain; C403235:Capn5 protein, mouse; C569870:CAPN2 protein, human; C058076:calpain inhibitor III
PDGF; Rictor; pulmonary hypertension; vascular smooth muscle cells