title: Fluid shear stress and sphingosine 1-phosphate activate calpain to promote membrane type 1 matrix metalloproteinase (MT1-MMP) membrane translocation and endothelial invasion into three-dimensional collagen matrices.
abstract: The vascular endothelium continually senses and responds to biochemical and mechanical stimuli to appropriately initiate angiogenesis. We have shown previously that fluid wall shear stress (WSS) and sphingosine 1-phosphate (S1P) cooperatively initiate the invasion of human umbilical vein endothelial cells into collagen matrices (Kang, H., Bayless, K. J., and Kaunas, R. (2008) Am. J. Physiol. Heart Circ. Physiol. 295, H2087-2097). Here, we investigated the role of calpains in the regulation of endothelial cell invasion in response to WSS and S1P. Calpain inhibition significantly decreased S1P- and WSS-induced invasion. Short hairpin RNA-mediated gene silencing demonstrated that calpain 1 and 2 were required for WSS and S1P-induced invasion. Also, S1P synergized with WSS to induce invasion and to activate calpains and promote calpain membrane localization. Calpain inhibition results in a cell morphology consistent with reduced matrix proteolysis. Membrane type 1-matrix metalloproteinase (MT1-MMP) has been shown by others to regulate endothelial cell invasion, prompting us to test whether calpain acted upstream of MT1-MMP. S1P and WSS synergistically activated MT1-MMP and induced cell membrane localization of MT1-MMP in a calpain-dependent manner. Calpain activation, MT1-MMP activation and MT1-MMP membrane localization were all maximal with 5.3 dynes/cm(2) WSS and S1P treatment, which correlated with maximal invasion responses. Our data show for the first time that 5.3 dynes/cm(2) WSS in the presence of S1P combine to activate calpains, which direct MT1-MMP membrane localization to initiate endothelial sprouting into three-dimensional collagen matrices.
authors: Department of Biomedical Engineering, Texas A&M University, College Station, Texas 77843.
authors: Hojin
authors: H
authors: Kang
authors: Department of Molecular & Cellular Medicine, Texas A&M Health Science Center, College Station, Texas 77843-1114.
authors: Hyeong-Il
authors: HI
authors: Kwak
authors: Department of Biomedical Engineering, Texas A&M University, College Station, Texas 77843.
authors: Roland
authors: R
authors: Kaunas
authors: Department of Molecular & Cellular Medicine, Texas A&M Health Science Center, College Station, Texas 77843-1114. Electronic address: kbayless@medicine.tamhsc.edu.
authors: Kayla J
authors: KJ
authors: Bayless
mesh_terms: D002154:Calpain / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology; D004730:Endothelium, Vascular / Q000473:pathology; D005109:Extracellular Matrix / Q000378:metabolism; D020868:Gene Silencing; D006801:Humans; D008246:Lysophospholipids / Q000378:metabolism*; D053511:Matrix Metalloproteinase 14 / Q000378:metabolism*; D008856:Microscopy, Fluorescence / Q000379:methods; D009389:Neovascularization, Pathologic; D033081:Shear Strength; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000378:metabolism; D013314:Stress, Mechanical; D013347:Subcellular Fractions / Q000378:metabolism; D014471:Umbilical Veins / Q000166:cytology
chemical_list: D008246:Lysophospholipids; C060506:sphingosine 1-phosphate; D002154:Calpain; D053511:Matrix Metalloproteinase 14; D013110:Sphingosine