title: Cytoskeletal rearrangement and signal transduction in TGF-beta1-stimulated mesangial cell collagen accumulation.
abstract: TGF-beta1 has been implicated in glomerular extracellular matrix accumulation, although the precise cellular mechanism(s) by which this occurs is not fully understood. The authors have previously shown that the Smad signaling pathway is present and functional in human glomerular mesangial cells and plays a role in activating type I collagen gene expression. It also was determined that TGF-beta1 activates ERK mitogen-activated protein kinase in mesangial cells to enhance Smad activation and collagen expression. Here, it was shown that TGF-beta1 rapidly induces cytoskeletal rearrangement in human mesangial cells, stimulating smooth muscle alpha-actin detection in stress fibers and promoting focal adhesion complex assembly and redistribution. Disrupting the actin cytoskeleton with cytochalasin D (Cyto D) selectively decreased basal and TGF-beta1-induced cell-layer collagen I and IV accumulation. The balance of matrix metalloproteinases (MMP) and inhibitors was altered by Cyto D or TGF-beta1 alone, increasing MMP activity, increasing MMP-1 expression, and decreasing tissue inhibitor of matrix metalloproteinase-2 expression. Cyto D also decreased basal and TGF-beta1-stimulated alpha1(I) collagen mRNA but did not inhibit TGF-beta-stimulated alpha1(IV) mRNA expression. A similar decrease in alpha1(I) mRNA expression caused by the actin polymerization inhibitor latrunculin B was partially blocked by the addition of jasplakinolide, which promotes actin assembly. The Rho-family GTPase inhibitor C. difficile toxin B or the Rho-associated kinase inhibitor Y-27632 also blocked TGF-beta1-stimulated alpha1(I) mRNA expression. Cytoskeletal disruption reduced Smad2 phosphorylation but had little effect on mRNA stability, TGF-beta receptor number, or receptor affinity. Thus, TGF-beta1-mediated collagen I accumulation is associated with cytoskeletal rearrangement and Rho-GTPase signaling.
authors.affiliation: Department of Pediatrics, Northwestern University Medical School, Chicago, Illinois, USA. s-hubchak@northwestern.edu
authors.forename: Susan C
authors.initials: SC
authors.lastname: Hubchak
authors.forename: Constance E
authors.initials: CE
authors.lastname: Runyan
authors.forename: Jeffrey I
authors.initials: JI
authors.lastname: Kreisberg
authors.forename: H William
authors.initials: HW
authors.lastname: Schnaper
mesh_terms: D000199:Actins / Q000378:metabolism; D015152:Blotting, Northern; D015153:Blotting, Western; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism*; D024042:Collagen Type I / Q000378:metabolism; D024141:Collagen Type IV / Q000378:metabolism; D015638:Cytochalasin D / Q000494:pharmacology; D003599:Cytoskeleton / Q000378:metabolism*; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005920:Glomerular Mesangium / Q000378:metabolism; D006801:Humans; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry; D020782:Matrix Metalloproteinases / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction; D013844:Thiazoles / Q000494:pharmacology; D053778:Thiazolidines; D013997:Time Factors; D019714:Tissue Inhibitor of Metalloproteinases / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D053773:Transforming Growth Factor beta1
chemical_list: D000199:Actins; D019086:Bridged Bicyclo Compounds, Heterocyclic; D024042:Collagen Type I; D024141:Collagen Type IV; D018076:DNA, Complementary; D012333:RNA, Messenger; D011994:Recombinant Proteins; C508960:TGFB1 protein, human; D013844:Thiazoles; D053778:Thiazolidines; D019714:Tissue Inhibitor of Metalloproteinases; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D015638:Cytochalasin D; D012313:RNA; D003094:Collagen; D020782:Matrix Metalloproteinases; C037068:latrunculin B