Search tips
Search criteria 


Logo of jcinvestThe Journal of Clinical Investigation
J Clin Invest. 1996 November 15; 98(10): 2218–2227.
PMCID: PMC507670

Osteopontin is produced by rat cardiac fibroblasts and mediates A(II)-induced DNA synthesis and collagen gel contraction.


Angiotensin II (AII) is a critical factor in cardiac remodeling which involves hypertrophy, fibroblast proliferation, and extracellular matrix production. However, little is known about the mechanism by which AII accelerates these responses. Osteopontin is an acidic phosphoprotein with RGD (arginine-glycine-aspartate) sequences that are involved in the vascular smooth muscle cell remodeling process. We identified the presence of osteopontin mRNA and protein in cultured rat cardiac fibroblasts and its prominent regulation by AII (10(-11) M). Osteopontin message levels were increased fourfold (P < 0.01) and protein fivefold (P < 0.05) at 24 h after addition of AII (10(-7) M). This response was inhibited by the AT1 receptor blocker, losartan. Osteopontin mRNA levels were increased in hypertrophied ventricles from animals with renovascular hypertension (1.6-fold, P < 0.05) and aortic banding (2.9-fold, P < 0.05). To examine the function of osteopontin, we determined its effects on (a) the ability of cardiac fibroblasts to contract three-dimensional collagen gels and (b) cardiac fibroblast growth. A monoclonal antibody against osteopontin partially blocked AII-induced three-dimensional collagen gel contraction by cardiac fibroblasts (64+/-4 vs. 86+/-5% in the presence of antibody, P < 0.05), while osteopontin itself promoted contraction of the gels by fibroblasts (71+/-5%, P < 0.05 compared with control). Either a monoclonal antibody against beta3 integrin which is a ligand for osteopontin or the RGD peptide blocked both AII and osteopontin-induced collagen gel contraction. Thus, the osteopontin RGD sequence binds to beta3 integrins on the fibroblast to promote fibroblast binding to collagen. All induced a threefold increase in DNA synthesis of cardiac fibroblasts, which was completely blocked by antibodies against osteopontin and beta3 integrin, or by RGD peptide, but not by controls. Thus, All-induced growth of cardiac fibroblasts also requires osteopontin engagement of the beta3 integrin. Taken together, these results provide the first evidence that osteopontin is a potentially important mediator of AII regulation of cardiac fibroblast behavior in the cardiac remodeling process.

Full Text

The Full Text of this article is available as a PDF (475K).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Denhardt DT, Guo X. Osteopontin: a protein with diverse functions. FASEB J. 1993 Dec;7(15):1475–1482. [PubMed]
  • Giachelli CM, Bae N, Almeida M, Denhardt DT, Alpers CE, Schwartz SM. Osteopontin is elevated during neointima formation in rat arteries and is a novel component of human atherosclerotic plaques. J Clin Invest. 1993 Oct;92(4):1686–1696. [PMC free article] [PubMed]
  • Liaw L, Almeida M, Hart CE, Schwartz SM, Giachelli CM. Osteopontin promotes vascular cell adhesion and spreading and is chemotactic for smooth muscle cells in vitro. Circ Res. 1994 Feb;74(2):214–224. [PubMed]
  • Chen Y, Bal BS, Gorski JP. Calcium and collagen binding properties of osteopontin, bone sialoprotein, and bone acidic glycoprotein-75 from bone. J Biol Chem. 1992 Dec 5;267(34):24871–24878. [PubMed]
  • Butler WT. The nature and significance of osteopontin. Connect Tissue Res. 1989;23(2-3):123–136. [PubMed]
  • Liaw L, Skinner MP, Raines EW, Ross R, Cheresh DA, Schwartz SM, Giachelli CM. The adhesive and migratory effects of osteopontin are mediated via distinct cell surface integrins. Role of alpha v beta 3 in smooth muscle cell migration to osteopontin in vitro. J Clin Invest. 1995 Feb;95(2):713–724. [PMC free article] [PubMed]
  • Choi ET, Engel L, Callow AD, Sun S, Trachtenberg J, Santoro S, Ryan US. Inhibition of neointimal hyperplasia by blocking alpha V beta 3 integrin with a small peptide antagonist GpenGRGDSPCA. J Vasc Surg. 1994 Jan;19(1):125–134. [PubMed]
  • Murry CE, Giachelli CM, Schwartz SM, Vracko R. Macrophages express osteopontin during repair of myocardial necrosis. Am J Pathol. 1994 Dec;145(6):1450–1462. [PubMed]
  • Williams EB, Halpert I, Wickline S, Davison G, Parks WC, Rottman JN. Osteopontin expression is increased in the heritable cardiomyopathy of Syrian hamsters. Circulation. 1995 Aug 15;92(4):705–709. [PubMed]
  • Singh K, Balligand JL, Fischer TA, Smith TW, Kelly RA. Glucocorticoids increase osteopontin expression in cardiac myocytes and microvascular endothelial cells. Role in regulation of inducible nitric oxide synthase. J Biol Chem. 1995 Nov 24;270(47):28471–28478. [PubMed]
  • Linz W, Schölkens BA, Ganten D. Converting enzyme inhibition specifically prevents the development and induces regression of cardiac hypertrophy in rats. Clin Exp Hypertens A. 1989;11(7):1325–1350. [PubMed]
  • Dahlöf B, Pennert K, Hansson L. Reversal of left ventricular hypertrophy in hypertensive patients. A metaanalysis of 109 treatment studies. Am J Hypertens. 1992 Feb;5(2):95–110. [PubMed]
  • Sadoshima J, Xu Y, Slayter HS, Izumo S. Autocrine release of angiotensin II mediates stretch-induced hypertrophy of cardiac myocytes in vitro. Cell. 1993 Dec 3;75(5):977–984. [PubMed]
  • Baker KM, Booz GW, Dostal DE. Cardiac actions of angiotensin II: Role of an intracardiac renin-angiotensin system. Annu Rev Physiol. 1992;54:227–241. [PubMed]
  • Morkin E, Ashford TP. Myocardial DNA synthesis in experimental cardiac hypertrophy. Am J Physiol. 1968 Dec;215(6):1409–1413. [PubMed]
  • Weber KT, Janicki JS, Shroff SG, Pick R, Chen RM, Bashey RI. Collagen remodeling of the pressure-overloaded, hypertrophied nonhuman primate myocardium. Circ Res. 1988 Apr;62(4):757–765. [PubMed]
  • Villarreal FJ, Kim NN, Ungab GD, Printz MP, Dillmann WH. Identification of functional angiotensin II receptors on rat cardiac fibroblasts. Circulation. 1993 Dec;88(6):2849–2861. [PubMed]
  • Matsubara H, Kanasaki M, Murasawa S, Tsukaguchi Y, Nio Y, Inada M. Differential gene expression and regulation of angiotensin II receptor subtypes in rat cardiac fibroblasts and cardiomyocytes in culture. J Clin Invest. 1994 Apr;93(4):1592–1601. [PMC free article] [PubMed]
  • Iwami K, Ashizawa N, Do YS, Graf K, Hsueh WA. Comparison of ANG II with other growth factors on Egr-1 and matrix gene expression in cardiac fibroblasts. Am J Physiol. 1996 Jun;270(6 Pt 2):H2100–H2107. [PubMed]
  • Schieffer B, Wirger A, Meybrunn M, Seitz S, Holtz J, Riede UN, Drexler H. Comparative effects of chronic angiotensin-converting enzyme inhibition and angiotensin II type 1 receptor blockade on cardiac remodeling after myocardial infarction in the rat. Circulation. 1994 May;89(5):2273–2282. [PubMed]
  • Buchanan TA, Sipos GF, Gadalah S, Yip KP, Marsh DJ, Hsueh W, Bergman RN. Glucose tolerance and insulin action in rats with renovascular hypertension. Hypertension. 1991 Sep;18(3):341–347. [PubMed]
  • Chomczynski P, Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. [PubMed]
  • Harpold MM, Evans RM, Salditt-Georgieff M, Darnell JE. Production of mRNA in Chinese hamster cells: relationship of the rate of synthesis to the cytoplasmic concentration of nine specific mRNA sequences. Cell. 1979 Aug;17(4):1025–1035. [PubMed]
  • Helfrich MH, Nesbitt SA, Horton MA. Integrins on rat osteoclasts: characterization of two monoclonal antibodies (F4 and F11) to rat beta 3. J Bone Miner Res. 1992 Mar;7(3):345–351. [PubMed]
  • Sollott SJ, Cheng L, Pauly RR, Jenkins GM, Monticone RE, Kuzuya M, Froehlich JP, Crow MT, Lakatta EG, Rowinsky EK, et al. Taxol inhibits neointimal smooth muscle cell accumulation after angioplasty in the rat. J Clin Invest. 1995 Apr;95(4):1869–1876. [PMC free article] [PubMed]
  • Guidry C, Grinnell F. Contraction of hydrated collagen gels by fibroblasts: evidence for two mechanisms by which collagen fibrils are stabilized. Coll Relat Res. 1987 Feb;6(6):515–529. [PubMed]
  • Gardner HA, Berse B, Senger DR. Specific reduction in osteopontin synthesis by antisense RNA inhibits the tumorigenicity of transformed Rat1 fibroblasts. Oncogene. 1994 Aug;9(8):2321–2326. [PubMed]
  • Brown LF, Berse B, Van de Water L, Papadopoulos-Sergiou A, Perruzzi CA, Manseau EJ, Dvorak HF, Senger DR. Expression and distribution of osteopontin in human tissues: widespread association with luminal epithelial surfaces. Mol Biol Cell. 1992 Oct;3(10):1169–1180. [PMC free article] [PubMed]
  • Harter LV, Hruska KA, Duncan RL. Human osteoblast-like cells respond to mechanical strain with increased bone matrix protein production independent of hormonal regulation. Endocrinology. 1995 Feb;136(2):528–535. [PubMed]
  • Everett AD, Tufro-McReddie A, Fisher A, Gomez RA. Angiotensin receptor regulates cardiac hypertrophy and transforming growth factor-beta 1 expression. Hypertension. 1994 May;23(5):587–592. [PubMed]
  • Mamuya WS, Brecher P. Fibronectin expression in the normal and hypertrophic rat heart. J Clin Invest. 1992 Feb;89(2):392–401. [PMC free article] [PubMed]
  • Boluyt MO, O'Neill L, Meredith AL, Bing OH, Brooks WW, Conrad CH, Crow MT, Lakatta EG. Alterations in cardiac gene expression during the transition from stable hypertrophy to heart failure. Marked upregulation of genes encoding extracellular matrix components. Circ Res. 1994 Jul;75(1):23–32. [PubMed]
  • Gillery P, Maquart FX, Borel JP. Fibronectin dependence of the contraction of collagen lattices by human skin fibroblasts. Exp Cell Res. 1986 Nov;167(1):29–37. [PubMed]
  • Mukherjee BB, Nemir M, Beninati S, Cordella-Miele E, Singh K, Chackalaparampil I, Shanmugam V, DeVouge MW, Mukherjee AB. Interaction of osteopontin with fibronectin and other extracellular matrix molecules. Ann N Y Acad Sci. 1995 Apr 21;760:201–212. [PubMed]
  • Cheresh DA, Smith JW, Cooper HM, Quaranta V. A novel vitronectin receptor integrin (alpha v beta x) is responsible for distinct adhesive properties of carcinoma cells. Cell. 1989 Apr 7;57(1):59–69. [PubMed]
  • Juliano RL, Haskill S. Signal transduction from the extracellular matrix. J Cell Biol. 1993 Feb;120(3):577–585. [PMC free article] [PubMed]
  • Hynes RO. Integrins: versatility, modulation, and signaling in cell adhesion. Cell. 1992 Apr 3;69(1):11–25. [PubMed]
  • Burgess ML, Carver WE, Terracio L, Wilson SP, Wilson MA, Borg TK. Integrin-mediated collagen gel contraction by cardiac fibroblasts. Effects of angiotensin II. Circ Res. 1994 Feb;74(2):291–298. [PubMed]
  • Knowlton KU, Michel MC, Itani M, Shubeita HE, Ishihara K, Brown JH, Chien KR. The alpha 1A-adrenergic receptor subtype mediates biochemical, molecular, and morphologic features of cultured myocardial cell hypertrophy. J Biol Chem. 1993 Jul 25;268(21):15374–15380. [PubMed]

Articles from The Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation