Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1988 Jul 1;107(1):307–319. doi: 10.1083/jcb.107.1.307

Diverse effects of fibronectin and laminin on phenotypic properties of cultured arterial smooth muscle cells

PMCID: PMC2115158  PMID: 2455726

Abstract

Plasma fibronectin promotes modulation of rat arterial smooth muscle cells from a contractile to a synthetic phenotype during the first few days in primary culture. This process includes cell adhesion and spreading, loss of myofilaments, and formation of a widespread rough endoplasmic reticulum and a prominent Golgi complex. The structural reorganization is accompanied by activation of overall RNA and protein synthesis. Moreover, the cells gain the ability to replicate their DNA and divide in response to platelet-derived growth factor. Here, it is demonstrated that the power of fibronectin to bring about this change in the differentiated properties of the smooth muscle cells resides in a 105-kD cell-binding fragment, whereas a 70-kD collagen-binding fragment and a 31-kD heparin-binding fragment are inactive in this respect. Laminin, another adhesive glycoprotein and a component of the basement membrane that normally surrounds arterial smooth muscle, was contrarily found to maintain the cells in a contractile phenotype. However, with increasing time more and more cells went through the modulation into a synthetic phenotype. This "catch-up" was counteracted by a peptide that contained the cell-attachment sequence of fibronectin (Arg-Gly-Asp-Ser). Hence, it is possible that the delayed modulation on laminin was due to production of fibronectin by the cells themselves. In support of this notion, fibronectin isolated from smooth muscle cultures was found to be as effective as plasma fibronectin in stimulating the phenotypic modulation. Moreover, using a combination of chemical, immunochemical, and immunocytochemical methods, it was demonstrated that the cells secreted fibronectin as well as laminin at an increasing rate during the first 4 d in primary culture and, notably, cells cultured on laminin produced more fibronectin than cells cultured on fibronectin. Newly synthesized fibronectin was incorporated into a network of pericellular and intercellular fibrils, whereas laminin formed a more diffuse layer covering the cells in a basement membrane-like manner. Taken together, the findings suggest diverse roles for fibronectin and laminin in the control of the differentiated properties of arterial smooth muscle cells. They further indicate that the ability of arterial smooth muscle cells to produce fibronectin and laminin early in primary culture is not directly related to the phenotypic state as determined morphologically and by measurement of overall rates of RNA and protein synthesis. This may be due to the cells being able to sense the macromolecular composition of the pericellular matrix and to modify their secretory activity accordingly.(ABSTRACT TRUNCATED AT 400 WORDS)

Full Text

The Full Text of this article is available as a PDF (4.2 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
  2. Carlsson R., Engvall E., Freeman A., Ruoslahti E. Laminin and fibronectin in cell adhesion: enhanced adhesion of cells from regenerating liver to laminin. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2403–2406. doi: 10.1073/pnas.78.4.2403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Chamley-Campbell J., Campbell G. R., Ross R. The smooth muscle cell in culture. Physiol Rev. 1979 Jan;59(1):1–61. doi: 10.1152/physrev.1979.59.1.1. [DOI] [PubMed] [Google Scholar]
  4. Couchman J. R., Hök M., Rees D. A., Timpl R. Adhesion, growth, and matrix production by fibroblasts on laminin substrates. J Cell Biol. 1983 Jan;96(1):177–183. doi: 10.1083/jcb.96.1.177. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Dufour S., Duband J. L., Thiéry J. P. Role of a major cell-substratum adhesion system in cell behavior and morphogenesis. Biol Cell. 1986;58(1):1–13. doi: 10.1111/j.1768-322x.1986.tb00483.x. [DOI] [PubMed] [Google Scholar]
  6. Graf J., Iwamoto Y., Sasaki M., Martin G. R., Kleinman H. K., Robey F. A., Yamada Y. Identification of an amino acid sequence in laminin mediating cell attachment, chemotaxis, and receptor binding. Cell. 1987 Mar 27;48(6):989–996. doi: 10.1016/0092-8674(87)90707-0. [DOI] [PubMed] [Google Scholar]
  7. Hedin U., Thyberg J. Plasma fibronectin promotes modulation of arterial smooth-muscle cells from contractile to synthetic phenotype. Differentiation. 1987;33(3):239–246. doi: 10.1111/j.1432-0436.1987.tb01563.x. [DOI] [PubMed] [Google Scholar]
  8. Heldin C. H., Johnsson A., Ek B., Wennergren S., Rönnstrand L., Hammacher A., Faulders B., Wasteson A., Westermark B. Purification of human platelet-derived growth factor. Methods Enzymol. 1987;147:3–13. doi: 10.1016/0076-6879(87)47094-8. [DOI] [PubMed] [Google Scholar]
  9. Heldin C. H., Wasteson A., Westermark B. Platelet-derived growth factor. Mol Cell Endocrinol. 1985 Mar;39(3):169–187. doi: 10.1016/0303-7207(85)90061-9. [DOI] [PubMed] [Google Scholar]
  10. Holderbaum D., Ehrhart L. A. Substratum influence on collagen and fibronectin biosynthesis by arterial smooth muscle cells in vitro. J Cell Physiol. 1986 Feb;126(2):216–224. doi: 10.1002/jcp.1041260210. [DOI] [PubMed] [Google Scholar]
  11. Hynes R. Molecular biology of fibronectin. Annu Rev Cell Biol. 1985;1:67–90. doi: 10.1146/annurev.cb.01.110185.000435. [DOI] [PubMed] [Google Scholar]
  12. Izzard C. S., Radinsky R., Culp L. A. Substratum contacts and cytoskeletal reorganization of BALB/c 3T3 cells on a cell-binding fragment and heparin-binding fragments of plasma fibronectin. Exp Cell Res. 1986 Aug;165(2):320–336. doi: 10.1016/0014-4827(86)90586-0. [DOI] [PubMed] [Google Scholar]
  13. Johansson S., Hök M. Substrate adhesion of rat hepatocytes: on the mechanism of attachment to fibronectin. J Cell Biol. 1984 Mar;98(3):810–817. doi: 10.1083/jcb.98.3.810. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Johansson S., Kjellén L., Hök M., Timpl R. Substrate adhesion of rat hepatocytes: a comparison of laminin and fibronectin as attachment proteins. J Cell Biol. 1981 Jul;90(1):260–264. doi: 10.1083/jcb.90.1.260. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kühl U., Timpl R., von der Mark K. Synthesis of type IV collagen and laminin in cultures of skeletal muscle cells and their assembly on the surface of myotubes. Dev Biol. 1982 Oct;93(2):344–354. doi: 10.1016/0012-1606(82)90122-1. [DOI] [PubMed] [Google Scholar]
  16. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  17. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  18. McDonald J. A., Quade B. J., Broekelmann T. J., LaChance R., Forsman K., Hasegawa E., Akiyama S. Fibronectin's cell-adhesive domain and an amino-terminal matrix assembly domain participate in its assembly into fibroblast pericellular matrix. J Biol Chem. 1987 Mar 5;262(7):2957–2967. [PubMed] [Google Scholar]
  19. Nilsson J., Sjölund M., Palmberg L., Thyberg J., Heldin C. H. Arterial smooth muscle cells in primary culture produce a platelet-derived growth factor-like protein. Proc Natl Acad Sci U S A. 1985 Jul;82(13):4418–4422. doi: 10.1073/pnas.82.13.4418. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Palmberg L., Sjölund M., Thyberg J. Phenotype modulation in primary cultures of arterial smooth-muscle cells: reorganization of the cytoskeleton and activation of synthetic activities. Differentiation. 1985;29(3):275–283. doi: 10.1111/j.1432-0436.1985.tb00327.x. [DOI] [PubMed] [Google Scholar]
  21. Perris R., Johansson S. Amphibian neural crest cell migration on purified extracellular matrix components: a chondroitin sulfate proteoglycan inhibits locomotion on fibronectin substrates. J Cell Biol. 1987 Dec;105(6 Pt 1):2511–2521. doi: 10.1083/jcb.105.6.2511. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Pierschbacher M. D., Ruoslahti E. Cell attachment activity of fibronectin can be duplicated by small synthetic fragments of the molecule. Nature. 1984 May 3;309(5963):30–33. doi: 10.1038/309030a0. [DOI] [PubMed] [Google Scholar]
  23. Ross R., Raines E. W., Bowen-Pope D. F. The biology of platelet-derived growth factor. Cell. 1986 Jul 18;46(2):155–169. doi: 10.1016/0092-8674(86)90733-6. [DOI] [PubMed] [Google Scholar]
  24. Ross R. The pathogenesis of atherosclerosis--an update. N Engl J Med. 1986 Feb 20;314(8):488–500. doi: 10.1056/NEJM198602203140806. [DOI] [PubMed] [Google Scholar]
  25. Ruoslahti E., Hayman E. G., Pierschbacher M. D. Extracellular matrices and cell adhesion. Arteriosclerosis. 1985 Nov-Dec;5(6):581–594. doi: 10.1161/01.atv.5.6.581. [DOI] [PubMed] [Google Scholar]
  26. Ruoslahti E., Hayman E. G., Pierschbacher M., Engvall E. Fibronectin: purification, immunochemical properties, and biological activities. Methods Enzymol. 1982;82(Pt A):803–831. doi: 10.1016/0076-6879(82)82103-4. [DOI] [PubMed] [Google Scholar]
  27. Schwartz S. M., Campbell G. R., Campbell J. H. Replication of smooth muscle cells in vascular disease. Circ Res. 1986 Apr;58(4):427–444. doi: 10.1161/01.res.58.4.427. [DOI] [PubMed] [Google Scholar]
  28. Sejersen T., Betsholtz C., Sjölund M., Heldin C. H., Westermark B., Thyberg J. Rat skeletal myoblasts and arterial smooth muscle cells express the gene for the A chain but not the gene for the B chain (c-sis) of platelet-derived growth factor (PDGF) and produce a PDGF-like protein. Proc Natl Acad Sci U S A. 1986 Sep;83(18):6844–6848. doi: 10.1073/pnas.83.18.6844. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sjölund M., Hedin U., Sejersen T., Heldin C. H., Thyberg J. Arterial smooth muscle cells express platelet-derived growth factor (PDGF) A chain mRNA, secrete a PDGF-like mitogen, and bind exogenous PDGF in a phenotype- and growth state-dependent manner. J Cell Biol. 1988 Feb;106(2):403–413. doi: 10.1083/jcb.106.2.403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Sjölund M., Madsen K., von der Mark K., Thyberg J. Phenotype modulation in primary cultures of smooth-muscle cells from rat aorta. Synthesis of collagen and elastin. Differentiation. 1986;32(2):173–180. doi: 10.1111/j.1432-0436.1986.tb00570.x. [DOI] [PubMed] [Google Scholar]
  31. Thyberg J., Palmberg L., Nilsson J., Ksiazek T., Sjölund M. Phenotype modulation in primary cultures of arterial smooth muscle cells. On the role of platelet-derived growth factor. Differentiation. 1983;25(2):156–167. doi: 10.1111/j.1432-0436.1984.tb01351.x. [DOI] [PubMed] [Google Scholar]
  32. Timpl R., Dziadek M. Structure, development, and molecular pathology of basement membranes. Int Rev Exp Pathol. 1986;29:1–112. [PubMed] [Google Scholar]
  33. Timpl R., Rohde H., Robey P. G., Rennard S. I., Foidart J. M., Martin G. R. Laminin--a glycoprotein from basement membranes. J Biol Chem. 1979 Oct 10;254(19):9933–9937. [PubMed] [Google Scholar]
  34. Voss B., Rauterberg J. Localization of collagen types I, III, IV and V, fibronectin and laminin in human arteries by the indirect immunofluorescence method. Pathol Res Pract. 1986 Oct;181(5):568–575. doi: 10.1016/S0344-0338(86)80151-0. [DOI] [PubMed] [Google Scholar]
  35. Warburton M. J., Ferns S. A., Rudland P. S. Enhanced synthesis of basement membrane proteins during the differentiation of rat mammary tumour epithelial cells into myoepithelial-like cells in vitro. Exp Cell Res. 1982 Feb;137(2):373–380. doi: 10.1016/0014-4827(82)90038-6. [DOI] [PubMed] [Google Scholar]
  36. Woods A., Couchman J. R., Johansson S., Hök M. Adhesion and cytoskeletal organisation of fibroblasts in response to fibronectin fragments. EMBO J. 1986 Apr;5(4):665–670. doi: 10.1002/j.1460-2075.1986.tb04265.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Yamada K. M., Akiyama S. K., Hasegawa T., Hasegawa E., Humphries M. J., Kennedy D. W., Nagata K., Urushihara H., Olden K., Chen W. T. Recent advances in research on fibronectin and other cell attachment proteins. J Cell Biochem. 1985;28(2):79–97. doi: 10.1002/jcb.240280202. [DOI] [PubMed] [Google Scholar]
  38. Yamada K. M., Kennedy D. W. Dualistic nature of adhesive protein function: fibronectin and its biologically active peptide fragments can autoinhibit fibronectin function. J Cell Biol. 1984 Jul;99(1 Pt 1):29–36. doi: 10.1083/jcb.99.1.29. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Yamada K. M., Yamada S. S., Pastan I. The major cell surface glycoprotein of chick embryo fibroblasts is an agglutinin. Proc Natl Acad Sci U S A. 1975 Aug;72(8):3158–3162. doi: 10.1073/pnas.72.8.3158. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES