Abstract
We have used an isometric force transducer to study contraction of two types of nonmuscle cells in tissue culture. This method permits the quantitative measurement of contractile force generated by cells of defined type under the influence of external agents while allowing detailed morphological observation. Chick embryo fibroblasts (CEF), which form a contractile network inside a collagen matrix, and human umbilical vein endothelial cells (HUVE), which are located in a monolayer on the surface of the collagen matrix, were studied. CEF and HUVE in 10% FCS produce a substantial tension of 4.5 +/- 0.2 x 10(4) dynes/cm2 and 6.1 x 10(4) dynes/cm2, respectively. Both cell types contract when stimulated with thrombin, generating a force per cell cross-sectional area of approximately 10(5) dynes/cm2, a value approximately an order of magnitude less than smooth muscle. The integrity of the actin cytoskeleton is essential for force generation, as disruption of actin microfilaments with cytochalasin D results in a rapid disappearance of force. Intact microtubules appear to reduce isometric force exerted by CEF, as microtubule-disrupting drugs result in increased tension. Contraction by HUVE precedes a dramatic rearrangement of actin microfilaments from a circumferential ring to stress fibers.
Full Text
The Full Text of this article is available as a PDF (1.5 MB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Adelstein R. S., Conti M. A. Phosphorylation of platelet myosin increases actin-activated myosin ATPase activity. Nature. 1975 Aug 14;256(5518):597–598. doi: 10.1038/256597a0. [DOI] [PubMed] [Google Scholar]
- Barak L. S., Yocum R. R., Nothnagel E. A., Webb W. W. Fluorescence staining of the actin cytoskeleton in living cells with 7-nitrobenz-2-oxa-1,3-diazole-phallacidin. Proc Natl Acad Sci U S A. 1980 Feb;77(2):980–984. doi: 10.1073/pnas.77.2.980. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bell E., Ivarsson B., Merrill C. Production of a tissue-like structure by contraction of collagen lattices by human fibroblasts of different proliferative potential in vitro. Proc Natl Acad Sci U S A. 1979 Mar;76(3):1274–1278. doi: 10.1073/pnas.76.3.1274. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bárány M., Close R. I. The transformation of myosin in cross-innervated rat muscles. J Physiol. 1971 Mar;213(2):455–474. doi: 10.1113/jphysiol.1971.sp009393. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chen L. B., Buchanan J. M. Mitogenic activity of blood components. I. Thrombin and prothrombin. Proc Natl Acad Sci U S A. 1975 Jan;72(1):131–135. doi: 10.1073/pnas.72.1.131. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Danowski B. A. Fibroblast contractility and actin organization are stimulated by microtubule inhibitors. J Cell Sci. 1989 Jun;93(Pt 2):255–266. doi: 10.1242/jcs.93.2.255. [DOI] [PubMed] [Google Scholar]
- Dennerll T. J., Joshi H. C., Steel V. L., Buxbaum R. E., Heidemann S. R. Tension and compression in the cytoskeleton of PC-12 neurites. II: Quantitative measurements. J Cell Biol. 1988 Aug;107(2):665–674. doi: 10.1083/jcb.107.2.665. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Elson E. L. Cellular mechanics as an indicator of cytoskeletal structure and function. Annu Rev Biophys Biophys Chem. 1988;17:397–430. doi: 10.1146/annurev.bb.17.060188.002145. [DOI] [PubMed] [Google Scholar]
- Fukui Y., Lynch T. J., Brzeska H., Korn E. D. Myosin I is located at the leading edges of locomoting Dictyostelium amoebae. Nature. 1989 Sep 28;341(6240):328–331. doi: 10.1038/341328a0. [DOI] [PubMed] [Google Scholar]
- Giloh H., Sedat J. W. Fluorescence microscopy: reduced photobleaching of rhodamine and fluorescein protein conjugates by n-propyl gallate. Science. 1982 Sep 24;217(4566):1252–1255. doi: 10.1126/science.7112126. [DOI] [PubMed] [Google Scholar]
- HIGTON D. I., JAMES D. W. THE FORCE OF CONTRACTION OF FULL-THICKNESS WOUNDS OF RABBIT SKIN. Br J Surg. 1964 Jun;51:462–466. doi: 10.1002/bjs.1800510616. [DOI] [PubMed] [Google Scholar]
- HOFFMANN-BERLING H. Adenosintriphosphat als Betriebsstoff von Zellbewegungen. Biochim Biophys Acta. 1954 Jun;14(2):182–194. doi: 10.1016/0006-3002(54)90157-2. [DOI] [PubMed] [Google Scholar]
- Harris A. K., Wild P., Stopak D. Silicone rubber substrata: a new wrinkle in the study of cell locomotion. Science. 1980 Apr 11;208(4440):177–179. doi: 10.1126/science.6987736. [DOI] [PubMed] [Google Scholar]
- Herlihy J. T., Murphy R. A. Length-tension relationship of smooth muscle of the hog carotid artery. Circ Res. 1973 Sep;33(3):275–283. doi: 10.1161/01.res.33.3.275. [DOI] [PubMed] [Google Scholar]
- Holzapfel G., Wehland J., Weber K. Calcium control of actin-myosin based contraction in triton models of mouse 3T3 fibroblasts is mediated by the myosin light chain kinase (MLCK)-calmodulin complex. Exp Cell Res. 1983 Oct;148(1):117–126. doi: 10.1016/0014-4827(83)90192-1. [DOI] [PubMed] [Google Scholar]
- Jaffe E. A., Nachman R. L., Becker C. G., Minick C. R. Culture of human endothelial cells derived from umbilical veins. Identification by morphologic and immunologic criteria. J Clin Invest. 1973 Nov;52(11):2745–2756. doi: 10.1172/JCI107470. [DOI] [PMC free article] [PubMed] [Google Scholar]
- James D. W., Taylor J. F. The stress developed by sheets of chick fibroblasts in vitro. Exp Cell Res. 1969 Jan;54(1):107–110. doi: 10.1016/0014-4827(69)90299-7. [DOI] [PubMed] [Google Scholar]
- Katsuragawa Y., Yanagisawa M., Inoue A., Masaki T. Two distinct nonmuscle myosin-heavy-chain mRNAs are differentially expressed in various chicken tissues. Identification of a novel gene family of vertebrate non-sarcomeric myosin heavy chains. Eur J Biochem. 1989 Oct 1;184(3):611–616. doi: 10.1111/j.1432-1033.1989.tb15057.x. [DOI] [PubMed] [Google Scholar]
- Kawamoto S., Adelstein R. S. Chicken nonmuscle myosin heavy chains: differential expression of two mRNAs and evidence for two different polypeptides. J Cell Biol. 1991 Mar;112(5):915–924. doi: 10.1083/jcb.112.5.915. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Korn E. D., Hammer J. A., 3rd Myosins of nonmuscle cells. Annu Rev Biophys Biophys Chem. 1988;17:23–45. doi: 10.1146/annurev.bb.17.060188.000323. [DOI] [PubMed] [Google Scholar]
- Laposata M., Dovnarsky D. K., Shin H. S. Thrombin-induced gap formation in confluent endothelial cell monolayers in vitro. Blood. 1983 Sep;62(3):549–556. [PubMed] [Google Scholar]
- MAJNO G., PALADE G. E. Studies on inflammation. 1. The effect of histamine and serotonin on vascular permeability: an electron microscopic study. J Biophys Biochem Cytol. 1961 Dec;11:571–605. doi: 10.1083/jcb.11.3.571. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Morel N. M., Dodge A. B., Patton W. F., Herman I. M., Hechtman H. B., Shepro D. Pulmonary microvascular endothelial cell contractility on silicone rubber substrate. J Cell Physiol. 1989 Dec;141(3):653–659. doi: 10.1002/jcp.1041410325. [DOI] [PubMed] [Google Scholar]
- Morel N. M., Petruzzo P. P., Hechtman H. B., Shepro D. Inflammatory agonists that increase microvascular permeability in vivo stimulate cultured pulmonary microvessel endothelial cell contraction. Inflammation. 1990 Oct;14(5):571–583. doi: 10.1007/BF00914277. [DOI] [PubMed] [Google Scholar]
- Pollard T. D., Doberstein S. K., Zot H. G. Myosin-I. Annu Rev Physiol. 1991;53:653–681. doi: 10.1146/annurev.ph.53.030191.003253. [DOI] [PubMed] [Google Scholar]
- Thornton S. C., Mueller S. N., Levine E. M. Human endothelial cells: use of heparin in cloning and long-term serial cultivation. Science. 1983 Nov 11;222(4624):623–625. doi: 10.1126/science.6635659. [DOI] [PubMed] [Google Scholar]
- Wysolmerski R. B., Lagunoff D. Involvement of myosin light-chain kinase in endothelial cell retraction. Proc Natl Acad Sci U S A. 1990 Jan;87(1):16–20. doi: 10.1073/pnas.87.1.16. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wysolmerski R. B., Lagunoff D. Regulation of permeabilized endothelial cell retraction by myosin phosphorylation. Am J Physiol. 1991 Jul;261(1 Pt 1):C32–C40. doi: 10.1152/ajpcell.1991.261.1.C32. [DOI] [PubMed] [Google Scholar]