Skip to main content
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1992 Oct 1;89(19):9069–9073. doi: 10.1073/pnas.89.19.9069

Glucocorticoid-induced formation of tight junctions in mouse mammary epithelial cells in vitro.

K S Zettl 1, M D Sjaastad 1, P M Riskin 1, G Parry 1, T E Machen 1, G L Firestone 1
PMCID: PMC50066  PMID: 1409603

Abstract

Phenotypically stable cultures of untransformed mouse mammary epithelial cells (denoted 31EG4) were established and utilized to investigate the lactogenic hormone (glucocorticoids, insulin, and prolactin) regulation of tight junction formation. When 31EG4 cells were grown on permeable supports for 4 days in medium containing the synthetic glucocorticoid dexamethasone and insulin, confluent cell monolayers obtained a transepithelial electrical resistance (TER) of 1000-3000 omega.cm2. In contrast, over the same time period, confluent monolayers treated with insulin or insulin and prolactin maintained a low TER (35-150 omega.cm2). Consistent with the formation of tight junctions, apical to basolateral paracellular permeability was decreased from 12% to 1% for [14C]mannitol and 3.3% to 0.3% for [3H]inulin when cells were cultured in dexamethasone. This effect of dexamethasone on TER required extracellular calcium, de novo protein synthesis, dose-dependently correlated with glucocorticoid receptor occupancy, and was not due to an increase in cell density. As shown by direct and indirect immunofluorescence microscopy, dexamethasone treatment did not modulate the production or location of filamentous actin, the tight junction protein ZO-1, or the cell adhesion protein E-cadherin. Our results suggest that glucocorticoids play a fundamental role in the function and maintenance of cell-cell contact in the mammary epithelia by inducing the formation of tight junctions.

Full text

PDF

Images in this article

Selected References

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

  1. Balda M. S., González-Mariscal L., Contreras R. G., Macias-Silva M., Torres-Marquez M. E., García-Sáinz J. A., Cereijido M. Assembly and sealing of tight junctions: possible participation of G-proteins, phospholipase C, protein kinase C and calmodulin. J Membr Biol. 1991 Jun;122(3):193–202. doi: 10.1007/BF01871420. [DOI] [PubMed] [Google Scholar]
  2. Duffey M. E., Hainau B., Ho S., Bentzel C. J. Regulation of epithelial tight junction permeability by cyclic AMP. Nature. 1981 Dec 3;294(5840):451–453. doi: 10.1038/294451a0. [DOI] [PubMed] [Google Scholar]
  3. Firestone G. L., John N. J., Yamamoto K. R. Glucocorticoid-regulated glycoprotein maturation in wild-type and mutant rat cell lines. J Cell Biol. 1986 Dec;103(6 Pt 1):2323–2331. doi: 10.1083/jcb.103.6.2323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Fuller P. J. The steroid receptor superfamily: mechanisms of diversity. FASEB J. 1991 Dec;5(15):3092–3099. doi: 10.1096/fasebj.5.15.1743440. [DOI] [PubMed] [Google Scholar]
  5. Gonzalez-Mariscal L., Chávez de Ramírez B., Cereijido M. Tight junction formation in cultured epithelial cells (MDCK). J Membr Biol. 1985;86(2):113–125. doi: 10.1007/BF01870778. [DOI] [PubMed] [Google Scholar]
  6. Gonzalez-Mariscal L., Contreras R. G., Bolívar J. J., Ponce A., Chávez De Ramirez B., Cereijido M. Role of calcium in tight junction formation between epithelial cells. Am J Physiol. 1990 Dec;259(6 Pt 1):C978–C986. doi: 10.1152/ajpcell.1990.259.6.C978. [DOI] [PubMed] [Google Scholar]
  7. Gumbiner B., Stevenson B., Grimaldi A. The role of the cell adhesion molecule uvomorulin in the formation and maintenance of the epithelial junctional complex. J Cell Biol. 1988 Oct;107(4):1575–1587. doi: 10.1083/jcb.107.4.1575. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Hammerton R. W., Krzeminski K. A., Mays R. W., Ryan T. A., Wollner D. A., Nelson W. J. Mechanism for regulating cell surface distribution of Na+,K(+)-ATPase in polarized epithelial cells. Science. 1991 Nov 8;254(5033):847–850. doi: 10.1126/science.1658934. [DOI] [PubMed] [Google Scholar]
  9. Madara J. L., Dharmsathaphorn K. Occluding junction structure-function relationships in a cultured epithelial monolayer. J Cell Biol. 1985 Dec;101(6):2124–2133. doi: 10.1083/jcb.101.6.2124. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Madara J. L. Intestinal absorptive cell tight junctions are linked to cytoskeleton. Am J Physiol. 1987 Jul;253(1 Pt 1):C171–C175. doi: 10.1152/ajpcell.1987.253.1.C171. [DOI] [PubMed] [Google Scholar]
  11. McRoberts J. A., Aranda R., Riley N., Kang H. Insulin regulates the paracellular permeability of cultured intestinal epithelial cell monolayers. J Clin Invest. 1990 Apr;85(4):1127–1134. doi: 10.1172/JCI114544. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. McRoberts J. A., Riley N. E. Regulation of T84 cell monolayer permeability by insulin-like growth factors. Am J Physiol. 1992 Jan;262(1 Pt 1):C207–C213. doi: 10.1152/ajpcell.1992.262.1.C207. [DOI] [PubMed] [Google Scholar]
  13. Mullin J. M., O'Brien T. G. Effects of tumor promoters on LLC-PK1 renal epithelial tight junctions and transepithelial fluxes. Am J Physiol. 1986 Oct;251(4 Pt 1):C597–C602. doi: 10.1152/ajpcell.1986.251.4.C597. [DOI] [PubMed] [Google Scholar]
  14. Ojakian G. K. Tumor promoter-induced changes in the permeability of epithelial cell tight junctions. Cell. 1981 Jan;23(1):95–103. doi: 10.1016/0092-8674(81)90274-9. [DOI] [PubMed] [Google Scholar]
  15. Ophir I., Cohen E., Bacher A., Ben-Shaul Y. Effect of protein synthesis inhibitors on formation and degradation of tight junctions in HT 29 adenocarcinoma cells. Eur J Cell Biol. 1989 Jun;49(1):116–122. [PubMed] [Google Scholar]
  16. Palant C. E., Duffey M. E., Mookerjee B. K., Ho S., Bentzel C. J. Ca2+ regulation of tight-junction permeability and structure in Necturus gallbladder. Am J Physiol. 1983 Sep;245(3):C203–C212. doi: 10.1152/ajpcell.1983.245.3.C203. [DOI] [PubMed] [Google Scholar]
  17. Pitelka D. R., Hamamoto S. T., Duafala J. G., Nemanic M. K. Cell contacts in the mouse mammary gland. I. Normal gland in postnatal development and the secretory cycle. J Cell Biol. 1973 Mar;56(3):797–818. doi: 10.1083/jcb.56.3.797. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Poyet P., Henning S. J., Rosen J. M. Hormone-dependent beta-casein mRNA stabilization requires ongoing protein synthesis. Mol Endocrinol. 1989 Dec;3(12):1961–1968. doi: 10.1210/mend-3-12-1961. [DOI] [PubMed] [Google Scholar]
  19. Reichmann E., Ball R., Groner B., Friis R. R. New mammary epithelial and fibroblastic cell clones in coculture form structures competent to differentiate functionally. J Cell Biol. 1989 Mar;108(3):1127–1138. doi: 10.1083/jcb.108.3.1127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Rodriguez-Boulan E., Nelson W. J. Morphogenesis of the polarized epithelial cell phenotype. Science. 1989 Aug 18;245(4919):718–725. doi: 10.1126/science.2672330. [DOI] [PubMed] [Google Scholar]
  21. Stevenson B. R., Anderson J. M., Braun I. D., Mooseker M. S. Phosphorylation of the tight-junction protein ZO-1 in two strains of Madin-Darby canine kidney cells which differ in transepithelial resistance. Biochem J. 1989 Oct 15;263(2):597–599. doi: 10.1042/bj2630597. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Stevenson B. R., Anderson J. M., Goodenough D. A., Mooseker M. S. Tight junction structure and ZO-1 content are identical in two strains of Madin-Darby canine kidney cells which differ in transepithelial resistance. J Cell Biol. 1988 Dec;107(6 Pt 1):2401–2408. doi: 10.1083/jcb.107.6.2401. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Stevenson B. R., Siliciano J. D., Mooseker M. S., Goodenough D. A. Identification of ZO-1: a high molecular weight polypeptide associated with the tight junction (zonula occludens) in a variety of epithelia. J Cell Biol. 1986 Sep;103(3):755–766. doi: 10.1083/jcb.103.3.755. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Strange R., Li F., Friis R. R., Reichmann E., Haenni B., Burri P. H. Mammary epithelial differentiation in vitro: minimum requirements for a functional response to hormonal stimulation. Cell Growth Differ. 1991 Nov;2(11):549–559. [PubMed] [Google Scholar]
  25. Webster M. K., Guthrie J., Firestone G. L. Glucocorticoid growth suppression response in 13762NF adenocarcinoma-derived Con8 rat mammary tumor cells is mediated by dominant trans-acting factors. Cancer Res. 1991 Nov 15;51(22):6031–6038. [PubMed] [Google Scholar]
  26. Webster M. K., Guthrie J., Firestone G. L. Suppression of rat mammary tumor cell growth in vitro by glucocorticoids requires serum proteins. Characterization of wild type and glucocorticoid-resistant epithelial tumor cells. J Biol Chem. 1990 Mar 25;265(9):4831–4838. [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES