Skip to main content
PMC home page
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
. 1991 Jun 15;88(12):5282–5286. doi: 10.1073/pnas.88.12.5282

Polarized intestinal hybrid cell lines derived from primary culture: establishment and characterization.

G W Aponte 1, A Keddie 1, G Halldén 1, R Hess 1, P Link 1
PMCID: PMC51856  PMID: 1711225

Abstract

A cell culture system has been developed that produces stable gastrointestinal (GI) polarized cell lines capable of maintaining hormone secretion. A spontaneously transformed rat mucosal epithelial cell was selected for hypoxanthine/guanine phosphoribosyltransferase deficiency and transfected with a plasmid conferring hygromycin resistance (BRIE 291 cells). Fusion of these cells with dispersed small intestinal epithelia cells resulted in hybrid cell lines that retained characteristic properties of the native GI cell more effectively than the transformed tumorigenic parental cell line. Hybrid hBRIE 380 cells are uniformly cuboidal with microvilli, contain villin, are contact inhibited, are anchorage dependent, require serum supplementation for growth, and are more sensitive to virus infection than the parental BRIE 291 cells. Fusion of BRIE 291 with dispersed pancreatic islet cells has resulted in a variety of pancreatic-hormone-producing cell lines. One of these, hybrid hBRIE 291-i2, forms confluent monolayers capable of synthesizing insulin-like immunoreactivity. These studies demonstrate that functionally polarized GI cells can be generated from primary cultures of nondividing committed epithelial cells by somatic cell hybridization and make feasible the selection and maintenance of specific GI epithelial cell types in confluent monolayer cultures.

Full text

PDF

Images in this article

5284Image
on p.5284
5284Image
on p.5284
5285Image
on p.5285
5285Image
on p.5285

Selected References

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

  1. Aponte G. W., Taylor I. L., Soll A. H. Primary culture of PYY cells from canine colon. Am J Physiol. 1988 Jun;254(6 Pt 1):G829–G836. doi: 10.1152/ajpgi.1988.254.6.G829. [DOI] [PubMed] [Google Scholar]
  2. Bergmann J. E., Fusco P. J. The M protein of vesicular stomatitis virus associates specifically with the basolateral membranes of polarized epithelial cells independently of the G protein. J Cell Biol. 1988 Nov;107(5):1707–1715. doi: 10.1083/jcb.107.5.1707. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bernard H. U., Krämmer G., Röwekamp W. G. Construction of a fusion gene that confers resistance against hygromycin B to mammalian cells in culture. Exp Cell Res. 1985 May;158(1):237–243. doi: 10.1016/0014-4827(85)90446-x. [DOI] [PubMed] [Google Scholar]
  4. Blay J., Brown K. D. Characterization of an epithelioid cell line derived from rat small intestine: demonstration of cytokeratin filaments. Cell Biol Int Rep. 1984 Jul;8(7):551–560. doi: 10.1016/0309-1651(84)90054-7. [DOI] [PubMed] [Google Scholar]
  5. Chantret I., Barbat A., Dussaulx E., Brattain M. G., Zweibaum A. Epithelial polarity, villin expression, and enterocytic differentiation of cultured human colon carcinoma cells: a survey of twenty cell lines. Cancer Res. 1988 Apr 1;48(7):1936–1942. [PubMed] [Google Scholar]
  6. Dudouet B., Robine S., Huet C., Sahuquillo-Merino C., Blair L., Coudrier E., Louvard D. Changes in villin synthesis and subcellular distribution during intestinal differentiation of HT29-18 clones. J Cell Biol. 1987 Jul;105(1):359–369. doi: 10.1083/jcb.105.1.359. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Dudouet B., Robine S., Huet C., Sahuquillo-Merino C., Blair L., Coudrier E., Louvard D. Changes in villin synthesis and subcellular distribution during intestinal differentiation of HT29-18 clones. J Cell Biol. 1987 Jul;105(1):359–369. doi: 10.1083/jcb.105.1.359. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Elsdale T., Bard J. Collagen substrata for studies on cell behavior. J Cell Biol. 1972 Sep;54(3):626–637. doi: 10.1083/jcb.54.3.626. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Emami S., Mir L., Gespach C., Rosselin G. Transfection of fetal rat intestinal epithelial cells by viral oncogenes: establishment and characterization of the E1A-immortalized SLC-11 cell line. Proc Natl Acad Sci U S A. 1989 May;86(9):3194–3198. doi: 10.1073/pnas.86.9.3194. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fujita T., Kobayashi S. Experimentally induced granule release in the endocrine cells of dog pyloric antrum. Z Zellforsch Mikrosk Anat. 1971;116(1):52–60. doi: 10.1007/BF00332857. [DOI] [PubMed] [Google Scholar]
  11. Fujita T., Kobayashi S. Structure and function of gut endocrine cells. Int Rev Cytol Suppl. 1977;(6):187–233. [PubMed] [Google Scholar]
  12. Gordon J. I. Intestinal epithelial differentiation: new insights from chimeric and transgenic mice. J Cell Biol. 1989 Apr;108(4):1187–1194. doi: 10.1083/jcb.108.4.1187. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hauri H. P., Sterchi E. E., Bienz D., Fransen J. A., Marxer A. Expression and intracellular transport of microvillus membrane hydrolases in human intestinal epithelial cells. J Cell Biol. 1985 Sep;101(3):838–851. doi: 10.1083/jcb.101.3.838. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Keddie B. A., Aponte G. W., Volkman L. E. The pathway of infection of Autographa californica nuclear polyhedrosis virus in an insect host. Science. 1989 Mar 31;243(4899):1728–1730. doi: 10.1126/science.2648574. [DOI] [PubMed] [Google Scholar]
  15. Kedinger M., Haffen K., Simon-Assmann P. Intestinal tissue and cell cultures. Differentiation. 1987;36(1):71–85. doi: 10.1111/j.1432-0436.1987.tb00182.x. [DOI] [PubMed] [Google Scholar]
  16. 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]
  17. McDonald K. Osmium ferricyanide fixation improves microfilament preservation and membrane visualization in a variety of animal cell types. J Ultrastruct Res. 1984 Feb;86(2):107–118. doi: 10.1016/s0022-5320(84)80051-9. [DOI] [PubMed] [Google Scholar]
  18. Moyer M. P., Aust J. B. Human colon cells: culture and in vitro transformation. Science. 1984 Jun 29;224(4656):1445–1447. doi: 10.1126/science.6328655. [DOI] [PubMed] [Google Scholar]
  19. Muggleton-Harris A. L., DeSimone D. W. Replicative potentials of various fusion products between WI-38 and SV40 transformed WI-38 cells and their components. Somatic Cell Genet. 1980 Nov;6(6):689–698. doi: 10.1007/BF01538968. [DOI] [PubMed] [Google Scholar]
  20. Pereira-Smith O. M., Smith J. R. Expression of SV40 T antigen in finite life-span hybrids of normal and SV40-transformed fibroblasts. Somatic Cell Genet. 1981 Jul;7(4):411–421. doi: 10.1007/BF01542986. [DOI] [PubMed] [Google Scholar]
  21. Pereira-Smith O. M., Smith J. R. Genetic analysis of indefinite division in human cells: identification of four complementation groups. Proc Natl Acad Sci U S A. 1988 Aug;85(16):6042–6046. doi: 10.1073/pnas.85.16.6042. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Pipeleers D. G., in't Veld P. A., Van de Winkel M., Maes E., Schuit F. C., Gepts W. A new in vitro model for the study of pancreatic A and B cells. Endocrinology. 1985 Sep;117(3):806–816. doi: 10.1210/endo-117-3-806. [DOI] [PubMed] [Google Scholar]
  23. Quaroni A. Development of fetal rat intestine in organ and monolayer culture. J Cell Biol. 1985 May;100(5):1611–1622. doi: 10.1083/jcb.100.5.1611. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Rindler M. J., Ivanov I. E., Plesken H., Rodriguez-Boulan E., Sabatini D. D. Viral glycoproteins destined for apical or basolateral plasma membrane domains traverse the same Golgi apparatus during their intracellular transport in doubly infected Madin-Darby canine kidney cells. J Cell Biol. 1984 Apr;98(4):1304–1319. doi: 10.1083/jcb.98.4.1304. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Rodriguez Boulan E., Sabatini D. D. Asymmetric budding of viruses in epithelial monlayers: a model system for study of epithelial polarity. Proc Natl Acad Sci U S A. 1978 Oct;75(10):5071–5075. doi: 10.1073/pnas.75.10.5071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Sargiacomo M., Lisanti M., Graeve L., Le Bivic A., Rodriguez-Boulan E. Integral and peripheral protein composition of the apical and basolateral membrane domains in MDCK cells. J Membr Biol. 1989 Mar;107(3):277–286. doi: 10.1007/BF01871942. [DOI] [PubMed] [Google Scholar]
  27. Simons K., Fuller S. D. Cell surface polarity in epithelia. Annu Rev Cell Biol. 1985;1:243–288. doi: 10.1146/annurev.cb.01.110185.001331. [DOI] [PubMed] [Google Scholar]
  28. Soll A. H., Yamada T., Park J., Thomas L. P. Release of somatostatinlike immunoreactivity from canine fundic mucosal cells in primary culture. Am J Physiol. 1984 Nov;247(5 Pt 1):G558–G566. doi: 10.1152/ajpgi.1984.247.5.G558. [DOI] [PubMed] [Google Scholar]
  29. Sugden B., Marsh K., Yates J. A vector that replicates as a plasmid and can be efficiently selected in B-lymphoblasts transformed by Epstein-Barr virus. Mol Cell Biol. 1985 Feb;5(2):410–413. doi: 10.1128/mcb.5.2.410. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Uchida S., Watanabe S., Aizawa T., Furuno A., Muto T. Polyoncogenicity and insulinoma-inducing ability of BK Virus, a human Papovavirus, in Syrian golden hamsters. J Natl Cancer Inst. 1979 Jul;63(1):119–126. [PubMed] [Google Scholar]
  32. Wigler M., Pellicer A., Silverstein S., Axel R. Biochemical transfer of single-copy eucaryotic genes using total cellular DNA as donor. Cell. 1978 Jul;14(3):725–731. doi: 10.1016/0092-8674(78)90254-4. [DOI] [PubMed] [Google Scholar]
  33. Worton R. G., Duff C. Karyotyping. Methods Enzymol. 1979;58:322–344. doi: 10.1016/s0076-6879(79)58148-8. [DOI] [PubMed] [Google Scholar]
  34. Yeh K. Y., Chopra D. P. Epithelial cell cultures from the colon of the suckling rat. In Vitro. 1980 Nov;16(11):976–986. doi: 10.1007/BF02619336. [DOI] [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