Skip to main content
NCBI 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
. 1987 Dec;84(23):8448–8452. doi: 10.1073/pnas.84.23.8448

Linoleic acid, but not cortisol, stimulates accumulation of casein by mouse mammary epithelial cells in serum-free collagen gel culture.

B K Levay-Young 1, G K Bandyopadhyay 1, S Nandi 1
PMCID: PMC299561  PMID: 3317407

Abstract

A two-step culture system has been developed to analyze the role of hormones in casein accumulation by mammary epithelial cells obtained from adrenalectomized and ovariectomized adult virgin mice. In the first step cells are grown inside collagen gel in medium containing insulin, epidermal growth factor (EGF), and linoleic acid for 9 days; these conditions stimulate very little casein accumulation. Following this growth phase the gels are released to float in medium containing insulin, prolactin, and linoleic acid. During this second phase the mammary cells will accumulate large amounts of casein, but only in the simultaneous presence of insulin, prolactin, and linoleic acid; in the absence of linoleic acid casein accumulation is greatly reduced. The casein accumulation is not dependent on the presence of the glucocorticoid cortisol and will occur in spite of the presence of the antiglucocorticoid agent RU 38 486. To determine if the response to cortisol observed in organ culture by other investigators might be mediated by stromal cells, epithelial cells were grown in collagen gel under fatty acid-free conditions and then cocultured with explants of mammary fat pads from adult virgin mice with or without mammary parenchyma. The cocultures were performed in fatty acid-free medium containing insulin and prolactin with or without cortisol. In the majority of experiments the mammary epithelial cells in the collagen gel accumulate more casein in the presence of cortisol than in its absence, irrespective of the presence of mammary parenchyma in the explant. Thus, mammary epithelial cells are directly dependent on insulin and prolactin for casein accumulation and indirectly dependent on cortisol by means of its effect on the stromal cells. This cortisol effect may be to cause release into the medium of linoleic acid or a metabolic product of linoleic acid from the stromal cells.

Full text

PDF
8448

Images in this article

8450Image
on p.8450

Selected References

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

  1. Bandyopadhyay G. K., Imagawa W., Wallace D., Nandi S. Linoleate metabolites enhance the in vitro proliferative response of mouse mammary epithelial cells to epidermal growth factor. J Biol Chem. 1987 Feb 25;262(6):2750–2756. [PubMed] [Google Scholar]
  2. Bolander F. F., Jr, Nicholas K. R., Topper Y. J. Retention of glucocorticoid by isolated mammary tissue may complicate interpretation of results from in vitro experiments. Biochem Biophys Res Commun. 1979 Nov 14;91(1):247–252. doi: 10.1016/0006-291x(79)90610-7. [DOI] [PubMed] [Google Scholar]
  3. Bolander F. F., Jr, Nicholas K. R., Van Wyk J. J., Topper Y. J. Insulin is essential for accumulation of casein mRNA in mouse mammary epithelial cells. Proc Natl Acad Sci U S A. 1981 Sep;78(9):5682–5684. doi: 10.1073/pnas.78.9.5682. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bolander F. F., Jr, Topper Y. J. Loss of differentiative potential of the mammary gland in ovariectomized mice: prevention and reversibility of the defect. Endocrinology. 1980 Nov;107(5):1281–1285. doi: 10.1210/endo-107-5-1281. [DOI] [PubMed] [Google Scholar]
  5. Chomczynski P., Qasba P., Topper Y. J. Essential role of insulin in transcription of the rat 25,000 molecular weight casein gene. Science. 1984 Dec 14;226(4680):1326–1328. doi: 10.1126/science.6390680. [DOI] [PubMed] [Google Scholar]
  6. Chomczynski P., Qasba P., Topper Y. J. Transcriptional and post-transcriptional roles of glucocorticoid in the expression of the rat 25,000 molecular weight casein gene. Biochem Biophys Res Commun. 1986 Jan 29;134(2):812–818. doi: 10.1016/s0006-291x(86)80493-4. [DOI] [PubMed] [Google Scholar]
  7. DEOME K. B., FAULKIN L. J., Jr, BERN H. A., BLAIR P. B. Development of mammary tumors from hyperplastic alveolar nodules transplanted into gland-free mammary fat pads of female C3H mice. Cancer Res. 1959 Jun;19(5):515–520. [PubMed] [Google Scholar]
  8. Durban E. M., Medina D., Butel J. S. Comparative analysis of casein synthesis during mammary cell differentiation in collagen and mammary gland development in vivo. Dev Biol. 1985 Jun;109(2):288–298. doi: 10.1016/0012-1606(85)90456-7. [DOI] [PubMed] [Google Scholar]
  9. ELIAS J. J. Cultivation of adult mouse mammary gland in hormone-enriched synthetic medium. Science. 1957 Oct 25;126(3278):842–843. doi: 10.1126/science.126.3278.842-a. [DOI] [PubMed] [Google Scholar]
  10. Edery M., Imagawa W., Larson L., Nandi S. Regulation of estrogen and progesterone receptor levels in mouse mammary epithelial cells grown in serum-free collagen gel cultures. Endocrinology. 1985 Jan;116(1):105–112. doi: 10.1210/endo-116-1-105. [DOI] [PubMed] [Google Scholar]
  11. Emerman J. T., Enami J., Pitelka D. R., Nandi S. Hormonal effects on intracellular and secreted casein in cultures of mouse mammary epithelial cells on floating collagen membranes. Proc Natl Acad Sci U S A. 1977 Oct;74(10):4466–4470. doi: 10.1073/pnas.74.10.4466. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Emerman J. T., Pitelka D. R. Maintenance and induction of morphological differentiation in dissociated mammary epithelium on floating collagen membranes. In Vitro. 1977 May;13(5):316–328. doi: 10.1007/BF02616178. [DOI] [PubMed] [Google Scholar]
  13. Enami J., Nandi S. A sensitive radioimmunoassay for a component of mouse casein. J Immunol Methods. 1977;18(3-4):235–244. doi: 10.1016/0022-1759(77)90177-6. [DOI] [PubMed] [Google Scholar]
  14. Fielder P. J., Talamantes F. The lipolytic effects of mouse placental lactogen II, mouse prolactin, and mouse growth hormone on adipose tissue from virgin and pregnant mice. Endocrinology. 1987 Aug;121(2):493–497. doi: 10.1210/endo-121-2-493. [DOI] [PubMed] [Google Scholar]
  15. Flynn D., Yang J., Nandi S. Growth and differentiation of primary cultures of mouse mammary epithelium embedded in collagen gel. Differentiation. 1982;22(3):191–194. doi: 10.1111/j.1432-0436.1982.tb01249.x. [DOI] [PubMed] [Google Scholar]
  16. Gagne D., Pons M., Philibert D. RU 38486: a potent antiglucocorticoid in vitro and in vivo. J Steroid Biochem. 1985 Sep;23(3):247–251. doi: 10.1016/0022-4731(85)90401-7. [DOI] [PubMed] [Google Scholar]
  17. Hinegardner R. T. An improved fluorometric assay for DNA. Anal Biochem. 1971 Jan;39(1):197–201. doi: 10.1016/0003-2697(71)90476-3. [DOI] [PubMed] [Google Scholar]
  18. ICHINOSE R. R., NANDI S. LOBULOALVEOLAR DIFFERENTIATION IN MOUSE MAMMARY TISSUES IN VITRO. Science. 1964 Jul 31;145(3631):496–497. doi: 10.1126/science.145.3631.496. [DOI] [PubMed] [Google Scholar]
  19. Imagawa W., Tomooka Y., Hamamoto S., Nandi S. Stimulation of mammary epithelial cell growth in vitro: interaction of epidermal growth factor and mammogenic hormones. Endocrinology. 1985 Apr;116(4):1514–1524. doi: 10.1210/endo-116-4-1514. [DOI] [PubMed] [Google Scholar]
  20. Imagawa W., Tomooka Y., Nandi S. Serum-free growth of normal and tumor mouse mammary epithelial cells in primary culture. Proc Natl Acad Sci U S A. 1982 Jul;79(13):4074–4077. doi: 10.1073/pnas.79.13.4074. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Juergens W. G., Stockdale F. E., Topper Y. J., Elias J. J. Hormone-dependent differentiation of mammary gland in vitro. Proc Natl Acad Sci U S A. 1965 Aug;54(2):629–634. doi: 10.1073/pnas.54.2.629. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kessler R. J., Vande Zande H., Tyson C. A., Blondin G. A., Fairfield J., Glasser P., Green D. E. Uncouplers and the molecular mechanism of uncoupling in mitochondria. Proc Natl Acad Sci U S A. 1977 Jun;74(6):2241–2245. doi: 10.1073/pnas.74.6.2241. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kratochwil K. Organ specificity in mesenchymal induction demonstrated in the embryonic development of the mammary gland of the mouse. Dev Biol. 1969 Jul;20(1):46–71. doi: 10.1016/0012-1606(69)90004-9. [DOI] [PubMed] [Google Scholar]
  24. Kulski J. K., Topper Y. J., Chomczynski P., Qasba P. An essential role for glucocorticoid in casein gene expression in rat mammary explants. Biochem Biophys Res Commun. 1983 Jul 18;114(1):380–387. doi: 10.1016/0006-291x(83)91638-8. [DOI] [PubMed] [Google Scholar]
  25. 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]
  26. Lee E. Y., Parry G., Bissell M. J. Modulation of secreted proteins of mouse mammary epithelial cells by the collagenous substrata. J Cell Biol. 1984 Jan;98(1):146–155. doi: 10.1083/jcb.98.1.146. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Levine J. F., Stockdale F. E. Cell-cell interactions promote mammary epithelial cell differentiation. J Cell Biol. 1985 May;100(5):1415–1422. doi: 10.1083/jcb.100.5.1415. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. McGrath M., Palmer S., Nandi S. Differential response of normal rat mammary epithelial cells to mammogenic hormones and EGF. J Cell Physiol. 1985 Nov;125(2):182–191. doi: 10.1002/jcp.1041250203. [DOI] [PubMed] [Google Scholar]
  29. Mehta N. M., Ganguly N., Ganguly R., Banerjee M. R. Hormonal modulation of the casein gene expression in a mammogenesis-lactogenesis culture model of the whole mammary gland of the mouse. J Biol Chem. 1980 May 25;255(10):4430–4434. [PubMed] [Google Scholar]
  30. Mills E. S., Topper Y. J. Some ultrastructural effects of insulin, hydrocortisone, and prolactin on mammary gland explants. J Cell Biol. 1970 Feb;44(2):310–328. doi: 10.1083/jcb.44.2.310. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Nagaiah K., Bolander F. F., Jr, Nicholas K. R., Takemoto T., Topper Y. J. Prolactin-induced accumulation of casein mRNA in mouse mammary explants: a selective role of glucocorticoid. Biochem Biophys Res Commun. 1981 Jan 30;98(2):380–387. doi: 10.1016/0006-291x(81)90851-2. [DOI] [PubMed] [Google Scholar]
  32. Oka T., Perry J. W. Studies on the function of glucocorticoid in mouse mammary epithelial cell differentiation in vitro. Stimulation of glucose 6-phosphate dehydrogenase. J Biol Chem. 1974 Jun 10;249(11):3586–3591. [PubMed] [Google Scholar]
  33. Oka T., Topper Y. J. Hormone-dependent accumulation of rough endoplasmic reticulum in mouse mammary epithelial cells in vitro. J Biol Chem. 1971 Dec 25;246(24):7701–7707. [PubMed] [Google Scholar]
  34. Ray D. B., Jansen R. W., Horst I. A., Mills N. C., Kowal J. A complex noncoordinate regulation of alpha-lactalbumin and 25 K beta-casein by corticosterone, prolactin, and insulin in long term cultures of normal rat mammary cells. Endocrinology. 1986 Jan;118(1):393–407. doi: 10.1210/endo-118-1-393. [DOI] [PubMed] [Google Scholar]
  35. Rocha V., Ringo D. L., Read D. B. Casein production during differentiation of mammary epithelial cells in collagen gel culture. Exp Cell Res. 1985 Jul;159(1):201–210. doi: 10.1016/s0014-4827(85)80049-5. [DOI] [PubMed] [Google Scholar]
  36. SOEMARWOTO I. N., BERN H. A. The effect of hormones on the vascular pattern of the mouse mammary gland. Am J Anat. 1958 Nov;103(3):403–435. doi: 10.1002/aja.1001030305. [DOI] [PubMed] [Google Scholar]
  37. Sakakura T., Nishizuka Y., Dawe C. J. Mesenchyme-dependent morphogenesis and epithelium-specific cytodifferentiation in mouse mammary gland. Science. 1976 Dec 24;194(4272):1439–1441. doi: 10.1126/science.827022. [DOI] [PubMed] [Google Scholar]
  38. Smith S., Watts R., Dils R. Quantitative gas-liquid chromatographic analysis of rodent milk triglycerides. J Lipid Res. 1968 Jan;9(1):52–57. [PubMed] [Google Scholar]
  39. Stockdale F. E., Topper Y. J. The role of DNA synthesis and mitosis in hormone-dependent differentiation. Proc Natl Acad Sci U S A. 1966 Oct;56(4):1283–1289. doi: 10.1073/pnas.56.4.1283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Taketani Y., Oka T. Hormonal regulation of the synthesis of casein and alpha-lactalbumin in a primary mammary cell culture system. Horm Metab Res. 1986 Feb;18(2):119–125. doi: 10.1055/s-2007-1012246. [DOI] [PubMed] [Google Scholar]
  41. Tonelli Q. J., Sorof S. Induction of biochemical differentiation in three-dimensional collagen cultures of mammary epithelial cells from virgin mice. Differentiation. 1982;22(3):195–200. doi: 10.1111/j.1432-0436.1982.tb01250.x. [DOI] [PubMed] [Google Scholar]
  42. Topper Y. J., Freeman C. S. Multiple hormone interactions in the developmental biology of the mammary gland. Physiol Rev. 1980 Oct;60(4):1049–1106. doi: 10.1152/physrev.1980.60.4.1049. [DOI] [PubMed] [Google Scholar]
  43. 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]
  44. Wicha M. S., Liotta L. A., Kidwell W. R. Effects of free fatty acids on the growth of normal and neoplastic rat mammary epithelial cells. Cancer Res. 1979 Feb;39(2 Pt 1):426–435. [PubMed] [Google Scholar]
  45. Wicha M. S., Lowrie G., Kohn E., Bagavandoss P., Mahn T. Extracellular matrix promotes mammary epithelial growth and differentiation in vitro. Proc Natl Acad Sci U S A. 1982 May;79(10):3213–3217. doi: 10.1073/pnas.79.10.3213. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Wiens D., Park C. S., Stockdale F. E. Milk protein expression and ductal morphogenesis in the mammary gland in vitro: hormone-dependent and -independent phases of adipocyte-mammary epithelial cell interaction. Dev Biol. 1987 Mar;120(1):245–258. doi: 10.1016/0012-1606(87)90122-9. [DOI] [PubMed] [Google Scholar]
  47. Yang J., Guzman R., Richards J., Imagawa W., McCormick K., Nandi S. Growth factor- and cyclic nucleotide-induced proliferation of normal and malignant mammary epithelial cells in primary culture. Endocrinology. 1980 Jul;107(1):35–41. doi: 10.1210/endo-107-1-35. [DOI] [PubMed] [Google Scholar]
  48. Yang J., Richards J., Guzman R., Imagawa W., Nandi S. Sustained growth in primary culture of normal mammary epithelial cells embedded in collagen gels. Proc Natl Acad Sci U S A. 1980 Apr;77(4):2088–2092. doi: 10.1073/pnas.77.4.2088. [DOI] [PMC free article] [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