The role of connective tissue elements in the hormonally induced functional defferentiation of mouse mammary gland in culture

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In mammary gland organ culture from midpregnant mice, we have established the method of measurement of casein and -lactalbumin synthesis. Furthermore we examined the production of collagen and its function in the hormone-dependent development of mammary gland in vitro.The measurement of collagen production by the hydroxyproline assay and by sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed that the accumulation of collagen type I and type III in cultured mammary explants increased with the addition of insulin, cortisol and prolactin to a chemically defined medium. When an analog of proline, L-azetidine-2-carboxylic acid (LACA), was added at a concentration of 80 g/ml with insulin, cortisol and prolactin at the beginning of culture, collagen production in cultured tissue was inhibited by 75% curing a 3-day incubation period. This agent also inhibited the synthesis of casein and -lactalbumin by about 77 and 70%, respectively. The inhibitory effect of LACA could be prevented by L-proline; concomitant addition of L-proline (80 g/ml) with LACA (80 g/ml) resulted in complete restoration of milk protein synthesis to normal levels. LACA, however, produced little inhibition of DNA synthesis in cultured tissue. These results suggest that collagen production may be involved in the phenotypic expression of milk protein genes during hormonal induction of mammary epithelial differentiation in-vitro.