Spermidine as a possible mediator of glucocorticoid effect on milk protein synthesis in mouse mammary epithelium in vitro.

Several lines of evidence are presented for the involvement of spermidine in milk protein synthesis during hormone-dependent differentiation of mouse mammary epithelium in vitro. (a) When mammary explants from midpregnant mice are cultured in the presence of insulin, hydrocortisone, and prolactin, synthesis of casein and cr-lactalbumin increases markedly during the initial 48 hours. The requirement for hydrocortisone, but not for insulin or prolactin, can be met by spermidine at a concentration as low as 10-d M. Thus the combination of insulin, spermidine, and prolactin results in an increase in synthesis of the milk proteins similar to that produced by the triple hormone combination. The related polyamines, spermine and putrescine, and divalent cations such as Mg2+ are ineffective. (b) The concentration of spermidine in mammary epithelium increases by about j-fold when mammary explants are incubated for 48 hours in the presence of insulin, hydrocortisone, and prolactin, the same combination of hormones which causes marked stimulation of milk protein synthesis. Significant increases in the spermidine concentration occur prior to the accelerated synthesis of milk proteins. The combination of insulin and prolactin, which results in a slight stimulation of milk protein synthesis, correspondingly effects a smaller increase in the intracellular concentration of spermidine. (c) Methylglyoxal bis(guanylhydrazone), a potent inhibitor of spermidine synthesis, at a concentration of 2 pM, prevents increases in both the cellular spermidine concentration and the milk protein synthesis that occur in the presence of the three hormones. The simultaneous presence of spermidine, but not spermine or putrescine, however, overcomes the inhibitory effect of methylglyoxal bis(guanylhydrazone) on milk protein synthesis. The mechanism whereby glucocorticoid, together with insulin and prolactin, causes the large increase in the concentration of spermidine in cultured mammary cells appears to be mediated through stimulation of the activity of S-adenosyl-r.-methionine decarboxylase, which plays a key role in the biosynthesis of spermidine. In mammary epithelium, glucocorticoid, in combination with insulin, markedly in-