| Heregulin-αwas a cytokine which was synthesized and secreted by mammary mesenchyme,adjacent to lobuloaveolar structures. It controlled the development and lactation of mammary gland.It influenced the development of duct in pregnancy and stimulated the proliferation anddifferentiation of mammary epithelial cells. At present, the research about Heregulin-αand itsreceptors ErbB2 and ErbB3 was only the expression of them in mRNA and the impact ofHeregulin-αon the proliferation of mammary epithelial cells and milk proteins expression inculture. There were no results about the expression of Heregulin-αand its receptors in protein andthe impacts of Heregulin-αon the development, lactation and involution of mammary gland. It wasalso not clear about the signal transduction pathway of Heregulin-αin mammary gland.In these experiments, for the first time, we investigated the expression and localization ofHeregulin-αand its receptors ErbB2 and ErbB3 which were produced by mouse mammary glanditself in virgin, pregnancy, lactation and involution systematically by confocal laser scanningmicroscope. These results showed that the expression of Heregulin-αwas nearly absent at virgin20d～40d, and from virgin 45d the expression of Heregulin-αupgraded; the expression increasedobviously, the highest level of expression at pregnancy 15d; from lactation, the expression wasdecreased sharply, becoming undetecTab, again at lactation 15d; in involution, the expression ofHeregulin-αstill absent untill at involution 6d, and its expression upgraded slightly. TheHeregulin-αexpression achieved another peak at involution 9d. At involution 12d, the expressionwas down to the level of Heregulin-αexpression in virgin. Heregulin-αwas found in adipocytes,ductal epithelial cells and basal lamina around the duct and only in virgin, pregnancy andinvolution expressed specially. In the whole development cycle, the expression of ErbB2 andErbB3 was similar with Heregulin-αand showed significant positive correlation. In virgin 35d, theexpression of ErbB2 and ErbB3 was nearly absent, and at virgin 20d～30d and 40～60d theexpression of ErbB2 and ErbB3 upgraded; in pregnancy, the expression of ErbB2 and ErbB3increased obviously, the highest level of expression at pregnancy 15d; from lactation, theexpression of ErbB2 and ErbB3 was decreased sharply, becoming undetecTab, again at lactation15d; in involution, the expression of ErbB2 and ErbB3 still absent untill at involution 6d itsexpression upgraded slightly, and the ErbB2 and ErbB3 expression achieved another peak atinvolution 9d. This peak is lower than the pregnancy peak. At involution 12d, the expression was down to the level of ErbB2 and ErbB3 expression in virgin. ErbB2 and ErbB3 were found inadipocytes, ductal epithelial cells and basal lamina around the duct and only in virgin, pregnancyand involution expressed specially. Heregulin-αand its receptors had a positive correlation, whichmeant that Heregulin-αenhanced ErbB2 and ErbB3 expression solitary. ErbB2 and ErbB3 had apositive correlation, which meant Heregulin-αaffected the development and physiology function ofmammary gland by binding ErbB2 and ErbB3.In order to clarify the biological function of Heregulin-αand ErbB2 and ErbB3 in thedevelopment cycle of mammary gland, explants of mammary gland were cultured in a syntheticmedium with various hormone supplements, with or without Heregulin-αin vitro. The paraffinsections of explants which were cultured for 6d in vitro showed that Heregulin-αstimulated theproliferation and differentiation of ductal epithelial cells in pregnancy and maintained themorphosis of mammary gland in lactation and induced mammary gland degeneration andrestitution in involution.To study the effect of Heregulin-αonβ-casein expression of pregnancy, lactation and involutionmammary epithelial cells, this experiment used Western Blot method. The results showed that theexpression ofβ-casein is unchanged in pregnancy. Heregulin-αinduced the secretion ofβ-casein todecrease the expression ofβ-casein in mammary epithelial cells in lactation. Heregulin-αrestainedthe secretion ofβ-casein to increase the expression ofβ-casein in mammary epithelial cells ininvolution.For studying the effect of Heregulin-αonβ-casein secretion of pregnancy, lactation andinvolution mammary gland, this experiment detected the the concentration ofβ-casein in mediumon 1d, 2d, 3d, 4d, 5d and 6d in culture by capillary electrophoresis. The results showed thatHeregulin-αincreased and maintained the secretion ofβ-casein for 5d in pregnancy; Heregulin-αinduced the secretion ofβ-casein obviously within 5d in lactation; Heregulin-αdecreased thesecretion ofβ-casein obviously in involution.In order to clarify the function of Heregulin-αon the expression of Rab3A in pregnancy,lactation and involution mammary gland, this experiment adopt at Western Blot method. Theresults showed that in pregnancy, lactation and involution, Heregulin-αinduced the expression ofRab3A to stimulate the proliferation and differentiation of ductal epithelial cells in pregnancy;Heregulin-αinduced Rab3A to increase the expression of secretedβ-casein in lactation;Heregulin-αinduced Rab3A to promote mammary gland apoptpsis and restitution in involution.In order to clarify the mechanism of Heregulin-αin mammary gland, Western Blot was used toresearch the phosphorylation of signal molecules in Heregulin-αsignal transduction pathway. Theresults showed that Heregulin-αactivated STAT5, p42/p44, p38 and PKC specifically to induce theproliferation and differentiation of ductal epithelial cells in pregnancy. Heregulin-αinduced thephosphorylation of STAT5 to enhance the secretion of milk protein in lactation. The active p38 andrestrained PKC maintained the morphosis of mammary gland in lactation. Heregulin-αinduced thephosphorylation of STAT3 specifically, and restrained PKC to induce the degeneration of mammary epithelial cells and restitution in involution.
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