Hormonal induction of functional differentiation and mammary-derived growth inhibitor expression in cultured mouse mammary gland explants

Abstract

A method for the cultivation of organ explants from abdominal mammary glands of virgin mice has been established. In a serum-free medium containing aldosterone, prolactin, insulin, and cortisol (APIH medium) mammary gland development was documented by lobuloalveolar morphogenesis. The hormonal requirements for in vitro expression of beta-casein and of the mammary-derived growth inhibitor (MDGI) were tested. To this end, a full length cDNA coding for mouse MDGI was prepared displaying strong homologies to a mouse heart fatty acid binding protein, which is also expressed in the mammary gland. MDGI and beta-casein transcripts were found to be absent in the mammary tissue from primed virgin mice, and were induced upon culture of mammary explants in the APIH medium. An immunohistochemical analysis with specific antibodies against MDGI and casein revealed a different pattern of expression for the two proteins. In the APIH medium, MDGI was expressed mainly in differentiating alveolar cells of the lobuloalveolar structures, whereas beta-casein was present in both ductules and alveoli. The relationship between functional differentiation and MDGI expression was further studied in explants from glands of late-pregnant mice. At this stage of development, MDGI is found both in ducts and in alveoli. If explants were cultured with epidermal growth factor (EGF) and insulin, the lobuloalveolar structure was still present, whereas MDGI disappeared. Reinduction of MDGI expression was achieved by subsequent PIH treatment. Independent on developmental stage, EGF strongly inhibits MDGI mRNA expression. It is concluded that MDGI-expression is associated with functional differentiation in the normal gland.

Vanadate disrupts mammary gland development in whole organ culture

“…Importantly, this culture system promotes mammary epithelial cell differentiation as exhibited by the development of lobuloalveoli and the expression of milk proteins, such as ␤-casein (Terry et al, 1977;Mehta et al, 1980;Ganguly et al, 1981;Plaut et al, 1993). The extent of morphological and functional differentiation exhibited by the control mammary glands in our experiments (Figures 2, 5, and 7) are similar to previous reports (Terry et al, 1977;Mehta et al, 1980;Ganguly et al, 1981;Binas et al, 1992;Plaut et al, 1993;Li et al, 1995). In addition to proliferation and differentiation, mammary glands in whole organ culture can be induced to involute, a process mediated by apoptosis (Atwood et al, 1995;Casey et al, 1996), by the removal of the lactogenic hormones and culturing only with insulin (Topper et al, 1975).…”

“…Three-week-old female BALB/c mice (Taconic, Germantown, NY) were primed by daily subcutaneous injections of 50 l of a solution containing 1 g 17␤-estradiol and 1 mg progesterone in peanut oil (EP solution) for nine days (Binas et al, 1992). Briefly, The EP solution was made by dissolving 17␤-estradiol and progesterone in 100% ethanol or pre-warmed peanut oil, respectively.…”

Developmental regulation of fatty acid binding protein in neural tissue

“…MDGI in normal mammary tissue is maximally expressed after cell proliferation has occurred and during the onset of lactation by the differentiated gland (Binas et al, 1992). This suggests a role as a signal for the end of cell division and the completion of tissue differentiation (Spener and Borchers, 1992).…”

Induction of Mammary Gland Differentiation in Transgenic Mice by the Fatty Acid-binding Protein MRG

“…In vitro studies of mouse and bovine MDGI suggest several functions of MDGI on growth and differentiation of mammary gland. These include 1) MDGI specifically inhibit the growth of normal mouse mammary epithelial cells and promote morphological differentiation: the appearance of bulbous alveolar-like structure and formation of fully developed lobuloalveolar structures (16); 2) selective inhibition of endogenous MDGI expression in mouse mammary epithelial cells by use of antisense oligonucleotides suppresses the formation of alveolar-liken structure and impairs ␤-casein synthesis in organ cultures (16); 3) increasing amounts of MDGI mRNA were detected in terminal parts of ducts and lobuloalveolar epithelial cells of differentiated glands and maximally expressed in the terminally differentiated state found just prior to lactation (14); and 4) MDGI expression in mouse mammary epithelium cells is hormonally regulated (41,42). Although a large body of evidence has suggested that MDGI promotes differentiation of mammary epithelial cells in vitro, there is a concern about its role as differentiating factor for mammary gland in vivo.…”