Role of DGAT1 in Mammary Gland Development and Cancer

描述（由申请人提供）：我们提出使用缺乏DGAT 1的小鼠分析三酰甘油（TG）代谢在乳腺上皮发育和乳腺癌中的作用 （Dgat 1-/-），一种从脂肪酸和甘油二酯底物合成TG的主要酶。 在正常乳腺发育中，TG在上皮中合成用于乳汁分泌，并储存在脂肪细胞中，脂肪细胞是乳腺基质中最丰富的细胞类型。 肥胖，其特征在于脂肪组织中过量的TG储存，正在研究作为乳腺癌的危险因素。 在动物研究和一些人类营养和流行病学研究中，高总脂肪摄入量与乳腺癌风险增加有关。 然而，这些影响的具体机制在很大程度上是未知的。 在Dgat 1-/-小鼠中，乳腺上皮生长和分化在妊娠期间受损。 近年来，乳腺基质在上皮生长中的作用已经在正常乳腺发育和癌症中得到证实。 乳腺移植实验表明，Dgat 1-/-小鼠上皮生长的减少是由基质效应局部介导的。 基于初步实验，与野生型相比，当将原发性乳腺癌细胞注射到Dgat 1-/-小鼠的乳腺脂肪垫中时，乳腺肿瘤的发展也减少了。 本提案中的实验将探索Dgat 1-/-乳腺间质中脂质代谢改变与正常和恶性上皮生长之间的联系机制。 我们推测，脂质代谢物或其他因素在乳腺DGAT 1的情况下发生改变，并损害乳腺上皮发育的信号传导。 我们将使用生物化学技术来测量Dgat 1-/-乳腺的脂质含量和组成。 我们还将进行微阵列实验，以分析与野生型相比，Dgat 1-/-乳腺脂肪细胞中的基因表达变化。 作为鉴定改变因子的补充方法，将开发上皮细胞和脂肪细胞的体外共培养系统。 为了探索DGAT 1缺陷在乳腺癌中的作用，我们将在Dgat 1-/-小鼠中使用乳腺癌细胞的原位移植物，并在Dgat 1-/-小鼠和两种乳腺肿瘤发病率增加的转基因小鼠模型之间进行杂交。 我们将测量肿瘤发生率、肿瘤负荷和形态学，并确定DGAT 1缺陷的保护作用是否对这些肿瘤模型之一具有特异性。 最后，基于显示Dgat 1-/-腹股沟淋巴结中白细胞分布改变的初步数据，我们将检验免疫系统在小鼠肿瘤排斥中起作用的假设。

DESCRIPTION (provided by applicant): We propose to analyze the role of triacylglycerol (TG) metabolism in mammary epithelial development and breast cancer using mice lacking DGAT1 (Dgat1-/-), a major enzyme that synthesizes TGs from fatty acids and diacylglycerol substrates. In normal mammary gland development, TGs are synthesized in the epithelium for milk secretion and stored in adipocytes, the most abundant cell type in the mammary stroma. Obesity, characterized by excess TG storage in adipose tissue, is under study as a risk factor for breast cancer. High intake of total fat has been associated with increased breast cancer risk in animal studies and in some nutrition and epidemiological studies in humans. However, specific mechanisms for these effects are largely unknown. In Dgat1-/- mice, mammary epithelial growth and differentiation are impaired during pregnancy. In recent years, the role of the mammary stroma in epithelial growth has been established in both normal breast development and cancer. Mammary transplantation experiments showed that the reduction in epithelial growth in Dgat1-/- mice is mediated locally by stromal effects. Based on preliminary experiments, breast tumor development is also reduced when primary breast cancer cells are injected into the mammary fat pad of Dgat1-/- mice compared to wild-types. Experiments in this proposal will explore mechanisms linking alteration of lipid metabolism in Dgat1-/- mammary stroma with both normal and malignant epithelial growth. We hypothesize that lipid metabolites or other factors are altered in mammary gland in the absence of DGAT1 and impair signaling for mammary epithelial development. We will use biochemical techniques to measure the lipid content and composition of Dgat1-/- mammary glands. We will also perform microarray experiments to analyze gene expression changes in Dgat1-/- mammary adipocytes compared to wild-types. As a complementary approach to identify altered factors, an in vitro system of co-culture of epithelium and adipocytes will be developed. To explore effects of DGAT1 deficiency in breast cancer, we will use orthotopic grafts of breast cancer cells in Dgat1-/- mice and crosses between Dgat1-/- mice and two transgenic mouse models with increased incidence of breast tumors. We will measure tumor incidence, tumor load, and morphology, and determine whether the protective effect of DGAT1 deficiency is specific for one of these tumor models. Finally, based on preliminary data showing altered distribution of leukocytes in Dgat1-/- inguinal lymph nodes, we will test the hypothesis that the immune system plays a role in tumor rejection by mice.