Expression data from in utero exposure to genistein, vinclozolin and the mixture of genistein and vinclozolin on the mammary gland
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ABSTRACT: Morphogenesis of the mammary gland relies on the precise developmental control of morphological elements including TEBs, ducts and lobules. In the peripubertal mammary gland, rising levels of ovarian hormones control this development through a tightly controlled genetic program where specific sets of genes are up-regulated. We used microarrays to detail the program of gene expression underlying different classes of up-regulated genes during the peripubertal process after administration of endocrine disruptors during the fetal and neonatal development.
Project description:Morphogenesis of the mammary gland relies on the precise developmental control of morphological elements including TEBs, ducts and lobules. In the peripubertal mammary gland, rising levels of ovarian hormones control this development through a tightly controlled genetic program where specific sets of genes are up-regulated. We used microarrays to detail the program of gene expression underlying different classes of up-regulated genes during the peripubertal process after administration of endocrine disruptors during the fetal and neonatal development. Rat mammary glands were selected at two peripubertal periods (days 35 and 50), after administration of genistein and vinclozolin during the fetal and neonatal development, for RNA extraction and hybridization on Affymetrix microarrays.. We sought to obtain homogeneous populations of mammary gland for each treatment, and at each developmental stage, in order to increase the temporal and specific effect resolution of time course and endocrine disruption.
Project description:Diethlstilbestrol (DES) is a synthetic estrogen prescribed to several millions of pregnant women worldwide. The risk for breast cancer after age 40 in women prenatally exposed to DES has been reported, however, the precise mechanism of susceptibilities to breast cancer remains to be resolved. To understand the mechanism of effect of intrauterine exposure to DES on human mammary glands at peripubertal stage, we investigated the global gene expression profile of terminal end buds (TEBs), the target of carcinogen, in rat mammary glamds at postnatal days (PND) 35 and 49, equivalent to peripubertal stage in human, exposed to low or high dose of DES at neonatal period. In all group, the number of TEBs was gradually increased and peaked at PND35 and decreased at PND49. At PND 35 and 49, low dose of DES-treated group showed highest number of TEBs. At PND35, β- and γ-casein mRNA expression inreased 8.19-fold and 26.05-fold, respectively, and the most significant network revealed by IPA analyses showed relevance of NF-κB in low dose of DES-treated group. The present findings suggested that deregulation of mammary gland differentiation and development-related genes group may contribute to the increase of number of TEBs at critical period of carcinogen susceptibilities.
Project description:Diethlstilbestrol (DES) is a synthetic estrogen prescribed to several millions of pregnant women worldwide. The risk for breast cancer after age 40 in women prenatally exposed to DES has been reported, however, the precise mechanism of susceptibilities to breast cancer remains to be resolved. To understand the mechanism of effect of intrauterine exposure to DES on human mammary glands at peripubertal stage, we investigated the global gene expression profile of terminal end buds (TEBs), the target of carcinogen, in rat mammary glamds at postnatal days (PND) 35 and 49, equivalent to peripubertal stage in human, exposed to low or high dose of DES at neonatal period. In all group, the number of TEBs was gradually increased and peaked at PND35 and decreased at PND49. At PND 35 and 49, low dose of DES-treated group showed highest number of TEBs. At PND35, β- and γ-casein mRNA expression inreased 8.19-fold and 26.05-fold, respectively, and the most significant network revealed by IPA analyses showed relevance of NF-κB in low dose of DES-treated group. The present findings suggested that deregulation of mammary gland differentiation and development-related genes group may contribute to the increase of number of TEBs at critical period of carcinogen susceptibilities. Experiment: We used laser capture microdissection to specifically isolate the cells from terminal end buds (TEBs) in rat mammary gland at 35 and 49 days after birth. Inbred Sprague-Dawley female rats were subcutaneously given 0.05ml sesame oil (group I, control) and either 1 μg (group II) or 100 μg (group III) diethlstilbestrol (DES, Sigma Chemical Co., St. Louis, Mo. USA) dissolved in 0.05 ml sesame oil within 24 hr of birth. Animal were weaned at 21 days of birth and each group of rats was autopsied at 35 and 49 days of birth. From group I (35 days of birth: n=2, 49 days of birth: n=2), group II (35 days of birth: n=2, 49 days of birth: n=2), group III (35 days of birth: n=2, 49 days of birth: n=2), we performed a hybridization, respectively.
Project description:The mammary gland is a highly dynamic organ that mainly develops during puberty. Based on morphology and proliferation analysis, mammary stem cells (MaSCs) are thought to be close to or reside in the terminal end buds (TEBs) during pubertal development. However, exclusive stem cell markers are lacking, and therefore the true identity of MaSCs, including their location, multiplicity, dynamics and fate during branching morphogenesis, has yet to be defined. To gain more insights into the molecular identity and heterogeneity of the MaSC pool, we performed single cell transcriptome sequencing of mammary epithelial cells micro-dissected from ducts and TEBs during puberty. These data show that the behaviour of MaSCs cannot be directly linked to a single expression profile. Instead, morphogenesis of the mammary epithelium relies upon a heterogeneous population of MaSCs that functions long-term as a single equipotent pool of stem cells.
Project description:Pregnancy-Induced Non-Coding RNA (PINC) is upregulated in alveolar cells of the mammary gland during pregnancy and persist in alveolar cells that remain in the regressed lobules following involution. Here, we show that in the post-pubertal mouse mammary gland, mPINC expression increases throughout pregnancy and then declines in early lactation, when alveolar cells undergo terminal differentiation. Accordingly, mPINC expression is significantly decreased when HC11 mammary epithelial cells are induced to differentiate and produce milk proteins. This natural reduction in mPINC levels may be necessary for lactation, as overexpression of mPINC in HC11 cells blocks lactogenic differentiation, while knockdown of mPINC enhances differentiation. HC11 mammary epithelial cells, with or without knockdown or over-expression of PINC, and with or without treatment by differentiation-inducing agents, were profiled for gene expression.
Project description:Pregnancy-Induced Non-Coding RNA (PINC) is upregulated in alveolar cells of the mammary gland during pregnancy and persist in alveolar cells that remain in the regressed lobules following involution. Here, we show that in the post-pubertal mouse mammary gland, mPINC expression increases throughout pregnancy and then declines in early lactation, when alveolar cells undergo terminal differentiation. Accordingly, mPINC expression is significantly decreased when HC11 mammary epithelial cells are induced to differentiate and produce milk proteins. This natural reduction in mPINC levels may be necessary for lactation, as overexpression of mPINC in HC11 cells blocks lactogenic differentiation, while knockdown of mPINC enhances differentiation.
Project description:The developing mammary gland contains distinct microenvironments that perform specialized functions for branching and ductal invasion. These microenvironments include the terminal end buds (TEBs) at the tips of invading primary ducts and the more differentiated proximal ducts that give rise to side branches. We have devised a novel microarray approach to identify genes that are expressed in the epithelium and stroma of these distinct microenvironments. Keywords: spotted oligonucleotide
Project description:Rodent studies have indicated that gestational and perinatal bisphenol A (BPA) exposure increase the risk of developing breast cancer during adulthood. In contrast, some dietary compounds such as genistein (GEN) and indole 3-carbinol (I3C) present potential protective effects against inducing hormone-dependent cancers, including that of the mammary gland. Thus, we aimed to evaluate the role of these dietary compounds on early mammary gland development and carcinogenesis in female Sprague-Dawley offspring. Pregnant Sprague-Dawley (SD) rats were treated with BPA at 25 or 250µg/kg b.w./day by gavage from gestational day (GD) 10 to 21 with or without dietary GEN (250 mg/kg chow, ~5.5 mg/kg b.w./day) or I3C (2000 mg/kg chow, ~45.0 mg/kg b.w./day). At post-natal day (PND) 21, some female offspring from different litters were euthanized for mammary gland development and gene expression analyses while other female offspring received a single dose of N-methyl-N-nitrosourea (MNU) for mammary carcinogenesis initiation. The findings this study indicated the prenatal exposure to BPA, GEN and I3C did not significantly alter ductal elongation, number of terminal end buds (TEB) or cell proliferation, and estrogen receptor alpha (ER-α) immunostaining expression in epithelial mammary cells at PND 21. BPA treatment modulated the expression of several genes, but these changes were not associated with a dose dependent response. Dietary GEN and I3C treatment causally and consistent with the mammary gland structures outcomes. Besides, maternal BPA exposure associated with dietary GEN and I3C did not alter the susceptibility to the mammary cancer development in adulthood when the carcinogen was administered in a window of immature mammary gland development.
Project description:7,12-Dimethylbenz[a]anthracene (DMBA)-induced rat mammary carcinoma is a well-recognized model, however, the genetic alterations during its carcinogenesis have yet to be determined. We used laser capture microdissection to specifically isolate cells from terminal end buds (TEBs), the origin of carcinoma, at 2 weeks after sesame oil- treatment (control) or DMBA-treatment (DMBA-TEBs), ductal carcinoma in situ (DCIS) and invasive mammary carcinoma (MC). Using an oligonucleotide microarray representing 20600 rat probe sequences, we analyzed gene expression profiles and validated mRNA and protein levels of genes of interest by real-time quantitative PCR, immunohistochemistry and immunofluorescence. The number of differentially expressed genes was smallest in DMBA-TEBs (63), followed by DCIS (798) and MC (981). Only the expression of PEP-19, an anti-apoptic gene, showed significant increases in DMBA-TEBs (4-fold), DCIS (10-fold) and MC (16-fold). MMP-13 expression was increased markedly in DCIS (19-fold) and MC (61-fold) while OPN expression was increased 6-fold in DCIS and 8-fold in MC. MMP-7 expression was increased 4-fold in MC. Nidogen-1; a participant in the assembly of basement membranes, TSP-2; an inhibitor of angiogenesis and COUP-TFI; a transcription repressor showed significant decreases in DCIS (4-, 9-, and 17-fold, respectively) and MC (10-, 37-, and 100-fold). Network analyses with IPA software revealed that the most significant network was centered on Akt groups in DCIS and ERK groups in MC. The present findings provide insights into the molecular changes and suggest the importance of PEP-19 overexpression very early on during mammary carcinogenesis. Keywords: Comparative experiments of sesame oil treatmentM-oM-<M-^HcontrolM-oM-<M-^Iand 7,12-Dimethylbenz[a]anthraceneM-oM-<M-^HDMBAM-oM-<M-^Itreatment group. Or comparative experiments of tissue type ExperimentM-oM-<M-^Z7,12-Dimethylbenz[a]anthracene (DMBA)-induced rat mammary carcinoma is a well-recognized model, however, the genetic alterations during carcinogenesis have yet to be determined. We used laser capture microdissection to specifically isolate the cells from terminal end buds (TEBs), the origin of MC, at 2 weeks after sesame oil-(control) or DMBA-treatment (DMBA-TEBs), ductal carcinoma in situ (DCIS) and invasive mammary carcinoma (MC). At 50 days after birth, inbred Sprague-Dawley female rats were given 1ml sesame oil (control) or 20 mg DMBA (Nacalai tesque, Inc. Kyoto Japan) dissolved in 1ml sesame oil, by gastric intubations. Animals in control group were euthanized after 2 weeks (W) and DMBA-treated group were euthanized after 2 weeks (W), 6W and 8W, respectively. From for 4 sample(control, DMBA-TEBs, DCIS, MC), we performed a hybridization repeatedly, respectively.
Project description:Mammary gland branching morphogenesis is thought to depend on the mobilization of the membrane-anchored matrix metalloproteinases, MT1-MMP and MT2-MMP, that drive epithelial cell invasion by remodeling the extracellular matrix and triggering associated signaling cascades. However, the roles that these proteinases play during mammary gland development in vivo remains undefined. A mammary gland branching program that occurs during the first 10 days of early postnatal development was used to characterize the impact of global Mt1-mmp or Mt2-mmp targeting on mammary gland morphogenesis. Transcriptome profiling of ductal networks and associated stroma was used to investigate the functional roles of MT2-MMP in the early postnatal mammary gland in an unbiased fashion.