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Mammary Epithelial Cells (mammary + epithelial_cell)
Kinds of Mammary Epithelial Cells Terms modified by Mammary Epithelial Cells Selected AbstractsCentrosome amplification induced by the antiretroviral nucleoside reverse transcriptase inhibitors lamivudine, stavudine, and didanosineENVIRONMENTAL AND MOLECULAR MUTAGENESIS, Issue 8 2009Mia Yu Abstract In cultured cells, exposure to the nucleoside reverse transcriptase inhibitor (NRTI) zidovudine (AZT) induces genomic instability, cell cycle arrest, micronuclei, sister chromatid exchanges, and shortened telomeres. In previous studies, we demonstrated AZT-induced centrosome amplification (>2 centrosomes/cell). Here, we investigate centrosome amplification in cells exposed to other commonly used NRTIs. Experiments were performed using Chinese Hamster ovary (CHO) cells, and two normal human mammary epithelial cell (NHMEC) strains: M99005 and M98040, which are high and low incorporators of AZT into DNA, respectively. Cells were exposed for 24 hr to lamivudine (3TC), stavudine (d4T), didanosine (ddI), and thymidine, and stained with anti-pericentrin antibody. Dose response curves were performed to determine cytotoxicity and a lower concentration at near plasma levels and a 10 fold higher concentration were chosen for the experiments. In CHO cells, there was a concentration-dependent, significant (P < 0.05) increase in centrosome amplification for each of the NRTIs. In NHMEC strain M99005, an NRTI-induced increase (P < 0.05) in centrosome amplification was observed for the high concentrations of each NRTI and the low doses of 3TC and ddI. In NHMEC strain M98040, the high doses of ddI and d4T showed significant increases in centrosome amplification. Functional viability of amplified centrosomes was assessed by arresting microtubule nucleation with nocodazole. In cells with more than two centrosomes, the ability to recover microtubule nucleation was similar to that of unexposed cells. We conclude that centrosome amplification is a consequence of exposure to NRTIs and that cells with centrosome amplification are able to accomplish cell division. Environ. Mol. Mutagen., 2009. © 2009 Wiley-Liss, Inc. [source] Circadian clock and cell cycle gene expression in mouse mammary epithelial cells and in the developing mouse mammary glandDEVELOPMENTAL DYNAMICS, Issue 1 2006Richard P. Metz Abstract Mouse mammary epithelial cells (HC-11) and mammary tissues were analyzed for developmental changes in circadian clock, cellular proliferation, and differentiation marker genes. Expression of the clock genes Per1 and Bmal1 were elevated in differentiated HC-11 cells, whereas Per2 mRNA levels were higher in undifferentiated cells. This differentiation-dependent profile of clock gene expression was consistent with that observed in mouse mammary glands, as Per1 and Bmal1 mRNA levels were elevated in late pregnant and lactating mammary tissues, whereas Per2 expression was higher in proliferating virgin and early pregnant glands. In both HC-11 cells and mammary glands, elevated Per2 expression was positively correlated with c-Myc and Cyclin D1 mRNA levels, whereas Per1 and Bmal1 expression changed in conjunction with ,- casein mRNA levels. Interestingly, developmental stage had differential effects on rhythms of clock gene expression in the mammary gland. These data suggest that circadian clock genes may play a role in mouse mammary gland development and differentiation. Developmental Dynamics 235:263,271, 2006. © 2005 Wiley-Liss, Inc. [source] Expression of the whey acidic protein (Wap) is necessary for adequate nourishment of the offspring but not functional differentiation of mammary epithelial cellsGENESIS: THE JOURNAL OF GENETICS AND DEVELOPMENT, Issue 1 2005Aleata A. Triplett Abstract Whey acidic protein (WAP) is the principal whey protein found in rodent milk, which contains a cysteine-rich motif identified in some protease inhibitors and proteins involved in tissue modeling. The expression of the Wap gene, which is principally restricted to the mammary gland, increases more than 1,000-fold around mid-pregnancy. To determine whether the expression of this major milk protein gene is a prerequisite for functional differentiation of mammary epithelial cells, we generated conventional knockout mice lacking two alleles of the Wap gene. Wap-deficient females gave birth to normal litter sizes and, initially, produced enough milk to sustain the offspring. The histological analysis of postpartum mammary glands from knockout dams does not reveal striking phenotypic abnormalities. This suggests that the expression of the Wap gene is not required for alveolar specification and functional differentiation. In addition, we found that Wap is dispensable as a protease inhibitor to maintain the stability of secretory proteins in the milk. Nevertheless, a significant number of litters thrived poorly on Wap-deficient dams, in particular during the second half of lactation. This observation suggests that Wap may be essential for the adequate nourishment of the growing young, which triple in size within the first 10 days of lactation. Important implications of these findings for the use of Wap as a marker for advanced differentiation of mammary epithelial cells and the biology of pluripotent progenitors are discussed in the final section. genesis 43:1,11, 2005. © 2005 Wiley-Liss, Inc. [source] Loss of RAB25 expression in breast cancerINTERNATIONAL JOURNAL OF CANCER, Issue 12 2006Ji-Ming Cheng Abstract A novel breast cancer cell line (RAO-3) was established by transduction of the Q61L mutant RAS into human mammary epithelial cells that were immortalized with catalytic subunit of telomerase (hTERT). The cells displayed anchorage-independent growth and proliferation, and formed human mammary spindle cell carcinoma when injected into nude mice. Chromosome locus 1q22-23 was partially duplicated and inverted on one of the 3 chromosomes present in the cell line. We report here that mutations of chromosome 1q22-23 locus have resulted in the loss of RAB25 expression in the breast cancer cell line. Transduction of RAB25 into the breast cancer cell line arrests anchorage-independent growth. We have also demonstrated loss of RAB25 in human breast tumor tissue. These data suggest that loss of RAB25 might contribute to tumorigenesis of breast cancer, and RAB25 is likely to be an important factor in the development of breast cancer. RAB25 could be used as biological marker of breast cancer and provides a target for gene replacement therapy. © 2006 Wiley-Liss, Inc. [source] TCDD causes suppression of growth and differentiation of MCF10A, human mammary epithelial cells by interfering with their insulin receptor signaling through c-Src kinase and ERK activationJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 6 2005Sujin Park Abstract One of the proposed mechanisms of carcinogenic action of TCDD (=dioxin) on breast cells is that it causes significant inhibition of proper differentiation of mammary duct epithelial cells and thereby increases the number of terminal end buds, which are susceptible to other carcinogens (Fenton et al., Toxicol Sci 2002;67:63,74; Brown et al., Carcinogenesis 1998; 19:1623,1629; Lamartiniere, J Mammary Gland Biol Neoplasia 2002;7:67,76). To address this topic, we selected MCF10A, a line of immortalized normal human breast epithelial cells as an in vitro model. An initial effort was made to optimize the cultural condition of MCF10A cells to promote the cell differentiation effect of insulin. Under this condition, TCDD clearly antagonized the action of insulin only in the presence of cholera toxin that is known to promote the differentiation of normal human breast epithelial cells. To test the hypothesis that TCDD-induced c-Src kinase activation is casually related to this compound's antagonistic action against insulin, we treated MCF10A cells with two c-Src blocking agents, an anti-Src antisense oligonucleotides blocker and a known specific inhibitor of c-Src kinase, PP-2 and studied the effect of insulin and TCDD on cell proliferation. The results showed that, in cells treated with either of these two c-Src blocking agents, the antagonistic effect of TCDD disappeared. It was also found that agents which specifically block the activation of ERK could also abrogate the action of TCDD to suppress insulin signaling. Together, these results indicate that the mechanism of the antagonistic action of TCDD on insulin signaling is mainly mediated through c-Src signaling through activation of ERK. © 2005 Wiley Periodicals, Inc. J Biochem Mol Toxicol 18:322,331, 2004; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20040 [source] Negative effects of the amino acids Lys, His, and Thr on S6K1 phosphorylation in mammary epithelial cellsJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 4 2008Rotem Ladovsky Prizant Abstract The role of essential amino acids (AA) on protein synthesis via the mTOR pathway was studied in murine mammary epithelial cells cultured under lactogenic conditions. Leu, Ile, and Val increased S6K1 phosphorylation compared to that measured in AA-deprived cells. Trp, Phe, and Met had no effect. Surprisingly, Lys, His, and Thr inhibited S6K1 phosphorylation in both murine and bovine mammary cells. Thr exhibited the most potent inhibition, being the only amino acid that competed with Leu's positive role. In non-deprived cells, there was no observable effect of Lys, His, or Thr on S6K1 phosphorylation at concentrations up to five times those in the medium. However, their addition as a mix revealed a synergistic negative effect. Supplementation of Lys, His, and Thr abrogated mTOR Ser 2448 phosphorylation, with no effect on Akt Ser 473,an mTORC2 target. This confirms specific mTORC1 regulation of S6K1 phosphorylation. The individual supplementation of Lys, His, and Thr maintained a low level of IRS-1 phosphorylation, which was dose-dependently increased by their combined addition. Thus, in parallel to inhibiting S6K1 activity, these AA may act synergistically to activate an additional kinase, phosphorylating IRS-1 via an S6K1-independent pathway. In cultures supplemented by Lys, His, and Thr, cellular protein synthesis decreased by up to 65%. A more pronounced effect was observed on ,-casein synthesis. These findings indicate that positive and negative signaling from AA to the mTOR pathway, combined with modulation of insulin sensitization, mediate the synthesis rates of total and specific milk proteins in mammary epithelial cells. J. Cell. Biochem. 105: 1038,1047, 2008. © 2008 Wiley-Liss, Inc. [source] TGF-, inhibits prolactin-induced expression of ,-casein by a Smad3-dependent mechanism,JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 5 2008Wen-Jun Wu Abstract Transforming growth factor-, (TGF-,) is a multifunctional growth factor, affecting cell proliferation, apoptosis, and extracellular matrix homeostasis. It also plays critical roles in mammary gland development, one of which involves inhibition of the expression of milk proteins, such as ,-casein, during pregnancy. Here we further explore the underlying signaling mechanism for it. Our results show that TGF-, suppresses prolactin-induced expression of ,-casein mRNA and protein in primary mouse mammary epithelial cells, but its effect on protein expression is more evident. We also find out that this inhibition is not due to the effect of TGF-, on cell apoptosis. Furthermore, inhibition of TGF-, type I receptor kinase activity by a pharmacological inhibitor SB431542 or overexpression of dominant negative Smad3 substantially restores ,-casein expression. By contrast, inhibition of p38 and Erk that are known to be activated by TGF-, does not alleviate the inhibitory effect of TGF-,. These results are consistent with our other observation that Smad but not MAPK pathway is activated by TGF-, in mammary epithelial cells. Lastly, we show that prolactin-induced tyrosine phosphorylation of Jak2 and Stat5 as well as serine/threonine phosphorylation of p70S6K and S6 ribosomal protein are downregulated by TGF-,, although the former event requires considerably long exposure to TGF-,. We speculate that these events might be involved in repressing transcription and translation of ,-casein gene, respectively. Taken together, our results demonstrate that TGF-, abrogates prolactin-stimulated ,-casein gene expression in mammary epithelial cells through, at least in part, a Smad3-dependent mechanism. J. Cell. Biochem. 104: 1647,1659, 2008. © 2008 Wiley-Liss, Inc. [source] Extracellular matrix regulates alpha s1-casein gene expression in rabbit primary mammary cells and CCAAT enhancer binding protein (C/EBP) binding activityJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 3 2001Geneviève Jolivet Abstract Previous studies have shown that both the signal transducer and activator of transcription 5 (STAT5) and the CCAAT enhancer binding proteins (C/EBPs) are involved in the regulation of casein gene expression by mammary epithelial cells. Prolactin (Prl) activation of STAT5 is necessary for casein gene expression. The extracellular matrix (ECM) regulates also casein gene expression. Here, we have investigated whether ECM regulates C/EBPs activity in primary rabbit mammary epithelial cells. Isolated primary mammary cells were cultured on plastic or on floating collagen I gel. Prolactin induced ,s 1-casein gene expression when cells were cultured on collagen but not on plastic. It is noteworthy that activated STAT5 was detected in both culture conditions. Several STAT5 isoforms (STAT5a, STAT5b, and other STAT5 related isoforms, some with lower molecular weight than the full-length STAT5a and STAT5b) were detected under the different culture conditions. However, their presence was not related to the expression of ,s 1-casein gene. The binding of nuclear factors to a C/EBP specific binding site and the protein level of C/EBP, differed in cells cultured on plastic or on collagen but these parameters were not modified by Prl. This suggests that C/EBP binding activity was regulated by ECM and not by Prl. Interestingly, these modifications were correlated to the expression of the ,s 1-casein gene. Hence, the activation of the ,s 1-casein gene expression depends on two independent signals, one delivered by Prl via the activation of STAT5, the other delivered by ECM via C/EBP. J. Cell. Biochem. 82:371,386, 2001. © 2001 Wiley-Liss, Inc. [source] Retinoic acid induces expression of the interleukin-1, gene in cultured normal human mammary epithelial cells and in human breast carcinoma linesJOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2002Limin Liu Retinoic acid (RA) and its derivatives inhibit the proliferation of normal human mammary epithelial cells (HMEC) and some breast carcinoma lines by mechanisms which are not fully understood. To identify genes that mediate RA-induced cell growth arrest, an HMEC cDNA library was synthesized and subtractive screening was performed. We identified the interleukin-1, (IL-1,) gene as an RA induced gene in HMEC. Northern blot analyses showed that the IL-1, gene was up-regulated as early as 2 h after RA treatment. Results from the treatment of HMEC with cycloheximide and actinomycin D indicated that the regulation of the IL-1, gene by RA occurred at the transcriptional level and that the IL-1, gene is a direct, downstream target gene of RA. To evaluate the effects of IL-1, on cell proliferation, the proliferation of HMEC was measured in the presence of RA or IL-1,, or both. Either RA or IL-1, could significantly inhibit the proliferation of HMEC. However, the addition of soluble IL-1 receptor antagonist (sIL-1ra) to the cell culture medium did not block RA-induced HMEC growth inhibition, whereas sIL-1ra did block the growth inhibition of HMEC by IL-1,. IL-1, expression was not observed in the three carcinoma cell lines, MCF-7, MDA-MB-231, and MDA-MB-468, as compared to the HMEC. Growth curves of the breast carcinoma cell lines showed strong inhibitory effects of RA and IL-1, on the growth of the estrogen receptor (ER) positive MCF-7 cell line, but only a small effect on the ER negative MDA-MB-231 cells. The expression of the IL-1, gene was also transcriptionally activated by RA in normal epithelial cells of prostate and oral cavity. Our results suggest that: (a) the IL-1, gene is a primary target of RA receptors in HMEC; (b) the enhanced expression of the IL-1, gene does not mediate the RA-induced growth arrest of HMEC; and (c) the expression of the IL-1, gene is low or absent in all three human breast carcinoma cell lines examined, but the defect in the IL-1, signaling pathway may be different in ER positive versus ER negative carcinoma cells. © 2002 Wiley-Liss, Inc. [source] Induction or suppression of expression of cytochrome C oxidase subunit II by heregulin , 1 in human mammary epithelial cells is dependent on the levels of ErbB2 expressionJOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2002Yanbo Sun The ErbB family of receptor kinases is composed of four members: epidermal growth factor receptor (EGFR/ErbB1), ErbB2/neu, ErbB3, and ErbB4. Amplification of the ErbB2/neu is found in about 30% of breast cancer patients and is associated with a poor prognosis. Heregulin (HRG) activates the ErbB2 via induction of heterodimerization with ErbB3 and ErbB4 receptors. With suppression subtractive hybridization, we demonstrated that the expression of cytochrome c oxidase subunit II (COXII) is HRG-responsive. Two nontransformed human mammary epithelial cell lines, the HB2 and the HB2ErbB2 (the HB2 engineered to overexpress ErbB2), displayed an opposite response to HRG-mediated regulation. HRG upregulated mRNA expression of COXII in the HB2 cells, but suppressed COXII expression in the HB2ErbB2 cells. A human breast cancer cell line (T47D), which expresses ErbB2 at a level similar to that of the HB2 cells, also responded to HRG by increasing COXII mRNA levels. Therefore, HRG regulation of COXII expression depends on the levels of ErbB2 expression. Furthermore, the expression of COXII was inversely correlated to the levels of ErbB2, i.e., the cells overexpressing ErbB2 exhibited lower COXII levels. HRG-evoked signal transduction differed between the cells with normal ErbB expression and the cells overexpressing ErbB2. The activation of both ERK and PI3-K was essential for HRG regulation of COXII, i.e., blockage of either pathway eliminated HRG-mediated alteration. This is the first report demonstrating that the expression of mitochondria-encoded COXII is HRG-responsive. The levels of ErbB2 expression are decisive for the diverse biological activities of HRG. © 2002 Wiley-Liss, Inc. [source] Autocrine TGF, signaling mediates vitamin D3 analog-induced growth inhibition in breast cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2001Limin Yang In this study, we address whether TGF, signaling mediates vitamin D3 analog-induced growth inhibition in nonmalignant and malignant breast cells. Normal mammary epithelial cells (184), immortalized nonmalignant mammary epithelial cells (184A1 and MCF10A), and breast cancer cells (early passage MCF7: MCF7E) were sensitive to the inhibitory effects of vitamin D3 analogs (EB1089 and MC1288) while late passage MCF7 breast cancer (MCF7L) cells were relatively resistant. A similar pattern of sensitivity to TGF, was observed with these cells. Thus, the sensitivity to the vitamin D3 analogs correlated with the sensitivity to TGF,. MCF7L TGF,RII-transfected cells, which have autocrine TGF, activity, were more sensitive to EB1089 than MCF7L cells. TGF, neutralizing antibody was found to block the inhibitory effects of these analogs. These results are consistent with the idea that autocrine TGF, signaling mediates the anti-proliferative effects of the vitamin D3 analogs in these cells. The expression of TGF, isoforms and/or TGF, receptors was induced by the analogs in the vitamin D3 and TGF, sensitive cells. Vitamin D3 analogs did not induce TGF, or TGF, receptor expression in the resistant MCF7L cells. Therefore, EB1089 induces autocrine TGF, activity through increasing expression of TGF, isoforms and/or TGF, receptors. In addition, EB1089 induced nuclear VDR protein levels in the sensitive 184A1 cells but not in the resistant MCF7L cells. 184A1 cells were more sensitive to EB1089-induced VDR-dependent transactivation than MCF7L cells as measured by a luciferase reporter construct containing the VDRE, indicating a defect of VDR signaling in MCF7L cells. Smad3, a TGF, signaling mediator, coactivated VDR-dependent transactivation in 184A1 cells but not in MCF7L cells. These results indicate that Smad3 coactivates VDR to further enhance TGF, signaling and vitamin D3 signaling in the sensitive 184A1 cells. The results also indicate that Smad3 is not of itself sufficient to coactivate VDR in TGF,/vitamin D3 resistant MCF7L cells and other factors are required. We found that the PI 3-kinase pathway inhibitor LY29004 inhibited the synergy of TGF, and EB1089 on VDR-dependent transactivation activity. This indicates that the crosstalk between TGF, and vitamin D signaling is also PI 3-kinase pathway dependent. © 2001 Wiley-Liss, Inc. [source] TNF, induces NF,B/p50 in association with the growth and morphogenesis of normal and transformed rat mammary epithelial cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 1 2001Linda M. Varela In contrast to the cytotoxic or cytostatic effect of TNF, on many breast cancer cell lines, TNF, stimulates growth and morphogenesis of normal rat mammary epithelial cells (MEC). The present studies were carried out to determine whether there are intrinsic differences between normal and malignant MEC which may explain the differing responsiveness to TNF,. Freshly isolated rat MEC organoids from normal mammary gland or 1-methyl-1-nitrosourea-induced mammary tumors were treated with TNF, for 21 days. Unexpectedly, TNF, stimulated growth and morphogenesis of both normal and transformed MEC in primary culture, although in transformed cells its effects were delayed and the majority of the colonies were histologically abnormal, with multiple cell layers and no lumen. Since NF,B is a key mediator of TNF, action and has been implicated in carcinogenesis, the expression of the p50, p52, p65, and c-rel NF,B proteins in normal and transformed MEC was determined. Expression of p52 was significantly reduced in tumor cells, and p50 was absent, although its putative precursor, p105 was abundant. There were no changes in the levels of p65 or c-rel. TNF, induced a pronounced and sustained increase of a p50 homodimeric NF,B/DNA complex in both normal and transformed MEC. However, in transformed MEC, NF,B binding was initially undetectable but then increased in response to TNF,. Thus, NF,B expression and DNA binding activity are altered during mammary carcinogenesis. In addition, the significant increase in NF,B/p50 DNA-binding was temporally coincident with TNF,-induced growth and morphogenesis, suggesting that it may play a significant role in both normal development and carcinogenesis. © 2001 Wiley-Liss, Inc. [source] Bioactivity of falcarinol and the influenceof processing and storage on its content in carrots (Daucus carota L)JOURNAL OF THE SCIENCE OF FOOD AND AGRICULTURE, Issue 10 2003Susanne L Hansen Abstract The concentration-dependent activity of the polyacetylene falcarinol ((9Z)-heptadeca-1,9-dien-4,6-diyn-3-ol), isolated from carrots, was investigated in a bioassay with primary mammary epithelial cells in collagen gels and compared with that of ,-carotene, the orange pigment in carrots. Falcarinol showed biphasic activity, having stimulatory effects between 0.01 and 0.05 µg ml,1 and inhibitory effects between 1 and 10 µg ml,1, whereas ,-carotene showed no effect in the concentration range 0.001,100 µg ml,1. The results are discussed in relation to the health-promoting effects of carrots and related vegetables. Falcarinol was quantified in the carrot cultivars Bolero, Rodelika and Fancy by analytical reverse phase HPLC, subjected to various processing and storage conditions in order to study how long-term storage, blanching, freezing and boiling influence the content of falcarinol. Long-term storage of raw carrot cubes (1 cm3) reduced the falcarinol content by almost 35%. A similar reduction was found in steam-blanched carrot cubes (1 cm3). Long-term storage at ,24 °C of steam blanched carrot cubes did not reduce the falcarinol content further. A reduction of almost 70% in the falcarinol content was found in carrot pieces boiled in water for 12 min compared with raw carrots. Copyright © 2003 Society of Chemical Industry [source] Retinoids and retinoic acid receptors regulate growth arrest and apoptosis in human mammary epithelial cells and modulate expression of CBP/p300MICROSCOPY RESEARCH AND TECHNIQUE, Issue 1 2002Eric C. Dietze Abstract Retinoids and retinoic acid receptors (RARs) are important mediators of normal epithelial cell homeostasis. To assess the role of retinoids and RARs in regulating growth arrest and apoptosis in benign and malignant mammary epithelial cells, two model systems were developed: 1) RAR function was suppressed in retinoid-sensitive normal human mammary epithelial cells (HMECs) by the dominant-negative retinoic acid receptor, RAR,403 (DNRAR), and 2) retinoid-resistant MCF-7 breast cancer cells were transduced with a functional RAR,2. Inhibition of RAR function by the DNRAR in HMECs resulted in retinoid-resistance, increased proliferation, and dysregulated growth when cells were cultured in reconstituted extracellular matrix (rECM). Expression of RAR,2 in MCF-7 cells resulted in sensitivity to retinoid-induced growth arrest and apoptosis. The CREB-binding protein (CBP) and the homologous protein p300 are tightly regulated, rate-limiting integrators of diverse signaling pathways and are recruited during retinoid-mediated transcriptional activation. The relationship between retinoid receptor expression, growth regulation, and transcriptional regulation of CBP/p300 is poorly understood. Inhibition of RAR function in HMECs by DNRAR suppressed expression of CBP/p300 and expression of RAR,2 in MCF-7 cells promoted induction of CBP/p300 when cells were treated with 1.0 ,M all- trans -retinoic acid (ATRA). These results suggest that ATRA and RARs regulate growth arrest of HMECs and modulate CBP/p300 protein expression. Since CBP and p300 are normally present in limiting amounts, their regulation by ATRA and RARs may be an important element in the control of transcriptional activation of genes regulating growth arrest and apoptosis. Microsc. Res. Tech. 59:23,40, 2002. © 2002 Wiley-Liss, Inc. [source] Expression of chromogranin/secretogranin mRNA in spontaneous mammary tumors in aging Fischer-344 ratsPATHOLOGY INTERNATIONAL, Issue 9 2001Shinobu Umemura There is a type of human breast cancer showing a neuroendocrine differentiation. Little is known, however, about the cell origin of this cancer or the process by which it expresses neuroendocrine features. Rat mammary tumors, either spontaneous or induced, have not been subjects for the investigation of aspects regarding the neuroendocrine differentiation of mammary epithelial cells. The aim of the present study was to show the potential of rat mammary tumors for expressing chromogranin (Cg)/secretogranin (Sg) mRNA. We examined CgA, SgI and SgII mRNA expression by reverse transcription,polymerase chain reaction in rat mammary adenocarcinoma and fibroadenoma which had arisen spontaneously in aging Fischer-344 rats. CgA and SgII mRNA were expressed in both mammary tumors, but SgI mRNA was not detected in either. The results of the present study show that rat mammary tumors can express chromogranin genes. [source] Improved proteome coverage by using high efficiency cysteinyl peptide enrichment: The human mammary epithelial cell proteomePROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 5 2005Tao Liu Abstract Automated multidimensional capillary liquid chromatography-tandem mass spectrometry (LC-MS/MS) has been increasingly applied in various large scale proteome profiling efforts. However, comprehensive global proteome analysis remains technically challenging due to issues associated with sample complexity and dynamic range of protein abundances, which is particularly apparent in mammalian biological systems. We report here the application of a high efficiency cysteinyl peptide enrichment (CPE) approach to the global proteome analysis of human mammary epithelial cells (HMECs) which significantly improved both sequence coverage of protein identifications and the overall proteome coverage. The cysteinyl peptides were specifically enriched by using a thiol-specific covalent resin, fractionated by strong cation exchange chromatography, and subsequently analyzed by reversed-phase capillary LC-MS/MS. An HMEC tryptic digest without CPE was also fractionated and analyzed under the same conditions for comparison. The combined analyses of HMEC tryptic digests with and without CPE resulted in a total of 14,416 confidently identified peptides covering 4294 different proteins with an estimated 10%,gene coverage of the human genome. By using the high efficiency CPE, an additional 1096 relatively low abundance proteins were identified, resulting in 34.3% increase in proteome coverage; 1390,proteins were observed with increased sequence coverage. Comparative protein distribution analyses revealed that the CPE method is not biased with regard to protein Mr,, pI, cellular location, or biological functions. These results demonstrate that the use of the CPE approach provides improved efficiency in comprehensive proteome-wide analyses of highly complex mammalian biological systems. [source] The influence of extracellular matrix and prolactin on global gene expression profiles of primary bovine mammary epithelial cells in vitroANIMAL GENETICS, Issue 1 2010L. G. Riley Summary An in vitro bovine mammosphere model was characterized for use in lactational biology studies using a functional genomics approach. Primary bovine mammary epithelial cells cultured on a basement membrane, Matrigel, formed three-dimensional alveoli-like structures or mammospheres. Gene expression profiling during mammosphere formation by high-density microarray analysis indicated that mammospheres underwent similar molecular and cellular processes to developing alveoli in the mammary gland. Gene expression profiles indicated that genes involved in milk protein and fat biosynthesis were expressed, however, lactose biosynthesis may have been compromised. Investigation of factors influencing mammosphere formation revealed that extracellular matrix (ECM) was responsible for the initiation of this process and that prolactin (Prl) was necessary for high levels of milk protein expression. CSN3 (encoding ,-casein) was the most highly expressed casein gene, followed by CSN1S1 (encoding ,S1-casein) and CSN2 (encoding ,-casein). Eighteen Prl-responsive genes were identified, including CSN1S1, SOCS2 and CSN2, however, expression of CSN3 was not significantly increased by Prl and CSN1S2 was not expressed at detectable levels in mammospheres. A number of novel Prl responsive genes were identified, including ECM components and genes involved in differentiation and apoptosis. This mammosphere model is a useful model system for functional genomics studies of certain aspects of dairy cattle lactation. [source] Latent transforming growth factor binding protein 4 (LTBP-4) is downregulated in human mammary adenocarcinomas in vitro and in vivo,APMIS, Issue 6 2007SUSANNE MAUEL Transforming growth factor beta (TGF-ß) is able to inhibit proliferation of epithelial cells and is involved in the carcinogenesis of human mammary tumours. Three latent transforming growth factor-ß binding proteins (LTBP-1, -3 and -4) are involved in TGF-ß function. The aim of the study was to analyze the expression profiles of TGF-ß 1 and 2 and LTBP-4 in human mammary carcinoma cell lines as well as in human mammary tumours. Expression analysis was performed at the transcription and protein level under in vivo and in vitro conditions. LTBP-4 expression was quantitatively analysed in human carcinomas of the mammary gland and in healthy mammary tissues of the same patients. Downregulation of LTBP-4 in all investigated human mammary tumours compared to normal tissues could be demonstrated. Results also revealed that protein levels of TGF-ß 1 are downregulated and of TGF-ß 2 are upregulated in human mammary carcinoma cell lines compared to primary (normal) human mammary epithelial cells. LTBP-4 reduction in neoplasms leads to a possible decrease of TGF-ß 1 extracellular deposition with reduced TGF-ß 1 bioavailability. TGF-ß 2 was upregulated, which indicates a possible compensatory mechanism. This study demonstrated a possible functional role of LTBP-4 for TGF-ß bioavailability with respect to carcinogenesis of human mammary tumours in vivo and in vitro. [source] The accumulation of ,-Tocopherol and Retinol in the milk of water buffalo is correlated with the plasma levels of triiodothyronineBIOFACTORS, Issue 3-4 2003M. S. Spagnuolo Abstract Milk is the most important source of Retinol and ,-Tocopherol for calves. These antioxidants save the food quality and prevent lipid oxidation in the mammary gland and the calf growing tissues. In Bubalus bubalis, seasonal changes for the plasma levels of both antioxidants were not found. The levels of Retinol and ,-Tocopherol in the milk were 2 and 1.7 times higher in winter than in summer, respectively. These levels were correlated with the plasma level of triiodothyronine, and markedly increased in cows injected with triiodothyronine in summer. The cytosol from alveolar epithelial cells of mammary glands was incubated with ,-Tocopherol and 3H-Retinol and, after gel filtration chromatography, both antioxidants were found associated with proteins migrating as a single peak of 33 kD. The amount of ,-Tocopherol and Retinol binding proteins was 1.5 and 2.3 times higher in winter than in summer respectively. The Retinol binding proteins migrated as two bands (33 and 16 kD) by electrophoresis in denaturing and reducing conditions. Our data suggest that triiodothyronine enhances the transport of both liposoluble antioxidants through the blood-mammary barrier, and demonstrate that proteins of the mammary epithelial cells are involved in such a transport. [source] Vitamin E inhibition of normal mammary epithelial cell growth is associated with a reduction in protein kinase C, activationCELL PROLIFERATION, Issue 6 2001P. W. Sylvester Tocopherols and tocotrienols represent the two subclasses within the vitamin E family of compounds. However, tocotrienols are significantly more potent than tocopherols in suppressing epidermal growth factor (EGF)-dependent normal mammary epithelial cell growth. EGF is a potent mitogen for normal mammary epithelial cells and an initial event in EGF-receptor mitogenic-signalling is protein kinase C (PKC) activation. Studies were conducted to determine if the antiproliferative effects of specific tocopherol and tocotrienol isoforms are associated with a reduction in EGF-receptor mitogenic signalling and/or PKC activation. Normal mammary epithelial cells isolated from midpregnant BALB/c mice were grown in primary culture, and maintained on serum-free media containing 10 ng/mL EGF as a mitogen, and treated with various doses (0,250 µm) of ,-, ,-, or ,-tocopherol or ,-, ,-, or ,-tocotrienol. Treatment with growth inhibitory doses of ,-tocopherol (100 µm), ,-tocotrienol (50 µm), or ,- or ,-tocotrienol (10 µm) did not affect EGF-receptor levels, EGF-induced EGF-receptor tyrosine kinase activity, or total intracellular levels of PKC,. However, these treatments were found to inhibit EGF-induced PKC, activation as determined by its translocation from the cytosolic to membrane fraction. Treatment with 250 µm,- or ,-tocopherol had no affect on EGF-receptor mitogenic signalling or cell growth. These findings demonstrate that the inhibitory effects of specific tocopherol and tocotrienol isoforms on EGF-dependent normal mammary epithelial cell mitogenesis occurs downstream from the EGF-receptor and appears to be mediated, at least in part, by a reduction in PKC, activation. [source] | ||||||||||||||||