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1.
Correlation of Physical and Biological Properties of Mouse Mammary Tumor Agent 总被引:2,自引:3,他引:2
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Dan H. Moore E. Y. Lasfargues Margaret R. Murray Cushman D. Haagensen E. C. Pollard 《The Journal of cell biology》1959,5(1):85-92
Biophysical procedures have been used to determine the size and structure of the biologically active agent responsible for the transmission, through milk, of mouse mammary adenocarcinoma. Filtration of milk from RIII high-breast-cancer mice through gradocol membranes with decreasing pore sizes indicated that a minimum of activity passed through intermediate pore sizes (100 to 160 mµ). Filtrates through smaller pores were significantly active. Milk treated with small doses of deuteron irradiation produced more tumors than the control, unirradiated milk; larger doses indicated a particle size much less than 100 mµ. Free diffusion experiments indicated that the activity was associated with particles of two different sizes. Altogether the data denoted the presence of a large agent about 100 mµ in diameter and a small agent 20 to 30 mµ in diameter or possibly smaller. Furthermore, the presence in the milk of an inhibitor 40 to 60 mµ is indicated by the results of all three approaches. The complex nature of the milk agent disclosed by the physical measurements agrees with the picture of one of the structures revealed by electron microscopy as well as with seemingly conflicting measurements reported in the literature. The large agent defined by these indirect methods corresponds to the whole particle seen in the electron microscope and the small agent corresponds to its internal core or nucleoid. It is suggested that the nucleoid is essentially a nucleic acid which may, in the absence of the "inhibitor," retain its activity after being stripped of its outer membrane or sac. 相似文献
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Growth and Differentiation of Primary Cultures of Mouse Mammary Epithelium Embedded in Collagen Gel 总被引:1,自引:0,他引:1
DARRAGH FLYNN JASON YANG S. NANDI 《Differentiation; research in biological diversity》1982,22(1-3):191-194
Mammary glands from BALB/cfC3H midpregnant (9–11 days) mice were dissociated with collagenase and pronase, separated on a Percoll gradient, and the epithelial cells were cultured inside collagen gel. The cell number increased three-to five-fold when cultured for 6–8 days in DME/F12 (1: 1) medium containing 3% swine serum, insulin (10 μg/ml), cortisol (1.0 μg/ml), prolactin (10 μg/ml), transferrin (10 μg/ml), and epidermal growth factor (0.01 μg/ml). The casein level, as determined by radioimmunoassay, at the end of this growth phase, was much lower than that present in freshly dissociated cells. In order to stimulate casein production, the gels were released from the sides of the plastic dish and allowed to float for eight days in Waymouth's medium, containing insulin (10 μg/ml), cortisol (5 μg/ml), prolactin (10 μg/ml), and 0.25% bovine serum albumin. The casein level at the end of this differentiation phase was found to be comparable to that seen in the original freshly dissociated cells. Cells grown in DME/F12 (1: 1) medium containing 3% swine serum, insulin (10 μg/ml), and transferrin (10 μg/ml) were still capable of undergoing casein production, indicating that the presence of exogenous lactogenic hormones such as cortisol and prolactin, as well as exogenous growth factors such as epidermal growth factor, is not necessary during the growth phase for subsequent casein production during the differentiation phase. Two factors that seemed more important for subsequent casein stimulation were: (1) releasing collagen gels at the beginning of the differentiation phase, and (2) switching to'differentiation' medium. This present two-step protocol has allowed primary cultures of dissociated midpregnant mouse mammary epithelial cells to undergo several rounds of division inside a collagen gel matrix and to be subsequently stimulated to produce the mammary-specific protein, casein. 相似文献
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D. L. Fine L. O. Arthur J. K. Plowman E. A. Hillman F. Klein 《Applied microbiology》1974,28(6):1040-1046
An in vitro system for production, purification, and concentration of mouse mammary tumor virus is described. Monolayer cultures of C(3)H mouse mammary tumor cells propagated at 34 C in roller bottles in the presence of dexamethasone, a glucocorticoid hormone, release B-type particles which possess ribonucleic acid and a ribonucleic acid-dependent deoxyribonucleic acid polymerase. One thousandfold concentration by ultracentrifugation with subsequent gradient fractionation yielded > 7 x 10(10) particles per ml in the 1.16- to 1.18-g/ml region. Mouse mammary tumor virus produced in this system was free of detectable C-type virus. 相似文献
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组织纤溶酶原激活剂突变体在转基因小鼠乳腺中的表达 总被引:1,自引:0,他引:1
组织纤溶酶原激活剂(tPA)是一种较理想的溶血栓药物,本研究采用其突变体——长效组织纤溶酶原激活剂(LAtPA)的cDNA作为目的基因,将它插入羊β-乳球蛋白(BLG)基因起始密码之前,使LAtPA的转录、翻译受控于BLG基因的5′、3′序列,再将所构建的BLG-LAtPA用显微注射方法建立转基因鼠,经点杂交筛选和Southern印迹鉴定,获得2只LAtPA基因整合阳性鼠,并在阳性母鼠乳汁中检测到有溶纤活性的LAtPA表达,表达水平1.5μg/ml。在这两只转基因鼠的9只F1代子鼠中,有5只是阳性的,tPA表达水平维持在1~2μg/ml,BLGtPA融合基因整合到小鼠基因组,能稳定地遗传给子代。 相似文献
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Milk Protein Synthesis, Gene Expression, and Hormonal Responsiveness in Primary Cultures of Mammary Cells from Lactating Sheep 总被引:1,自引:0,他引:1
Thomas T. Wheeler Megan R. Callaghan Stephen R. Davis Colin G. Prosser Richard J. Wilkins 《Experimental cell research》1995,217(2)
A ruminant mammary cell culture that accurately reproduces mammary function in vitro would be a valuable tool in studies of ruminant lactation, With this in mind, we have examined milk protein synthesis and secretion, milk protein mRNA abundance, and hormonal responsiveness in primary cultures of mammary acini from lecturing sheep. α- and β-casein protein synthesis, β-lactoglobulin synthesis, and α-casein, β-casein, and β-lactoglobulin secretion are maintained at high levels for 8 h in culture, but then decline to approximately 25% of maximal rates between 8 and 24 h in culture, whereas synthesis of other proteins remains unaltered. The relative abundance of α-S1-casein, β-lactoglobulin, and α-lactalbumin mRNAs similarly decline between 8 and 24 h in culture. Extracellular labeled α-casein is increased fourfold in the presence of fetal calf serum (FCS). In total, FCS alters the abundance of 47 of 68 secreted proteins detected by two-dimensional electrophoresis. However, FCS and lactogenic/galactopoietic hormones had no effect on the rate of decline of mammary function and did not promote any regaining of function when present for up to 9 days in culture. These results suggest that providing its limitations are recognized, this primary cell culture system may be useful in studying some aspects of ruminant mammary function in vitro. 相似文献
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The revitalization of mixed strain dried starter cultures at 22 and 32 C in sterile skim milk was materially accelerated when the substrate was fortified with 0.2% pancreas-extract solids. At 22 C, all cultures grew up satisfactorily in 18 hr, and in unfortified milk none of the cultures reached comparable growth in this period. When the cultures were grown at 32 C, the dried cultures developed adequately in 7.5 hr, but required 9 to 10 hr in plain milk. Culture growth was enhanced in milk containing pancreas extract to the extent that the amount of dried culture required to produce adequate acidity in normal incubation times could be markedly reduced. At 32 C, certain cultures could be reduced to 12.5% of recommended amounts, and at 22 C certain ones could be reduced by 50%. Revitalization of the dried cultures in milk containing pancreas extract did not affect the growth of subcultures in plain milk. Also, when dried cultures initiated growth in fortified milk at 32 C their subsequent growth at 22 C in milk alone was not affected. The faster rate of culture growth in milk containing pancreas extract should permit, with more certainty, the establishment of active mother and bulk starters. Furthermore, economy of dried cultures, as well as of time, could be realized by the use of fortified milk. 相似文献
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Ester Production by Pseudomonas fragi. II. Factors Influencing Ester Levels in Milk Cultures 总被引:6,自引:3,他引:3
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Cultures of Pseudomonas fragi were grown at 21 C in sterile homogenized milk and reconstituted skim milk media supplemented with ethyl alcohol. Quantitative determinations of ethyl butyrate and ethyl hexanoate by gas-liquid chromatography showed definite increases in the concentrations of the two esters produced in these media in comparison to media not supplemented with ethyl alcohol. Supplementation with butyric acid in addition to ethyl alcohol generally elevated the ethyl butyrate concentration and usually depressed the cell count slightly. Aeration of any of the media during growth tended to reduce the cell population slightly. A relationship between increase in cell number and increase in concentration of esters during the growth of the culture was observed. Media containing high concentrations of ethyl alcohol plus milk fat or low-molecular-weight fatty acids were conducive to the production of a fruity aroma by P. fragi. 相似文献
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myo-Inositol Transport in Mouse Astroglia-Rich Primary Cultures 总被引:1,自引:1,他引:0
Heinrich Wiesinger 《Journal of neurochemistry》1991,56(5):1698-1704
Uptake of radiolabeled myo-inositol was studied in astroglia-rich primary cultures derived from neonatal mouse brains. The uptake was saturable in the presence of Na+ with a Km of 25 microM and a Vmax of 60 pmol.min-1.(mg protein)-1, suggesting a high-affinity transport system for myo-inositol in astroglial cells. In addition, a Na(+)-independent, nonsaturable component was found. Carrier-mediated uptake was not inhibited by cytochalasin B (50 microM), but was reduced by depolarizing concentrations of K+ and, to different extents, in the presence of phloretin, ouabain, or amiloride (1 mM each). scyllo-Inositol, glucose, and galactose also reduced myo-inositol uptake; inhibition by the two hexoses was not reversed in the presence of 0.4 mM sorbinil. On the other hand, uptake of 2-deoxyglucose was not inhibited by high concentrations of myo-inositol. Preincubation of the cells with glucose-free or inositol-free medium stimulated uptake of myo-inositol and preincubation with 25 mM glucose in the presence of 0.4 mM sorbinil had no effect on the rate of uptake. The results suggest that myo-inositol is taken up into the astroglial cells by a transport mechanism that is distinct from that of glucose and probably is an active one. Sorbitol pathway activity does not interfere with myo-inositol uptake. 相似文献
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《Journal of receptor and signal transduction research》2013,33(1):91-111
AbstractThe recent demonstration in our laboratory of at least two specific estrogen binding sites in the rat uterus prompted us to investigate similar heterogeneity of binding sites in a trans-plantable ovarian dependent mouse mammary tumor (MXT-3590). Saturation analysis of cytoplasmic (protamine sulfate or hydroxylapatite exchange assay) or crude nuclear fractions (protamine sulfate precipitated nuclear exchange assay) revealed two binding components: type I which conforms to the classically described estrogen receptor and type II which has a lower affinity for estradiol but a greater capacity than type I sites. Exposure of cytosol to charcoal partially removes bound 3H-estradiol from type II sites but not from type I sites. Type II sites are specific for estrogens and do not translocate from the cytoplasmic to the nuclear compartment. Although Type II sites undergo dissociation on prelabeled sucrose density gradients, they are readily demonstrable by postlabeling sucrose density gradient fractions and hydroxylapatite adsorption. Since the presence of type II sites interferes with the measurement of the estrogen receptor (type I) which may also undergo dissociation on sucrose gradients, we recommended that the technique of postlabeling be used for the sucrose gradient analysis of type I and II sites. In addition, saturation assays should be performed over a wide range of 3H-es-tradiol concentrations (0.1–120 nM) for proper evaluation of both sites. These considerations may contribute to more accurate predictions about the response of breast cancers to endocrine therapies. 相似文献
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小鼠乳腺细胞凋亡及瘦素对凋亡的影响 总被引:1,自引:0,他引:1
目的系统的研究小鼠乳腺发育周期中乳腺细胞凋亡情况,并阐明瘦素对乳腺细胞凋亡的影响。方法以小鼠乳腺为实验材料,采用TUNEL法系统地研究小鼠乳腺在青春期、妊娠期、泌乳期和退化期的整个发育周期中的细胞凋亡情况,并通过培养基中添加瘦素的方法研究瘦素对乳腺细胞凋亡的影响。结果在青春期50~60d、妊娠期10~16d、退化期1~10d检测到较多的细胞凋亡,其中退化期细胞凋亡最为显著。添加瘦素培养的乳腺细胞凋亡信号明显增多。结论小鼠乳腺发育不同时期细胞凋亡同结构和功能发育之间相互联系。同时通过小鼠乳腺组织体外培养的方法,证明瘦素在退化期乳腺组织中可明显诱导组织凋亡。 相似文献
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Involvement of a Plasmid in Production of Ropiness (Mucoidness) in Milk Cultures by Streptococcus cremoris MS 总被引:6,自引:3,他引:3
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Curing and genetic transfer experiments showed that lactose-fermenting ability (Lac+) and the ability to produce mucoidness in milk cultures (Muc+) in Streptococcus cremoris MS were coded on plasmids. The Lac+ phenotype was associated with a 75.8-megadalton plasmid, pSRQ2201. The Muc+ phenotype was associated with a 18.5-megadalton plasmid, pSRQ2202. The Lac plasmid, pSRQ2201, was first conjugatively transferred from S. cremoris MS to Lac−S. lactis ML-3/2.2. Later, the Muc plasmid, pSRQ2202, was conjugatively transferred from Lac− Muc+S. cremoris MS04 to Lac+ nonmucoid S. lactis transconjugant ML-3/2.201. Subsequently, pSRQ2201 and pSRQ2202 were cotransferred from Lac+ Muc+S. lactis transconjugant ML-3/2.202 to Lac−, nonmucoid, malty S. lactis 4/4.2 and S. lactis subsp. diacetylactis SLA3.25. Transconjugants showing pSRQ2201 were Lac+; those containing pSRQ2202 were Muc+. With the transfer of pSRQ2202, the transconjugants S. lactis ML-3/2.202 and S. lactis subsp. diacetylactis SLA3.2501 not only acquired the Muc+ phenotype but also resistance to bacteriophages, which were lytic to the respective parent strains S. lactis ML-3/2.201 and S. lactis subsp. diacetylactis SLA3.25. 相似文献
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Hyoung H. Kim Shannon M. Grande John G. Monroe Susan R. Ross 《Journal of virology》2012,86(24):13232-13240
Many receptors in hematopoietic cells use a common signaling pathway that relies on a highly conserved immunoreceptor tyrosine-based activation motif (ITAM), which signals through Src family tyrosine kinases. ITAM-bearing proteins are also found in many oncogenic viruses, including the mouse mammary tumor virus (MMTV) envelope (Env). We previously showed that MMTV Env expression transformed normal mammary epithelial cells and that Src kinases were important mediators in this transformation. To study how ITAM signaling affects mammary cell transformation, we utilized mammary cell lines expressing two different ITAM-containing proteins, one encoding a MMTV provirus and the other a B cell receptor fusion protein. ITAM-expressing cells were resistant to both serum starvation- and chemotherapeutic drug-induced apoptosis, whereas cells transduced with these molecules bearing ITAM mutations were indistinguishable from untransduced cells in their sensitivity to these treatments. We also found that Src kinase was activated in the MMTV-expressing cells and that MMTV-induced apoptosis resistance was completely restored by the Src inhibitor PP2. In vivo, MMTV infection delayed involution-induced apoptosis in the mouse mammary gland. Our results show that MMTV suppresses apoptosis through ITAM-mediated Src tyrosine kinase signaling. These studies could lead to the development of effective treatment of nonhematopoietic cell cancers in which ITAM-mediated signaling plays a role. 相似文献
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Kavanaugh CJ Desai KV Calvo A Brown PH Couldrey C Lubet R Green JE 《Transgenic research》2002,11(6):617-633
Breast cancer is a leading cause of cancer morbidity and mortality. Given that the majority of human breast cancers appear to be due to non-genetic factors, identifying agents and mechanisms of prevention is key to lowering the incidence of cancer. Genetically engineered mouse models of mammary cancer have been important in elucidating molecular pathways and signaling events associated with the initiation, promotion, and the progression of cancer. Since several transgenic mammary models of human breast cancer progress through well-defined cancer stages, they are useful pre-clinical systems to test the efficacy of chemopreventive and chemotherapeutic agents. This review outlines several oncogenic pathways through which mammary cancer can be induced in transgenic models and describes several types of preventive and therapeutic agents that have been tested in transgenic models of mammary cancer. The effectiveness of farnesyl inhibitors, aromatase inhibitors, differentiating agents, polyamine inhibitors, anti-angiogenic inhibitors, and immunotherapeutic compounds including vaccines have been evaluated in reducing mammary cancer and tumor progression in transgenic models. 相似文献