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1.
Kapalavayi Nagaiah Franklyn F. Bolander Kevin R. Nicholas Toshiyuki Takemoto Yale J. Topper 《Biochemical and biophysical research communications》1981,98(2):380-387
The role of glucocorticoid in the prolactin-induced accumulation of casein mRNA in mammary explants from midpregnant mice has been studied after an initial 4-day incubation to allow the level of messenger to decline to undetectable levels. Subsequent culture for 3 days: 1) with insulin and glucocorticoid did not result in detectable accumulation of messenger; 2) with insulin and prolactin resulted in a very small accumulation; 3) with insulin, glucocorticoid and prolactin elicited a 20-fold greater accumulation of casein mRNA than the system with only insulin and prolactin. Therefore, although glucocorticoids are not an absolute requirement for casein gene expression in mouse mammary tissue, they are necessary for massive accumulation of casein mRNA induced by prolactin. It appears that this dependence is not a result of either mRNA stabilization or alteration in prolactin receptors. By contrast, stimulation of total epithelial RNA synthesis by prolactin does not have this glucocorticoid dependency. 相似文献
2.
Mammary explants from pregnant ewes were cultured in the presence of insulin, cortisol and prolactin, either alone, or in combination. After 2 d of culture, total RNA was extracted from explants and the content of beta-lactoglobulin mRNA was estimated using a specific labelled cDNA probe. The mRNA for beta-lactoglobulin was only deinduced slowly in the absence of hormone during the culture. Prolactin alone induced the accumulation of the mRNA. Insulin and cortisol added together were also stimulatory, but they only moderately amplified the prolactin effect. Beta-lactoglobulin gene in ewes, is therefore, controlled by the lactogenic hormones which also induce casein gene expression. The amplitude of the stimulation was unexpectedly low. This seems due in part to the fact that the gene was only deinduced weakly. In this respect, beta-lactoglobulin gene appears to be less dependent on lactogenic hormones under these experimental conditions than casein genes. 相似文献
3.
The differential actions of cortisol on the synthesis and turnover of alpha-lactalbumin and casein and on accumulation of their mRNA in mouse mammary gland in organ culture. 总被引:1,自引:0,他引:1
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Cortisol was previously shown to exert different, concentration-dependent, effects on the accumulation of casein and alpha-lactalbumin in mammary glands from mid-pregnant mice cultured in the presence of insulin and prolactin [Ono & Oka (1980) Cell 19, 473-480]. The present study demonstrated that the addition of 30nM-cortisol to the medium containing insulin and prolactin resulted in a marked enhancement of the rate of synthesis of both alpha-lactalbumin and casein in cultured tissue. The addition of 3 microM-cortisol in combination with insulin and prolactin caused a marked decrease in the rate of alpha-lactalbumin synthesis, but increased casein synthesis substantially. Similar changes were also observed in the amount of translatable mRNA for alpha-lactalbumin and casein in mammary explants cultured with insulin, prolactin and the two concentrations of cortisol. The study of the turnover of the milk proteins in cultured explants showed that virtually all of the casein synthesized remained intact in tissue explants cultured with 3 microM cortisol, whereas about 45% of casein disappeared in 40h from explants cultured with 30nM-cortisol. In contrast, the two concentrations of cortisol did not differentially affect the disappearance of alpha-lactalbumin, which was about 55% in 40h. These results indicate that the concentration-dependent differential actions of cortisol on the accumulation of alpha-lactalbumin and casein are exerted through its effects on the rate of synthesis and turnover of the two proteins as well as on the accumulation of their mRNA species. 相似文献
4.
Cortisol 21-mesylate, an alkylating derivatives of cortisol, was previously shown to exert an anti-glucocorticoid action in rat hepatoma cell culture (Simons, Thompson and Johnson 1980). In this study the effect of cortisol 21-mesylate on milk protein synthesis induced in cultured mouse mammary gland by glucocorticoid, insulin, and prolactin was investigated. Addition of cortisol 21-mesylate at concentrations ranging from 10(-8) M to 10(-6) M produced no inhibition of casein synthesis that was induced by glucocorticoid, insulin and prolactin in mammary explants from midpregnant mice. On the other hand, cortisol 21-mesylate in combination with insulin and prolactin stimulated casein synthesis in cultured tissue. The potency of cortisol mesylate was about 1/10 to 1/30th of that of cortisol. Cortisol 21-mesylate, like cortisol, also augmented the accumulation of alpha-lactalbumin in midpregnant rat mammary tissue cultured in the presence of insulin and prolactin. A cell-free competition study of glucocorticoid receptors using cytoplasmic extracts from mouse mammary tissue showed that cortisol 21-mesylate competitively inhibited the binding of dexamethasone on glucocorticoid receptors. The apparent affinity of cortisol 21-mesylate for glucocorticoid receptors is about 1/10th of that of cortisol. These results indicate that cortisol 21-mesylate acts as a glucocorticoid but not as an antiglucocorticoid in the mammary gland. 相似文献
5.
Role of prolactin and glucocorticoids in the expression of casein genes in rabbit mammary gland organ culture. Quantification of casein mRNA 总被引:11,自引:0,他引:11
Milk synthesis is initiated solely by prolactin in the pseudopregnant rabbit and glucocorticoids potentiate this action of prolactin. In organ culture, prolactin, in the presence or in the absence of insulin, enhances casein synthesis and cortisol (inactive alone) amplifies this action. Measurements of casein mRNA concentration in total cellular RNA, by hybridization with DNA complementary to casein mRNA, revealed that the stimulation of casein synthesis by the glucocorticoid is accompanied by an increase in the amount of casein mRNA. A systematic comparison of variations of these two parameters indicated that the major effect of glucocorticoids on lactogenesis in the rabbit at this stage of mammary gland development is mediated through an increase in the quantity of casein mRNA available for translation. No simultaneous control of casein mRNA translation by cortisol was observed. 相似文献
6.
When mammary gland explants from mid-pregnant rats were incubated with insulin (5 μg/ml) and [3H]cortisol (5 μg/ml) for one day, the tissue accumulated 1.69 μg cortisol/g wet tissue. During a second incubation with insulin and prolactin (5 μg/ml), only 20% of the steroid was lost per day. Such retention of glucocorticoid had an important biological consequence: the tissue exposed for one day to insulin and cortisol showed a transient stimulation of casein synthesis during a subsequent, five-day incubation with insulin and prolactin. No casein synthesis was detected, if the first culture medium contained only insulin. In conclusion, mammary gland explants from mid-pregnant rats require a glucocorticoid for casein synthesis, but this requirement may be obscured if the explants are initially incubated in medium containing cortisol, since they are capable of accumulating and retaining this steroid. Similar interpretative difficulties may arise in studies on other steroid-tissue relationships. 相似文献
7.
The effects of insulin, cortisol, prolactin, 3,3',5-triiodo-L-thyronine (L-T3) and progesterone on the synthesis of total protein and casein in mammary explants from pregnant goats were studied. In the absence of hormones and in the presence of insulin plus cortisol the rate of incorporation of 14C-leucine into proteins that were precipitated with the anti-casein antibody decreased during culture. The addition of prolactin to hormonal combination of insulin and cortisol caused large stimulation of rates of casein synthesis. Maximum incorporation of leucine was attained between 3 and 5 days of culture in the presence of 0.5 microgram ml-1 of prolactin. Prolactin stimulated-casein and total protein synthesis were not consistently affected by the addition of L-T3 or progesterone. The inhibition of DNA synthesis by hydroxyurea or cytosine-arabinofuranoside had no effect on casein synthesis in mammary explants from pregnant goats. 相似文献
8.
Glucocorticoids increase insulin binding and the amount of insulin-receptor mRNA in human cultured lymphocytes. 总被引:1,自引:0,他引:1
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Y Shibasaki H Sakura M Odawara M Shibuya Y Kanazawa Y Akanuma F Takaku M Kasuga 《The Biochemical journal》1988,249(3):715-719
The effect of steroid hormones on insulin binding and the amount of insulin-receptor mRNA was examined in IM-9 lymphocytes. Cortisol and cortexolone, but not oestrogen, increased both the binding of insulin and the amount of insulin-receptor mRNA in a time- and dose-dependent manner. Cortisol was most potent, and induced a 2-fold increase in insulin binding and a 4-fold increase in mRNA. The elevation in binding was due to an increased number of insulin receptors at the cell surface. The increase in mRNA involved all four of the insulin-receptor mRNAs and could not be inhibited by cycloheximide. The cortisol-induced increase in mRNA was associated with a 3-4-fold increase in the synthesis of pro-receptor. The relative potency of the three steroids indicated that these effects were mediated by an interaction with the glucocorticoid receptor. The results of this study suggest that cortisol can increase the number of insulin receptors at the cell surface by increasing the amounts of insulin-receptor mRNA and the synthesis de novo of insulin receptors. 相似文献
9.
Stabilization of casein mRNA by prolactin and glucocorticoids. 总被引:5,自引:0,他引:5
Prolactin injected into pseudopregnant rabbits led to a parallel enhancement of casein synthesis and casein mRNA concentration. When this stimulation was followed by a withdrawal of prolactin obtained by injections of bromocriptine, the rate of casein synthesis progressively diminished. In the presence of endogenous prolactin after the initial stimulation, the decline of casein synthesis was delayed. Hydrocortisone acetate injected with bromocriptine after the initial stimulation by prolactin was able to maintain a high rate of casein synthesis. Measurements of casein mRNA concentration by hybridization with casein cDNA indicated that in all cases the amount of casein mRNA was correlated with the magnitude of casein synthesis. This suggests that the lactogenic hormones, prolactin and glucocorticoids, which were previously demonstrated to be responsible for the enhancement of casein mRNA concentration are involved in their stabilization. 相似文献
10.
Prolactin induction of casein mRNA in organ culture. A model system for studying peptide hormone regulation of gene expression 总被引:17,自引:0,他引:17
The peptide hormone, prolactin, when added to organ explants of rat mammary gland, rapidly (within 1 h) induced the accumulation of casein mRNA. Casein mRNA sequences, as determined by hybridization with a specific cDNA probe, were shown to increase for up to 48 h after prolactin addition. The magnitude of this response was dependent upon the day of pregnancy at which the tissue was placed in culture. Maximal levels of induction (as great as 45-fold) were obtained using tissue from 15-day pregnant rats. Further data indicate that two steroid hormones, hydrocortisone and progesterone, were able to modulate the prolactin-induced accumulation of casein mRNA. The continuous presence of hydrocortisone was not necessary for prolactin induction of casein mRNA. However, the presence of hydrocortisone was required for maximal accumulation of casein mRNA. The induction of casein mRNA by prolactin was inhibited in a dose-dependent manner by the simultaneous addition of progesterone to the organ culture. Thus, hydrocortisone appears to potentiate the prolactin induction of casein mRNA, whereas progesterone is able to prevent casein mRNA accumulation. Since mammary gland organ culture is performed in a serum-free, chemically defined medium, this system allows a detailed examination of the mechanims by which a peptide hormone regulates the rapid accumulation of a specific mRNA. 相似文献
11.
12.
The study of differentiative potential of the lactating mouse mammary gland in organ culture 总被引:1,自引:0,他引:1
The organ culture of the mammary gland of lactating mice was used to examine the response of the differentiated gland to lactogenic stimuli, insulin, cortisol, and prolactin. Time course studies showed that casein synthesis in cultured tissue decreased rapidly during the first 2 d despite the presence of the three hormones, but on the 3rd d tissue cultured with either insulin and prolactin or all three hormones regained the ability to synthesize milk proteins, casein, and alpha-lactalbumin: a greater increase occurred in the three hormone system. The delayed addition of prolactin on Day 2 to the culture system containing insulin and cortisol also stimulated casein synthesis. The addition of cytarabine, which inhibited insulin-dependent cell proliferation in cultured explants, did not block the rebound of milk protein synthesis. These results indicate that in the presence of insulin, cortisol, and prolactin mammary epithelial cells in culture first lose and then regain the ability of synthesizing milk protein without requiring the formation of new daughter cells. 相似文献
13.
The interaction of cortisol and prostaglandins on the phenotypic expression of the alpha-lactalbumin gene in the mouse mammary gland in culture 总被引:1,自引:0,他引:1
N Terada L J Leiderman T Oka 《Biochemical and biophysical research communications》1983,111(3):1059-1065
Addition of cortisol at concentrations above 300 nM selectively inhibited the synthesis of alpha-lactalbumin and the accumulation of its mRNA in the mouse mammary gland cultured in the presence of insulin and prolactin, whereas the same treatment augmented casein synthesis and the accumulation of casein mRNA. Prostaglandin E2 or F2 alpha reversed the inhibitory effects of cortisol in a dose-dependent manner, without affecting casein production. The levels of prostaglandin E2 or F2 alpha in tissue explants cultured with insulin and prolactin increased about 2.6-fold over those in uncultured tissue, and the addition of cortisol decreased these levels approximately 2-fold. These results indicate the ability of prostaglandins to counteract the inhibitory effect of cortisol on the alpha-lactalbumin gene expression in the mouse mammary gland. 相似文献
14.
Glucocorticoid-dependent uteroglobin synthesis and uteroglobulin mRNA levels in rabbit lung explants cultured in vitro 总被引:3,自引:0,他引:3
In rabbit lung explants cultured in vitro in a synthetic medium, the synthesis of the protein uteroglobin decayed progressively becoming virtually undetectable between 24-48 h of culture. Addition of glucocorticoids to the medium maintained the synthesis of uteroglobin. This glucocorticoid effect was dose-dependent with optima at about 0.1 microM and 1 microM for dexamethasone and cortisol respectively. Estradiol, progesterone, triiodothyronine, insulin or 10% calf serum added to the medium were ineffective in maintaining uteroglobin synthesis. Actinomycin D (10 micrograms/ml) added to the medium inhibited the effect of cortisol on uteroglobin synthesis. After 24 h of culture, both the relative levels of uteroglobin mRNA, measured by molecular hybridization, and uteroglobin synthesis were correlatively higher (up to 10-fold) in glucocorticoid-treated than in control explants. 相似文献
15.
Summary The organ culture of the mammary gland of lactating mice was used to examine the response of the differentiated gland to lactogenic
stimuli, insulin, cortisol, and prolactin. Time course studies showed that casein synthesis in cultured tissue decreased rapidly
during the first 2 d despite the presence of the three hormones, but on the 3rd d tissue cultured with either insulin and
prolactin or all three hormones regained the ability to synthesize milk proteins, casein, and α-lactalbumin: a greater increase
occurred in the three hormone system. The delayed addition of prolactin on Day 2 to the culture system containing insulin
and cortisol also stimulated casein synthesis. The addition of cytarabine, which inhibited insulin-dependent cell proliferation
in cultured explants, did not block the rebound of milk protein synthesis. The results indicate that in the presence of insulin,
cortisol, and prolactin mammary epithelial cells in culture first lose and then regain the ability of synthesizing milk protein
without requiring the formation of new daughter cells. 相似文献
16.
Khalidah Al-Sarraj John Newbury David A. White R. John Mayer 《The Biochemical journal》1979,182(3):837-845
1. Explants of mammary gland from mid-pregnant rabbits were cultured in medium 199 containing insulin, prolactin and cortisol, and specific anti-casein immunoglobulin G was used to measure the amount, rate of synthesis and rate of degradation of casein in the explants in the presence of hormones and after removal of hormones from previously stimulated tissue. 2. The amount of casein in particle-free supernatants prepared from mammary explants was measured by ;rocket' immunoelectrophoresis. 3. The rate of incorporation of l-[4,5-(3)H]leucine into casein was measured after isolation of the casein by immunoadsorbent chromatography and polyacrylamide-gel electrophoresis in the presence of urea and sodium dodecyl sulphate. 4. Casein accumulates in mammary explants in the presence of insulin, prolactin and cortisol, but not in the absence of hormones. Removal of hormones after 24h in culture results in a decrease in the rate of accumulation of casein in the explants. 5. Casein-synthetic rate increases in mammary explants in the presence of insulin, prolactin and cortisol, but not in the absence of hormones. Removal of hormones after 24h in culture results in continued casein synthesis at approx. 30% of the rate in the presence of hormones. The synthetic rate does not decrease to values observed in explants cultured throughout in the absence of hormones. 6. Casein is not degraded in mammary explants during a phase of rapid casein accumulation (36-72h) in the presence of hormones. Furthermore casein is not degraded when hormones are removed from the tissue after between 36 and 72h in culture. 7. Casein is glycosylated in mammary explants; the extent of glycosylation parallels the rate of synthesis. The glycosylated protein is rapidly secreted from the tissue. 8. The results are consistent with the notion that after hormonal stimulation mammary explants from mid-pregnant rabbits synthesize, glycosylate and rapidly secrete casein. Removal of hormones decreases the synthetic rate of casein, but does not cause the accumulation of a pool of degradable casein in the lobuloalveolar cells. 相似文献
17.
The dose-response relationship between cortisol and the accumulation of the two milk proteins, casein and α-lactalbumin, was studied in organ culture of mammary gland from midpregnant mice. The accumulation of casein was low in culture with insulin but was enhanced by the further addition of prolactin. Further increases in casein were effected by the addition of cortisol in increasing concentrations up to 3 × 10?6 M, which was optimal for the accumulation of this protein. The content of α-lactalbumin in explants was similarly low in culture with insulin alone, but, in contrast, was increased to a maximal level by the addition of insulin and prolactin. The addition of cortisol up to 3 × 10?8 M with insulin and prolactin did not further increase the level of α-lactalbumin; in fact, at concentrations above 3 × 10?7 M the steroid caused progressive inhibition of the accumulation of this protein in cultured explants. Studies of the appearance of casein and α-lactalbumin in incubation medium during organ culture revealed the presence of substantial amounts of these milk proteins. During the first 2 days of culture with insulin, prolactin and 3 × 10?6 M cortisol, the amount of α-lactalbumin in culture medium was almost equal to the level found in tissue, whereas in the presence of 3 × 10?8 M cortisol, or in the absence of exogenous steroid, over 70% of total α-lactalbumin was retained in tissue. The observed difference in the amount of α-lactalbumin in culture medium can, however, only partially account for the inhibitory effect of high doses of cortisol on the accumulation of α-lactalbumin in cultured mammary explants. In contrast to α-lactalbumin, the relative amount of casein in culture medium containing insulin and prolactin was smaller—19% of total casein synthesized—and was further reduced to 16% and 11% of the total in the presence of 3 × 10?8 M and 3 × 10?6 M cortisol, respectively. The above results indicate that cortisol exerts dose-dependent differential actions on the accumulation of casein and α-lactalbumin in mouse mammary epithelium in vitro. 相似文献
18.
A number of growth factors can maintain hormonally-responsive epithelium in murine mammary explants as well as insulin, but only insulin can promote the synthesis of casein and alpha-lactalbumin, in the presence of glucocorticoid and prolactin. Anti-insulin receptor serum can elicit these effects of insulin on milk protein gene expression. The anti-serum is unique in its ability to mimic the developmental role of insulin in murine mammary epithelium. 相似文献
19.
20.
Hormonal regulation of the synthesis of casein and alpha-lactalbumin in a primary mammary cell culture system 总被引:1,自引:0,他引:1
The addition of 5 micrograms/ml of both insulin and prolactin, 3 microM cortisol and 5% fetal bovine serum stimulated casein synthesis during a 5 day culture of mammary epithelium from lactating mice using a floating collagen gel as a culture substratum. Omission of any of the three hormones or serum decreased casein synthesis substantially. The use of 10% serum or the attached gel culture system also decreased casein synthesis. Cells cultured with the combination of the three hormones and 5% serum contained a low level of casein mRNA on day 2, but it increased to much higher levels on day 4 and 5, amounting to over 30% of total mRNA on day 5. In contrast to casein synthesis, the maximal increase in alpha-lactalbumin synthesis required the presence of 0.03 microM cortisol. The combination of insulin, prolactin and 3 microM cortisol or insulin and prolactin elicited smaller increases. The translatable mRNA for alpha-lactalbumin in cells cultured with insulin, cortisol and prolactin for 5 days was detected, but not in cells with insulin and cortisol. Both a high and low concentration of cortisol in combination with insulin increased prolactin binding capacity of cultured cells to the same extent, whereas cells cultured with insulin alone contained much lower levels of prolactin binding. The difference in the capacity of prolactin binding between cells cultured with insulin alone and those cultured with insulin and cortisol correlated well with their ability to synthesize casein in response to prolactin. 相似文献