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K R Nicholas

Publications and source records attributed to K R Nicholas.

35 records · Page 2Linked to original sources
PubMed1987

A novel whey protein synthesized only in late lactation by the mammary gland from the tammar (Macropus eugenii).

A major whey protein which appears in milk from the tammar wallaby (Macropus eugenii) only during the second half of lactation (late lactation protein-A, LLP-A) was purified to apparent homogeneity by ion-exchange chromatography and gel filtration. An Mr of 21,600 +/- 2000 was calculated from its amino acid composition. A computer-based comparison of the sequence of the first 69 amino acid residues with the Atlas of Protein Sequence data base showed no significant homology with known proteins. Antiserum to LLP-A was prepared in rabbits, and single radial immunodiffusion was used to measure the amounts of LLP-A in milk during the first 40 weeks of lactation. LLP-A was first detected at 26 weeks; thereafter its concentration increased abruptly, to reach a maximum of 26 g/l at approx. 36 weeks of lactation. Explants prepared from mammary gland biopsies at 20 and 35 weeks of lactation were exposed to [3H]amino acids for 8 h; immunoprecipitation of tissue extracts showed that, whereas the rate of casein synthesis was the same at both stages of lactation, LLP-A was synthesized only by the 35-week mammary gland.

Amino Acid Sequence↗
PubMed1987

Pituitary-induced lactation in mammary gland explants from the pregnant tammar (Macropus eugenii): a negative role for cyclic AMP.

1. alpha-Lactalbumin and casein have been isolated from tammar milk. 2. alpha-Lactalbumin was induced in mammary explants by culture with anterior pituitary. 3. Casein was induced maximally in the presence of a physiological concentration of prolactin alone. 4. Progesterone did not inhibit the prolactin-induced synthesis of casein, alpha-lactalbumin, galactosyltransferase or fatty acids. 5. Both dibutyryl cAMP and a combination of cholera toxin and IBMX did significantly inhibit the induction of casein and alpha-lactalbumin. 6. Progesterone withdrawal is not a component of the lactogenic trigger in this marsupial but cAMP may be a common intracellular signal for negative control of lactogenesis in both marsupials and eutherians.

Animals↗
PubMed1985

Prolactin-dependent accumulation of alpha-lactalbumin in mammary gland explants from the pregnant tammar wallaby (Macropus eugenii).

The minimal hormonal requirements for the in-vitro accumulation of alpha-lactalbumin have been investigated in a marsupial, the tammar (Macropus eugenii). Mammary gland explants from 24-day pregnant tammars cultured in medium containing bovine insulin, cortisol and ovine prolactin showed a progressive increase in accumulation of alpha-lactalbumin during 4 days of incubation. No increment was observed if prolactin was omitted from the medium. However, a similar rate of increase was observed after 3 days of culture in medium containing prolactin alone. This induction of alpha-lactalbumin was maximal at a prolactin concentration of approximately 0.02 mg/l, which corresponds to physiological levels during pregnancy and early lactation. The absence of an effect of bovine insulin on tammar explants is not due to a general unresponsiveness to this hormone since insulin-stimulated DNA synthesis and amino acid uptake was evident after 3 days of culture. The inclusion of tri-iodothyronine and raised concentrations of cortisol in culture media have been shown to modulate alpha-lactalbumin synthesis in eutherian mammals but were without effect in the tammar. In addition, increased levels of progesterone did not inhibit the induction of alpha-lactalbumin, confirming an earlier in-vivo study suggesting that progesterone withdrawal may not be the lactogenic trigger in this species. Thus the pregnant tammar is the only species yet described in which alpha-lactalbumin is induced maximally in vitro in response to a single hormone.

Animals↗
PubMed1983

Essentiality of insulin and prolactin for accumulation of rat casein mRNAs.

Fetal calf serum (20%) does not substitute for insulin in terms of enhanced accumulation of rat casein mRNAs, but it can sustain the mammary tissue in culture. This property of fetal calf serum was utilized to show that in the presence of cortisol and 1) insulin or 2) prolactin, the delayed addition of prolactin or insulin, respectively, produced similar enhancement of 25 K and 42 K casein mRNAs. The results indicate that both insulin and prolactin are required for accumulation of rat casein mRNAs, the accumulation is independent of the sequence of addition of these hormones, and that the response cannot be ascribed entirely to the hormone added last.

Animals↗
PubMed1983

A unique and essential role for insulin in the phenotypic expression of rat mammary epithelial cells unrelated to its function in cell maintenance.

Mammary explants from pregnant rats can be induced in regard to casein synthesis and alpha-lactalbumin activity when cultured in the presence of hydrocortisone, prolactin and levels of insulin approaching physiological concentrations. No detectable induction occurs in the absence of insulin. Although epidermal growth factor and multiplication stimulating activity, in the presence of hydrocortisone, can maintain the initial level of NADH-cytochrome c reductase as well as insulin, neither can substitute effectively for insulin in the induction of the milk proteins. Proinsulin, nerve growth factor, platelet-derived growth factor and fibroblast growth factor are also ineffective substitutes for insulin in this regard. Whereas prolonged tissue exposure to multiplication stimulating activity, hydrocortisone and prolactin does not result in induction of alpha-lactalbumin activity, subsequent addition of insulin leads to prompt response. The results suggest that the ability of insulin to function as a unique, essential factor in the induction of rat milk proteins is independent of its cell-maintenance activity. Thus, in addition to its well established functions in metabolic processes, insulin appears to play a vital role in certain developmental processes.

Animals↗
PubMed1983

Anti-insulin receptor serum mimics the developmental role of insulin in mouse mammary explants.

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.

Animals↗
PubMed1983

Deoxyribonucleic acid synthesis-dependent casein gene expression: species differences.

Mammary gland explants from mature virgin mice, rats, and rabbits exhibit an increased rate of both DNA and casein syntheses when cultured in the presence of specific combinations of insulin, hydrocortisone, and PRL. If cytosine arabinoside, a potent inhibitor of DNA synthesis, is added to the culture medium, casein synthesis is inhibited in explants from mice but not in those from rats or rabbits. This inhibition is at the level of accumulation of casein mRNA; an 89% reduction of stimulated levels was observed. The nature of this block was investigated further by examining the general response of mature virgin mouse mammary explants to hydrocortisone and PRL, hormones considered essential for casein gene expression in this species. Cytosine arabinoside did not prevent an increment in either hydrocortisone-induced NADH-cytochrome c-reductase or PRL-induced total RNA synthesis. Previous work has shown that certain insulin-induced responses are also unaffected. Taken collectively, these results suggest that the lesion induced by cytosine arabinoside inhibition of DNA synthesis is distal to the receptor for one or more of these hormones. The necessity for coupling DNA synthesis with overt differentiation in the mature virgin mouse, but not in the rat or rabbit, is one of numerous examples of species variation in regard to the interaction of hormones with the mammary gland.

Animals↗
PubMed1981

Alpha-Lactalbumin and lactose concentrations in rat milk during lactation.

Homogeneous rat alpha-lactalbumin was prepared from whey by chromatography on DEAE-Sephadex A-50 and Ultrogel AcA 44. Two biologically active forms of alpha-lactalbumin were apparent after ion-exchange chromatography, but on gel filtration the combined forms were eluted as a single peak with a molecular weight of approx. 33000. The molecular weight when determined by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis was 15100. Antiserum to alpha-lactalbumin was prepared from rabbits, and single radial immunodiffusion was used to measure the concentration of alpha-lactalbumin in milk expressed from rats during lactation and for 2 days after the cessation of lactation. A significant positive correlation (r = + 0.89) between the concentrations of alpha-lactalbumin and lactose was obtained for the first 20 days of lactation. This is consistent with the suggestion that alpha-lactalbumin may control the concentration of lactose in milk. However, a significant negative correlation (r = -0.91) between the concentration of alpha-lactalbumin and lactose was obtained for 2 days after the cessation of lactation on day 20.

Animals↗
PubMed1981

Progesterone control of the initiation of lactose synthesis in the rat.

The in vitro incorporation of [14c]glucose into lactose in mammary tissue, the concentration of lactose in the mammary tissue and the concentration of lactose in the mammary secretion were determined during late pregnancy and lactation in the rat. These changes were related to the decline in blood progesterone during late pregnancy. The incorporation of [14C]glucose into lactose was detected on day 20 of pregnancy; it increased gradually until day 22 and then increased rapidly just prior to term (day 23 of pregnancy) to reach mean +/- s.e.m. maximum hourly values of 12.0 +/- 1.5 cpm x 10(4)/g by day 3 of lactation, and then declined to lower values (6.1 +/- 0.6 cpm x 10(4)/g) by day 20 of lactation. The concentration of lactose in both in the mammary tissue and in the mammary secretion increased rapidly over the last 24h of pregnancy and then more gradually after birth to reach mean +/- s.e.m. maximum values of 6.85 +/- 1.11 mg/g tissues and 43.1 +/-0 2.1 g/l respectively on day 15 of lactation. The decline in plasma progesterone to low levels between days 21 and 22 of pregnancy preceded the rapid increase in the concentration of lactose in the mammary tissue. A similar relationship was observed between the decline in progesterone and the increase in lactose in mammary tissue in rats Caesarean-sectioned on day 19 of pregnancy, and the administration of progesterone immediately following Caesarean section significantly depressed the accumulation of lactose in the mammary tissue. The results support the involvement of progesterone withdrawal in lactogenesis in the rat and indicate that [14C]glucose incorporation into lactose and the concentration of lactose both in the mammary tissue and in mammary secretion are useful indicators of the synthetic activity of the mammary gland.

Animals↗
PubMed1981

Progressive changes in plasma progesterone, prolactin and corticosteroid levels during late pregnancy and the initiation of lactose synthesis in the rat.

The relationship between progesterone, prolactin, corticosteroids and corticosteroid binding globulin (CBG) activity in plasma and the initiation of lactation were studied in normal parturient rats, and rats either ovariohysterectomized or Caesarean-sectioned on day 19 of gestation. In chronically cannulated rats the decline in plasma progesterone to low values (less than 10 micrograms/l) in normal parturient rats 20 h before term and in Caesarean-sectioned rats 10-14 h after surgery was closely related to an increase in plasma prolactin. However, in ovariohysterectomized rats the levels of progesterone declined abruptly to 20 micrograms/l within 30 min of surgery and prolactin remained low (5 micrograms/l) for 4-8 h and then gradually increased during the subsequent 16-18 h. Lactose concentrations in mammary tissue of rats killed at 0, 12, 18, 24, 36 and 48 h after surgery was low (less than 0.14 mg/g tissue) up to 12 h, and increased to reach maximum values at 36h and 48 h after surgery in ovariohysterectomized and Caesarean-sectioned rats respectively. The concentration of corticosteroids and CBG capacity in the plasma was 250-550 micrograms/l and 250-480 micrograms corticosterone bound per litre, respectively, in rats killed during the last 4 days of gestation. However, both the concentration of corticosteroids and the CBG capacity, within the individual cannulated rats, remained relatively constant during late gestation. These findings support the proposal that progesterone withdrawal is the lactogenic trigger and suggest the sequential involvement of prolactin. An increase in the concentration of free corticosteroids in late pregnancy and a related stimulatory role in the lactogenic mechanism were not established.

Adrenal Cortex Hormones↗
PubMed1981

The foetoplacental unit and the initiation of lactation in the rat.

Rats were either Caesarean-sectioned, ovariohysterectomized or ovariectomized on day 19 of gestation and the role of prolactin and corticosteroids in the initiation of lactation was studied by administering 2-bromo-alpha-ergocryptine (CB 154) and prolactin, and by using adrenalectomy and foetectomy. The concentrations of corticosteroids and prolactin in the plasma and the weight and lactose content of the inguinal mammary glands were determined 48 h after Caesarean section, and 24 h after ovariohysterectomy and ovariectomy. The ranges in concentrations of corticosteroids, prolactin and mammary lactose were 207-348 micrograms/l plasma, 21.9-65.3 micrograms/l plasma and 1.21-4.50 mg/g tissue, respectively, the Caesarean-sectioned, ovariohysterectomized and ovariectomized rats. The administration of CB 154 after either Caesarean section, ovariohysterectomy or ovariectomy decreased the concentration of prolactin to less than 8.8 micrograms/l. Whereas CB 154 significantly depressed the lactose content of the mammary tissue in Caesarean-sectioned and ovariohysterectomized rats (0.42 and 0.31 mg/g tissue, respectively), no effect was observed in ovariectomized rats (1.39 mg/g tissue). The administration of ovine prolactin reversed the inhibitory effects of CB 154. Furthermore, in two rats which were not ovariectomized and foetectomized, CB 154 did not inhibit lactose accumulation in the mammary tissue (2.02 mg/g tissue). Adrenalectomy of ovariohysterectomized rats decreased corticosteroid concentrations from 348 to 14 microgram/l plasma and the lactose content of mammary tissue from 1.31 to 0.19 mg/g tissue. Whereas adrenalectomy of ovariectomized rats decreased corticosteroid concentrations to 53 micrograms/l, the lactose content of mammary tissue (1.82 mg/g tissue) remained within the range for ovariohysterectomized rats. These finding show that hormones produced in late pregnancy by the foetus and placenta can support lactogenesis in the rat in the absence of maternal prolactin and corticosteroids.

Adrenal Cortex Hormones↗
PubMed1981

Insulin is essential for accumulation of casein mRNA in mouse mammary epithelial cells.

In the presence of cortisol and prolactin, insulin at concentrations as low as 1 ng/ml significantly stimulates casein synthesis in mammary explants from midpregnant mice; maximal synthesis occurs at 10 ng/ml. However, in the absence of insulin, no detectable immunoprecipitable casein is produced. Insulin also supports enhanced accumulation of casein mRNA in the presence of cortisol and prolactin; neither epidermal growth factor nor somatomedin C has this effect. These inductive actions of insulin are not secondary to a general maintenance effect on the mammary epithelial cell; insulin, epidermal growth factor, and somatomedin C can support the accumulation of RNA in rough endoplasmic reticulum equally well. In addition, these effects do not reflect a specific insulin requirement for prolactin sensitivity; epidermal growth factor can support prolactin-induced total RNA synthesis as well as insulin can. The results demonstrate that, although insulin, epidermal growth factor, and somatomedin C can all function as cell maintenance agents, only insulin, together with cortisol and prolactin, can induce casein mRNA accumulation.

Animals↗
PubMed1981

The induction of alpha-lactalbumin in rat mammary explants in the absence of exogenous prolactin: effects of progesterone and estrogen.

Mammary explants from both pregnant and virgin rats cultured in medium containing insulin, hydrocortisone and prolactin show a progressive increase in alpha-lactalbumin activity. However, when cultured with insulin and hydrocortisone only, explants from pregnant, but not those from virgin rats show an induction of alpha-lactalbumin-like activity similar to that seen when prolactin is present. The activity induced in the absence of exogenous prolactin corresponds to newly synthesized alpha-lactalbumin molecules, since 1) the activity is suppressed completely by rabbit anti-rat alpha-lactalbumin serum and 2) culture in the presence of [3H]-amino acids generates [3H]-alpha-lactalbumin, identified by SDS-polyacrylamide gel electrophoresis. Mammary tissue from virgin rats can be rendered responsive to insulin and hydrocortisone in this regard by administration in vivo of either progesterone or estrogen, but not by desoxycorticosterone acetate. This ability to convert virgin rat mammary cells to a state in which they are independent of exogenous prolactin in terms of alpha-lactalbumin synthesis represents a heretofore unreported effect of the female sex steroids.

Animals↗