The effect of hypophysectomy and subsequent replacement therapy with sheep prolactin or bovine growth hormone on the lactose synthetase activity of rabbit mammary gland.
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ABSTRACT:
SUBMITTER: Jones EA
PROVIDER: S-EPMC1174549 | biostudies-other | 1972 Dec
REPOSITORIES: biostudies-other
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Project description:1. The particulate form of lactating bovine mammary lactose synthetase activity is shown to be more highly organized than previously reported. 2. A novel method of shattering frozen mammary tissue with effective cell disruption is described. 3. The apparent subcellular distribution of lactose synthetase was shown to reflect the method of homogenization. 4. After mild homogenization particles associated with a high content of intact lactose synthetase activity sedimented in the lysosome size range between 5x10(4) and 3x10(5)g-min. 5. Lactose synthetase was dissociated and solubilized by VirTis homogenization and ultrasonic treatment. The activities and behaviour of UDP-galactose hydrolase, succinate dehydrogenase, beta-glucuronidase and phosphodiesterase I were compared. 6. Inhibition of UDP-galactose hydrolase by UTP and alpha-lactalbumin was observed.
| S-EPMC1186771 | biostudies-other
Project description:The augmentation of lactose synthetase activity during late pregnancy and lactation was measured by using both a tissue-culture assay and a cell-free assay. The results indicated at least a 100-fold augmentation in specific activity between late pregnancy and lactation. The cell-free assay indicated that the activities of both subunits of this enzyme had increased to 20-30% of the value during lactation by the last day of pregnancy. The tissue-culture assay, however, showed activities only 3-4% of the maximum at the time of parturition. This suggests that not all the enzyme present in the tissue before lactation commenced was active. Since at all stages of pregnancy and lactation the B subunit, alpha-lactalbumin (which is also a milk protein), was rate-limiting, it is suggested that the rate of lactose synthesis may be linked to the rate of milk-protein synthesis. Both subunits of lactose synthetase could be induced in tissue culture by the hormones insulin+hydrocortisone+prolactin. Of the three hormones, prolactin appeared to be the ;trigger' that induced the synthesis of these proteins if the tissue had been stimulated previously by insulin+hydrocortisone.
| S-EPMC1184648 | biostudies-other
Project description:1. The conditions that promoted the solubilization of particulate lactose synthetase were effective for solubilizing the thiamine pyrophosphatase of the Golgi apparatus but differed from those effective for beta-glucuronidase or acid phosphatase of lysosomes. 2. Lactose synthetase-containing particles did not bind Mg(2+) or Cs(+) ions, suggesting that they are not related to endoplasmic reticulum membranes. 3. Intact lactose synthetase and thiamine pyrophosphatase particles banded isopycnically at a density of 1.143 in a sucrose gradient. The dissociated ;A' sub-unit of lactose synthetase, UDP-galactose hydrolase, p-nitrophenyl phosphate acid phosphatase, alkaline phosphatase and phosphodiesterase I were associated with particles of a broad density range from 1.12 to 1.20. Lysosomal enzymes beta-glucuronidase, arylsulphatase and beta-glycerophosphate acid phosphatase were associated with particles of density 1.20, 1.175 and 1.15 respectively. 4. Rate-zonal sedimentation studies indicated that lactose synthetase particles have S(20,w) values exceeding 24000s, corresponding to spherical particles of diameter exceeding 5.4x10(-5)cm. 5. Electron micrographs of lactose synthetase particles purified over 20-fold revealed small spherical bodies (0.1-0.5mu) resembling lysosomes, the smaller of which were attached to membranes, and larger heterogeneous spherical or oval bodies (0.7-1.8mu) resembling lipofuscin secretory granules. 6. The relationship between lactose synthetase particles and the Golgi origin of secretion granules is discussed.
| S-EPMC1186772 | biostudies-other
Project description:1. Some of the kinetic properties of the particulate lactose synthetase of mouse mammary gland were investigated and shown to resemble those that have been reported for the soluble enzyme. Typical values for intact preparations were 2.3mum for the apparent K(m) for alpha-lactalbumin at 40mm-glucose and 1.7mm for the apparent K(m) for glucose at the endogenous concentration of alpha-lactalbumin. 2. Digitonin treatment increased total assayable activity approximately twofold but almost eliminated the endogenous activity found in the absence of alpha-lactalbumin, these findings being consistent with the location of endogenous activity within Golgi vesicles. 3. From the properties of the particulate fraction from lactating mice it was deduced that the effective endogenous alpha-lactalbumin concentrations were in the range 1-10mum. 4. The concentration of alpha-lactalbumin was not significantly different in particles isolated at various stages of pregnancy and early lactation. 5. The implications of these results for the control of lactose synthetase in vivo are discussed.
| S-EPMC1178352 | biostudies-other
Project description:PI 3-kinase enhancer A (PIKE-A) is critical for the activation of Akt signalling, and has an essential function in promoting cancer cell survival. However, its physiological functions are poorly understood. Here, we show that PIKE-A directly associates with both signal transducer and activator of transcription 5a (STAT5a) and prolactin (PRL) receptor, which is essential for PRL-provoked STAT5a activation and the subsequent gene transcription. Depletion of PIKE-A in HC11 epithelial cells diminished PRL-induced STAT5 activation and cyclin D1 expression, resulting in profoundly impaired cell proliferation in vitro. To confirm the function of PIKE-A in PRL signalling in vivo, we generated PIKE knockout (PIKE-/-) mice. PIKE-/- mice displayed a severe lactation defect that was characterized by enhanced apoptosis and impaired proliferation of mammary epithelial cells. At parturition, STAT5 activation and cyclin D1 expression were substantially reduced in the mammary epithelium of PIKE-/- mice. The defective mammary gland development in PIKE-/- mice was rescued by overexpression of a mammary-specific cyclin D1 transgene. These data establish a critical function for PIKE-A in mediating PRL functions.
| S-EPMC2837170 | biostudies-literature
Project description:Prolactin receptors present in the particulate fraction of lactating pig mammary gland were solubilized by 7.5mM-3-[(3-cholamidopropyl)dimethylammonio]-1-propane-su lph onic acid (Chaps) and purified by affinity chromatography on prolactin coupled to Affi-Gel 10. Nearly 30% of the particulate receptors were solubilized by the detergent and over a 1000-fold purification from homogenates was achieved. A water-soluble fraction rich in receptors was observed during the preparation of membranes, although this fraction has not yet been purified. Prolactin binding to the receptors was a time-dependent, reversible and saturable reaction in particulate, Chaps-solubilized and purified receptors. In all forms, receptors showed the same specificity to peptide hormones. Prolactin and human growth hormone bound to the same receptors, whereas bovine growth hormone, follicle-stimulating hormone, luteinizing hormone, thyroid-stimulating hormone and insulin failed to bind. After solubilization, the dissociation constant (Kd) for prolactin was decreased 5-fold from 9.8 X 10(-11) M in the particulate receptors to 1.8 X 10(-11) M in solubilized and purified receptors, being due principally to an increase in the association rate constant from 1.0 X 10(9)M-1 X h-1 to (3.9-4.6) X 10(9)M-1 X h-1, respectively, with the dissociation rate constant remaining unchanged at (1.1-1.3) X 10(-2)h-1. Isoelectric focusing of the prolactin-receptor complex revealed two peaks, one at a pI of 5.5-5.6 and the other at 5.2-5.3. Microsomal receptors were covalently cross-linked to 125I-labelled ovine prolactin with ethylene glycol bis(succinimidyl succinate) and analysed by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Autoradiography of the gel revealed a major subunit of Mr 28 000-35 000 and a minor one of Mr 67 000-69 000. Anti-(prolactin receptor) antibodies raised against rabbit mammary gland prolactin receptors were equally effective in inhibiting prolactin binding to particulate, solubilized and affinity-purified receptors, suggesting that purified prolactin receptors have a structure indistinguishable immunologically from particulate receptors and rabbit mammary gland prolactin receptors. The present demonstration shows that particulate prolactin receptors from a domestic animal can be solubilized and purified without losing the original properties of high affinity and binding specificity for hormones.
| S-EPMC1144528 | biostudies-other
Project description:RNA-Seq enables the generation of extensive transcriptome information providing the capability to characterize transcripts (including alternative isoforms and polymorphism), to quantify expression and to identify differential regulation in a single experiment. Our aim in this study was to take advantage of using RNA-Seq high-throughput technology to provide a comprehensive transcriptome profiling of the sheep lactating mammary gland. Eight ewes of two dairy sheep breeds with differences in milk production traits were used in this experiment (four Churra and four Assaf ewes). Milk samples from these animals were collected on days 10, 50, 120 and 150 after lambing to cover the various physiological stages of the mammary gland across the complete lactation. RNA samples were extracted from milk somatic cells. The RNA-Seq dataset was generated using an Illumina HiSeq 2000 sequencer. The information reported here will be useful to understand the biology of lactation in sheep, providing also an opportunity to characterize their different patterns on milk production aptitude.
| S-EPMC4932878 | biostudies-literature
Project description:During lactation, large amounts of calcium are exported from the mammary gland into milk to ensure skeletal growth of the offspring. Recent studies revealed that serotonin (5-HT) is essential to stimulate skeletal calcium resorption for milk synthesis. Our objective was to explore the correlation between circulating 5-HT and serum calcium and parathyroid hormone-related protein (PTHrP) concentrations around parturition in dairy goats. We also investigated the effect of 5-HT on PTHrP expression in cultured primary goat mammary epithelial cells (GMEC). Blood samples of multiparous Guanzhong dairy goats were collected on day -5 to 3 postpartum for analysis of serum concentrations of calcium, 5-HT, and PTHrP. Results revealed that from day -3 to 0 postpartum serum calcium and 5-HT concentrations decreased progressively, but serum PTHrP concentration only had a sharp drop in the postpartum period sampled. Correlation analysis of circulating 5-HT and serum calcium and PTHrP concentrations on day 1 and 2 postpartum revealed that low serum 5-HT concentration was positively correlated with serum total calcium or PTHrP concentration. By knocking down tryptophan hydroxylase-1 (TPH1) or adding 5-hydroxytryptophan (5-HTP) to decrease or increase the levels of 5-HT in GMEC, we observed that 5-HTP increased PTHrP expression in a dose-dependent manner and siTPH1 decreased PTHrP protein expression. Furthermore, 5-HT increased mRNA abundance of calcium-sensing receptor (CaSR) in a dose-dependent manner and decreased the expression of plasma membrane Ca2+ ATPase-1 (PMCA1). Taken together, 5-HT seems to induce PTHrP expression in goat mammary cells during and after parturition. These findings suggest that increasing 5-HT biosynthesis could be a potential therapeutic target for prevention of hypocalcemia in dairy goats.
| S-EPMC6093522 | biostudies-literature
Project description:Using microsomes prepared from rabbit mammary gland, the dissociation of prolactin (PRL) from its receptor was determined in the presence of peptide hormones or various concentrations of PRL. Among the hormones tested, PRL (ovine, mouse and bullfrog), human growth hormone and human placental lactogen each accelerated the dissociation of PRL in a manner proportional to their receptor-binding activities. Hormone-dependent dissociation was observed at higher concentrations than those at which the binding of PRL was completely inhibited by lactogenic hormones. In the concentration range 0.1 ng/ml-10 micrograms/ml, PRL increased the rate of dissociation in a logarithmic concentration-dependent manner. It was concluded that the dissociation of PRL from its receptor caused by lactogenic hormones is dependent on the hormone concentration. Arrhenius plot analysis revealed that PRL changed the frequency factor for the dissociation reaction. PRL in the medium inhibited the re-association of dissociated PRL. The data also suggested that PRL regulates the rate of dissociation by interacting with the PRL-receptor complex.
| S-EPMC1151627 | biostudies-other
Project description:BackgroundNeoplasms of the mammary gland are among the most common diseases in female domestic dogs (Canis familiaris). It is assumed that reproductive hormones influence tumorigenesis in this species, although the precise role of the endocrine milieu and reproductive state is subject to continuing discussion. In line with this, a recent systematic review of available data on the development of mammary neoplasms revealed weak evidence for risk reduction after neutering and an effect of age at neutering. Investigation of several hormone receptors has revealed decreased expression of estrogen receptor-alpha (ERα, ESR1), progesterone (P4) receptor (PGR), prolactin (PRL) receptor (PRLR) and growth hormone receptor (GHR) associated with neoplastic differentiation of mammary tissues. In other studies, increased levels of estrogens, progesterone and prolactin were found in serum and/or tissue homogenates of dogs with malignant neoplasms. However, the association between these entities within one animal population was never previously examined. Therefore, this study investigated the association between circulating serum concentrations of estradiol-17β, progesterone and prolactin, and gene expression of ERα (ESR1), ERβ (ESR2), PGR, PRLR, PRL and GHR, with respect to reproductive state (spayed vs. intact) and cycle stage (anestrus vs. diestrus). Additionally, the expression of E-cadherin (CDH-1) was evaluated as a possible indicator of metastatic potential.ResultsFor all receptors, the lowest gene expression was found in malignant tumors compared to normal tissues of affected dogs. Steroid levels were not influenced by their corresponding receptor expression in mammary neoplasms, but increased PRL levels were negatively associated with low PRLR gene expression in malignant tumors. The expression of CDH-1 was influenced by tumor malignancy and cycle stage, i.e., the highest gene expression was found in benign mammary tumors in diestrous dogs compared to normal and malignant mammary tissues of anestrous and spayed dogs.ConclusionsHerein, it has been confirmed that transformation towards malignant neoplasms is associated with significant reduction of gene expression of particular hormone receptors. Only PRLR in malignant tumors seems to be influenced by circulating PRL levels. In dogs, CDH-1 can be used as a prognostic factor; its expression, however, in benign tumors is influenced by cycle stage.
| S-EPMC4570623 | biostudies-literature