Isolation and partial characterization of prolactin from equine pituitary gland (hypophysis)

Highly purified equine prolactin was prepared from equine pituitary glands (hypophysis) by serial extractions with water at pH 5.5, 0.1 M (NH4)2SO4 at pH 4.0, and 0.25 M (NH4)2SO4 at pH 5.5 to remove other hormones, and then finally with 70% ethanol at pH 9.3 to 10.0 to extract prolactin. Preliminary purification of the extract involved salting out other substances with 0.1% NaCl at pH 9.0. Prolactin was precipitated out by adding three times the volume of 95% ethanol at 4 C. This prolactin preparation had a biological potency of 24 IU/mg. Further purification by isoelectric focusing on a pH gradient of 5 to 7 gave three prolactin components with the following characteristics: isoelectric point 5.8, 5.7, and 5.25; biological potencies (IU/mg) 35.6, 19.6, and 11.3. The major component had a molecular weight of 25,000, an isoelectric point of 5.8, and a biological potency of 35.6 IU/mg. Antiserum produced against this component did not cross-react with equine follicular stimulating hormone, luteinizing hormone, and growth hormone, but did cross-react with ovine and bovine prolactin. Human and murine prolactin had little cross-reactivity with the equine prolactin antiserum.     

Comparison of estrogen responsive genes in the mouse uterus, vagina and mammary gland

Female reproductive organs are mainly regulated by estrogen and progesterone. Specifically, the uterus, vagina and mammary gland show organ-specific mitosis and morphological changes during proliferative events, such as estrous cycle, gestation and lactation. The mechanism underlying these organ-specific estrogen-dependent events is still unknown. We examined, therefore, global gene expression in the mature uterus, vagina and mammary gland of ovariectomized adult mice 6 hr after an injection of 5 microg/kg 17beta-estradiol (E2) using a microarray method in order to identify primary E2-responsive genes. Half of the E2 up-regulated genes in the uterus were similar to those in the vagina. E2 up-regulated the expression of Insulin-like growth factor 1 (Igf-1) genes in the uterus and vagina. In the vagina, E2 up-regulated the expression of IGF binding proteins (Igfbp2 and Igfbp5). In the mammary gland, unlike the uterus and vagina, no gene showed altered expression 6 hr after the E2 exposure. These results suggest that expression of Igf-1 and morphogenesis genes is regulated by E2 in an organ-specific manner, and it is supported by the results of BrdU labeling showing E2-induced mitosis in the uterus and vagina except the mammary gland. The differences in organ specificity in response to E2 may be attributed by differences in gene expression regulated by E2 in female reproductive organs. The candidate estrogen-responsive genes in the uterus and vagina identified by profiling provide an important foundation understanding functional mechanisms of estrogen regulating morphogenesis and maintenance of each reproductive organ.     

Salsolinol is present in the bovine posterior pituitary gland and stimulates the release of prolactin both in vivo and in vitro in ruminants

The aims of the present study were to determine whether salsolinol (SAL), a dopamine-related compound, is present in the bovine posterior pituitary (PP) gland, and to clarify the effect of SAL on the secretion of prolactin (PRL) in ruminants. SAL was detected in extract of bovine PP gland using high-pressure liquid chromatography with electrochemical detection (HPLC-EC). A single intravenous (i.v.) injection of SAL (5 and 10mg/kg body weight) significantly and dose-dependently stimulated the release of PRL in goats (P<0.05). Plasma PRL levels reached a peak 10min after the injection, then gradually returned to basal values in 60-80min. The PRL-releasing pattern was similar to that in response to sulpiride (a dopamine receptor antagonist). The intracerebroventricular (i.c.v.) injection of 1mg of SAL had no significant effect on the release of PRL in calves, however, 5mg significantly stimulated the release (P<0.05) with peak values reached 30-40min after the injection. Moreover, SAL significantly stimulated the release of PRL from cultured bovine anterior pituitary cells at doses of 10(-6) and 10(-5)M, compared to control cells (P<0.05). Taken together, our data clearly show that SAL is present in extract of the PP gland of ruminants, and has PRL-releasing activity both in vivo and in vitro. Therefore, this endogenous compound is a strong candidate for the factor having PRL-releasing activity that has been previously detected in extract of the bovine PP gland.     

The effect of alkaloids and seed extracts of endophyte-infected tall fescue on prolactin secretion in an in vitro rat pituitary perfusion system.

The objective of this research was to measure the effects of endophyte-infected tall fescue seed extract and various alkaloids associated with the endophyte on in vitro prolactin secretion by rat hemipituitaries. Rat anterior pituitaries (AP) were dissected into halves and placed in temperature-controlled culture chambers (37 degrees C). The tissue was perfused with culture media at a flow rate of 12 mL/h. After perfusion for at least 90 min with control media, AP halves were exposed to their respective treatments for 15 min before they were returned to the control media. The treatments for Exp. 1 were .01 micrograms of alpha-ergocryptine/mL of culture medium, .01 microgram of ergonovine/mL of culture medium, .01 gram-equivalents of endophyte-infected tall fescue seed/mL of culture medium, and .01 gram-equivalents of endophyte free tall fescue seed/mL of culture medium. Treatments for Exp. 2 consisted of 10(-4), 10(-6), and 10(-8) M concentrations of perloline, N-formyl loline, N-acetyl loline, N-methyl loline, and alpha-ergocryptine. alpha-Ergocryptine suppressed (P less than .10) prolactin secretion in both experiments. Ergonovine and perloline both stimulated (P less than .10) prolactin secretion. The loline alkaloids (N-formyl loline, N-acetyl loline, N-methyl loline) had no effect on prolactin secretion. The endophyte-infected seed extract treatment suppressed (P less than .10) prolactin secretion. The endophyte-free seed extract treatment had no effect on prolactin secretion. In Exp. 2, prolactin secretion from AP responded to alpha-ergocryptine treatment in a dose-dependent fashion.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of phenothiazine on plasma prolactin levels in non-pregnant mares

Sixteen non-pregnant pony mares were divided into four groups of similar age and bodyweight (bwt). Groups were randomly assigned to one of four treatments consisting of oral administration of perphenazine (0.5 and 1.0 mg/kg bwt, phenothiazine (10 mg/kg bwt) and a control group. Blood samples were taken by jugular venepuncture and plasma prolactin concentrations measured using an homologous assay for equine prolactin. Analysis of variance was conducted on data designed as a split plot over time. Perphenazine given orally (0.5 and 1.0 mg/kg bwt) increased plasma prolactin concentrations when measured 3 and 6 h following feeding (P less than 0.05). Prolactin concentrations returned to normal by 11 h post drug administration. There was no response in plasma prolactin concentrations following oral phenothiazine treatment (10 mg/kg bwt). Perphenazine at the 1.0 mg/kg bwt level was discontinued after two days due to two mares exhibiting signs of hyperesthesia.     

Relationships among prolactin binding, prolactin concentrations in plasma and metabolic activity of the porcine mammary gland

The current studies were designed to investigate relationships among prolactin (PRL) binding, PRL concentrations in plasma and metabolic activity of porcine mammary glands. Preliminary studies revealed specific high-affinity binding of oPRL to porcine mammary gland. Conditions for optimal specific binding were similar to those observed for other species. To address the main objectives of the study, four mammary biopsies and blood samples were obtained from each of four gilts during lactogenesis and lactation (d-11, 4, 21 and 42 of lactation) to measure in vitro rates of metabolic activity, PRL binding to mammary membranes and PRL concentrations in plasma. Metabolic activity, as measured by oxidation of glucose or acetate to CO2 and incorporation into lipid, was low during pregnancy, increased two- to five-fold on d 4, and then paralleled the lactation curve for sows. There were highly significant positive correlations between PRL binding and all measures of mammary metabolism when data from pregnancy and lactation were utilized. Coefficients were positive but generally not statistically significant when lactation data only were utilized. During lactation, significant negative correlations were observed between concentrations of PRL in plasma and PRL binding and between PRL in plasma and mammary metabolic rate. These data provide evidence that binding of PRL to its receptor is an important effector of milk production in sows. Furthermore, oPRL is a suitable ligand to quantify PRL binding to porcine mammary tissue.     

The effect of xylazine on basal serum prolactin concentrations in male rats

In this study the effects of the major tranquilizer, xylazine, on serum prolactin level were studied in male rats. The results showed that, unlike all other tranquilizers, xylazine produced significant decrease (P less than 0.05) in serum prolactin levels. Xylazine at doses of 2.5, 5, and 10 mg/kg body weight (b.w.) induced significant decrease in serum prolactin levels 1 h after its intramuscular injection which remained significantly low for 4 h. The inhibition produced by 5 mg/kg and the 10 mg/kg b.w. doses was observed to be stronger in both magnitude and duration than that produced by the lower dose (2.5 mg/kg). This inhibitory effect of xylazine on prolactin levels may be attributable to its dopaminergic and/or alpha adrenergic antagonist activity. However, this point needs further investigation.     

母马生长激素细胞和催乳激素细胞的形态计测学研究

利用免疫组织化学和形态计测学的方法 ,观察了 1 8匹成熟雌性蒙古马的脑垂体前叶生长激素细胞和催乳激素细胞的数量和面积 ,同时利用放射免疫分析方法检测了这两种激素的血浆水平。结果表明 ,每个马脑垂体前叶中 ,生长激素细胞的平均数量为 6 .42× 1 8,每个细胞的平均面积为 82 .40μm2 ;催乳激素细胞的平均数量为 6 .0 7× 1 0 8,每个细胞的平均面积为 47.31μm2 。生长激素的血浆含量平均为 2 .84ng/ m L,但个体差异较大 ,变异系数高达 78.5 % ,催乳激素的血浆含量平均为 7.2 6 ng/ m L。本研究结果揭示 :母马脑垂体生长激素细胞和催乳激素细胞的数量和面积并不是决定母马这两种激素血中浓度的唯一重要因素 ;生长激素血中浓度上的个体差异 ,可能与其搏动性分泌形式有关     

催乳素对奶牛脂肪细胞脂代谢关键基因的作用

催乳素(prolactin,PRL)是启动和维持泌乳的重要激素,在泌乳期间能调节机体将营养物质和能量优先流向乳腺,用于乳脂和乳蛋白的合成。本试验采用体外分离培养奶牛前脂肪细胞,诱导分化为成熟脂肪细胞,加入PRL探讨其对脂肪细胞脂代谢相关基因的作用。结果显示,奶牛白色脂肪细胞存在催乳素受体(prolactin receptor,PRLR),且在PRL质量浓度为75μg/L,作用时间为9h时,PRLR表达量达到最高,此时调节脂合成的核转录因子SREBPE-1c及下游控制脂合成的基因FAS和LPL均显著降低,75μg/L组与对照组相比差异极显著(P0.01);控制脂分解的基因HSL显著升高,75μg/L组与对照组相比差异显著(P0.05);细胞培养上清液中的甘油含量显著升高,75μg/L组与对照组相比差异显著(P0.05)。结果表明,PRL能直接作用于奶牛白色脂肪细胞,降低脂合成作用并促进脂分解作用。     

Dynamic computed tomographic evaluation of the pituitary gland in healthy dogs

OBJECTIVE: To determine the contrast enhancement pattern of the pituitary gland in healthy dogs via dynamic computed tomography (CT). ANIMALS: 17 dogs. PROCEDURE: With each dog in sternal recumbency, transverse CT scans were made perpendicular to the skull base from the rostral clinoid processes to the dorsum sellae. At the position of the image that contained the largest cross section of the pituitary gland, a series of 9 to 11 scans was made during and after IV injection of contrast medium (dynamic CT scans). The contrast enhancement pattern of the pituitary gland and surrounding arteries was assessed visually and by use of time-density curves. RESULTS: After strong enhancement of the maxillary arteries, the intracavernous parts of the internal carotid arteries, and the communicating arteries of the arterial cerebral circle, there was a strong enhancement of the central part of the pituitary gland followed by enhancement of its peripheral part. On the last images of the dynamic series of the pituitary gland, the central part was hypodense, compared with the peripheral part. Time-density curves confirmed an early, strong enhancement of the central part and a delayed, less strong enhancement of the peripheral part of the gland. CONCLUSIONS AND CLINICAL RELEVANCE: The difference in enhancement between the central and peripheral parts of the pituitary gland was attributable to a difference in vascularization of the neurohypophysis and adenohypophysis, respectively. Distortion or disappearance of the strong central enhancement (pituitary flush) may be used for the detection and localization of pituitary abnormalities in the adenohypophysis.     

大鼠不同时期乳腺组织中雌激素孕激素及催乳素的变化规律

为探讨雌激素(E2)、孕激素(P)及催乳素(Prl)在大鼠不同时期乳腺组织中的变化规律,选雌性SD大鼠42只,分别为处女鼠、妊娠鼠(6d、12d、18 d)、泌乳鼠(6d、12d、18 d),每个时期6只.在指定的时间点处死大鼠取乳腺组织.用放免(RIA)组织匀浆中E2、P及Prl水平进行定量研究,用统计学方法进行处理,观察这三种激素的变化规律.结果显示E2从处女期到整个妊娠期,其呈现下降趋势.而分娩后E2水平急剧升高,后又有所下降,但维持在较高水平;在乳腺发育不同时期,P在乳腺组织中整体水平一直呈现下降趋势;Prl水平以处女期最高;随着妊娠进行逐渐降低,整个乳腺发育过程中Prl整体上几乎一直呈下降趋势.表明E2、P及Prl在促进乳腺发育及维持和启动泌乳方面发挥了重要的作用.     

Identification and characterisation of side population cells in the canine pituitary gland

To date, stem/progenitor cells have not been identified in the canine pituitary gland. Cells that efficiently exclude the vital dye Hoechst 33342 can be visualised and identified using fluorescence activated cell sorting (FACS) as a 'side population' (SP), distinct from the main population (MP). Such SPs have been identified in several tissues and display stem/progenitor cell characteristics. In this study, a small SP (1.3%, n=6) was detected in the anterior pituitary glands of healthy dogs. Quantitative PCR indicated significantly higher expression of CD34 and Thy1 in this SP, but no differences in the expression of CD133, Bmi-1, Axin2 or Shh. Pro-opiomelanocortin (POMC) and Lhx3 expression were significantly higher in the MP than in the SP, but no differences in the expression of Tpit, GH or PRL were found. The study demonstrated the existence of an SP of cells in the normal canine pituitary gland, encompassing cells with stem cell characteristics and without POMC expression.     

紫苏子油纳米乳诱导小鼠乳腺癌细胞EMT-6凋亡试验

采用滴定法制备紫苏子油纳米乳,用激光粒度测定仪分析其粒径,并进行其对体外培养的小鼠乳腺癌细胞EMT-6的凋亡诱导试验.结果显示,制备的紫苏子油纳米乳外观澄明,平均粒径为(50.3±1.1)nm.与紫苏子油软胶囊相比,质量浓度为100～800 mg/L时的紫苏子油纳米乳较紫苏子油软胶囊对EMT-6细胞的诱导凋亡作用均强,差异显著(P<0.05).透射电镜下,EMT-6细胞染色质边集,胞质内出现空泡,形成凋亡小体.800 mg/L剂量紫苏子油纳米乳处理EMT-6细胞48 h后,凝胶电泳可见明显的"阶梯状"DNA条带,为180～200 bp整数倍递增分布,表明紫苏子油纳米乳具有明显的诱导EMT-6细胞凋亡的作用.     

Inhibition of prolactin in the last trimester of gestation decreases mammary gland development in gilts

Prolactin is required for mammary development in various species but its possible role for mammogenesis in pigs is not known. The goal of the present study was therefore to determine the effect of prolactin inhibition by bromocriptine during the last third of gestation on mammary gland development in gilts. Twenty-eight primigravid gilts were assigned as controls (n = 15) or received 10 mg of bromocriptine orally thrice daily (n = 13) from d 70 to 110 of gestation. Jugular blood samples were collected on d 70 of gestation and every 8 d thereafter and were assayed for prolactin, IGF-I, estradiol, and progesterone. Gilts were slaughtered on d 110 of gestation and fetuses were counted and weighed. One row of mammary glands was used for dissection of parenchymal and extraparenchymal tissues and for determination of DNA, RNA, dry matter, protein, and fat contents. Tissue from the other row was used for measures of prolactin receptor number and affinity. Concentrations of prolactin were drastically reduced throughout the bromocriptine treatment period (P < .001), whereas there was no overall treatment effect on progesterone and IGF-I levels (P > .10). Total weight and extraparenchymal tissue weight of the mammary glands were unaffected by treatment (P > or = .1), but weight of parenchymal tissue, total DNA, and total RNA decreased (P < .01) with bromocriptine treatment. Percentages of fat and dry matter in parenchymal tissue increased with bromocriptine treatment (P < .01) and the percentage of protein decreased (P < .01). Number of prolactin receptors in parenchymal tissue decreased with bromocriptine treatment (P < .001) and receptor affinity increased (P < .001). Average fetal weight was lower in gilts receiving bromocriptine than in control gilts (P = .05), but fetal number did not differ (P > .1). These results clearly demonstrate that prolactin is essential for normal mammary gland development and can affect fetal growth during the last third of gestation in gilts.