The biological assay of chicken pituitary gonadotrophins

The properties of gonadotrophins prepared from chicken pituitary glands have been examined by assay methods specific for follicle‐stimulating hormone and luteinising hormone and by two assays which measure the combined effects of these hormones.

Valid parallel line assays were obtained when different preparations of chicken gonadotrophins were compared with standard preparations of human and ovine gonadotrophins. The augmentation and ovarian ascorbic acid depletion assays, which use mice and rats as test animals, were less sensitive to the chicken material than the method which depends on the uptake of ³²P by the testes of 1‐d‐old chicks. It is suggested that this might reflect a species specificity in response.     

Effects of ageing and gonadal steroid hormones on prolagtin concentration in the anterior pituitary of the chicken

1. Effects of ageing and gonadal steroid hormones on pituitary prolactin (PRL) concentration in the chicken were investigated.

2. No difference was found in the qualitative electrophoretic patterns of the anterior pituitary homogenates at different ages and between the sexes.

3. Pituitary PRL concentrations in males were generally higher than in females.

4. Pituitary PRL concentrations in castrated immature cockerels were higher than in intact birds, while PRL concentration was depressed by the injection of testosterone propionate or oestradiol benzoate.

5. Prolactin concentration in intact immature cockerels was decreased by daily injection of testosterone or oestradiol for 3 weeks but not by injection of the hormones for 1 week.     

The chicken pituitary-specific transcription factor PIT-1 is involved in the hypothalamic regulation of pituitary hormones

Pit-1 is a pituitary-specific POU-domain DNA binding factor, which binds to and trans-activates promoters of growth hormone- (GH), prolactin- (PRL) and thyroid stimulating hormone-beta- (TSHbeta) encoding genes. Thyrotropin-releasing hormone (TRH) is located in the hypothalamus and stimulates TSH, GH and PRL release from the pituitary gland. In the present study, we successfully used the cell aggregate culture system for chicken pituitary cells to study the effect of TRH administration on the ggPit-l* (chicken Pit-1), GH and TSHbeta mRNA expression in vitro. In pituitary cell aggregates of 11-day-old male broiler chicks the ggPit-l * mRNA expression was significantly increased following TRH administration, indicating that the stimulatory effects of TRH on several pituitary hormones are mediated via its effect on the ggPit-l* gene expression. Therefore, a semiquantitative RT-PCR method was used to detect possible changes in GH and TSHbeta mRNA levels. TRH affected both the GH and TSHbeta mRNA levels. The results of this in vitro study reveal that ggPit-1 * has a role in mediating the stimulatory effects of TRH on pituitary hormones like GH and TSHbeta in the chicken pituitary.     

Influence of catecholamines, prostaglandins and thyroid hormones on growth hormone secretion by chicken pituitary cells in vitro

In young chickens plasma concentrations of growth hormone (GH) are depressed by prostaglandins (PG) E1 and E2, epinephrine, norepinephrine, alpha 2 and beta agonists or thyroid hormones. A primary culture of chicken adenohypophyseal cells was used to examine the direct effects of these agents at the level of the pituitary as evaluated by GH release in the presence and absence of growth hormone releasing factor (GRF). Following collagenase dispersion and culture (preincubation, 48 hr) cells were exposed (incubation, 2 hr) to test agents, except for thyroid hormones which were added during the preincubation, and incubation period. Growth hormone release was increased (P less than .05) in the presence of PGE1 (10(-8)M by 34%; 10(-7)M by 54%), PGE2 (10(-8)M by 29%; 10(-7)M by 29%), PGF2 alpha (10(-8)M by 28%), and the beta agonist isoproterenol (10(-7)M by 46%). Basal GH release from chicken pituitary cells was not affected by dopamine, norepinephrine, epinephrine, thyroxine (T4), triiodothyronine (T3), or alpha adrenergic agonists. Growth hormone releasing factor stimulated GH release was not affected by the presence of prostaglandins E1, E2 or F2 alpha in the incubation media. However, GRF stimulated GH release was reduced by high doses of catecholamines: dopamine (10(-6)M by 34%), norepinephrine (10(-6)M by 74%), epinephrine (10(-8)M by 47%; 10(-7)M by 41%; 10(-6)M by 89%), and by the alpha 1 adrenergic agonist, phenylephrine (10(-7)M by 52%), the alpha 2 agonist, clonidine (10(-8)M by 34%; 10(-7)M by 83%) and the beta agonist, isoproterenol (10(-7)M by 64%).(ABSTRACT TRUNCATED AT 250 WORDS)     

Satellite cell proliferation in response to pituitary hormones

Proliferation of rat skeletal muscle satellite cells was studied in vitro, and their ability to respond to a variety of protein hormones was examined, including: growth hormone, prolactin, luteinizing hormone, thyrotropin and fibroblast growth factor. All experiments were conducted in serum-free medium to avoid complicating interactions with serum constituents such as other hormones or binding proteins. Dose-response curves were constructed for each protein and encompassed the physiological range plus concentrations two to three orders of magnitude greater than the physiological range. Of the proteins examined, the only one shown to have the ability to directly stimulate satellite cell proliferation was fibroblast growth factor. None of the anterior pituitary protein hormones had the ability to interact directly with satellite cells to stimulate proliferation in vitro. Therefore, satellite cells seem to be selective in their response to protein hormones, and the classes identified thus far are hormones in the insulin-like growth factor family and fibroblast growth factor. These two classes of protein hormones are quite different and would not be expected to act through a common pathway. Consequently, we have proposed a dual regulatory system that may allow for local as well as systemic stimulation of satellite cells.     

鸡α干扰素基因的原核表达及其活性测定

应用RT-PCR技术,从被刺激诱导的鸡脾脏淋巴细胞中扩增鸡α-干扰素(ChIFN-α)基因cDNA,经克隆和测序后,构建原核表达重组质粒pGEX-proChIFN-α(全长序列)和pET-ChIFN-α(不含信号肽序列),并分别转化相应的表达菌.重组菌经IPTG诱导,SDS-PAGE结果表明,ChIFN-α基因均获得表达,重组蛋白大小分别为45 ku、26ku左右,表达的蛋白以包涵体形式存在.表达产物经变性、提纯、复性,重组蛋白的含量约为1 mg/mL.复性后的ChIFN-α能够在鸡胚成纤维细胞上抑制H5N1禽流感病毒的复制;用水泡性口炎病毒检测结果表明,重组ChIFN-α具有较高的抗病毒作用,其生物学活性达到2×106 u/mg.     

Regulation of pituitary somatotroph differentiation by hormones of peripheral endocrine glands

Anterior pituitary somatotroph differentiation occurs during chick embryonic and rat fetal development. A number of findings support the hypothesis that differentiation of these growth hormone (GH) producing cells in the chick and the rat is regulated by adrenal glucocorticoids and thyroid hormones. Somatotroph differentiation can be induced in cultures of chick embryonic and rat fetal pituitary cells with adrenal glucocorticoids and this effect can be modulated by concomitant treatment with thyroid hormones. Plasma levels of thyroid hormones, corticosterone and adrenocorticotropic hormone increase during development, consistent with the ontogeny of somatotrophs. Treatment of chick embryos or rat fetuses with glucocorticoids in vivo induces premature somatotroph differentiation, indicating that the adrenal gland, and ultimately anterior pituitary corticotrophs, may function to regulate pituitary GH cell differentiation during development. Administration of thyroid hormones in vivo also increases somatotrophs prematurely, and administration of the thyroid hormone synthesis inhibitor methimazole inhibits somatotroph differentiation in vivo, suggesting that endogenous thyroid hormone synthesis contributes to normal somatotroph differentiation. Our working model for the regulation of somatotroph differentiation during normal development includes modulation by elements of the hypothalamo-pituitary-adrenal and hypothalamo-pituitary-thyroid axes. Additional research is reviewed defining the mechanism of action for these peripheral hormones in induction of pituitary GH gene expression during development.     

镉对鸡垂体线粒体结构和ATP酶的影响

镉是一种重要的环境污染物,动物镉中毒的主要原因是饲料中镉含量超标[1].进入机体中的镉将长期蓄积,对多器官和系统产生损伤.长期以来,人们对其致肝、肾、睾丸等器官及其线粒体损伤的研究较多,但对鸡垂体细胞线粒体损伤及垂体内酶活性的影响未见报道.     

Serum concentrations of pituitary and adrenal hormones in female pigs exposed to two photoperiods

Serum concentrations of pituitary and adrenal hormones were determined in lactating sows and ovariectomized (OVX) gilts exposed to 8 h (8L:16D) or 16 h of light (16L:8D). In addition serum prolactin (PRL) concentrations were determined after a thyrotropin releasing hormone (TRH) challenge. At 103 +/- 2 d of gestation or 3 wk after ovariectomy of nulliparous gilts on d 7 to 9 of the estrous cycle (d - 10), blood samples were collected from jugular vein cannulae at 30-min intervals for 8 h beginning at 0800 h. Immediately after the last sample, 13 sows and five OVX gilts were assigned to 8L:16D and 14 sows and five OVX gilts were assigned to 16L:8D/d and placed in two identical chambers in the farrowing house. Blood sampling was repeated on d 7, 14 and 21 of lactation in the sows and on d 7, 14, 21 and 28 in the OVX gilts. In Exp. 1, serum cortisol (C) concentrations were similar for sows exposed to 8L:16D (n = 7) and 16L:8D (n = 6) treatments, whereas in Exp. 2, serum C concentrations for sows exposed to 8L:16D (n = 6) were lower than those exposed to 16L:8D (n = 6) on d 7, 14 and 21. Photoperiod failed to influence serum concentrations of PRL, luteinizing hormone (LH) and growth hormone in the lactating sows or PRL in the OVX gilts. Photoperiod also failed to affect mean basal serum concentrations, peak height and peak frequency for PRL and LH in the lactating sows or for PRL in the OVX gilts.(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparison of the fractionation and assay of domestic duck and fowl pituitary gonadotrophins

1. Pituitary glycoproteins from domestic ducks and fowls were fractionated to separate luteinising hormone (LH) and follicle stimulating hormone (FSH) activities using the same chromatographic steps. 2. Fractions were bioassayed for LH using the release of progesterone from fowl granulosa cells and for thyroid stimulating hormone (TSH) by measuring the release of thyroxine in 3-d-old chicks. Follicle stimulating hormone activity was measured either in a cockerel-testes radioreceptor assay or by the release of oestrogen from cultured rat Sertoli cells. 3. Fractions containing predominantly FSH or LH activity were isolated from the fowl glycoproteins. Duck gonadotrophin did not occur in fractions corresponding to those containing fowl FSH. 4. Duck gonadotrophin was found in a fraction corresponding with the most highly purified fowl LH fraction. A duck LH fraction was found with little FSH activity for which there was no corresponding fowl LH fraction. 5. It is concluded that domestic fowl and duck gonadotrophins have different chromatographic properties. Further study is required to determine whether the purified duck gonadotrophin preparation comprises proteins with similar physico-chemical properties but with separate FSH and LH biological activities.     

甘氨酰谷氨酰胺对鸡生长、内源激素及相关基因表达的影响

9日龄优质粤黄鸡360只，随机分为4组，每组90只，通过饮水添加不同剂量的甘氨酰谷氨酰胺（Glycyl—Glu—tamine，Gly—Gln，剂量分别为0％、0．01％、0．05％和0．1％），观察Gly—Gln对粤黄鸡生长性能、内源激素和肝脏GHRmRNA、肌肉IGF—IR mRNA的影响。结果显示：一定剂量的Gly—Gln显著提高了试验组鸡的平均体重、胸肌率和腿肌率，显著降低了腹脂率；添加Gly—Gln的各试验组鸡血清中IGF-1、TSH、T3、T4水平均明显升高，而血清胰岛素水平降低；添加Gly—Gln的各试验组鸡肝脏GHRmRNA表达量略高于对照组，但差异不显著，而肌肉IGF—IRmRNA表达量低于对照组，其中0．01％和0．1％Gly—Gin组差异显著。以上结果提示，Gly—Gin可通过提高鸡血液IGF一1、TSH、T3、T4水平，降低血液胰岛素浓度而发挥其促生长和改善胴体品质的作用。     

饲料中鸡源性成分PCR-DHPLC检测方法的建立

以鸡组织为材料,提取鸡组织的线粒体DNA,以鸡12SrRNA基因为靶基因设计引物,PCR反应扩增片段230bp;用13种畜禽和水产动物线粒体DNA验证该PCR引物的特异性,结果只有鸡线粒体DNA为阳性;PCR-DHPLC检测灵敏度在DNA水平上达到了2.04mg/L;在随机采集的22份饲料样品中,检出8份鸡源性成分样品。结果表明,本研究建立的PCR-DHPLC方法可特异、灵敏地对饲料中的鸡源性成分进行快速准确检测。     

Gonadotrophin assay of cephalic and caudal lobes of anterior pituitary from growing turkeys

Pituitaries from male and female turkeys reared under natural daylight were collected every 2 weeks. Each pituitary was divided into the cephalic and caudal lobe and then homogenised and frozen until assayed. Gonadotrophic activity was determined by the P32 uptake of day‐old chick testes. Significant differences were found associated with the age of the birds, but not with either the sex or the lobe of the gland, nor any of the possible interactions. The peak activity was reached when the birds were 22 weeks of age, and a slight but non‐significant decrease took place until 34 weeks of age when the experiment was terminated.

A significant correlation (P≤0.01) between the amount of naturally occurring daylight and anterior pituitary gonadotrophin concentration of 0.86 was found. The interpretation of this correlation is discussed.     

酶联免疫法测定鸡肉中尼卡巴嗪残留

为了建立鸡肉中尼卡巴嗪残留标志物4,4’-二硝基均二苯脲（DNC）残留酶联免疫（ELISA）检测方法。将N-琥珀酰-L-丙氨酰-L-丙氨酰-L-丙氨酸4-硝基苯胺（SAN）与载体蛋白偶联作为抗原免疫动物,获得抗DNC抗体,在此基础上建立了鸡肉中尼卡巴嗪残留的检测方法。人工抗原中SAN与载体蛋白的结合比约为12：1,血清效价1：2000倍,鸡肉中检测限为9．2μg／kg,添加回收率在49．4％～118％之间,批内、批间变异系数均小于20％。该方法在灵敏度、精密度和准确度方面均能满足兽药残留筛选检测要求。     

A rapid and simplified technique for the assay of chicken hemolytic complement.

The technique described is a modification of a qualitative hemolytic radial diffusion technique. The test involves the use of sensitized sheep erythrocytes that have been incorporated into agarose. Tube dilutions were made of chicken serum and samples of each dilution were placed into wells cut in the agarose. The test is quantitative for hemolytic complement in that the highest dilution showing visible hemolysis of sensitized erythrocytes in agarose is determined to be the endpoint for that serum sample. The test as compared with the standard tube assay was determined to be less sensitive by approximately one dilution. The advantages of speed, simplicity, and cost more than offset the decrease in sensitivity of the test.     

鸡奇异变形杆菌外膜蛋白的抗血清制备及其荧光标记抗体检测方法的建立

为建立鸡奇异变形杆菌(P.mirabilis)直接荧光标记抗体检测方法,本研究提取P.mirabilis外膜蛋白(OMP),以OMP免疫健康新西兰家兔制备兔抗鸡P.mirabilis OMP高免血清,纯化的IgG,用FITC标记得到兔抗鸡P.mirabilis OMP-IgG的荧光标记抗体,建立直接荧光标记抗体检测方法。应用该方法检测临床样品184份,检测结果与细菌分离法和微量平板凝集比较,结果表明提取的OMP含有6种成分,分子量分别为67.0ku、63.1ku、57.5ku、53.7ku、43.0ku和31.6ku,经过优化的OMP-IgG荧光标记抗体最佳工作浓度为0.25mg/mL,最佳感作时间为30min。临床检测结果显示该方法具有简便、快速、敏感和特异性强等优点,为鸡P.mirabilis病的流行病学调查和临床快速诊断提供了技术保障。     

Effects of anaesthesia and manual restraint on the plasma concentrations of pituitary and adrenocortical hormones in ferrets

Two experiments were carried out to investigate the effect of sampling techniques on the plasma concentrations of pituitary and adrenocortical hormones in ferrets (Mustela putorius furo). In the first experiment blood was collected on two occasions from 29 ferrets which were either manually restrained or anaesthetised with isoflurane. In the second experiment eight intact ferrets were fitted with jugular catheters and blood was collected on four occasions, just before and as soon as possible after they had been manually restrained or anaesthetised with medetomidine or isoflurane; blood was also collected 10 and 30 minutes after the induction of anaesthesia. Medetomidine anaesthesia had no effect on the plasma concentrations of pituitary and adrenocortical hormones. Isoflurane anaesthesia resulted in a significant increase in the plasma concentration of alpha-melanocyte-stimulating hormone (alpha-MSH) directly after the induction of anaesthesia. Manual restraint resulted in a significant increase in the plasma concentrations of cortisol and adrenocorticotrophic hormone (ACTH) and a decrease in the plasma concentration of alpha-MSH.