Enzymatic O-glycosylation of kappa-caseinomacropeptide by ovine mammary Golgi membranes

The results of subcellular fractionation of sheep mammary gland membranes indicate that N-acetylgalactosaminyl polypeptide transferase and galactosyl-N-acetylgalactosaminyl transferase, which are involved in the assembly of disaccharide units of kappa-casein, are localized chiefly in Golgi membranes. The glycosyltransferase activities incorporating N-acetyl [1-14C] galactosamine and [U-14C] galactose from uridine diphosphate N-acetyl [1-14C] galactosamine and uridine diphosphate [U-14C] galactose, respectively, were measured after membrane solubilization with Triton X-100 either with unglycosylated caseinomacropeptide, or with this polypeptide containing the N-acetylgalactosamine side chain residues (desialylated and degalactosylated caseinomacropeptide). Radioactive N-acetylgalactosamine was incorporated in the unglycosylated acceptor peptide, and the glycosidic bonds in the product were alkali labile, suggesting that they were linked to the hydroxyamino acid residues. In addition radioactive N-acetylgalactosamine was released after alpha N-acetyl-D-galactosaminidase treatment of labelled caseinomacropeptide. [U-14C] galactose was incorporated in the desialylated and degalactosylated acceptor peptide. Reductive alkaline treatment of [U-14C] galactose peptide resulted in the release of a major product, the chromatographic properties of which in TLC were identical with authentic galactosyl (1 leads to 3) N-acetylgalactosaminitol. The structure of the labelled disacchariditol determined after periodate oxidation (two equivalents) by gas liquid chromatography-mass spectrometry revealed that the [U-14C] galactose was linked to position C-3 on the N-acetylgalactosaminyl-residue. The anomery of the galactose, as determined by a chemical method, indicates unambiguously a beta configuration.     

Co-expression of the proto-oncogene fos (c-fos) and an embryonic interferon (ovine trophoblastin) by sheep conceptuses during implantation.

Expression of the c-fos proto-oncogene by ovine conceptuses was analyzed by Northern and slot blots and indirect immunohistofluorescence in relation to the expression of the embryonic interferon-alpha (oTP) during implantation. c-fos was expressed initially in the trophoblast, and then in the allantois, when this tissue began to develop (day 17). In the embryonic tissues, the c-fos proto-oncogene was weakly expressed up to day 22 and increased thereafter. In the trophoblast, the expression of c-fos proto-oncogene was transient, occurring when the oTP gene was transcribed at a maximal level at the beginning of implantation (days 14-15), and decreased thereafter, following the pattern of oTP gene expression. This decline is due essentially to the arrest of c-fos and oTP gene expression by the trophoblastic cells which established cellular contacts with the uterine epithelium during the implantation process.     

In vitro fertilization with normal development in the sheep

Ovine tubal (n = 87) and ovarian in vitro matured oocytes (n = 99) were fertilized in vitro with ejaculated spermatozoa capacitated for 8 h in modified defined medium buffered with Hepes. High levels of fertilization were obtained as assessed by development to two-to six-cell stage within 40 h (75. 8% for ovulated and 62. 6% for in vitro matured oocytes). Electron microscope analysis of oocytes 20–22 h after insemination indicated that in vitro fertilization approximated the in vivo events. Embryos (two- to six-cell) were transferred surgically to the oviducts of pseudopregnant rabbits. Three days later, 42 (from ovulated oocytes) and 15 (from in vitro matured oocytes) embryos were recovered; 26 (61. 9%) and 10 (66. 6%), respectively, had cleaved at least once. Embryos incubated in vivo (n = 20 from ovulated oocytes; n = 9 from in vitro matured oocytes) were transferred surgically to the uteri of seven and four recipient ewes resulting in four and two pregnancies, respectively, from which three and one, respectively, have been maintained ( > 3 months). The first lamb resulting from the in vitro fertilization of an ovulated oocyte was born. In addition, six embryos (two- to four-cell) from tubal oocytes and ten embryos (two- to six-cell) from in vitro matured oocytes were directly transferred to the oviducts of two and three ewes, respectively. Two pregnancies resulting from in vitro matured fertilized oocytes are in progress ( > 3 months).     

Distribution of corticotropin-releasing factor in ovine brain determined by radioimmunoassay

We have developed and used a sensitive and specific radioimmunoassay to demonstrate the presence of CRF-like immunoreactivity in extra-hypothalamic areas of ovine brain. Synthetic CRF displaced antibody bound tracer at an ED50 value of 200 pg and there was no cross-reactivity with LHRH, TRH, ACTH, beta-endorphin and several other peptides. Displacement of bound 125I-CRF by brain extracts exhibited curves parallel to synthetic CRF standards. Highest concentrations (1 ng/mg tissue) of CRF-like immunoreactivity were found in the median eminence but surprisingly, high concentrations of CRF-like immunoreactivity were found in frontal, parietal, occipital and particularly temporal areas of cerebral cortex. Much lower concentrations were found in other brain areas including the basal ganglia, limbic system and brain stem.     

The seasonal occurrence of ticks (Acari: Ixodidae) on sheep and in the field in the Judean area of Israel

A 2 year survey of ixodid ticks in the Judean area of Israel between 1983 and 1985 showed that sheep were parasitized by the following species:Rhipicephalus sanguineus group (30.6%),Rhipicephalus bursa (25.3%),Haemaphysalis cretica (20.5%),Haemaphysalis otophila (20.1%) andHyalomma anatolicum excavatum (3.1%). Three other species,Hyalomma marginatum rufipes, Boophilus annulatus andAmblyomma lepidum were also present but in very low numbers. A total of 50 816 specimens (78.1% adult ticks) were collected from the sheep. In the field, a total of 4780 ticks (8.7% adults) were collected and 64.3% consisted ofR. bursa. Other species found wereH. cretica (14.5%),H. a. excavatum (13.7%),H. otophila (3.4%),R. sanguineus group (1.9%) and 1.7% wasIxodes eldaricus. The number of ticks collected varied from year to year and seasonal activity was observed in all species except forH. a. excavatum. In all species, males outnumbered females, with the exception ofH. cretica. The seasonal occurrence, tick ecology and disease relationship is discussed for each species.     

催乳素对培养的绵羊睾丸间质细胞睾酮分泌的影响

在绵羊睾丸间质细胞体外无血清长期培养的条件下,研究了催乳素对睾丸间质细胞睾酮分泌的调节作用。实验结果表明,催乳素可增强细胞对人绒毛膜促性腺激素(hCG)刺激的反应。催乳素的这种作用呈双相调节。睾酮分泌量显著高于hCG和催乳素单独作用时的总和。在hCG存在下,不同的底物转化为睾酮的量不同。其中雄烯二酮和孕酮转化为睾酮的方式存在着双相性。脱氢表雄酮转为睾酮的量少,不存在双相性,而与其剂量成正比。催乳素在hCG存在下可调节底物转化为睾酮。低剂量的催乳素(1ng/ml)可使一定剂量的孕酮(10～30ng/ml)转化为睾酮的量明显增加,而高剂量的催乳素(>10ng/ml)却明显地抑制孕酮转化为睾酮。催乳素可明显地抑制雄烯二酮转化为睾酮,与剂量无关。可见催乳素对于孕酮和雄烯二酮这两个关键底物转化为睾酮的调节是不同的。催乳素增强hCG刺激睾酮分泌的作用可能部分是通过其促进孕酮转化为睾酮来实现的。     

An optimal control problem for ovine brucellosis with culling

A mathematical model is used to study the dynamics of ovine brucellosis when transmitted directly from infected individual, through contact with a contaminated environment or vertically through mother to child. The model developed by Aïnseba et al. [A model for ovine brucellosis incorporating direct and indirect transmission, J. Biol. Dyn. 4 (2010), pp. 2–11. Available at http://www.math.u-bordeaux1.fr/～pmagal100p/papers/BBM-JBD09.pdf. Accessed 3 July 2012] was modified to include culling and then used to determine important parameters in the spread of human brucellosis using sensitivity analysis. An optimal control analysis was performed on the model to determine the best way to control such as a disease in the population. Three time-dependent controls to prevent exposure, cull the infected and reduce environmental transmission were used to set up to minimize infection at a minimum cost.     

Biochemical composition of ovine follicular fluid in relation to follicle size

The aim of this study was to biochemically characterize ovine follicular fluid and to relate possible changes in composition to follicular size. Ovaries were collected from adult and cycling non-pregnant slaughtered sheep (Ovis aries) during breeding season. A total of 104 pairs of ovaries were investigated and these data were then compared. Follicular fluid was aspirated from small (< 2 mm), medium (2-4 mm) and large (> 4 mm) nonatretic ovarian follicles. The follicular fluid was centrifuged at 4 degrees C and 5000 g for 30 min to remove any cells and stored at -80 degrees C prior to assay. Follicular fluid samples were analyzed for glucose, total protein, cholesterol, triglycerides, lactate, urea, creatinine, sodium, potassium, chloride, calcium, phosphorus, magnesium, acid phosphatase, alkaline phosphatase, and lactate dehydrogenase. Data were analyzed by the linear regression model. As follicles became larger, the concentrations of glucose and cholesterol significantly (P < 0.05) increased while those of triglycerides, lactate, alkaline phosphatase and lactate dehydrogenase significantly (P < 0.05) decreased.     

New in vivo and ex vivo models for the experimental study of sheep scrapie: development and perspectives

Sheep scrapie is a prototypical transmissible spongiform encephalopathy (TSE), and the most widespread of these diseases. Experimental study of TSE infectious agents from sheep and other species essentially depends on bioassays in rodents. Transmission of natural sheep scrapie to conventional mice commonly requires one or two years. In an effort to develop laboratory models in which investigations on the sheep TSE agent would be facilitated, we have established mice and cell lines that were genetically engineered to express ovine PrP protein and examined their susceptibility to the infection. A series of transgenic mice lines (tgOv) expressing the high susceptibility allele (VRQ) of the ovine PrP gene from different constructs was expanded. Following intracerebral inoculation with natural scrapie isolates, all animals developed typical TSE neurological signs and accumulated abnormal PrP in their brain. The survival time in the highest expressing tgOv lines ranged from 2 to 7 months, depending on the isolate. It was inversely related to the brain PrP content, and essentially unchanged on further passaging. Ovine PrP transgene expression thus enhanced scrapie disease transmission from sheep to mice. Such tgOv mice may bring new opportunities for analysing the natural variation of scrapie strains and measuring infectivity. As no relevant cell culture models for agents of naturally-occurring TSE exist, we have explored various strategies in order to obtain stable cell lines that would propagate the sheep agent ex vivo without prior adaptation to rodent. In one otherwise refractory rabbit epithelial cell line, a regulable expression of ovine PrP was achieved and found to enable an efficient replication of the scrapie agent in inoculated cultures. Cells derived from sheep embryos or from tgOv mice were also used in an attempt to establish permissive cell lines derived from the nervous system. Cells engineered to express PrP proteins of a specified sequence may thus represent a promising strategy to further explore, at the cellular level, various aspects of TSE diseases.