Plasma Levels of Progesterone and Oestradiol-17 β in Reindeer (Rangifer Tarandus Tarandus) During Pregnancy

The plasma concentrations of progesterone and oestradiol-17p during pregnancy and the first 20 days after parturition were estimated in reindeer. The concentration of progesterone in the period 75–25 days prior to parturition was significantly higher than in the period 200– 75 days prior to parturition (P < 0.001). During the last 25 days before parturition the concentration decreased significantly. The concentration of oestradiol-17β was in most cases below 70 pg/ml until 50 days prior to parturition. During the last 25 days of pregnancy there was a significant increase in oestradiol-17β and the ratio progesterone/ oestradiol-17β was markedly lower than in the period 75–25 days before calving.Key words: reindeer, gestation, plasma, progesterone, oestradiol-17β     

The effect of a bacterial endotoxin or cloprostenol on the clinical status and hormonal levels in 80-100 days pregnant gilts

An experiment was conducted to examine the effect of a lipopoly-saccharide (LPS) of Salmonella typhimurium on the luteal function in 80 days pregnant gilts. Four animals were i.v. injected with 2 μg LPS/kg body weight and 3 animals were i.m. injected with 500 μg cloprostenol (CP). Gilts which maintained pregnancy after the initial injection were reinjected with GP around day 100. Clinical observations were made and plasma levels of 15-keto-13,14-dihydro-PGF₂α, progesterone, oestradiol-17β and oestrone sulphate were analysed by radioimmunoassay.The LPS induced a characteristic clinical endotoxemia. All LPS treated gilts maintained pregnancy until day 100 when 1 gilt aborted, 1 was emergency slaughtered and 2 reinjected. The comparative injections of CP induced abortion! within 48 h in 2 of 3 gilts at 80 days and in all reinjected animals at 100 days of pregnancy. Progesterone decreased immediately after both LPS and CP injections. In non-aborting gilts, the progesterone decrease had a transient character. The PGF_2α metabolite levels responded to LPS by a dramatic surge of approximately 4 h duration. All abortions were accompanied by a massive release of PGF_2α reaching peak levels during expulsion of the foetuses. Oestradiol-17ß and oestrone sulphate followed an ascendent pattern between days 80 and 100. Occasional transient decreases in oestradiol-17ß or increases in oestrone sulphate levels after LPS and CP injections were observed in several animals. Abortions were followed by a sharp decrease of both oestrogens. Post-abortum reproductive disorders occurred frequently. Endocrine changes associated with post-abortum ovarian activity were relevant to the clinical and morphological observations. The relationship between the stage of pregnancy in the pig and its endocrine response to abortifacient agents as well as some foetopathic effects of the endotoxin are discussed.     

Urinary oestrogen excretion in the goat during the oestrous cycle and after administration of pregnant mare serum gonadotrophin (PMSG)

Two goats were placed in cages constructed for urine collection. Samples were taken daily and stored in the frozen state until analysis. The urine was processed according to a method designed for determination of oestrone and oestradiol-17 α in bovine urine (Lunaas unpublished).     

The absorption of anabolic agents from pellets implanted at the base of the ear in sheep

Twenty wether sheep were allocated to seven groups and received implants near the base of one ear with pellets containing: for group 1, (OE) 20 mg oestradiol-17 beta alone; for group 2, (TBA/OE) 20 mg oestradiol-17 beta intimately mixed with 140 mg trenbolone acetate; for group 3, (T/OE) 20 mg oestradiol-17 beta mixed with 200 mg testosterone; for group 4, (P/OE) 20 mg oestradiol-17 beta mixed with 200 mg progesterone; for group 5, (TBA/OE2) 20 mg oestradiol-17 beta in one ear and 140 mg trenbolone in the other ear; for group 6, (TBA/OE1) 20 mg oestradiol-17 beta and 140 mg trenbolone as separate pellets in one ear; group 7 sheep received implants of carrier material and served as controls. The concentrations of steroids were measured in plasma samples collected from both jugular veins during the 16-week period after implantation. The absorption of oestradiol-17 beta was slower and more sustained from the pellets in which it was mixed with other steroids (groups 2, 3 and 4) than from the pellets containing oestradiol-17 beta alone (groups 1, 5 and 6). The concentration of each steroid in plasma was higher in the jugular vein ipsilateral to the implant than in the vein on the opposite side. The difference between the concentrations in the two veins was used to calculate the biological half-lives of the steroids; for oestradiol-17 beta and trenbolone the mean values ranged from 1.8 to 6.8 min and from 3 to 4 min, respectively, and for testosterone and progesterone the mean values were 4.7 and 3.5 min, respectively.     

The development of an ELISA-assay for semi-quantitative detection of dihydrogriesenin, a sesquiterpene lactone from Geigeria

Certain species of Geigeria contain sesquiterpene lactones which cause vomiting disease in sheep. Dihydrogriesenin (DHG), a sesquiterpene lactone from G. asperta, contains an alpha-methylene function which can spontaneously react with thiol groups on proteins to form a covalent adduct. A specific antiserum against a DHG-protein adduct can be used to determine the fate of DHG in poisoned animals. The preparation of such an antiserum is reported in this paper. DHG was reacted with cysteine and subsequently coupled to serum albumin using the carbodiimide reaction. When rabbits were immunized with one such conjugate (DHG-bovine serum albumin), it was found that the carrier determinants were immunodominant. A DHG-specific anti-serum of sufficient (ELISA) titre could, however, be obtained by alternating serum albumin carriers for DHG in booster immunizations. The ELISA antigen-antibody reaction could be inhibited by prior reaction of the antisera with cysteinyl-DHG in solution.     

Production of a highly group-specific monoclonal antibody against zearalenone and its application in an enzyme-linked immunosorbent assay

A monoclonal antibody (mAb) against zearalenone (ZEN) was produced using ZEN-carboxymethoxylamine and -BSA conjugates. Antibody produced by one clone showing a very high binding ability was selected and found to have a higher affinity for ZEN compared to a commerciall ZEN antibody. We developed two direct competitive ELISA systems using the selected antibody (ZEN-coated and anti-ZEN antibody-coated ELISA). Quantitative ranges for the anti-ZEN antibody-coated ELISA and ZEN-coated ELISA were from 25 to 750 ppb and from 12.5 to 100 ppb, respectively. The detection limit of both methods as measured with standard solutions was 10 ppb. The intra-plate and inter-well variation of both ELISAs were less than 10%. The IC₅₀ values for α-zearalenol, β-zearalenol, α-zearalanol, and β-zearalanol compared to ZEN were 108.1, 119.3, 114.1, and 130.3% for the ZEN-coated ELISA. These values were 100.7, 120.7, 121.6, and 151.6% for the anti-ZEN antibody-coated ELISA. According to the anti-ZEN antibody-coated ELISA, the average recovery rates of ZEN from spiked animal feed containing 150 to 600 ng/mL of ZEN ranged from 106.07 to 123.00% with 0.93 to 2.28% coefficients of variation. Our results demonstrate that the mAb developed in this study could be used to simultaneously screen for ZEN and its metabolites in feed.     

Clinical and endocrinological studies in primiparous post partum sows. Effects of lactation length and litter size

The object of this investigation was to determine the relationships between clinical findings and hormonal patterns in primiparous sows with different lactation length and litter size during lactation, weaning and to the first oestrus. Seven pairs of primiparous full sib sows were used to determine the effect of lactation length with normal litter size. One sow of each pair was assigned to nurse the piglets for 3 weeks (group A) while the other nusred for 5 weeks (group B). Another 8 primiparous sows (group C) were assigned to nurse 2–4 piglets during a 5-week lactation period. Oestrus detection was performed twice daily and laparoscopic examination every 2 weeks. If the sows did not come in oestrus within 3 weeks after weaning they were slaughtered. Peripheral plasma levels of progesterone, oestradiol-17β and LH were estimated by radioimmunoassays throughout the experimental period.     

Studies of electrophoresis on cellulose acetate membrane of serum proteins from normal horses, sheep and pigs

A method for the rapid electrophoresis on a cellulose acetate membrane of serum proteins from horses, sheep and pigs is discussed.The various main globulin fractions in the serum of these animals were experimentally identified.Normal values for the percentage composition of serum from normal horses, sheep and pigs were calculated.In the horse there was great individual variation in the shape of the β-fraction, assumed to be due to different transferrin types. The mean value for β-globulin of 19.5 % in the horse was higher than for the other two species.The albumin percentage was highest in the sheep and lowest in the pig, 48.5 % and 43.2 % respectively. The sheep had the highest γ-globulin percentage, 22.8 %, while the horse had the lowest with 19.0 %.Finally the values were compared with corresponding figures reported by other authors and the results discussed.     

Luteinizing hormone (LH) response to different doses of synthetic gonadotropin releasing hormone (GnRH) in prepubertal gilts

The object of this investigation was to study luteinizing hormone (LH) response to different doses of synthetic gonadotropin-releasing hormone (GnRH) in prepubertal gilts. Four crossbred prepubertal gilts, 128–134 days old and body weight 57–63 kg, were used in this study. Four doses, 0. 5, 25 and 125 μg, of GnRH were administered via a jugular vein catheter in a latin square design. Each treatment consisted of 3 injections at 90 min intervals. Frequent blood samples were taken during a period of 90 min before up to 90 min after treatment. Total LH responses were measured from post-treatment samples as the area under the curve above base level obtained from pre-treatment samples. A positive relationship between GnRH dose and LH release was obtained in all gilts, except for 1 treatment given to a gilt with high plasma level of oestradiol-17β on the day of treatment. This study has demonstrated the responsiveness of the pituitary gland by LH release to different doses of GnRH in 4.5-month-old prepubertal gilts.     

Immunization against gnRH in mature mares: antibody titres,ovarian function,hormonal levels and oestrous behaviour

The aim of the present study was to investigate the effect of active immunization against GnRH in mature Standardbred mares (three experimental and one control mare) on antibody titres, ovarian function, hormonal levels and oestrous behaviour. The mares were individually teased with a stallion once each day. During the first part of the experiment (period I: late April until November), blood was sampled every third day during the first 3 months, thereafter once per week. In the second part of the experiment (period II: December until August), sampling was carried out every second week. Progesterone, oestradiol-17beta and LH were analysed. Rectal gynaecological examination was made with the same intervals as the blood samplings and included palpation of the genital organs and ultrasonography. The experimental mares were immunized against GnRH with a GnRH-BSA conjugate. Equimune (Vetrepharm, Bracetown, Business Park, Clonee, Co. Meath, Republic of Ireland) was used as adjuvant. The mares were immunized on four occasions (20-30 day intervals) and GnRH antibody titre was determined. All immunized mares produced antibodies against GnRH but the maximum titres as well as the duration of a greater than 10% binding capacity varied between the mares (1 : 1600 to 1 : 50 000; 5 to 12 months, respectively). After the first injection, all mares showed one oestrus and ovulated at the regular time. In two of the mares, the immunization resulted in ovarian atrophy. Their hormone levels of progesterone, oestradiol- 17beta and progesterone decreased to basal levels and the cyclical hormone pattern was interrupted from approximately 30 days onwards. They continued to show oestrous signs but with irregular durations and intervals. The third mare showed ovarian suppression only for short periods and not in both ovaries at the same time; the hormone levels were basal for only about 20 days (days 50-70) and the mare ovulated on day 75 after start of immunization. The other mares ovulated after 13.5 and 15 months, respectively. It is concluded that the effect of immunization against GnRH in mature mares was individual concerning antibody titre response and the suppression of ovarian activity and hormone levels. Mares with totally inactive ovaries continued to show oestrous signs but with irregular intervals and durations.     

Peptococcus (S. Micrococcus) indolicus. The demonstration of two varieties of hemolysin forming strains.

On the basis of biochemical and serological criteria 2 hemolysin forming varieties of peptococci were identified as Peptococcus indolicus.Of a total of 16 hemolytic strains examined 9 originated from the vagina of clinically healthy cows, 4 from mastitis secretions from dry cows, 2 from the interdigital skin of clinically healthy sows, and 1 from a subcutaneous abscess in a pig.Two strains were designated a-hemolytic and 14 β-hemolytic.On blood agar plates colonies of the α-hemolytic variety were surrounded by narrow zones of almost complete hemolysis, while colonies of the β-hemolytic variety were surrounded by broad zones of incomplete hemolysis. The hemolysins were termed α- and β-hemolysin, respectively.The β-hemolysin, but not the α-hemolysin, could be demonstrated in cultures grown in liquid media. The β-hemolysin was found to be filtrable, relatively thermoresistant, and non-dermonecrotic.By gel diffusion analyses the 2 α-hemolytic strains were referred to Serotype C. Ten of the β-hemolytic strains belonged to Serotype C, 2 to Type B, 1 to Type D, and 1 to Type E.     

Passive immunization with antiserum to adrenocorticotropin increases weight gain in normal female rats.

Passive immunization against ACTH was used to test the hypothesis that growth in female rats is constrained by physiological concentrations of glucocorticoids. When animals were stressed by 15-min exposure to ether before blood sampling by cardiac puncture, serum concentrations of corticosterone were lower (P less than .05) in immunized rats than in stressed controls. The maximum effect was apparent 2 h after injection of ACTH antiserum, and no effect was apparent 6 h after injection. To examine the effects of ACTH immunization on growth, rats received daily injections of either saline, sheep immunoglobulin G, or ACTH antiserum, 2 h before the afternoon peak in plasma concentrations of corticosterone. After 7 d of treatment, rats treated with ACTH antiserum had gained 37% more body weight than saline-injected controls, and this effect was accompanied by a 59% reduction in peak plasma concentrations of corticosterone. Immunoglobulin G purified from normal sheep serum had no effect on weight gain. It is concluded that growth rate in normal female rats can be stimulated through the suppression of adrenal activity.     

Passive immunization of cyclic mares against androgen: gonadotropin and progesterone concentrations and estrous characteristics

Antiserum generated in a horse against testosterone conjugated to bovine serum albumin (BSA) was administered to six lighthorse mares (androgen-immunized mares) 1 to 3 d before a prostaglandin-induced estrus and twice again at 2-d intervals. Six control mares were administered antiserum generated against BSA on the same schedule. Relative to testosterone, cross-reactivities of other steroids with the testosterone antiserum were (%): dihydrotestosterone, 52; 5 alpha-androstane-3 alpha,17 beta-diol, 8.6; androst-4-ene-3,17-dione, 1.2; and all others tested less than .1. Tritiated testosterone binding in plasma increased (P less than .01) in androgen-immunized mares within 1 h and remained elevated (P less than .01) relative to controls for greater than 21 d. There was no effect (P greater than .10) of passive immunization against androgen on interval to estrus after prostaglandin injection, duration of estrus, ovarian volume, number of palpable follicles or follicular volume during estrus. In contrast, concentrations of luteinizing hormone (LH) were higher (P less than .05) in androgen-immunized mares than in control mares during estrus and early diestrus. Concentrations of follicle stimulating hormone (FSH) and progesterone at those times were not affected (P greater than .10). From these data, we conclude that androgens in the mare during estrus may be involved with the regulation of LH secretion. In contrast, no involvement with FSH secretion was apparent under these short-term conditions.     

Endocrine Changes after Mating in Pregnant and Non-Pregnant Llamas and Alpacas

Plasma concentrations of oestradiol-17ß, progesterone, 15-keto–dihydro–PGF_2α and luteinizing hormone (LH) were monitored in llamas and alpacas after mating with an intact male. Concentrations of LH and PGF_2α metabolite were high immediately after copulation. Ovulation occurred in 92% of the animals. The first significant increases in progesterone were recorded on day 4 after mating. In non-pregnant animals the lifespan of the corpus luteum was estimated to be 8–9 days. Luteolysis occurred in association with the release of PGF_2α. In pregnant animals, a transient decrease in progesterone concentrations was observed between days 8 and 18 in both species. No significant changes in PGF_2α secretion were registered during this period. Oes– tradiol–17ß concentrations were high on the day of mating, declined to low values on day 4, and started to increase again on day 8. Peak values after luteolysis in non-pregnant animals were significantly higher than those registered in pregnant ones. Furthermore, concentrations of oestradiol-17ß were elevated for a longer period in non–pregnant than in pregnant animals. The results suggest that progesterone from the corpus luteum exerts a negative influence on follicular activity in pregnant animals by reducing oes– tradiol-17ß secretion.     

Agarose gel electrophoretic fractionation of serum proteins in adult cattle. I. A study of clinically healthy cows.

Blood samples were taken from 25 clinically healthy Swedish red-and-white dairy cows on 11 occasions in the course of about one year. The blood serum proteins were studied, chiefly by agarose gel electrophoresis, in respect of relative and absolute concentrations. The following parameters were studied: Total protein, albumin, total globulin, A/G quotient, α₁-, α₂-, inter-α-β-, β₁-, β₂- and γ-globulin. The material was analysed statistically in respect of variation between and within cows, and of age, pregnancy and seasonal variation. The variation between cows was found to be significantly greater than that between different samplings of the same cow. Older cows exhibited significantly higher γ-globulin concentrations. The variation with season and stage of pregnancy was less pronounced for most serum protein fractions.     

抑制素α主动和被动免疫对哈萨克羊生殖激素含量的影响

为了研究抑制素α（INHα）主动和被动免疫对哈萨克羊生殖激素含量的影响，本试验在对抑制素α重组质粒表达菌株进行诱导表达的基础上，将经纯化、鉴定的抑制素α重组蛋白免疫接种新疆双峰骆驼，制备驼抗抑制素α多克隆抗体，并对其进行纯化，检测抗体效价，验证抗体的特异性。之后选择3~5岁、发情时间相近并处于间情期的45只成年哈萨克羊随机分为3组，分别作为抑制素α多克隆抗体免疫组（A组）、抑制素α重组蛋白免疫组（B组）及对照组（C组），每隔10 d连续进行3次加强免疫，应用ELISA法检测在绵羊繁殖活动中具有重要功能的5种生殖激素：促卵泡素（FSH）、促黄体素（LH）、孕酮素（P₄）、雌激素（E₂）、抑制素（INH），并检测血液生化指标。结果显示，IPTG的最佳诱导浓度是0.6 mmol/L，在4 h时诱导出产量较高的抑制素α包涵体蛋白，纯化后的抑制素α重组蛋白纯度较高，并具有较好的免疫原性，经进一步验证发现，制备的抑制素α抗体效价为1：512 000，该抗体可与抑制素α重组蛋白特异性结合。说明成功制备了具有免疫原性的抑制素α重组蛋白和高效价的驼抗抑制素α多克隆抗体。免疫后A组LH、P₄含量和B组FSH、LH、P₄、E₂、INH含量与C组相比差异不显著（P>0.05），而A组FSH含量和E₂含量显著高于C组（P<0.05），INH含量显著低于C组（P<0.05）。通过血液生化指标检测发现，抑制素α蛋白和抑制素α抗血清两种免疫制剂免疫后，试验动物均没有出现不良症状。说明两种抑制素α抗原均可对哈萨克羊血液生殖激素的分泌产生良好效果，相比之下抑制素α抗血清免疫效果更佳。     

Estrogen receptor independent neurotoxic mechanism of bisphenol A, an environmental estrogen

Bisphenol A (BPA), a ubiquitous environmental contaminant, has been shown to cause developmental toxicity and carcinogenic effects. BPA may have physiological activity through estrogen receptor (ER) -α and -β, which are expressed in the central nervous system. We previously found that exposure of BPA to immature mice resulted in behavioral alternation, suggesting that overexposure of BPA could be neurotoxic. In this study, we further investigated the molecular neurotoxic mechanisms of BPA. BPA increased vulnerability (decrease of cell viability and differentiation, and increase of apoptotic cell death) of undifferentiated PC12 cells and cortical neuronal cells isolated from gestation 18 day rat embryos in a concentration-dependent manner (more than 50 µM). The ER antagonists, ICI 182,780, and tamoxifen, did not block these effects. The cell vulnerability against BPA was not significantly different in the PC12 cells overexpressing ER-α and ER-β compared with PC12 cells expressing vector alone. In addition, there was no difference observed between BPA and 17-β estradiol, a well-known agonist of ER receptor in the induction of neurotoxic responses. Further study of the mechanism showed that BPA significantly activated extracellular signal-regulated kinase (ERK) but inhibited anti-apoptotic nuclear factor kappa B (NF-κB) activation. In addition, ERK-specific inhibitor, PD 98,059, reversed BPA-induced cell death and restored NF-κB activity. This study demonstrated that exposure to BPA can cause neuronal cell death which may eventually be related with behavioral alternation in vivo. However, this neurotoxic effect may not be directly mediated through an ER receptor, as an ERK/NF-κB pathway may be more closely involved in BPA-induced neuronal toxicity.     

不同处理方法对巴美肉羊同期发情效果的比较

旨在研究不同处理方法对巴美肉羊同期发情效果的影响。采用CIDR(孕酮阴道栓)+PMSG(孕马血清促性腺激素)+PGF2α(氯前列烯醇)和CIDR+PMSG2种不同处理方法对内蒙古巴彦淖尔市养殖场中本地绵羊(即巴美肉羊)进行同期发情处理,比较二者的发情率。结果表明,CIDR+PMSG+PGF2α处理组试验羊的发情率为74%,CIDR+PMSG处理组试验羊的发情率为94%。由该试验结果可以得出,采用CIDR+PMSG处理方法对巴美肉羊进行同期发情处理效果更佳。     

Detection and characterization of Clostridium perfringens in the feces of healthy and diarrheic dogs

Clostridium perfringens has been implicated as a cause of diarrhea in dogs. The objectives of this study were to compare 2 culture methods and to evaluate a multiplex polymerase chain reaction (PCR) assay to detect C. perfringens toxin genes alpha (α), beta (β ), beta 2 (β2), epsilon (ɛ), iota (ι), and C. perfringens enterotoxin (cpe) from canine isolates. Fecal samples were collected from clinically normal non-diarrheic (ND) dogs, (n = 105) and diarrheic dogs (DD, n = 54). Clostridium perfringens was isolated by directly inoculating stool onto 5% sheep blood agar (SBA) and enrichment in brain-heart infusion (BHI) broth, followed by inoculation onto SBA. Isolates were tested by multiplex PCR for the presence of α, β, β2, ɛ, ι, and cpe genes. C. perfringens was isolated from 84% of ND samples using direct culture and from 87.6% with enrichment (P = 0.79). In the DD group, corresponding isolation rates were 90.7% and 93.8% (P = 0.65). All isolates possessed the α toxin gene. Beta (β), β2, ɛ, ι, and cpe toxin genes were identified in 4.5%, 1.1%, 3.4%, 1.1%, and 14.8% of ND isolates, respectively. In the DD group, β and β2 were identified in 5%, ɛ and ι were not identified, and the cpe gene was identified in 16.9% of isolates. Enrichment with BHI broth did not significantly increase the yield of C. perfringens, but it did increase the time and cost of the procedure. C. perfringens toxin genes were present in equal proportions in both the ND and DD groups (P ≤ 0.15 to 0.6). Within the parameters of this study, culture of C. perfringens and PCR for toxin genes is of limited diagnostic usefulness due to its high prevalence in normal dogs and the lack of apparent difference in the distribution of toxin genes between normal and diarrheic dogs.     

The development of a sensitive enzyme immunoassay for the determination of estrone and estradiol-17beta in bovine blood plasma based on the same homologous combination with antiserum and steroid-enzyme conjugate

The development of a sensitive enzymeimmunoassay (EIA) for the determination of estrone (E1) and estradiol-17beta (E2 beta) in bovine plasma is described. The assay is a homologous double-antibody EIA with E2beta 17hemisuccinate (HS) as hapten for the immunoreactive reagent. The antiserum was raised against E2beta 17HS bovine serum albumin conjugate in the rabbit, and E2beta 17HS-horseradish peroxidase was used as steroid-enzyme conjugate. Each estrogen EIA was distinguished only by using the each working standard and sample for the EIA. Bovine plasma E1 and E2beta were extracted and purified before EIA. The antiserum was used at 1:1,750,000 dilutions for EIA. Estrone and E2beta showed high cross-reactivity with the antiserum (E1: 350.7%, E2beta:100%). The sensitivities were <0.03 pg/well for E1 and <0.12 pg/well for E2beta. Recovery rates of E1 and E2beta added to bovine blood plasma were 94.5% and 93.9%, respectively. The precision for EIA of estrogens was below 9.7%. The profiles of either estrogen as determined by EIA corresponded closely well with follicle dynamics in the cow during the estrous cycles and with placental function in pregnant animals. In conclusion, our new EIA can be applied with sufficient sensitivities, recovery and precision for the routine analysis of E1 and E2beta concentrations in bovine plasma.