Ghrelin对雌性大鼠促性腺激素分泌及其mRNA表达的影响

【目的】探明Ghrelin在中枢水平上对雌性大鼠促性腺激素分泌及其mRNA表达的影响。【方法】用1 nmol Ghrelin对不同发情周期(发情前期、发情期、发情后期、间情期)雌性大鼠及用不同剂量(0.3,1和3 nmol)Gh-relin对发情后期雌性大鼠分别进行侧脑室注射,15 min后断头采血,离心血清,同时采集脑垂体,采用酶免法测定血清促黄体素(LH)、促卵泡素(FSH)的质量浓度,用实时荧光定量PCR法检测LH mRNA、FSH mRNA在垂体中的表达。【结果】①在发情前期和间情期,Ghrelin可显著抑制雌性大鼠LH的分泌和LH mRNA的表达(P0.05);Ghre-lin对整个发情周期中FSH的分泌不产生影响,仅在发情期抑制FSH mRNA的表达(P0.05)。②在发情后期,0.3和1 nmol的Ghrelin对大鼠促性腺激素分泌和mRNA表达均无影响,但3 nmol的Ghrelin可显著抑制LH、FSH的分泌和LH mRNA的表达(P0.05)。【结论】Ghrelin在中枢水平上对FSH、LH的释放及FSH mRNA和LH mR-NA的表达均有抑制作用,但存在一定的剂量差异,并受发情周期的调节。     

促黄体素对牛卵母细胞体外核成熟的影响

为了研究促黄体素(luteinizing hormone,LH)在牛卵母细胞成熟培养体系中的时间依从性,试验通过向牛卵母细胞体外成熟培养的不同时间段添加LH来观察其核成熟的情况。首先用无LH的基础成熟液I和添加LH的成熟液Ⅱ分别培养,在成熟的不同时间点(3、6、9h)用醋酸地衣红染色观察各自的生发泡破裂率。然后在体外成熟培养的不同时间段(0～3、0～6、0～9、0～24h)添加LH,24h后统计各自的第一极体率。结果表明,添加LH组在6h的生发泡破裂率显著低于对照1组(P0.05),24h添加LH组的第一极体率显著高于对照2组(P0.05)。因此,添加LH能延迟牛卵母细胞的核成熟,成熟培养中24h全程添加LH较不添加组能显著提高核成熟率。     

屡配不孕母牛发情开始后八天内血清促黄体素（LH）含量的变化

本实验采用放射免疫测定技术(RIA)分析了9头屡配不孕母牛和6头正常母牛于发情开始后8天内的血清促黄体素(LH)的含量变化。测定结果表明:屡配不孕母牛的LH排卵峰值低于正常母牛,其峰值分别为2.71±1.00ng/ml和4.70±1.71ng/ml,差异显著(P<0.025)。为了初步验证LH峰值与屡配不孕之间的关系,对20头屡配不孕母牛在发情开始至配种前肌肉注射外源性LH180IU,其配种后60天的受胎率为70％(14/20),而未用LH处理的11头屡配不孕母牛的受胎率为18％(2/11),两者差异极显著(P<0.01)。上述实验结果提示:体内LH排卵峰值降低可能是导致乳牛屡配不孕的重要原因之一。     

Role of ghrelin in regulating rabbit ovarian function and the response to LH and IGF-I

The aim of these in vivo and in vitro studies was to examine the role of ghrelin in the control of plasma hormone concentrations, the proliferation, apoptosis and secretory activity of ovarian granulosa cells and the response of these cells to hormonal treatments. Female rabbits were injected with ghrelin (10 μg/animal/day for one week before ovulation induced by 25 IU PMSG and 0.25 IU LHRH). On the day of ovulation, blood samples were collected and analyzed for concentrations of progesterone (P₄), testosterone (T), estradiol (E₂), estrone-sulphate (ES), insulin-like growth factor I (IGF-I) and leptin (L) by RIA. Some control and ghrelin-treated animals were killed in the periovulatory period, their ovaries were weighed and granulosa cells were isolated and cultured for 2 d. Cell proliferation (expression of PCNA) and apoptosis (expression of TdT) were evaluated by immunocytochemistry and TUNEL respectively. Secretion of P₄, T, E₂, IGF-I, and prostaglandin F (PGF) by granulosa cells cultured with and without LH or IGF-I (1, 10 or 100 ng/ml medium) was assessed by RIA. The remaining control and treated animals were kept until parturition, while the number, viability and body weight of pups were recorded.     

能量对初情期前母猪卵巢LHR和FSHR mRNA表达的影响

对9头50日龄、体质量为30 kg的初情期前杜×长×大三元杂交母猪进行长期日粮能量差异饲养试验.饲养结束后屠宰,取卵巢液氮保存.半定量RT-PCR检测每头母猪卵巢LH和FSH受体mRNA的含量.结果,高能组卵巢LH和FSH受体mRNA的含量显著高于中能组和低能组(P<0.05);而低能组显著低于中能组和高能组(P<0.05).表明,能量水平高的日粮可以显著地促进初情期前卵巢上LH和FSH受体在mRNA水平的表达,而能量水平不足的日粮则不利于这种表达.本文首次报道了日粮能量水平对母猪卵巢LH和FSH受体mRNA表达的影响.     

抑制素被动免疫对黄体中期山羊生殖激素分泌的影响

为了探讨抑制素在山羊生殖激素分泌调节中的作用 ,本试验用抗抑制素血清对黄体中期雌山羊进行了被动免疫试验。将酪 30 -抑制素 α(1,30 ) NH2 与兔血清白蛋白螯合 ,用去势雄山羊制备抗抑制素血清 (INH- AS) ,最终滴度为 1∶96 0 0 0。对照血清来自牛血清白蛋白免疫的去势雄山羊。 12只黄体期日本 SHIBA山羊 ,肌注 PGF2α(2 mg/只 )进行同期发情处理 ,处理后 3～ 5 d母羊出现发情。在发情后 10 d(发情当天为 0 d) ,将母羊分为 2组 ,每组 6只。试验组颈静脉注射 INH- AS(2 0 m L /只 ) ,并在注射前 (0 h)和注射后 6、12、2 4、4 8、72、96、12 0 h采取外周血样 (肝素抗凝 ) ;对照组注射对照血清 (2 0 m L/只 )。抗凝血样离心 (4℃、170× g、15 min)分离血清 ,- 2 0℃保存待测。血清 FSH和 L H含量用NIDDK放免试剂盒测定 ,其中 FSH标记纯品为 NIDDK- o FSH- I- 1,标准品为 NIDDK- o FSH- RO- 1,一抗为 NIDDK-抗- o FSH- 1;L H标记纯品为 NIDDK- o L H- I- 3,标准品为 NIDDK- o L H- RP- 2 4 ,一抗为抗 - o L H- YM。二抗为山羊抗家兔Ig G。血清 E2 和 P含量用1 2 5I放免盒测定。测 E2 时 ,为除去游离脂肪 ,样品用 0 .5 m L乙腈 +2 m L乙烷处理。试验结果表明 ,注射 INH- AS后 12 h,血清 FSH含量显著提高     

超排方法及兔龄对家兔超排效果的影响

采用国产促卵泡激素（FSH），促黄体激素（LH）和人绒毛膜促性腺激素（HCG），分别以FSH LH（A法），FSH HCG（B法）两种不同的处理方法，对不同年龄的家兔进行超排处理，旨在选择对家兔有效的超排方法，比较兔龄对超排效果的影响。结果表明，用A法对经产母兔和青年母兔处理后的排卵数分别为68枚，46枚；回收胚胎数分别为36枚，15枚；有效胚胎数分别为27枚，10枚，用B法对经产母兔和青年母兔处理后的排卵数分别为93枚，71枚；回收胚胎数分别为59枚，37枚，有效胚胎数分别为45枚。26枚。以上指标均以FSH HCG组最高，经产兔超排效果较青年兔好。     

番鸭就巢期生殖激素水平的变化规律

对番鸭就巢期不同时间的血浆生殖激素水平进行测试,揭示了不同就巢时间血浆生殖激素浓度变化的规律,并对就巢与激素之间的相关性进行分析。结果表明:(1)抱窝期血浆PRL含量(0.68±0.11ng/mL)极显著高于醒抱期的含量(0.33±0.04ng/mL)(P<0.01);(2)醒抱前后血浆LH含量差异很大,醒抱后LH含量(33.17±15.49mIU/mL)极显著高于抱窝期含量(5.56±1.99mIU/mL)(P<0.01);(3)抱窝期血浆E2含量(0.9±0.65ng/mL)极显著低于醒抱期含量(1.9±1.17ng/mL)(P<0.01);(4)PRL与LH、PRL与E2、LH与E2之间的相关系数分别为-0.923、-0.666和0.641。     

LRH─A3、hCG和PMSG对体外培养条件下牦牛垂体组织LH和FSH分泌的调节

促卵泡素３号（ＬＲＨ－Ａ３）、人绒毛膜促性腺激素（ｈＣＧ）和孕马血清促性腺激素（ＰＭＳＧ）等３种外源性激素均可增加体外培养条件下牦牛垂体组织分泌ＬＨ和ＦＳＨ的能力。培养液中ＬＨ和ＦＳＨ含量与加入的ＬＲＨ－Ａ３量呈正相关，与加入的ＰＭＳＧ和ｈＣＧ的量无显著关系     

Radioimmunoassay of Bovine,Ovine and Porcine Luteinizing Hormone with a Monoclonal Antibody and a Human Tracer

Forsberg, M., R. Tagle, A. Madej, J.R. Molina and M.-A. Carlsson. Radioimmunoassay of bovine, ovine and porcine luteinizing hormone with a monoclonal antibody and a human tracer. Acta vet. scand. 1993, 255-262.– A radioimmunoassay for bovine (bLH), ovine (oLH) and porcine (pLH) luteinizing hormone was developed using a human ¹²⁵ ILH tracer from a commercial kit and a monoclonal antibody (518B7) specific for LH but with low species specificity. Standard curves demonstrated similar binding kinetics when bLH, oLH and pLH were incubated with tracer and antibody for 2 h at room temperature. A 30-min delay in the addition of the tracer gave sufficient sensitivity when analysing pLH. Separation of antibody-bound LH from free hormone was achieved by using second antibody-coated micro Sepharose beads. The assay was validated and the performance compared with that of an RIA currently in use for determination of bLH and oLH (coefficient of correlation: 0.99 and 0.98). Regardless of the standards used, intra-assay coefficients of variation were <10% for LH concentrations exceeding 1 µg/L. The inter-assay coefficients of variation were <15%. The assay was used for clinical evaluation demonstrating the pre-ovulatory LH surge in two cyclic cows, LH pulsatility in an oophorectomized ewe and LH response to GnRH injection in a boar.