Cloning of cDNAs encoding the three pituitary glycoprotein hormone beta subunit precursor molecules in the Japanese toad,Bufo japonicus

Complementary DNAs encoding precursor molecules of the beta subunits of three pituitary glycoprotein hormones (LH, FSH, and TSH) of the Japanese toad (Bufo japonicus) were isolated and sequenced. Unexpectedly large numbers of single nucleotide substitutions were found in all three beta subunit cDNAs. The eight isolated LH beta precursor cDNA clones were classified into six forms of nucleotide sequence, with four nucleotide substitutions each in the apoprotein coding region and in the 3' untranslated region (UTR). In the deduced amino acid sequence, the LH beta subunit showed two forms with a single amino acid substitution. The seven isolated FSH beta subunit cDNAs were classified into two forms, which differed from each other at 11 positions in the 3' UTR. The six isolated TSH beta subunit clones were classified into four forms with 2 and 5 nucleotide substitutions in the signal peptide and apoprotein coding regions, respectively. However, all the substitutions in the apoprotein coding region were silent. The substitution in the signal peptide coding region could produce three forms of signal peptide. Amino acid sequence comparison revealed that the toad LH beta subunit is more similar to the fish GTH II beta subunit than to mammalian and avian LH beta subunits. We found that the toad LH beta subunit molecule is a partial chimera of LH and FSH; amino acid residues located in 36th to 42nd and 96th to 99th are identical or similar to those of not LH- but FSH-beta subunit in mammalian, whereas it is more similar to LH- than FSH-beta subunit in total. We also found that the toad FSH beta subunit is more similar to the fish GTH II beta subunit than to the fish GTH I beta subunit and that the toad TSH beta subunit is more similar to tetrapod TSH beta subunits than to fish TSH beta subunits.     

Cloning of a protein binding to the most proximal Pit-1 binding element of prolactin gene from human pituitary cDNA library

Activin is produced in mammalian ovarian follicles and is known to function as a paracrine as well as autocrine factor for folliculogenesis and oogenesis. We investigated the functional mechanism of activin using a hormone-supplemented serum-free culture system of granulosa cells isolated from diethylstilbestrol (DES)-primed 21-day-old rats. Recombinant human-activin A appeared to induce CycD2 and to act synergistically with FSH to promote G1/S transition and cell proliferation starting from 12 h after stimulation, accompanied by an increase of the hyperphosphorylated retinoblastoma protein (ppRb). Cells from unprimed rats gave similar results. FSH, in contrast, showed no CycD2-inducing activity, but turned out to modulate CycD2/cdk4 complex formation and enhance ppRb formation in conjunction with activin. These findings showed that the induction of CycD2 by activin and the synergistic effect of activin with FSH on ppRb formation play important roles in promoting G1/S transition in rat primary granulosa cells.     

Comparison of day 3 FSH serum values as determined by six different immunoassays

BACKGROUND: The accurate assessment of FSH concentration is important for evaluating ovarian function prior to IVF. However, a number of different assay techniques are currently in use, leading to inconsistencies in the hormone data being reported. To address this problem, we measured FSH concentration using a number of commercially available systems. METHODS: Day 3 serum FSH levels were measured in 215 healthy fertile women using six different immunoassays: Coatria (125)I (Bio-Mérieux), ACS-180 (Bayer Diagnostics), Advia-Centaur (Bayer Diagnostics), Vitros ECi (Ortho-Clinical Diagnostics), Architect i2000 (Abbott) and Elecsys 2010 (Roche Diagnostics). RESULTS: According to the immunoassay, means +/- SD of FSH concentrations were: 6.5 +/- 2.2 mIU/ml for Coatria (125)I, 6.8 +/- 2.7 mIU/ml for Advia-Centaur, 6.7 +/- 3.0 mIU/ml for Vitros ECi, 7.6 +/- 3.0 mIU/ml for ACS-180, 8.2 +/- 3.3 mIU/ml for Architect i2000 and 8.8 +/- 3.0 mIU/ml for Elecsys 2010. CONCLUSION: Day 3 FSH values determined by six different immunoassays were significantly different (P < 0.01, paired t-test). Physicians must take care when interpreting results from different clinical laboratories.     

Biological versus chronological ageing of oocytes,distinguishable by raised FSH levels in relation to the success of IVF treatment

BACKGROUND: The present study addresses the issue of biological ageing of the oocyte (as indicated by basal serum FSH levels) versus chronological ageing. METHODS: 1019 infertile but ovulating women were studied in their first cycle of IVF treatment. A series of logistic regression models were developed to assess statistical significance of effects of age and FSH on implantation rates and live babies born. RESULTS: The number of oocytes retrieved and embryos available for transfer declined with increasing age and basal serum FSH concentrations. Fertilizing ability of oocytes increased with advancing age but was not affected by FSH concentrations. Although the number of oocytes or embryos available for transfer had no independent effect on implantation rates, the implanting ability of fertilized oocytes (embryos) was inversely related to increasing age and independently to FSH. The chance of a baby being born, however, was determined more by age than by serum FSH. CONCLUSIONS: Ovarian ageing affecting oocyte quality and fecundity can occur independently of chronological age. This has important practical implications whereby serum basal FSH measurement may be a valuable prognostic index, though chronological age remains important.     

Differential patterns of inhibin secretion in response to gonadotrophin stimulation in normal men

Inhibin B is produced by the testis, and its constituent alpha and beta B subunits have been localized immunohistochemically to Leydig as well as Sertoli cells in both rodent and human testes. Whether Leydig cells contribute to circulating inhibin B concentrations, however, is uncertain. We have investigated this by selectively stimulating Leydig and Sertoli cells with hCG and FSH, respectively. The study was a randomized crossover trial, investigating responses to 225 IU recombinant FSH or 3000 IU hCG administered s/c 4-6 weeks apart. Ten normal men were recruited to participate. Blood was taken twice before treatment and after 8, 24, 48, 72 and 96 h. Serum was assayed for FSH, LH and testosterone by radioimmunoassay (RIA); inhibin B and pro-alpha C inhibin forms by ELISA. Administration of hCG, but not FSH, caused a rapid increase in blood testosterone levels, which reached a maximum after 72 h (22.2 +/- 2.7-50.1 +/- 4.5 nmol/L, p < 0.001). Inhibin B concentrations in blood were unchanged following either treatment. Conversely, pro-alpha C concentrations increased following both treatments. FSH administration resulted in a gradual increase in pro-alpha C concentrations (369 +/- 18 pg/mL pre-treatment to 453 +/- 33 pg/mL after 96 h, p=0.013). Administration of hCG resulted in a more rapid response, with pro-alpha C concentrations rising from 384 +/- 23 pg/mL pre-treatment to a peak at 48 h of 535 +/- 45 pg/mL (p=0.007). This response was more rapid than that of testosterone. These results demonstrate that adult human Leydig, as well as Sertoli, cells secrete inhibin alpha subunit in response to gonadotrophin stimulation but provide no evidence for the secretion of inhibin B from Leydig cells. The lack of change in inhibin B secretion in response to FSH suggests that more prolonged or intense stimulation of Sertoli cells may be required for secretion of the dimeric form.     

坤泰胶囊联合唑吡坦治疗肾虚肝郁型更年期综合征睡眠障碍的疗效观察

目的 探讨坤泰胶囊联合唑吡坦治疗肾虚肝郁型更年期综合征睡眠障碍的临床效果。方法 选取2022年1月—2022年12月济宁医学院附属医院收治的120例肾虚肝郁型更年期综合征睡眠障碍患者，随机分为对照组与治疗组，每组各60例。对照组患者睡前口服酒石酸唑吡坦片，起始剂量5 mg/次，1次/d，最大剂量10 mg。治疗组在对照组基础上口服坤泰胶囊，4粒/次，3次/d。两组患者治疗12周。观察两组患者临床疗效，比较治疗前后两组患者中医证候积分和匹兹堡睡眠质量指数量表（PSQI）评分，及雌二醇（E₂）、卵泡刺激素（FSH）和促黄体生成素（LH）水平。结果 治疗后，治疗组患者总有效率较对照组显著升高（98.33% vs 88.33%，P＜0.05）。治疗后，两组患者中医证候积分及PSQI评分均显著低于治疗前（P＜0.05），且治疗组评分显著低于对照组（P＜0.05）。治疗后，两组患者FSH和LH水平显著低于治疗前，而E₂水平显著高于治疗前（P＜0.05），且治疗组患者性激素水平显著好于对照组（P＜0.05）。结论 坤泰胶囊联合唑吡坦治疗肾虚肝郁型更年期综合征睡眠障碍能有效调控患者性激素水平，改善中医证候及提高睡眠质量。     

长期停服棉酚男子血液性激素的变化

本文测定了16名长期停服棉酚男子血清FSH、LH和睾丸酮(T)的水平。停服棉酚后,精子计数为零的7名受试者,血清FSH和LH水平显著高于正常对照组;血清T略低于正常对照组,而T/LH值则显著低于正常对照组。表明睾丸功能受到不同程度的损伤。另外9名停服棉酚后,精子计数完全恢复者,除血FSH高于对照组外,其余指标与正常对照组比较均无显著性差异。提示当棉酚剂量尚未使生精过程受到不可逆性损伤时,其间质细胞的功能受损可能不大。     

Refractoriness to follicle-stimulating hormone in rat ovarian granulosa cells--a concentration- and time-dependent phenomenon

Granulosa cells from pre-ovulatory ovarian follicles of rats were exposed, in vitro, to one or two pulses of follicle-stimulating hormone (FSH) in a superfusion apparatus. The superfusate was analysed for cyclic adenosine-3',5' monophosphate (cAMP) and steroids. In experiments with two consecutive FSH pulses, the response to the second pulse (100 ng/ml) was inversely related to the concentration of the first FSH pulse (2.5-100 ng/ml). Within certain limits, a lower total amount of cAMP and progesterone was accordingly released in response to the two FSH pulses when the concentration of the first pulse was increased. This refractoriness declined within a few hours as shown in experiments with extended pulse intervals. Two short and separate FSH pulses also evoked a higher combined response than did a single long FSH pulse lasting the whole time period. In these cases the cells thus responded with a lower total cAMP and progesterone release when exposed to a higher amount of FSH. Clinical trials on i.m. versus pulsatile i.v. FSH administration to anovulatory women have shown a similar relationship between FSH dose and effect, though other parameters were measured than those in the present study.     

Intratesticular steroids and gonadotrophin receptor concentrations in the testes of immature unilaterally cryptorchid rats

Testicular descent was prevented unilaterally in newborn rats. In this experimental model of cryptorchidism the first morphological changes are noted in the abdominal testis at 16 days of age. Testicular weight was increased in the abdominal testis at 16 days of age, was unaffected at 20 days but was decreased at 30 days of age. The concentrations of LH and FSH receptors, and the hCG-stimulated progesterone and testosterone secretion in vivo were analysed in scrotal and abdominal testes at different ages. LH- and FSH-receptor content per testis were unaffected by cryptorchidism at 12, 16 and 20 days of age but were depressed markedly at 30 days of age. At 20 days of age (but not earlier) hCG-stimulated progesterone was reduced while testosterone secretion remained unchanged. It is concluded that the early functional changes in abdominal testes are not related to changes in gonadotrophin receptor content.     

Acute effects of cadmium on preovulatory serum FSH, LH, and prolactin levels and on ovulation and ovarian hormone secretion in estrous rats

On the day of diestrus II CFY rats were given 5, 10, or 15 mg/kg cadmium chloride (CdCl₂) or 1.0 mL/kg of 0.9% NaCl. On the next day a group of animals was anesthetized with pentobarbital and blood was collected from the aorta at 13:00, 15:00, 16:30, or 18:00 h. for FSH, LH, prolactin (PRL), progesterone (P), and estradiol-17β (E₂) determination. On the day of the expected estrus, the second group of animals was anesthetized with pentobarbital and cannulas were inserted in one of the femoral arteries and veins, and in one of the utero-ovarian veins. Five-minute blood fractions were collected from the ovary for 40 min, and following the first blood samples, 10 IU hCG was injected iv. Ovarian venous outflow and blood pressure were continuously recorded. From the blood fractions, P and E₂ were determined, and their secretion rates were calculated. In a third group of treated animals, the ovaries were excised for histological examination, and oviducts were flushed for counting oocytes. CdCl₂ in the dose of 10 or 15 mg/kg increased the PRL serum levels at 13:00 h; it diminished FSH serum levels in the dose of 10 mg/kg and LH serum levels in the doses of 10 and 15 mg/kg at 15:00 h. The decrease in LH levels continued until 16:30 h in the dose of 10 mg/kg CdCl₂. In estrous animals, CdCl₂ did not influence the blood pressure and ovarian blood flow. In animals receiving 10 or 15 mg/kg CdCl₂, a decrease in basal secretion of P occurred. The hCG induced a rapid rise in P secretion of controls; the 5 mg/kg CdCl₂ diminished the effect of hCG, and the 10 and 15 mg/kg completely abolished it, but failed to influence the basal secretion of E₂ and its response to hCG. CdCl₂ prevented ovulation in 40% to 50% of the animals; however, when ovulation occurred, normal oocyte numbers were found. No alteration could be observed in the light microscopic histology of ovary.