犏牛和牦牛ESCO2基因的序列特征及其在睾丸中的表达对比分析

旨在克隆犏牛和牦牛姐妹染色单体内聚建立蛋白2（establishment of sister chromatid cohesion N-acetyltransferase 2,ESCO2）基因,并分析其在不同发育阶段睾丸中的表达与定位,为进一步解析ESCO2在减数分裂过程中的作用机制提供理论依据。本研究以健康雄性犏牛及牦牛为试验动物,根据年龄分为胎牛组（5~6月龄）、幼年组（1~2岁）和成年组（3~4岁）,每组各3头。通过RT-PCR技术克隆犏牛和牦牛ESCO2基因并进行生物信息学分析,采用实时荧光定量PCR （quantitative real-time PCR,qRT-PCR）检测ESCO2基因在犏牛不同组织中的表达谱,比较分析ESCO2在犏牛和牦牛不同时期睾丸中的表达规律,利用免疫组织化学（immunohistochemistry,IHC）技术检测ESCO2蛋白的细胞定位和表达差异。结果显示,犏牛ESCO2基因（GenBank登录号：MW198470） CDS区为1 833 bp,编码610个氨基酸,与牦牛相比,犏牛ESCO2序列第301~319位多19个氨基酸,另有3个氨基酸突变;犏牛ESCO2蛋白序列与黄牛的同源性高于其他哺乳动物;ESCO2可能与SMC3、SMC1A、PDS5A、PDS5B、STAG2等蛋白相互作用,互作蛋白功能与姐妹染色单体凝聚、减数分裂细胞周期、DNA修复、细胞分裂和染色体重构等生物学过程相关。ESCO2在犏牛各组织中均有表达,但在睾丸中的相对表达水平显著高于其它组织（P<0.05）;在犏牛睾丸中的表达随年龄增长呈上升趋势,幼年和成年时期犏牛睾丸中ESCO2的表达显著低于同时期牦牛（P<0.05）;IHC染色结果发现,雄性犏牛减数分裂阻滞于初级精母细胞,ESCO2蛋白在犏牛初级精母细胞中无表达并与牦牛存在差异。本研究结果表明,犏牛与牦牛的ESCO2基因、蛋白序列差异较大,且在睾丸发育过程中的表达模式差异显著,这可能是引起雄性犏牛减数分裂阻滞及不育的原因之一,其具体作用机制有待进一步研究。     

FGF9调节性腺分化研究进展

性别决定过程是双潜能胚胎发育成睾丸或卵巢的过程.通过小鼠基因敲除试验以及在遗传水平分析性反转病人证明该过程是由多个基因调控的.论文综述了睾丸发育通路中信号分子成纤维细胞生长因子9(FGF9)在性腺分化中的作用.FGF9基因参与调节早期睾丸发育的3个重要事件,为调节性腺体腔上皮细胞增殖,促进Sertoli前体细胞形成,调节中肾细胞迁移,影响睾丸索的形成,FGF9诱导FGFR2在核内定位,并维持Sox9的表达.     

Study on Expression of Bcl-2 and PCNA Genes in Testicular Tissue of Chickens,Quails and Chicken-quail Hybrids

In order to investigate the molecular mechanism of stunted growth testis of chicken (♂) and quail (♀) hybrids. 84 testicular tissue samples of New Roman cocks, male Korean quails and chicken-quail hybrids at different developmental stages were collected,the mRNA expression of Bcl-2 and PCNA genes in testicular tissue of cocks, quails and chicken-quail hybrids at different growth stages were detected using Real-time PCR. The results showed that the Bcl-2 and PCNA genes mRNA expression patterns in testes of chickens and quails at different growth stages were similar.The Bcl-2 gene was fluctuated in a whole, and decreased to the lowest value, then recovered rapidly and remained at a high level. The PCNA gene had a very significant peak value.Compared with the chicken and quail,the expression of Bcl-2 and PCNA genes mRNA in hybrids had no obvious change, which indicated that the pattern of the Bcl-2 gene regulation apoptosis in testis was different from chicken and quail at the hybrid testicular development process, and there was no obvious cell appreciation process.It was suggested that the molecular influencing factors of testicular dysplasia of chicken and quail hybrids were related to the abnormal expression of Bcl-2 and PCNA genes mRNA in testes.The results provided an important reference for the further study of the mechanism of testicular dysplasia between chicken and quail interspecific hybrids.     

睾丸生理功能的植物性神经调节研究进展

研究发现除了内分泌调控外,睾丸神经对睾丸的发育、雄激素的分泌以及精子的发生也发挥重要的调节作用,与睾丸的内分泌调节功能相比较人们对植物性神经在睾丸生理活动调节方面的认识还很少。论文从睾丸的植物性神经分布、精索神经对睾丸发育和睾丸生理功能的影响以及睾丸不同发育时期胆碱能受体和肾上腺素能受体的分布、神经递质NE和Ach对睾丸发育和功能的影响等方面做了详细综述,并对今后的研究方向和研究意义做了展望。     

日粮添加不同水平芦丁对热应激小鼠睾丸组织的影响

本试验旨在探讨日粮添加不同水平芦丁对缓解热应激小鼠睾丸组织损伤的影响。将30只5周龄体重相近（20～22 g）的ICR系雄性小鼠随机分为5组,各组小鼠分别饲喂基础日粮添加0（CON组）、0（HS组）、250（HS+R250）、500（HS+R500）和1 000（HS+R1000） mg·kg^-1芦丁的日粮,饲养10 d后,除对照组（CON）外,所有小鼠在每天10：00至14：00之间置于42℃恒温箱中连续处理8 d后屠宰取样分析。结果显示：1）与CON组相比,HS组小鼠睾丸指数无显著变化（P>0.05）,而日粮添加250 mg·kg^-1芦丁显著提高热应激小鼠睾丸指数（P<0.05）;小鼠睾丸组织切片结果显示,与CON组相比,HS组小鼠生精小管直径和横截面积显著降低（P<0.05）,生精细胞脱落比率显著增加（P<0.05）;与HS组相比,HS+R250组小鼠生精小管横截面积显著增加（P<0.05）,生精细胞脱落比率显著降低（P<0.05）,HS+R500组生精细胞脱落比率也显著降低（P<0.05）,HS+R1000组小鼠生精小管直径显著增加（P<0.05）;小鼠睾丸指数、生精小管直径及生精小管脱落比率在芦丁处理组与CON组之间无显著差异（P>0.05）,但HS+R250组小鼠睾丸生精小管横截面积显著高于CON组（P<0.05）。2）与CON组相比,HS组小鼠睾丸组织丙二醛（MDA）含量显著升高（P<0.05）,总抗氧化能力（T-AOC）与还原型谷胱甘肽（GSH）含量以及血红素酶-1（HO-1） mRNA的表达量均显著降低（P<0.05）;日粮添加250 mg·kg^-1芦丁显著降低热应激小鼠睾丸组织中MDA含量（P<0.05）,提高T-AOC与GSH含量（P<0.05）,并增强核因子E2相关因子2（Nrf2）、HO-1和谷胱甘肽过氧化物酶（GSH-Px） mRNA的表达量（P<0.05）,而添加500和1 000 mg·kg^-1芦丁也显著提高了GSH含量（P<0.05）;与CON组相比,除HS+R250组Nrf2 mRNA表达量显著高于CON组（P<0.05）外,芦丁组热应激小鼠睾丸组织抗氧化相应基因与酶活指标均无显著性变化（P>0.05）。3）热应激小鼠睾丸中核因子-κB （NF-κB）、TOLL样受体4（TLR-4）和Bax的mRNA表达量显著增加（P<0.05）,而Bcl-2表达量显著降低（P<0.05）;日粮添加250 mg·kg^-1芦丁显著降低NF-κB、TLR-4、髓样分化因子88（MyD88）、白细胞介素-Iβ（IL-1β）和Bax mRNA表达量（P<0.05）,增强Bcl-2的表达水平（P<0.05）;添加500 mg·kg^-1芦丁显著降低NF-κB和TLR-4表达量（P<0.05）,而1 000 mg·kg^-1芦丁能够显著增加Bcl-2的表达（P<0.05）;小鼠睾丸组织中免疫和增值凋亡相关基因的表达量在添加芦丁组及CON组之间无显著性差异（P>0.05）。结果提示,日粮添加适宜剂量芦丁具有改善热应激小鼠睾丸组织形态及功能的效果,其机制可能与芦丁通过Nrf2信号通路缓解氧化应激,通过TLR-4/NF-κB信号通路抑制炎症反应及调控Bax/Bcl-2 mRNA的表达密切相关。本试验条件下日粮添加250 mg·kg^-1芦丁的效果较好。     

关中奶山羊睾丸不同发育阶段代谢物的差异分析

【目的】 试验旨在从代谢组学角度探究关中奶山羊睾丸发育代谢机制。【方法】 选择1月龄断奶雄性关中奶山羊和24月龄体成熟的雄性关中奶山羊各6只,采集睾丸组织,使用液相色谱-质谱（LC-MS）技术对睾丸组织进行化学检测,通过多维和单维分析相结合的方式对两组不同代谢物进行化学检测、对比与鉴别,通过富集差异代谢物分析筛选关中奶山羊发育代谢中的潜在关键通路。【结果】 在1和24月龄关中奶山羊睾丸组织中共筛选出334个差异代谢物,其中显著上调137个,显著下调197个。对前36个差异代谢物进行比较鉴定发现,分别为脂质和类脂质分子、有机酸及其衍生物、未分类化合物和苯丙烷和聚酮化合物四大类。经相关性分析发现,在1和24月龄关中奶山羊睾丸发育过程中,脂质和类脂质分子与甘油磷脂呈最大正相关,羧基及其衍生物与甘油磷脂呈最大负相关。对不同的代谢物进行富集分析,筛选出7条潜在的重要代谢通路,分别为肿瘤中的胆碱代谢、精氨酸和脯氨酸代谢、肿瘤中的中心碳代谢、铁死亡、谷胱甘肽代谢、氨酰-tRNA生物组成以及牛磺酸和亚牛磺酸代谢。【结论】 本试验从代谢组学角度揭示了1和24月龄关中奶山羊睾丸的差异代谢物及代谢机制,为今后哺乳动物睾丸的发育研究奠定了基础。     

Effect of a single injection of gonadotropin-releasing hormone (GnRH) and human chorionic gonadotropin (hCG) on testicular blood flow measured by color doppler ultrasonography in male Shiba goats

Although color Doppler ultrasonography has been used to evaluate testicular blood flow in many species, very little has been done in goat. Eight male Shiba goats were exposed to a single intramuscular injection of either gonadotropin-releasing hormone (GnRH group; 1 µg/kg BW) or human chorionic gonadotropin (hCG group; 25 IU/kg BW). Plasma testosterone (T), estradiol (E2) and inhibin (INH) were measured just before (0 hr) and at different intervals post injection by radioimmunoassay. Testis volume (TV) and Doppler indices, such as resistive index (RI) and pulsatility index (PI) of the supratesticular artery, were measured by B-mode and color Doppler ultrasonography, respectively. The results indicated an increase in testicular blood flow in both groups, as RI and PI decreased significantly (P<0.05), but this increase was significant higher and earlier in hCG group (1 hr) than in the GnRH group (2 hr). A high correlation was found for RI and PI with both T (RI, r= −0.862; PI, r= −0.707) and INH in the GnRH group (RI, r=0.661; PI, r=0.701). However, a significant (P<0.05) correlation was found between E2 and both RI (r= −0.610) and PI (r= −0.763) in hCG group. In addition, TV significantly increased and was highly correlated with RI in both groups (GnRH, r= −0.718; hCG, r= −0.779). In conclusion, hCG and GnRH may improve testicular blood flow and TV in Shiba goats.     

嫁虫戚精巢的组织学研究

本文研究了嫁虫戚雄性生殖系统的组织学。嫁虫戚的精巢结构同贻贝、栉孔扇贝的相似,也是由外膜及其内的无数生殖小管构成。外膜较薄,由平滑肌和少量结缔组织组成,只在局部区域含有单层柱状上皮细胞。生殖小管间充填着结缔组织及来自外膜的薄层肌肉,结缔组织中含有血管。生殖小管由位于基膜上的生殖上皮及由生殖上皮增殖出的精原细胞、初级精母细胞、次级精母细胞、精细胞和精子组成。精原细胞较大,圆形或椭圆形,核较大,与细胞同形,染色质呈颗粒状靠近核膜分布,核仁1个,圆形,明显。初级精母细胞和次级精母细胞结构相似。精细胞圆形,核染色深。精子呈长棒状,悬浮于生殖小管腔内。     

Pubertal development of male African catfish,Clarias gariepinus. In vitro steroidogenesis by testis and interrenal tissue and plasma levels of sexual steroids

In fish, sex steroids initiate and/or accelerate the maturation of the brain-pituitary-gonad axis. In order to obtain information on the steroid milieu during the pubertal development of male African catfish, we have monitored the conversion of [³H]-pregnenolone and [³H]-androstenedione by testis and [³H]-pregnenolone by interrenal tissue fragmentsin vitro. Pubertal development occurs between two and six months of age. Testicular development proceeds through four main stages that are characterised histologically by the presence of spermatogonia (stage I), spermatogonia and spermatocytes (stage II), spermatogonia, spermatocytes and spermatids (stage III), and all germ cells including spermatozoa (stage IV). 11β-Hydroxyandrostenedione and cortisol were the main products of testes and interrenal tissue, respectively, in all stages of the pubertal development; a change in the steroidogenic pattern was not observed during this period. The interrenal tissue displayed no significant conversion of [³H]-pregnenolone to 11-oxygenated androgens. Blood plasma was analyzed for the presence of five androgens; testosterone, 11β-hydroxytestosterone, 11β-hydroxyandrostenedione, androstenetrione, and 11-ketotestosterone. 11-Ketotestosterone was the quantitatively dominating androgen in the circulation at all stages of development, which was more pronounced in stages III and IV. The obvious differences between thein vitro andin vivo results, namely 11β-hydroxyandrostenedione being the main testicular productvs. 11-ketotestosterone dominating in the blood, may indicate that 11β-hydroxyandrostenedione is converted to 11-ketotestosterone at extratesticular sites.