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1.
利用RT-PCR和Western-blotting技术分析了胆固醇侧链裂解酶（P450scc）、3β-羟甾脱氢酶（3β-HSD）、C17-20裂解酶（P450c17）、StAR mRNA和蛋白在成年昆明小白鼠心脏、脾、肝、肾中的表达.结果显示,StAR mRNA及其蛋白、P450scc mRNA在以上组织中均有表达;3β-HSD在肾中有较强的表达,在其他组织的表达较弱;P450c17在上述组织中无明显的表达,但在睾丸中有明显的表达.这表明,心脏、脾、肝、肾均具有合成类固醇的能力,但合成类固醇的类型存在一定的差异.  相似文献   

2.
睾丸间质细胞（Leydig cells,LCs）的主要功能是合成和分泌睾酮。在睾丸间质细胞内,以胆固醇为原料,位于线粒体外膜上的类固醇合成急性调节蛋白（steroidogenic acute regulatory protein,StAR）促进胆固醇向线粒体内膜转运,在线粒体内膜胆固醇侧链裂解酶（cholesterol side-chain cleavage cytochrome,P450scc）的催化下生成孕烯醇酮,而后通过光面内质网的羟基类固醇脱氢酶（3β-hydroxysteroid dehydrogenase,3β-HSD）和转运蛋白（translocator protein,TSPO）的共同作用合成睾酮。因此,睾丸间质细胞合成和分泌睾酮与线粒体密切相关,线粒体结构和功能的完整性直接影响睾酮的生物合成,而位于线粒体上的StAR和P450scc是睾酮合成的关键调控因子。睾酮能够促进雄性生殖器官发育成熟并维持其功能,对促进蛋白质合成（如肌肉、骨骼及生殖器官的蛋白质合成）具有重要意义。近年来,通过维持线粒体结构完整性和改善线粒体氧化损伤、线粒体生物发生等功能进而促进睾酮的合成已成为睾酮合成机制的研究热点,受到国内外学者的广泛关注。作者介绍了睾丸间质细胞内睾酮合成的分子机制及影响睾酮合成的重要因子,综述了睾丸间质细胞线粒体结构、线粒体氧化损伤、线粒体调控的细胞凋亡和线粒体的生物发生等对睾酮合成的影响,阐述了线粒体与睾酮合成之间的关系,为改善睾丸间质细胞线粒体结构和功能从而促进睾酮合成提供依据,对于深入了解雄性动物的睾酮合成调节和提高雄性动物的繁殖性能具有重要的意义。  相似文献   

3.
为了探讨脱氢表雄酮(DHEA)对大鼠原代卵巢颗粒细胞激素生成及相关甾体生成酶的影响,采用机械法分离培养大鼠原代卵巢颗粒细胞,放射免疫分析法检测培养液中雌二醇、孕酮和睾酮的含量;Real-time PCR法检测类固醇代谢途径中3β-羟基固醇脱氢酶(3β-HSD)、17β-羟基固醇脱氢酶(17β-HSD)、CYP450 mRNA的表达变化。结果显示:10μmol/L DHEA处理48 h后可显著促进大鼠原代卵巢颗粒细胞雌二醇(P<0.05)、孕酮(P<0.01)和睾酮的分泌量(P<0.01);显著促进3β-HSD(P<0.01)和17β-HSD(P<0.05)mRNA的表达,CYP450 mRNA表达略有降低,但无统计学意义。说明外源性DHEA处理可能通过调节3β-HSD、17β-HSD和CYP450等相关酶的基因表达,进而影响大鼠原代卵巢颗粒细胞雌激素和雄激素的分泌。  相似文献   

4.
试验旨在研究精索上神经（superior spermatic nerve,SSN）对睾丸功能的影响,25只成年Wistar大鼠随机分为2组,试验组左右睾丸切除精索上神经,手术30 d后两组大鼠同时处死取样分析。结果显示,睾丸去精索上神经不影响睾丸指数的大小但使附睾尾精子数极显著降低（P＜0.01）,平均日增重显著增加（P＜0.05）。增殖细胞核抗原（PCNA）免疫组织化学染色结果显示,睾丸去精索上神经不影响曲细精管精原细胞和初级精母细胞的增殖,但影响曲细精管的形态和生殖细胞的规则排列。RT-PCR结果显示,睾丸去精索上神经明显上调β1AR mRNA的表达,下调β2AR mRNA的表达;另外切除精索上神经不影响睾丸3β-羟胆固醇脱氢酶（3β-HSD）mRNA表达,但显著抑制类固醇快速调节蛋白（StAR）、胆固醇侧链裂解酶（P450scc）mRNA表达（P＜0.05）。结果表明,支配睾丸的精索上神经通过调节肾上腺素能受体β1AR和β2AR的表达及StAR和P450scc影响睾丸睾酮的合成和精子的发生。  相似文献   

5.
为建立成年小鼠睾丸间质细胞分离纯化方法,并对分离纯化的睾丸间质细胞进行睾酮分泌功能检测,对成年小鼠睾丸组织进行Ⅰ型胶原酶消化、Percoll分离液密度梯度离心,分离成年小鼠睾丸间质细胞.细胞纯度用3β-羟类固醇脱氢酶(3β-hydroxy-steroid-dehydrogenase,3β-HSD)染色鉴定.将分离纯化的睾丸间质细胞进行体外培养,在基础睾酮和人绒毛膜促性腺激素(hCG)刺激培养条件下,用放射免疫分析法对培养上清中的睾酮含量进行检测.结果显示,通过该方法能获得高纯度成年小鼠睾丸间质细胞(＞95％),培养的睾丸间质细胞具有分泌基础睾酮以及对hCG刺激反应的能力.提示采用该方法对成年小鼠睾丸间质细胞进行分离纯化具有高效性、可用性,通过该方法获得的睾丸间质细胞是体外研究药物对睾酮分泌功能影响的良好模型.  相似文献   

6.
本试验旨在研究RNA干扰抑制素α亚基(Inhibinα-subunit,INHα)基因和添加抑制素A(InhibinA)对绵羊颗粒细胞雌激素(Estrogen,E2)和孕酮(Progesterone,P)分泌及相关基因表达的影响,以探索抑制素在绵羊颗粒细胞E2和P分泌中的作用。从1.0~1.5岁小尾寒羊的卵泡(3~7mm)中分离培养颗粒细胞,分为2个处理组:干扰组(脂质体介导法转染颗粒细胞)和InhibinA添加组(200ng·mL~(-1))。利用ELISA试剂盒检测E2和P的分泌,荧光定量RT-PCR检测E2和P的分泌相关基因(CYP11、3β-HSD和CYP19)的表达量。结果表明,与对照组相比,干扰组siRNA转染颗粒细胞48h后,INHα基因的抑制率达87%,E2和P的分泌量显著降低(P0.05);InhibinA添加组E2和P的分泌水平显著升高(P0.05);干扰组3β-HSD和CYP19的mRNA表达量显著降低(P0.05),CYP11的表达量显著升高(P0.05);InhibinA添加组CYP19、CYP11和3β-HSD的mRNA表达水平均显著升高(P0.05)。综上表明,抑制素在绵羊颗粒细胞中起关键调控作用,通过调节绵羊颗粒细胞类固醇激素的分泌参与调控卵泡发育和排卵过程。  相似文献   

7.
为探寻绵羊卵巢颗粒细胞分泌孕酮与激素合成相关基因之间的关系,体外培养了绵羊卵巢颗粒细胞并进行鉴定后,经FSH、LH处理后添加不同浓度(1,10,58,100,145nmol/L)的食欲素A分别处理0,24,48,72h后提取蛋白,运用Western blot技术检测绵羊卵巢颗粒细胞分泌孕酮过程中的关键蛋白STAR、3β-HSD和P450(CYP11)的表达量变化情况。结果表明:绵羊卵巢黄体化颗粒细胞添加食欲素A后同一时间STAR、3β-HSD和P450(CYP11)的表达量随食欲素浓度的增加呈逐渐升高然后下降的趋势,且浓度在10nmol/L处表达量达到最高;添加同一浓度的食欲素后,随着作用时间的增加STAR及3β-HSD的表达量呈逐渐升高然后下降的趋势,且时间在24h时表达量达到最高,但P450(CYP11)的浓度随时间的增加而降低。结论:绵羊卵巢颗粒细胞分泌孕酮受食欲素A的浓度及时间变化影响,并进一步证实了食欲素A与孕酮的分泌周期性变化密切相关。  相似文献   

8.
高效氯氰菊酯通过ERK1/2影响小鼠睾丸睾酮合成   总被引：2，自引：0，他引：2  
为了阐明氯氰菊酯（CP）影响睾酮合成的机制,本研究以成年SPF昆白小鼠为试验动物,通过连续7 d灌喂不同浓度的高效氯氰菊酯（β-CP）,采用RT-PCR、Western blot和放射免疫分析技术分析了β-CP对睾丸重量、血浆类固醇水平、类固醇合成酶、StAR蛋白以及ERK1/2活性的影响。结果显示：随着β-CP浓度的增加,小鼠睾丸的重量无明显变化,但小鼠血浆睾酮的水平逐渐降低;P450sccmRNA、3β-HSDmRNA和StAR mRNA的水平则无明显变化,而P450 c17 mRNA的表达明显下降;StAR蛋白的水平也呈下降趋势,但ERK1/2活性则逐渐升高。以上结果表明,β-CP通过激活ERK1/2而级联抑制P450 c 17 mRNA的转录和StAR前体蛋白的裂解,从而降低睾酮的合成。  相似文献   

9.
本研究旨在探究促卵泡素(follicle stimulating hormone,FSH)处理对体外培养的牛有腔卵泡颗粒细胞和膜细胞类固醇激素合成相关基因表达的影响。采集牛卵巢表面直径9～11 mm的有腔卵泡,用含不同浓度FSH的DMEM/F12体外培养牛有腔卵泡24 h。提取卵泡颗粒细胞、膜细胞RNA并反转录成cDNA,利用实时荧光定量PCR检测卵泡颗粒细胞、膜细胞中类固醇激素合成酶基因(CYP11A1、3β-HSD、CYP17A1、CYP19A1、17β-HSD)和促性腺激素受体基因(FSHR、LHR)的表达水平。结果显示,FSH处理上调了颗粒细胞中CYP11A1、3β-HSD和CYP19A1基因表达,其中,25 ng/mL FSH处理极显著上调了CYP11A1基因表达(P0.01),10 ng/mL FSH处理显著上调了3β-HSD基因表达(P0.05),50 ng/mL FSH处理显著上调了CYP19A1基因表达(P0.05);50 ng/mL FSH处理显著或极显著上调了膜细胞中CYP11A1、3β-HSD和CYP17A1基因表达(P0.05;P0.01),但在10和25 ng/mL FSH处理组中CYP11A1、3β-HSD和CYP17A1基因表达显著或极显著下调(P0.05;P0.01)。对FSHR、LHR基因研究结果显示,不同浓度FSH处理对颗粒细胞中FSHR、LHR基因的表达均无显著影响(P0.05),只有25和50 ng/mL FSH处理显著或极显著上调了膜细胞中LHR基因表达水平(P0.05;P0.01),且不同处理组之间膜细胞中CYP11A1、3β-HSD和CYP17A1基因的表达变化与LHR基因表达变化趋势较一致。结果表明,FSH处理可提高牛有腔卵泡颗粒细胞中CYP11A1、3β-HSD和CYP17A1基因的表达,膜细胞中CYP11A1、3β-HSD和CYP17A1基因对LH的刺激更敏感,FSH可能通过影响LHR基因的表达来调节膜细胞中类固醇合成酶基因的表达。  相似文献   

10.
《中国兽医学报》2016,(2):343-347
为追寻能量代谢与激素合成相关基因之间潜在的联系,本试验在秋季经公羊试情后,于0、3、6、9、12d选取不同发情阶段的蒙古绵羊,分别获取黄体和卵巢非黄体组织,通过Western Blot和RT-qPCR技术,研究了绵羊卵巢周期性变化中黄体形成过程的关键基因StAR、3β-HSD和P450(CYP11)与能量代谢通路AMPK-UCP1调控的关系。试验结果提示,StAR、3β-HSD和P450(CYP11)参与了绵羊卵巢周期性变化并与孕酮的分泌协调一致,与此同时AMPK-UCP1通路参与黄体周期性变化的调控,进一步证实了能量代谢平衡与黄体的周期性变化密切相关。  相似文献   

11.
Expression of mRNAs encoding cytochrome P450 side-chain cleavage (P450scc), cytochrome P450 17 -hydroxylase (P450c17), and cytochrome P450 aromatase (P450arom) were characterized by the RT-PCR technique and concentrations of progesterone (P4), testosterone (T0) and estradiol (E2) were measured by radioimmunoassay during follicular development of prepubertal goats. Synthesis of mRNAs encoding P450scc and P450c17 began in preantral follicles, but mRNA encoding P450arom was not detectable until early antral formation. While mRNA for P450scc was expressed in both theca and granulosa cells, mRNA for P450c17 was expressed only in theca cells while P450arom mRNA only in granulosa cells. In nonatretic follicles from prepubertal ovaries, the relative quantity of mRNA expression of all the three enzymes increased with follicle size; however, while the concentration of P4 and E2 increased, that of T0 decreased with follicle size. While expression of mRNA encoding P450scc was unaffected, that of P450c17 mRNA decreased to the lowest level and mRNA for P450arom became undetectable following atresia; accordingly, while the concentration of P4 increased in the atretic medium follicles, that of T0 and E2 decreased to the lowest level after atresia. While the adult follicular stage follicles showed a similar cytochrome expression as the nonatretic follicles of prepubertal goats, the former contained higher levels of E2 and P4 than the latter. The presence of corpus luteum in an ovary decreased expression of P450scc, significantly in large follicles while it increased concentration of P4. These findings indicated that (1) similar to other species, changes in follicular steroid production in goats were explained in large measure by changes in steroidogenic enzyme expression; (2) while mRNA expression was similar, activities of some of the steroidogenic enzymes may differ between sexually mature and immature goats.  相似文献   

12.
Testosterone secretion and the expression and relative contents of steroidogenic acute regulatory (StAR) protein and steroidogenic enzymes cholesterol side-chain cleavage cytochrome P450 (P450scc), 3β-hydroxysteroid dehydrogenase /Δ5 → Δ4 − isomerase (3β-HSD), and 17α-hydroxylase cytochrome P450/C17–20 lyase (P45017α) were determined in testicular tissues of bulls treated with a LHRH agonist. Testis morphology and spermatogenesis were also examined. In Experiment 1, bulls (30-mo-old) received no treatment (control, n = 7) or were implanted for 10 days with the LHRH agonist deslorelin (n = 7). Bulls were castrated on Day 10 and testis tissues prepared for Western and Northern blotting. At castration, bulls implanted with deslorelin had greater plasma testosterone (5-fold) and testis content of testosterone (10-fold) compared with control bulls. Relative content (per μg total testis protein or RNA) of StAR protein, 3β-HSD, P450scc, and mRNA for P45017α in bulls treated with deslorelin ranged from 3- to 6-fold that of control bulls. In Experiment 2, bulls (20-mo-old) were left untreated (control, n = 6) or implanted with deslorelin (n = 12) for 120 days. On Day 120, bulls were castrated and right testis tissues prepared for morphology. Testis volume and weight were increased (P < 0.01) in bulls treated with deslorelin compared with control bulls. Stereological analysis revealed that this increase occurred in all compartments (seminiferous epithelium, lumen and interstitium) studied, but was significant (P < 0.01) only for the seminiferous epithelium. Absolute numbers of round spermatids per testis were increased (P < 0.05) in bulls treated with deslorelin compared with control bulls. Increased testosterone secretion in bulls treated with deslorelin was associated with increased testicular StAR protein and steroidogenic enzymes. Bulls treated long-term with deslorelin had a faster rate of testis growth and increased daily sperm production at the end of the experiment.  相似文献   

13.
Twenty-one wild male Japanese black bears (Ursus thibetanus japonicus) were captured in the summer-autumn of 1998-2000 in the vicinity of Neo Village, Gifu Prefecture. Testes were measured, and testicular samples were biopsied and observed histologically. Four steroidogenic enzymes, i.e., cholesterol side-chain cleavage cytochrome P450 (P450scc), 3beta-hydroxysteroid dehydrogenase (3betaHSD), 17-alpha hydroxylase cytochrome P450 (P450c17), and aromatase cytochrome P450 (P450arom) were immunolocalized. Serum testosterone concentrations were measured by radioimmunoassay. Testis size changed little from 1-3 years of age, increased rapidly at 4 years, and attained its peak at 5 years. Serum testosterone concentrations ranged from 0.05 to 1.78 ng/m l, and the mean +/- standard deviation was 0.43 +/- 0.48 ng/ml. Age of sexual maturation in wild male Japanese black bears was estimated to be 3-4 years. Seasonal changes in spermatogenesis were obvious; active in June, July and August, degenerated by September. Leydig cells, Sertoli cells and germ cells have the capability of synthesizing androgen, and Leydig cells, Sertoli cells, spermatids and spermatogonia have the capability of synthesizing estrogen in Japanese black bears.  相似文献   

14.
Nineteen cycling ewes underwent transrectal ultrasonography of ovaries followed by ovariectomies during the growth phase of the first follicular wave of the interovulatory interval or the proestrus/estrus phase of the cycle. Quantitative ultrasonographic characteristics of the antrum and follicular wall in a total of forty-three ovine antral follicles were examined for correlations with the protein expression of three steroidogenic enzymes (cytochrome P450 17α-hydroxylase, CYP17; cytochrome P450 aromatase, CYP19; and 3β-hydroxysteroid dehydrogenase, 3β-HSD) determined by densitometric analysis of immunohistochemical slides, follicular dimensions, granulosa layer thickness and the percentage of apoptotic granulosa cells. Significant correlations were found between echotextural attributes of ovine antral follicles and the percentage of apoptotic granulosa cells, CYP17 expression (theca), CYP19 expression (granulosa) and 3β-HSD expression (theca cells). Computer-aided analyses of ultrasonographic images can be beneficial to the development of assisted reproductive technologies and diagnosis of hormonal imbalances without the need for ovarian biopsies or hormone assays.  相似文献   

15.
本研究采用低氧(5％O2)条件培养水牛内膜细胞,探讨低氧对水牛内膜细胞雄激素合成的影响.结果 显示,低氧能显著提高水牛内膜细胞雄激素合成相关基因(CYP11A1、CYP17A1和3β-HSD)表达和睾酮分泌水平(P＜0.05);低氧能显著增强水牛内膜细胞对促黄体素(LH)的敏感性(P＜0.05).结果 表明,低氧(5％...  相似文献   

16.
The purpose of this study was to explore the variations in the circulating leptin concentrations of the wild ground squirrels in relation to seasonal changes in testicular activities. Hematoxylin-eosin staining showed all types of elongated spermatids and spermatogenic cells existed in the testis in April, while the primary spermatocytes and spermatogonia were most advanced stages of germ cells in June. In addition, the primary spermatocytes, secondary spermatocytes, and spermatogonia were most advanced stages of germ cells in September. The highest circulating leptin concentration was consistent with the maximum body weight results from accumulation of adipose tissue in September. The mRNA expression level of leptin receptor (Ob-R) and STAT3 was lowest in June, raised in September, and remained increased in April. Ob-R and STAT3 were stronger staining in the Leydig cells in July. Moreover, the concentrations of testosterone (T) showed the maximum values in April, the minimum values in June, and significant increases in September. Furthermore, it is worth noting that the levels of T increased with the mRNA levels of Ob-R, STAT3, StAR, and testicular steroidogenic enzymes (3β-HSD, P450c17, and P450scc). Moreover, RNA-seq analyses of testis during the different periods showed that a total of 4209 genes were differentially expressed genes (DEGs); further analysis revealed that DEGs related with the Jak/STAT pathways and reproduction were altered. Taken together, the results suggested that the leptin regulated testicular function through the Jak/STAT pathways and testicular steroidogenic factor expressions.  相似文献   

17.
The objective of this study was to investigate immunolocalization of steroidogenic enzymes in G?ttingen miniature (GM) pig testes. Testes of 6 adult GM pigs were obtained in September 1996 (n=2), February (n=2) and June (n=2), 1997. Steroidogenic enzymes were immunolocalized using polyclonal antisera raised against bovine adrenal cholesterol side-chain cleavage cytochrome P450 (P450scc), human placental 3beta-hydroxysteroid dehydrogenase (3betaHSD), porcine testicular 17alpha-hydroxylase cytochrome P450 (P450c17), and human placental aromatase cytochrome P450 (P450arom). Histologically, all types of spermatogenic cells including mature-phase spermatozoa in seminiferous tubules were observed in all testes throughout the year. Moreover, P450scc, 3betaHSD, P450c17and P450arom were identified in Leydig cells but not in Sertoli cells of all testes. These results suggested that adult GM pig testes have the ability to produce germ cells throughout the year, and the synthesis of progestin, androgen and estrogen occurs in the Leydig cells of GM pig testes.  相似文献   

18.
Although circulating progesterone (P(4)) levels tend to change with the season, little is known about the seasonal changes of P(4) synthesis-related proteins in the corpus luteum (CL) of mares. To examine these changes, seventy-four ovaries containing a CL were collected from Anglo-Norman mares at a local abattoir in Kumamoto, Japan (~N32°), five times during one year. The stages of the CLs were classified as early, mid and regressed by macroscopic observation of the CL and follicles. The mid CL, which had the highest P(4) concentration, was used to evaluate the seasonal changes in P(4) synthesis. The luteal P(4) concentration and mRNA expression of luteinizing hormone receptor (LHCGR) were lowest during early winter and highest during late winter. The mRNA expressions of steroidogenic acute regulatory protein (StAR), P450 cholesterol side-chain cleavage enzyme (P450scc) and 3β-hydroxysteroid dehydrogenase/Δ5-Δ4 isomerase (3β-HSD) were lowest during early winter and increased during late winter. These results suggest that P(4) synthesis in the CL is affected by the seasonal changes in the mRNA expressions of P(4) synthesis-related proteins in mares.  相似文献   

19.
The raccoon is a seasonal breeder with a mating season in the winter. In a previous study, adult male raccoons exhibited active spermatogenesis with high plasma testosterone concentrations, in the winter mating season. Maintenance of spermatogenesis generally requires high testosterone, which is produced by steroidogenic enzymes. However, even in the summer non-mating season, some males produce spermatozoa actively despite low plasma testosterone concentrations. To identify the factors that regulate testosterone production and contribute to differences in spermatogenetic activity in the summer non-mating season, morphological, histological and endocrinological changes in the testes of wild male raccoons should be known. In this study, to assess changes in the biosynthesis, metabolism and reactivity of testosterone, the localization and immunohistochemical staining intensity of four steroidogenic enzymes (P450scc, P450c17, 3βHSD, P450arom) and the androgen receptor (AR) were investigated using immunohistochemical methods. P450scc and P450c17 were detected in testicular tissue throughout the year. Seasonal changes in testosterone concentration were correlated with 3βHSD expression, suggesting that 3βHSD may be important in regulating the seasonality of testosterone production in raccoon testes. Immunostaining of P450arom and AR was detected in testicular tissues that exhibited active spermatogenesis in the summer, while staining was scarce in aspermatogenic testes. This suggests that spermatogenesis in the raccoon testis might be maintained by some mechanism that regulates P450arom expression in synthesizing estradiol and AR expression in controlling reactivity to testosterone.  相似文献   

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