糖皮质激素诱导大鼠睾丸间质细胞凋亡的研究

目的 :研究糖皮质激素诱导不同发育阶段培养的大鼠睾丸间质细胞凋亡状况。方法 :以Annexin V FITC和PI双标细胞 ,应用激光共聚焦显微镜和流式细胞仪检测。　结果 :ILC、ALC经皮质酮处理后 ,其凋亡量显著高于对照组 ;PLC经皮质酮处理后 ,其凋亡量与对照组无显著差异。　结论 :糖皮质激素能诱导大鼠睾丸未成熟型和成熟型间质细胞凋亡 ,而不能诱导前体型间质细胞凋亡。     

热应反应对感染性休克大鼠肺损伤的影响

目的 研究热应激反应对感染性休克大鼠肺损伤的影响。方法 Ｗｉｓｔａｒ大鼠２４只随机分为三组。分别经肺动脉注射ＮＳ０．５ｍｌ（１组）或内毒素（ＬＰＳ）１５ｍｇ．ｋｇ＾－１（Ⅱ组）或热应激处理１５分钟恢复２４小时后注射ＬＰＳ１５ｍｇ．ｋｇ＾－１（Ⅲ组）,然后取注射ＮＳ或ＬＰＳ前和后１、１．５、２、２．５、３．４小时末的动脉血分离血浆测定ＴＮＦ－α,同时用四导生理仪记录其平均肺动脉压（ｍＰＡＰ）,平均动     

大鼠内源性糖皮质激素对间质细胞功能的影响

本文旨在评价大鼠体内糖皮质激素（大鼠体内是皮质酮）对间质细胞中睾酮合成和１１－β羟类固醇脱氢酶（１１β－ＨＳＤ）活性的影响。为观察内源性皮质酮对间质细胞中睾酮合成的影响，本文测定了切除肾上腺雄性大鼠（４５天龄）血清皮质酮、睾酮，ＬＨ含量和经ＬＨ刺激的纯化间质细胞中睾酮生成量及纯化间质细胞中１１β－ＨＳＤ活性水平。结果表明，切除肾上腺后大鼠血清睾酮水平高于手术对照组及切除后补充皮质酮大鼠。切除肾上腺后大鼠和切除后补充皮质酮大鼠血清ＬＨ水平均无明显变化，表明在上述二实验组中，由于内源性皮质酮变化而导致的间质细胞中睾酮生成的变化不依赖ＬＨ的存在。切除肾上腺后大鼠其纯化的间质细胞睾酮生成量比对照组增加一倍。但经补充皮质酮后睾酮生成量被抑制到对照组水平以下，表明给切除肾上腺后大鼠补充皮质酮能明显地抑制睾酮生成。切除肾上腺后大鼠间质细胞中１１β－ＨＳＤ活性降低，并且这一降低可经补充皮质酮得到恢复。上述结果表明生理性浓度皮质酮对间质细胞中睾酮合成行使直接的负性控制，同时诱导细胞内１１β－ＨＳＤ活性，由此保护间质细胞中睾酮合成免于受到糖皮质激素的过度抑制。     

激素调节大鼠间质细胞中11β—羟类固醇脱氢酶活性

糖皮质激素（大鼠体内是皮质酮）抑制睾丸间质细胞中睾酮合成。１１β－羟类固醇脱氢酶（１１β－ＨＳＤ）通过控制细胞内糖皮质激素浓度调节睾酮合成,并且其活性又能被大鼠内源性皮质酮诱导,由此保护了间质细胞中睾酮的生物合成,使之免于受到糖皮质激素的过度抑制。本研究旨在观察大鼠间质细胞中受１１β－ＨＳＤ调节的两种激素———皮质酮和睾酮是否对培养的大鼠间质细胞中１１β－ＨＳＤ活性及其ｍＲＮＡ表达具有调节作用。结果表明,皮质酮能增加切除肾上腺大鼠培养的间质细胞中１１β－ＨＳＤ氧化酶活性及其ｍＲＮＡ表达。睾酮则明显抑制培养的大鼠间质细胞中１１β－ＨＳＤ氧化酶活性及其ｍＲＮＡ表达。     

损伤大鼠坐骨神经诱导运动神经元凋亡的初步报告

目的探索成年大鼠坐骨神经高位损伤后是否出现运动神经元细胞的凋亡。方法取鼠龄为５～６周的成年雄性ＳＤ大鼠２４只,体重１２０～１５０ｇ。大鼠分组：（１）正常对照组,６只大鼠;（２）实验组：１８只大鼠。高位切断并结扎其坐骨神经,按手术先后随机分成术后５、１４、２１天３个不同时间组。按术后不同时间处死动物。用４％多聚甲醛经心脏灌注后,切取Ｌ４～６节段腰髓,制成石蜡切片。采用原位末端脱氧核糖核苷酸转移酶介导的ｄＵＤＰ标记法标记凋亡的神经元。结果对照组未出现阳性标记的细胞。神经损伤后第５天有１只大鼠伤侧前角出现３个阳性标记的凋亡运动神经元。伤后第１４天、２１天出现凋亡神经元的数目分别为２．２±１．２及５．２±２．３个（ｘ±ｓｘ,下同）。结论当成年大鼠坐骨神经切断并被结扎后,由于阻断了神经营养因子的逆行运输,可以导致运动神经元的凋亡。     

大鼠内源性糖皮质激素对间质细胞功能的影响

本文旨在评价大鼠体内糖皮质激素（大鼠体内是皮质酮）对间质细胞中睾酮合成和１１－β羟类固醇脱氢酶（１１β－ＨＳＤ）活性的影响，为观察内源性皮质酮对间质细胞中睾酮合成的影响，本文测定了切除肾上腺雄性大鼠（４５天龄）血清皮质酮睾酮，ＬＨ含量和经ＬＨ刺激的纯化间质细胞中质酮生成量及纯化间质细胞中１１β－ＨＳＤ活性水平，结果表明，切除肾上腺后大鼠血清睾酮水平高于手术对照组及切除后补充皮质酮大鼠。切除肾上腺后     

大鼠间质细胞中11β-羟类固醇脱氢酶mRNA的表达

间质细胞中１１β－羟类固醇脱氢酶（１１β－ＨＳＤ）通过控制细胞内糖皮质激素（大鼠体内是皮质酮）浓度调节睾酮合成。大鼠出生后２６天内，其未成熟间质细胞不含１１β－ＨＳＤ，随着间质细胞成熟，１１β－ＨＳＤ逐渐产生，成年大鼠间质细胞中１１β－ＨＳＤ活性最高。１１β－ＨＳＤ控制着间质细胞中糖皮质激素浓度，生理浓度糖皮质激素同样调节着１１β－ＨＳＤ活性水平。本研究观察了三个不同年龄组大鼠和切除肾上腺后及切除后补充皮质酮大鼠，间质细胞中１１β－ＨＳＤｍＲＮＡ表达情况，发现：出生２１天大鼠间质细胞中无１１β－ＨＳＤｍＲＮＡ的表达，４５天大鼠有１１β－ＨＳＤｍＲＮＡ表达，９０天大鼠表达量最大，切除肾上腺后大鼠间质细胞中１１β－ＨＳＤｍＲＮＡ表达降低，切除后经皮质酮补充大鼠１１β－ＨＳＤｍＲＮＡ表达量回复到对照组水平。结果表明三个不同年龄组大鼠间质细胞中１１β－ＨＳＤｍＲＮＡ表达情况和以往采用免疫组织化学及生物化学方法测定抗原含量或酶活性的结果是一致的。内源性及生理浓度的皮质酮对１１β－ＨＳＤｍＲＮＡ表达的调节也表现出类似的一致性     

人绒毛膜促性腺激素对大鼠睾丸LH/hCG受体影响的研究

８０只Ｗｉｓｔａｒ大鼠分为对照组、单剂人绒毛膜促性腺激素（ｈＣＧ）注射组、间隔３日双剂ｈＣＧ注射组及连续多剂ｈＣＧ注射组。每只大鼠每天注射的ｈＣＧ剂量为１００ＩＵ，分别在１、３、５、７、１０天分批处死动物，用放射配体分析法测定睾丸组织中１２５Ｉ－ｈＣＧ结合位点。结果ｈＣＧ注射后２４小时ＬＨ／ｈＣＧ的结合位点较对照组降低了９９．３８％，单剂组的ｈＣＧ结合率１０天恢复至正常水平；双剂组再次注射后第１０天恢复至８４．４％，而连续多剂组一直处于低水平（０．７％～１．４％）。     

医源性肝外胆管损伤的处理

目的 总结医源性胆管损伤的处理经验。方法 对５４例肝外胆管损伤处理进行回顾性分析。结果 ５４例中６例为腹腔镜胆囊切除术损伤,４８例为开腹胆囊切除术损伤。术中及时发现１８例（３３．３％）。术后２４ｈ后发现２４例（４４．４％）,手术１个月以后发现１２例（２２．３％）。５４例经道次处理后２３例（４２．６％）治愈,２８例（５１．９％）经过２次以上胆管手术,死亡３例（５．６％）。结论 正确掌握胆管损伤的处理     

溶脲脲原体感染动物模型的建立──在泌尿生殖系中的分布

将１００只雄性ＳＤ大鼠分为３组。Ａ组４５只；Ｂ组２２只；Ｃ组３３只。Ａ组每只动物经膀胱注入血清８型溶脲脲原体（ｕｒｅａｐｌａｓｍａｕｒｅａｌｙｔｉｃｕｍ，ＵＵ）菌株（１９６０），浓度为１０５ＣＣＵ／ｍｌ。Ｂ组处理方法同Ａ组，在接种３个月后给予美满霉素（２０～１００ｍｇ／ｋｇ体重），连续１４天。Ｃ组为对照组，每只动物膀胱注入等量溶脲脲原体液体培养基。结果表明，在Ａ组动物的膀胱（８７．５％）、睾丸（６２．５％）、附睾（５５．０％）、精囊（６５．０％）和前列腺（５７．５％）中均检出溶脲脲原体。Ｂ组溶脲脲原体检出率显著低于Ａ组。Ｃ组上述器官溶脲脲原体培养均为阴性。     

皮质酮诱导大鼠Leydig细胞凋亡是一经caspase-3激活的过程

目的 超生理剂量的皮质酮(大鼠体内的糖皮质激素)能诱导大鼠Leydig细胞凋亡。但有关皮质酮诱导Leydig细胞凋亡的细胞内机制尚不清楚。本研究旨在观察皮质酮是否经caspase-3激活的途径诱导大鼠Leydig细胞凋亡。方法 采用Western Blot方法检测不同时间点上经皮质酮处理的大鼠Leydig细胞中caspase-3酶原及裂解的caspase-3酶表达情况。运用荧光发光法检测不同时间点上经皮质酮处理的人鼠Leydig细胞中caspase-3酶活性。结果 caspase-3酶原表达水平在皮质酮处理6h时开始上升，12h及24h时表达量下降，而具生物活性的、裂解的caspase-3酶于12h时开始出现，24h时的表达水平最为显著。caspase-3酶活性在皮质酮处理12h时明显升高，以24h时最为显著。Caspase-3抑制剂DEVD-CHO对经皮质酮处理12、24及48h的Leydig细胞中的caspase-3酶活性均具有明显的抑制作用，加caspase-3抑制剂的处理组其细胞基因组DNA电泳未见有凋亡特征性的梯状条带。结论 皮质酮诱导的大鼠Leydig细胞凋亡是一经caspase-3激活的过程。     

Involvement of nuclear factor-kappa B on corticosterone- induced rat Leydig cell apoptosis

Aim:To investigate the activation of nuclear factor-kappa B(NF-kappa B)and its function in glucocorticoid-inducedLeydig cell apoptosis.Methods:The Leydig cells were isolated from male Sprague-Dawley rats(90 days of age)andwere incubated with corticosterone(CORT,glucocorticoid in rat)for 6 h,12 h and 24 h,respectively.The P65subunit of NF-kappa B(NF-kappa B/P65)in nuclei and the inhibitor of NF-kappa B(Ikappa B)in cytoplasm wereanalyzed by Western-blotting.The Leydig cells were treated with anti-Fas antibody for 3 h followed by Westernblotting to assay the changes of NF-kappa B/P65 in nuclei and in cytoplasm.The role of NF-kappa B in CORT-induced Leydig cell apoptosis was evaluated by observing the effects of NF-kappa B/P65 overexpression and inhibit-ing activation of NF-kappa B by 100μmol/L Pyrrolidine dithiocarbamate(PDTC)on this apoptosis.Results:Thetreatment of Leydig cells with CORT increased the levels of NF-kappa B/P65 in nuclei and decreased the levels ofIkappa B in cytoplasm.Following the Leydig cells were treated with anti-Fas antibody,the levels of NF-kappaB/P65was increased in nuclei and decreased in cytoplasm.The CORT-induced Leydig cell apoptosis was inhibited byoverexpressed NF-kappaB/P65 and was enhanced by incubation with PDTC.Conclusion:NF-kappa B is activatedby increased FasL/Fas in CORT-induced Leydig cell apoptosis.NF-kappa B may play an anti-apoptotic role in thisapoptosis.(Asian J Androl 2006 Nov;8:693-702)     

Erratum

To evaluate the effect of estrogen and androgen levels on erythrocyte deformability in endocrinological erectile dysfunction patients. Methods: The estrogen level, androgen level, IR of 30 psychogenic and 15 endocriological ED were studied and the correlation between the estrogen and androgen levels and RI were analyzed. Results: There is a negative correlation between the androgen and estrogen levels and IR;     

Decreased steroidogenesis and cAMP production in vitro by leydig cells isolated from rats made hypothyroid during adulthood

The Leydig cell function of adult male rats made hypothyroid with 6-propyl-2-thiouracil (6-PT, 0.1% w/v in drinking water for 1 month) was studied and compared with that of age-matched controls. After 6-PT treatment, a slight, non-significant decrease in serum testosterone was observed, but no changes in testis weight or number of Leydig cells were noted. The in vitro function of Leydig cells was therefore investigated during incubation for 3 h in the presence or absence of several stimuli: LH (30 mIU/mL), forskolin (FK 1 microM), isobutylmethylxanthine (IBMX, 100 microM), GnRH (100 nM) or FK 1 microM + IBMX 100 microM. Irrespective of the stimulus, cells from hypothyroid rats secreted less cyclic AMP, 17-hydroxyprogesterone, androstenedione and testosterone. No differences in LH receptors were noted between the groups. Prolonged incubation with triiodothyronine (5-250 ng/mL) or thyroxine (5-250 ng/mL) for 3, 16, 24 or 48 h did not affect testosterone secretion in either group; however, administration of IGF-I (8 ng/mL for 24 h) resulted in increased spontaneous and stimulated testosterone production in both groups. However, when hypothyroid animals were supplemented in vivo with thyroxine a full recovery of Leydig cell function in vitro was noted. In conclusion: (1) Leydig cells from rats made hypothyroid during adulthood produce less testosterone in vitro, both spontaneously and in response to cAMP and non-cAMP-mediated stimuli; (2) this is due to a reduction in cAMP production and in the activity of the enzymes in the androgen biosynthetic pathway, and not to changes in LH receptors; (3) direct administration of thyroid hormones did not improve testosterone secretion in either group, while incubation with IGF-I did.     

Effects of varicocele on testosterone, apoptosis and expression of StAR mRNA in rat Leydig cells

The aim of this study was to explore the effects of varicocele on the morphology and function of Leydig cells in the rat testis. Forty male Sprague-Dawley rats were divided into two groups: the experimental group underwent surgery to create a left varicocele (VC), and the control group underwent a sham operation. Serum testosterone and intratesticular testosterone levels were measured using a radioimmunoassay after 4 and 8 weeks of operation. Leydig cells were studied for apoptosis and expression of steroidogenetic acute regulatory (StAR) protein mRNA levels. Serum testosterone levels declined after 4 and 8 weeks of operation but were not significant (P>0.05). However, the intratesticular testosterone levels after 8 weeks were significantly decreased compared with the control group (P<0.01). The mean apoptosis index of Leydig cells in the experimental group was significantly higher than that in the control group after 4 or 8 weeks (P<0.01). StAR mRNA levels in the Leydig cells of the experimental group were significantly lower compared to those of the control group (P<0.01). Our data show that varicocele did impair Leydig cell function by increasing apoptosis and suppressing the expression of the StAR protein.     

Prenatal exposure to dexamethasone alters Leydig cell steroidogenic capacity in immature and adult rats.

This study examines the effects of prenatal exposure to dexamethasone (DEX) on postnatal testosterone production in male rats. Pregnant female rats were treated on gestation days 14-19 with DEX (100 microg/kg body weight per day; n = 9) or vehicle (n = 9). Results show that 35-day-old male offspring from DEX-treated pregnant females (n = 42) had decreased levels of serum testosterone (45.6% lower, P < .05) compared with control offspring (n = 43), although serum luteinizing hormone (LH) levels were not significantly altered. These findings suggest that a direct programming of developing gonadal cells occurs in response to high levels of maternal glucocorticoid. Indeed, testosterone production was significantly reduced in Leydig cells isolated from immature offspring of DEX-treated pregnant females compared with controls (48.3%, P < .001), and LH stimulation of these cells did not compensate for the lowered steroidogenic capacity. The hypothalamic-pituitary-adrenal axis was also affected, because significant reductions in both serum adrenocorticotropic hormone (ACTH; 26.2%, P < .001) and corticosterone (CORT; 32.3%, P < .001) were measured in DEX-exposed immature male offspring. In contrast, adult male offspring from DEX-treated dams had significantly higher levels of serum ACTH (39.2%, P <. 001) and CORT (37.8%, P < .001). These same animals had higher serum testosterone (31.6%, P < or = .05) and a significant reduction in serum LH (30.8%, P < .001). Moreover, Leydig cells isolated from these adult offspring exhibited an increased capacity for testosterone biosynthesis under basal (38.6%, P < .001) and LH-stimulated conditions (33.5%, P < .001). In summary, sustained changes in steroidogenic capacity were observed in male rats exposed to high levels of glucocorticoid during prenatal development. More specifically, DEX exposure in utero perturbed Leydig cell testosterone production in both pubertal and adult rats.     

二氢青蒿素对胰腺癌生长的抑制作用

目的 探讨二氢青蒿素在体外、体内对胰腺癌的生长抑制作用.方法 通过MTT法检测二氢青蒿素对胰腺癌细胞株生长的抑制作用,Annexin V-FITC/PI染色流式细胞术检测细胞凋亡;Western blot检测BxPC-3细胞中增殖、凋亡相关蛋白的表达.监测给药后胰腺癌裸鼠移植瘤体积的变化,并通过对肿瘤组织标本Ki-67染色和TUNEL染色检测肿瘤细胞增殖和凋亡情况.结果 MTT结果显示,二氢青蒿素可抑制体外培养的胰腺癌细胞BxPC-3和AsPC-1的增殖,诱导细胞凋亡,且呈剂量依赖性.二氢青蒿素亦可通过抑制增殖和诱导凋亡而抑制胰腺癌的体内生长.Western blot检测BxPC-3细胞中蛋白的表达水平,结果显示二氢青蒿素上调增殖相关蛋白p21WAF1、下调PCNA的表达;上调凋亡相关蛋白Bax、下调Bcl-2的表达,且可增加caspase-9的活化水平.结论 二氢青蒿素在体内外对胰腺癌均有抗肿瘤作用,是胰腺癌治疗的潜在药物.     

P38MAPK信号通路抑制剂SB203580对肝细胞缺氧再灌注影响的实验研究

目的 观察P38MAPK抑制剂SB203580在人肝癌细胞系HepG2细胞和人正常细胞系L02细胞缺氧再灌注过程中的作用.方法 实验分为正常对照组、缺氧对照组、SB203580＋正常培养组、SB203580＋缺氧培养组,缺氧培养24h、复氧1h后,分别应用Western Blot、MTT、划痕实验、Transwell实验、AnnexinV-FITC/PI双染实验检测细胞中P38蛋白表达情况和细胞增殖、迁移、侵袭和凋亡的变化.结果 与正常对照组相比,缺氧培养组HepG2细胞的凋亡率、P38MAPK磷酸化水平增加;SB203580＋缺氧培养组HepG2细胞的凋亡率相对于缺氧培养组增加,P38MAPK磷酸化水平降低;划痕实验48 h后,缺氧对照组HepG2细胞明显向划痕的中央迁移,而SB203580＋缺氧培养组HepG2细胞迁移受到抑制;侵袭实验24h后,SB203580＋缺氧培养组HepG2细胞的侵袭能力较缺氧对照组降低（P＜0.05）;AnnexinV-FITC/PI双染实验显示与缺氧对照组相比,L02细胞SB203580＋缺氧培养组凋亡率降低,而HepG2细胞SB203580＋缺氧培养组凋亡率则增加（P ＜0.01）;MTT结果显示实验组HepG2细胞和LO2细胞增殖抑制率受到影响,并出现时间浓度依赖关系.结论 SB203580通过特异性阻断P38MAPK信号转导通路,对缺氧培养的正常肝细胞LO2细胞产生保护作用,同时对缺氧培养的肝癌细胞HepG2具有促进凋亡、抑制增殖以及抑制细胞迁移和降低侵袭能力的作用.     

皮质酮诱导的Leydig细胞凋亡中Egr的表达及其作用

目的评价皮质酮是否通过活化钙调神经磷酸酶（CaN）来诱导Leydig细胞中转录因子Egr-2及Egr-3的表达及两种Egr对皮质酮诱导的Leydig细胞凋亡的影响。方法采用RT-PCR方法检测CaN的抑制剂环孢菌素A（CsA）对经皮质酮处理的Leydig细胞中Egr-2及Egr-3的mRNA表达水平；用Annexin-V-FITC和PI双标法评价Egr-2和Egr-3的表达载体对Leydig细胞凋亡率的影响。结果皮质酮能诱导Leydig细胞中Egr-2及Egr-3的表达，且该诱导作用可被CsA抑制；过量表达的Egr-2及Egr-3均可使经皮质酮诱导的Leydig细胞凋亡率增加，并以Egr-3的作用较为显著。结论高浓度的皮质酮可通过活化CaN诱导Leydig细胞中Egr．2和Egr．3的表达，两种Egr对经皮质酮诱导的Leydig细胞的凋亡均具有促进作用。     

17 beta-estradiol inhibition of Leydig cell regeneration in the ethane dimethylsulfonate-treated mature rat.

This study was designed to determine the effects of 17 beta-estradiol (E2) on Leydig cell development in the rat. Mature (60 to 65 days old) male rats received a single intraperitoneal injection of ethane dimethylsulfonate (EDS, 100 mg/kg body weight); untreated rats served as controls. In one series of experiments, groups of EDS-treated rats also received daily injections of either E2 (25 micrograms/100 g body weight), human chorionic gonadotropin (hCG, 20 IU/day), a combination of the two, or vehicle only (EDS controls). Animals were killed on days 2, 4, 10, 16, 24, 30, and 36 after EDS treatment. In another series of experiments, groups of EDS-treated rats received daily injections of hCG and E2 during days 0 through 5, 5 through 30, or 16 through 30 after EDS treatment, and were killed on day 30. In both series of experiments, the steroidogenic capacity and hCG binding capacity of the Leydig cells were examined in short-term in vitro incubations using collagenase-dispersed interstitial cells. Testes were also prepared and examined histologically by light and electron microscopy. E2 treatment of animals during the initial 5 days after EDS administration had no effect on the regeneration of interstitial cells and Leydig cells. Treatment with E2 during days 5 to 30 post-EDS blocked the regeneration of Leydig cells and thereby significantly reduced the increase in interstitial cell numbers. Finally, when E2 treatment was delayed until 16 days post-EDS, there was no significant reduction in the regeneration of interstitial or Leydig cells. These data suggest that an important developmental process that is necessary for Leydig cell regeneration occurs between days 5 and 16 post-EDS.(ABSTRACT TRUNCATED AT 250 WORDS)