N-硝基-L-精氨酸甲酯对大鼠实验性隐睾生精细胞凋亡及Bax表达的影响

目的研究N-硝基-L-精氨酸甲酯(L-NAME)对大鼠实验性隐睾生精细胞凋亡及凋亡相关基因Bax表达的影响。方法手术建立大鼠单侧隐睾模型,实验分为隐睾组、隐睾 L-NAME组[术后腹腔注射溶于生理盐水的L-NAME 50 mg/(kg.d)]、隐睾 生理盐水组、假手术组,7 d后采用化学比色法检测睾丸组织NOS活性,流式细胞术(FCM)和原位缺口末端标记法(TUNEL)检测生精细胞凋亡,RT-PCR和免疫组织化学法检测Bax基因表达变化。结果隐睾组和隐睾 生理盐水组之间检测指标无明显差异;隐睾 L-NAME组隐睾凋亡细胞百分比和生精细胞凋亡指数均低于隐睾组及隐睾 生理盐水组,高于假手术组(P<0.05);Bax mRNA和蛋白表达水平隐睾组及隐睾 生理盐水组最高,假手术组次之,隐睾 L-NAME组最低。结论隐睾生精细胞Bax表达增加,L-NAME可通过下调Bax的表达抑制隐睾生精细胞凋亡。     

N-硝基-L-精氨酸甲酯对氧惊厥发作的影响

目的:探讨一氧化氮合酶(NOS)抑制剂N-硝基-L-精氨酸甲酯(L-NAME)对脑型氧中毒(氧惊厥)发作的影响。方法:SD大鼠40只,分为氧惊厥组、生理盐水对照组、L-NAME组和氮-氧混合气组。L-NAME在高压氧暴露前10min腹腔注射用药。观察各组大鼠氧惊厥始发时间(ITC)、惊厥严重程度(SOC)和存活时间(ST)。结果:氧惊厥和生理盐水对照组的ITC、SOC、ST无显著性差异,应用L-NAME后,ITC、ST明显延长(P<0.01),SOC显著下降(P<0.01)。结论:高压氧暴露前10min应用L-NAME可延长氧惊厥发作的潜伏期和生存时间,降低氧惊厥的严重程度,具有预防氧惊厥的作用。     

L-NAME对实验性大鼠隐睾生精细胞凋亡的抑制作用

目的：研究N-硝基-L-精氨酸甲酯(L-NAME)对实验性大鼠隐睾生精细胞凋亡的影响及其作用机制。方法：36只雄性SD大鼠戊巴比妥钠(50mg／kg)腹腔内注射麻醉，下腹正中切口切断左侧睾丸引带，用5-0尼龙线将左侧睾丸固定于腹后壁，切断右侧睾丸引带后将右侧睾丸还纳入阴囊后关腹，术后第一天将大鼠随机分为两组，     

硝普钠和L-硝基-精氨酸甲酯对大鼠睾丸生精细胞凋亡的影响

目的　观察硝普钠 (SNP)和L 硝基 精氨酸甲酯 (L NAME)对大鼠睾丸生精细胞凋亡的影响。　方法雄性SD大鼠分为 4组 :分别于腹腔内注射SNP、L NAME、SNP +L NAME与生理盐水 ,每天 1次 ,共 12d。颈动脉放血处死动物。流式细胞术 (FCM)结合TUNEL法检测睾丸生精细胞的凋亡。　结果　FCM结合TUNEL法检测发现SNP组亚单倍体峰 (AP峰 )及凋亡指数 (AI)较生理盐水组明显增高 ,L NAME组AP峰及AI显著低于生理盐水组 (P<0 0 1)。SNP和L NAME混合注射时 ,AP峰及AI与生理盐水组无显著性差异 (P >0 0 5 )。　结论　SNP促进生精细胞的凋亡 ,L NAME抑制生精细胞的凋亡 ,其对生精细胞凋亡的调节可能是由一氧化氮 (NO)介导的。     

Nω-硝基-L-精氨酸甲酯对大鼠严重烫伤血脑屏障损伤及脑水肿的影响

目的:观察Nω-硝基-L-精氨酸甲酯(L-NAME)对严重烫伤大鼠血脑屏障(BBB)通透性的影响,探讨NO在严重烫伤中损害BBB的机制.方法:实验SD大鼠分为烫伤组和L-NAME组后又各分为烫伤后1～24 h等5组.检测大鼠脑组织伊文蓝(EB)含量及脑百分含水量;观察BBB超微结构变化.结果:烫伤组与正常组比较,脑EB含量明显增高,具有显著差异.用L-NAME治疗后在1～12 h均可见明显的疗效,脑EB含量与同组比较明显下降.电镜显示:烫伤3 h以后脑毛细血管周围出现空泡化,内皮肿胀,基底膜厚薄不均;经L-NAME治疗后脑毛细胞血管基本恢复到正常.结论:严重烫伤可导致脑BBB通透性增高,早期应用L-NAME能明显减轻烫伤对脑BBB的损伤.     

一氧化氮合酶抑制剂N-硝基-L-精氨酸甲基酯对大鼠淋巴管运动的影响

目的研究一氧化氮合酶抑制剂N-硝基-L-精氨酸甲基酯对大鼠肠系膜淋巴管作用。方法股静脉注入N-硝基-L-精氨酸甲基酯,活体动态观察其2小时内淋巴管运动的变化;用HE染色、酶组化染色研究其组织学改变。结果发现静注N-硝基-L-精氨酸甲基酯可使淋巴管管径变细,运动频率增加,L-精氨酸可暂时逆转该变化;镜下发现淋巴管内皮细胞及管壁上神经纤维都含有一氧化氮合酶阳性反应产物,注N-硝基-L-精氨酸甲基酯组小肠组织HE切片出现固有层中央乳糜管扩张。说明一氧化氮合酶在淋巴循环中可能具有重要调节作用。     

硝普钠和Nω-单硝基-L-精氨酸甲酯对去垂体大鼠生精能力的影响

目的：阐明NO对大鼠生精能力的影响及其作用机制。方法：大鼠去垂体,观察硝普钠(SNP)与N^ω-单硝基-L-精氨酸甲酯(L-NAME)对大鼠生精功能的直接作用。用放射免疫法测定血清内卵泡刺激素(FSH)、黄体生成素(LH)及睾酮含量;Greiss法测定血清NOx^-含量;流式细胞术测定各生精细胞DNA含量,计算1c、2c及4c期细胞的百分比。结果：各组血清LH、FSH和睾酮浓度显著下降或逐渐下降。SNP组血清内NOx^-含量显著升高,L—NAME组血清内NOx^-含量显著降低;对照组、L-NAME组和SNP L—NAME混合注射组的细胞分布均以1c为主;而SNP组的细胞以2c为主。结论：体内高浓度的糖皮质激素可屏蔽LH和FSH的分泌,进一步抑制睾酮的分泌;SNP与L-NAME可通过NO直接影响生精细胞的凋亡。     

N-硝基-L-精氨酸甲酯对大鼠后肢动脉生成过程中eNOS、Ki67及CD11b表达的影响

目的:探讨大鼠后肢动脉生成过程中eNOS、Ki67和CD11b的表达及N-硝基-L-精氨酸甲酯(L-NAME)对eNOS、Ki67、CD11b表达的影响.方法:18只健康SD大鼠,雌雄性各半,随机分为假手术组、模型组及L-NAME组.模型组采用大鼠股动脉结扎诱导动脉生成动物模型;L-NAME组在建立动物模型的基础上,腹腔注射L-NAME干预;假手术组除不结扎股动脉外,其他步骤与模型组相同.采用免疫荧光组织化学显色法检测血管eNOS、Ki67、CD11b等蛋白的表达.结果:eNOS主要表达于血管内皮细胞,Ki67、CD11b可表达于血管壁各层,模型组eNOS、Ki67、CD11b免疫荧光强度较假手术组强,L-NAME组eNOS、Ki67、CD11b免疫荧光强度较模型组弱.结论:在大鼠后肢动脉生成过程中,eNOS、Ki67和CD11b蛋白在血管大量表达,L-NAME可下调eNOS的表达并抑制血管增殖及巨噬细胞的浸润.     

SB203580对大鼠隐睾生精细胞凋亡的影响

目的:探讨SB203580对大鼠隐睾所致生精细胞凋亡过程中的作用。方法:雄性SD大鼠随机分为隐睾组、隐睾 SB203580(p38MAPK抑制剂)组、隐睾 溶媒组、假手术组。复制单侧隐睾模型,术后分别腹腔内注射溶媒或SB203580。7 d后快速断颈椎处死,TUNEL原位末端标记法结合流式细胞术(FCM)检测各组右侧睾丸生精细胞凋亡,免疫组化观察各组右侧睾丸磷酸化p38MAPK蛋白表达的变化。结果:FCM及TUNEL法均显示隐睾 SB203580组生精细胞凋亡指数低于隐睾组和隐睾 溶媒组,同时磷酸化p38MAPK蛋白表达减少。结论:SB203580能降低p-p38MAPK蛋白表达并抑制热压诱导的生精细胞凋亡。     

Sperm retrieval techniques in rats with suppressed spermatogenesis by experimental cryptorchidism

OBJECTIVE: Our aim was to assess the suppression of spermatogenesis and sperm retrieval rate after testicular sperm extraction (TESE) or testicular sperm aspiration (TESA) in adult rats with surgically induced cryptorchidism. METHODS: Adult rats were submitted to TESE and TESA procedures after 15 days of induced cryptorchidism. After spermatozoa retrieval, the testicles were extracted, weighed and a morphological analysis by conventional light microscopy was done. The numbers of spermatozoa retrieved in both TESA and TESE were rated and compared. RESULTS: Histological analysis of the testicles revealed Sertoli cell-only syndrome in 60% of the testicles, and maturation arrest in the remaining cryptorchid testicles. Significant differences were seen in the number of spermatozoa retrieved (P < 0.05) between cryptorchidic and control rats. When sperm retrieval techniques were compared, no differences were detected in the number of spermatozoa obtained (P > 0.05). CONCLUSIONS: It seems that a 15 day period of cryptorchidism is enough to induce spermatogenesis disorders. No differences were detected in the number of spermatozoa retrieved in the right or left testicles, irrespective of the testicular pole. Furthermore, and even more importantly, no differences in the retrieval rate were seen between the two techniques.     

Single administration of butylparaben induces spermatogenic cell apoptosis in prepubertal rats

Parabens are p-hydroxybenzoic acid ester compounds widely used as preservatives in foods, cosmetics, toiletries and pharmaceuticals. Some parabens, including butylparaben, exert an estrogenic activity as determined by in vitro estrogen receptor assay and in vivo uterotrophic assay, and adversely affect endocrine secretion and male reproductive function. We conducted a research study to evaluate the acute effects of butylparaben on testicular tissues of prepubertal rats. Three-week-old male rats (n = 8) were given a single dose of 1000 mg/kg butylparaben. The rats were sacrificed under anesthesia at 3, 6 and 24 h after administration, and their testes were collected for histopathological examination. The study revealed progressive detachment and sloughing of spermatogenic cells into the lumen of the seminiferous tubules at 3 h, and this effect was enhanced at 6 h after administration. Thin seminiferous epithelia and wide tubular lumina were seen at 24 h in the butylparaben-treated group, compared to the control. In order to clarify whether sloughed spermatogenic cells underwent apoptosis, TUNEL assay was carried out. We found a significant increase in the number of apoptotic spermatogenic cells in all the treated groups, compared to the controls and a maximal number of apoptotic cells were detected at 6 h after administration. In semithin sections, apoptotic cells were easily detected by their prominent basophilia and condensed chromatin, mainly found in spermatocytes. Ultrastructurally, the condensed chromatin and shrunken cytoplasm and nucleus, hallmarks of apoptotic cell death, were observed in butylparaben-treated groups. These observations lead us to postulate that butylparaben, similar to other estrogenic compounds, also induces spermatogenic cell apoptosis.     

Effects of aminoguanidine and L-NAME on histamine-induced blood pressure drop in the rat

Mean arterial blood pressure changes in response to i.v. administration of histamine were monitored in the anaesthetized rat in the absence or presence of the diamine oxidase (DAO) inhibitor aminoguanidine (AMG, 10 mg kg^?1). AMG prolonged the duration of the transient drop in blood pressure induced by a bolus injection of histamine (0.05 mg kg^?1) by 34%. In animals pretreated with AMG, no potentiation of the decrease in pressure in response to a 10 min infusion of histamine was observed. However, when infusion was stopped, the time needed for pressure recovery was twice as long in animals treated with AMG as in controls. Blood samples were taken prior to infusion and during the recovery phase and the quantities of histamine were determined by liquid chromatography. The prolonged recovery phase observed in animals pretreated with AMG was associated with five times higher levels of histamine. The duration of histamine-induced hypotension (0.01 mg kg^?1) was 50% shorter in the presence of the nitric oxide synthase inhibitor L -NAME (10 mg kg^?1). We suggest that DAO, through elimination of histamine from the bloodstream, is important for the recovery from histamine-induced hypotension, and that the duration of histamine-induced pressure drop is influenced by formation of nitric oxide.     

低剂量棉酚与甾体激素联合用药对大鼠生精细胞凋亡的影响

采用检测细胞凋亡的微核 (MN)、原位末端标记检测 (ISEL)及单细胞凝胶电泳 (SCGE)等方法 ,研究低剂量棉酚与激素组合用药对大鼠生精细胞凋亡的影响 ,分析不同剂量棉酚 (高剂量 10 0mg/(kg·d)、中剂量 6 0mg/(kg·d)及低剂量 12mg/(kg·d) )及维持量棉酚对生精细胞凋亡及DNA的作用。结果表明不同剂量的棉酚G与单剂量激素H(甲基睾丸素 2 0mg/(kg·d)、炔雌醇 10 0 (g/(kg·d) )合并用药 6周后 ,生精细胞的凋亡系数和微核率随棉酚剂量的增大而增加。高和中剂量棉酚组的凋亡系数分别为 3 5 2 0± 0 4 6 5和 2 0 2 0± 0 0 17,显著高于对照组 (0 2 33± 0 0 88)(P <0 0 5 )。微核率在高剂量组为 2 7 6 33± 3 75 5 ,显著高于对照组 (5 6 0 0± 1 137) (P <0 0 5 ) ;而低剂量组 (3 5 6 7±0 86 9)反而显著低于对照组。服低剂量棉酚G +H 6周后继续服单独低剂量棉酚 (12mg/(kg·d) ) 12周 ,单细胞凝胶电泳未见生精细胞出现DNA单链断裂的彗星状细胞核 ,与高、中剂量组较多的出现率形成明显对照。结果提示低剂量棉酚与甾体激素组合用药在睾丸生精过程中互为调节 ,棉酚引起的生精细胞凋亡效应可在甾体激素的调节下降低而起保护作用     

褪黑激素对小鼠睾丸生精细胞凋亡和睾丸间质细胞nNOS表达的影响

目的：观察外源性褪黑激素(MT)对睾丸生精细胞凋亡和nNOS表达的影响,探讨MT诱导生精细胞凋亡的可能机制。方法：20d和30d小鼠,应用化学发光法检测血清睾酮浓度,TUNEL测定生精细胞凋亡,免疫组化,SABC法观察nNOS在睾丸中的表达。结果：20d和30d实验组血清中睾酮水平明显下降,TUNEL阳性生精细胞数显著增多,TUNEL阳性生精细胞数与睾酮浓度呈负相关;20d实验小鼠中睾丸间质nNOS阳性细胞数与对照组相比明显减少,且与睾酮浓度呈正相关,而与TUNEL阳性生精细胞数呈负相关。结论：MT具有促进生精细胞凋亡的作用,与睾酮分泌受抑制有关;青春期前MT引起生精细胞凋亡可能还与nNOS表达有关。     

Time response of rat testicular alterations induced by cryptorchidism and orchiopexy

Cryptorchidism is one of the main risk factors for infertility and testicular cancer. Orchiopexy surgery corrects cryptorchidism effects. Different models of cryptorchidism developed in the rat include surgery. We assessed testicular alterations in rats submitted to surgical cryptorchidism and examined their potential for reversibility at different time points in order to verify time dependency effect(s) on the recovery of the undescended testes. Cryptorchidism was induced in 3‐week‐old rats. Animals were euthanized 3, 6 or 11 weeks after surgery to evaluate the morphological progression of cryptorchidism‐induced germinative epithelial alterations. Other groups underwent orchiopexy 3, 5 or 9 weeks after surgical cryptorchidism, before or after puberty. Animals were euthanized 3 or 8 weeks after orchiopexy. Controls underwent sham surgery at the same time points as the surgical groups. Cryptorchid testes showed decreased weight, germinative epithelial degeneration, apoptosis and vacuolation, corresponding to impairment of spermatogenesis and of Sertoli cells. Some tubules has a Sertoli cell‐only pattern and atrophy. The intensity of damage was related to the duration of cryptorchidism. After orchiopexy, spermatogenesis completely recovered only when testicular relocation occurred before puberty and the interval for recovery was extended. These results indicate that age, sexual maturity and extension of germ cell damage were relevant for producing germ cell restoration and normal spermatogenesis. We provide original observations on the time dependency of testicular alterations induced by cryptorchidism and their restoration using morphologic, morphometric and immunohistochemical approaches. It may be useful to study germ cell impairment, progression and recovery in different experimental settings, including exposure to exogenous chemicals.     

白细胞介素1受体拮抗剂对大鼠角膜移植物中细胞凋亡的影响

目的:观察急性排斥反应期大鼠角膜移植物中细胞的凋亡,探讨白细胞介素1受体拮抗剂(IL-1ra)对细胞凋亡的影响。方法:建立大鼠穿透性角膜移植模型。实验分为4组,即正常Wistar大鼠(A组),同基因(Wistar大鼠→Wistar大鼠)角膜移植组(B组),同种异体(Wistar大鼠→SD大鼠)角膜移植生理盐水处理组(C组)及IL-1ra治疗组(D组)。应用TUNEL染色法,检测术后7、10及14d各组角膜植片中细胞的凋亡,并对染色结果采用全自动图像分析系统处理后,计算阳性单位(PU)值。制备正常角膜及角膜植片的电镜标本,在透射电镜下观察角膜植片中细胞超微结构的变化。结果:(1)C组角膜植片的平均存活时间为(10.38±1.85)d;而D组为(13.56±1.94)d,二者差异具有统计学意义(P<0.01)。(2)与正常角膜和未发生排斥反应的角膜植片相比较,发生排斥反应的角膜植片中细胞的超微结构改变主要表现为细胞凋亡和细胞坏死共存的特征。(3)正常角膜上皮细胞层中仅有极少数凋亡的细胞,基质层及内皮细胞层中几乎无凋亡的细胞。术后1周,B、C和D组的角膜植片中的各层中均可见散在的细胞凋亡,各组的平均PU值差异无统计学意义(P>0.05)。急性排斥期C组和D组的角膜植片在伤口附近及中央区凋亡的细胞均明显增多,尤其是C组,凋亡的细胞主要集中在上皮细胞基底层及浅层基质。结论:在角膜移植免疫排斥反应中细胞凋亡发挥着重要作用。IL-1ra可通过抑制角膜植片中细胞的凋亡进而抑制角膜移植免疫排斥反应,而延长植片的存活时间。     

L-NAME对沙土鼠脑缺血再灌流后海马区星形胶质细胞活化的作用机制研究

目的 探讨一氧化氮合酶(NOS)抑制剂左旋硝基精氨酸甲酯(L-NAME)对沙土鼠脑缺血再灌注后海马区胶质纤维酸性蛋白(GFAP)合成的影响。方法 钳夹沙土鼠的双侧颈总动脉制造脑缺血模型，应用免疫荧光法染色，观察GFAP密度的变化及分布。结果 脑缺血再灌流后海马区GFAP合成增加，GFAP阳性细胞主要分布在放射层及分子层，L-NAME能减少海马区GFAP的合成。结论 L-NAME是NOS强有力的抑制剂，L-NAME可能通过抑制NO的产生抑制了GFAP的合成。     

Xanthine oxidase inhibitors suppress testicular germ cell apoptosis induced by experimental cryptorchidism

Apoptotic degeneration of germ cells in cryptorchid testis is associated with an increased level of reactive oxygen species, and may be prevented by antioxidant treatment. The objective of this study was to investigate whether xanthine oxidase inhibitors could confer such protection. Unilateral cryptorchidism was surgically induced in the immature rats, which were then left untreated or treated with xanthine oxidase inhibitors, and the testes were evaluated 7 days after the operation. In the control group, the weight of the cryptorchid testis was decreased to 47% of that of the contralateral scrotal testis. However, administration of a xanthine oxidase inhibitor allopurinol (> or = 1 mg/kg/day) significantly attenuated weight reduction of the cryptorchid testis (68-77% of the contralateral scrotal testis, P < 0.01 versus control). Another highly specific xanthine oxidase inhibitor, BOF-4272, also attenuated cryptorchidism-induced testis regression. The effects of allopurinol were associated with decreased apoptosis in germ cells as evaluated by in-situ staining of fragmented DNA. In testicular cells cultured at 37 degrees C, either allopurinol or BOF-4272 prevented DNA cleavage characteristic of apoptosis. In conclusion, xanthine oxidase inhibitors can inhibit germ cell apoptosis induced by experimental cryptorchidism, and may be considered for treatment of male infertility associated with heat stress.