衰老对大鼠阴茎海绵体NOSⅠ的表达和NOS活性的影响

目的 :探讨衰老对大鼠阴茎海绵体一氧化氮合酶Ⅰ (NOSⅠ)mRNA、蛋白的表达和NOS活性的影响。　方法 :30只雄性SD大鼠按不同月龄分为成年组、老年组和衰老组 ,应用Western印迹、RT PCR方法分别检测不同年龄组阴茎海绵体NOSⅠ蛋白及mRNA的表达 ;用紫外分光光度计测定不同年龄组阴茎海绵体NOS的活性。　结果 :成年组NOSⅠ 蛋白的表达量最高 ,老年组和衰老组显著降低 ,分别为成年组的 75 .6 %和 6 1.2 % ;NOSⅠmRNA的表达与蛋白表达的变化一致 ;老年组NOS活性与成年组差异无显著性 (P >0 .0 5 ) ,衰老组NOS活性明显降低 ,是成年组的70 .4 % ,并且差异非常显著 (P <0 .0 1)。　结论 :衰老引起NOSⅠ 蛋白及mRNA的表达降低和NOS活性的显著降低 ,可能是老年性阴茎勃起功能障碍的主要机制之一。     

大鼠阴茎海绵体NOSⅠ、NOSⅢ mRNA和蛋白表达研究

目的 :探讨正常大鼠阴茎海绵体中一氧化氮合酶 (NOS) Ⅰ和 Ⅲ mRNA和蛋白的表达。　方法 :取雄性SD大鼠阴茎组织 ,应用高度特异性抗体经免疫沉淀、Western印迹分析技术分别检测NOSⅠ 、NOS Ⅲ 蛋白的表达 ;RT PCR方法检测NOSⅠ和NOS Ⅲ mRNA的表达。　结果 :正常大鼠阴茎海绵体组织中有NOSⅠ 和NOS ⅢmRNA和蛋白的表达 ,并且NOSⅢmRNA和蛋白的表达明显多于NOSⅠmRNA和蛋白的表达。　结论 :不同的NOS亚型可能共同参与阴茎勃起的调控。     

不同剂量香烟烟雾提取物对阴茎海绵体NOS活性和Cx43蛋白的影响

目的:探讨不同剂量的香烟烟雾提取物对雄性大鼠勃起功能的影响,进一步研究吸烟导致ED的发病机制。方法:75只健康雄性SD大鼠(8周龄),随机分为5组(15只/组)。A组为对照组;B组为皮下注射二甲基亚砜(DMSO)组;C组为皮下注射香烟烟雾提取物(CSE)低剂量组;D组为皮下注射CSE中剂量组;E组为皮下注射CSE高剂量组。应用SD雄性大鼠建立皮下注射CSE模型60 d后,经皮下注射阿朴吗啡(APO)后观察大鼠阴茎勃起情况,处死大鼠取阴茎海绵体组织。一部分标本采用激光共聚焦扫描显微镜方法检测缝隙连接蛋白43(Cx43)的表达;另一部分标本采用比色法测定大鼠阴茎海绵体组织中NOS活性。结果:不同剂量CSE组阴茎勃起次数、阴茎海绵体组织NOS活性和海绵体平滑肌中Cx43表达与DMSO组和对照组比较均明显减少(P<0.05)。实验组中,阴茎勃起次数、阴茎海绵体组织NOS活性和Cx43表达均随着CSE剂量的增加而减少。DMSO组与对照组比较差异无统计学意义(P>0.05)。结论:CSE使阴茎海绵体组织NOS活性明显降低、海绵体平滑肌中Cx43蛋白表达明显减少并且严重影响阴茎勃起功能,并且CSE剂量越大,其影响越明显。提示,Cx43蛋白表达下调、NOS活性降低可能是CSE导致ED的发病机制之一。     

甲状腺素对大鼠阴茎海绵体内NOS及CO含量影响的研究

目的:建立甲状腺功能亢进及甲状腺功能减退Wistar大鼠动物模型,检测其阴茎海绵体内NOS及内源性一氧化碳(CO)的含量,探讨甲状腺素对大鼠勃起功能的影响及内源性CO在阴茎海绵体勃起过程中的作用,进一步讨论甲状腺素对人类勃起功能的影响。方法:将50只3月龄雄性Wistar大鼠随机均分为甲亢组、甲亢治疗组、甲减组、甲减治疗组及正常对照组。用紫外分光光度计分别测定阴茎海绵体内NOS及CO的含量。结果:无论甲状腺素增多及减少都会使大鼠阴茎海绵体NOS含量降低(P<0.01),并且甲减组阴茎海绵体内NOS活性低于甲亢组(P<0.01)。无论甲状腺素增多还是减少都会使大鼠阴茎海绵体内CO含量降低(P<0.01),并且甲亢组阴茎海绵体内CO活性低于甲减组(P<0.01)。在对甲减组及甲亢组进行治疗后其CO及NOS的含量得到提高,与正常对照组无显著差异(P>0.05)。结论:甲状腺功能紊乱情况下阴茎海绵体中NOS和CO的浓度均减低;甲状腺功能紊乱被及时纠正后阴茎海绵体中CO及NOS的含量可恢复到正常水平。在相同条件下甲状腺功能低下对性功能的损害强于甲状腺功能亢进对勃起功能的损害。     

当归对大鼠海绵体神经损伤后阴茎NOS活性的影响

目的 :探讨海绵体神经 (CN)钳夹损伤后阴茎组织一氧化氮合酶 (NOS)活性的变化及局部使用当归注射液对其活性的影响。　方法 :30只SD大鼠随机平均分成当归组、对照组和正常组 ,对前两组大鼠通过手术建立双侧CN钳夹损伤模型 ,经耻骨上在CN周围每天分别注射 2 5 %当归注射液或相当剂量生理盐水 (10ml/kg) ,正常组只注射生理盐水而不行手术 ;两周后用分光光度法检测阴茎组织NOS活性。　结果 :NOS活性均值当归组为 (1.36 0±0 .139)U/mg·prot,达正常组的 77.4% ,而对照组为 (0 .945± 0 .0 98)U/mg·prot,只有正常组的 5 3.8% ,二者比较有极显著性差异 (P <0 .0 1)。　结论 :CN损伤可致阴茎组织NOS活性降低 ,局部使用当归能够一定程度地避免其活性下降。     

糖尿病性阴茎勃起功能障碍大鼠阴茎海绵体一氧化氮合酶活性的测定

目的　探讨糖尿病 (DM )性阴茎勃起功能障碍 (ED)的发病机理。　方法　SD大鼠注射链脲佐菌素建立DM动物模型后 ,注射阿朴吗啡观察 6周、8周及 12周大鼠阻茎勃起情况 ,筛选DM性ED大鼠模型 ,测定其阴茎海绵体组织一氧化氮合酶 (NOS)的活性。　结果　DM性ED大鼠阴茎海绵体组织NOS活性与对照组相比显著降低 (P <0 0 0 1或P <0 0 1) ,随DM病程延长 ,NOS活性明显下降 (P <0 0 1)。　结论　DM严重影响阴茎勃起功能 ,海绵体组织NOS活性降低可能是其发病机理之一。     

高脂血症大鼠阴茎海绵体中VEGF含量变化对NOS的影响

目的 探讨高脂血症引起大鼠阴茎海绵体中血管内皮生长因子(VEGF)含量的变化,以及这种变化对一氧化氮合酶(NOS)活性的影响.方法 将32只雄性wistar大鼠随机分为2组,高脂血症组和正常对照组各16只,通过建立大鼠高脂血症模型,于实验开始后10d和20d,分别随机取各组1/2大鼠观察其勃起功能;采用免疫组化方法观察大鼠阴茎海绵体内VEGF,并用图像分析法测定其含量;用分光光度法分别测定海绵体匀浆中NOS活性.结果 造模开始后第10天,高脂血症组和正常对照组相比,大鼠阴茎海绵体内VEGF含量、NOS活性及阴茎勃起次数差异均有显著性(P＜0.05):造模开始后第20天,两组的VEGF含量、NOS活性及阴茎勃起次数差异均有显著性(P＜0.05).高脂血症组VEGF含量、NOS活性及阴茎勃起次数,第20天均明显低于第10天,两两比较差异均有显著性(P＜0.05).结论 高脂血症大鼠阴茎海绵体中VEGF明显减少,内皮细胞增殖下降,进而NOS下降,这可能是高脂血症引起勃起功能障碍(ED)的发病机制之一.     

2型糖尿病大鼠血浆同型半胱氨酸与阴茎海绵体内NOS和内源性CO的相关性研究

目的:研究2型糖尿病性大鼠血浆同型半胱氨酸(Hcy)与阴茎海绵体内NOS和内源性CO的相关性。方法:选取3月龄雄性Wistar大鼠50只,随机选取10只为对照组(A组);高糖高脂饲料饲养4周后从其他40只大鼠中筛选出30只构建成功的糖尿病(DM)大鼠模型,随机分成3组:DM大鼠组(B组);胰岛素治疗组(C组)和叶酸+维生素B12治疗组(D组)。8周及12周后注射阿朴吗啡观察各组大鼠阴茎勃起情况。12周后测各组大鼠血浆总Hcy含量及阴茎海绵体内NOS活性和CO含量。结果:与A组比较,B组大鼠血浆Hcy浓度明显升高,阴茎勃起功能明显降低,阴茎海绵体NOS活性和CO含量均下降,差异有显著性(P<0.01)。2型DM大鼠中高Hcy血症发生率为55%。与B组比较,C组和D组中大鼠血浆Hcy浓度显著下降,阴茎勃起功能、阴茎海绵体NOS活性均升高(P<0.01),Hcy与NOS(rA=-0.89,rB=-0.76,rC=-0.91,rD=-0.91)及CO含量(rA=-0.82,rB=-0.77,rC=-0.93,rD=-0.81)均呈负相关。结论:2型DM大鼠血浆中的高Hcy可能是引起阴茎海绵体NOS活性下降、CO含量下降,进而导致DM ED发病的分子机制之一。胰岛素、叶酸和维生素B12可以改善DM大鼠的勃起功能,提高阴茎海绵体NOS活性和CO含量。     

高脂血症对大鼠阴茎海绵体中NOS及CO含量影响的研究

目的 探讨高脂血症对大鼠阴茎海绵体中一氧化氮合成酶(NOS)活性及一氧化碳(CO)浓度的影响及两者之间的关系.方法 将32只雄性wistar大鼠随机分为2组:高脂血症组和正常对照组,每组各16只大鼠.通过建立大鼠高脂血症模型,于实验开始后10d和20d,分别随机取各组1/2大鼠观察其勃起功能;用分光光度法分别测定海绵体匀浆中NOS活性和CO浓度.结果 在第10天及第20天,高脂血症组和正常对照组相比,大鼠阴茎海绵体内NOS活性、CO浓度及阴茎勃起次数差异均有统计学意义(P<0.05);而且第20天高脂血症组NOS活性、CO浓度及阴茎勃起次数,均明显低于第10天,两两比较差异均有统计学意义(P<0.05).结论 高脂血症大鼠阴茎海绵体中NOS及CO下降,这可能是高脂血症引起勃起功能障碍的发病机制之一.     

降钙素基因相关肽对糖尿病性ED大鼠阴茎海绵体平滑肌细胞表型转化的影响

目的:了解降钙素基因相关肽(CGRP)对糖尿病性ED大鼠阴茎海绵体平滑肌细胞表型转化的影响。方法:利用链脲佐菌素建立糖尿病及糖尿病性ED大鼠模型。阴茎海绵体平滑肌细胞原代培养,并进行免疫细胞化学染色鉴定。实验分为2组:正常对照组和糖尿病性ED大鼠组。不同浓度(0、10,60,100 nmol/L)CGRP作用24h后,利用qRT-PCR检测各组细胞碱性调宁蛋白和骨桥蛋白mRNA的表达。结果:各组原代培养阴茎海绵体平滑肌细胞α-肌动蛋白阳性细胞率为(95.94±0.03)%。与正常对照组比较,糖尿病组ED大鼠阴茎海绵体平滑肌细胞碱性调宁蛋白mRNA表达显著减少(4.41±0.29 vs 10.35±0.62,P<0.01),而骨桥蛋白mRNA表达水平显著上调(5.28±0.32 vs 1.32±0.24,P<0.01)。当CGRP作用的终浓度为100 nmol/L时,糖尿病组大鼠阴茎海绵体平滑肌细胞经CGRP作用后,与未经其作用相比,碱性调宁蛋白mRNA表达显著上调(6.90±0.22 vs 4.41±0.29,P<0.01),而骨桥蛋白mRNA表达水平显著减少(3.26±0.31 vs 5.28±0.32,P<0.01)。结论:CGRP可使糖尿病性ED大鼠阴茎海绵体平滑肌细胞表型从合成型向收缩型转化。     

Effect of aging on expression of nitric oxide synthase I and activity of nitric oxide synthase in rat penis

Aim: To investigate the effect of aging on the expression of nitric oxide synthase I (NOS I) and the activity of NOS in rat penis. Methods: Sixty male rats from 3 age groups (adult, old and senescent) were investigated. The expression of NOS I protein and mRNA in rat penis were detected by Western blot and RT-PCR respectively and the NOS activity, with ultraviolet spectrophotometry. Results: In the old and senescent group, NOS I protein expression was significantly decreased as compared with the adult. NOS I mRNA expression was well correlated with the protein expression. NOS activity was not statistically different between the adult and old groups, but it was significantly reduced in the senescent compared with the adult group (P<0.01). Conclusion: The aging-induced decreases in NOS I expression and NOS activity may be one of the main mechanisms leading to erectile dysfunction in the senescent rats.     

Histopathological changes and nitric oxide synthase activity in corpus cavernosum from rats with neurogenic erectile dysfunction

OBJECTIVE: To investigate changes in histology and nitric oxide synthase (NOS) activity in cavernosal tissues from rats with neurogenic erectile dysfunction induced experimentally. MATERIALS AND METHODS: Twenty-four adult male Sprague-Dawley rats were divided equally into three groups and underwent a sham operation (control, group 1), unilateral (group 2) or bilateral (group 3) cavernosal nerve resection. Three months later they were killed and the cavernosal tissues analysed histologically by light and transmission electron microscopy, with NOS activity detected using an NADPH-diaphorase staining technique. RESULTS: On light and electron microscopy, while penile nerves and cavernosal smooth muscle cells had a normal morphological appearance in the eight control rats, there were degenerative changes of the myelinated penile nerves and axonal fibrosis in groups 2 and 3. However, these changes were not significant. Using NADPH-diaphorase staining, NOS activity was detected in all three groups in endothelial cells and cavernosal structures. However, the staining was more intense in endothelial cells and cavernosal muscles of rats in group 2 than in the other groups. CONCLUSION: NOS activity was increased in the cavernosal tissue after cavernosal denervation, but the pharmacological action of nitric oxide may be impaired.     

大鼠肝缺血／再灌注后肝组织一氧化氮和不同亚型一氧化氮合酶的变化

目的探讨一氧化氮（NO）和一氧化氮合成酶（NOS）在肝缺血／再灌注（I／R）过程中的变化和作用。方法健康雄性SD大鼠24只，随机分为3组（每组8只）：①正常对照组，术中只分离肝周围韧带，不做肝门阻断及再灌注。②I/R组，进行45min的部分肝门阻断及60min的再灌注。③L-精氨酸（L—Arg）组，缺血前20min经阴茎背静脉注射L—Arg（300mg／kg），余同②组。实验结束后，取下腔静脉血2ml，并迅速切取缺血肝组织。检测血清丙氨酸转氨酶（ALT）、门冬氨酸转氨酶（AST）、乳酸脱氢酶（LDH）；测定肝组织中超氧化物歧化酶（SOD）、丙二醛（MDA）、黄嘌呤氧化酶（XOD）、一氧化氮（NO）和一氧化氯合成酶（NOS）等指标；观察光镜和电镜下肝组织学变化。结果与正常对照组相比，I／R组iNOS升高，NO降低；L-Arg组NO、eNOS均高于I／R组。2、3组比1组大鼠的肝组织病理损害重、肝功能差，L—Arg组病理损害较I／R组明显减轻、肝功能改善。结论NO对大鼠肝I／R损伤具有保护作用．不同亚型NOS的变化参与其中。     

大鼠脊髓损伤后一氧化氮合酶基因表达的变化

目的　探讨大鼠脊髓损伤后3种类型一氧化氮合酶（NOS）mRNA表达的变化规律。方法　成年SD大鼠36只,随机分为种类6组,每组6只大鼠。建立大鼠脊髓压迫伤模型,以逆转录-聚合酶链反应（RT-PCR）法测定伤段脊髓组织神经型（nNOS）、诱导型（iNOS）及内皮型（eNOS）一氧化氮合酶的mRNA表达情况。结果　脊髓压迫伤后nNOSmRNA及NOSRNA表达增强,伤后6h达到高峰0.633±0.012、1.236±0.207;iNOSmRNA表达亦增高,但在伤后24h才达到高峰1.043±0.049。结论　脊髓损伤后NOSmRNA的表达增强,但不同类型的NOSmRNA变化规律不同,增强或抑制不同NOSmRNA的表达可能减轻脊髓继发性损伤。     

Renal cortical nitric oxide synthase activity during maturational growth in the rat

The present study was designed to test the hypothesis that growth from puberty to adulthood in the rat is associated with an increase in renal cortical nitric oxide synthase (NOS) activity that results in an augmented impact of nitric oxide (NO) on hemodynamic function. Two groups of male Sprague-Dawley rats were studied: juvenile rats (approximately 2 months old) and mature rats (approximately 5 months old). NOS activity, measured as -nitro-L-arginine (NNA)-sensitive (3)H-L-citrulline production from (3)H-L-arginine, was significantly higher in the renal cortex of mature rats (57+/-2 pmol/h per mg protein) than in juveniles (42+/-3 pmol/h per mg protein). Additional animals from each group were anesthetized to determine the acute impact of NOS inhibition on arterial pressure and renal cortical blood flow, measured by single-fiber Doppler flowmetry. Cortical blood flow was higher in mature rats than in juveniles, averaging 22+/-2 and 16+/-1 perfusion units, respectively. NOS inhibition (10 mg/kg NNA i.v.) decreased renal cortical blood flow in mature rats by 35+/-7%, but only by 9+/-4% in juvenile animals. These data support the hypothesis that maturational growth in the rat is associated with augmented NOS activity coupled with an increased tonic influence of NO on renal cortical blood flow.     

Effects of icariin on erectile function and expression of nitric oxide synthase isoforms in castrated rats

Aim： To investigate the effect of icariin on erectile function and the expression of nitric oxide synthase （NOS） isoforms in castrated rats. Methods： Thirty-two adult male Wistar rats were randomly divided into one shamoperated group （A） and three castrated groups （B, C and D）. One week after surgery, rats were treated with normal saline （groups A and B） or oral icariin （1 mg/[kg·day] for group C and 5 mg/[kg·day] for group D） for 4 weeks. One week after treatment, the erectile function of the rats was assessed by measuring intracavernosal pressure （ICP） during electrostimulation of the cavernosal nerve. The serum testosterone （ST） levels, the percent of smooth muscle （PSM） in trabecular tissue, and the expression of mRNA and proteins of neuronal nitric oxide synthase （nNOS）, inducible nitric oxide synthase （iNOS）, endothelial nitric oxide synthase （eNOS） and phosphodiesterase V （PDES） in corpus cavernosum （CC） were also evaluated. Results： ICP, PSM, ST and the expression of nNOS, iNOS, eNOS and PDE5 were significantly decreased in group B compared with those in group A （P 〈 0.01）. However, ICE PSM and the expression of nNOS and iNOS were increased in groups C and D compared with those in group B （P 〈 0.05）. Changes in ST and the expression of eNOS and PDE5 were not significant （P 〉 0.05） in groups C and D compared with those in group B. Conclusion： Oral treatment with icariin （〉 98.6 % purity） for 4 weeks potentially improves erectile function. This effect is correlated with an increase in PSM and the expression of certain NOS in the CC of castrated rats. These results suggest that icariin may have a therapeutic effect on erectile dysfunction.     

Gene transfer of endothelial nitric oxide synthase partially restores nitric oxide synthesis and erectile function in streptozotocin diabetic rats

PURPOSE: We determined whether adenoviral gene transfer of endothelial nitric oxide synthase (eNOS) to the penis of streptozotocin induced diabetic rats could improve the impaired erectile response. MATERIALS AND METHODS: Two experimental groups of animals were transfected with adenoviruses, including streptozotocin (Sigma Chemical Company, St. Louis, Missouri) diabetic rats with AdCMVbetagal and streptozotocin diabetic rats with AdCMVeNOS. At 1 to 2 days after transfection these study animals underwent cavernous nerve stimulation to assess erectile function and their responses were compared with those of age matched control rats. In control and transfected streptozotocin diabetic rats eNOS and neuronal NOS (nNOS) were examined by Western blot analysis. Constitutive and inducible NOS activities were evaluated in the presence and absence of calcium by L-arginine to L-citrulline conversion and nitrate plus nitrite levels were measured. In control and streptozotocin diabetic penes beta-galactosidase activity and localization were determined. RESULTS: After transfection with AdCMVbetagal beta-galactosidase was localized to the endothelium and smooth muscle cells of the streptozotocin diabetic rat penis. Streptozotocin diabetic rats had a significant decrease in erectile function, as determined by peak and total intracavernous pressure (area under the curve) after cavernous nerve stimulation compared with control rats. Streptozotocin diabetic rats transfected with AdCMVeNOS had peak intracavernous pressure and area under the curve similar to those in control animals. This change in erectile function was a result of eNOS over expression with an increase in eNOS protein expression and constitutive NOS activity as well as an increase in nitric oxide biosynthesis, as reflected by an increase in cavernous nitrate plus nitrite formation. There was no change in nNOS protein expression or calcium independent conversion of NOS (inducible NOS activity). CONCLUSIONS: Adenoviral gene transfer of eNOS significantly increased peak and total intracavernous pressure to cavernous nerve stimulation in streptozotocin diabetic rats to a value similar to the response observed in control rats. Our results suggest that eNOS contributes significantly to the physiology of penile erection. These data demonstrate that in vivo adenoviral gene transfer of eNOS can physiologically improve erectile function in the streptozotocin diabetic rat.