中电导型钙依赖钾通道在糖尿病大鼠阴茎海绵体中的表达及意义

目的 探讨糖尿病对大鼠阴茎海绵体中电导钙激活性钾通道蛋白(IKCa)表达的影响以及意义.方法 选用SD雄性大鼠35只,随机选择25只用于制作糖尿病动物模型,剩余10只用于空白对照.造模成功后饲养8周,注射阿朴吗啡(APO),观察大鼠阴茎勃起,同时采用逆转录-聚合酶链反应(RT-PCR)和Western blot技术检测IKCa mRNA和蛋白在大鼠海绵体的表达.结果 DM组大鼠阴茎勃起和IKCa mRNA及蛋白表达均显著低于STZ组和NDM组(P<0.05),STZ组和NDM组之间大鼠阴茎勃起情况和IKCa mRNA及蛋白表达无统计学意义(P>0.05).绪论糖尿病可降低大鼠阴茎勃起功能,IKCa在糖尿病大鼠海绵体表达明显降低.糖尿病对大鼠阴茎勃起功能的影响与阴茎海绵体IKCa表达减少密切相关.     

可分泌表达人降钙素基因相关肽重组腺相关病毒对糖尿病大鼠阴茎勃起的作用

目的:探讨重组腺相关病毒(rAAV)介导人降钙素基因相关肽(hCGRP)基因转移在糖尿病大鼠阴茎海绵体平滑肌分泌表达及其对阴茎勃起的作用。方法:建立链佐脲菌素(STZ)糖尿病大鼠模型,随机分为3组,分别将VssHGCMV-hCGRP、VssCMV-GFP和rAAV空病毒液注射于阴茎海绵体。在注射后5 d,采用SMUP-PC型生物信号处理系统检测阴茎背神经电刺激诱发的阴茎勃起反应及海绵体内压(ICP)变化。切取海绵体组织,通过免疫组化技术和激光共聚焦显微镜分别检测hCGRP和绿色荧光蛋白(GFP)表达,以放射免疫法检测组织中环磷酸腺苷(cAMP)和环磷酸鸟苷(cGMP)变化。结果:在VssCMV-GFP转染后5 d,显示阴茎海绵体内几乎所有组织均有广泛的GFP表达,而rAAV空病毒转染的海绵体则无GFP表达。VssHGCMV-hCGRP转染STZ诱导的糖尿病大鼠后5d,电刺激阴茎背神经可诱发明显的阴茎勃起,监测ICP明显增高[(60.5±4.5)mm Hg,1 mm Hg=0.133kPa],而对rAAV空病毒转染的对照组STZ糖尿病大鼠以同样的参数电刺激阴茎背神经则无勃起反应,ICP无明显增加[(22.3±1.3)mm Hg],两组差异有显著性(P<0.01)。免疫组化观察显示在VssHGCMV-hCGRP转染的STZ糖尿病大鼠阴茎海绵体组织中hCGRP表达增强,同时当电刺激阴茎背神经诱发勃起反应时,海绵体内cAMP和cGMP水平均升高,分别为(48.4±6.5)nmol/L和(21.2±13.6)nmol/L,较rAAV空病毒组[(16.7±2.5)nmol/L和(0.42±0.12)nmol/L]明显增高(P<0.01)。结论:经阴茎海绵体内注射重组腺相关病毒VssHGCMV-hCGRP在糖尿病大鼠阴茎海绵体内获得了hCGRP转基因高效表达,其可增加阴茎背神经电刺激诱发的阴茎ICP和勃起反应。     

糖尿病大鼠阴茎海绵体组织nNOS神经变化的实验研究

目的 :研究糖尿病对大鼠阴茎海绵体组织nNOS神经纤维的影响。　方法 :34只SD雄性大鼠分为 6周组(17只 )和 8周组 (17只 ) ,其中每组各 11只腹腔注射链脲佐菌素制备糖尿病模型 ,其余各 6只注射柠檬酸缓冲液作为空白对照 ,分别在注射 6周和 8周后 ,观察大鼠阴茎勃起功能 ,并采用免疫组化SP法检测糖尿病大鼠阴茎组织nNOS神经纤维的数量。　结果 :2组糖尿病大鼠的阴茎勃起率分别为 37.5 %和 14 .3% ,差异有显著性 (P <0 .0 5 ) ,均明显低于对照组 (P <0 .0 1)。 2组糖尿病大鼠nNOS神经纤维的表达明显低于对照组 (P <0 .0 1)。 8周组中糖尿病大鼠阴茎的勃起功能和nNOS神经纤维的数量明显低于 6周组 ,分别为 (37.6 0± 6 .76 )条和 (2 8.0 0±5 .2 9)条 ,即随着病程的延长 ,勃起功能和nNOS神经纤维的表达下降 (P <0 .0 5 ) ,而对照组大鼠nNOS神经纤维的数量差异无显著性 [(83.0 0± 3.2 2 )vs(81.0 0± 3.6 1) ]。　结论 :糖尿病严重影响勃起功能和nNOS神经纤维的表达。糖尿病大鼠阴茎海绵体组织中nNOS神经纤维的数量明显减少 ,且与病程正相关 ,这可能是糖尿病性勃起功能障碍的发病机理之一。     

糖尿病鼠周围神经和阴茎nNOS的表达及其与阴茎勃起功能的关系

目的研究糖尿病(Diabetes mullitus,DM)对大鼠坐骨神经、阴部神经及阴茎海绵体中神经型一氧化氮合酶(nNOS)变化及其与勃起功能障碍的相关性。方法16只雄性大鼠随机分为2组：12周组和16周组各8只,腹腔注射链脲佐菌素制备糖尿病模型：另取8只注射枸檬酸缓冲液作为空白对照,分别在注射12周和16周后,观察大鼠阴茎勃起功能,采用免疫组织化学法检测DM大鼠坐骨神经、阴部神经和阴茎海绵体nNOS阳性神经纤维的表达。结果两组大鼠的阴茎勃起次数明显低于对照组大鼠,(P＜0．01)；16W组大鼠阴茎勃起次数也明显低于12周组大鼠(P＜0．05)。两组DM大鼠nNOS坐骨神经、阴部神经和阴茎海绵体中nNOS的表达明显低于对照组(P＜0．01)；16W糖尿病大鼠nNOS阳性纤维的表达也明显低于12周组。两组大鼠对照组间无显著差异。结论糖尿病性阴茎勃起功能障碍与nNOS阳性神经纤维数量的减少相关,可能为糖尿病性勃起功能障碍的发病机理之一。     

糖尿病性勃起功能障碍大鼠阴茎组织内Cx43和eNOS表达的研究

目的:探讨糖尿病(DM)性勃起功能障碍(DMED)与阴茎组织中连接蛋白(Cx43)和eNOS水平变化的相关性。方法:25只SD大鼠腹腔内注射四氧嘧啶建立DM模型,8周后注射阿朴吗啡观察大鼠阴茎勃起情况,筛选出DMED大鼠,应用RT-PCR和免疫组化法分别检测阴茎组织中Cx43和eNOSmRNA表达水平。结果:DMED大鼠阴茎海绵体组织eNOSmRNA表达(0.155±0.157)与DM组(0.508±0.242)比较,显著降低(P<0.01),而Cx43mRNA则显著升高(0.993±0.157vs0.545±0.138,P<0.01),二者存在负相关性(r=-0.987)。免疫组化检测也显示DMED组大鼠阴茎组织平滑肌细胞中eNOS蛋白表达减少,而Cx43蛋白水平增多。结论:阴茎组织内eNOS水平下降可能是DM患者发生ED的主要机制之一;同时,Cx43的表达呈代偿性增加,其功能有待进一步探讨。     

胰岛素治疗糖尿病大鼠阴茎勃起功能障碍的研究

目的 观察糖尿病(diabetes mellitus,DM)大鼠胰岛素治疗后阴茎勃起功能的情况及阴茎海绵体细胞凋亡、Bcl-2和Bax的基因表达.方法 成年雄性Wistar大鼠50只.腹腔内注射链尿佐菌素(STZ)建立糖尿病动物模型,成模后8周根据阴茎勃起功能的情况,随机分成未行胰岛素治疗组(DM组)和胰岛素治疗组.胰岛素治疗组大鼠治疗16周后,处死并取阴茎海绵体,测血糖及糖化血红蛋白,用原位末端标记法(TUNEL)检测细胞凋亡,western blot方法检测阴茎组织Bcl-2、Bax蛋白的表达.结果 DM组大鼠阴茎勃起功能明显低于胰岛素组.胰岛素组血糖及糖化血红蛋白下降(P＜0.001),DM组大鼠较胰岛素治疗组大鼠阴茎海绵体凋亡细胞数明显增多,Bax表达增强,Bcl-2表达减弱.结论 DM大鼠胰岛素治疗后,阴茎海绵体细胞凋亡率减少,阴茎勃起功能改善,Bcl-2和Bax可能参与了DM大鼠阴茎海绵体细胞凋亡的基因调控.     

高血压大鼠海绵体平滑肌细胞间连接的变化

目的:比较自发性高血压大鼠(SHR)和正常血压大鼠阴茎海绵体平滑肌细胞间连接改变及与阴茎勃起功能的关系。方法:注射阿朴吗啡(APO)观察14周龄SHR(SHR组,n=5)、W istar-Kyoto大鼠(WKY组,n=5)阴茎勃起情况,用透射电镜观察其阴茎海绵体平滑肌细胞间连接超微结构,RT-PCR测定海绵体平滑肌细胞Connexin 43的mRNA表达,免疫组化观察Connexin 43蛋白表达。结果:SHR组大鼠阴茎勃起次数明显低于WKY组(P<0.05),电镜发现SHR组大鼠阴茎海绵体平滑肌细胞间大量胶原纤维增生,Connexin 43蛋白及其mRNA表达较WKY组显著降低(P<0.05)。结论:高血压影响阴茎勃起功能,阴茎海绵体平滑肌细胞间连接的病理改变可能是高血压性勃起功能障碍的发病机制之一。     

P-Erk1/2和P-Akt1在高血压大鼠阴茎海绵体中的表达

目的:了解磷酸化Erk1/2(P-Erk1/2)和磷酸化Akt1(P-Akt1)在自发性高血压大鼠(SHR)和正常血压大鼠阴茎海绵体中的表达及与勃起功能的关系。方法:健康成年雄性SPF级SHR与对照组WKY大鼠各8只,14周龄,体重250～300g。麻醉后颈动脉和海绵体内插管连续监测平均动脉压(MAP)和海绵体内压(ICP),利用电刺激海绵体神经,记录ICP/MAP比值变化;免疫组织化学及RT-PCR技术检测P-Erk1/2和P-Akt1在大鼠阴茎海绵体的表达。结果:3V和5V电刺激海绵体神经后SHR组ICP/MAP比值(0.26±0.06、0.28±0.04)均较WKY组(0.46±0.12、0.76±0.13)显著降低(P均<0.05),P-Erk1、P-Erk2mRNA和P-Erk1/2蛋白的相对表达量在SHR组(0.81±0.05、0.91±0.06、54.22±10.05)较WKY组(0.42±0.04、0.68±0.14、7.05±1.45)显著升高(P均<0.05);P-Akt1mRNA和P-Akt1蛋白的相对表达量在SHR组(0.90±0.05、11.17±2.21)与WKY组(0.92±0.06、10.91±1.86)无显著差异(P均>0.05)。结论:高血压性勃起功能障碍的发生与阴茎海绵体P-Erk1/2的过度表达有关,而与P-Akt1的表达水平无明显相关。     

PDE_5基因特异性siRNA改善糖尿病性勃起功能障碍大鼠勃起功能的研究

目的 探讨人鼠PDEs基因siRNA对糖尿病性勃起功能障碍(DM ED)大鼠PDE,基因表达的抑制作用及其对DMED大鼠勃起功能的影响.方法 构建可同时表达2条针对大鼠PDEs基因的shRNA重组腺病毒;50只雄性SD大鼠随机取10只作为正常对照.其余建立DM ED人鼠模型,随机分为实验组、阴性对照组和空白对照组,分别将重组腺病毒rAd-rPDEs-shRNA、腺病毒rAd-mock及生理盐水注射于阴茎海绵体.注射后第7天,电刺激盆神经测定各组大鼠阴茎海绵体内压(ICP)及平均周围动脉压(MAP),然后取海绵体组织通过RT-PCR和Western blot分别检测PDE5基因mRNA及蛋白的表达.结果 ICP/MAP实验组和正常对照组显著高于阴性对照组和窄白对照组(P＜0.05),实验组和正常对照组间无显著性差异(P＞0.05);RT-PCR和Western blot 分析,实验组大鼠PDE5基因mRNA和蛋白表达均显著低于阴性对照组和空白对照组(P＜0.05).结论 PDE5基因shRNA重组腺病毒转染可以改善DMED大鼠的勃起功能,为DMED治疗方法的探索提供了新的思路.     

人胰岛素样生长因子-1基因治疗老龄大鼠阴茎勃起功能障碍的研究

目的探讨阴茎海绵体内注射胰岛素样生长因子1(hIGF-1)基因提高老年性大鼠阴茎勃起功能的可能性. 方法9月龄＞500g SD大鼠35只随机分为3组,注射PCDB-hIGF-1组15只,注射PCDB组10只,空白对照组10只.注射后1～2 d测量大鼠海绵体内压(ICP),收集大鼠阴茎海绵体组织,Trizol方法提取总RNA,RT-PCR检测hIGF-1基因mRNA的表达,免疫组化检测hIGF-1蛋白表达.结果注射PCDB-hIGF-1组大鼠ICP(149±10)高于注射PCDB空质粒组(93±11)和对照组(89±12),差异有统计学意义(P＜0.05);RT-PCR检测注射PCDB-hIGF-1组检测到392 bp的特异性条带,注射PCDB组和对照组未检测到特异性条带;注射PCDB-hIGF-1组免疫组化均可见hIGF-1蛋白的阳性表达.结论hIGF-1基因在大鼠阴茎海绵体中有大量表达,PCDB质粒介导的海绵体内hIGF-1基因注射可部分恢复老龄大鼠的勃起功能.     

Antifibrogenic role of valproic acid in streptozotocin induced diabetic rat penis

We investigated the therapeutic effects of valproic acid (VPA) on erectile dysfunction and reducing penile fibrosis in streptozocin (STZ)‐induced diabetic rats. Eighteen male rats were divided into three experimental groups (Control, STZ‐DM, STZ‐DM plus VPA) and diabetes was induced by transperitoneal single dose STZ. Eight weeks after, VPA and placebo treatments were given according to groups for 15 days. All rats were anesthetised for the measurement of in vivo erectile response to cavernous nerve stimulation. Afterward penes were evaluated histologically in terms of immune labelling scores of endothelial nitric oxide synthase (eNOS), vascular endothelial growth factor (VEGF) and transforming growth factor‐β1 (TGF‐β1). Slides were also evaluated in terms of collagen/smooth muscle ratio and penile apoptosis. After the treatment with VPA, erectile responses were found as improved when compared with STZ‐DM rats but not statistically meaningful. eNOS and VEGF immune expressions diminished in penile corpora of STZ‐DM rats and improved with VPA treatment. VPA led to decrease in TGF‐β1 expression and collagen content of diabetic rats’ penes. Penile apoptosis was not diminished with VPA. In conclusion, VPA treatment seems to be effective for reducing penile fibrosis in diabetic rats and more prolonged treatment period may enhance erectile functions.     

Dietary resistant maltodextrin ameliorates testicular function and spermatogenesis in streptozotocin–nicotinamide‐induced diabetic rats

This study investigated the effect of resistant maltodextrin (RMD) on reproduction in streptozotocin (STZ)–nicotinamide‐induced type 2 diabetic male rats. Forty male rats were induced with diabetes by a single intraperitoneal injection of STZ (50 mg kg^?1) and nicotinamide (100 mg kg^?1). Five groups were analysed in total: normal, diabetic rats without RMD, diabetic rats with RMD 1.2 g per 100 g diet (1×), with RMD 2.4 g per 100 g (2×), and with RMD 6.0 g per 100 g (5×). The groups of diabetic rats with the RMD supplement, compared to those without supplement, showed improved plasma glucose control, attenuated insulin resistance and recovery of testosterone level and spermatogenesis stage. The STZ–nicotinamide‐induced diabetes mellitus (DM) caused a significant reduction in serum testosterone, testis androgen receptor (AR), steroidogenic acute regulatory protein (StAR) and 3β‐hydroxysteroid dehydrogenase (3β‐HSD) protein, but a statistical recovery in each of these was observed in the 5× group. TUNEL‐positive cells were observed in the diabetic without RMD group, and RMD treatment reduced apoptotic germ cells. The expression of Bax/Bcl2 was induced in the diabetic group and also significantly reduced in the 5× group. Dietary RMD may improve metabolic control in STZ–nicotinamide‐induced diabetic rats and attenuate hyperglycaemia‐related impaired male reproduction and testicular function.     

神经生长因子在糖尿病神经病理性疼痛大鼠脊髓和背根神经节中的表达

目的 观察糖尿病早期大鼠脊髓背角及背根神经节神经生长因子(NGF)的表达及其与疼痛行为的相关性.方法 50只SD大鼠随机分为制模组(DM组,n=40)和止常对照组(C组,n=10).DM组又均分为制模1周组(DM1组)、制模2周组(DM2组)、制模4周组(DM4组)和制模8周组(DM8组)四个亚组.按55 mg/kg一次性腹腔注射链脲佐菌素(STZ)建立糖尿病大鼠模型.在制模前及制模后1、2、4、8周利用von Frey hairs测定大鼠机械痛阈,并通过蛋白质免疫印迹法测定每个时间点大鼠脊髓背角和背根神经节NGF的表达水平.结果 糖尿病大鼠机械痛阈在制模1周后即明显下降,并持续至8周;分别存制模2周和1周后脊髓背角和背根神经节内NGF表达水平出现显著下调,并分别持续至4周和8周.大鼠NGF表达水平与机械痛阈存在正相天性.结论 糖尿病早期大鼠外州及中枢神经系统NGF表达即出现下调,并与疼痛相关.     

糖尿病大鼠痛性周围神经病变时脊髓小胶质细胞的活化

目的 评价糖尿病大鼠痛性周围神经病变时脊髓小胶质细胞的活化.方法 SD大鼠25只,2月龄,雌雄不限,腹腔注射1%链脲佐菌素60 mg/kg以制备大鼠糖尿病痛性周围神经病变模型,取造模成功的10只SD大鼠作为糖尿病痛性周围神经病变组(DM组),另取10只同月龄SD大鼠作为对照组(NC组).分别于注射前(T1)、注射后2、7、14、21、28 d(T2～6)时称量体重,取尾静脉血0.1 ml测定血糖水平,于T1、T3～6时测定大鼠右后爪机械缩足反应阈值;注射28 d时麻醉大鼠,取L4.5脊髓制备切片,采用免疫组化法检测脊髓小胶质细胞补体受体3(CR3)的表达.结果 与NC组比较,DM组T2～6时血糖升高、体重下降,T4～6时机械缩足反应阈值降低,T6时小胶质细胞CR3表达上调(P<0.05或0.01);与T1时比较,DM组T2～6时血糖升高、体重下降,T6时机械缩足反应阈值降低(P<0.01).结论 脊髓小胶质细胞的活化与大鼠糖尿病痛性周围神经病变的发病有关.     

Effects of different n6/n3 ratios and supplementation with DHA and EPA on the testicular histology and lipogenesis in streptozotocin‐treated rats

The aim of this study was to investigate the possible influence of different dietary n6/n3 ratios and DHA/EPA addition on the testis histology, antioxidative status and lipogenesis of streptozotocin‐treated rats. Diabetes was induced by a single intraperitoneal injection of streptozotocin (55 mg/kg). The rats were divided into five groups: CON (n6/n3 ratio, 7), CON‐DM1 (STZ; n6/n3 ratio, 7), N3‐DM1 (STZ; n6/n3 ratio, 1), N6‐DM1 (STZ; n6/n3 ratio, 60) and DHA‐DM1 (STZ; n6/n3 ratio, 1; added DHA/EPA). Antioxidative status was improved in the DHA‐DM1 group. Seminiferous tubule diameter, testicular pathohistological scoring and total lipid content were improved in the N6‐DM1 group compared to the other streptozotocin‐treated groups. Streptozotocin treatment induced changes in testis fatty acid profile depending on n6/n3 ratio. The fatty acid profile of N6‐DM1 group was characterised by similar or increased values for n6 fatty acids compared to the CON group, while the DHA‐DM1 group had the lowest content of n6 fatty acids. The content of n3 fatty acids was increased in the N3‐DM1 and DHA‐DM1 groups. These results suggest that a n6/n3 ratio could significantly influence testicular antioxidative status, histology and lipogenesis and that these effects vary depending on the supplemented fatty acid.     

糖尿病性阴茎勃起功能障碍大鼠模型海绵体超微结构的研究

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制造ＤＭ 动物模型后,注射阿朴吗啡观察６ 周、８ 周及１２ 周大鼠阴茎勃起情况,筛选ＤＭ 性ＥＤ 大鼠模型,观察其阴茎海绵体组织超微结构的改变。　结果　ＤＭ 性ＥＤ 大鼠模型阴茎海绵体内皮细胞及平滑肌细胞超微结构均有明显的病理改变：线粒体退变、内质网扩张,糖原颗粒、吞饮小泡及微丝减少。此外,还可见大量间质组织增生及微血管腔闭塞。随ＤＭ 病程不同,其改变程度不同,８ 周时以内皮细胞损害为明显,１２ 周时以平滑肌细胞损害为明显。　结论　ＤＭ 严重影响阴茎勃起功能,海绵体组织超微结构的病理改变可能是ＤＭ 性ＥＤ 发病机理之一。     

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。     

Exogenously administered adenosine attenuates renal damage in streptozotocin-induced diabetic rats

Background: Diabetic nephropathy (DNP) is one of the most serious complications of diabetes mellitus (DM). In the present study, we investigated the potential of adenosine as a therapeutic candidate for preventing DNP.

Methods: Twenty-one adult male rats were included in the study. Fourteen rats were administered a single dose of 60?mg/kg streptozotocin (STZ) to induce diabetes. Seven rats served as normal control group. Diabetic rats were randomly divided into two groups: one group was treated with 1?mL/kg saline/day (DM?+?saline) and the other group was treated with 5?mg/kg/day adenosine (DM?+?adenosine) for 6?weeks. After 6?weeks, biochemical parameters including urea, creatinine, blood urea nitrogen (BUN), kidney injury molecule-1 (KIM-1) and tumor necrosis factor-α (TNF-α) were measured in plasma samples. Also, kidneys were removed for histopathological assessment.

Results: Both of plasma KIM-1 and TNF-α levels were significantly higher in DM?+?saline group compared to controls. However, treatment of diabetic rats with adenosine significantly decreased the plasma KIM-1 and TNF-α levels compared to DM?+?saline group. Significant histopathological changes were observed in diabetic rats whereas adenosine treatment effectively prevented these changes.

Conclusions: The findings of the present study suggest that adenosine may be a useful therapeutic agent for preventing DNP.