糖尿病大鼠膀胱纤维化及氧化应激状态的研究
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摘要
研究目的探讨糖尿病膀胱发生纤维化的机制以及氧化应激在糖尿病膀胱进程中的作用,同时观察生长因子抑制剂(太得恩)在改善糖尿病膀胱病变进程中的作用。
研究材料、方法
一建立糖尿病大鼠模型
取30只3月龄(200±10g)雄性Wistar大鼠随机分为3组,分笼饲养。分别为对照组(n=10);糖尿病组(n=10);治疗组(n=10)。建模成功后饲养4周,然后对治疗组大鼠给予太得恩药物(100mg.Kg~(-1).d~(-1),溶于2ml花生油中,胃管注入)治疗4周,对照组及糖尿病组大鼠给予同等剂量花生油赋形剂。以上动物共饲养8周。8周后处死完整切取膀胱组织。
二糖尿病膀胱中纤维化的检测
1采用碱水解法检测膀胱组织中羟脯氨酸的含量
采用碱水解法检测各组膀胱组织中羟脯氨酸的含量,用方差分析比较各组之间羟脯氨酸的表达差异,探讨糖尿病病变时膀胱纤维化的改变。
2采用免疫组化、RT-PCR及ELISA方法检测膀胱组织中TGFβ1、bFGF的表达分别采用免疫组化、RT-PCR及ELISA方法检测各组膀胱组织中TGFβ1、bFGF的表达水平,用方差分析比较各组之间TGFβ1、bFGF的表达差异,探讨糖尿病病变时膀胱组织中TGFβ1、bFGF表达的变化。
三糖尿病膀胱中氧化应激状态的检测
1采用生化方法检测各组膀胱组织中过氧化氢酶(CAT)活性、超氧化物歧化酶(SOD)活性、丙二醛(MDA)水平
2采用免疫组化方法检测膀胱组织中可诱导一氧化氮合酶(iNOS)水平采用免疫组化方法检测各组膀胱组织中的iNOS表达水平,用方差分析比较各组之间iNOS的表达差异。
结果
1各组大鼠体重及血糖变化
糖尿病组大鼠和治疗组大鼠体重(240.82±10.32g,245.76±9.87g)明显低于对照组(340.34±10.12g)(P＜0.01);血糖(25.46±4.45mmol/L,25.41±4.45mmol/L)明显高于对照组(4.36±3.46mmol/L)(P＜0.01),说明糖尿病大鼠模型诱导成功。
2羟脯氨酸结果
糖尿病组羟脯氨酸含量(2.7917±1.1642μg/mg膀胱重量)比对照组(1.5326±0.5405μg/mg膀胱重量)显著升高(P＜0.05),治疗组羟脯氨酸含量(1.1745±0.7043μg/mg膀胱重量)比糖尿病组显著降低(P＜0.05)。
3 TGFβ1及bFGF免疫组化结果
免疫组化结果显示TGFβ1和bFGF阳性表达主要分布在平滑肌细胞胞浆,糖尿病组阳性TGFβ1积分光密度值(48.81±5.05)比对照组(3.73±2.26)显著升高(P＜0.05),治疗组阳性TGFβ1积分光密度值(18.65±4.60)比糖尿病组(48.81±5.05)显著降低(P＜0.05);糖尿病组阳性bFGF积分光密度值(58.40±7.04)比对照组(4.83±3.16)显著升高(P＜0.05),治疗组阳性bFGF积分光密度值(7.75±4.10)比糖尿病组(58.40±7.04)显著降低(P＜0.05)。
4 TGFβ1及bFGF的表达
糖尿病组TGFβmRNA表达中位数(0.422±0.127)及蛋白表达(21.436±1.65pg./μgprotein)比对照组(0.1 45±0.052)及(7.159±1.956pg./μgprotein)显著升高(P＜0.05),治疗组TGFβmRNA表达中位数(0.281±0.066)及蛋白表达(14.641±1.938pg./μgprotein)比糖尿病组(0.422±0.127)及(21.436±1.65 Pg./μg protein)显著降低(P＜0.05);糖尿病组bFGFmRNA表达中位数(0.489±0.096)及蛋白表达(25.968±1.648pg./μg protein)比对照组(0.118±0.021)及(7.952±0.854 pg./μg protein)显著升高(P＜0.05),治疗组bFGFmRNA表达中位数(0.097±0.011)及蛋白表达(12.911±0.984pg./μgprotein)比糖尿病组(0.489±0.096)及(25.968±1.648 pg./μg protein)显著降低(P＜0.05)。
5氧化应激水平
糖尿病组膀胱组织中的CAT活性(11.458±1.688 U/mgprotein)和SOD活性(7.159±2.956 U/mgprotein)与对照组(16.099±1.767 U/mgprotein,21.436±5.645U/mgprotein)相比显著降低(P＜0.05),糖尿病组中MDA水平(15.968±1.648nmol/mgprotein)与对照组(7.952±0.854 nmol/mgprotein)相比显著升高(P＜0.05),免疫组化研究显示糖尿病组膀胱组织中iNOS水平(67.5±10.6)与对照组(0.0±0.0)相比显著升高(P＜0.05)。治疗组膀胱组织中的CAT活性(14.472±1.529 U/mgprotein)和SOD活性(16.641±4.938 U/mgprotein)与糖尿病组(11.458±1.688 U/mgprotein,7.159±2.956 U/mgprotein)相比显著升高(P＜0.05);治疗组中MDA水平(10.911±0.984 nmol/mgprotein)与糖尿病组(15.968±1.648 nmol/mgprotein)相比显著降低(P＜0.05);免疫组化研究显示治疗组膀胱组织中iNOS水平(13.2±5.4)与糖尿病组(67.5±10.6)相比显著降低(P＜0.05).
结论
1糖尿病膀胱发生明显纤维化。
2 TGFβ1及bFGF在糖尿病膀胱中表达明显增加,可能是导致糖尿病膀胱病变的一个重要因素。
3 CAT、SOD活性明显降低,MDA水平及iNOS表达明显增加,氧化应激状态的异常可能是导致糖尿病膀胱病变的一个重要因素。
4生长因子抑制剂能改善糖尿病膀胱纤维化及氧化应激状态。
Objective:To discuss the pathogenesis of bladder fibrosis and investigate oxidative stress in the bladder 8 weeks after diabetes induction and evaluate the effect of Tadenan on improving bladder fibrosis and oxidative stress status due to diabetes.
Materials and Methods:
1.To set up the model of diabetic cystopathy in Wistar rats:30 Wistar rats(3months, 200±10g)were divided into three groups:control(n=10),streptozotocin-induced diabetic group(n=10),treatment(n=10;diabetic rats were fed with Tadenan100 mg.Kg~(-1).day~(-1)).8 weeks later,the bladders were dissected.
2.Measurement of hydroxyproline using a method based on alkaline hydrolysis, Variance analysis compares the difference of results of each group.
3.Measurement of the expression of TGFβ1 and bFGF by immunohistochemistry, RT-PCR and ELISA in the bladder,Variance analysis compares the difference of results of each group.
4.Measurement of catalase(CAT),superoxide dismutase(SOD),mark for lipid peroxidation,maleic dialdehyde(MDA)by chemical methods and the levels of inducible nitric oxide synthase(iNOS)using immunohistochemistry.
Results:
1.Bodyweight and blood glucose in each group:bodyweight and blood glucose of control group,diabetic group and treatment group are 340.34±10.12g, 4.36±3.46mmol/L,240.82±10.32g,25.46±4.45mmol/L,245.76±9.87g, 25.41±4.45mmol/L,which illustrated the rats in diabetic and treatment group had successfully induced into diabetic rats.
2.The content of hydroxyproline in control,diabetic and treatment group are 1.5326±0.5405μg/mg bladder weight,2.7917±1.1642μg/mg bladder weight, 1.1745±0.7043μg/mg bladder weight.There is great difference in diabetic group vs control(P＜0.05)and in treatment vs diabetic group(P＜0.05).
3.Immunohistochemical results in each group:the IOD of TGF and bFGF in control, diabetic and treatment group are 3.73±2.26,4.83±3.16;48.81±5.05,58.40±7.04; 18.65±4.60,7.75±4.10.There is great difference in diabetic group vs control(P＜0.05) and in treatment group vs diabetic group(P＜0.05).
4.Expression of TGF and bFGF in each group:the median expression of TGF mRNA and protein in diabetic group(0.422±0.127;21.436±1.65 pg./μgprotein)are significantly higher than that in control(0.145±0.052;7.159±1.956 pg./μgprotein) (P＜0.05),the median expression of TGF mRNA and protein in treatment group(0.281±0.066;14.641±1.938 pg./μgprotein)are significantly lower than that in diabetic group(0.422±0.127;21.436±1.65 pg./μgprotein)(P＜0.05).The median expression of bFGF mRNA and protein in diabetic group(0.489±0.096;25.968±1.648 pg./μg protein)are significantly higher than that in control(0.118±0.021;7.952±0.854 pg./μg protein)(P＜0.05),the median expression of bFGF mRNA and protein in treatment group(0.097±0.011;12.911±0.984 pg./μgprotein)are significantly lower than that in diabetic group(0.489±0.096;25.968±1.648 pg./μg protein)(P＜0.05).
5.The CAT and SOD activity(11.458±1.688 U/mgprotein,7.159±2.956 U/mgprotein) significantly decreased from diabetic group compared with control group(16.099±1.767 U/mgprotein,21.436±5.645 U/mgprotein)(P＜0.05),MDA levels(15.968±1.648 nmol/mgprotein)significantly increased from diabetic group compared with control group(7.952±0.854 nmol/mgprotein)(P＜0.05), Immunohistochemical studies showed a statistically significant increased number of iNOS-positive cells in diabetic group(67.5±10.6)compared with control group(0.0±0.0)(P＜0.05).The CAT and SOD activity(14.472±1.529 U/mgprotein, 16.641±4.938 U/mgprotein)significantly increased from treatment group compared with diabetic group(11.458±1.688 U/mgprotein,7.159±2.956 U/mgprotein)(P＜0.05), MDA levels(10.911±0.984 nmol/mgprotein)significantly decreased from treatment group compared with diabetic group(15.968±1.648 nmol/mgprotein)(P＜0.05), Immunohistochemical studies showed a statistically significant decreased number of iNOS-positive cells in treatment group(13.2±5.4)compared with diabetic group(67.5±10.6)(P＜0.05).
Conclusions:
1.Bladder fibrosis occurred easily during diabetic cystopathy.
2.Expression of TGFβ1 and bFGF were increased significantly in diabetic bladder, which maybe an important factor in diabetic cystopathy.
3.Activities of CAT and SOD were decreased significantly and the level of MDA and expression of iNOS were increased significantly in diabetic bladder,which indicated oxidative stress played an important role in diabetic cystopathy.
4.Tadenan could effectively slow down the process of bladder fibrosis and improve oxidative stress status due to diabetes.
研究材料、方法
一建立糖尿病大鼠模型
取30只3月龄(200±10g)雄性Wistar大鼠随机分为3组,分笼饲养。分别为对照组(n=10);糖尿病组(n=10);治疗组(n=10)。建模成功后饲养4周,然后对治疗组大鼠给予太得恩药物(100mg.Kg~(-1).d~(-1),溶于2ml花生油中,胃管注入)治疗4周,对照组及糖尿病组大鼠给予同等剂量花生油赋形剂。以上动物共饲养8周。8周后处死完整切取膀胱组织。
二糖尿病膀胱中纤维化的检测
1采用碱水解法检测膀胱组织中羟脯氨酸的含量
采用碱水解法检测各组膀胱组织中羟脯氨酸的含量,用方差分析比较各组之间羟脯氨酸的表达差异,探讨糖尿病病变时膀胱纤维化的改变。
2采用免疫组化、RT-PCR及ELISA方法检测膀胱组织中TGFβ1、bFGF的表达分别采用免疫组化、RT-PCR及ELISA方法检测各组膀胱组织中TGFβ1、bFGF的表达水平,用方差分析比较各组之间TGFβ1、bFGF的表达差异,探讨糖尿病病变时膀胱组织中TGFβ1、bFGF表达的变化。
三糖尿病膀胱中氧化应激状态的检测
1采用生化方法检测各组膀胱组织中过氧化氢酶(CAT)活性、超氧化物歧化酶(SOD)活性、丙二醛(MDA)水平
2采用免疫组化方法检测膀胱组织中可诱导一氧化氮合酶(iNOS)水平采用免疫组化方法检测各组膀胱组织中的iNOS表达水平,用方差分析比较各组之间iNOS的表达差异。
结果
1各组大鼠体重及血糖变化
糖尿病组大鼠和治疗组大鼠体重(240.82±10.32g,245.76±9.87g)明显低于对照组(340.34±10.12g)(P＜0.01);血糖(25.46±4.45mmol/L,25.41±4.45mmol/L)明显高于对照组(4.36±3.46mmol/L)(P＜0.01),说明糖尿病大鼠模型诱导成功。
2羟脯氨酸结果
糖尿病组羟脯氨酸含量(2.7917±1.1642μg/mg膀胱重量)比对照组(1.5326±0.5405μg/mg膀胱重量)显著升高(P＜0.05),治疗组羟脯氨酸含量(1.1745±0.7043μg/mg膀胱重量)比糖尿病组显著降低(P＜0.05)。
3 TGFβ1及bFGF免疫组化结果
免疫组化结果显示TGFβ1和bFGF阳性表达主要分布在平滑肌细胞胞浆,糖尿病组阳性TGFβ1积分光密度值(48.81±5.05)比对照组(3.73±2.26)显著升高(P＜0.05),治疗组阳性TGFβ1积分光密度值(18.65±4.60)比糖尿病组(48.81±5.05)显著降低(P＜0.05);糖尿病组阳性bFGF积分光密度值(58.40±7.04)比对照组(4.83±3.16)显著升高(P＜0.05),治疗组阳性bFGF积分光密度值(7.75±4.10)比糖尿病组(58.40±7.04)显著降低(P＜0.05)。
4 TGFβ1及bFGF的表达
糖尿病组TGFβmRNA表达中位数(0.422±0.127)及蛋白表达(21.436±1.65pg./μgprotein)比对照组(0.1 45±0.052)及(7.159±1.956pg./μgprotein)显著升高(P＜0.05),治疗组TGFβmRNA表达中位数(0.281±0.066)及蛋白表达(14.641±1.938pg./μgprotein)比糖尿病组(0.422±0.127)及(21.436±1.65 Pg./μg protein)显著降低(P＜0.05);糖尿病组bFGFmRNA表达中位数(0.489±0.096)及蛋白表达(25.968±1.648pg./μg protein)比对照组(0.118±0.021)及(7.952±0.854 pg./μg protein)显著升高(P＜0.05),治疗组bFGFmRNA表达中位数(0.097±0.011)及蛋白表达(12.911±0.984pg./μgprotein)比糖尿病组(0.489±0.096)及(25.968±1.648 pg./μg protein)显著降低(P＜0.05)。
5氧化应激水平
糖尿病组膀胱组织中的CAT活性(11.458±1.688 U/mgprotein)和SOD活性(7.159±2.956 U/mgprotein)与对照组(16.099±1.767 U/mgprotein,21.436±5.645U/mgprotein)相比显著降低(P＜0.05),糖尿病组中MDA水平(15.968±1.648nmol/mgprotein)与对照组(7.952±0.854 nmol/mgprotein)相比显著升高(P＜0.05),免疫组化研究显示糖尿病组膀胱组织中iNOS水平(67.5±10.6)与对照组(0.0±0.0)相比显著升高(P＜0.05)。治疗组膀胱组织中的CAT活性(14.472±1.529 U/mgprotein)和SOD活性(16.641±4.938 U/mgprotein)与糖尿病组(11.458±1.688 U/mgprotein,7.159±2.956 U/mgprotein)相比显著升高(P＜0.05);治疗组中MDA水平(10.911±0.984 nmol/mgprotein)与糖尿病组(15.968±1.648 nmol/mgprotein)相比显著降低(P＜0.05);免疫组化研究显示治疗组膀胱组织中iNOS水平(13.2±5.4)与糖尿病组(67.5±10.6)相比显著降低(P＜0.05).
结论
1糖尿病膀胱发生明显纤维化。
2 TGFβ1及bFGF在糖尿病膀胱中表达明显增加,可能是导致糖尿病膀胱病变的一个重要因素。
3 CAT、SOD活性明显降低,MDA水平及iNOS表达明显增加,氧化应激状态的异常可能是导致糖尿病膀胱病变的一个重要因素。
4生长因子抑制剂能改善糖尿病膀胱纤维化及氧化应激状态。
Objective:To discuss the pathogenesis of bladder fibrosis and investigate oxidative stress in the bladder 8 weeks after diabetes induction and evaluate the effect of Tadenan on improving bladder fibrosis and oxidative stress status due to diabetes.
Materials and Methods:
1.To set up the model of diabetic cystopathy in Wistar rats:30 Wistar rats(3months, 200±10g)were divided into three groups:control(n=10),streptozotocin-induced diabetic group(n=10),treatment(n=10;diabetic rats were fed with Tadenan100 mg.Kg~(-1).day~(-1)).8 weeks later,the bladders were dissected.
2.Measurement of hydroxyproline using a method based on alkaline hydrolysis, Variance analysis compares the difference of results of each group.
3.Measurement of the expression of TGFβ1 and bFGF by immunohistochemistry, RT-PCR and ELISA in the bladder,Variance analysis compares the difference of results of each group.
4.Measurement of catalase(CAT),superoxide dismutase(SOD),mark for lipid peroxidation,maleic dialdehyde(MDA)by chemical methods and the levels of inducible nitric oxide synthase(iNOS)using immunohistochemistry.
Results:
1.Bodyweight and blood glucose in each group:bodyweight and blood glucose of control group,diabetic group and treatment group are 340.34±10.12g, 4.36±3.46mmol/L,240.82±10.32g,25.46±4.45mmol/L,245.76±9.87g, 25.41±4.45mmol/L,which illustrated the rats in diabetic and treatment group had successfully induced into diabetic rats.
2.The content of hydroxyproline in control,diabetic and treatment group are 1.5326±0.5405μg/mg bladder weight,2.7917±1.1642μg/mg bladder weight, 1.1745±0.7043μg/mg bladder weight.There is great difference in diabetic group vs control(P＜0.05)and in treatment vs diabetic group(P＜0.05).
3.Immunohistochemical results in each group:the IOD of TGF and bFGF in control, diabetic and treatment group are 3.73±2.26,4.83±3.16;48.81±5.05,58.40±7.04; 18.65±4.60,7.75±4.10.There is great difference in diabetic group vs control(P＜0.05) and in treatment group vs diabetic group(P＜0.05).
4.Expression of TGF and bFGF in each group:the median expression of TGF mRNA and protein in diabetic group(0.422±0.127;21.436±1.65 pg./μgprotein)are significantly higher than that in control(0.145±0.052;7.159±1.956 pg./μgprotein) (P＜0.05),the median expression of TGF mRNA and protein in treatment group(0.281±0.066;14.641±1.938 pg./μgprotein)are significantly lower than that in diabetic group(0.422±0.127;21.436±1.65 pg./μgprotein)(P＜0.05).The median expression of bFGF mRNA and protein in diabetic group(0.489±0.096;25.968±1.648 pg./μg protein)are significantly higher than that in control(0.118±0.021;7.952±0.854 pg./μg protein)(P＜0.05),the median expression of bFGF mRNA and protein in treatment group(0.097±0.011;12.911±0.984 pg./μgprotein)are significantly lower than that in diabetic group(0.489±0.096;25.968±1.648 pg./μg protein)(P＜0.05).
5.The CAT and SOD activity(11.458±1.688 U/mgprotein,7.159±2.956 U/mgprotein) significantly decreased from diabetic group compared with control group(16.099±1.767 U/mgprotein,21.436±5.645 U/mgprotein)(P＜0.05),MDA levels(15.968±1.648 nmol/mgprotein)significantly increased from diabetic group compared with control group(7.952±0.854 nmol/mgprotein)(P＜0.05), Immunohistochemical studies showed a statistically significant increased number of iNOS-positive cells in diabetic group(67.5±10.6)compared with control group(0.0±0.0)(P＜0.05).The CAT and SOD activity(14.472±1.529 U/mgprotein, 16.641±4.938 U/mgprotein)significantly increased from treatment group compared with diabetic group(11.458±1.688 U/mgprotein,7.159±2.956 U/mgprotein)(P＜0.05), MDA levels(10.911±0.984 nmol/mgprotein)significantly decreased from treatment group compared with diabetic group(15.968±1.648 nmol/mgprotein)(P＜0.05), Immunohistochemical studies showed a statistically significant decreased number of iNOS-positive cells in treatment group(13.2±5.4)compared with diabetic group(67.5±10.6)(P＜0.05).
Conclusions:
1.Bladder fibrosis occurred easily during diabetic cystopathy.
2.Expression of TGFβ1 and bFGF were increased significantly in diabetic bladder, which maybe an important factor in diabetic cystopathy.
3.Activities of CAT and SOD were decreased significantly and the level of MDA and expression of iNOS were increased significantly in diabetic bladder,which indicated oxidative stress played an important role in diabetic cystopathy.
4.Tadenan could effectively slow down the process of bladder fibrosis and improve oxidative stress status due to diabetes.
引文
1 Olapade-Olaopa EO, Morley RN. Diabetic cystopathy presenting as primary acute urinary retention in a previously undiagnosed young male diabetic patient. Diabetic Complications, 1997,11(6): 350-1.
2 Deveaud CM, Macarak EJ, Kucich U, et al. Molecular analysis of collagens in bladder fibrosis. J Urol, 1998,160(4): 1518-27.
3 Howard PS, Renfrow D, Schechter NM, et al. Mast cell chymase is a possible mediator of neurogenic bladder fibrosis. Neurol Urodyn, 2004, 23(4): 374-82.
4 Kim JC, Yoon JY, Seo SI, et al. Effects of partial bladder outlet obstruction and its relief of types I and II collagen and detrusor contractility in the rat. Neurol Urodyn 2000,19: 29-42.
5 Kim JC, Seo SIL, Park YH, et al. Changes in detrusor and urinary growth factors according to detrusor function after partial bladder outlet obstruction in the rat. Urology, 2001, 57(2): 371-5.
6 Nakae M, Kamiya H, Naruse K, et al. Effects of basic fibroblast growth factor on experimental diabetic neuropathy in rats. Diabetes, 2006, 55(5): 1470-77.
7 Wang Z-J, Ikeda K, Wada Y, et al. Expression and location of basic fibroblast growth factor in diabetic rat prostate. BJU Int, 2000, 85: 945-52.
8 Roberts AB, McCune BK, Sporn MB. TGF-beta: regulation of extracellular matrix. Kidney Int, 1992,41(3):557-9.
9 Miyazono K, Hellman U, Wernstedt C, Heldin CH. Latent high molecular weight complex of transforming growth factor beta 1. Purification from human platelets and structural characterization. J Biol Chem, 1988, 263(13): 6407-15.
10 Border WA, Ruoslahti E. Transforming growth factor-b in disease: The dark side of tissue repair. J Clin Invest, 1992, 90(1): 1-7.
11 Darius J, Bagli L, John G, et al. Basic fibroblast growth factor in the urine of children with voiding pathology. J Urol, 1997, 158(3 pt 2): 1123-1127.
12 Laurence S, BaskinR, Ronald S, et al. Growth factors in bladder wound healing. J Urol,1997,157(6):2388-2395.
13孙小兵,李金良,陈雨历,等.TGF-β1与神经性膀胱逼尿肌纤维化.中华小儿外科杂志,2002,23(6):502-504.
14杨秀颖,杜冠化.组织羟脯氨酸含量微量测定方法及应用.中国药理学通报,2004,20(7):836-837.
15Slatter DA,Bolton CH,Bailey AJ.The importance of lipid-derived malondialdehyde in diabetes mellitus.Diabetologia,2000,43(5):550-557.
16Kfistal BS,Koopmans S J,Jackson CT,et al.Oxidant mediated repression of the mitochondrial transcription in diabetic rats.Free Rad Biol Med,1997,22(5):813-822.
17Bonnefont-Rousselot D,Bastard JP,Jaudon MC,et al.Consequences of the diabetic status on the oxidant/antioxidant balance.Diabetes Metab,2000,26(3):163-176.
18Munzel T,Heitzer T,Harrison DG.The physiology and pathophysiology of nitric oxide/superoxide system.Herz,1997,22(3):158-172.
19Beshay E,and Prud homme G.Inhibitors of phosphodiesterase isoforms Ⅲ or Ⅳsuppress islet-cell nitric oxide production.Lab Invest,2001,81(8):1109-1117.
20Romaneko A,Morimura K,Waniuchi H,et al.Increased oxidative stress with gene alteration in urinary bladder urothelium after Chemobyl accident.Int J Cancer,2000,86(8):790-798.
21Yablonsky F,Nicolas V,Riffaud JP,et al.Antiproliferative effect of Phygeum africanum on rat prostate fibroblasts.J Urol,1997;157(6):2381-7.
22Kindt E,Gueneva-Boucheva K,Rekhter MD,et al.Determination of hydroxyproline in plasma and tissue using electrospray mass spectrometry.J Pharm Biomed Anal,2003,33(5):1081-92.
23Howard PS,Kcich U,Coplen DE,et al.Transforming growth factor-beta-1-induced hypertrophy and matrix expression in human bladder smooth muscle cells.Urology,2005,66(6):1349-53.
24李京红,何冰,翁保迎.转化生长因子β1单克隆抗体对大鼠肺纤维化的治疗观察.中华结核和呼吸杂志,1997,20:347-349.
25Morrissey J,Guo G,Moridaira K,et al.Transforming growth factor-beta induces renal epithelial jagged-1 expression in fibrotic disease.J Am Soc Nephrol,2002,13(6):1499-508.
26刘维田,王吉耀,朱无难,等.转化生长因子β1对纤维母细胞Ⅰ、Ⅲ型前胶原及胶原酶mRNA表达的影响.中华肝脏病杂志,1996,4:97-100.
27Ebner R,Chen RH,Lawler S,et al.Determination of type Ⅰ receptor specificity by the type Ⅱ receptor for TGFI3or activin.Science,1993,262(5135):900-950.
28Chiarelli F,Santilli F,Mohn A.Role of growth factors in the development of diabetic complications.Horm Res,2000,53(2):53-67.
29Koo HP,Santarosa RP,Buttyan R,et al.Early molecular changes associated with streptozotocin-induced diabetic bladder hypertrophy in the rat.Urol Res 1993,21(6):375-81.
30Darius J,Bagli L,John G,et al.Basic fibroblast growth factor in the urine of children with voiding pathology.J Urol,1997,158(3 pt 2):1123-1127.
31Laurence S,Baskin R,Ronald S,et al.Growth factors in bladder wound healing.J Urol,1997,157(6):2388-2395.
32Majors A,Ehrhart LA.Basic fibroblast growth factor in the extracellular matrix suppresses collagen synthesis and type III procollagen mRNA levels in arterial smooth muscle cell cultures.Arterioscler Thromb,1993,13(5):680-6.
33Kakkar R,Kalra J,Mantha SV,et al.Lipid peroxidation and activity of antioxidant enzymes in diabetic rats.Mol Cell Biochem,1995,151(2):113-119.
34Pryor WA,and Godberg SS.Noninvasive measures of oxidative stress status in humans.Free Rad Biol Med,1991,10(3-4):177-184.
35Xu X,Cubeddu LX,and Malave A.Expression of inducible nitric oxide synthase in primary culture of rat bladder smooth muscle cells by plasma from cyclophosphamide-treated rats.Eur J Pharm,2001,416(1-2):1-9.
36Hunt JV,Smith CCT,Wolff SP.Autoxidative glycation and possible involvement of peroxide and free radicals in LDL modification by glucose.Diabetes,1990,39:1420-1424.
37 Gillery P,Monboiss JC,Maquart FX,et al.Glycation of protein as a source of superoxide.Diabetes Metab,1988,14:25-30.
38 Mullarkey CJ,Edelstein D,Brownlee M.Free radical generation by early glycation products:a mechanism for accelerated atherogenesis in diabetes.Bioch Biophys Res Commun,1990,173(3):932-939.
39 Wong A,Dukic-Stefanovic S,Gasic-Milenkovic J,et al.Anti-inflammatory antioxidants attenuate the expression of inducible nitric oxide synthase mediated by advanced glycation endproducts in murine microglia.Eur J Neurosci,2001,14(12):1961-1967.
40 Lin CH,Lin YF,Chang MC,et al.Advanced glycation end products induce nitric oxide synthase expression in C6 glioma cells:involvement of a p38 MAP kinase-dependent mechanism.Life Sci,2001,69(21):2503-2515.
41 Rojas A,Caveda L,Romay C,et al.Effect of advanced glycosylation end products on the induction of nitric oxide synthase in murine macrophages.Biochem Biophys Res Comm,1996,225(2):358-362.
42 Szolnoki E,Reichart E,Marchal S,et al.The effect of Pygeum africanum on fibroblast growth factor(FGF)and transforming growth factor beta(YGF betal/LAP)expression in animal model.Acta Microbiol Immuno Hung,2001,48(1):1-9.
43 王国民,王杭,张永康.Tadenan对实验兔膀胱出口部分梗阻所致膀胱功能障碍的治疗作用.中华泌尿外科杂志,2001,22(9):517.519.
44 Persson K,Andersson KE.Nitric oxide and relaxation of pig lower urinary tract.Br J Pharmocol,1992,106(2):416-422.
45 Michael A,Pfeifer P,Mary P,et al.Perspectives in diabetes clinical trials of diabetic neuropathy past,present and future.Diabetes,1995,44:1355.
46 Luheshi GN,Zar MA.The effect of streptozotocin-induced diabetes on cholinergic motor transmission in the rat urinary bladder.Br J Pharmacol,1991,103(3):1657-62.
47 Steers WD,Mackway-Gerardi AM,Ciambotti J,et al.Alterations in neural pathways to the urinary bladder of the rat in response to streptozotocin-induced diabetes.J Auton Nerv Syst,1994,47:83-94.
48 Nadelhaft I,Vera PL,Steinbacher B.Hypertrophic neurons innervating the urinary bladder and colon of the streptozotocin-induced diabetic rat.Brain Res,1993,609:277-283.
49 Luheshi GN,Zar MA.The effect of strptozotocin-induced diabetes on cholinergic motor transmission in the rat urinary bladder.Br J Pharmacol,1991,103:1657-1662.
50 Dahlatrand(2,Dahlstrom A,Ahlman H,et al.Effect of substance P on detrusor muscle rabbits with alloxan induced hyperglycemia:possible role of 5-hydorxytryptamine for diabetic bladder dysfunction.J Urol,2002,168:303-307.
51 Kamata K,Inoue K,Kasuya Y.Changes in contractile responses of the urinary bladder to substance P in streptozotocin-induced diabetic rats.Gen Pharmacol,1993,24:547-535.
52 Changolkar AK,Hypolite JA,Disanto M,et al.Diabetes induced decrease in detrusor smooth muscle force is associated with oxidative stress and overactivity of aldose reductase.J Urol,2005,173:309-313.
53 Yat-Ching Tong,Juei-Tang Cheng.Changes in bladder nerve-growth factor and p75NTR genetic expression in streptozotocin-induced diabetic rats.BJU International,2005,96(6):1392-1396.
54 龚宇,熊恩庆,金锡御,等.糖尿病大鼠膀胱壁CGRP神经改变的研究.中华泌尿外科杂志,2000,21:634-635.
55 Li yongzhi,Shi benkang,Zhang jianping,Ran lingxia,Bian wei,Zhu yaofeng,Zhang keqin,Vincent laudon.Expression of transforming growth factor β1 gene,basic fibroblast growth gene and hydroxyproline in diabetes-induced bladder dysfunction in a rat model.Neurourol Urodyn.2007,27(3):254-259.
56 王东文,双卫兵,高宏飞,等.糖尿病大鼠逼尿肌线粒体琥珀酸脱氢酶及其超微结构的改变.中华实验外科杂志,2005,22:1362-1363.
57 Latifpour J,Nishimoto T,Marian MJ,et al.Differential regulation of bladder beta-adrenergic and muscarinic cholinergic receptors in experimental diabetes.Diabetes,1991,0:1150-1156.
58 Ichiyanagi N,Tsujii T,Masude H,et al.changed responsiveness of the detrusor in rabbits with alloxan induced hyperglycemia:possible role of 5-hydroxytryptamine for diabetic bladder dysfunction.J Urol,2002,168:303-307.
59 Waring JV,Wendt R.Effects of strptozotocin-induced diabetes mellitus on intracellular calcium and contraction of longitudinal smooth muscle from rat urinary bladder.J Urol,2000,163:323.
60 Longhurst PA.Urinary bladder function 6 months after the onset of diabetes in the spontaneously diabetic BB rat.J Urol,1991,145:417-422.
61 Longhuret PA,Broteke TP,Leggett RE,et al.The influence of streptozotocin-induced diabetes mellitus on the sensitivity of rat urinary bladder body and base strips to changes inextracellular calcium.Gen Pharmacol,1992,23:83-88.
62 Saffore LY,Firouz D,Venk K,et al.Protein kinase C may be elevated in diabetic bladder muscle.Diabetes,2004,53:613.
63 Nakamura J,Kato K,H amada Y,et al.A protein kinase C-beta-selective inhibitor ameliorates neural dysfunction in streptozotocin-induced diaebetic rats.Diabetes,1999,48:2090-2095.
64 张德宪,林君丽,于青云,等.温阳利水法治疗糖尿病神经原性膀胱.山东中医杂志,1998,17:403-404.
65 史本康,李永智,张建平,边炜,王海峰.糖尿病大鼠膀胱中P物质、神经生长因子表达及氧化应激状态的研究.中华泌尿外科杂志.2008,29(1):49-53.
66 Antonio C.New insights on oxidatant stress and diabetic complications may lead to.a causal antioxidant therapy.Diabetes Care,2003,26:1589-1596.
67 Virag L,Salzman AL,Szabo C.Poly synthetase activation mediates mitochondrial injury during oxidant-induced cell death.J Immunol,1998,161:3753-3759.
68 邹大进,陈月.糖尿病微血管并发症的发病机制及治疗.中华糖尿病杂志,2005,13(5):393-395.
69 Kazumasa Torimoto,Matthew O Fraser,Yoshihiko Hirao.Urethral dysfunction in diabetic rats.J Urol,2004,171(5):1959-64.
70 Zhongguang Yang,Paul C Dolber,Matthew O Fraser.Diabetic urethropathy compounds the effects of diabetic cystopathy.J Urol,2007,178(11):2213-9.
71 Torimoto K, Hirao Y, Matsuyoshi H, et al. Alpha1-adrenergic mechanism in diabetic urethral dysfunction in rats. J Urol, 2005,173(3): 1027.
73 Sato M, Shirai M. Diabetic urinary bladder and urethral dysfunction. Nippon Rinsho, 1997, 55(suppl):876-80.
74 Rapidi CA, Karandreas N, Katsifotis C, et al. A combined urodynamic and electrophysiological study of diabetic cystopathy. Neurourol Urodyn,2006,25(1):32-8.
75 Soylu A, Akinci A, Yilmaz U, et al. Sympathetic skin responses in Type-1 diabetic children: relationship to urodynamic findings. Neurourol Urodyn, 2006, 25(3):243-8.
2 Deveaud CM, Macarak EJ, Kucich U, et al. Molecular analysis of collagens in bladder fibrosis. J Urol, 1998,160(4): 1518-27.
3 Howard PS, Renfrow D, Schechter NM, et al. Mast cell chymase is a possible mediator of neurogenic bladder fibrosis. Neurol Urodyn, 2004, 23(4): 374-82.
4 Kim JC, Yoon JY, Seo SI, et al. Effects of partial bladder outlet obstruction and its relief of types I and II collagen and detrusor contractility in the rat. Neurol Urodyn 2000,19: 29-42.
5 Kim JC, Seo SIL, Park YH, et al. Changes in detrusor and urinary growth factors according to detrusor function after partial bladder outlet obstruction in the rat. Urology, 2001, 57(2): 371-5.
6 Nakae M, Kamiya H, Naruse K, et al. Effects of basic fibroblast growth factor on experimental diabetic neuropathy in rats. Diabetes, 2006, 55(5): 1470-77.
7 Wang Z-J, Ikeda K, Wada Y, et al. Expression and location of basic fibroblast growth factor in diabetic rat prostate. BJU Int, 2000, 85: 945-52.
8 Roberts AB, McCune BK, Sporn MB. TGF-beta: regulation of extracellular matrix. Kidney Int, 1992,41(3):557-9.
9 Miyazono K, Hellman U, Wernstedt C, Heldin CH. Latent high molecular weight complex of transforming growth factor beta 1. Purification from human platelets and structural characterization. J Biol Chem, 1988, 263(13): 6407-15.
10 Border WA, Ruoslahti E. Transforming growth factor-b in disease: The dark side of tissue repair. J Clin Invest, 1992, 90(1): 1-7.
11 Darius J, Bagli L, John G, et al. Basic fibroblast growth factor in the urine of children with voiding pathology. J Urol, 1997, 158(3 pt 2): 1123-1127.
12 Laurence S, BaskinR, Ronald S, et al. Growth factors in bladder wound healing. J Urol,1997,157(6):2388-2395.
13孙小兵,李金良,陈雨历,等.TGF-β1与神经性膀胱逼尿肌纤维化.中华小儿外科杂志,2002,23(6):502-504.
14杨秀颖,杜冠化.组织羟脯氨酸含量微量测定方法及应用.中国药理学通报,2004,20(7):836-837.
15Slatter DA,Bolton CH,Bailey AJ.The importance of lipid-derived malondialdehyde in diabetes mellitus.Diabetologia,2000,43(5):550-557.
16Kfistal BS,Koopmans S J,Jackson CT,et al.Oxidant mediated repression of the mitochondrial transcription in diabetic rats.Free Rad Biol Med,1997,22(5):813-822.
17Bonnefont-Rousselot D,Bastard JP,Jaudon MC,et al.Consequences of the diabetic status on the oxidant/antioxidant balance.Diabetes Metab,2000,26(3):163-176.
18Munzel T,Heitzer T,Harrison DG.The physiology and pathophysiology of nitric oxide/superoxide system.Herz,1997,22(3):158-172.
19Beshay E,and Prud homme G.Inhibitors of phosphodiesterase isoforms Ⅲ or Ⅳsuppress islet-cell nitric oxide production.Lab Invest,2001,81(8):1109-1117.
20Romaneko A,Morimura K,Waniuchi H,et al.Increased oxidative stress with gene alteration in urinary bladder urothelium after Chemobyl accident.Int J Cancer,2000,86(8):790-798.
21Yablonsky F,Nicolas V,Riffaud JP,et al.Antiproliferative effect of Phygeum africanum on rat prostate fibroblasts.J Urol,1997;157(6):2381-7.
22Kindt E,Gueneva-Boucheva K,Rekhter MD,et al.Determination of hydroxyproline in plasma and tissue using electrospray mass spectrometry.J Pharm Biomed Anal,2003,33(5):1081-92.
23Howard PS,Kcich U,Coplen DE,et al.Transforming growth factor-beta-1-induced hypertrophy and matrix expression in human bladder smooth muscle cells.Urology,2005,66(6):1349-53.
24李京红,何冰,翁保迎.转化生长因子β1单克隆抗体对大鼠肺纤维化的治疗观察.中华结核和呼吸杂志,1997,20:347-349.
25Morrissey J,Guo G,Moridaira K,et al.Transforming growth factor-beta induces renal epithelial jagged-1 expression in fibrotic disease.J Am Soc Nephrol,2002,13(6):1499-508.
26刘维田,王吉耀,朱无难,等.转化生长因子β1对纤维母细胞Ⅰ、Ⅲ型前胶原及胶原酶mRNA表达的影响.中华肝脏病杂志,1996,4:97-100.
27Ebner R,Chen RH,Lawler S,et al.Determination of type Ⅰ receptor specificity by the type Ⅱ receptor for TGFI3or activin.Science,1993,262(5135):900-950.
28Chiarelli F,Santilli F,Mohn A.Role of growth factors in the development of diabetic complications.Horm Res,2000,53(2):53-67.
29Koo HP,Santarosa RP,Buttyan R,et al.Early molecular changes associated with streptozotocin-induced diabetic bladder hypertrophy in the rat.Urol Res 1993,21(6):375-81.
30Darius J,Bagli L,John G,et al.Basic fibroblast growth factor in the urine of children with voiding pathology.J Urol,1997,158(3 pt 2):1123-1127.
31Laurence S,Baskin R,Ronald S,et al.Growth factors in bladder wound healing.J Urol,1997,157(6):2388-2395.
32Majors A,Ehrhart LA.Basic fibroblast growth factor in the extracellular matrix suppresses collagen synthesis and type III procollagen mRNA levels in arterial smooth muscle cell cultures.Arterioscler Thromb,1993,13(5):680-6.
33Kakkar R,Kalra J,Mantha SV,et al.Lipid peroxidation and activity of antioxidant enzymes in diabetic rats.Mol Cell Biochem,1995,151(2):113-119.
34Pryor WA,and Godberg SS.Noninvasive measures of oxidative stress status in humans.Free Rad Biol Med,1991,10(3-4):177-184.
35Xu X,Cubeddu LX,and Malave A.Expression of inducible nitric oxide synthase in primary culture of rat bladder smooth muscle cells by plasma from cyclophosphamide-treated rats.Eur J Pharm,2001,416(1-2):1-9.
36Hunt JV,Smith CCT,Wolff SP.Autoxidative glycation and possible involvement of peroxide and free radicals in LDL modification by glucose.Diabetes,1990,39:1420-1424.
37 Gillery P,Monboiss JC,Maquart FX,et al.Glycation of protein as a source of superoxide.Diabetes Metab,1988,14:25-30.
38 Mullarkey CJ,Edelstein D,Brownlee M.Free radical generation by early glycation products:a mechanism for accelerated atherogenesis in diabetes.Bioch Biophys Res Commun,1990,173(3):932-939.
39 Wong A,Dukic-Stefanovic S,Gasic-Milenkovic J,et al.Anti-inflammatory antioxidants attenuate the expression of inducible nitric oxide synthase mediated by advanced glycation endproducts in murine microglia.Eur J Neurosci,2001,14(12):1961-1967.
40 Lin CH,Lin YF,Chang MC,et al.Advanced glycation end products induce nitric oxide synthase expression in C6 glioma cells:involvement of a p38 MAP kinase-dependent mechanism.Life Sci,2001,69(21):2503-2515.
41 Rojas A,Caveda L,Romay C,et al.Effect of advanced glycosylation end products on the induction of nitric oxide synthase in murine macrophages.Biochem Biophys Res Comm,1996,225(2):358-362.
42 Szolnoki E,Reichart E,Marchal S,et al.The effect of Pygeum africanum on fibroblast growth factor(FGF)and transforming growth factor beta(YGF betal/LAP)expression in animal model.Acta Microbiol Immuno Hung,2001,48(1):1-9.
43 王国民,王杭,张永康.Tadenan对实验兔膀胱出口部分梗阻所致膀胱功能障碍的治疗作用.中华泌尿外科杂志,2001,22(9):517.519.
44 Persson K,Andersson KE.Nitric oxide and relaxation of pig lower urinary tract.Br J Pharmocol,1992,106(2):416-422.
45 Michael A,Pfeifer P,Mary P,et al.Perspectives in diabetes clinical trials of diabetic neuropathy past,present and future.Diabetes,1995,44:1355.
46 Luheshi GN,Zar MA.The effect of streptozotocin-induced diabetes on cholinergic motor transmission in the rat urinary bladder.Br J Pharmacol,1991,103(3):1657-62.
47 Steers WD,Mackway-Gerardi AM,Ciambotti J,et al.Alterations in neural pathways to the urinary bladder of the rat in response to streptozotocin-induced diabetes.J Auton Nerv Syst,1994,47:83-94.
48 Nadelhaft I,Vera PL,Steinbacher B.Hypertrophic neurons innervating the urinary bladder and colon of the streptozotocin-induced diabetic rat.Brain Res,1993,609:277-283.
49 Luheshi GN,Zar MA.The effect of strptozotocin-induced diabetes on cholinergic motor transmission in the rat urinary bladder.Br J Pharmacol,1991,103:1657-1662.
50 Dahlatrand(2,Dahlstrom A,Ahlman H,et al.Effect of substance P on detrusor muscle rabbits with alloxan induced hyperglycemia:possible role of 5-hydorxytryptamine for diabetic bladder dysfunction.J Urol,2002,168:303-307.
51 Kamata K,Inoue K,Kasuya Y.Changes in contractile responses of the urinary bladder to substance P in streptozotocin-induced diabetic rats.Gen Pharmacol,1993,24:547-535.
52 Changolkar AK,Hypolite JA,Disanto M,et al.Diabetes induced decrease in detrusor smooth muscle force is associated with oxidative stress and overactivity of aldose reductase.J Urol,2005,173:309-313.
53 Yat-Ching Tong,Juei-Tang Cheng.Changes in bladder nerve-growth factor and p75NTR genetic expression in streptozotocin-induced diabetic rats.BJU International,2005,96(6):1392-1396.
54 龚宇,熊恩庆,金锡御,等.糖尿病大鼠膀胱壁CGRP神经改变的研究.中华泌尿外科杂志,2000,21:634-635.
55 Li yongzhi,Shi benkang,Zhang jianping,Ran lingxia,Bian wei,Zhu yaofeng,Zhang keqin,Vincent laudon.Expression of transforming growth factor β1 gene,basic fibroblast growth gene and hydroxyproline in diabetes-induced bladder dysfunction in a rat model.Neurourol Urodyn.2007,27(3):254-259.
56 王东文,双卫兵,高宏飞,等.糖尿病大鼠逼尿肌线粒体琥珀酸脱氢酶及其超微结构的改变.中华实验外科杂志,2005,22:1362-1363.
57 Latifpour J,Nishimoto T,Marian MJ,et al.Differential regulation of bladder beta-adrenergic and muscarinic cholinergic receptors in experimental diabetes.Diabetes,1991,0:1150-1156.
58 Ichiyanagi N,Tsujii T,Masude H,et al.changed responsiveness of the detrusor in rabbits with alloxan induced hyperglycemia:possible role of 5-hydroxytryptamine for diabetic bladder dysfunction.J Urol,2002,168:303-307.
59 Waring JV,Wendt R.Effects of strptozotocin-induced diabetes mellitus on intracellular calcium and contraction of longitudinal smooth muscle from rat urinary bladder.J Urol,2000,163:323.
60 Longhurst PA.Urinary bladder function 6 months after the onset of diabetes in the spontaneously diabetic BB rat.J Urol,1991,145:417-422.
61 Longhuret PA,Broteke TP,Leggett RE,et al.The influence of streptozotocin-induced diabetes mellitus on the sensitivity of rat urinary bladder body and base strips to changes inextracellular calcium.Gen Pharmacol,1992,23:83-88.
62 Saffore LY,Firouz D,Venk K,et al.Protein kinase C may be elevated in diabetic bladder muscle.Diabetes,2004,53:613.
63 Nakamura J,Kato K,H amada Y,et al.A protein kinase C-beta-selective inhibitor ameliorates neural dysfunction in streptozotocin-induced diaebetic rats.Diabetes,1999,48:2090-2095.
64 张德宪,林君丽,于青云,等.温阳利水法治疗糖尿病神经原性膀胱.山东中医杂志,1998,17:403-404.
65 史本康,李永智,张建平,边炜,王海峰.糖尿病大鼠膀胱中P物质、神经生长因子表达及氧化应激状态的研究.中华泌尿外科杂志.2008,29(1):49-53.
66 Antonio C.New insights on oxidatant stress and diabetic complications may lead to.a causal antioxidant therapy.Diabetes Care,2003,26:1589-1596.
67 Virag L,Salzman AL,Szabo C.Poly synthetase activation mediates mitochondrial injury during oxidant-induced cell death.J Immunol,1998,161:3753-3759.
68 邹大进,陈月.糖尿病微血管并发症的发病机制及治疗.中华糖尿病杂志,2005,13(5):393-395.
69 Kazumasa Torimoto,Matthew O Fraser,Yoshihiko Hirao.Urethral dysfunction in diabetic rats.J Urol,2004,171(5):1959-64.
70 Zhongguang Yang,Paul C Dolber,Matthew O Fraser.Diabetic urethropathy compounds the effects of diabetic cystopathy.J Urol,2007,178(11):2213-9.
71 Torimoto K, Hirao Y, Matsuyoshi H, et al. Alpha1-adrenergic mechanism in diabetic urethral dysfunction in rats. J Urol, 2005,173(3): 1027.
73 Sato M, Shirai M. Diabetic urinary bladder and urethral dysfunction. Nippon Rinsho, 1997, 55(suppl):876-80.
74 Rapidi CA, Karandreas N, Katsifotis C, et al. A combined urodynamic and electrophysiological study of diabetic cystopathy. Neurourol Urodyn,2006,25(1):32-8.
75 Soylu A, Akinci A, Yilmaz U, et al. Sympathetic skin responses in Type-1 diabetic children: relationship to urodynamic findings. Neurourol Urodyn, 2006, 25(3):243-8.