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[摘要] 目的 探討2型糖尿病胰岛素抵抗大鼠前列腺微血管新生与良性前列腺增生症发生发展的关系。 方法 ①正常对照组大鼠（Normal组，n=15）：8～10周龄正常雄性Wistar大鼠；②单纯良性前列腺增生组大鼠（BPH组，n=15）：手术去势后外源性给予高剂量雄激素；③胰岛素抵抗前列腺增生组大鼠（GK BPH组，n=15）：8～10周龄自发型非肥胖型2型糖尿病Wistar大鼠（GK大鼠），手术去势后外源性给予高剂量雄激素；④胰岛素抵抗前列腺增生血糖干预组大鼠（GK BPH PH组，n=15）：8～10周龄GK大鼠手术去势后外源性给予高剂量雄激素，同时给予盐酸吡格列酮灌胃。光化学法检测大鼠空腹血糖水平；ELISA双抗体夹心法检测大鼠血清胰岛素水平及前列腺DHT含量；real-time PCR法检测大鼠前列腺VEGF、Ang-1、Ang-2 mRNA表达量；免疫组化S-P法检测前列腺组织CD31表达并根据阳性结果计数MVD。 结果 ①GK BPH、GK BPH PH组空腹血糖水平及胰岛素抵抗指数高于Normal组及BPH组（P<0.05）；GK BPH组指标高于GK BPH PH组（P<0.05）；Normal组及BPH组上述指标差异无统计学意义（P>0.05）。BPH组、GK BPH组、GK BPH PH组前列腺组织中DHT含量与Normal组相比明显增高（P<0.05）；GK BPH组、GK BPH PH组高于BPH组，且GK BPH组高于GK BPH PH组（P<0.05）；②各组前列腺组织中VEGF、Ang-1、Ang-2 mRNA表达量及MVD计数结果为GK BPH组>GK BPH PH组>BPH组>Normal组，组间差异有统计学意义（P<0.05）；③GK BPH组、GK BPH PH组大鼠前列腺组织中DHT含量与胰岛素抵抗指数呈正相关（P<0.05），相关性显著。 结论 2型糖尿病伴胰岛素抵抗时，可通过增加前列腺组织DHT的含量从而上调VEGF、Ang-1、Ang-2的表达，促进前列腺组织局部血管新生以加速良性前列腺增生症的进展。
[关键词] 胰岛素抵抗；前列腺增生症；血管内皮生长因子；血管生成素；微血管密度
[中图分类号] R691.9 [文献标识码] A [文章编号] 1673-9701（2018）08-0035-05
Experiment of microvascular angiogenesis of prostate in rats with insulin resistance
MI Yang1 YUAN Xiaobin2 ZHANG Bin2 WANG Dongwen2 ZHANG Xuhui2
1.First School of Clinical Medicine of Shanxi Medical University， Taiyuan 030001， China； 2.Department of Urology， First Hospital of Shanxi Medical University， Taiyuan 030001， China
[Abstract] Objective To investigate the association between the microvascular angiogenesis and the occurrence and progress of benign prostatic hyperplasia（BPH） in type-2 diabetic rats with insulin resistance（IR）. Methods ①Rats of normal group（n=15）： normal male Wistar rats aged 8 to 10 weeks. ②Rats of BPH group（n=15）： high-dose exogenous androgen was given after surgical castration. ③Rats with IR and BPH（GK BPH） group（n=15）： high-dose exogenous androgen was given to spontaneous non-obese type-2 diabetic Wistar rats（GK rats） aged 8 to 10 weeks. ④Rats of IR and BPH with blood glucose intervened（GK BPH PH） group（n=15）： high-dose exogenous androgen was given to GK rats aged 8 to 10 weeks and pioglitazone hydrochloride was given by intragastric administration. The level of fasting blood glucose in rats was assessed by photochemical method. The levels of serum insulin and DHT in prostate of rats were assessed by double antibody sandwich ELISA. The expression of VEGF， Ang-1 and Ang-2 in prostate of rats was assessed by real-time. The expression of CD31 in prostate tissue was assessed by the S-P immunohistochemical method and MVD was counted based on the positive results. Results ①The level of fasting blood glucose and the IR index in GK BPH group and GK BPH PH group were higher than those in normal group and BPH group（P<0.05）. Those in GK BPH group were higher than GK BPH PH group（P<0.05） while there was no statistical difference between normal group and BPH group（P>0.05）. The level of DHT in the prostate tissue of BPH group， GK BPH group and GK BPH PH group was higher than that in normal group（P<0.05）. That in GK BPH group and GK BPH PH group was higher than BPH group and that in GK BPH group was higher than GK BPH PH group（P<0.05）. ②The levels of mRNA of VEGF， Ang-1 and Ang-2 and the MVD count were： GK BPH group>GK BPH PH group>BPH group>Normal group， with statistical differences between each other（P<0.05）. ③The level of DHT in the prostate tissue of GK BPH group and GK BPH PH group was significantly and positively associated with the IR index（P<0.05）. Conclusion When IR occurred in type-2 diabetes， the level of DHT in prostate tissue could be increased to up-regulate the expression of VEGF， Ang-1 and Ang-2 and promote the microvascular angiogenesis in prostate tissue so as to accelerate the progress of benign prostatic hyperplasia. [Key words] Insulin resistance； Prostatic hyperplasia； Vascular endothelial growth factor； Angiogenin； Microvessel density
雄激素是前列腺组织中重要的激素，目前研究认为其与良性前列腺增生症（benign prostatic hyperplasia，BPH）的发生发展密不可分[1]。前列腺血管内皮细胞是雄激素在前列腺组织中发挥生物學效应的主要靶点之一[2，3]，前列腺增生腺体中的血管过度生成可能与腺体的增生密切相关[4]。相关研究已证实，2型糖尿病合并胰岛素抵抗（insulin resistance，IR）患者前列腺生长速度及前列腺体积与单纯BPH患者相比明显增高，但其具体机制尚未明确[5]。本研究通过构建2型糖尿病胰岛素抵抗合并BPH及单纯BPH大鼠模型，比较各模型组与正常大鼠前列腺组织中（dihydrotestosterone，DHT）、血管生成素-1（Angiopoietin-1，Ang-1）、血管生成素-2（Angiopoietin-2，Ang-2）、血管内皮生长因子（vascular endothelial growth factor，VEGF）及微血管密度（microvessel density，MVD）计数差异，探讨2型糖尿病胰岛素抵抗与BPH发生发展的相关性。
1材料与方法
1.1 实验动物及饲养条件
2型糖尿病胰岛素抵抗大鼠模型：使用8～10周龄，体重250～300 g，SPF级雄性非胰岛素依赖型非肥胖型2型糖尿病Wistar大鼠（GK大鼠），购自于江苏省常州市卡文斯实验动物有限公司，许可证号SCXK（苏）2011-003；其余各组选用同周龄SPF级雄性Wistar大鼠，购自于山西医科大学动物实验中心，许可证号：SYXK（晋）2003-013。所有大鼠均饲养于山西医科大学动物实验中心SPF级实验室，标准化饲养房喂养，饲喂标准颗粒饲料，自由饮水。饲养环境：T 18℃～22℃，H 50%～70%，日照时间为8：00-20：00。
1.2 主要试剂
盐酸吡格列酮（Sigma公司，PHR1632）；丙酸睾酮（Sigma公司，BP720）；苯甲酸雌二醇（Sigma公司，E8875）；大鼠胰岛素ELISA定量试剂盒（Thermo Fisher公司，ERINS）；大鼠DHT ELISA定量试剂盒（IBL公司，DB52021）；兔抗大鼠CD31单克隆抗体（R
[关键词] 胰岛素抵抗；前列腺增生症；血管内皮生长因子；血管生成素；微血管密度
[中图分类号] R691.9 [文献标识码] A [文章编号] 1673-9701（2018）08-0035-05
Experiment of microvascular angiogenesis of prostate in rats with insulin resistance
MI Yang1 YUAN Xiaobin2 ZHANG Bin2 WANG Dongwen2 ZHANG Xuhui2
1.First School of Clinical Medicine of Shanxi Medical University， Taiyuan 030001， China； 2.Department of Urology， First Hospital of Shanxi Medical University， Taiyuan 030001， China
[Abstract] Objective To investigate the association between the microvascular angiogenesis and the occurrence and progress of benign prostatic hyperplasia（BPH） in type-2 diabetic rats with insulin resistance（IR）. Methods ①Rats of normal group（n=15）： normal male Wistar rats aged 8 to 10 weeks. ②Rats of BPH group（n=15）： high-dose exogenous androgen was given after surgical castration. ③Rats with IR and BPH（GK BPH） group（n=15）： high-dose exogenous androgen was given to spontaneous non-obese type-2 diabetic Wistar rats（GK rats） aged 8 to 10 weeks. ④Rats of IR and BPH with blood glucose intervened（GK BPH PH） group（n=15）： high-dose exogenous androgen was given to GK rats aged 8 to 10 weeks and pioglitazone hydrochloride was given by intragastric administration. The level of fasting blood glucose in rats was assessed by photochemical method. The levels of serum insulin and DHT in prostate of rats were assessed by double antibody sandwich ELISA. The expression of VEGF， Ang-1 and Ang-2 in prostate of rats was assessed by real-time. The expression of CD31 in prostate tissue was assessed by the S-P immunohistochemical method and MVD was counted based on the positive results. Results ①The level of fasting blood glucose and the IR index in GK BPH group and GK BPH PH group were higher than those in normal group and BPH group（P<0.05）. Those in GK BPH group were higher than GK BPH PH group（P<0.05） while there was no statistical difference between normal group and BPH group（P>0.05）. The level of DHT in the prostate tissue of BPH group， GK BPH group and GK BPH PH group was higher than that in normal group（P<0.05）. That in GK BPH group and GK BPH PH group was higher than BPH group and that in GK BPH group was higher than GK BPH PH group（P<0.05）. ②The levels of mRNA of VEGF， Ang-1 and Ang-2 and the MVD count were： GK BPH group>GK BPH PH group>BPH group>Normal group， with statistical differences between each other（P<0.05）. ③The level of DHT in the prostate tissue of GK BPH group and GK BPH PH group was significantly and positively associated with the IR index（P<0.05）. Conclusion When IR occurred in type-2 diabetes， the level of DHT in prostate tissue could be increased to up-regulate the expression of VEGF， Ang-1 and Ang-2 and promote the microvascular angiogenesis in prostate tissue so as to accelerate the progress of benign prostatic hyperplasia. [Key words] Insulin resistance； Prostatic hyperplasia； Vascular endothelial growth factor； Angiogenin； Microvessel density
雄激素是前列腺组织中重要的激素，目前研究认为其与良性前列腺增生症（benign prostatic hyperplasia，BPH）的发生发展密不可分[1]。前列腺血管内皮细胞是雄激素在前列腺组织中发挥生物學效应的主要靶点之一[2，3]，前列腺增生腺体中的血管过度生成可能与腺体的增生密切相关[4]。相关研究已证实，2型糖尿病合并胰岛素抵抗（insulin resistance，IR）患者前列腺生长速度及前列腺体积与单纯BPH患者相比明显增高，但其具体机制尚未明确[5]。本研究通过构建2型糖尿病胰岛素抵抗合并BPH及单纯BPH大鼠模型，比较各模型组与正常大鼠前列腺组织中（dihydrotestosterone，DHT）、血管生成素-1（Angiopoietin-1，Ang-1）、血管生成素-2（Angiopoietin-2，Ang-2）、血管内皮生长因子（vascular endothelial growth factor，VEGF）及微血管密度（microvessel density，MVD）计数差异，探讨2型糖尿病胰岛素抵抗与BPH发生发展的相关性。
1材料与方法
1.1 实验动物及饲养条件
2型糖尿病胰岛素抵抗大鼠模型：使用8～10周龄，体重250～300 g，SPF级雄性非胰岛素依赖型非肥胖型2型糖尿病Wistar大鼠（GK大鼠），购自于江苏省常州市卡文斯实验动物有限公司，许可证号SCXK（苏）2011-003；其余各组选用同周龄SPF级雄性Wistar大鼠，购自于山西医科大学动物实验中心，许可证号：SYXK（晋）2003-013。所有大鼠均饲养于山西医科大学动物实验中心SPF级实验室，标准化饲养房喂养，饲喂标准颗粒饲料，自由饮水。饲养环境：T 18℃～22℃，H 50%～70%，日照时间为8：00-20：00。
1.2 主要试剂
盐酸吡格列酮（Sigma公司，PHR1632）；丙酸睾酮（Sigma公司，BP720）；苯甲酸雌二醇（Sigma公司，E8875）；大鼠胰岛素ELISA定量试剂盒（Thermo Fisher公司，ERINS）；大鼠DHT ELISA定量试剂盒（IBL公司，DB52021）；兔抗大鼠CD31单克隆抗体（R