缬沙坦对自发性高血压大鼠肾脏水通道蛋白-1、2、5表达的影响

发布时间：2018-03-25 05:29

本文选题：自发性高血压　切入点：缬沙坦　出处：《遵义医学院》2017年硕士论文

【摘要】：目的:观察缬沙坦对自发性高血压大鼠(SHR)血压的影响,并检测大鼠肾脏水通道蛋白-1、2、5(AQP1、AQP2、AQP5)m RNA和蛋白的表达变化,为缬沙坦治疗高血压的药效学机制提供新的理论依据,并为抗高血压药的研发提供可能的作用靶点。方法:自发性高血压大鼠(SHR)和Wistar Kyoto(WKY)大鼠分别12只,随机分为缬沙坦灌胃组(SHR-V组、WKY-V组)和生理盐水灌胃组(SHR-N组、WKY-N组),每组各6只。灌胃剂量:缬沙坦为每天10mg/kg体重,生理盐水为换算出的不含缬沙坦的相应体积。每周称量大鼠体重并以此调整一次药物剂量,用无创血压仪测量尾动脉收缩压一次,持续至第40周时腹主动脉采血并收集肾脏,用ELISA和化学发光法分别测定血浆精氨酸加压素(AVP)和血管紧张素Ⅱ(AngⅡ)浓度,用苏木素-伊红(HE)染色,观察各组大鼠肾脏形态学变化,用实时荧光定量PCR和Western blot分别检测肾脏AQP1,2,5 m RNA和蛋白的表达变化,用免疫组织化学(IHC)法观测肾脏AQP1,2,5蛋白的定位和表达。结果:1.SHR血压值在第12周开始灌胃时明显高于WKY大鼠;在第16周时SHR-V组血压值比SHR-N组已有明显降低(P0.05);第40周时,血压进一步降低,但仍明显高于WKY-N组和WKY-V组,而WKY-N组与WKY-V组之间无明显差异(P0.05)。2.第40周血浆AngⅡ浓度:SHR-V组比SHR-N组、WKY-V比WKY-N组均明显升高(P0.05);SHR-N组比WKY-N组、SHR-V组比WKY-V组均略有升高,但无统计学意义。血浆AVP浓度:SHR-V组比SHR-N组明显降低(P0.05);WKY-V组比WKY-N组明显升高(P0.05);而SHR-N组比WKY-N组、SHR-V组比WKY-V组均明显降低(P0.05)。3.第40周肾脏HE染色显示,WKY-N组与WHY-V组肾小球毛细血管壁薄,基底膜未见明显增厚,肾小管腔刷状缘及肾小囊腔清晰可见。SHR-N组可见部分肾小球增大,肾小管管腔及毛细血管扩张,部分肾小球萎缩,囊腔内分叶状增多,毛细血管结构不清,系膜细胞及基质增生,肾小管间质增多,管腔变窄,SHR-V组中上述病变明显减轻。4.第40周肾脏RT-PCR结果显示,SHR-V组比SHR-N组AQP1,2,5 m RNA相对表达量均明显升高(P0.05);WKY-V组比WKY-N组均无明显改变(P0.05);SHR-N组比WKY-N组AQP1,5明显降低(P0.05),而AQP2则明显升高(P0.05);SHR-V组比WKY-V组AQP1,5无明显变化(P0.05),而AQP2则明显升高(P0.05)。5.第40周肾脏Western blot结果显示,SHR-V组比SHR-N组AQP1,2,5相对灰度值均明显升高(P0.05);WKY-V组比WKY-N组AQP1,2,5均无明显改变(P0.05);SHR-N组比WKY-N组AQP1,5明显降低(P0.05),而AQP2则无明显改变(P0.05);SHR-V组比WKY-V组AQP1,2,5均无明显改变(P0.05)。6.第40周肾脏IHC结果显示,AQP1在皮质主要表达于近端小管,在髓质主要表达于髓袢降支细段,WKY-N组平均光密度值皮质明显高于髓质(P0.05),WKY-V组以及SHR-V组皮质均明显低于髓质(P0.05),SHR-N组则无明显差异(P0.05);AQP2在皮质主要表达于连接小管和皮质集合管,在髓质主要表达于髓质集合管,WKY-N组、WKY-V组、SHR-N组以及SHR-V组皮质平均光密度值均明显低于髓质(P0.05);AQP5在皮质主要表达于连接小管和皮质集合管,在髓质主要表达于髓质集合管,WKY-N组、WKY-V组、SHR-N组以及SHR-V组皮质平均光密度值均明显低于髓质(P0.05)。结论:1.在自发性高血压后期,SHR肾脏内局部RAS更加活跃,AVP的体液调节作用相对增强,缬沙坦可以使SHR的体液调节效应进一步加强。2.血压的长期变化过程会引起大鼠肾脏AQP1在皮质与髓质间的重新分布,缬沙坦可以改变SHR AQP1皮质与髓质间的平均分布状态,使髓质表达增多。3.AQP2在大鼠肾脏的分布皮质低于髓质,缬沙坦能够显著提高SHR肾脏内AQP2 m RNA的表达,而蛋白表达升高相对缓慢。4.AQP5在大鼠肾内的分布皮质低于髓质,缬沙坦能显著提高SHR肾脏AQP5m RNA和蛋白的表达量。
[Abstract]:Objective: To observe the effect of valsartan on spontaneously hypertensive rats (SHR) the influence of blood pressure, and the detection of renal aquaporins in rats with -1,2,5 (AQP1, AQP2, AQP5) expression of M protein and RNA, provide a new theoretical basis for the efficacy of valsartan in the treatment of hypertension mechanism and provide potential targets for the development of resistance the treatment of hypertension. Methods: spontaneously hypertensive rats (SHR) and Wistar Kyoto (WKY) rats were 12 rats were randomly divided into valsartan gavage group (SHR-V group, WKY-V group) and saline group (SHR-N group, WKY-N group), 6 rats in each group. Intragastric dose of valsartan for: every 10mg/kg weight, normal saline was calculated without the corresponding volume weight per week. Valsartan weighed rats and to adjust doses, with non-invasive blood pressure measurement of systolic blood pressure once, continued to fortieth weeks of abdominal aorta and kidney were collected by ELISA and chemical. Measured plasma arginine vasopressin chemiluminescence (AVP) and angiotensin II (Ang II) concentration, with hematoxylin eosin (HE) staining, observe the morphological changes of kidney of rats were detected the expression change of M renal AQP1,2,5 RNA and protein by real-time quantitative PCR and Western blot, with immunohistochemical (IHC) localization and expression of AQP1,2,5 protein in kidney was observed. Results: 1.SHR blood pressure values in twelfth week mice was significantly higher than that of WKY rats; at sixteenth weeks, the blood pressure in the SHR-V group was significantly lower than SHR-N group (P0.05); for fortieth weeks, the blood pressure decreased further, but still significantly higher than WKY-N group and WKY-V group, but no significant difference between group WKY-N and group WKY-V (P0.05).2. fortieth week plasma Ang II: SHR-V group than in the SHR-N group, the WKY-V ratio was significantly increased in the WKY-N group (P0.05); SHR-N group than in the WKY-N group, SHR-V group than in the WKY-V group were slightly increased, but no statistical significance Yi. The concentration of plasma AVP in SHR-V group was significantly lower than that of group SHR-N (P0.05); WKY-V group was significantly higher than that in WKY-N group (P0.05); and SHR-N group than in the WKY-N group, SHR-V group than in WKY-V group was significantly lower (P0.05).3. fortieth weeks of renal HE staining showed that WKY-N group and WHY-V group glomerular capillary wall thin basement membrane was thicker, tubular brush border and renal cysts visible.SHR-N group showed partial glomerular enlargement, renal tubular lumen and telangiectasia, partial glomerular atrophy, lobulated cystic cavity increased, capillary structure is not clear, mesangial cells and matrix hyperplasia, tubulointerstitial increased lumen narrowing in the SHR-V group, the lesion was reduced fortieth weeks of.4. kidney RT-PCR results showed that the SHR-V AQP1,2,5 M group than in the SHR-N group the relative expression of RNA were significantly increased (P0.05); WKY-V group than in the WKY-N group showed no significant change (P0.05); SHR-N group was significantly reduced than the WKY-N group AQP1,5 Low (P0.05), while AQP2 increased significantly (P0.05); SHR-V group than in the WKY-V group there was no significant change in AQP1,5 (P0.05), and AQP2 (P0.05).5. significantly increased fortieth weeks of renal Western blot results showed that the SHR-V group than in the SHR-N group relative gray scale values of AQP1,2,5 were significantly increased (P0.05); group WKY-V than WKY-N group AQP1,2,5 showed no significant change (P0.05); SHR-N group was significantly lower than in group WKY-N (P0.05), AQP1,5 and AQP2 were not significantly changed (P0.05); SHR-V group than in WKY-V group AQP1,2,5 showed no significant change (P0.05.6.) fortieth weeks of renal IHC showed that AQP1 was mainly expressed in the cortex in the proximal tubule, in the medulla is mainly expressed in the thin descending limb of Henle, WKY-N group was significantly higher than that of the average optical density of cortical medulla (P0.05), WKY-V group and SHR-V group were significantly lower than that of the medulla cortex (P0.05), there was no obvious difference between the SHR-N group (P0.05); AQP2 was mainly expressed in the cortex in the connecting tubule and cortical collecting duct in the medulla the main The expression in the medullary collecting duct, WKY-N group, WKY-V group, SHR-N group and SHR-V group, the average optical density value of cortex was significantly lower than that of the medulla (P0.05); AQP5 was mainly expressed in the cortex in the connecting tubule and cortical collecting duct in the medulla is mainly expressed in the medullary collecting duct, WKY-N group, WKY-V group, SHR-N group and SHR-V group of cortex the average optical density values were significantly lower than that of the medulla (P0.05). Conclusion: 1. in the late SHR in the kidney of spontaneously hypertensive, local RAS was more active, relatively enhanced AVP humoral regulation effect of valsartan can make SHR humoral regulation effect to further strengthen the long-term changes of blood pressure of.2. process will cause redistribution in AQP1 rat kidney cortex and medulla the average distribution of valsartan can change the SHR AQP1 between the cortex and medulla, the increased expression of.3.AQP2 in cortex medulla distribution of rat kidney medulla below, valsartan can significantly improve SHR in the kidney of AQP 2, the expression of M RNA was increased, while protein expression increased slowly. The distribution of.4.AQP5 in rat kidneys was lower than that in medulla. Valsartan significantly increased the expression of AQP5m RNA and protein in SHR kidney.

【学位授予单位】：遵义医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R544.1

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