發(fā)布時間：2018-06-03 22:19

  本文選題：腎素-血管緊張素系統(tǒng) + 非肽類腎素抑制劑　； 參考：《河北聯(lián)合大學》2014年碩士論文

【摘要】：目的腎素-血管緊張素系統(tǒng)（Renin-angiotensin system，RAS）失調(diào)在高血壓及高血壓腎臟損傷的發(fā)生發(fā)展中起重要作用。腎素（renin）是啟動RAS第一步的限速酶，對RAS的活化有至關(guān)重要的作用。本研究通過觀察阿利克侖（Aliskiren）對人血管緊張素原-腎素（Angiotensinogen-renin，AGT-REN）雙轉(zhuǎn)基因高血壓小鼠血壓、腎損傷指標以及循環(huán)和腎臟局部RAS成分表達的變化，探討非肽類腎素抑制劑阿利克侖對高血壓及腎臟損傷的影響及其作用機制。 方法1動物分組及給藥方法：12只10月齡雄性AGT-REN雙轉(zhuǎn)基因高血壓小鼠隨機分為高血壓對照組（HT組）和阿利克侖治療組（HT+Aliskiren組），每組6只，另選擇背景相同的6只野生型C57B6雄性小鼠作為正常對照組（WT）。給藥方法：單次給藥采用灌胃經(jīng)口途徑，分別給予5mg/kg、10mg/kg、20mg/kg阿利克侖，監(jiān)測給藥后24h內(nèi)血壓；持續(xù)給藥通過皮下埋置微量滲透泵，以20mg/kg/d對HT+Aliskiren組小鼠給予阿利克侖處理28d，其余兩組給予生理鹽水作為空白對照，期間監(jiān)測血壓，28d后取材，進行各項指標檢測。2動脈血壓監(jiān)測：采用無創(chuàng)尾套法測量各組小鼠在清醒與安靜狀態(tài)下給藥前后的平均動脈壓（MAP），觀察阿利克侖單次給藥后24h內(nèi)血壓以及持續(xù)給藥28d內(nèi)隔日血壓的變化。3腎功能損傷指標檢測：①考馬斯亮藍（CBB）結(jié)合法測定24h尿蛋白含量；②全自動生化分析儀測定血清肌酐、尿素；③光鏡（HE、Masson染色）和電鏡觀察腎臟組織病理形態(tài)改變。4氧化應激反應指標檢測：化學發(fā)光法測定腎組織勻漿超氧化物歧化酶（SOD）活性、丙二醛（MDA）含量以及蛋白免疫印跡法（Western blot, WB）檢測腎組織p47phox蛋白表達變化。5RAS成分表達變化檢測：ELISA法測定血清和腎組織勻漿AngⅡ和Ang(1-7)含量；WB檢測腎組織ACE、ACE2蛋白表達。 結(jié)果1阿利克侖的降壓效果：與WT小鼠比較，HT組小鼠MAP明顯升高；單次和持續(xù)給藥實驗均顯示，HT+Aliskiren組小鼠MAP均較HT組明顯降低。2HT小鼠24h尿蛋白、血清尿素、肌酐均較WT小鼠明顯升高；在HT+Aliskiren組小鼠上述腎功能損傷指標較HT組明顯降低。3病理形態(tài)學結(jié)果顯示：光鏡下HT組小鼠腎臟實質(zhì)厚度較WT組明顯變薄，HT+Aliskiren組上述病理損傷得到改善。與WT小鼠比較，HT組小鼠腎小球硬化面積評分和腎間質(zhì)損傷評分明顯增加，HT+Aliskiren組評分結(jié)果較HT組降低。電鏡下，HT小鼠腎小球基底膜增厚，足突融合甚至消失，腎小管上皮細胞胞漿內(nèi)線粒體水腫、空泡變性，間質(zhì)可見大量膠原纖維沉積，；HT+Aliskiren組小鼠腎臟上述病變較HT組明顯減輕。4與WT小鼠比較，HT小鼠腎臟組織SOD活性降低，MDA生成增加，p47phox蛋白表達明顯增多；與HT組小鼠比較，HT+Aliskiren組小鼠腎臟組織SOD活性增加，MDA的生成減少，p47phox蛋白表達明顯減少。5與WT組比較，HT組小鼠ACE蛋白表達增加，ACE2蛋白表達下降，AngⅡ/Ang(1-7)比值增加；與HT組小鼠相比，阿利克侖可下調(diào)HT小鼠腎組織ACE蛋白表達，上調(diào)ACE2表達，降低血漿和腎組織AngⅡ/Ang(1-7)比值。 結(jié)論1阿利克侖具有良好的降壓效果及腎臟保護作用。2阿利克侖通過抑制AngⅡ的生成，降低腎臟局部AngⅡ/Ang(1-7)比值，維持腎臟局部RAS穩(wěn)態(tài)平衡。3腎素抑制劑從源頭上抑制RAS激活，但對ACE2-Ang(1-7)無明顯抑制作用，且可上調(diào)腎臟ACE2蛋白表達，這一作用可能與減少了AngⅡ生成有關(guān)。4阿利克侖可通過抑制NADPH氧化酶表達，，降低氧化應激水平而發(fā)揮腎臟保護作用。
[Abstract]:Objective Renin - angiotensin system ( RAS ) imbalance plays an important role in the development of hypertension and hypertensive kidney injury . Renin is a limiting enzyme for the first step of RAS , which plays an important role in the activation of RAS .

Method 1 Animal grouping and administration were divided into two groups : hypertension control group ( HT group ) and aliskiren group ( HT + Aliskiren group ) .
The mean arterial pressure ( MAP ) before and after administration of 5 mg / kg / d to HT + Aliskiren group was monitored by subcutaneous implantation of micro osmotic pump . The average arterial pressure ( MAP ) before and after administration of the rats in conscious and quiet state was measured by noninvasive method .
( 2 ) measuring serum creatinine and urea by a full - automatic biochemical analyzer ;
The activity of superoxide dismutase ( SOD ) , the content of malondialdehyde ( MDA ) and the expression of p47phox protein were detected by Western blot and WB .
The expression of ACE and ACE2 protein in renal tissue was detected by WB .

Results Compared with WT mice , MAP in HT group increased significantly compared with WT mice .
Both single and continuous administration showed that the MAP in HT + Aliskiren group was significantly lower than that in HT group . 24h urinary protein , serum urea and creatinine of 2HT mice were significantly higher than those in WT mice .
The results showed that the renal parenchyma thickness of HT group mice was significantly lower than that in WT group and HT + Aliskiren group was significantly higher than that in HT group .
Compared with WT mice , the activity of SOD in renal tissue of HT mice decreased , MDA formation increased , and the expression of p47phox protein increased significantly .
Compared with the HT group , the activity of SOD in kidney of HT + Aliskiren group increased , the expression of MDA was decreased , the expression of p47phox protein decreased significantly . Compared with WT group , the expression of ACE protein in HT group increased , the expression of ACE2 protein decreased , and the ratio of Ang 鈪

本文編號：1974430