本文關(guān)鍵詞： 肥胖相關(guān)性腎病 過氧化物酶體增殖物激活受體-γ 脂聯(lián)素 氧化應(yīng)激 AMPK NOX-4 小鼠　出處：《天津醫(yī)科大學(xué)》2015年碩士論文　論文類型：學(xué)位論文

【摘要】：目的:肥胖越來越被認(rèn)為是腎臟疾病的主要危險(xiǎn)因素。我們之前的研究表明PPARγ激動(dòng)劑吡格列酮可通過升高單純肥胖小鼠血清脂聯(lián)素水平而起到腎臟保護(hù)作用,而有研究表明脂聯(lián)素有抗氧化應(yīng)激作用,但PPARγ激動(dòng)劑的腎臟保護(hù)作用是否與抗氧化應(yīng)激有關(guān)尚不清楚。本研究探討PPARγ激動(dòng)劑對(duì)單純肥胖小鼠腎組織AMPK的影響及其抗氧化應(yīng)激作用。方法:6周齡雄性ob/ob小鼠24只和其同系C57BL/6J小鼠8只,適應(yīng)性飼養(yǎng)2周,測(cè)量體重并留尿測(cè)量24h尿微量白蛋白,確定ob/ob小鼠體重及24h尿微量白蛋白與C57BL/6J小鼠有顯著差異。按照體質(zhì)量分層抽樣,將ob/ob小鼠隨機(jī)分為3組:模型組(M組,n=8)、小劑量治療組(T1組,n=8)、大劑量治療組(T2組,n=8);C57BL/6J小鼠作為對(duì)照組(C組,n-8)。治療組分別給予吡格列酮灌胃,其中T1組為吡格列酮2.25mg·kg-1·d-1,T2組為吡格列酮6.75mg·kg-1·d-1,C組和M組小鼠給予等體積蒸餾水灌胃。每周稱量小鼠體重一次,根據(jù)體重調(diào)整給藥劑量,連續(xù)12周。灌胃前后分別采集血液標(biāo)本以測(cè)定小鼠血糖,血清ADP,并留尿液測(cè)定24小時(shí)尿m ALB。12周后處死小鼠,摘取腎臟,稱量右腎濕重。光鏡下觀察小鼠腎臟病理改變,并測(cè)量腎小球直徑;免疫組化方法檢測(cè)腎組織內(nèi)AMPK及NOX-4的表達(dá)。比較四組小鼠各個(gè)指標(biāo)間的差異,并將血清ADP、24h尿微量白蛋白、腎組織AMPK及NOX4表達(dá)水平分別與其他指標(biāo)做相關(guān)性分析。采用SPSS 17.0統(tǒng)計(jì)軟件進(jìn)行數(shù)據(jù)處理和統(tǒng)計(jì)學(xué)分析,P0.05為差異具有統(tǒng)計(jì)學(xué)意義。結(jié)果:1小鼠的一般情況1.1實(shí)驗(yàn)前兩種小鼠體重及尿蛋白的比較:6周齡時(shí)ob/ob小鼠平均體質(zhì)量已明顯高于C57BL/6J小鼠(P0.01),適應(yīng)性飼養(yǎng)2周后,ob/ob小鼠體質(zhì)量不僅明顯高于C57BL/6J小鼠(P0.01),且平均體重已經(jīng)超過后者的50%(20%),即達(dá)到小鼠肥胖標(biāo)準(zhǔn),ob/ob小鼠尿微量白蛋白明顯高于C57BL/6J小鼠(P0.01),雖然兩種小鼠血糖存在差異(P0.05),但都尚未達(dá)到糖尿病標(biāo)準(zhǔn),可排除因糖尿病導(dǎo)致的蛋白尿,確定ob/ob小鼠蛋白尿是由肥胖造成。1.2各組小鼠實(shí)驗(yàn)前后體重:分組后M組和T1、T2組小鼠體質(zhì)量均超過C組小鼠平均體質(zhì)量的20%,即達(dá)到小鼠肥胖標(biāo)準(zhǔn)(P0.01),M組、T1組、T2組小鼠體質(zhì)量無顯著差異(P0.05)。實(shí)驗(yàn)結(jié)束時(shí)M組和T1、T2組小鼠體質(zhì)量超過C組小鼠更為明顯(P0.01),T2組較T1組小鼠體質(zhì)量高(P0.01)。各組小鼠實(shí)驗(yàn)后體質(zhì)量均高于實(shí)驗(yàn)前(P0.01)2血清脂聯(lián)素濃度:2.1各組前后比較:實(shí)驗(yàn)后M組和T1、T2組小鼠血清脂聯(lián)素水平均較實(shí)驗(yàn)前降低(P0.01),C組前后無明顯變化(P0.05)。2.2各組之間比較:實(shí)驗(yàn)前M組和T1、T2組小鼠血清ADP較C組降低(P0.01),實(shí)驗(yàn)后T1、T2組小鼠血清ADP高于M組(P0.01),且T2組高于T1組(P0.01)。3 24h尿微量白蛋白:3.1各組前后比較:實(shí)驗(yàn)后M組和T1、T2組小鼠24h尿微量白蛋白均較實(shí)驗(yàn)前增加(P0.01),C組前后無明顯變化(P0.05)。3.2各組之間比較:實(shí)驗(yàn)前后M組和T1、T2組小鼠24h尿微量白蛋白均明顯高于C組(P0.01);實(shí)驗(yàn)后T1、T2組小鼠24h尿微量白蛋白較M組明顯降低(P0.01),且T2組明顯低于T1組(P0.01)。4腎臟病理改變:4.1腎臟濕重(右腎):實(shí)驗(yàn)后M組和T1、T2組小鼠腎臟濕重明顯高于C組(P0.05),T1組及T2組右腎濕重明顯小于M組(P0.01),T2組右腎濕重明顯小于T1組(P0.01)。4.2腎組織病理改變:M組小鼠腎小球鮑曼氏囊增寬,基底膜增厚,系膜細(xì)胞和系膜基質(zhì)增生,腎小球體積增大,部分伴有局灶節(jié)段性腎小球硬化,輕度腎小管萎縮和間質(zhì)纖維化。T1、T2組病變程度較M組減輕。4.3腎小球直徑:實(shí)驗(yàn)后M組和T1、T2組小鼠腎小球直徑明顯大于C組(P0.05),T1組及T2組腎小球直徑明顯小于M組(P0.01),T2組腎小球直徑明顯小于T1組(P0.01)。5腎組織AMPK表達(dá):可見AMPK在腎小球、腎小管及腎間質(zhì)均有表達(dá);M組和T1、T2組小鼠腎組織AMPK表達(dá)較C組明顯減少(P0.01),T1、T2組小鼠腎組織AMPK表達(dá)明顯高于M組(P0.01),T2組明顯高于T1組(P0.05)。6腎組織NOX-4表達(dá):可見NOX-4在腎小球、腎小管及腎間質(zhì)均有表達(dá);M組和T1、T2組小鼠腎組織NOX-4表達(dá)明顯高于C組(P0.01),T1、T2組小鼠腎組織NOX-4表達(dá)明顯低于M組(P0.01),T2組明顯低于T1組(P0.05)。7直線相關(guān)分析:7.1尿微量白蛋白與體質(zhì)量、腎小球直徑、右腎濕重及NOX-4表達(dá)水平呈正相關(guān)(r1=0.741;r2=0.936;r3=0.877;r4=0.878,P0.001),與血清ADP及AMPK表達(dá)水平呈負(fù)相關(guān)(r5=-0.915;r6=-0.759,P0.001)。7.2小鼠血清脂聯(lián)素與體質(zhì)量、腎小球直徑、右腎濕重、尿微量白蛋白、NOX-4呈負(fù)相關(guān)(r7=-0.772;r8=-0.821;r5=-0.915;r9=-0.958,P0.001),與AMPK表達(dá)水平呈正相關(guān)(r10=0.878,P0.001)。7.3腎組織AMPK表達(dá)水平與體質(zhì)量、腎小球直徑、右腎濕重、尿微量白蛋白及腎組織NOX4表達(dá)水平呈負(fù)相關(guān)(r11=-0.627;r12=-0.711;r13=-0.743;r6=-0.759;r14=-0.884,P0.001),與血清脂聯(lián)素濃度呈正相關(guān)(r10=0.878,P0.001)。7.4腎組織NOX-4表達(dá)水平與體質(zhì)量、腎小球直徑、右腎濕重、尿微量白蛋白呈正相關(guān)(r15=0.671;r16=0.843;r17=0.794;r4=0.878,P0.001),與血清脂聯(lián)素濃度及腎組織AMPK表達(dá)水平呈負(fù)相關(guān)(r9=-0.958;r14=-0.884,P0.001)。結(jié)論:1.肥胖小鼠血清ADP水平下降,腎組織AMPK表達(dá)降低,NOX-4表達(dá)增加,氧化應(yīng)激增強(qiáng),尿m ALB增加。2.PPARγ激動(dòng)劑吡格列酮可升高肥胖小鼠血清ADP水平及腎組織AMPK表達(dá),減少NOX-4表達(dá),改善肥胖小鼠的氧化應(yīng)激狀態(tài),從而減少尿蛋白,起到腎臟保護(hù)作用。3.相對(duì)于小劑量PPARγ激動(dòng)劑,大劑量PPARγ激動(dòng)劑更能在肥胖時(shí)對(duì)腎臟起到保護(hù)作用。
[Abstract]:Objective: obesity is increasingly considered as major risk factors for kidney disease. Our previous study showed that PPAR gamma agonist pioglitazone by elevated serum adiponectin levels in simple obese mice and play a role in renal protection, and studies have shown that adiponectin has anti oxidative stress, but the PPAR gamma agonist renal protective effect and oxidative stress is unclear. This study investigated the PPAR gamma agonist on the influence of obesity mice kidney AMPK and oxidative stress. Methods: 6 Zhou Lingxiong 24 ob/ob mice and its homologous C57BL/6J 8 rats, 2 week adaptive feeding, measuring the weight and keep the microalbuminuria measurement of urinary 24h, determine the weight of ob/ob mice 24h and microalbuminuria and C57BL/6J mice have significant differences. According to the body weight of stratified sampling, the ob/ob mice were randomly divided into 3 groups: model group (group M, n=8), low dose treatment Group (group T1, n=8), high dose treatment group (group T2, n=8); C57BL/6J mice as control group (group C, n-8). The treatment group were given pioglitazone orally, group T1, 2.25mg - kg-1 - D-1 pioglitazone group, T2 6.75mg kg-1 D-1 pioglitazone group, C and M mice were given the same volume of distilled water. The weight of mice weighed every week once, according to the weight to adjust the dosage for 12 weeks. Blood samples were collected before and after intragastric administration with serum glucose, serum ADP, m were determined in 24 hour urine urine ALB.12 weeks after the mice were sacrificed, the removal of the kidney, called the right amount the wet weight of kidney. Renal pathological changes were observed under light microscope, and measured the expression of glomerular diameter; immunohistochemical method of AMPK and NOX-4 in renal tissue. The difference of each index in the detection of mice between four groups, and the serum ADP, 24h urinary albumin, renal tissue AMPK and NOX4 levels and other indicators respectively. Do. Correlation analysis. Data processing and statistical analysis were conducted using SPSS 17 statistical software for P0.05, the difference was statistically significant. Results: comparison of body weight and urine protein in mice 1 mice in general 1.1 before the experiment two: 6 week old ob/ob mice have significantly higher than the average body weight of C57BL/6J mice (P0.01) after 2 weeks. Adaptive feeding, the body weight of ob/ob mice not only significantly higher than that of C57BL/6J mice (P0.01), and the average weight of more than 50% of the latter (20%), which reached the mouse obese standard, ob/ob mice urinary albumin was significantly higher than that of C57BL/6J mice (P0.01), while the two differences of blood glucose in mice (P0.05), but have not yet reached the standard of diabetes that can be excluded because of diabetes leads to proteinuria, proteinuria is determined ob/ob mice.1.2 mice before and after the experiment the weight caused by obesity: M group and T1 group, the body mass of mice in T2 group were higher than the average in C group 浣撹川閲忕殑20%,鍗寵揪鍒板皬榧犺偉鑳栨爣鍑，

本文編號(hào)：1535055