發(fā)布時間：2018-04-05 22:06

  本文選題：心房顫動　切入點(diǎn)：糖尿病　出處：《天津醫(yī)科大學(xué)》2017年博士論文

【摘要】：目的:糖尿病是心房顫動(房顫)的獨(dú)立的危險因素,但目前其發(fā)病機(jī)制尚不明確。最新證據(jù)顯示,線粒體功能障礙參與了房顫發(fā)生的病理生理過程。二肽基肽酶-4(DPP-4)抑制劑被發(fā)現(xiàn)具有改善線粒體功能的作用。在本研究中,我們探討DPP-4抑制劑阿格列汀對心房線粒體重構(gòu)的影響,以及對與代謝應(yīng)激相關(guān)房顫的預(yù)防作用。方法:健康日本大耳白兔用于建立家兔糖尿病模型,隨機(jī)分為三組:對照組(Control,C組),經(jīng)耳緣靜脈注射生理鹽水4ml;糖尿病組(Diabetes Mellitus,D組),經(jīng)耳緣靜脈注射四氧嘧啶120mg/kg;糖尿病+阿格列汀組(Diabetes+Alogliptin,DA組),口服阿格列汀12.5mg/Kg/d。每組30只,自由飲食,飼養(yǎng)8周。造模成功后,各組中隨機(jī)抽取10只用于以下三部分實(shí)驗(yàn):一、心臟超聲、血流動力學(xué)、血液學(xué)檢查、病理學(xué)和分子生物學(xué)實(shí)驗(yàn):心臟超聲測定各心腔大小、心室壁厚度及射血分?jǐn)?shù);血流動力學(xué)測定主動脈內(nèi)壓、心室內(nèi)壓及心室內(nèi)壓最大上升和下降速率(±dp/dtmax);血液學(xué)檢查測定包括部分生化指標(biāo)、胰島素(INS)、胰高血糖素樣肽-1(GLP-1),以及超氧化物歧化酶(SOD)、超敏C反應(yīng)蛋白(hs-CRP)、丙二醛(MDA)、8-羥基脫氧鳥苷(8-OHDG)等氧化應(yīng)激和炎癥指標(biāo);病理學(xué)檢查應(yīng)用HE和Masson染色評價心肌細(xì)胞橫截面積和膠原容積分?jǐn)?shù),透射電鏡觀察細(xì)胞核、肌絲及線粒體形態(tài)學(xué);分子生物學(xué)應(yīng)用Western blot方法測定TGF-β1、NF-κBp65蛋白表達(dá)水平,以及線粒體生物合成信號通路、動力學(xué)相關(guān)蛋白和ATP合酶亞基ATP5A的蛋白的表達(dá),應(yīng)用RT-PCR的方法測定電子傳遞鏈復(fù)合體Ⅰ-Ⅴ亞基的m RNA水平,以及PGC-1α、NRF-1、Tfam的m RNA表達(dá)水平。二、心臟電生理實(shí)驗(yàn):利用Langendorff離體灌流模型測定心房各點(diǎn)的有效不應(yīng)期(AERP)、有效不應(yīng)期離散度(AERPD)、房間傳導(dǎo)時間(IACT),計算房顫誘發(fā)率。三、線粒體功能實(shí)驗(yàn),測定線粒體呼吸控制率(RCR)、膜電位(Δψ)及ROS生成速率。結(jié)果:1.心臟超聲結(jié)果顯示,與C組相比,D組和DA組左心房內(nèi)徑(LAD)、室間隔厚度(IVS)和左室后壁(PWLV)均增加(P0.01),而在DA組LAD、IVS及PWLV較D組降低(P0.01);三組間EF值及血流動力學(xué)參數(shù)無統(tǒng)計學(xué)差異。2.與C組相比,D組血清INS(8.01±1.66 mmol/L vs 17.36±3.35 mmol/L,P0.01)和GLP-1(0.35±0.07 pmol/L vs 1.21±0.38 pmol/L,P0.01)水平降低,而阿格列汀治療后GLP-1的濃度(0.80±0.19 pmol/L vs 0.35±0.07 pmol/L,P0.01)較D組明顯升高。3.D組與C組相比,MDA、8-OHDG和hs-CRP濃度升高(P0.01),而SOD活性降低(P0.05),阿格列汀治療后降低了D組MDA、8-OHDG和hs-CRP的濃度(P0.01),但對SOD活性無明顯影響。4.與C組相比,D組心肌橫截面積、膠原容積分?jǐn)?shù)增大(P0.05),而阿格列汀治療后顯著抑制了這種病理改變(P0.05)。5.電生理研究顯示,D組AERPD較C組和DA組明顯延長(P0.01);D組IACT在基礎(chǔ)起搏周長150ms、200ms及250ms時較C組和DA組均明顯延長(P0.01)。與C組相比,D組房顫的誘發(fā)率從1.33%上升至5.56%,而DA組房顫的誘發(fā)率為2.44%,升高幅度明顯低于D組(P0.05)。6.線粒體功能實(shí)驗(yàn)顯示,D組RCR值及ΔΨm較C組明顯降低(P0.05),而阿格列汀治療后逆轉(zhuǎn)了這種病例生理改變(P0.05或0.01);線粒體ROS生成速率在D組明顯高于C組(P0.05),阿格列汀顯著抑制了糖尿病線粒體ROS的生成速率(P0.05)。7.Western blot結(jié)果顯示,D組TGF-β1和NF-κBp65蛋白表達(dá)水平明顯高于C組,阿格列汀治療后顯著抑制了這兩種蛋白的表達(dá);D組線粒體生物合成相關(guān)蛋白APN、p AMPK、PGC-1α、NRF1、Tfam蛋白表達(dá)水平較C組下調(diào);而與D組比較,DA組中這些蛋白表達(dá)水平均上調(diào);D組線粒體分裂蛋白Drp1、融合蛋白Mfn1及OPA1表達(dá)下調(diào),而DA組中蛋白表達(dá)水平較D組均上調(diào);三組間AMPK和ATP5A的蛋白表達(dá)水平無顯著差異。8.RT-PCR實(shí)驗(yàn),D組PGC-1α和Tfam的m RNA表達(dá)水平低于C組和DA組,而NRF1則在三組間無差異;D組復(fù)合體Ⅰ、Ⅱ、Ⅳ亞基NDUFB8、SDHB、MTCO1的m RNA表達(dá)水平明顯低于C組和DA組,而D組復(fù)合體Ⅲ亞基UQCRC2的m RNA表達(dá)水平高于C組和DA組;三組間ATP5A的m RNA表達(dá)水平無統(tǒng)計學(xué)差異。結(jié)論:DPP-4抑制劑改善糖尿病心房基質(zhì),對房顫具有潛在的預(yù)防作用,其機(jī)制涉及減輕線粒體氧化應(yīng)激,改善線粒體呼吸功能和促進(jìn)線粒體生物合成。
[Abstract]:Objective: diabetes is atrial fibrillation (AF) independent risk factors, but its pathogenesis is still not clear. Recent evidence suggests that mitochondrial dysfunction is involved in the pathophysiology of atrial fibrillation. Two dipeptidyl peptidase -4 inhibitors (DPP-4) were found to have improved mitochondrial function. In this study, we investigate the effects of DPP-4 inhibitors on atrial remodeling in A Glenn Dean mitochondria associated with metabolic stress, as well as for the prevention of atrial fibrillation. Methods: the rabbits were used to establish the rabbit model of diabetes, were randomly divided into three groups: control group (Control, group C), intravenous injection of saline 4ml the diabetic group (Diabetes; Mellitus, D group), intravenous injection of alloxan diabetes + four 120mg/kg; A Glenn Dean group (Diabetes+Alogliptin, group DA), oral A Glenn Dean 12.5mg/Kg/d. each group had 30 rats, free diet, feeding 8 weeks made. After the success of the model in each group, 10 rats were randomly selected for the following three parts: an experiment, echocardiography, hemodynamics, blood examination, pathology and molecular biology experiment: cardiac ultrasound determination of size of the cardiac chambers, ventricular wall thickness and ejection fraction; hemodynamic measurements of aortic pressure, intraventricular pressure and ventricular the maximum rate of pressure rise and fall (+ dp/dtmax); blood tests including determination of some biochemical indexes (INS), insulin, glucagon like peptide -1 (GLP-1), and superoxide dismutase (SOD), high sensitive C reactive protein (hs-CRP), malondialdehyde (MDA), 8- hydroxy deoxyguanosine glucoside (8-OHDG), oxidative stress and inflammation index; pathological examination by HE and Masson staining were used to evaluate the myocardial cell cross-sectional area and collagen volume fraction, TEM observation of nuclear, myofilament and mitochondrial morphology; molecular biology application of Western blot method for the determination of TGF- beta 1, NF- 魏Bp65铔嬬櫧琛ㄨ揪姘村鉤,浠ュ強(qiáng)綰跨矑浣撶敓鐗╁悎鎴愪俊鍙烽，

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