本文選題：內(nèi)質(zhì)網(wǎng)應(yīng)激 + 非諾貝特��； 參考：《安徽醫(yī)科大學(xué)》2015年碩士論文

【摘要】：目的：1.觀察高脂飲食(HFD)能否誘導(dǎo)SD大鼠胸主動(dòng)脈血管環(huán)舒縮功能變化及其病理學(xué)改變,以及非諾貝特(FF)是否能夠保護(hù)血管。2.研究棕櫚酸(PA)是否減弱正常雌性大鼠內(nèi)皮依賴(lài)性血管舒張反應(yīng)及誘導(dǎo)小鼠胸主動(dòng)脈內(nèi)皮細(xì)胞(MAEC)產(chǎn)生內(nèi)質(zhì)網(wǎng)應(yīng)激(ERS),以及非諾貝特酸(FA)的保護(hù)作用是否與緩解內(nèi)質(zhì)網(wǎng)應(yīng)激有關(guān)。方法：1.體內(nèi)實(shí)驗(yàn)部分：依據(jù)隨機(jī)分配原則,將雌性大鼠隨機(jī)分為正常飲食組(SCD)、高脂飲食組(HFD)和HFD加非諾貝特治療組(HFD+FF),以5個(gè)月高脂飲食建立SD大鼠高脂血癥模型,非諾貝特灌胃治療2個(gè)月。每周稱(chēng)量大鼠體重,測(cè)定血清生化指標(biāo)TG、TC、HDL-C、LDL-C,氣相色譜法(GC)檢測(cè)FFA、PA含量,硝酸還原酶法檢測(cè)內(nèi)皮功能紊亂指示劑NO生成變化；HE染色研究胸主動(dòng)脈病理學(xué)改變,Weigert染色檢測(cè)胸主動(dòng)脈彈力纖維變化,油紅O染色檢測(cè)脂質(zhì)沉積；RT-PCR分析胸主動(dòng)脈中CHOP及GRP78基因表達(dá)的改變；Western blot法檢測(cè)內(nèi)質(zhì)網(wǎng)應(yīng)激標(biāo)志物CHOP、GRP78的蛋白表達(dá)以及ERK、ERK磷酸化,JNK、JNK磷酸化和eNOS和eNOS的磷酸化水平變化；血管張力實(shí)驗(yàn)觀察梯度濃度乙酰膽堿(ACh)對(duì)血管環(huán)的作用變化。2.體外實(shí)驗(yàn)部分：以正常雌性SD大鼠血管環(huán)為研究對(duì)象,不同濃度棕櫚酸和非諾貝特孵育,觀察棕櫚酸和非諾貝特對(duì)乙酰膽堿(ACh)誘導(dǎo)的血管舒張反應(yīng)的影響。將體外培養(yǎng)的MAEC分為正常對(duì)照組(CON),4-PBA組(PBA),棕櫚酸組(PA),4-PBA+棕櫚酸組(PA),棕櫚酸+非諾貝特酸干預(yù)組(FA+PA),衣霉素陽(yáng)性對(duì)照組(TM), MTT法研究棕櫚酸組和非諾貝特酸組細(xì)胞活力變化；硝酸還原酶法檢測(cè)非諾貝特對(duì)棕櫚酸作用后培養(yǎng)液NO生成的變化；RT-PCR分析各組CHOP, GRP78, IRE1α和XBP1-s基因水平變化；Western blot法檢測(cè)內(nèi)質(zhì)網(wǎng)應(yīng)激標(biāo)志物CHOP、GRP78的蛋白表達(dá)以及ERK、ERK磷酸化,JNK、JNK磷酸化和eNOS和eNOS的磷酸化變化。結(jié)果：1.體內(nèi)實(shí)驗(yàn)部分：5個(gè)月HFD飲食后,與對(duì)照組比較①HFD組大鼠體重明顯高于SCD組；②HFD組大鼠血清TC、TG、LDL-C顯著增高,HDL-C顯著降低；③HFD組大鼠血清FFA、PA水平顯著高于正常組；④HFD組血清NO水平顯著高于對(duì)照組；⑤HE及Weigert染色顯示：HFD組胸主動(dòng)脈管壁增厚,彈力纖維排列紊亂,局部有斷裂現(xiàn)象；⑥油紅O染色提示HFD組血管脂質(zhì)生成顯著增加；⑦HFD組胸主動(dòng)脈組織CHOP和GRP78基因表達(dá)顯著增加；⑧CHOP、 GRP78、ERK磷酸化及JNK磷酸化表達(dá)明顯高于SCD組,eNOS磷酸化表達(dá)顯著降低；⑨HFD組胸主動(dòng)脈內(nèi)皮依賴(lài)性血管舒張反應(yīng)顯著降低；⑩非諾貝特灌胃治療2個(gè)月,與HFD組相比,顯著降低大鼠體重,逆轉(zhuǎn)血脂水平改變,明顯改善胸主動(dòng)脈病理學(xué)變化,CHOP和GRP78基因表達(dá)顯著降低,各蛋白表達(dá)CHOP、GRP78、p-ERK、 p-JNK明顯降低,p-eNOS顯著增加；舒張反應(yīng)明顯改善。2.體外實(shí)驗(yàn)部分：與CON組相比,①不同濃度PA作用正常雌性SD大鼠血管環(huán)3 h后,內(nèi)皮依賴(lài)性血管舒張變化明顯減弱,以0.2 mmol/L PA減弱最為明顯；內(nèi)質(zhì)網(wǎng)應(yīng)激抑制劑4-PBA或FA預(yù)處理后,血管環(huán)舒張反應(yīng)明顯改善；TM孵育血管環(huán)90 min后,舒張反應(yīng)顯著降低②不同濃度PA對(duì)MAEC活力明顯減弱,以0.5mmol/L PA作用24 h最明顯；③PA處理顯著減少細(xì)胞上清液NO的水平,FA孵育后逆轉(zhuǎn)PA作用下NO水平變化；④PA顯著誘導(dǎo)CHOP、GRP78、IRE1α、XBP1-s基因表達(dá);⑤CHOP、GRP78、p-ERK、p-JNK表達(dá)顯著增加,p-eNOS表達(dá)顯著降低；⑥內(nèi)質(zhì)網(wǎng)應(yīng)激抑制劑4-PBA或FA預(yù)處理后,各基因表達(dá)顯著降低,CHOP、GRP78、p-ERK、p-JNK蛋白表達(dá)明顯降低,p-eNOS顯著增加；⑦內(nèi)質(zhì)網(wǎng)應(yīng)激陽(yáng)性對(duì)照物TM組CHOP、GRP78基因和蛋白水平均明顯升高。結(jié)論：1.5個(gè)月的高脂飲食可以成功構(gòu)建雌性大鼠高脂血癥模型且減弱胸主動(dòng)脈內(nèi)皮依賴(lài)的血管舒張反應(yīng),非諾貝特可改善其變化,其機(jī)制可能與緩解內(nèi)質(zhì)網(wǎng)應(yīng)激,增加eNOS的表達(dá)和磷酸化有關(guān)。2.棕櫚酸可抑制MAEC增殖,引起內(nèi)質(zhì)網(wǎng)應(yīng)激,非諾貝特(酸)可能是通過(guò)促進(jìn)細(xì)胞增殖、抑制內(nèi)質(zhì)網(wǎng)應(yīng)激、改善內(nèi)皮依賴(lài)性血管舒張反應(yīng)來(lái)保護(hù)內(nèi)皮細(xì)胞的。
[Abstract]:Objective: 1. to observe whether the high fat diet (HFD) can induce the changes in the vasoconstrictor function of the thoracic aorta of SD rats and its pathological changes, and whether FF can protect the blood vessel.2. to study whether palmitic acid (PA) attenuated the endothelium dependent vasodilatation of normal female rats and induced the production of MAEC in the thoracic aorta of mice. Endoplasmic reticulum stress (ERS) and the protective effect of fenofibrate (FA) were associated with endoplasmic reticulum stress. Methods: 1. in vivo, the female rats were randomly divided into normal diet group (SCD), high fat diet group (HFD) and HFD plus fenofibrate (HFD+FF), and SD rats were established by high fat diet for 5 months. Hyperlipidemia model, fenofibrate gavage for 2 months. Weighing rats weekly, serum biochemical indexes TG, TC, HDL-C, LDL-C, gas chromatography (GC) were used to detect FFA, PA content, and nitric acid reductase assay for endothelial dysfunction indicator NO formation change; HE staining for pathological changes in thoracic aorta, Weigert staining for thoracic aorta Changes in elastic fiber, oil red O staining and lipid deposition; RT-PCR analysis of CHOP and GRP78 gene expression in thoracic aorta; Western blot method for detecting endoplasmic reticulum stress markers CHOP, GRP78 protein expression, ERK, ERK phosphorylation, JNK, JNK phosphorylation and phosphorylation level; vascular tension test observed gradient concentration. The effect of degree acetylcholine (ACh) on vascular ring changes in vitro.2. experimental part: the effects of palmitic acid and fenofibrate on the vasodilatation induced by acetylcholine (ACh) were observed in normal female SD rats. The effects of palmitic acid and fenofibrate on the vasodilatation induced by acetylcholine (ACh) were observed. The MAEC in vitro was divided into normal control group (CON), 4 Group -PBA (PBA), palmitic acid group (PA), 4-PBA+ palmitic acid group (PA), palmitic acid + fenofibric acid intervention group (FA+PA), mycophencin positive control group (TM), MTT method to study the change of cell vitality in palmitic acid group and fenofibrate group; nitrate reductase method was used to detect the change of NO generation after the action of palmitic acid on fenofibric acid; RT-PCR analysis of CHOP in each group. GRP78, IRE1 alpha and XBP1-s gene level changes; Western blot assay was used to detect endoplasmic reticulum stress markers CHOP, GRP78 protein expression, ERK phosphorylation, JNK, JNK phosphorylation, eNOS and phosphorylation. Results: 1. after 5 months of diet, the body weight of the rats was significantly higher than that of the control group. (2) the serum TC, TG, LDL-C and HDL-C significantly decreased in the HFD group, and the level of FFA and PA in the HFD group was significantly higher than that in the normal group; (4) the serum NO level of the group HFD was significantly higher than that of the control group. The expression of CHOP and GRP78 in thoracic aorta was significantly increased in group HFD, and the expression of CHOP, GRP78, ERK phosphorylation and JNK phosphorylation was significantly higher than that of group SCD, and the expression of phosphorylation of eNOS was significantly lower in HFD, and the vascular endothelial dependent blood Guan Shuzhang reaction in thoracic aorta decreased significantly in HFD group; After 2 months of treatment, compared with group HFD, the body weight, the change of blood lipid level, the pathological changes of the thoracic aorta, the expression of CHOP and GRP78 genes were significantly reduced, the expressions of CHOP, GRP78, p-ERK, p-JNK were significantly decreased and p-eNOS increased significantly, and the diastolic reaction obviously improved the experimental part of.2. in vitro: compared with the CON group, (1) no After 3 h of normal female SD rats with the same concentration of PA, endothelium dependent vasodilatation decreased obviously, and the most obvious decrease was 0.2 mmol/L PA. After endoplasmic reticulum stress inhibitor 4-PBA or FA preconditioning, the vasodilatation reaction of vascular ring was obviously improved; after TM incubated with 90 min, the diastolic reaction was significantly reduced by PA to MAEC alive at different concentrations. The strength was obviously weakened and the 0.5mmol/L PA was the most obvious effect of 24 h; (3) PA treatment significantly reduced the level of NO in the cell supernatant, and FA was incubated to reverse the change of NO level under the action of PA; (4) PA significantly induced CHOP, GRP78, IRE1 alpha, gene expression; After pretreatment with 4-PBA or FA, the expression of each gene was significantly reduced. The expression of CHOP, GRP78, p-ERK, p-JNK protein was significantly decreased, and p-eNOS was significantly increased; and the level of CHOP, GRP78 gene and protein in TM group of endoplasmic reticulum stress positive control was significantly increased. Conclusion: high fat diet in 1.5 months could successfully construct hyperlipidemia model and weaken in female rats. The endothelium-dependent vasodilatation response of the thoracic aorta, fenofibrate can improve its changes, its mechanism may be associated with alleviating endoplasmic reticulum stress, increasing the expression of eNOS and phosphorylation of.2. palmitic acid to inhibit MAEC proliferation, causing endoplasmic reticulum stress, fenofibrate (acid) may be by promoting cell proliferation, inhibiting endoplasmic reticulum stress and improving endothelium dependence. The vasodilatation response is used to protect endothelial cells.
【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類(lèi)號(hào)】：R589

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相關(guān)期刊論文 前1條

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本文編號(hào)：1997624