【摘要】：目的：觀察Daxx對(duì)Chol:MβCD誘導(dǎo)RAW264.7細(xì)胞荷脂與凋亡的影響，為延緩動(dòng)脈粥樣硬化的病理改變提供理論依據(jù)。 方法：人源性Daxx基因重組質(zhì)粒穩(wěn)定轉(zhuǎn)染RAW264.7細(xì)胞，應(yīng)用RT-PCR和Western blot檢測(cè)穩(wěn)定轉(zhuǎn)染細(xì)胞內(nèi)Daxx的表達(dá)。然后用Chol:MβCD與RAW264.7細(xì)胞共同孵育48h，建立RAW264.7細(xì)胞荷脂模型，油紅O染色和酶熒光法檢測(cè)細(xì)胞內(nèi)荷脂蓄積情況。MTT檢測(cè)細(xì)胞活性，流式細(xì)胞術(shù)觀察細(xì)胞凋亡。Westernblot測(cè)定ASK1，JNK，Caspase3的表達(dá)情況。 結(jié)果：轉(zhuǎn)染細(xì)胞經(jīng)G418篩選14天后，RT-PCR和Western blot檢測(cè)轉(zhuǎn)染細(xì)胞內(nèi)Daxx明顯高表達(dá)，表明穩(wěn)定轉(zhuǎn)染成功。Chol:MβCD與RAW264.7細(xì)胞共同孵育48h后，結(jié)果發(fā)現(xiàn)Daxx高表達(dá)組細(xì)胞膽固醇蓄積明顯，并且細(xì)胞活性下調(diào)，凋亡細(xì)胞數(shù)目增加；同時(shí)Daxx轉(zhuǎn)染細(xì)胞組ASK1，JNK，Caspase3蛋白表達(dá)明顯增高。 結(jié)論：Daxx可介導(dǎo)Chol:MβCD誘導(dǎo)的巨噬細(xì)胞凋亡，其凋亡通路可能與Daxx上調(diào)ASK1表達(dá)，繼之增加下游JNK活性，，進(jìn)一步激活Caspase3有關(guān)。
[Abstract]:Aim: to observe the effects of Daxx on lipid loading and apoptosis of RAW264.7 cells induced by Chol:M 尾 CD, and to provide a theoretical basis for delaying the pathological changes of atherosclerosis. Methods: the recombinant plasmid of human Daxx gene was transfected into RAW264.7 cells stably. RT-PCR and Western blot were used to detect the expression of Daxx in the transfected cells. Then Chol:M 尾 CD was incubated with RAW264.7 cells for 48h to establish the model of RAW264.7 cell lipids. Oil red O staining and enzyme fluorescence assay were used to detect the accumulation of lipid in the cells. MTT was used to detect cell activity, flow cytometry was used to observe apoptosis. Westernblot was used to detect ASK1,. Expression of JNK,Caspase3. Results: after being screened by G418 for 14 days, the expression of Daxx in transfected cells was detected by RT-PCR and Western blot, indicating that the transfected cells were stably transfected successfully. Chol:M 尾 CD was incubated with RAW264.7 cells for 48 h. The results showed that the accumulation of cholesterol was obvious, the cell activity was down-regulated and the number of apoptotic cells was increased in the high expression group of Daxx. At the same time, the expression of ASK1,JNK,Caspase3 protein was significantly increased in Daxx transfected cells. Conclusion: Daxx can mediate the apoptosis of macrophages induced by Chol:M 尾 CD, and its apoptotic pathway may be related to the up-regulation of ASK1 expression by Daxx, the increase of downstream JNK activity and the further activation of Caspase3.
【學(xué)位授予單位】：南華大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R363

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 李明;丁健;繆澤鴻;;未折疊蛋白反應(yīng)的信號(hào)轉(zhuǎn)導(dǎo)[J];生命科學(xué);2008年02期

本文編號(hào)：2369789