本文選題：rHDL + CACs衰老　； 參考：《北京協(xié)和醫(yī)學(xué)院》2011年碩士論文

【摘要】：目的 循環(huán)成血管細胞(circulating angiogenic cells, CACs)參與新生血管的形成和動脈修復(fù)。高密度脂蛋白(HDL)可提高CACs的活性,但其中的調(diào)節(jié)機制迄今仍不清楚。本研究旨在闡述rHDL是通過什么分子機制來影響CACs衰老的。方法 1、從健康志愿者外周血中分離單個核細胞,并誘導(dǎo)分化培養(yǎng)為CACs。 2、CACs與rHDL共培養(yǎng)后(加或不加雷帕霉素),用β-半乳糖苷酶染色法檢測CACs衰老程度。 3、Western Blot口免疫沉淀法檢測：nTOR磷酸化、(?)nTOR-rictor復(fù)合物形成和mTOR-rictor依賴的Akt活化。 4、免疫細胞組織化學(xué)法、RT-PCR及Western Blot檢測人端粒酶逆轉(zhuǎn)錄酶(hTERT)的核轉(zhuǎn)位,試劑盒檢測核端粒酶活性。結(jié)果 1、rHDL可減弱CACs的衰老,然而此效應(yīng)可被哺乳動物雷帕霉素靶蛋白(mTOR)抑制劑雷帕霉素所阻斷。 2、rHDL能促使mTOR發(fā)生磷酸化、mTOR-rictor復(fù)合物形成和mTOR-rictor依賴的Akt活化；促使人端粒酶逆轉(zhuǎn)錄酶(hTERT)的核轉(zhuǎn)位增加及核端粒酶活性增強。 3、用SiRNA抑制rictor基因表達后,(?)nTOR-rictor復(fù)合物形成和Akt活化均被阻斷,同時rHDL對CACs衰老的抑制作用和核端粒酶活性的提高作用均被抑制。結(jié)論 rHDL能促使(?)nTOR的持久磷酸化和(?)nTOR-rictor復(fù)合物形成,并能通過mTORC2信號通路的激活而抑韋CACs的衰老。
[Abstract]:Purpose Circulating angiogenic cells, CACs) is involved in angiogenesis and arterial repair. High density lipoprotein (HDL) can improve the activity of CACs, but the regulatory mechanism is still unclear. The aim of this study was to elucidate the molecular mechanism by which rHDL affects CACs senescence. Method 1. Mononuclear cells were isolated from the peripheral blood of healthy volunteers and differentiated into CACs. (2) after co-culture with rHDL (with or without rapamycin), 尾 -galactosidase staining was used to detect the senescence of CACs. 3Western Blot immunoprecipitation assay was used to detect the formation of nTOR-rictor complex and the activation of mTOR-rictor dependent Akt. 4. The nuclear translocation of human telomerase reverse transcriptase (hTERT) was detected by RT-PCR and Western Blot, and the nuclear telomerase activity was detected by kit. Result 1rHDL can attenuate the senescence of CACs, but this effect can be blocked by rapamycin, an inhibitor of rapamycin target protein mTORin in mammals. 2rHDL could induce the formation of phosphorylated mTOR-rictor complex and the activation of mTOR-rictor dependent Akt, and increase the nuclear translocation and nuclear telomerase activity of human telomerase reverse transcriptase (hTERT). 3. The inhibition of rictor gene expression by SiRNA and the inhibition of rHDL on CACs senescence and the increase of nuclear telomerase activity were all blocked. Conclusion RHDL can promote the sustained phosphorylation of nTOR and the formation of nTOR-rictor complex, and inhibit the senescence of CACs through the activation of mTORC2 signaling pathway.
【學(xué)位授予單位】：北京協(xié)和醫(yī)學(xué)院
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R363

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