本文選題：E-選擇素 + 細(xì)胞間粘附分子-1�。� 參考：《福建醫(yī)科大學(xué)》2008年碩士論文

【摘要】： 目的:探討E-選擇素(E-selectin)和細(xì)胞間粘附分子-1(ICAM-1)反義寡核苷酸(asODN)及其與辛伐他汀(simvastatin)聯(lián)合應(yīng)用對(duì)人臍靜脈內(nèi)皮細(xì)胞(HUVEC) ICAM-1和E-selectin表達(dá)的抑制和對(duì)外周血單個(gè)核細(xì)胞粘附的影響。 方法:原代培養(yǎng)人臍靜脈內(nèi)皮細(xì)胞(HUVEC),并傳代;應(yīng)用脂質(zhì)體轉(zhuǎn)染法將人工合成E-selectin asODN和ICAM-1 asODN分別導(dǎo)入HUVECs;實(shí)驗(yàn)共分8組,分別為空白對(duì)照(basal)組、TNF-α誘導(dǎo)組、liposome對(duì)照組、liposome-control asODN組、裸asODN (nude asODN)組、脂質(zhì)體asODN (lipo-asODN)組、simvastatin處理組、lipo-asODN與simvastatin聯(lián)合組,除空白組外,其余7組均以TNF-α誘導(dǎo)內(nèi)皮損傷;以流式細(xì)胞技術(shù)測(cè)定HUVECs細(xì)胞膜E-selectin和ICAM-1蛋白的表達(dá);以RT-PCR半定量技術(shù)測(cè)定HUVECs E-selectin mRNA和ICAM-1 mRNA的表達(dá);以細(xì)胞粘附試驗(yàn)測(cè)定HUVECs與單個(gè)核細(xì)胞的粘附率;應(yīng)用辛伐他汀預(yù)先處理HUVECs,再用E-selectin lipo-asODN和ICAM-1 lipo-asODN轉(zhuǎn)染HUVECs,觀察E-selectin asODN和ICAM-1 asODN與辛伐他汀聯(lián)合應(yīng)用對(duì)HUVECs E-selectin和ICAM-1的表達(dá)以及對(duì)HUVECs與外周血單個(gè)核細(xì)胞粘附抑制的影響。 結(jié)果:⑴與TNF-α誘導(dǎo)組相比,E-selectin asODN和ICAM-1 asODN以及E-selectin lipo-asODN和ICAM-1 lipo-asODN均明顯降低了HUVECs相應(yīng)的E-selectin和ICAM-1蛋白及mRNA的表達(dá)(P0.01);⑵E-selectin asODN和ICAM-1 lipo-asODN與simvastatin聯(lián)合應(yīng)用可降低對(duì)HUVECs相應(yīng)的E-selectin和ICAM-1的蛋白及mRNA表達(dá);⑶lipo-asODN與simvastatin聯(lián)合組與1ipo-asODN組比較,E-selectin表達(dá)水平顯著下降(P0.01),而ICAM-1的表達(dá)水平下降無(wú)顯著意義(P0.05),表明E-selectin lipo-asODN與simvastatin聯(lián)合作用有更好的抑制效果。⑷與TNF-α刺激組相比,lipo-asODN組及1ipo-asODN與simvastatin聯(lián)合組HUVECs與單個(gè)核細(xì)胞粘附率均顯著下降(P0.01),尤其以HUVECs與CD3+T淋巴細(xì)胞的粘附率下降最為明顯,但1ipo-asODN與simvastatin聯(lián)合組與1ipo-asODN組比較,未見(jiàn)明顯變化(P0.05)。 結(jié)論:E-selectin asODN和ICAM-1 asODN可抑制HUVECs相應(yīng)的蛋白及mRNA的表達(dá),并可抑制HUVECs與單個(gè)核細(xì)胞的粘附,尤其對(duì)HUVECs與CD3+ T細(xì)胞粘附的抑制作用更為明顯。1ipo-asODN與simvastatin聯(lián)合應(yīng)用比單獨(dú)1ipo-asODN的抑制作用更加明顯。本研究為E-selectin asODN和ICAM-1 asODN應(yīng)用于臨床抗動(dòng)脈粥樣硬化的基因治療的前景開(kāi)拓新的思路和提供理論依據(jù)。
[Abstract]:Aim: to investigate the inhibition of ICAM-1 and E-selectin expression in human umbilical vein endothelial cells (HUVECs) by Eselectin E-selectin (E-selectin) and intercellular adhesion molecule-1 (ICAM-1) antisense oligodeoxynucleotides (ASODN) and its combination with simvastatin in human umbilical vein endothelial cells (HUVECs) and the effects on the adhesion of peripheral blood mononuclear cells (PBMC). Methods: human umbilical vein endothelial cells (HUVECs) were cultured and subcultured, the synthetic E-selectin asODN and ICAM-1 asODN were transfected into HUVECs by liposome transfection, and were divided into 8 groups: TNF- 偽 induced liposome-control asODN group and naked asODN nude asODN group. The endothelial damage was induced by TNF- 偽 in the liposome asODN liposome lipo-asODN group and simvastatin treated group, and the expression of E-selectin and ICAM-1 protein in the HUVECs cell membrane was measured by flow cytometry. The expression of HUVECs E-selectin mRNA and ICAM-1 mRNA was detected by RT-PCR semi-quantitative technique, and the adhesion rate of HUVECs to mononuclear cells was measured by cell adhesion test. HUVECs were pretreated with simvastatin and transfected with E-selectin lipo-asODN and ICAM-1 lipo-asODN. The effects of E-selectin asODN and ICAM-1 asODN combined with simvastatin on the expression of HUVECs E-selectin and ICAM-1 and the inhibition of HUVECs adhesion to peripheral blood mononuclear cells were observed. Results compared with TNF- 偽 induced group, the expression of E-selectin asODN and ICAM-1 asODN, E-selectin lipo-asODN and ICAM-1 lipo-asODN, E-selectin and ICAM-1 protein of HUVECs and the expression of mRNA, P0.01E-selectin asODN and ICAM-1 lipo-asODN combined with simvastatin decreased the corresponding E-selectin and ICAM-1 protein and mRNA expression of HUVECs. The expression of E-selectin in 3lipo-asODN combined with simvastatin group was significantly lower than that in 1ipo-asODN group, while the expression level of ICAM-1 was not significantly decreased in 1ipo-asODN group. It indicated that the combined effect of E-selectin lipo-asODN and simvastatin was better than that of TNF- 偽 stimulation group. 4. The inhibitory effect was better than that of TNF- 偽 -stimulated group and 1ipo-asODN and simvastatin group. In the combined group, the adhesion rate of HUVECs to mononuclear cells decreased significantly, especially the adhesion rate of HUVECs to CD3 T lymphocytes. However, there was no significant difference between 1ipo-asODN and simvastatin group and 1ipo-asODN group. Conclusion asODN and ICAM-1 asODN can inhibit the expression of HUVECs protein and mRNA, and inhibit the adhesion of HUVECs to mononuclear cells. In particular, the inhibitory effect on HUVECs adhesion to CD3 T cells was more obvious. 1Ipo-asODN combined with simvastatin had more obvious inhibitory effect than 1ipo-asODN alone. This study provides new ideas and theoretical basis for the application of E-selectin asODN and ICAM-1 asODN in gene therapy against atherosclerosis.
【學(xué)位授予單位】：福建醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2008
【分類號(hào)】：R392;R543

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