【摘要】：目的：探討內(nèi)皮細(xì)胞活化對(duì)血管平滑肌細(xì)胞增殖及分泌的的影響及其中機(jī)制。 方法：(1)以不同濃度和不同干預(yù)時(shí)間的TNF-α刺激人臍靜脈內(nèi)皮細(xì)胞(HUVEC),收集細(xì)胞上清液,用ELISA法測(cè)定內(nèi)皮細(xì)胞IL-6的分泌,TUNEL法檢測(cè)內(nèi)皮細(xì)胞的凋亡；(2) HUVEC培養(yǎng)液中分別加入10ng/mlTNF-α或不加入TNF-α24h后,收集上清離心,用0.22um的微孔濾膜過(guò)濾,4℃保存?zhèn)溆?分別為A液和B液；(3)將臍靜脈平滑肌細(xì)胞接種于培養(yǎng)板中分為4組：①刺激組：按1：1比例加入A液和DMEM培養(yǎng)液,②拮抗組：在刺激組基礎(chǔ)上加入抗IL-6單抗,③對(duì)照組：按1：1比例加入B液和DMEM培養(yǎng)液,④空白組：僅加入DMEM培養(yǎng)液。分組后立即采集少量細(xì)胞上清液以ELISA法測(cè)定ICAM-1、 IL-8水平。繼續(xù)培養(yǎng)24h后,再次采集上清液測(cè)定ICAM-1和IL-8水平,并收集血管平滑肌細(xì)胞以MTT法檢測(cè)平滑肌細(xì)胞增殖的情況。 結(jié)果：1、TNF-α對(duì)血管內(nèi)皮細(xì)胞分泌和凋亡的影響：(1) TNF-α濃度和作用時(shí)間對(duì)血管內(nèi)皮細(xì)胞分泌IL-6的影響：課題對(duì)照組：(31.2±2.9) pg/ml、5ng/ml組：(95.4±4.9) pg/ml、10ng/ml組：(122.7±6.3) pg/ml、20ng/ml組：(132.5±7.1) pg/ml、40ng/ml組：(79.1±5.9) pg/ml;時(shí)間效應(yīng)：對(duì)照組：(12.4±1.6) pg/ml、6h組：(79.3±5.5) pg/ml、12h組：(97.5±3.9) pg/ml、24h組：(122.7±6.3) pg/ml、48h組：(108.0±4.4) pg/ml;(2) TNF-α對(duì)血管內(nèi)皮細(xì)胞凋亡的影響各時(shí)間點(diǎn)細(xì)胞凋亡指數(shù)測(cè)定：對(duì)照組：(0.029±0.004)、12h組：(0.140±0.005)、24h組：(0.255±0.006)、48h組：(0.329±0.013)。2、內(nèi)皮細(xì)胞條件培養(yǎng)液對(duì)平滑肌細(xì)胞增殖和分泌的影響：(1)內(nèi)皮細(xì)胞條件培養(yǎng)液對(duì)平滑肌細(xì)胞增殖的影響：四組細(xì)胞的OD值依次為(1.349±0.115)、(1.176±0.093)、(1.012±0.087)和(0.798±0.014),組間比較有顯著性差異(F=22.315,P0.05)；(2)內(nèi)皮細(xì)胞條件培養(yǎng)液對(duì)平滑肌細(xì)胞分泌的影響：四組細(xì)胞上清液中ICAM-1和IL-8均較基線值有所增加,其中刺激組ICAM-1和IL-8分別較基線升高(0.222±0.044) ng/ml和(2.996±0.059) ng/ml,均高于拮抗劑組、對(duì)照組和空白組(P0.05)。 結(jié)論：TNF-α誘導(dǎo)內(nèi)皮細(xì)胞活化,通過(guò)刺激IL-6釋放,促進(jìn)血管平滑肌細(xì)胞增殖和ICAM-1和IL-8等細(xì)胞因子的分泌。
[Abstract]:Aim: to investigate the effect of endothelial cell activation on proliferation and secretion of vascular smooth muscle cells and its mechanism. Methods: (1) the supernatant of human umbilical vein endothelial cells (HUVEC),) was collected by TNF- 偽 stimulation with different concentration and different intervention time. ELISA assay was used to detect the secretion of IL-6 in endothelial cells and Tunel method was used to detect the apoptosis of endothelial cells. (2) after 10 ng / ml TNF- 偽 or no TNF- 偽 was added to the HUVEC medium for 24 hours, the supernatant was collected and centrifuged, and then stored at 4 鈩，

本文編號(hào)：2238593