【摘要】：目的:探討白細(xì)胞分化抗原36(cluster of differentiation 36,CD36)是否參與了棕櫚酸誘導(dǎo)的HepG2細(xì)胞炎癥反應(yīng)。方法:給予HepG2細(xì)胞不同濃度的棕櫚酸(0.00、0.08、0.16、0.32、0.64 mmol/L)處理24 h,使用實(shí)時(shí)熒光定量PCR方法檢測(cè)CD36和炎癥因子的mRNA表達(dá)水平,Western blot檢測(cè)CD36蛋白表達(dá)水平。然后利用小RNA干擾技術(shù),構(gòu)建低表達(dá)CD36的HepG2細(xì)胞(CD36RNAi HepG2)和對(duì)照細(xì)胞(NCi HepG2)模型。通過實(shí)時(shí)熒光定量PCR檢測(cè)棕櫚酸負(fù)荷的NCi HepG2和CD36RNAi HepG2中炎癥因子的表達(dá)情況,熒光探針DCFH DA法檢測(cè)細(xì)胞內(nèi)活性氧含量,Western blot檢測(cè)細(xì)胞核內(nèi)的核轉(zhuǎn)錄因子-κB的蛋白表達(dá)水平。結(jié)果:棕櫚酸能夠上調(diào)HepG2細(xì)胞中CD36及炎癥因子——腫瘤壞死因子-α(tumor necrosis factor-α,TNF-α)和白介素-6(interleukin-6,IL-6)的表達(dá)。棕櫚酸濃度分別為0.00、0.08、0.16、0.32及0.64 mmol/L時(shí),CD36的mRNA表達(dá)相對(duì)值分別為(1.001±0.078)、(1.510±0.341)(q=4.763,P=0.007)、(1.780±0.070)(q=7.301,P=0.000)、(2.510±0.141)(q=14.16,P=0.000)及(2.103±0.150)(q=10.34,P=0.000)。TNF-α的mRNA表達(dá)相對(duì)值分別為(1.000±0.098)、(1.571±0.177)(q=1.598,P=0.141)、(1.725±0.274)(q=2.016,P=0.071)、(2.113±0.341)(q=3.102,P=0.011)及(5.282±0.855)(q=11.940,P=0.000)。IL-6的mRNA表達(dá)相對(duì)值分別為(0.999±0.047)、(1.791±0.596)(q=1.486,P=0.168)、(2.119±0.294)(q=2.107,P=0.061)、(2.808±0.147)(q=3.398,P=0.007)及(6.916±1.284)(q=11.130,P=0.000)。抑制HepG2細(xì)胞中CD36表達(dá),可以顯著降低棕櫚酸所誘導(dǎo)的細(xì)胞內(nèi)活性氧(reactive oxygen species,ROS)增加和核轉(zhuǎn)錄因子-κB在核內(nèi)分布的增強(qiáng),降低HepG2細(xì)胞內(nèi)炎癥因子表達(dá)水平。NCi HepG2、NCi HepG2+棕櫚酸組及CD36 RNAi HepG2+棕櫚酸組的ROS相對(duì)含量分別為(1.000±0.113)、(1.968±0.293)(與NCi組相比,t=6.888,P=0.000)、(0.519±0.129)(與NCi+棕櫚酸組相比,t=10.106,P=0.000)。NCi HepG2、NCi HepG2+棕櫚酸組及CD36 RNAi HepG2+棕櫚酸組的核轉(zhuǎn)錄因子-κB的蛋白相對(duì)值分別為(0.997±0.093)、(2.443±0.085)(與NC相比,t=19.892,P=0.000)、(1.607±0.107)(與NCi+棕櫚酸組相比,t=10.607,P=0.000)。抑制HepG2細(xì)胞中CD36表達(dá)后,棕櫚酸濃度分別為0.08、0.16、0.32及0.64 mmol/L時(shí),NCi HepG2及CD36 RNAi HepG2組的TNF-α的表達(dá)水平分別為(1.004±0.113)和(0.185±0.071)(t=10.716,P=0.000);(1.009±0.160)和(0.221±0.028)(t=8.389,P=0.001);(1.008±0.163)和(0.173±0.011)(t=8.780,P=0.001);(1.009±0.163)和(0.147±0.013)(t=9.013,P=0.000)。抑制HepG2細(xì)胞中CD36表達(dá)后,棕櫚酸濃度分別為0.08、0.16、0.32及0.64 mmol/L時(shí),NCi HepG2及CD36 RNAi HepG2組的IL-6的表達(dá)水平分別為(1.044±0.347)和(0.207±0.033)(t=4.121,P=0.015);(1.006±0.139)和(0.198±0.007)(t=10.110,P=0.001);(1.001±0.052)和(0.125±0.006)(t=28.443,P=0.000);(1.012±0.188)和(0.114±0.015)(t=8.367,P=0.001)。結(jié)論:棕櫚酸能夠上調(diào)HepG2細(xì)胞CD36表達(dá),促進(jìn)HepG2炎癥反應(yīng);抑制CD36可能通過減少氧化應(yīng)激、抑制核轉(zhuǎn)錄因子-κB信號(hào)通路,進(jìn)而改善棕櫚酸誘導(dǎo)的HepG2細(xì)胞炎癥。
[Abstract]:Aim: to investigate the role of leukocyte differentiation antigen 36 (cluster of differentiation 36 (CD36) in palmitic acid-induced inflammation of HepG2 cells. Methods: HepG2 cells were treated with different concentrations of palmitic acid (0.000. 08 ~ 0.16 ~ 0. 32 ~ 0. 64 mmol/L) for 24 h. The expression level of CD36 and mRNA of inflammatory factors was detected by real-time fluorescence quantitative PCR. CD36 protein expression was detected by, Western blot. Then HepG2 cells (CD36RNAi HepG2) and control cells (NCi HepG2) with low expression of CD36 were constructed by using small RNA interference technique. The expression of inflammatory factors in palmitic acid-loaded NCi HepG2 and CD36RNAi HepG2 was detected by real-time fluorescence quantitative PCR, and the expression of nuclear transcription factor- 魏 B was detected by fluorescence probe DCFH DA method. The active oxygen species (Ros) content in the cells was detected by, Western blot. Results: palmitic acid could up-regulate the expression of CD36, tumor necrosis factor- 偽 (TNF- 偽) and interleukin-6 (interleukin-6,IL-6) in HepG2 cells. 媯曟閰告祿搴﹀垎鍒負(fù)0.00,0.08,0.16,0.32鍙，

本文編號(hào)：2265650