体外高表达肿瘤坏死样因子配体1A的巨噬细胞对肝星状细胞活化与增殖的影响  

The effects of macrophages with high expression of TL1A on activation and proliferation of hepatic stellate cells in vitro

作　　者：罗雨欣 郭金波[1] 尹凤荣[1] 霍晓霞[1] 郑力搏[1] 张红[1] 张晓岚[1] Luo Yuxin;Guo Jinbo;Yin Fengrong;Huo Xiaoxia;Zheng Libo;Zhang Hong;Zhang Xiaolan(Department of Gastroentology, the Second Hospital of Hebei Medical University, Shijiazhuang 050035, Chin)

机构地区：[1]河北医科大学第二医院消化内科,石家庄050035

出　　处：《中华肝脏病杂志》2018年第5期347-352,共6页Chinese Journal of Hepatology

基　　金：河北医科大学第二医院科学研究基金项目（2h2017052）

摘　　要：目的 探讨体外高表达肿瘤坏死样因子配体1A （TL1A）的巨噬细胞对肝星状细胞（HSCs）的活化、增殖的影响. 方法 分离、分化并活化C57BL/6野生型（WT）与髓系高表达TL1A的转基因（M-Tg）小鼠骨髓来源的巨噬细胞（BMMs）和腹腔巨噬细胞（PMs）;并分离WT小鼠原代HSCs;收取活化后的巨噬细胞培养上清液为条件培养基（CM）干预HSCs,应用免疫荧光法及real-time Q-PCR方法检测HSCs的活化、CCK-8法及BrdU法检测HSCs增殖情况,酶联免疫吸附试验法测定各组巨噬细胞培养上清液中白细胞介素1β和血小板衍生生长因子（PDGF） BB的含量. 结果 应用BMMs来源的CM干预HSCs,免疫荧光法分别检测第2、4、6天α-平滑肌肌动蛋白（α-SMA）表达量,第2天和第6天时各组间差异无统计学意义,P＞0.05;第4天时CM/Tg组明显高于CM/WT组,P＜0.01;PMs来源的CM干预HSCs的检测结果与上述趋势一致.应用BMMs来源的CM干预HSCs,real-time Q-PCR法分别检测第2、4、6天α-SMA mRNA表达量,第2天时各组差异无统计学意义,P＞0.05;第4、6天时α-SMA mRNA进一步升高,且CM/Tg组明显高于CM/WT组,P＜0.05;PMs来源的CM培养HSCs的检测结果与上述趋势一致.CCK-8法和BrdU法检测HSCs增殖速度结果显示CM/Tg组明显高于CM/WT组,P＜0.01;应用PMs来源的CM干预HSCs,检测结果与上述趋势一致.对于BMMs、脂多糖＋干扰素γ/Tg组培养上清液中白细胞介素1 β和PDGF-BB表达水平显著高于脂多糖＋干扰素γ/WT组,P＜0.01;PMs的培养上清液检测结果与上述趋势一致. 结论 高表达TL1A的巨噬细胞可能通过IL-1β和PDGF-BB分泌增加促进HSCs的活化与增殖.Objective To explore the effects of macrophages with high expression of TL1A on the activation and proliferation of HSCs in vitro.Methods The Bone marrow-derived macrophages （BMMs） and peritoneal macrophages （PMs） from wild type （WT） and myeloid-overexpressed TL1A transgenic mice were isolated,differentiated and activated.HSCs were harvested from activated macrophages culture supernatant （CM）.HSCs were detected by immunofluorescence and real-time Q-PCR.And the proliferation was detected by CCK-8 and BrdU assay kit.The levels of IL-1β and PDGF-BB in macrophage culture supernatants were determined by enzyme-linked immunosorbent assay （ELISA）.Results BMMs-derived CM-intervention HSCs were used to detect the expression of α-smooth muscle actin （α-SMA） on the 2nd,4th and 6th day respectively by immunofluorescence method.There was no significant difference between the two groups on the 2 nd and the 6th day,P 〉 0.05;On day 4,the CM/Tg group was significantly higher than that of CM/WT group,P 〈 0.01;the results of CMs derived from PMs were consistent with the above trend.The expression of α-SMA mRNA on the 2nd,4th and 6th day was detected by real-time Q-PCR method using BM-derived CMs.No significant difference was found between the groups on the 2nd day （P 〉 0.05）.α-SMA mRNA increased further on the 4th and 6th day,and the level of CM/Tg in CM/Tg group was significantly higher than that in CM/WT group （P 〈 0.05）.The detection results of CMs derived from PMs were consistent with the above trend.The results of CCK-8 assay and BrdU assay showed that the proliferation rate of HSCs in CM Tg group was significantly higher than that in CM/WT group （P 〈 0.01）.The CMs derived from PMs were used to interfere with HSCs.And the results were consistent with the above trend.For BMMs,the levels of IL-1 β and PDGF-BB in the lipopolysaccharide （LPS） ＋ IFNγ/Tg culture supernatant were significantly higher than those in the LPS＋IFNγ/WT group （P 〈 0.01）.For the culture supe

关 键 词：巨噬细胞 肝星状细胞 肿瘤坏死因子样配体1A 白细胞介素1Β 血小板衍生生长因子-BB 

分 类 号：R575.2[医药卫生—消化系统]