摘要
目的：研究白细胞介素-32（Interleukin-32， IL-32）可否诱导肝星状细胞LX-2表达基质金属蛋白酶9（matrix metalloproteinase 9， MMP-9）。方法用不同浓度的IL-32γ与LX-2细胞共培养，收集细胞培养上清，用TRizol试剂提取细胞总RNA，用定量 PCR检测MMP-9 RNA表达水平， MMP-9蛋白质表达水平用酶联免疫吸附试验（ enzyme-linked immunosorbent assay， ELISA）试剂盒检测。接下来将Nuclear factor kappa B （ NF-κB）亚基p50、 p65真核表达载体pCMV-p50、 pCMV-p65单独或联合转染LX-2细胞，48 h后收集细胞培养上清，酶联免疫吸附测定（ enzyme linked immunosorbent assay， ELISA）检测MMP-9蛋白质表达水平。将不同浓度的NF-κB抑制剂SN50加入IL-32γ与LX-2细胞共培养的培养液中，48 h后收集细胞培养上清，用ELISA检测MMP-9蛋白质表达水平。结果 IL-32γ可以诱导人LX-2细胞表达MMP-9，且其表达水平随IL-32γ浓度增加而增强； NF-κB亚基p50、 p65可以诱导LX-2细胞MMP-9表达增高； NF-κB抑制剂SN50阻断NF-κB通路可降低IL-32γ诱导的MMP-9表达。结论 IL-32γ通过活化NF-κB通路诱导LX-2细胞表达MMP-9， IL-32可能通过诱导肝星状细胞表达MMP-9参与肝纤维化形成。
Objective To investigate interlukin-32γ （ IL-32γ） -induced expression of matrix metalloproteinase （ MMP-9 ） in LX-2 cells. Methods LX-2 cells were treated with different concen-trations of IL-32γ. The supernatants were harvested, total RNA extracted by using TRizol agent and subjected to quantitative PCR for detection of the expression of MMP-9 mRNA. The expression level of MMP-9 protein was detected by enzyme-linked immunosorbent assay （ ELISA） . Subsequently, LX-2 cells were transfected with pCMV-p50 and pCMV-p65 alone or combined. Forty-eight h later, cell supernatants were collected and the MMP-9 protein expression was measured by ELISA. Then different concentrations of NF-κB inhibitor SN50 were added to the media of LX-2 cells in the pres- ence of IL-32γ. After 48 h, ELISA was used to detect the MMP-9 expression at protein lev-el. Results IL-32γ could induce the expression of MMP-9 in a dose-dependent manner in LX-2 cells. NF-κB subunits, p50 and/or p65 also induced the MMP-9 expression in LX-2 cells. The MMP-9 expression induced by IL-32γ was diminished after treatment with the NF-κB inhibitor SN50. Conclusion IL-32γ could induce the expression of MMP-9 in LX-2 cells through activation of NF-κB signaling pathway. It may be implicated in the pathogenesis of liver fibrosis through induc-tion of the MMP-9 expression in LX-2 cells.
出处
《医学分子生物学杂志》
CAS
2016年第3期149-152,共4页
Journal of Medical Molecular Biology
基金
国家自然科学基金（No.81401306,81373152）,广州医科大学附属第三医院博士启动基金（No.2014Y02）
