关键词: 抑制剂, 核因子-κB, 白细胞介素, 细胞培养, 反转录聚合酶链反应, 人

Abstract: Objective To investigate the inhibition effects of nuclear factor-κB(NF-κB) on cytokine expression of cultured human corneal fibroblasts in vitro. Methods The cultured human corneal fibroblasts were used in the present experiment. Cell viability insensitive to emodin on cultured human corneal fibroblasts in vitro was assessed using the MTT assay.Cells were randomly divided into three groups：control group（group 0 hour，EM>n/EM>=6）,lipopolysaccharidel (LPS)group （EM>n/EM>=24）and emodin pretreatment group（EM>n/EM>=24）. Cells of the emodin pretreatment group were incubated with emodin for 30 minutes before LPS challenged. The cultured human corneal fibroblasts were then challenged with LPS, the expression of interleukin-8(IL-8) mRNAs was detected by reverse transcription polymerase chain reaction analysis (RT-PCR) and the proteins of IL-8 secreted from these cells were assessed by enzymelinked immunosorbent assays (ELISA). Western blotting analysis was used for detecting the protein expression of inhibitor of nuclear factor-κB-α（IκB-α）induced by LPS.At the same time，the effects of emodin on these expressions were also assessed in fibroblasts. Results The concentration of emodin used in this study was safe to these cultured human corneal fibroblasts in vitro.Both the protein release and mRNA expression of IL-8 in corneal fibroblasts increased significantly after challenged with LPS, however, the protein level of IκB-α significantly decreased after treated with LPS. These results indicated that LPS could mediate the activation of NF-κB and upregulate the expression of cytokines in corneal fibroblasts. Pretreated with emodin 30 min before challenged with LPS, the activation of NF-κB induced by LPS was markedly inhibited, and the

Key words: Inhibitor, Nuclear factor-κB, Interleukin, Cell culture, RT-PCR, Human