TNF-α对未分化及成熟3T3-L1脂肪细胞中TSG-6基因表达的调节作用

Regulative role of tumor necrosis factor-α on TSG-6 gene expression in both undifferenti-ated and matured 3T3-L1 adipocytes

目的:探讨TNF鄄α对未分化及成熟3T3鄄L1脂肪细胞中TSG鄄6基因(tumornecrosisfactoralpha鄄stimulatedgene鄄6)表达水平的调节作用。方法:体外培养3T3鄄L1脂肪细胞,在3T3鄄L1细胞生长至完全融合后2天(第0天)或细胞诱导分化的第10天,应用重组TNF鄄α(0.1、1.0、10.0ng/ml)干预未分化(第0天)或已分化成熟(第10天)的脂肪细胞,采用RT鄄PCR技术检测干预后不同时间(0、0.5、2.0、6.0、12.0、24.0h)脂肪细胞中TSG鄄6基因的表达水平。结果:①不同浓度TNF鄄α对未分化脂肪细胞中TSG鄄6基因的表达水平均具抑制作用。TNF鄄α浓度0.1ng/ml时,TSG鄄6基因表达水平2h内下降67.30%,24h时下降76.05%;TNF鄄α浓度1.0ng/ml时,TSG鄄6基因表达水平0.5h内下降52.79%,24h时下降99.20%;TNF鄄α浓度10.0ng/ml时,TSG鄄6基因表达水平0.5h内下降96.14%,其表达几乎被完全抑制;②不同浓度TNF鄄α均能显著抑制成熟脂肪细胞中TSG鄄6基因的表达。TNF鄄α浓度0.1ng/ml时,TSG鄄6基因表达水平6h内下降33.73%,12h内下降97.39%;TNF鄄α浓度1.0ng/ml时,TSG鄄6基因表达水平6h内下降78.68%,并持续至24h;TNF鄄α浓度10.0ng/ml时,TSG鄄6基因表达水平2h内下降96.27%,TSG鄄6基因的表达几乎被完全抑制。

Objective: To explore the regulative role of tumor necrosis factor-α(TNF-α) on tumor necrosis factor alpha-stimulated gene-6(TSG-6) gene expression in both undifferentiated and matured 3T3-L1 adipocytes. Methods: 3T3-L1 preadipocytes were culturedand differentiated into the matured adipocytes. Recombinant TNF-α at different concentrations(0.1,1.0,10.0 ng/ml) was added to the culture medium 2 days post-confluent undifferentiated adipocytes(Day 0) or fully differentiated adipocytes (Day 10) for various times(0, 0.5, 2.0, 6.0, 12.0, 24.0 h). Total RNA from these adipocytes was extracted and the expression of TSG-6 mRNA was evaluated by RT-PCR. Results:①Treatment of undifferentiated 3T3-L1 adipocytes with TNF-α at different concentrations (0.1, 1.0, 10.0 ng/ml) resulted in the significant decrease in the expression of TSG-6 mRNA. When 0.1 ng/ml TNF-α was applied,the levels of TSG-6 gene expression significantly decreased by 67.30% at 2 h, and 76.05% at 24 h. When 1.0 ng/ml TNF-α was used, the levels of TSG-6 gene expression significantly decreased by 52.79% at 0.5 h, and 99.20% at 24 h. When the cells were treated with 10.0 ng/ml TNF-α, TSG-6 gene expression was almost fully inhibited(96.14% decreased) at 0.5 h. ②Treatment of matured 3T3-L1 adipocytes with TNF-α at different concentrations(0.1,1.0,10.0 ng/ml) also resulted in the significant decrease in the expression of TSG-6 mRNA. When the cells were treated with 0.1 ng/ml TNF-α, the levels of TSG-6 gene expression decreased by 33.73% at 6 h, and 97.39% at 12 h. When treated with 1.0 ng/ml TNF-α, the levels of TSG-6 gene expression decreased by 78.68% at 6 h, which sustained until 24 h. When treated with 10.0 ng/ml TNF-α, the levels of TSG-6 gene expression decreased by 96.27% at 2 h and TSG-6 gene expression was almost fully inhibited. Conclusion: TNF-α can downregulate the mRNA expression of TSG-6 in both undifferentiated and matured adipocytes. The inhibitive effect of TNF-α on TSG-6 gene expression is in dose and time-dependent manner.