目的探索三氧化二砷（As2O3）对肺癌A549细胞和血管内皮细胞Notch通路相关分子表达的影响。方法体外培养人肺癌A549细胞和人脐静脉内皮细胞（HUVECs）,分别用0（空白对照）、0.5、1.0、2.0、4.0、8.0μmol·L-1的As2O3进行干预,24 h后收集细胞,定量PCR检测A549细胞Dll4 m RNA的水平、HUVECs Dll4和Notch-1 m RNA的水平。结果 A549细胞的Dll4 m RNA水平在0～2.0μmol·L-1组逐渐升高,2.0～8.0μmol·L-1组逐渐降低,且8.0μmol·L-1组的表达水平显著低于空白对照组（P〈0.01）;HUVECs的Dll4 m RNA水平在0～1.0μmol·L-1组逐渐升高,1.0～8.0μmol·L-1组逐渐降低,且4.0和8.0μmol·L-1组的表达水平显著低于空白对照组（P〈0.01）;HUVECs的Notch-1 m RNA水平随给药浓度的增加而升高,各浓度组均高于空白对照组（P〈0.01）。结论在体外条件下,As2O3在较低浓度能促进A549细胞和HUVECs Dll4 m RNA表达,而在较高浓度则抑制其表达;As2O3能浓度依赖性提高HUVECs的Notch-1 m RNA表达水平。
Background The transcription factor, repressor of GATA-3 （ROG）, can simultaneously suppress the expression of T helper cells （Thl and Th2） cytokines. Since the suppression of Th2 cytokines by GATA-3 is well understood, it is postulated that there are other molecular targets of ROG that can suppress the expression of the Thl cytokines. We hypothesized that ROG might suppress the stimulators of T lymphocyte cytokines such as CD3, CD28, and inducible costimulator （ICOS）, or indirectly enhance the expression of cytokine suppressors such as T lymphocyte-associated antigen-4 （CTLA-4） and CD45. The objective of this study was to clarify the molecular targets of ROG involved in suppressing Thl or Th2 cytokines. Methods Real-time quantitative PCR （RT-PCR） and Western blotting were performed to evaluate the mRNA and protein levels of CD3, CD28, ICOS, CTLA-4, and CD45 in Thl and Th2 cells during various levels of ROG expression. Enzyme-linked immunosorbent assay （ELISA） was used to measure the levels of interferon-y （IFN-y） and intedeukin （IL）-4 in culture media of Thl and Th2 cells. Results The results showed that the mRNA and protein levels of ROG were relatively low in Thl and Th2 cells （P 〈0.01）. After ROG-pcDNA3.1 transfection, the mRNA and protein level of ROG was significantly elevated, while the expression of ICOS, IFN-y, and IL-4 was markedly down-regulated （P 〈0.01）. Conversely, transfection of ROG-siRNA led to inhibition of ROG expression and up-regulation of ICOS, IFN-y and IL-4 （P 〈0.01）. However, the expression levels of CD3, CD28, CTLA-4 and CD45 did not change in either ROG-pcDNA3.1 or ROG-siRNA-transfected Thl and Th2 cells （P 〉0.05）. Conclusion It is concluded that ROG can inhibit the expression of Thl and Th2 cytokines by down-regulating the expression of ICOS, which might be a potential molecular target for asthma treatment.
