Involvement of membrane microdomains in oxidative stress-induced invasion of hepatoma cells

研究了膜微结构域在氧化应激诱导的肝癌细胞侵袭过程中的作用。我已经报道过活性氧可以促进肝癌细胞的侵袭，并且肝细胞生长因子（HGF）与其受体c-met之间的自分泌途径参与了这一促进。本研究观察到缺氧培养条件下侵袭活性降低，但不改变HGF基因表达，提示氧化应激诱导肝癌细胞侵袭变化存在新的调控途径。AH109A细胞在其细胞膜上表达整合素α2、α5、α6、β1，这些整合素被证实定位于其膜微域。甲基-β-环糊精影响AH109A的侵袭，表明膜微结构域参与了AH109A的侵袭。因此，氧化应激特别是缺氧对肝癌细胞侵袭的影响被广泛研究。缺氧条件下培养的AH109A细胞侵袭活性降低，但对正常细胞单层的粘附活性不降低，HGF的生成不减少。缺氧培养条件下，AH109A细胞中c-met的表达水平没有变化。最近报道c-met与整合素的关联在HGF促进细胞运动中起重要作用。这些结果表明，缺氧可能会影响AH109A细胞膜微域c-met与整合素的结合，从而改变其侵袭活性。虽然需要进一步的研究，如c-met和整合素的共免疫沉淀试验，来阐明缺氧诱导的侵袭活性降低的确切机制，但本研究获得了膜微结构域可能参与氧化应激诱导的肝癌侵袭变化的一些证据。

The possible involvement of membrane microdomains in oxidative stress-induced change in invasion of hepatoma cells was investigated. I have already reported that reactive oxygen species could promote hepatoma cell invasion and that the autocrine pathway between hepatoyte growth factor (HGF) and its receptor, c-met, is involved in this promotion. In this research, the decrease in the invasive activity under the hypoxia culture condition without changing HGF gene expression was observed, suggesting the presence of the novel regulation pathway in oxidative stress-induced change in hepatoma cell invasion. AH109A cells expressed integrins α2, α5, α6, β1 on their cell membrane and these integrins were proved to be localized in their membrane microdomains. Moreover, methyl-β-cyclodextrin affected AH109A invasion, suggesting the involvement of membrane microdomain in their invasion. So, the effect of oxidative stress, especially hypoxia, on hepatoma cell invasion was extensively investigated. AH109A cells, when cultured under the hypoxia condition, showed decreased invasive activities, but they did not show decreased adhesive activities to the normal cell monolayer and decreased HGF production. The expression level of c-met in AH109A cells did not change under the hypoxia culture condition. Recently the association of c-met with integrins is reported to be important in the promotion of cell motility by HGF. These results suggest that hypoxia may affect the association of c-met with integrins in membrane microdomains of AH109A cells, thus changing their invasive activities. Although further studies, such as co-immunoprecipitation assay of c-met and integrins, are thought to be needed to clarify the precise mechanism for hypoxia-induced reduction in invasive activities, some evidences for the possible involvement of membrane microdomains in oxidative stress-induced change in hapatoma invasion were obtained by this research.