微囊藻毒素LR对人HL7702细胞蛋白磷酸酶2A的影响

摘要

微囊藻毒素能显著抑制蛋白磷酸酶2A的活性,是公认的一类肝毒素.为进一步研究微囊藻毒素作用于离体细胞后细胞中PP2A相关指标的变化,实验用不同浓度的MC-LR对人正常肝细胞HL7702染毒24h后检测细胞中PP2A亚基蛋白和mRNA水平,并检测3、6、12和24h作用后细胞内PP2A的酶活性.结果发现,除B55α蛋白表达有明显的下降趋势且1000 nmol/L组有显著性降低外,其他亚基的蛋白没有明显改变;15种PP2A亚基mRNA水平在100 nmol/L组都显著升高,而500和1000 nmol/L组分别有7种和6种亚基的mRNA表现出显著性降低.用不同浓度MC-LR染毒6h后,细胞内PP2A酶活有显著性降低,并且呈现剂量依赖性关系,而3、12和24h染毒后酶活性没有明显变化.以上结果表明:一定浓度的MC-LR作用24h后影响了HL7702细胞PP2A相关亚基的转录,但没有明显改变蛋白质水平;MC-LR对细胞内PP2A的酶活抑制作用与染毒时间密切相关.%Microcystins (MCs) are hepatotoxins which inhibit the activity of protein phosphatase 2A (PP2A) notably.The cells were incubated with different concentrations of MC-LR to determine the effect of MCs on PP2A in HL7702 respectively for 24h.The mRNA and protein levels of PP2A subunits were detected.Furthermore, the activity of PP2A was assayed after 3h, 6h, 12h and 24h exposure.The protein levels were not changed significantly except B55α which decreased concentration-dependently and significantly in 1000 nmol/L.For mRNA levels, 15 subunits increased in 100 nmol/L group, whereas 7 subunits in 500 nmol/L group and 6 subunits in 1000 nmol/L group decreased.The activity of PP2A was inhibited significantly and concentration-dependently after 6h exposure while there were no changes in 3, 12 and 24h exposure.In conclusion, MC-LR affects the transcription of PP2A subunits after 24h exposure, without concomitant changes in the interrelated proteins.Exposure time might be a determinant when considering the effect of MC-LR on PP2A activity.