鞘氨醇激酶对胃癌细胞凋亡作用的初步研究

Effect of sphingosine kinase on the apoptosis of gastric carcinoma cells

目的:研究鞘氨醇激酶(SPK)对胃癌细胞凋亡的作用。方法:氯化铯密度梯度离心法纯化携带人野生型(rAd-SPKWT)及突变体(rAd-SPKDN)SPK的腺病毒,用噬斑分析和快速CPE法测定病毒感染滴度。以腺病毒为载体将SPK基因导入胃癌细胞BGC-823。分别以Westernblot、酶活性法检测外源基因表达和SPK酶活性。结果:获得了高滴度的rAd-SPKWT、rAd-SPKDN腺病毒;腺病毒能高效感染胃癌细胞BGC-823;外源SPK在胃癌细胞能有效表达,rAd-SPKWT增强SPK活性,rAd-SPKDN抑制SPK活性;高表达SPK可以抑制胃癌细胞对5-FU的敏感性,阻断SPK可以增强其对5-FU的敏感性。结论:SPK可以抑制胃癌细胞凋亡,SPK有可能成为胃癌新的治疗靶点。

Objective： To study the role of sphingosine kinase （SPK） in the regulation of the apoptosis of gastric carcinoma cells. Methods： The adenovirus carrying the wild type SPK gene （SPK^WT） and SPK mutant （ SPK^DN ） were constructed respectively ; viral infection titer was determined with plaque assay and CPE method. The effect of the adenoviral mediated-expression of these genes on the BGC-823 cells was evaluated, and we detect gene expression by Western blot and SPK enzyme activity assay. Results： Re-adenoviruses were amplified and purified; rAd-SPK^WT and rAd-SPK^DN can infect BGC-823 cells efficiently; Exogenous SPK may effectively express in BGC-823 cells, SPK enzyme activity assay suggested wide type SPK gene could increase SPK activity and mutant SPK gene could decrease SPK activity respectively; SPK could inhibit the apoptosis induced by 5-FU in the cells of BGC-823. Condusion：SPK could inhibit the apoptosis of the cells of BGC-823, which maybe a new target for tumor treatment.