摘要
目的分析磷脂酰肌醇激酶3抑制剂2-(4-吗啉基)-8-苯基-4氢-1-苯并吡喃-4酮(LY294002)联合利妥昔单抗注射液对白血病T细胞的作用机制。方法Jurtak细胞人外周血白血病T细胞按照排板顺序分为对照组、LY294002组、利妥昔单抗组、联合用药组。对照组不加药物,LY294002组加40μmol/L LY294002,利妥昔单抗组加375 mg/mm^(2)利妥昔单抗注射液,联合用药组加40μmol/L LY294002和375 mg/mm^(2)利妥昔单抗注射液。作用48 h后,用MTT比色测定3个干预组和对照组中Jurkat细胞的抑制情况,流式细胞仪测定Jurkat细胞的生长周期情况,Transwell和划痕实验检测Jurkat细胞侵袭能力和转移能力,RT-q PCR法测定PI3k、Akt mRNA的表达,使用蛋白质印迹法测定Caspase-3、Bcl-2蛋白和P-gp表达。结果分别干预24、48 h后,与对照组相比,3个干预组增殖率和被侵袭细胞数均显著减少,联合用药组也显著低于LY294002组和利妥昔单抗注射液组,差异均有统计学意义(P<0.05),而LY294002组和利妥昔单抗注射液组相比差异无统计学意义(P>0.05)。与干预24 h相比,干预48 h后,LY294002组、利妥昔单抗注射液组和联合用药组增殖率和被侵袭细胞数均显著减少,差异均有统计学意义(P<0.05)。与对照组相比,3个干预组P13K和Akt mRNA表达、Bcl-2和P-gp蛋白表达和S期细胞占比均显著降低,Caspase-3蛋白表达和G_(0)/G_(1)期细胞占比显著增加,联合用药组P13K和Akt mRNA表达、Bcl-2和P-gp蛋白表达和S期细胞占比也显著低于LY294002组和利妥昔单抗注射液组,Caspase-3蛋白表达和G_(0)/G_(1)期细胞占比也显著低于该两组,差异均有统计学意义(P<0.05),而LY294002组和利妥昔单抗注射液组相比差异无统计学意义(P>0.05)。结论相比单独用药,LY294002联合利妥昔单抗注射液用药对诱导Jurkat细胞凋亡,抑制P13K Akt信号转导通路的活性,抑制细胞的侵袭、增殖,有明显的促进作用。
Objective To analyze the mechanism of phosphatidylinositol kinase 3 inhibitor 2-(4-morpholine)-8-phenyl-4 h-1-benzo pyran 4 ketone(LY294002)combined with rituximab injection on leukemia T cells.Methods Jurtak cell human peripheral blood leukemia T cells were divided into control group,LY294002 group,rituximab group,and combination group according to the order of platooning.The control group received no drug,the LY294002 group added 40μmol/L LY294002,and the rituximab group added 375 mg/mm^(2) rituximab injection and the combination group was given 40μmol/L LY294002 and 375 mg/mm^(2) rituximab injection.After 48 hours of intervention,the inhibition of Jurkat cells in the three groups of experimental group and control group was measured by MTT colorimetry,the growth cycle of Jurkat cells was measured by flow cytometry,the invasion and metastasis ability of Jurkat cells were measured by Transwell and scratch test,the expression of PI3K Akt mRNA was detected by RT-qPCR,Caspase-3,Bcl-2 and P-gp were detected by Western blotting.Results After 24 and 48 hours of intervention,compared with the control group,the proliferation rate and the number of invaded cells in the 3 intervention groups were significantly reduced,the combination group was also significantly lower than LY294002 group and rituximab group,the differences were statistically significant(P<0.05),however,there was no statistically significant difference between the LY294002 group and the rituxan group(P>0.05).Compared with the intervention for 24 h,the proliferation rate and the number of invaded cells in the LY294002 group,rituximab group and combined group were significantly reduced after the intervention for 48 h,the differences were statistically significant(P<0.05).Compared with the control group,the expression of P13K and Akt mRNA,Bcl-2 and P-gp protein expression and the proportion of S-phase cells in the three intervention groups were significantly reduced,and the expression of Caspase-3 protein and the proportion of cells in G_(0)/G_(1) phase increased significantly,and the expression of P13K and Akt mRNA,Bcl-2 and P-gp protein expression and the proportion of S-phase cells in the combination group were also significantly lower than that of the LY294002 group and rituxima group,and the expression of Caspase-3 protein and the proportion of cells in G_(0)/G_(1) phase were also significantly lower than those of the two groups,the differences were statistically significant(P<0.05),however,there was no statistically significant difference between the LY294002 group and the rituxan group(P>0.05).Conclusion LY294002 combined with meiluo has a significant promoting effect on inducing apoptosis of Jurkat cells,inhibiting the activity of P13K Akt signal transduction pathway,and inhibiting the invasion and proliferation of cells,compared with the drug.
作者
李玉
董彦成
李小平
LI Yu;DONG Yan-cheng;LI Xiao-ping(Department of Hematology,the 920th Hospital of the Joint Logistics Support Force of the People's Liberation Army,Kunming Yunnan 650032,China)
出处
《临床和实验医学杂志》
2022年第6期569-573,共5页
Journal of Clinical and Experimental Medicine
基金
云南省科技厅科技计划项目(编号:202102AA100011)。
