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目的研究信号转导中的蛋白激酶C(PKC)途径与磷酸肌醇3激酶(PI3K)在人外周血γδT细胞分泌Th1型细胞因子IL-2与IFN-γ中的作用。方法人PBMC先用PI3K抑制剂Ly294002、PKC抑制剂Rottlerin或NF-κB抑制剂TPCK分别预处理1h,再加入结核杆菌低分子量多肽抗原(Mtb-Ag)和rhIL-2刺激3 d,最后用低浓度PMA+Ionomycin短时再刺激后检测γδT细胞IL-2和IFN-γ分泌;并同时检测αβT细胞作为对照。结果无抑制剂的对照组γδT和αβT细胞中分泌IL-2的比例分别为44.5%和15.2%,分泌IFN-γ的比例分别为51.1%和10.7%;Ly294002、Rottlerin、TPCK组分泌IL-2的γδT和αβT细胞均显著减少(P<0.05或P<0.01),后二组分泌IFN-γ的γδT和αβT细胞亦显著减少(P<0.05或P<0.01);但Ly294002组分泌IFN-γ的γδT和αβT细胞反而上升至77.2%和22.3%(P<0.01)。结论PKC途径活化促进人γδT细胞分泌IL-2与IFN-γ;PI3K亦促进人γδT细胞分泌IL-2但在其分泌IFN-γ中起负向作用。
Objective To investigate the role of protein kinase C (PKC) pathway and phosphoinositide 3 kinase (PI3K) signaling in the secretion of Th1 cytokines IL-2 and IFN-γ from human peripheral blood γδT cells. Methods Human PBMCs were pretreated with PI3K inhibitor Ly294002, PKC inhibitor Rottlerin or NF-κB inhibitor TPCK for 1 h, then treated with Mtb-Ag and rhIL-2 for 3 days, respectively. The concentrations of PMA + Ionomycin were detected by RT-PCR. The levels of IL-2 and IFN-γ in γδT cells were detected by RT-PCR. ΑβT cells were also detected as control. Results The percentages of secreted IL-2 in γδT and αβT cells without control were 44.5% and 15.2%, respectively, and those secreting IFN-γ were 51.1% and 10.7% respectively. Ly294002, Rottlerin and TPCK secreted IL-2 (P <0.05 or P <0.01). The γδT and αβ T cells secreting IFN-γ in the latter two groups also decreased significantly (P <0.05 or P <0.01). However, IFN-γ secreted by Ly294002 ΓδT and αβT cells increased to 77.2% and 22.3%, respectively (P <0.01). Conclusion Activation of PKC promotes the secretion of IL-2 and IFN-γ by human γδT cells. PI3K also promotes the secretion of IL-2 by human γδT cells but plays a negative role in the secretion of IFN-γ.