Objective:To investigate the effect of ketamine on the differentiation of human T helper(TH) cells and activity of T-bet and GATA3 in vitro. Methods: Ten male healthy volunteers (aged 20~45y,BMI 18~25 kg/m2) were enrolled in this study. Tweenty milliliter peripheral venous blood was collected from each volunteer and peripheral blood mononuclear cells(PBMC) were isolated and incubated in the presence of phorbol-myristate-acetate (PMA)(25 ng/ml) plus ionomycin (1 -滋g/ml) and various concentrations of ketamine(0,50,100,200 -滋mol/L). TH subsets,supernatant interferon γ (IFN-γ) and interleukin 4 (IL-4) levels,and the DNA binding activity of intracellular transcriptional factors T-bet and GATA3 were detected and analyzed after four hours culture. Results: TH1 and TH2 cell numbers,IFN-γ and IL-4 levels were significantly increased by PMA and ionomycin stimulation. Ketamine dose-dependently decreased TH1 and TH2 cell numbers,IFN-γ,and IL-4 levels,but increased the ratio of TH1/TH2 and IFN-γ/IL-4 in the presence of PMA and ionomycin. T-bet and GATA3 activities in PBMC were significantly increased by PMA and ionomycin stimulation. Ketamine dose-dependently decreased T-bet and GATA3 activity,but increased the ratio of T-bet/GATA3 activity in the presence of PMA and ionomycin. Conclusion: Ketamine can suppress TH cell differentiation and subsequent cytokines. But it increase TH1/TH2 ratio after PMA and ionomycin stimulation,which may be related to its regulation on T-bet and GATA3 activity.