Skip to main content
Fig. 5 | Journal of Hematology & Oncology

Fig. 5

From: Mesenchymal stromal cells as vehicles of tetravalent bispecific Tandab (CD3/CD19) for the treatment of B cell lymphoma combined with IDO pathway inhibitor d-1-methyl-tryptophan

Fig. 5

IDO pathway inhibitor D-1MT enhances the cytotoxicity induced by MSC-secreting Tandab (CD3/CD19) in vitro. a IDO mRNA expression in MSCs stimulated with IFN-γ. MSCs were cultured in the absence or presence of exogenous IFN-γ (20 ng/mL). Cells were harvested after 2 days in culture, total mRNA was extracted, and IDO message was assayed by real-time PCR. b IDO protein was determined by Western blot analysis. MSCs were harvested after cultured for 48 h, as in a, and total cell lysates were assayed for IDO protein. Results are representative of three independent experiments. c To determine enzyme activity of IDO, MSCs were cultured with or without IFN-γ (20 ng/mL) stimulation for 48 h. IDO enzyme activity was evaluated by spectrophotometric detection of the tryptophan metabolite, kynurenine, a product of IDO catabolism. d Influence of D-1MT on the proliferation of cells (Raji, PBMCs and MSCs). Cells were seeded in 96-well plates and treated with different concentrations of D-1MT (0–2000 μM) for 72 h. Cell proliferation was determined by MTT assay, and the y axis represents cell proliferation as a percent of the control. e D-1MT promotes the specific lysis of Raji cells induced by MSC-secreting Tandab (CD3/CD19). MSCs, Raji cells, and PBMCs were co-cultured as mentioned above with or without D-1MT (1 mM) for 24, 48, and 72 h. The residual Raji cells were detected by FACS with FITC-conjugated anti-CD19 antibodies. D-1MT, d-1-methyl-tryptophan. f Determination of kynurenine in the supernatant from the co-culture system. g Analysis of anergy-associated genes expression of CD98 (left panel) and Jumonji (right panel). The cells in the inserts were harvested in the indicated time. Total mRNA was extracted, and the messages of CD98 and Jumonji were assayed by real-time PCR. NS, normal saline. h Relative proliferation of T cells. The cells were harvested in the indicated time. Proliferation of T cells was detected using BrdU flow kit. The proliferation of T cells in PR group (without MSCs) was served as a reference. *P < 0.05 compared with the control group; ***P < 0.001 compared with the control group. Data shown are the mean ± SD of the three repeated experiments

Back to article page