T-bet are required for radiofrequency ablation-induced anti-tumor immune response

Abstract

Objective To investigate the role of transcription factor T-box expressed in T cells (T-bet) in radiofrequency ablation (RFA)-induced anti-tumor immune response. Methods C57BL/6 mice were inoculated with B16 melonoma cells on bilateral flanks, and RFA was performed for tumors on the right flank when the tumor volume reached about 500 mm3. The number and subset of the lymphocytes infiltrating into the tumor on the left flank were analyzed by flow cytometry 3 days after RFA. The mRNA levels of several cytokines were tested by quantitative real-time PCR. The expression of T-bet in T cells and NK cells were also analyzed by FCM. In addition, we established RFA treatment model in T-bet-/- mice. The infiltrating immune cells were analyzed. The growth curve of the left tumor and survival curve of mice were also compared to untreated wild-type mice. Results RFA induced an inhibition of tumor growth on the contralateral flank and prolonged the survival of tumor-bearing mice. The survival time of the treated group and the untreated group was (29.0±1.4) and (21.0±0.7) days respectively P=0.000). In the control and RFA-treated group, the number of CD45+ , CD3+ , CD4+ FoxP3-, CD8+ and NK (NK1.1+ ) cells infiltrating into the tumor on the left flank was 1.40±0.32 vs. 6.22±0.80, 0.46±0.08 vs. 2.06±0.11, 0.14±0.11 vs. 0.53±0.20, 0.20±0.03 vs. 0.68±0.10 and 0.05±0.02 vs. 0.17±0.03 (×106), respectively (P=0.000). The number of CD4+ FoxP3+ was 0.06±0.01 vs. 0.07±0.02 (×106) (P=0.978). Compared to the control group, the relative mRNA level of interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α) and IL-2 was 6.50±0.51, 5.02±0.48 and 4.12±0.31 in the RFA-treated group (P=0.000). The ratio of CD4+ T, CD8+ T and NK cell expressing T-bet was (22.30±0.60)% vs. (42.60±3.10)%, (28.40±2.20)% vs. (48.30±2.50)% and (9.10±0.20)% vs. (19.10±1.30)% in the control and RFA-treated groups, respectively (P=0.000). In T-bet-/- mice, the survival time was (21.00±1.20) d in RFA-treated group, and (21.00±0.56) d in the control. There is no significant difference between the two groups (P=0.981). In the control and RFA-treated group, the number of CD45+ , CD3+ , CD4+ FoxP3-, CD4+ FoxP3+ , CD8+ and NK (NK1.1+ ) cells infiltrating into the tumor on the left flank was 1.46±0.59 vs. 1.55±0.21, 0.43±0.04 vs. 0.45±0.10, 0.20±0.03 vs. 0.22±0.04, 0.09±0.01 vs. 0.11±0.01, 0.15±0.03 vs. 0.13±0.09 and 0.04±0.02 vs. 0.04±0.01 (×106), respectively (P=0.454, 0.982, 0.800, 0.987, 0.462, 0.278). Conclusion T-bet plays an important role in anti-tumor immune responses induced by RFA. Key words: Radiofrequency ablation; Anti-tumor immunity; Transcription factor; T-bet