To investigate the therapeutic efficacy of lipoteichoic acid of Bifidobacterium (BLTA) in combination with 5-fluorouracil (5-FU) treatment on the mice bearing inoculated hepatoma-22 (H₂₂) cells and the effects of BLTA on immunological regulation of organism, and explore its mechanisms.

Tumor-bearing mice were treated with 5-FU alone, BLTA alone or BLTA in combination with 5-FU. The tumor size were observed and measured regularly. The growth inhibiting rate (IR) of tumor was detected. MTT assay was used to evaluate the proliferation of T lymphocytes and splenic NK cell and CTL activity. Enzyme linked immunosorbent assay (ELISA) was used to detect the change of IFN-Γ. FCM was used to detect T subgroup ratio of spleen cells of tumor-bearing mice. Expression change of mRNA and proteins of Foxp3 and TIM-3 were detected by Real-Time-PCR and Western blot in tumor-bearing mice tumor tissue.

Both 5-FU and BLTA had inhibition effect on tumor-growth. While in the 5-FU+BLTA group, the inhibition of tumor growth was more significant, with increased T lymphocyte proliferation and IFN-Γproduction of spleen cells. Spleen cells of tumor-bearing mice had high CD4⁺CD25⁺regulatory T cell (CD4⁺CD25⁺T_reg) ratio and high mRNA and proteins expression of Foxp3 and TIM-3, but in the BLTA and 5-FU group, CD4⁺CD25⁺T_reg ratio degraded, with down regulation mRNA and proteins expression of Foxp3 and TIM-3. But CD4⁺ T cells also decreased in spleen cells of tumor-bearing mice by alone 5-FU treated, splenic NK cell and CTL activity also degraded, while CD4⁺ T cells and splenic NK cell and CTL activity significantly increased by BLTA treated. BLTA in combination with 5-FU could also enhance the ratio of CD4⁺ T cells and splenic NK cell and CTL activity.

The present study suggested that BLTA in combination with 5-FU could enhance antitumor effect, with inhibiting TIM-3/TIM-3L pathway, cutting down immunosuppressive activity of CD4⁺CD25⁺ T_reg and enhancing cell-mediated immunity.