Supercritical CO2 extracts of propolis inhibits tumor proliferation and Enhances the immunomodulatory activity via activating the TLR4-MAPK/NF-κB signaling pathway.

Propolis is a natural immunomodulator with anticancer activity. This study investigated the immunomodulatory mechanism and anti-tumor activity of supercritical CO₂ extracts of propolis (SEP) in tumor-bearing immunosuppression mice. We used cyclophosphamide (CTX) to construct the immunosuppressive mice model and then inoculated them with CT26 cells to build the CT26 tumor-bearing immunosuppression mice model. Upon treatment with SEP, tumor proliferation in mice was markedly suppressed, with tumor volumes decreasing from 1881.43 mm³ to 1049.95 mm³ and weights reducing from 2.07 g to 1.13 g. Concurrently, the immune system recovered well, and the spleen and thymus indexes increased significantly. The total T lymphocyte (CD3⁺ T cell) contents in the spleen and blood recovered from 11.88 % to 21.19 % and 15.32 % to 22.19 %, respectively. In addition, the CD4⁺ /CD8⁺ ratio has returned to a healthy level, 3.12 in the spleen and 5.42 in the blood. The levels of IL-1β, IL-6, and TNF-α were increased by 2.17, 2.76, and 7.15 times in the spleen, 2.76, 1.92, and 3.02 times in the serum. Moreover, the western blot results showed that SEP treatment increased the expression of toll-like receptor 4 (TLR4) and the phosphorylation of p38, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p65. These results indicated that SEP activated the immune activity of RAW 264.7 macrophages through the TLR4-mitogen-activated protein kinase (MAPK)/nuclear factor kappa B (NF-κB) signaling pathway to exert immunomodulatory function and inhibit tumor proliferation. This study facilitated the further application of SEP as a potential immunomodulatory and anti-tumor functional food.