Immunostimulatory activity of sea buckthorn polysaccharides via TLR2/4-mediated MAPK and NF-κB signaling pathways in vitro and in vivo.

SP0.1-1, derived from Sea buckthorn (Hippophae rhamnoides L.), has been discovered to exhibit unique antioxidant activity. In this study, we investigated the immunomodulatory activity and mechanisms of SP0.1-1 on macrophage RAW 264.7 cells in vitro and immunosuppressive mice induced by cyclophosphamide in vivo. The results indicated SP0.1-1 strengthened the immune functions via promoting the proliferation of RAW264.7 cells and phagocytic activity, along with stimulating the release of NO, ROS and cytokines including TNF-α, IL-6, IL-1β and IFN-γ. Western blot and molecular docking analysis demonstrated that SP0.1-1 attached to the prime receptors TLR2 and TLR4 in RAW264.7 cells, and triggered the activation of MyD88-mediated MAPK and NF-κB signaling pathways, thereby exerting the immune response in RAW264.7 cells. However, the intervention of specific inhibitors against TLR2, TLR4, JNK, ERK, p38 and NF-κB blocked the TLR-mediated MAPK and NF-κB signaling pathways and downregulated the levels of NO and the aforementioned cytokines, thus suppressing the activation of macrophages. Therefore, it can be speculated that SP0.1-1 activated the macrophages principally via the TLR2/4-MyD88-mediated MAPK and NF-κB signaling pathways. Additionally, SP0.1-1 could protect against the cyclophosphamide-induced immunosuppression in mice, manifested by the improvement of body weight, immune organ indices, phagocytic index, and the relievement of spleen damage, along with the enhancement of cytokines TNF-α, IL-6, IFN-γ and immunoglobulin IgG and IgM. These findings will shed light on the molecular mechanism of SP0.1-1 on the immunoregulatory effect, and lay the foundation for exploiting a potential immunostimulatory agent of SP0.1-1.