A glucose-rich heteropolysaccharide from Marsdenia tenacissima (Roxb.) Wight et Arn. and its zinc-modified complex enhance immunoregulation by regulating TLR4-Myd88-NF-κB pathway

A previously unreported immunological polysaccharide (MTP70–1) was obtained from Marsdenia tenacissima (Roxb.) Wight et Arn. MTP70–1 (2738 Da) is a heteropolysaccharide that mainly consists of (1 → 5)-linked-L-Araf, t-D-Glcp, (1 → 3,5)-linked-L-Araf, (1 → 4)-linked-D-Galp, (1 → 6)-linked-D-Glcp, and (1 → 3,6)-linked-D-Manp. In vitro cell assays revealed that MTP70–1 exhibits moderate immunomodulatory effects at the cellular level, and MTP70–1 was further modified with zinc to improve these effects. These modifications enhanced the immunomodulatory effects of MTP70–1, as phagocytosis was enhanced, the secretion of cytokines (TNF-α, IL-6, IL-1β, and IL-18) was increased, and the generation of chemokines (NO and ROS) in macrophages was enhanced. The intracellular mechanism by which MTP70–1 and MTP70-Zn activate macrophages was further revealed to be closely related to the TLR4-Myd88-NF-κB signaling pathway. In addition, a microscale thermophoresis binding (MST) assay confirmed that Zn modification can effectively enhance the binding affinity of MTP70–1 for TLR4. Ultimately, better immune-enhancing activity was attained with MTP70-Zn than MTP70–1. The immune-enhancing activity of MTP70-Zn was further demonstrated through zebrafish assays, which revealed that MTP70-Zn can effectively enhance the proliferation of macrophages and neutrophils.