We have characterized and reported the immunostimulating properties of a novel polysaccharide - (1,4)-α-D-glucan (RR1)- isolated from the medicinal plant Tinospora cordifolia [23]. This novel glucan is water soluble and having (1, 4)-α-D-glycosidic linkages in the main chain and (1, 6)-α-D-glycosidic linked side chains at an interval of 6, 7 glucose units. The signaling mechanism of the novel (1,4)-a-D-glucan (RR1) was investigated in macrophages to evaluate its immunostimulating properties. When RAW264.7 macrophages were incubated with RR1 at 4°C, the novel glucan inhibited the phagocytosis of unopsonized zymosan A bioparticles in a dose-dependent manner. RR1 also inhibited the binding and internalization of opsonized zymosan A bioparticles, although at a lower level than laminarin. Incubation of macrophages with anti-CD11b mAb followed by RR1 failed to show any inhibitory effect on RR1-induced TNF-α synthesis confirming that complement receptor 3 (CR3) is not involved in the opsonic binding and internalization of RR1 in macrophages unlike zymosan A. The anti-CD11b mAb has significant inhibitory effect on the zymosan A-induced tumor necrosis factor (TNF)-α synthesis. RR1 induced TNF-α synthesis in macrophages in a dose-dependent manner which can be completely inhibited by the NF-κB inhibitor caffeic acid phenethyl ester (CAPE) or curcumin. RR1 activated NF-κB in a time- and dose-dependent manner and this modulation of nuclear NF-κB activity is associated with the degradation of I-κB a thus facilitating the translocation of NF-κB into the nucleus. RR1-induced NF-κB activity peaks at 8 h of RR1 stimulation while I-κB a degradation occurred within 1 h of stimulation. RR1-induced NF-κB activation occurred through TLR6 signaling as evidenced by the synthesis of IL-8 in TLR6-transfected HEK293 cells. These results show that the novel (1,4)-α-D glucan from Tinospora cordifolia activates the immune system through the activation of macrophages that occurs through TLR6 signaling, NF-κB translocation and cytokine production. (Supported by Miami Children’s Hospital Foundation research funds).

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