Immunomodulatory effects of Radix isatidis polysaccharides in vitro and in vivo

Radix isatidis (R. isatidis) is a commonly used traditional Chinese herbal medicine, which has been used for thousands of years in China and is believed to have the pharmacological properties of heat-clearing and detoxification. Heat-clearing and detoxification are theories of traditional Chinese medicine meaning that R. isatidis could treat febrile disease by clearing heat and reducing swelling. Polysaccharides isolated from R. isatidis by water extraction and alcohol precipitation have exhibited numerous biological activities, including antiviral and immunomodulatory effects. The present study was performed to investigate the immunomodulatory effects of water-soluble R. isatidis polysaccharides (RIPs) on RAW264.7 macrophages and murine splenocytes, and attempt to preliminarily identify the mechanism of immunomodulation. In vitro, RIPs had a low cytotoxicity, as shown by CellTiter 96(R) AQ(ueous) One Solution Cell Proliferation Assay. RAW264.7 cells treated with different concentrations of RIP displayed different morphological changes, from a round shape and aggregation to polygonal shape and dispersion in a dose-dependent manner. In the 5 mg/ml RIP-treated group, the changes of morphology were as same as the lipopolysaccharide-treated group. RIP also significantly enhanced the release of nitric oxide as shown by Griess method, and the secretion of TNF-alpha and IL-6 in RAW264.7 cells was confirmed by ELISA assay. Western blotting revealed a significant increase of toll-like receptor-4 (TLR-4) in RIP-treated RAW264.7, suggesting that TLR-4 may be associated with the immunomodulatory mechanism of RIP. Animal experiments also demonstrated through ELISA assays a significant increase in IFN-gamma and IL-10 levels after the splenocytes of RIP-immunized mice were stimulated by inactivated herpes simplex virus type 2. The immune function of RIP-immunized mice was improved. The present study suggested that RIP could be potentially used as a novel immunomodulator.