Polysaccharides extracted from the leaves of Plantago palmata Hook.f. induce nitric oxide and tumor necrosis factor-α production by interferon-γ-activated macrophages

Ethnopharmacological data from eastern African traditional uses of Plantago palmata leaves suggest that some therapeutical activities could be dependent on their content in polysaccharides (PS). To further investigate immunomodulatory properties of these PS, they were extracted in water from leaves by maceration either at (i) 50°C without filtration (PS50); (ii) 50°C and filtration (PS50F); (iii) 100°C, without filtration (PS100); or 100°C and filtration (PS100F). The extract PS50 was further fractionated by high performance liquid chromatography (gel permeation chromatography) in three fractions namely F1, F2, and F3. The immunomodulatory properties of these four crude extracts and PS50 fractions were investigated by the measurement of nitric oxide (NO), tumor necrosis factor-alpha (TNF-α), and interleukin-10 (IL-10) production using interferon gamma-(IFN-γ)-activated macrophages and non-activated macrophages as control. Non-activated J774 cell line macrophages, treated with any of the PS extracts, did produce neither NO, nor TNF-α, nor IL-10. In contrast, IFN-γ-activated J774 macrophages synergized with PS50, but not with the three other crude extracts, to produce both NO and TNF-α, but not IL-10. Immunomodulatory effects due to traces of lipopolysaccharides (LPS) in PS extracts and fractions were ruled out by the use of macrophages from C3H/Hej mice known to be very low responders to LPS and similar results were obtained. In addition, F2 fraction from PS50 was particularly active in enhancing NO and TNF-α (but not IL-10) production by IFN-γ-activated C3H/Hej macrophages. These results suggest that PS from P. palmata leaves possess immunomodulatory properties by stimulating NO and TNF-α production by activated macrophages.