Role for toll-like receptor 4 in TNF-alpha secretion by murine macrophages in response to polysaccharide Krestin, a Trametes versicolor mushroom extract

Woody fungi and yeast preparations show promise in cancer treatment by activating anti-tumor immune responses. Macrophages (J774A.1) were treated with PSK, Reishi extract, scleroglucan or vehicle control. Pre-incubation with TLR4 blocking antibody inhibited TNF-alpha secretion by both J774A.1 cells and primary splenocytes but had inconclusive effect on scleroglucan-induced secretion of TNF-alpha. PSK induced TNF-alpha and IL-6 secretion by wild type but not by TLR4-deficient peritoneal macrophages. We conclude that constituents from PSK act as ligands for TLR4 receptors leading to induction of TNF-alpha and IL-6 inflammatory cytokines. Receptor-mediated differences may be due to structural differences in beta glucans or non-glucan constituents.

TLR4 blocking antibody inhibits PSK-induced TNF-alpha secretion by mouse splenocytes. Splenocytes were harvested from healthy C57Bl/6 mice and treated with media (control), PSK (125 μg/mL), PSK (125 μg/mL) after pretreatment with TLR4 blocking antibody, or pretreated with TLR4 blocking antibody alone. Supernatants were collected and frozen at −80 °C until time of assay for TNF concentration by ELISA. Results represent mean TNF concentration detected in supernatants from triplicate cultures.Figure optionsDownload as PowerPoint slide