Figure 1.

LF 15-0195 prevents maturation and function of dendritic cells. (a) Phenotypic analysis of LF-treated dendritic cells (DC). Bone-marrow-derived DC were cultured in the presence of granulocyte-macrophage colony-stimulating factor (10 ng/ml) and IL-4 (10 ng/ml) for 7 days. Control mature DC (upper panels) were activated using tumor necrosis factor alpha (TNFα)/lipopolysaccharide (LPS) in the last 24-hour culture. DC (lower panel) were treated by addition of LF (10 ng/ml) in the culture medium from day 4 onwards, and fresh medium was added every 24 hours. DC were stained with FITC-conjugated mAbs and analyzed by flow cytometry. Results represent one of three experiments (n = 4 per group/experiment). (b) LF regulates cytokine expression in DC. DC were treated with LF as in (a). The supernatants of DC culture were collected and used to measure IL-12 and IL-10 levels by ELISA as described in Materials and methods. *P < 0.05, comparing untreated control DC.(c) LF inhibits DC allostimulatory capacity in a mixed leukocyte reaction. DC were pretreated with LF and subsequently stimulated with 10 ng/ml TNFα/LPS as described in (a). DBA/1 control DC and LF-treated DC, at indicated concentrations, were used as stimulators, and BALB/c splenocytes (1 × 10⁵/well) were used as responders. Stimulators and responders were cocultured, and proliferation was assessed as described in Materials and methods. Data shown are representative of three independent experiments (n = 4 per group/experiment).P < 0.05, comparing untreated control DC.(d) LF-treated DC regulate T helper cell deviation. LF-treated DC and PBS-treated control DC (10⁶) (DBA/1) were subsequently cultured with allogeneic (BALB/c) T cells (10⁷) for 48 hours. Supernatants were collected from the cultures and interferon gamma (IFNγ; Th1) and IL-4 cytokine (Th2) levels were measured by ELISA. Results represent one of three experiments (n = 4 per group/experiment). P < 0.05, comparing untreated control DC.