Fig 7. Stimulation of CD40 expression and ISRE driven luciferase signals by infected red blood cells (iRBCs), parasite DNA/RNA, or parasite GPI.
(A) Flow cytometry counts of RAW Lucia cells containing N67 iRBCs 1 h and 8 h post incubation. The top two panels are RBCs, and the bottom panels are iRBCs. SSC, side-scattered light; CF-SE, signals of carboxyfluorescein succinimidyl ester (CF-SE) labeled RBCs or iRBCs (B) CD40 protein expression in RAW Lucia cells after incubation with RBCs or iRBCs. The antibody was against an epitope at C-terminus of the CD40 protein. Anti-β-actin was used as protein loading control. (C) The same as (B) but in DC2.4 cells. (D) ISRE promoter driven luciferase signals in RAW Lucia cells after ingestion with RBCs or iRBCs. (E) CD40 protein expression in RAW Lucia cells stimulated with parasite genomic DNA or RNA with or without the presence of lipofectamine. (F) Luciferase activities driven by ISRE promoter after stimulation with parasite DNA or RNA as in (E). (G) CD40 protein expression in RAW Lucia cells stimulated with parasite DNA or RNA with or without DNase or RNase treatment, respectively. EDTA and/or lipofectamine (Lipo) were used as buffer controls. (H) Luciferase activities driven by ISRE promoter after stimulation with parasite DNA or RNA with or without DNase or RNase treatment, respectively, as in (G). (I) CD40 protein expression in RAW Lucia cells stimulated with LPS, Pam3CSK4, and Plasmodium falciparum GPI (glycophosphatidylinositol) dissolved in ethanol (marked with EH) or H2O. (J) Protein band intensities in (I) relative to non-stimulated in ethanol (NoneEH = 1). (K) Luciferase activities driven by ISRE promoter in RAW Lucia cells stimulated with LPS, Pam3CSK4, and P. falciparum GPI as in (I). For (D), (F), (H), and (K), data are means+s.d. from three experiments; t-test, *P<0.05; **P<0.01; ***P<0.001. For all the experiments, 5 × 105 cells were used.