Figure 3

Staphylococcal LTA inhibits LPS-induced B cell proliferation through interaction with accessory cells in an indirect manner. (a) Splenocytes were stained with PE-conjugated rat anti-mouse TLR2 antibody and TLR2 surface expression on gated CD11c⁺, F4/80⁺, B220⁺, or CD3⁺ cells was analyzed by flow cytometry. The gray-filled area indicates histograms of the isotype control. One representative result from three similar results is shown. (b) Splenocytes were incubated with Alexa Fluor^® 546-conjugated Sa.LTA for 1 h at 4 °C and the binding of Sa.LTA to CD11c⁺, F4/80⁺, B220⁺, or CD3⁺ cells was analyzed by flow cytometry. The number indicates mean fluorescence intensity (MFI). One representative result from three similar results is shown. (c) Splenic B cells were negatively purified through magnetic cell sorting as described in the Materials and Methods. Whole splenocytes or purified B cells were incubated with Sa.LTA and/or LPS at the indicated concentrations for 72 h and cell proliferation was determined by [³H]-thymidine incorporation. Data are expressed as the average cpm ± standard deviation of three separate experiments. *Indicates P < 0.05 compared with LPS treatment group without Sa.LTA.