In vitro TLR-mediated downmodulation of CD5 expression on leukemic cells. (A) TCL1^tg/tgTIR8^−/− mice were monitored during time (months indicated) by flow cytometry analysis of Peripheral Blood (PB) to detect leukemic clone (CD19⁺CD5⁺). 2 representative analysis (lower panel with CD5 downmodulation) of TCL1^tg/tg/TIR8^−/− mice are shown (n = 6). (B) CD19⁺enriched splenocytes from TCL1^tg/tgTIR8^wt/wt and TCL1^tg/tg/TIR8^−/− mice were cultured in vitro with increasing amounts of LPS or anti-IgM for 24 hours and analyzed by flow cytometry CD25 and CD86 expression. One representative experiment is shown (n = 5 each group). (C) CD19⁺ enriched splenocytes from TCL1^tg/tgTIR8^wt/wt mice were stained with CFSE and cultured in vitro with increasing amounts of LPS for 5 days. Flow cytometry analysis of CFSE dilution revealed a peak of proliferation in leukemic cells that was augmented by LPS treatment. One representative plot shown (n = 3). (D) CD19⁺enriched splenocytes (the same as in panel B) from TCL1^tg/tgTIR8^wt/wt and TCL1^tg/tg/TIR8^−/− mice were cultured in vitro with increasing amounts of LPS or anti-IgM for 24 hours and analyzed by flow cytometry for CD19 and CD5 expression. One representative experiment is shown (n = 5 each group). Mean fluorescence intensity (MFI) of CD19+CD5+ cells are indicated.