CP-25 inhibits the functions of activated human B cells through regulating BAFF-TRAF2-NF-κB and TNF-alpha-TRAF2-NF-κB signaling

OBJECTIVE This study was to investigate the effects of CP-25 on the functions of activated human B cells through regulating BAFF and TNF-alpha signaling.METHODS B cells from peripheral blood mononuclear cells（PBMCs） of normal human were isolated using magnetic cell separation（MACS） by a positive selection.B cells(107 cells·mL;) were stimulated by BAFF(100 ng·mL;)or TNF-alpha(100 ng·mL;) for two hours,and then were treated with CP-25(10-5 mol·L;) or Rituximab(5 μg·mL;) or Etanercept(10 μg·mL;).B cell proliferation was detected by CCK-8.B cell subsets and BAFF receptors（BAFFR,BCMA and TACI） were analyzed by flow cytometry.The expression of TNFR1 and TNFR2 on B cells was analyzed by flow cytometry.The expression of MKK3,MKK6,P-p38,P-p65,TRAF2 and p100/52 was analyzed by Western blotting.RESULTS CP-25 inhibited B cells proliferation stimulated by BAFF or TNF-alpha.CP-25,Rituximab and Etanercept reduced the percentage and numbers of CD19;B cells,CD19;CD20;B cells,CD19;CD27;B cells and CD19;CD20;CD27;B cells induced by BAFF or TNF-alpha.CP-25 down-regulated the high expression of BAFFR,BCMA and TACI stimulated by BAFF or TNF-alpha.CP-25,Rituximab and Etanercept down-regulated significantly the expression of TNFR1 and TNFR2 on B cell stimulated by BAFF or TNF-alpha.CP-25,Rituximab and Etanercept down-regulated the expression of MKK3,P-p38,P-p65,TRAF2 and p52 in B cells stimulated by BAFF and the expression of TRAF2 and P-p65 in B cells stimulated by TNF-alpha.CONCLUSION CP-25 regulated moderately activated B cells function by by regulating the classical and alternative NF-κB signaling pathway mediated by BAFF and TNF-alpha-TRAF2-NF-κB signaling pathway.This study suggests that CP-25 may be a promising anti-inflammatory immune and soft regulation drug.