Fig. 5
Downregulation of CX3CR1 increases the expression of CCL2/R2 and proinflammatory cytokines TNFα and IL-1β. a Representative Western blot and quantification for CCL2 (15 kDa) in control siRNA and CX3CR1 siRNA-transfected THP-1 cells (mean ± SEM, n = 3 cultures, three wells per culture were pooled). Expression of CCL2 is significantly elevated in CX3CR1 siRNA-transfected THP-1 cells relative to controls. *p < 0.01, Student’s t test. b Representative Western blot and quantification for CCR2 (42 kDa) in control and CX3CR1 siRNA-transfected THP-1 cells ± pre-treatment with either SB203580 (p38 MAP Kinase inhibitor) or PD98059 (MEK inhibitor). c Quantification of CCR2 expression (pg/ml) using ELISA (mean ± SEM, n = 3 cultures). Transfection of THP-1 cells with CX3CR1 siRNA with PD98059 pre-treatment results in a significant increase in CCR2 expression. This is not observed following pre-treatment with SB203580. **p < 0.01 relative to control siRNA transfected. d, e TNFα (d) and IL1β (e) quantification by ELISA of THP-1 culture medium following transfection with CX3CR1 siRNA or 3 h stimulation with recombinant CCL2 (10, 50 and 100 ng/ml). CX3CR1 downregulation and all concentrations of CCL2 significantly increase TNFα and IL1β expression (mean ± SEM, n = 3 cultures). *p < 0.05 and**p < 0.01 relative to control, Student’s t test