Figure 7.

A Novel DPP4/CD32b/NF-κB Circuit in CRP-Rich Diabetic Condition

(A) Immunofluorescence staining shows the co-localization of DPP4 and CD32b in the diabetic injured kidney of CRP^tg db/db mice at 24 weeks old (original magnification ×400). (B) The predicted binding site of NF-κB on the evolutionarily conserved region of DPP4 in human (yellow highlighted at right lower panel) and mouse genomes by ECR browser, where ChIP assay shows that NF-κB physically binds DPP4 promoter in response to CRP (10 μg/mL) in HK-2 cells at 1 h. (C) Co-immunoprecipitation (coIP) assay demonstrates that CRP triggers the binding of DPP4 to CD32b in HK-2 cells at 24 h after stimulation. (D and E) Real-time PCR (D) and western blot (E) analysis detect that DPP4 inhibition suppresses CRP-induced CD32b expression in HK-2 cells. (F and G) Real-time PCR (F) and western blot (G) analysis show the addition of either NF-κB inhibitor BAY11-7085 (NF-κBi, 10 μM), DPP4i, or CD32b-neutralizing antibody (CD32bi, 10 μg/mL) also blocks the activation of CRP-induced DPP4/CD32b/NF-κB signaling circuit in HK-2 cells. Data represent the mean ± SEM for three independent experiments. ∗∗p < 0.01, ∗∗∗p < 0.001 versus control; ^#p < 0.05, ^##p < 0.01 versus CRP-stimulated HK-2 cells.