Bradykinin mediates the secretion of coagulation factor VIII by mouse dendritic cells via bradykinin 2 receptor (B2R) activation

Recent experimental evidences proved that kinins, which are a family of octa‐ to decapeptides structurally related to bradykinin (BK), have modulatory effects on multiple players of the immune system, including macrophages, dendritic cells (DC), T and B lymphocytes, and induce the activation, proliferation, migration, and effector functions of these cells. Kinins exert their biological activities by activating two pharmacologically different heterotrimeric G‐protein coupled receptors: B1R, or B2R. Herein we report, for the first time, that BK peptide (RPPGFSPFR) induces the production of coagulation Factor VIII by mouse DCs in response to the activation of B2R. C57BL/6J wild type mice were treated with: 1) BK, 2) Captopril. an ACE (angiotensin‐converting enzyme) inhibitor expected to enhance the endogenous BK and 3) a combination of both drugs (BK+CAP) for 12 hours. Following treatment, the DCs were purified from control (untreated) and from the BK/CAP/CAP+BK treated mice by immunomagnetic sorting with anti‐mouse CD11c biotin‐conjugated antibodies. The purified DCs were shown to have a significant dose‐response increase in the expression of MHC class II molecules (MHC‐II), and the co‐stimulatory molecules B7‐1 and B7‐2 (p<0.05) with the highest expression detected in the DCs purified from mice treated with BK+CAP. The activation of DCs was further positively correlated with the increase in the B2R expression. The activation of DCs in response to the BK/CAP/BK+CAP treatment was further supported by two comparative proteomics approaches: 1) 2D‐DIGE protein expression profiling and 2) label‐free 1D SDS‐PAGE coupled with nanoLC/MS/MS analysis of tryptic peptides derived from DCs protein lysates. Comparative Ingenuity Pathway Analysis (IPA) identified the upregulated cellular pathways in the BK/CAP/BK+CAP treated DCs. This analysis highlighted a wide spectrum of molecules involved in the DCs migration/chemotaxis, MHC‐I and MHC‐II expression, and antigen presentation pathways, inflammation and cytokines secretion. Moreover, both the comparative proteomics and the transcriptomic (mRNA) analyses demonstrated the significant upregulation of the coagulation pathway, positively correlated with the activation of DCs purified from treated mice. In particular, coagulation factor VII was found to be upregulated by any of the BK/CAP/BK+CAP treatments using proteomics approaches, while coagulation factors V, VIII, and XIIIa1—together with thrombin receptors PAR2 and PAR3—were observed to be upregulated by the same treatment using transcriptomic analysis. Production of coagulation factors VIII and V by BK‐activated DCs was validated by RT‐PCR, and by immunofluorescence studies.Pharmacological modulation of DC by BK and captopril (ACEI): comparative proteomics profiling by 2D‐DIGE electrophoresis followed by nanoLC MS/MS