Abstract P197: Extracellular Ubiquitin Blocks CXCL12-Induced Proliferation of Cardiac Fibroblasts
CXCR4 receptors mediate in part hypertension-induced cardiac fibrosis. This suggests that endogenous CXCR4-receptor agonists stimulate cardiac fibroblast (CF) proliferation; however, this possibility is unexplored. Although CXCL12 (aka SDF-1α) is the best described endogenous CXCR4 agonist, ubiquitin1-76(U-76) exists in the extracellular compartment (10 to 100 nM) and is reported to be a CXCR4 agonist. Therefore, we investigated the ability of both CXCL12 and U-76 to stimulate growth of rat CFs. Low concentrations of CXCL12 (10 nM x 4 days) increased CF proliferation (from 38,973 ± 384 to 44,429 ± 774 cells/well, n=6, p<0.0001), and this effect was augmented by sitagliptin, a dipeptidyl peptidase 4 (DPP4) inhibitor (% increase by CXCL12: without sitagliptin, 14 ± 2 (n=6); with 1 μM of sitagliptin, 38 ± 9, n=6, p<0.0001). This finding is consistent with the known ability of DPP4 to metabolize CXCL12, i.e., sitagliptin inhibits the metabolism of CXCL12 and enhances CXCL12-induced effects. Not only did U-76 at low physiological concentrations (10 nM) not stimulate CF proliferation, U-76 surprisingly nearly abolished CF proliferation induced by CXCL12 + sitagliptin (% increase by CXCL12 + sitagliptin: without U-76, 58 ± 5, n=12; with U-76, 10 ± 7, n=12, p<0.0001). Ubiquitin1-74 (U-74), a metabolite of U-76, inhibited the pro-growth effects of CXCL12 + sitagliptin 10-fold more potently than did U-76. CFs expressed insulin degrading enzyme (IDE) mRNA (qRT-PCR), protein (western blot), and activity (IDE activity assay) and inhibition of IDE with the newly discovered potent IDE inhibitor 6bK (1 μM) prevented U-76 from blocking the growth effects of CXCL12 + sitagliptin. Analysis by mass spectrometry (selected ion monitoring) demonstrated that CFs converted U-76 to U-74 and this conversion was attenuated by 6bK. Conclusions: CFs express IDE and therefore convert U-76 to U-74; U-74 blocks CXCR4 receptors thus protecting against CXCL12 + DPP4 inhibitor induced CF proliferation. Implications: In patients with low IDE activity (due to genes or drugs) who are treated with DPP4 inhibitors, U-74 (CXCR4 antagonist) levels would be low and CXCL12 (CXCR4 agonist) levels would be high, thus possibly creating a “perfect storm” for CF over-proliferation and cardiac fibrosis.
Author Disclosures: E.K. Jackson: None. E. Mi: None. V.B. Ritov: None. D.G. Gillespie: None.
- © 2015 by American Heart Association, Inc.