Background: High blood pressure (HBP) is an important risk factor for cardiac, renal and vascular dysfunction. Excess inflammation is the major pathogenic mechanism for HBP-induced target organ damage (TOD). N-acetyl-Ser-Asp-Lys-Pro (Ac-SDKP), a tetrapeptide specifically degraded by angiotensin converting enzyme (ACE), reduces inflammation, fibrosis and TOD induced by high blood pressure. Hypothesis: Ac-SDKP exerts its anti-inflammatory effects by inhibiting: 1) differentiation of bone marrow stem cells (BMSC) to macrophages, 2) activation and migration of macrophages and 3) release of the pro-inflammatory cytokine TNF- by activated macrophages. Methods: BMSC were freshly isolated from mouse tibias and femurs. Differentiation of murine BMSC to macrophages was analyzed by flow cytometry using F4/80 as a marker of macrophage maturation. Macrophage migration was measured in a modified Boyden chamber. TNF- release by activated macrophages was measured by enzyme-linked immunosorbent assay (ELISA). Myocardial macrophage activation in Angiotensin-II (Ang II) treated mice was detected by Western blotting of Mac-2 (galectin-3) protein. Interstitial collagen deposition was measured by Picrosirius red staining. Results: Ac-SDKP (10 nM) reduced differentiation of cultured BMSC to mature macrophages by 24.5% [F4/80 positivity: 14.09 ± 1.06 mean fluorescent intensity (MFI) for vehicle and 10.63 ± 0.35 for Ac-SDKP; p < 0.05]. Ac-SDKP also decreased galectin-3 and macrophage colony-stimulating factor (M-CSF)-dependent macrophage migration. In addition, Ac-SDKP decreased secretion of TNF- by macrophages stimulated with bacterial LPS. In Ang II-treated hypertensive mice, Ac-SDKP reduced expression of galectin-3, a protein produced by infiltrating macrophages in the myocardium and interstitial collagen deposition. Conclusion: This study demonstrates that part of the anti-inflammatory effect of Ac-SDKP is due to its direct effect on BMSC differentiation and macrophage activation, reinforcing the protective properties of Ac-SDKP in high blood pressure-induced inflammation and organ damage.