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A7613 - Stromal Derived Factor-1 Is Important for Mesenchymal Stromal Cell Function: Impact on Cell Response During Sepsis
Author Block: M. Kwon1, K. Tsoyi1, S. Ghanta2, X. Liu3, M. A. Perrella3; 1Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Boston, MA, United States, 2Pediatric Newborn Medicine, Brigham and Women's Hospital, Boston, MA, United States, 3Pulmonary and Critical Care Medicine, Brigham and Womens Hospital, Boston, MA, United States.
Rationale: Sepsis is a complex disease process, characterized by a systemic inflammatory response to a severe infection. Mesenchymal stromal cells (MSCs) have beneficial effects in animal models of sepsis, including the ability to modulate the immune response and improve survival in mice. Stromal derived factor-1 (SDF-1) is important for mobilization of neutrophils from the bone marrow during sepsis, and this neutrophil response is critical for the host to clear the infection. We are investigating the importance of SDF-1 in MSC function, and its role in the interaction of MSCs with neutrophils during sepsis.
Methods: MSCs were harvested from compact bones of mice, and SDF-1 was silenced using SDF-1 shRNA (shSDF-1), versus a scrambled shRNA (shSCR) control. MSCs were treated with hydrogen peroxide (H2O2), and cell survival was measured (tetrazolium MTT assay). MSC migration was assessed using a transwell system. Phagocytosis of Escherichia [E.] coli bacteria was assessed in isolated neutrophils exposed to shSDF-1 or shSCR MSCs. AMD3100, an antagonist of a receptor of SDF-1, was also used in phagocytosis assays. Sepsis was induced in mice using cecal ligation and puncture (CLP, 2/3 cecal ligation, 21-gauge 2 holes). 500,000 MSCs were administered intravenously 2 and 24 hours after CLP, and survival was assessed over 7 days.
Results: SDF-1 expression was silenced in MSCs, and cell function assessed. When MSCs were exposed to oxidative stress (H2O2), cell survival was significantly worse in shSDF-1 MSCs. Moreover, shSDF-1 MSCs had a 3-fold decrease in migration compared with shSCR MSCs. Co-culture of neutrophils with shSCR MSCs increased neutrophil phagocytosis 9-fold compared with neutrophils alone, whereas neutrophil phagocytosis was blunted with shSDF-1 MSCs. Conditioned medium from MSCs was also able to increase E. coli phagocytosis by neutrophils, and neutrophils exposure to AMD3100 abrogated the phagocytic response. Next, we assessed the therapeutic impact of shSDF-1 MSCs in vivo. After CLP-induced sepsis, mice received vehicle (phosphate buffered saline), shSCR MSCs, or shSDF-1 MSCs. Mice treated with vehicle alone had a dramatic death response to CLP, with 10% survival at 7 days. Injection of shSCR MSCs led to a dramatic increase in mouse survival (70%) at 7 days, while the survival of mice receiving shSDF-1 MSCs was significantly diminished (33%).
Conclusion: These data support an important role for SDF-1 in MSC function, including an effect on cell survival, migration, and promoting phagocytosis of bacteria by neutrophils. Silencing of SDF-1 in MSCs results in decreased therapeutic benefit in experimental sepsis.