1. We used EGFP (Enhanced Green Fluorescent Protein) labelled bone marrow to transplant into mice with middle cerebral artery occlusion (stroke model) to study the contribution of BM derived cells to tissue repair. Granulocyte colony-stimulating factor(G-CSF) induces proliferation of bone marrow derived cells. G-CSF is neuroprotective after experimental brain injury, but the mechanisms involved remain unclear. Stem cell factor (SCF) is a cytokine important for the survival and differentiation of hematopoietic stem cells. Its receptor (ckit or CD117) is present in some endothelial cells.
We aim ed to determine whether the combination of G-CSF/SCF induces angiogenesis in the central nervous system by promoting entry of endothelial precursors into the injured brain and causing them to proliferate there. We induced permanent middle cerebral artery occlusion in female mice that previously underwent sex-mismatched bone marrow transplantation from enhanced green fluorescent protein (EGFP) expressing mice. G-CSF/SCF treatment reduced infarct volumes by more than 50% and resulted in a 1.5-fold increase in vessel formation in mice with stroke, a large percentage of which contain endothelial cells of bone marrow origin. Most cells entering the brain maintained their bone marrow identity and did not trans-differentiate into neural cells. G-CSF/SCF treatment also led to a 2-fold increase in the number of newborn cells in the ischemic hemisphere. These findings suggest that G-CSF/SCF treatment might help recovery through induction of bone marrow derived angiogenesis, thus improving neuronal survival and functional outcome. (Blood. 2008;111:5544-5552. We continued our studies to analyze the relationship between the mobilization of BM derived cells and brain repair in stroke. We found that TGF-alpha has a significant effect in recruiting neural stem cells to the site of injury and in the induction of neural differentiation. We have recently published these observation in the journal """"""""Neuroscience"""""""" (Transforming growth factor induces angiogenesis and neurogenesis following stroke;e-published,doi:10.1016/ j.neuroscience. 2009.05.050) 2. We have initiated several studies to examine the effect of intravenously injected bone marrow stromal cells in disease models. We used CLP (cecal ligation and puncture) to induce sepsis in mice. Acute kidney injury (AKI) in septic shock is associated with a high mortality in ICU patients. Bone marrow stromal cells (BMSCs), which are often referred to as mesenchymal stem cells, have been shown to improve outcome in a number of different animal injury models by altering inflammation, apoptosis, and necrosis. In this study we set out to determine whether BMSCs can attenuate the severity of sepsis-induced AKI. Sepsis was induced in C57/BL6 mice using cecal ligation and puncture (CLP). BMSCs were obtained from the bone marrow of 6-8 week old mice. Intravenous administration of BMSCs (1 million cells/mouse) immediately before CLP surgery resulted in significantly longer survival and improved kidney, liver and pancreatic function. Such organ protection was not obtained with either hematopoietic stem cells or necrotic BMSCs. Searching for a mechanism of action, we found that 24 hours after CLP the serum levels of proinflammatory cytokines (TNF-α, IL-6), peritoneal and kidney vascular permeability, splenic apoptosis and blood bacterial count were significantly reduced in BMSC-treated animals vs controls; the level of IL-10, an anti-inflammatory cytokine, was not affected. Six hours after their injection, fluorescently-labeled BMSCs were detected mostly in the lung. By prelabeling BMSCs with quantum dots and later performing immunostaining with a macrophage marker (Iba-1), we found the BMSCs adjacent to macrophages in the lung. Some were in the spleen, and rare cells were seen in the kidney;24 hours after they were injected, we found few BMSCs in any organ. The positive effect of BMSC treatment was still present in Rag-/- mice;in NK cell depleted mice, and in IFN-γ-/- mice, suggesting that B,T and NK cells, and IFN-γdo not play a significant role in mediating the effects of the BMSCs. On the other hand, macrophage depletion or pretreatment with IL-10 or IL10 receptor antibodies eliminated the beneficial effects of BMSCs in the sepsis model. Our results suggest that BMSCs may act on tissue macrophages resulting in enhanced production of IL-10, a potent anti-inflammatory cytokine.We used a variety of transgenic mice to determine the mechanism of the effect and concluded that Prostaglandin E2 plays a crucial role in the immunomodulation that we observed.We published our findings in Nature Medicine (Vol.15, 1, p42, 2009).
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