We are interested in bone marrow stem cells and are trying to find out if and how they can repair tissues, including cells of the immune system itself. Once we learn how these cells can repair tissues or how they can reprogram immune cells we hope to be able to use that knowledge to help patients to heal diseased organs and/or to fight autoimmune diseases. We try to track bone marrow cells by a variety of techniques that include detecting the Y chromosome or the green fluorescent protein in gender mismatched, green BM transplanted mice. We also try to use transgenic mouse models to follow the fate of cells that originate in the bone marrow and express certain stem cell specific markers, such as CD45 - a marker of hematopoetic cells. To study the immune function we use available mouse models that exhibit immune deficiencies, such as the NOD (non-obese diabetic) mice, that also develop a problem in producing saliva - a syndrome that is similar to Sjogren's syndrome in people.? ? We performed a large series of experiments starting 3 years ago using GFP bone marrow transplanted mice that underwent brain surgery (middle cerebral artery occlusion ? MCAO; stroke). Following surgery we used different growth factors injected locally; peripheral bone marrow stimulation; and a combination of the above to study the possible role of circulating bone marrow cells in CNS regeneration after stroke. We performed behaviour tests at different time points; injected growth factors into the stroke site; perfused all (60) mice; saved all organs. We have been sectioning and evaluating the brains following multiple immunostainings and in situ hybridizations ? in correlation with the motor tests. We are currently studying the effect of granulocyte stimulatory factor and stem cell factor (these increase the number of bone marrow derived stem cells in the circulation) in revascularization of the brain after injury.? In collaboration with Denis Faustman and Simon Tran we are analyzing the salivary glands of NOD mice that received healthy gender mismatched bone marrow or gender mismatched CD45- splenocytes. We found that injecting splenocytes along with complete Freund?s adjuvant restores salivary function in NOD mice. However, there are only a few Y chromosome containing salivary cells in these mice suggesting a different mechanism being responsible for the reversal of the autoimmune process. We continue to investigate how this effect occurs.? ? In collaboration with Bob Redman, we are studying whether gender mismatched BM cells injected into the salivary ducts of rats can be incorporated into the gland; if they could function as secretory cells and if their number will increase following injury to the gland. We set up the technique to detect the Y chromosome in the glands and to be able to double label it with salivary cell markers (such as cytokeratin; cadherin; mucin). The rat Y chromosome detection is much more difficult than in the mouse due to the lack of a repeat sequence that is specific to the Y. We can now successfully detect the donor cells and are in the process of studying their differentiation.? ? In collaboration with Andras Bratincsak (left NIH), and Michael Brownstein (NIMH and Venter Institute) we developed a transgenic mouse (CRE-CD45) model that enables us to track cells of the hematopoetic line to study their fate choices. We have now crossed this mouse with an EGFP-Lox mouse and in the resulting offspring, all cells that ever expressed CD45 (considered a hematopoetic cell marker) will recombine the DNA in a way that they will express GFP no matter what tissue they end up in. After the first generation crosses we found that the subsequent generations of the crossed mice develop random recombinations - which was a serious setback. We have now started a new colony and are using the first generation of mice for our experiments.? ? We recently started to study the interaction between bone marrow stromal cells and the immune system. We are using in vivo and in vitro model systems including mouse models of autoimmune and inflammatory diseases (autoimmune gastritis; pemphigus; autoimmune - type1 - diabetes). We are also studying the effect of a variety of cytokines and chemochines on the immunomodulatory effect of BM stromal cells.
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