PROJECT SUMMARY/ABSTRACT The proposed Mentored Research Scientist Development Award (K01) will provide the candidate, Dr. Alix Berglund, DVM, PhD, with the necessary knowledge, training, and experience to become an independent translational biomedical researcher in the fields of immunology and mesenchymal stem cell (MSC) biology. MSCs are a promising cell source for treating inflammatory and immune-mediated diseases. Allogeneic therapy would provide cost-effective and efficient treatment, but is currently hindered by recipient immune rejection of donor MSCs expressing mismatched-major histocompatibility complex (MHC) molecules. MSCs are strongly immunomodulatory, however, and manipulation of MHC expression may be sufficient to allow donor MSCs to evade recipient immune responses. The central hypothesis of this proposal, which is supported by strong preliminary research, is that treating MSCs with transforming growth factor-β2 (TGF-β2) downregulates MHC I and MHC II gene transcription thereby reducing the in vivo immunogenicity of MHC-mismatched MSCs. The aims of this project are to 1) Identify how TGF-β2 downregulates MHC expression in MSCs and 2) Determine how TGF-β2 treatment affects MSC immunogenicity in vivo. Both murine and equine MSCs will be utilized to elucidate TGF-β2 signaling pathways to increase translational potential to humans (Aim 1) and non-inflammatory and inflammatory murine models will be used to analyze how the immune system responds to TGF-β2-treated MSCs in vivo (Aim 2). Completion of the proposed research is a first step towards improving the efficacy and safety of allogeneic MSCs for clinical use. To accomplish these aims, Dr. Berglund will build on her background in mesenchymal stem cell biology, immunology, and large animal models by developing expertise in molecular immunology techniques, in vivo alloimmune response analysis, and the utilization of murine models. Dr. Berglund's mentoring team has combined expertise in immunology, mesenchymal stem cells, and genetics to facilitate her transition to an independent translational biomedical researcher. This work will be completed primarily at the North Carolina State University College of Veterinary Medicine, which is eminently qualified to train translational clinician scientists, with additional work at the University of North Carolina-Chapel Hill.

PROJECT NARRATIVE Allogeneic mesenchymal stem cells (MSCs) are a promising cell source for treating inflammatory and immune- mediated diseases, but are targeted for death by the recipient immune system due to mismatched-major histocompatibility complex (MHC) molecules expressed on the cell surface. The aim of this proposal is to analyze how transforming growth factor-β2 (TGF-β2) regulates MHC expression in MSCs and how the recipient immune system responds to TGF-β2-treated MSCs in vivo. Results from the completion of these studies will support the generation of allogeneic donor MSCs that can be used “off-the-shelf” manner for safe and efficacious treatment.