Although cell engraftment in a human X-SCID fetus has been achieved by in utero transplantation (IUT), B cell engraftment was limited. We have shown that allogeneic BM cells given via IUT to SCID mice results in high multilineage engraftment. B cell reconstitution was vigorous. High basal immunoglobulin (Ig) levels were detected in reconstituted recipients. While T cell independent B cell function was intact, T cell dependent Ig isotype switching and germinal center (GC) formation was defective. The intrathymic and peripheral negative selection of potentially autoreactive T cells was markedly impaired. We will focus upon several seminal issues in the field. Because B cell engraftment and function are needed to correct SCID, aim 1B will focus upon donor B cell competition using knockout or transgenic (tr) mice with various stages of host B cell developmental arrest later than the pro-B cell defect in SCID. Poor Ig switching and GC formation can be due to defects in T helper dependent B cell activation, intrinsic B cell dysfunction or poor follicular dendritic cell function (aim 1A). Each will be investigated. High basal Ig levels may be a consequence of failed negative selection of autoreactive B cells. Studies are proposed using tr donors and IUT recipients to physically track B cell clonal deletion (aim 1C).
In aim 2 A, intrathymic and peripheral T cell clonal deletion will be tracked using tr donors whose T cell receptor recognizes host alloantigen-bearing cells, a relevant stimulus for modeling negative selection in human IUT recipients. Despite impaired negative selection, recipients had little evidence of GVHD. Peripheral mechanism(s) that may be responsible for downregulating donor anti-host alloreactivity will be sought (aim 2B). Because the thymus is uniquely influenced by donor BM-derived precursors cells in utero, the migration of BM cells into the thymus will be tracked using tr BM cells (aim 2A). A critical issue is whether IUT is advantageous as compared to post- natal BMT for the treatment of SCID. BMT for SCID in humans is performed without or with myeloablation. Experiments are proposed to determine the extent to which immune restoration is facilitated by IUT vs. post-natal BMT in SCID recipients that receive no conditioning therapy or either one of two commonly used conditioning regimens. Data generated by this proposal will provide vital information relevant to the choice of IUT vs. post-natal BMT and will increase our fundamental understanding of factors that influence T- and B-cell development in SCID.
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