Immune Injury by Allogeneic CD4⁺ T Cells Leads to Host Hematopoietic Stem Cell Dormancy and Prevents Engraftment of Donor Cells.

Abstract 2366

The bone marrow (BM) is a complex microsystem that supports lifelong blood production and dynamically responds to injury, blood cell loss and senescence. At steady-state most hematopoietic stem cells (HSC) are quiescent. However, in situations of increased demand, their activation is triggered by an array of signals, such as cytokines released during infections. The susceptibility of BM to injury by T cells (TC) is long recognized, e.g. in aplastic anemia, and BM failure syndromes, and the inhibitory effect of IFNg on the proliferative activity of (HSC) is established. How donor TC given as part of an allogeneic hematopoietic cell transplantation (HCT) affect the recipient BM microenvironment and influence HSC activity has not been elucidated. Yet, graft TC are believed to be essential to overcome host barriers, especially following lower intensity (non-myeloablative) conditioning that spares host HSC and residual host immunity. We examined in an MHC-matched, minor-antigen-mismatched mouse model of non-myeloablative HCT the impact of graft TC subsets on HSC and hematopoietic reconstitution. BALB.K (H2^k) and BALB.B (H2^b) mice were prepared with sublethal radiation at a dose that allows donor cells (AKR/J [H2^k] and C57BL/6 [H2^b]) to engraft but also permits survival of host HSC and immune populations. Grafts consisted of purified HSC (KTLS; cKit⁺Thy1.1^loLin⁻Sca-1⁺) given alone or supplemented with donor TC or TC subsets. Rather than improve engraftment, donor TC, specifically CD4⁺ cells, resulted in detainment of host HSC in their niches and consequent failure of donor HSC to engraft. These events appeared to be initiated specifically by conventional alloreactive donor CD4⁺CD25⁻ cells. These CD4⁺CD25⁻ cells interacted with immune-competent host cells, stimulating CD11c⁺ dendritic cells to upregulate the activation markers CD40 and MHC II. Following their activation dendritic cells produced excessive amounts of IL-12. IL-12, in turn, activated donor CD4⁺ cells to produce IFNg. This Th1-response was prominent in BM but absent in the spleen. In this Th1-cytokine environment, high levels of quiescent host HSC were observed, arrested at the short-term HSC stage, while more mature, multipotent progenitors were absent. Moreover, HSC arrest correlated with marrow aplasia, analogous to that seen in BM failure syndromes. This hypocellularity was only observed in recipients of CD4⁺ containing grafts, while recipients given HSC alone or HSC in combination with CD8⁺ TC had prompt blood regeneration. When resting HSC were FACS-purified from the BM of recipients of HSC+CD4⁺ and infused into secondary recipients they promptly expanded and gave rise to all blood lineages. Although recipients of HSC+CD4⁺ cells ultimately recovered blood formation, B lymphopoiesis was markedly delayed. No clinical signs of graft-versus-host disease were noted in these mice. Lymphocytes that recovered in these recipients of HSC+CD4⁺ cells were of host type only. In contrast, recipients of pure HSC demonstrated prompt donor cell engraftment, and mixed chimerism in all lymphoid lineages, including T cells. Our model corroborates the importance of the cytokine microenvironment in hematopoietic reconstitution and marrow failure post-transplant. Moreover, our findings underscore the importance of IFNg in BM aplasia and support the idea that IFNg can, under physiologic conditions, exert regulatory effects on hematopoiesis by controlling the exit of ST-HSC out of the marrow niche. Our studies shed new light on understanding the physiology of engraftment and marrow failure after HCT using nonmyeloablative conditioning, and indicate that the composition of the graft is of critical importance due to its effects on the marrow microenvironment We postulate that in some naturally occurring settings of BM inflammation (e.g., infections) detainment of resting HSC in their niche may provide protection of hematopoiesis and that comparable events are relevant for engraftment post-HCT.