Survival and Function of MiHA Epitope-Specific Host CD8 TM Cells Following Ablative Conditioning and HCT Alwi M. Shatry, Derry C. Roopenian, Robert B.

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Survival and Function of MiHA Epitope-Specific Host CD8 TM Cells Following Ablative Conditioning and HCT Alwi M. Shatry, Derry C. Roopenian, Robert B. Levy Biology of Blood and Marrow Transplantation Volume 13, Issue 3, Pages 293-298 (March 2007) DOI: 10.1016/j.bbmt.2006.12.440 Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 1 Recipients containing CD8 TM specific for the single MiHA epitope resist MHC matched allogeneic marrow engraftment. (A) Generation of anti-H60 CD8 TM. B6 mice sensitized 2× with B6 marrow-derived DC pulsed with H60 were analyzed for the presence of circulating CD8+H60TCR+ T cells 23 days following a booster injection. Gated CD8+ H60TCR+ cells were analyzed for the expression of the TM markers CD44 and Ly 6C (A, histograms). (B). One week post-HCT, peripheral donor chimerism (Ly 5.2, gfp) was analyzed in DC-H60 sensitized B6 (CD45.1) mice ablatively conditioned (9.0 Gy) and transplanted with B6 H60 congenic 5 × 106 TCD-BM. Biology of Blood and Marrow Transplantation 2007 13, 293-298DOI: (10.1016/j.bbmt.2006.12.440) Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 2 Resistance by recipients with CD8 TM specific for a single epitope against MiHA-mismatched marrow allografts. (A) B6 naïve mice or mice containing CD8+H60+ TM (B6DCH60) were transplanted with 5 × 106 BALB.B BMC. Peripheral blood donor chimerism (Ly 9.1) was analyzed at 1 week post-HCT. (B) Recipient spleen cells were assayed for the presence of committed progenitors (CFU-IL3) 1 week post-HCT (2 independent experiments). Biology of Blood and Marrow Transplantation 2007 13, 293-298DOI: (10.1016/j.bbmt.2006.12.440) Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 3 Resistance to MHC-matched allogeneic marrow in DC-H60 sensitized B6 mice is evident 2 weeks post-HCT. Naïve or B6 mice sensitized with marrow-derived DC pulsed with the H60 peptide were irradiated and transplanted as described in Figure 2. Peripheral blood donor chimerism (Ly9.1+ or gfp+) was analyzed 2 weeks post-HCT. Numbers represent % of total cells gated by forward and side angle scatter from pooled mice (n = 3-4/group). Biology of Blood and Marrow Transplantation 2007 13, 293-298DOI: (10.1016/j.bbmt.2006.12.440) Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 4 CD8+H60TCR+ cell presence following ablative TBI conditioning and synerneic BMT. Naïve (A) or B6 mice sensitized (B) with BALB.B × 106 syngeneic B6 Ly 5.1 BM-TCD. Fifteen days post-BMT, peripheral blood was analyzed for recipient (Ly 5.2) CD8+H60TCR+ cells. Ly 5.2+ cells were gated (inset) and analyzed for the presence of CD8 and H60 staining in naïve (A) or primed recipients (B). Numbers represent % CD8+H60TCR+ in gated cells. Biology of Blood and Marrow Transplantation 2007 13, 293-298DOI: (10.1016/j.bbmt.2006.12.440) Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 5 Memory cells present in sensitized recipients following ablative conditioning and HCT are capable of mediating effector repsonses. Sensitized or unsensitized B6 mice were first lethally (9.0 Gy) irradiated, and 24 hours later transplanted with 5 × 106 syngeneic TCD-BM. Twelve days following this first HCT, the mice were irradiated with 5.5 Gy and transplanted 1 day later with 1 × 107 BALB.B or B6-gfp TCD-BM. Ten days following this second transplant, mice were sacrificed and splenic CFU-IL3 analyzed. Legend represents donor-recipient combinations in 2nd transplant. Biology of Blood and Marrow Transplantation 2007 13, 293-298DOI: (10.1016/j.bbmt.2006.12.440) Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions