Importance of thymus to maintain operational tolerance to fully allogeneic cardiac grafts Masanori Niimi, MD, PhD, Nozomu Shirasugi, MD, Yosifumi Ikeda,

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Importance of thymus to maintain operational tolerance to fully allogeneic cardiac grafts Masanori Niimi, MD, PhD, Nozomu Shirasugi, MD, Yosifumi Ikeda, MD, Shigenao Kan, MD, Hiroshi Takami, MD, Kimikazu Hamano, MD, PhD The Annals of Thoracic Surgery Volume 72, Issue 3, Pages 735-739 (September 2001) DOI: 10.1016/S0003-4975(01)02887-9

Fig 1 Depletion and recovery of CD4+ cells in the spleen after injection of anti-CD4. CBA mice were treated with depleting anti-CD4 monoclonal antibody (YTA3.1) (50 μg/dose) on two consecutive days. CD4+ cells in the spleen were counted by fluorescence activated cell sorter. Percentage of CD4+ cells compared to the original number was presented by mean ± standard deviation. The Annals of Thoracic Surgery 2001 72, 735-739DOI: (10.1016/S0003-4975(01)02887-9)

Fig 2 Operational tolerance was induced after pretreatment with donor-specific blood transfusion (DST) plus anti-CD4. CBA mice were pretreated with anti-CD4 (50 μg/dose) in combination with 0.25 mL of donor-specific blood transfusion 4 weeks before transplantation of a C57BL/10 cardiac graft. The recipients accepted the graft indefinitely in their abdomen. Second C57BL/10 or Balb/c cardiac grafts were transplanted into the neck of the recipients 100 days after the first transplantation. ∗p < 0.001 between the two groups. (MST = median survival time.) The Annals of Thoracic Surgery 2001 72, 735-739DOI: (10.1016/S0003-4975(01)02887-9)

Fig 3 Regulatory cells were generated 100 days after transplantation in the mice pretreated with donor-specific blood transfusion (DST) plus anti-CD4. CBA mice were pretreated with anti-CD4 (50 μg/dose) in combination with 0.25 mL of donor-specific blood transfusion 4 weeks before transplantation of a C57BL/10 cardiac graft. The recipients accepted their graft indefinitely. Splenocytes (5 × 107) from the recipient or naive mice were adoptively transferred into the naive CBA mice. C57BL/10 cardiac grafts were transplanted into the second recipient 1 day after the adoptive transfer. ∗p less than 0.001 between the two groups. (MST = median survival time.) The Annals of Thoracic Surgery 2001 72, 735-739DOI: (10.1016/S0003-4975(01)02887-9)

Fig 4 C57BL/10 splenocytes (1 × 107) were stained with carboxyfluorescein succinimidyl ester (CFSE) before injection and injected intravenously into CBA mice. Spleens and thymus were sampled and fluorescence-positive cells were counted by fluorescence activated cell sorter. Data were presented as the mean of three mouse samples ± standard deviation and are representative of four independent experiments. The Annals of Thoracic Surgery 2001 72, 735-739DOI: (10.1016/S0003-4975(01)02887-9)