Figure 1.

Ex vivo or in vivo depletion of CD45RC^highT cells associated with a suboptimal dose of rapamycin induces long-term survival in a model of aGVHD in F1 Lew/BN rats. (A) Schematic of the protocol: 8-week-old F1 male Lew/BN (RT1^l/n) rats, obtained following crossing of Lew (RT1^l) female and BN (RT1ⁿ) male rats, were irradiated 4.5 Gy 1 day before being transferred with total T cells or CD45RC^high-depleted T cells enriched from Lew splenocytes. (B) Splenocytes were stained for CD45RC with an anti-CD45RC mAb (OX32) (y-axis) before sort and after depletion of CD45RC^high cells and cell sorting using the anti-CD45RC mAb OX22 clone binding a distinct epitope than OX32. (C) F1 rats received either 20 × 10⁶ F1 or Lew T cells or 15 × 10⁶ Lew T cells. Results are expressed in percentage of weight loss for each recipient and percentage of survival. Two-way repeated measures ANOVA and log rank (Mantel Cox) tests. *P < .05; ***P < .001. Two independent experiments. (D) Schematic of the protocol: Lew or Lew/BN F1 T cells enriched from splenocytes were transferred (20 × 10⁶) into irradiated (d-1) F1 rats. Recipients were treated with anti-rat CD45RC mAbs (n = 6-8) or isotype control mAbs (n = 6-8) every 2.5 days for 30 days in association with suboptimal doses (0.4 or 0.8 mg/kg) of rapamycin each day for 10 days. Results are expressed in mean percentage of weight loss (E) and survival (F). Four independent experiments. (G) Animals were assessed at different times after injection for a clinical score of aGVHD (see supplemental Table 1) and euthanized at score 10. Two independent experiments. (H) Histological analysis of small intestine (ileum) and scoring of lesions was performed at day 28 in rats treated with isotype control or anti-CD45RC mAb and rapamycin or in long survivors rats at day 150 (original magnification ×5 and ×20). The black squares indicate the zone of ×20 magnification. Results are expressed in mean ± SEM. **P < .01; ***P < .001. depl., depleted; FSC, forward scatter; p.o., per os (orally).