Intrapulmonary Immune Regulation by Pre-Transplant Infusion of Recipient-Derived Regulatory T Cells in a Rat Model of Ex Vivo Lung Perfusion Followed by Transplantation

Purpose Regulatory T cells (Tregs) are a potential therapeutic option in lung transplantation (LTx). We have shown that expanded recipient Tregs can be seeded pre-transplant in the donor lung graft using normothermic ex vivo lung perfusion (EVLP). Here, we examined whether this therapy can alleviate lung injury early post-transplant. Methods Study design is shown in Fig A. CD4 + CD25 hi Tregs were isolated from Wistar Kyoto rats (WKy, RT1 l ) and expanded for 7d-14d. Fischer 344 (F344 RT1 lv1 ) heart-lung blocks were flushed and placed on EVLP. Lungs were perfused with Steen TM solution (37°C, 4h). Expanded Tregs (5-120 × 10 6 ) were infused after 1h. Control lungs received no Tregs. The right lung was used for assessment at end-EVLP, and left LTx into WKy recipients was performed after 60min cold preservation. Necropsy was performed on day 3 post-LTx. Results Wet-dry ratio was lower in Treg-treated grafts compared to controls (Fig B). Transcripts for FoxP3, IL-10, CTLA-4 and GITR were elevated in Treg-treated grafts (Fig B). Tregs isolated from the right donor lung at end-EVLP suppressed CD4 + CD25 − T cell proliferation in vitro (Fig C). On day 3 post-LTx, ICAM1 and CD44 (adhesion molecules) expression on both CD4+ and CD8+ T cells were lower in Treg-treated grafts compared to untreated controls (Fig D). Histologically, both Treg-treated and control grafts had minor lung injury and cellular rejection on day 3 post-LTx: acute lung injury scores (4.10 ± 1.66 and 3.88 ± 1.36), ISHLT A grades (0.60 ± 0.516 and 0.625 ± 0.744), B grades (0.400 ± 0.516 and 0.313 ± 0.458). Conclusion Expanded recipient Tregs directly delivered to the lung allograft prior to transplantation exert immune regulatory effects in vivo and in vitro . Further work is needed to optimize this pre-transplant therapeutic approach.