The use of mouse models offers a feasible alternative to human observations, when hypothesis-driven studies are needed, but mouse-in-mouse systems do not always reflect the pathology of human diseases. In many aGVHD models, the effector cell is based on infusion of murine splenocytes which may behave differently to human effector cells; furthermore, conventional mice are not well aligned to the study of human cell therapy products. The introduction of the interleukin (IL)-2 receptor gamma mutation onto the non-obese diabetic

(NOD)-severe compromised immunodeficient (SCID) background has allowed for the development ITF2357 clinical trial of refined mouse models. NOD-SCID IL-2rγnull (NSG) mice are deficient for T, B and NK cell activity and allow engraftment of high levels of human peripheral blood mononuclear cells (PBMC) [29]. The NSG model offers an opportunity to examine human donor cells in combination with clinical cell therapeutics. Using a humanized NSG mouse model of aGVHD, this study sought to examine the effect of human MSC cell therapy, and to investigate the possible therapeutic mechanisms involved. Human MSC cell therapy significantly prolonged the survival of

NSG mice with aGVHD, reducing target organ pathology. MSC therapy did not interfere with donor PBMC engraftment or involve the induction of donor T Anti-infection Compound Library ic50 cell apoptosis, anergy or regulatory cell expansion, but rather the direct inhibition of both donor CD4+ T cell proliferation and tumour necrosis factor (TNF)-α production. All procedures involving animals or human material were carried out by licensed personnel according to approved guidelines. Ethical approval for all work was received from the ethics committee of National University of Ireland (NUI) Maynooth. A humanized mouse model of aGVHD was adapted and optimized from a protocol described by Pearson et al. [29]. NOD.Cg-PrkdcscidIL2tmlWjl/Szj mice (NOD-SCID IL-2rγnull) (NSG) (Jackson Laboratories, Bar Harbour, ME, USA) were exposed to a conditioning dose of 2·4 Gray (Gy) of whole-body gamma irradiation. Human PBMC from healthy volunteer donors were isolated by Ficoll-density

centrifugation and administered intravenously (i.v.) to NSG mice (6·3 × 105 g−1) via the tail vein 4 h following irradiation. Negative control mice received a sham infusion of phosphate-buffered saline (PBS) alone. Signs of aGVHD occurred typically between days 12 and 15 post-PBMC transfusion. Carnitine palmitoyltransferase II In some mice, conventional human mesenchymal stem cell (MSC) (4·4 × 104 g−1) therapy was administered on day 7 post-PBMC transfusion. In other groups, interferon (IFN)-γ stimulated MSC (4·4 × 104 g−1) were administered concurrent with PBMC on day 0. The level of human cell chimerism was analysed by flow cytometry (days 4, 8 and 12), examining the expression of CD45+ cells and the ratios between human CD4 and CD8 T cells. aGVHD development was determined by examining features daily including body weight, ruffled fur, locomotor activity, posture and diarrhoea.