Immunosuppressive Effects of Multipotent Mesenchymal Stromal Cells on Graft-Versus-Host Disease in Rats Following Allogeneic Bone Marrow Transplantation.

Abstract

The major obstacle to successful bone marrow transplantation (BMT) is graft versus host disease (GVHD). Multipotent mesenchymal stromal cells (MSC) have shown immunosupressive effects. The aim of this study was to investigate the effect of MSC on the prevention of GVHD in a rat BMT model. We prepared rat GVHD model using allogeneic BMT+4x108 donor splenocyte infusion (DSI). Allogeneic BMT were performed from Spraque Dawley (SD) rats to female Wistar rats by applying 1x108 mononucleer cells (MNC) via tail vein. The recipient animals were conditioned with a myeloablative regimen consisted of 950 cGy total body irradiation prior to BMT. GVHD prophylactic regimen was performed 3 mg/kg/day cyclosporine-A (CsA)+0.25 mg/kg methotrexate (Mtx) IP +1., +3., +6. days. The rats were divided into three control group (CG) and four study group (SG) (n=7). CG-I; only allogeneic BMT+DSI (GVHD model), CG-II; only myeloablative regimen, CG-III; only inspection. GVHD model was performed in all SGs and then SG-I receiving only GVHD prophylactic regimen, SG-II receiving only MSC, SG-III receiving GVHD prophylactic regimen+MSC, SG-IV receiving MSC after GVHD findings developed. Gross (diarrhea, lose weighting, skin findings), histopathological, and immunophenotypical (CD4, CD8, CD25 and plasma IL-2 levels) examinations of GVHD following allogeneic BMT, and the survival analysis of all groups were performed. The survival following allogeneic BMT was significant longer (p<0.05) in SG-I, SG-II and SG-IV than in CG-I (only GVHD model). However, the survival of SG-III (both GVHD prophylactic regimen and MSC administration) was not significant longer than in CG-I (only GVHD model). Gross and histopathological findings of GVHD was significant lower (p<0.05) in SG-I and SG-II than in CG-I (only GVHD model). The expression of CD25 was showed significant increase (p<0.001) in all SGs and CG-I (only GVHD model) than in CG-III (healthy control). Plasma IL-2 levels were showed significant increase in SG-I (p=0.032), SG-II (p=0.018) and SG-IV (p=0.032) than in CG-I (healthy control). This increase was very prominent (p<0.001) in CG-I (only GVHD model) than in CG-III (healthy control). The ratio of CD4/CD8 expression was showed significant lower in SG-I (P=0.008) than in CG-III (healthy control). In contrast, this expression was showed significant increase in SG-II and SG-IV (p=0.014 for both) than in CG-III (healthy control). Finally, clinical use of MSC in both prophylactic and after GVHD developed was as effective as GVHD prophylactic regimen in preventing GVHD. However, MSC combined with GVHD prophylactic regimen was caused strong immunosuppression, and therefore this combination may results in early mortality. This study may form the basis for use of MSC in the prophylaxis and the treatment of GVHD. Our results are encouraging in the use of MSC in GVHD further to clinical trials.