ALBERT

Description: Chronic graft-versus-host-disease (cGVHD) is a major complication of allogeneic hematopoietic stem cell transplantation (allo-HSCT), with highly variable clinical presentations. The pathophysiology of cGVHD remains to be further understood. The utilization of murine models to study cGVHD has contributed to the understanding of cGVHD, and highlights its mechanistic complexity. Here, we report a new murine cGVHD modle with obvious lung tissue damage in these mice resulted in the development of bronchiolitis obliterans (BO), which is pathopneumonic for cGVHD. Recipients 8 weeks BALB/c (H-2d) female mice, which irradiated with 700cGy dose of linear accelerator, then were injected with bone-marrow cells (1×107) and spleen cells (5×106) from (C57BL/6 x BALB/c) F1 donors (H-2bd haplotype) male mice. The observed items post-transplantation included hematopoietic reconstruction, implant, and general condition. Clinical scores were assessed every 3 days after +14 d. At + 135 d, mice were killed to evaluate the pathological changes of major target organs. Mice in transplantation groups were in hematopoietic reconstruction at +7 d, and all survived to the end point (+135 d). Chromosomes of recipient mice were in full donor chimera. Clinical scores of cGVHD group have been more than 0.6 since +90 d. These mice develop pathologic manifestations in several organs including lung, skin, and liver. Biopsy-proven BO incidence was 100%, and pathological scores of cGVHD group were 5.33±1.55, which significantly higher than those of transplantation control groups (P 〈 0.05). To date, most models involve parent-into-F1 combinations the transfer of MHC-mismatched cells resulted in a phenotype that resembles clinical systemic lupus erythematosus (SLE), termed SLE-cGVHD. However, some of the models do not use any pretransplantation conditioning and no obvious lung tissue damage. We exchanged parent for recipients, and provide a total boby irradiation as pretransplantation conditioning. New MHC-mismatched murine cGVHD model was easily obtained, and more relevancy to the clinical features of cGVHD. This model provides an ideal study model of clinical cGVHD pathogenesis and treatment strategies. NOTES: The project was sponsored by grants from National Natural Science Foundation (No. 81270648 and 30972790), National Public Health Grand Research Foundation (No.201202017), and Guangdong Natural Science Foundation (No. S2012010009560). Disclosures: No relevant conflicts of interest to declare.

Description: Acute graft-versus-host disease (aGVHD) is still an important complication after allogeneic hematopoietic stem cell transplantation. Dendritic cells (DCs) are crucial for initiating cell-mediated adaptive immune responses or maintaining immune tolerance, which play key role on inducting of GVHD. We have reported a subset of dendritic cells (DCs), CD8α+ Jagged2high CD11bhigh regulatory dendritic cells (DCregs), which also express MHC molecules and low lever of costimulatory molecules, induce the production of regulatory T cells and inhibit the proliferation function of lymphocyte. In this study, we examined the role of the CD8α+ Jagged2high CD11bhighregulatory dendritic cells (DCregs) in murine major histocompatibility complex (MHC)-incompatible model of aGVHD in alloHSCT. BALB /c (H-2d) mice bone marrow cells (BMCs) were isolated and were induced to DCs generation by 4 cytokines, SCF (100ng/mL), IL-4 (20ng/mL), Flt3 ligand (25ng/mL), and GM-CSF (5ng/mL) for 3 days. The new DCs were selected by flow cytometry, and were co-cultured with Bone marrow mesenchymal stem cells (BMSCs) for 10 days. Characteristics of such regulatory dendritic cells and immune tolerance features were detected before and after co-culture by FCM analysis, ELISA kit, and mixed lymphocyte reaction. Male 8-week-old C57BL / 6 (H-2b) mice were used as donor, and female 8-week-old BALB / c (H-2d) were used as recipient. The recipient mice received SSD100, 30×30 radiation field, 700cGy total body irradiation (TBI) pretreatment, and grouped as follows: 1.normal control group, 2.TBI control group, 3. transplantat control group, 4. aGVHD group: 1 × 107 BMC and 1 × 107 spleen cells (SPC) were injected through caudal vein 5. DCregs groups: 1 × 107 BMC, 1 × 107 spleen cells (SPC) and 1 × 106DCregs were injected through caudal vein. Evaluate the severity of GVHD according to clinical manifestations (weekly the GVHD ratings), the survival time and histopathological examination. The results showed that novel DCs were transformed into CD8α+Jagged2highCD11bhigh Dcregs. The expression of CD86, CD80, CD40, and MHC-II were evidently decreased, while that of CD205, Jagged1 and Jagged2 were significantly increased on DCregs (P

Description: Abstract 4709 Objective: Acute graft-versus-host disease (aGVHD) confines the wider application of allogeneic bone marrow transplantation (allo-BMT), but recently studies indicate that it is possible to reduce the incidence and severity of aGVHD while preserving the GVT by using bortezomib. In current study, we explored the changes of T cell subsets after allo-BMT administrated with bortezomib immediately, in order to establish the mechanism about bortezomib attenuation aGVHD. Materials and Methods: BALB/c mouse were injected of 0.5 mL PBS containing C57BL/6 2×107 nucleated BM cells plus 1×107 splenocytes followed a single dose of lethal total body irradiation (TBI, 0.7 Gy/min, 8.0 Gy) with or without bortezomib at 1.0 mg/kg. The level of CD4+ CD25+ Foxp3+ regulator T cells is quantified by flow cytometry, and the cytokine level of IL-2 and IL-4 is quantified by ELISA. Results: Bortezomib remarkably reduce aGVHD severity and prolonged the surviving time. Along with bortezomib injection, the level of CD4+ CD25+ Foxp3+ regulator T cell is significantly increased, the cytokine level of IL-2 is decreased but IL-4 is increased. Conclusion: Bortezomib inhibit aGVHD through shifting the combination of T cell subpopulations with up regulation CD4+CD25+ Foxp3+ regulator T cells lead to reset Th1/Th2 cytokine balance. Disclosures: No relevant conflicts of interest to declare.

Description: Abstract 4086 Background: Cutaneous chronic graft versus host disease (cGVHD) is an increasingly common complication in long term survivors of hematopoietic stem cell transplantation, but currently available therapies have demonstrated limited efficacy. Furthermore, Mesenchymal Stromal Cells (MSCs) have been reported to be effective in various immune-mediated disease models, but their therapeutic potentials versus cutaneous cGVHD have not been determined. Methods: We enrolled 23 patients suffering from cutaneous cGVHD who had failed to respond to conventional immunosuppressive therapy or relapsed after reduction of prednisone. They received intravenous in vitro expanded bone marrow (BM)-derived MSCs. The median follow-up period was 90 days (range 60–146 days).The dermal manifestations of cGVHD were monitored, and a score was given to the cutaneous response by the physician. B lymphocyte subsets and ratio of CD4/CD8 were detected by flow cytometry before the first and after the last MSC infusion. The plasmatic levels of TNF-α, sICAM-1 and Th1 or Th2 factors, including IL-2 and IL-4, were measured by enzyme-linked immunosorbent assay (ELISA). Results: Nineteen patients (82.61%) had skin symptom abatement after MSCs treatment with the skin scores improved according to the NIH criteria. Most patients had healing of skin ulceration, regression on skin findings and increased flexibility of involved joint. Steroid sparing or discontinuation of immunosuppressive medications was possible in all of the patients. Clinical improvement was accompanied by increasing levels of CD19+CD5+ B cells and elevated ratio of CD4/CD8. In addition, after the last MSCs treatment, the patients had significantly higher levels of IL-2 compared to before the first MSC infusion, while they had lower levels of TNF-α, sICAM-1 and IL-4. Conclusion: MSCs infusion represents an effective treatment option for cutaneous chronic GVHD. This is of high interest in patients suffering from cutanous cGVHD resistant to conventional immunosuppressive therapy. MSCs alter allo-reactivity by affecting CD19+CD5+ B cells, possibly normalize the ratio of CD4/CD8 and Th1/Th2 and decreased the level of proinflammatory cytokine TNF-α and proadhesive cytokine sICAM-1. Our study demonstrates CD19+CD5+ B cells could be a possible target for therapeutic intervention in cutaneous cGVHD. Disclosures: No relevant conflicts of interest to declare.