CD8α⁺ Jagged2^high CD11b^high Regulatory Dendritic Cells Prevent Acute Gvhd

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⁺ Jagged2^high CD11b^high regulatory dendritic cells (DC_regs), 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⁺ Jagged2^high CD11b^highregulatory dendritic cells (DCregs) in murine major histocompatibility complex (MHC)-incompatible model of aGVHD in alloHSCT.

BALB /c (H-2^d) 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-2^b) mice were used as donor, and female 8-week-old BALB / c (H-2^d) 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⁺Jagged2^highCD11b^high 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 <0.05). Meanwhile, the percentage of treated DCs cells were increased in the G2 and S phase. Compared with DC control group, IL-6 and IL-12 expression were decreased significantly (P <0.05), IL-10 and TGF-beta expression was significantly elevated (P <0.05). CD8α⁺Jagged2^highCD11b^highDcregs have stronger inhibitory effect of lymphocyte proliferation (P <0.05).The mice of transplant control group were observed to +60d and no GVHD clinical manifestation was observed. The clinical manifestations of aGVHD group is fatigue, piloerection, arched, progressive weight loss, hair removal, repeated skin ulcers, perianal redness and secretions, and all died within +40 d , while the median survival time was 27 days. The median survival of the DCregs group was +33 d, and two group of +30 d survival rates were 20% and 100%, respectively. aGVHD score (P<0.05) and histopathological changes at +33 d of DCregs group were better than the aGHVD group.

CD8α⁺CD11b⁺jagged2^highregulatory dendritic cells can reduce the symptom of GVHD, delay the death of aGVHD model, but the effect duration was short, and might need maintained by infusion repeatedly. The immune tolerance mechanisms of this novel DCregs might be relate to up-regulated Jagged1 and Jagged2 expression and T cell Notch signaling pathway activation.

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).