Generation and functional assessment of nonhuman primate regulatory dendritic cells and their therapeutic efficacy in renal transplantation

Abstract

Nonhuman primates (NHP) are important pre-clinical models for evaluation of the safety and efficacy of the most promising potential therapeutic advances in organ transplantation based on rodent studies. Although rare, dendritic cells (DC) play important roles in preservation of self tolerance and DC with immunoregulatory properties (regulatory DC; DCreg) can promote transplant tolerance in rodents when adoptively transferred to allograft recipients. NHP DCreg can be generated ex vivo from bone marrow precursors or blood monocytes of cynomolgus or rhesus macaques or baboons. NHP DCreg generated in the presence of anti-inflammatory factors that confer stability and resistance to maturation, subvert alloreactive T cell responses. When infused into rhesus renal allograft recipients before transplant, they safely prolong MHC mis-matched graft survival, associated with attenuation of anti-donor immune reactivity. In this concise review we describe the properties of NHP DCreg and discuss their influence on T cell responses, alloimmunity and organ transplant survival.

Keywords: Dendritic cells; Immune regulation; Nonhuman primates; Tolerance; Transplantation.

Fig. 1.

Phenotypic characteristics of control DC and DCreg propagated from rhesus macaque CD14⁺ blood monocytes (mobilized in leukapheresis products). DC were propagated in GM-CSF + IL-4 from immunobead-isolated CD14⁺ monocytes, in the absence (control DC) or presence of VitD3/IL-10 (DCreg). Phenotypic data obtained using flow cytometry are shown for unstimulated control DC and DCreg, and also for both populations stimulated for 24 h with a potent, pro-inflammatory cytokine cocktail (“cytokine stimulation”). Note the resistance of DCreg to phenotypic maturation (upregulation of MHC class II [HLA-DR], CD80, CD86 and CD83) compared with control DC, and the comparatively high ratios of programed death-ligand 1 (PD-L1):CD80 and PD-L1:CD86 expressed by DCreg. MFI=mean fluorescence intensity. Reproduced with permission from The American Journal of Transplantation (Ezzelarab MB et al, Am J Transplant, 2013;13:1989–2005; Fig. 1A, page 1992).

Fig. 2.

Influence of donor-derived, or donor alloAg-pulsed, recipient-derived DCreg on life-sustaining MHC-mismatched renal allograft survival in rhesus macaques. Donor-derived DCreg were infused 1 week before transplant or donor alloAg-pulsed recipient-derived DCreg infused 1 day before transplant, in combination with a minimal immunosuppressive regimen comprising co-stimulation blockade (CTLA4Ig) and rapamycin. All immunosuppressive therapy was stopped on day 180 post-transplant. Actuarial graft survival data using Kaplan-Meier analysis. Further details of the experimental procedures are provided in [42] and [48].

Fig. 3.

Donor-derived DCreg infusion enhances concomitant cytotoxic T lymphocyte Ag 4 (CTLA4) and programed cell death protein-1 (PD1) expression on donor-reactive CD4⁺ and CD8⁺ Tmem in renal-allograft recipient monkeys. Recipient PBMC were obtained 28 days after transplant and cocultured with either donor or 3^rd party stimulators for 5 days. Ex-vivo-stimulated CD95⁺ Tmem in recipients with DCreg infusion exhibited significantly higher donor-reactive CTLA4⁺PD1⁺ cells after transplant, compared to control group recipients (no cell infusion). Graph bars represent means + SD. Data are from 4 graft recipients in each group.