T Cell Repertoire Maturation Induced by Persistent and Latent Viral Infection Is Insufficient to Induce Costimulation Blockade Resistant Organ Allograft Rejection in Mice

Abstract

CD28:CD80/86 pathway costimulation blockade (CoB) with the CD80/86-specific fusion protein CTLA4-Ig prevents T cell-mediated allograft rejection in mice. However, in humans, transplantation with CoB has been hampered by CoB-resistant rejection (CoBRR). CoBRR has been attributed in part to pathogen-driven T cell repertoire maturation and resultant heterologous alloreactive memory. This has been demonstrated experimentally in mice. However, prior murine models have used viral pathogens, CoB regimens, graft types, and/or antigen systems atypically encountered clinically. We therefore sought to explore whether CoBRR would emerge in a model of virus-induced memory differentiation designed to more closely mimic clinical conditions. Specifically, we examined mouse homologs of clinically prevalent viruses including murine polyomavirus, cytomegalovirus, and gammaherpesvirus 68 in the presence of clinically relevant maintenance CoB regimens using a fully MHC-mismatched, vascularized allograft model. Infected mice developed a significant, sustained increase in effector memory T cells consistent with that seen in humans, but neither developed heterologous alloreactivity nor rejected primarily vascularized heterotopic heart transplants at an increased rate compared with uninfected mice. These results indicate that memory acquisition alone is insufficient to provoke CoBRR and suggest that knowledge of prior latent or persistent viral infection may have limited utility in anticipating heterologous CoB-resistant alloimmunity.

Undernutrition and Hypoleptinemia Modulate Alloimmunity and CMV-specific Viral Immunity in Transplantation

“…To measure mCMV-specific cytokine production by CD8 + T cells, a viral peptide stimulation assay was performed as previously described. 21 Briefly, mouse splenocytes were retrieved and incubated with 2 µg/mL of a peptide mixture (JPT Peptide Technologies GmbH, Berlin, Germany) corresponding to the following open reading frames and sequences from mCMV: M38, SSPPMFRV; M45, HGIRMASFI; IE3, RALEYKNL; and M139, TVYGFCLL. After addition of brefeldin A (eBioscience, San Diego, CA) in the presence of Golgi stop for 4 h, cells were washed, stained for interferon-gamma (IFN-γ) and tumor necrosis factor-alpha (TNF-α), and assessed by flow cytometry.…”

“…20 mCMV Infection Infection of C57BL/6 mice with murine cytomegalovirus (mCMV) was accomplished by intraperitoneal (IP) injection of 1 × 10 5 plaque-forming unit of virus suspended in 200 µL PBS. 21 Mice receiving sham infections were injected IP with 200 µL of PBS. Mice were euthanized on either day 5 or day 7 postinfection or post sham infection.…”

“…For the assessment of mCMVspecific T-cell cytokine production, cells were stained for surface CD3, CD4, and CD8, fixed using a BD Cytofix Fixation/Permeabilization Buffer Set (BD Biosciences), and stained for intracellular IFN-γ (BioLegend; clone XMG1.2) and TNF-α (BioLegend; clone MP6XT22). 21 Labeling of cells was performed at 4°C and was assessed using an LSR Fortesa flow cytometer (BD Biosciences). 21 The following gating strategy was used: T cells were selected on the basis of CD3 staining, and those cells were distinguished on the basis of CD4 and CD8 T cells.…”

“…21 Labeling of cells was performed at 4°C and was assessed using an LSR Fortesa flow cytometer (BD Biosciences). 21 The following gating strategy was used: T cells were selected on the basis of CD3 staining, and those cells were distinguished on the basis of CD4 and CD8 T cells. Memory T-cell subsets were defined by their expression of CD44 and CD62L: naive T cells were defined as CD44 -CD62L + , T CM cells as CD44 + CD62L + , and T Eff cells as CD44 + CD62L -.…”

Optimal Immunosuppression Strategy in the Sensitized Kidney Transplant Recipient

“…However, the risk of acute rejection, graft loss, and death were comparable [93,94] and despite the similar risk of graft and patient survival and less nephrotoxicity, widespread adoption of belatacept amongst all transplant recipients has been limited by concerns regarding higher rates of acute rejection and PTLD [95][96][97]. Subsequent to this, the understanding of the mechanisms of costimulation blockade resistant rejection (CoBRR) has improved [98][99][100][101]. Additionally, there is developing clinical evidence of significant dampening effects on humoral responses induced by costimulation blockade [90,91,102].…”