[Background] The generation of high affinity antibody and B cell memory requires the interaction of antigen-specific B and T cells in lymphoid follicles and the formation of germinal centers (GC) which is tightly regulated by follicular helper (Tfh) T cells. While increasing evidence has implicated humoral responses in the immunopathogenesis of chronic rejection, the relevance and role of Tfh cells remains under studied. Here, we tested the effect of belatacept alone on desensitization.

[Methods] Female rhesus macaques received skin transplantation (n=4). Grafts were all rejected within 2 weeks without treatment. As of POD 106, half of recipients were treated with belatacept (20mg/kg weekly × 8 weeks). Tfh cells (PD-1hiICOShiCD4+), Treg (CD4⁺CD39⁺FoxP3⁺) and clonal expansion of B cells (Ki67⁺CD20⁺IgG⁺) were measured in draining lymph nodes. Plasma cells (CD38⁺CD138⁺CD20^loCD19⁺CD27⁺) in the bone marrow and serum DSA were measured upon completion of Belatacept treatment.

[Results] Unlike the huge impact on Tfh cell population seen with belatacept given with T cell depletion, belatacept alone does not completely suppress the Tfh cell (PD-1⁺ICOS⁺) population in the lymph node but shows a trend of reduction (as % of CD4 cell; 27.2±8.06 vs. 17.4±4.38%) compared to date-matched sensitized/untreated rhesus macaques. No obvious reduction of IgG⁺CD20⁺ proliferation was seen (as % of IgG⁺CD20⁺; 3.32±1.54 vs. 4.33±4.0). Unexpectedly, the Treg (as % of total CD4; CD39⁺FoxP3⁺) cell population showed strong trend of reduction after belatacept treatment (as % of CD4 cell; 0.57±0.11 vs. 0.12±0.06%). Unfortunately, serum DSA level was not reduced (% MFI reduction, 18.76±6.81 vs. 24.05±16.66%) and similar level of plasma cells (CD38⁺CD138⁺) was found in the bone marrow (as % of live CD20^loCD19⁺ cell; 1.545±0.47 vs. 1.06 ± 0.01%) compared to untreated control. Finally, we confirmed that the germinal center reaction measured in situ (CD20⁺PD-1⁺CD4⁺) was only moderately reduced with pre-emptive belatacept alone treatment compared to healthy control (area of PD-1⁺CD4⁺; 357.34 +292.49 vs. 555.57+322.98 mm²; p=0.53).

[Conclusions] Costimulation through CD28 on T cells with CD80/86 on B cells are known to contribute to upregulation of the transcription factor Bcl-6, a signature transcription factors for the Tfh cell development. However, targeting CD28 signal via Belatacept (CTLA4-Ig) does not affect Tfh cell population and germinal center response in sensitized macaques.

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