Toll-Like Receptor 2 and 4 Are Expressed on Transplanted T-Cell Subsets and Influence Outcome after Allogeneic Unrelated Peripheral Blood Stem Cell Transplantation.

Introduction: Recently, Toll-like receptor (TLR) 2 and 4 have been identified as the most important receptors for LPS, which is contained in the cell wall of gram-negative bacteria and is known to be a main inducer of graft versus host disease (GVHD). The role of TLR expressing T-cells within the graft for the induction of GVHD in patients after unrelated peripheral blood stem cell transplantation (PBSCT) is unknown.

Methods and patients: We therefore determined by flow cytometry expression of TLR 2 and 4 on T-cells within the graft of 63 patients receiving unrelated PBSCT after intensive conditioning followed by cyclosporine A and methotrexate as GVHD prophylaxis. Additionally, donor specific single nucleotide polymorphisms (SNP) for TLR2 (R753Q), TLR4 (D299G) and TLR4 (Y135A) were determined. The data were finally correlated with clinical endpoints.

Results: As expected, TLRs were not expressed on T-cells in peripheral blood of healthy donors (TLR 2: <1.0%, TLR4 <0.5% of T-cells, n=6). In contrast we detected a distinct up-regulation of these receptors on T-cells within the grafts. TLR2 and TLR4 expression on CD4⁺ T-cells ranged from 1.2%–12.3% (median 3.1%) for TLR2 and from 1.2%–12.0% (median 3.7%) for TLR4. Among the CD8⁺ T-cells 0.9%–16.1% (median 3.3%) expressed TLR2 and 0.7%–13.3% (median 3.5%) expressed TLR4. The SNP for TLR2 (R753Q) and TLR4 (D299G) was found in 8.6% and 10.3% of the allogeneic donors, respectively but did neither correlate with the expression levels of TLR on T-cells nor with clinical endpoints. Treatment-related mortality from infections was observed in 10 patients (16%). Interestingly, higher expression of TLR2 and 4 on CD4⁺ but not CD8⁺ T-cells was significantly associated with an increased cumulative incidence of fatal infections (38% vs. 8% p=0.03 for TLR2 and 37% vs. 9% p=0.007 for TLR4). Neither the overall CD3⁺, CD4⁺ and CD8⁺ cell dose nor the expression of TLR2 and 4 on CD4⁺ and CD8⁺ T-cells showed a significant association with the incidence of acute or chronic GVHD or relapse.

Conclusion: These data suggest a previously unrecognised up-regulation of TLR, on CD4⁺ and CD8⁺ T-cells contained in G-CSF mobilized apheresis products. Whether these phenotypic changes impact on T cell function or patient outcome warrants further investigation.