Background: Nuclear factor κB (NF-κB) overactivation plays a crucial role in T-helper 2 (Th2)-biased allergic airway inflammation by increased activation and decreased apoptosis of CD4+ T cells. We have shown that targeted NF-κB suppression in dendritic cells by adenoviral gene transfer of a novel mutated inhibitor of NF-κB (IκBα) (AdIκBαM) contributes to T-cell tolerance, but the immunosuppressive action of AdIκBαM on memory (CD45RO+) CD4+ T cells remains enigmatic.
Methods: CD45RO+ T cells from Dermatophagoides farinaei-sensitized asthmatic patients, untransfected or transfected with AdIκBαM or AdLacZ (β-galactosidase) for 24 h, were stimulated with anti-CD3 (1.0 μg/ml) plus anti-CD28 (0.5 μg/ml) monoclonal antibody for an additional 24 h. IκBαM transgene expression and NF-κB activation were detected by polymerase chain reaction (PCR), reverse transcription-PCR (RT-PCR), Western blot analysis, and electrophoretic mobility shift assay. Phenotype and apoptosis were measured by flow cytometry, annexin V binding, and terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling analyses. Cytokine production and cell proliferation were determined using enzyme-linked immunosorbent assay and [3H] thymidine incorporation.
Results: A unique 801-bp IκBαM cDNA and a dose-dependent increase in IκBαM transgene expression were observed in AdIκBαM-transfected CD45RO+ T cells. Significantly, AdIκBαM inhibited CD3/CD28-mediated NF-κB activation in CD45RO+ T cells, leading to evident apoptosis, reduction of eotaxin, RANTES, Th1 [interferon (IFN)-γ and interleukin (IL)-2], and Th2 (IL-4, IL-5, and IL-13 despite a slight decrease in IL-10) cytokines and secondary proliferative response. AdIκBαM also upregulated cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) and downregulated CD69 besides no change in CD28.
Conclusion: IκBαM might be beneficial to augment memory CD4+ T-cell tolerance through modulating B7-CD28/CTLA-4 co-stimulatory pathways and NF-κB-dependent cytokine profiles in allergic inflammatory diseases including asthma.