Aim: To study the regulation of superantigen in expression and function of CD226 on NK cells.
Methods: Double fluorescent staining and flow cytometry analysis were employed to detect the expression of CD226 on NK cells from human peripheral blood mononuclear cells (PBMC) stimulated by staphylococcus enterotoxin A (SEA) or SEB. (51)Cr release assay was performed to compare the cytotoxicities of NK cells in different activation models. Laser scanning confocal microscope was used to observe the distribution of CD226 on NK cells activated by superantigens at killing stage.
Results: In resting NK cells, the percentage of CD56(+) NK cells and CD56(+) CD226(+) cells were 12.3% and 1.4% respectively, and the cytotoxicity of NK cells against K562 cells was (3.2+/-0.2)% at the ratio of 5:1. After PBMC were stimulated by 0.1 microg/mL SEA and SEB for 1 day, the percentage of CD56(+) NK cells was 13.5% and 14.1% respectively, and the percentage of CD56(+) CD226(+) cells were increased. Interestingly, CD226 was mainly expressed on CD56(dim) NK cells. On day 2 after SEA/SEB stimulation, the proportions of CD56(+) CD226(+) cells among CD56(+) cells were 69.1% and 64.3% in SEA and SEB group respectively. On day 3 after SEA/SEB stimulation, the expression of CD226 on NK cells decreased. Furthermore, the cytotoxicity of NK cells stimulated by SEA or SEB for 3 days against K562 cells were much higher than that of the resting NK cells as well as NK cells cultured without SAg for the same culture time (P<0.05), and reached to the peak at day 2 (82.3%+/-6.9% and 80.6%+/-7.5%, respectively). Additionally, we observed that CD226 molecules were colocalized with LFA-1 at the interface of NK cells which contacted K562 cells.
Conclusion: The cytotoxicity of NK cells enhanced by SEA or SEB may be correlation with the increased expression of CD226 molecule, and CD226 may be involved in synapse formation of NK cells at killing stage.