446 Staphylococcus aureus isolated from patients with atopic dermatitis accumulates in lysosome and induces IL-1a via TLR9 in keratinocytes

It becomes evident that staphylococcus aureus (S.a) is frequently detected in skin of patients with atopic dermatitis (AD), and increased occupancy rate of S.a on skin surface is deeply involved in AD flare. Genotypes of S.a separated from skin of patients with AD seems to be different from S.a separated from a healthy controls, because S.a from AD skin was reported to have a capacity to secrete more toxins. In addition, patients with AD are likely to develop particular types of skin infections by S.a such as impetigo. However, the mechanism of impaired skin defense against to S.a and the aggravation of dermatitis by S.a in patients with AD is still unknown. To reveal the characteristic of S.a from patients with AD and the relationship between AD activity and S.a, we analyzed immunological response against the stimulation by a standard strain S.a and S.a derived from the surface of AD skin (Sa-AD) by using keratinocyte cell line (HaCat cell). We cultured HaCat cells with heat-killed S.a using strains of a S.a standarad strain and Sa-ADs. After the culture, immunostaining shows that S.a-AD are strongly agglutinated inside cytoplasm especially located in lysosome. This phenomenon was not seen in S.a standard strain. To investigate the effect of agglutination of S.a-AD in lysosome, we quantified the cytokine production from S.a stimulated HaCat cell by using Cytometric Bead Array. Compated to S.a standard strain, HaCaT cells stimulated by S.a-AD induced significantly high amount of Interleukin 1 alpha (IL-1α), which was diminished by proteinase K treated S.a-AD with loss of agglutination in lysosome. Moreever, induction of IL-1α was inhibited by Toll-like receptor9 (TLR9) antagonist, inhibitory oligodeoxynucleotide. In summary, our study showed that S.a-AD accmulates in lysosome and induce IL-1α via TLR9 in HaCaT cells.