An Important Role of alpha-Hemolysin in Extracellular Vesicles on the Development of Atopic Dermatitis Induced by Staphylococcus aureus

Abstract

Skin barrier disruption and dermal inflammation are key phenotypes of atopic dermatitis (AD). Staphylococcus aureus secretes extracellular vesicles (EVs), which are involved in AD pathogenesis. Here, we evaluated the role of EVs-associated alpha-hemolysin derived from S. aureus in AD pathogenesis. alpha-hemolysin production from S. aureus was detected using western blot analyses. The cytotoxic activity of alpha-hemolysin on HaCaT keratinocytes was evaluated by measuring cell viability after treating cells with soluble and EVs-associated alpha-hemolysin. To determine the type of cell death, HaCaT keratinocytes were stained with annexin V and 7-AAD. The in vivo effects of alpha-hemolysin were evaluated by application of soluble and EV-associated alpha-hemolysin on the mouse skin. The present study showed that increased alpha-hemolysin was produced by S. aureus colonized on AD patients compared to healthy subjects. alpha-hemolysin production was also related to AD severity. In addition, EV-associated alpha-hemolysin was more cytotoxic to HaCaT keratinocytes than soluble alpha-hemolysin, and alpha-hemolysin-negative EVs did not induce keratinocyte death. EV-associated alpha-hemolysin induced necrosis, but soluble alpha-hemolysin induced apoptosis of keratinocytes. In vivo, skin barrier disruption and epidermal hyperplasia were induced by soluble and EV-associated alpha-hemolysin. However, AD-like dermal inflammation was only caused by EV-associated alpha-hemolysin. Moreover, neither skin barrier disruption nor AD-like skin inflammation was induced by alpha-hemolysin-negative EVs. Taken together, alpha-Hemolysin secreted from S. aureus, particularly the EV-associated form, induces both skin barrier disruption and AD-like skin inflammation, suggesting that EV-associated alpha-hemolysin is a novel diagnostic and therapeutic target for the control of AD.