Advancing human skin models by integrating skin microbes for next-generation research

The skin barrier comprises interdependent physical, chemical, immunological, and microbial components, of which the latter is constituted by a community of microbes residing on the skin surface that restricts the expansion of opportunistic pathogens, modulates keratinocyte signaling pathways, and fosters immune tolerance. However, molecular and cellular dynamics of host-microbe interactions remain incompletely characterized, partly due to the limited availability of physiologically relevant and robust preclinical models. We aimed to establish 3D human skin equivalents (HSEs) in co-culture with representative skin commensals to investigate host responses across an in vitro cohort of six biological replicates. Well-characterized HSEs were inoculated with Staphylococcus aureus, Staphylococcus epidermidis, and Cutibacterium acnes. A 48-hour co-culture period enabled microbial expansion, during which S. aureus exhibited the most substantial outgrowth, and strain-dependent variability was observed for S. epidermidis. Assessment of epidermal morphogenesis revealed that S. aureus exerted largest structural impact, whereas C. acnes promoted keratinocyte proliferation. Furthermore, S. aureus elicited a pro-inflammatory response, characterized by elevated secretion of IL-8 and CXCL1. In conclusion, we developed a reproducible experimental framework dissecting host-microbe interactions in HSEs to demonstrate that S. aureus induced substantial alterations in epidermal architecture and inflammatory signaling, underscoring its pathogenic potential in cutaneous environments.