In Vitro Analysis of the Dynamic Role of the Bacterial Virulence Factors in Skin Wound Healing

The skin acts as a primary barrier against environmental insults and maintains homeostasis. Injury initiates a wound healing cascade of hemostasis, inflammation, proliferation, and remodeling. In chronic wounds, persistent microbial colonization and inflammation disrupt this process, with bacterial virulence factors acting as key drivers. While the microbiome's role in chronic wounds is recognized, the effects of individual virulence factors on acute repair remain unclear. Therefore, this study investigated the differential effects of virulence factors derived from five skin-associated bacterial species on acute wound healing dynamics. In this context, virulence factors from <i>Pseudomonas aeruginosa</i>, <i>Staphylococcus aureus</i>, <i>Streptococcus pyogenes</i>, <i>Lactobacillus plantarum</i>, and <i>Enterococcus faecalis</i> were tested on HDF-n cell viability and wound closure, with multiplex ELISA used to assess inflammatory mediator secretion and underlying mechanisms. Overall, virulence factors were generally well tolerated across concentrations (0.05-16 µg/mL) and time points (24, 48 h), with cell viability >80%, except for <i>S. aureus</i>, which reduced viability to ~70% at higher concentrations by 48 h. Wound healing responses varied markedly: <i>S. aureus</i> significantly impaired closure in a dose-dependent manner (~10% closure at 16 µg/µL, 48 h), and <i>E. faecalis</i> similarly delayed repair. In contrast, factors from <i>P. aeruginosa</i>, <i>S. pyogenes</i>, and <i>L. plantarum</i> showed neutral or mildly pro-healing effects. Notably, co-treatment with <i>S. pyogenes</i> partially rescued impairment caused by <i>S. aureus</i> and <i>E. faecalis</i>. Collectively, these findings highlight that bacterial virulence factors have variable impacts on acute wound healing. These findings suggest potential therapeutic applications through targeted modulation or combinations of bacterial factors.