Comprehensive Analysis of Zinc Derivatives Pro-proliferative, Anti-Apoptotic and Antimicrobial Effect on Human Fibroblasts and Keratinocytes in a Simulated, Nutrient-deficient Environment In Vitro

Zinc as therapeutic agent in skin and wound care has been known for centuries, but its role is controversial and comprehensive investigations in nutrient-deficient environments are lacking. We aimed to provide a broad analysis of different zinc derivatives on proliferation, apoptosis and antimicrobial properties in a simulated nutrient-deficient environment in vitro. Human fibroblasts (CRL2522) and keratinocytes (HaCaT) were treated with a broad concentration range (10 – 0.0001 µg/mL) of zinc-sulfate (ZnSO(4)), -gluconate (ZnGluc) and -histidine (ZnHis) for 1-6 days under nutrient-deficient media conditions. Cell proliferation was investigated by XTT assay. Targeted analyzes in proliferation (E2F1, PCNA) and apoptosis (TP53) associated genes were performed via qRT-PCR and apoptosis was determined via FACS (annexin V/7-AAD staining). Antimicrobial efficacy was investigated using a quantitative suspension method against S. aureus, P. aeruginosa, E. coli, and C. albicans. The results indicated that 0.1 to 0.001 µg/mL Zn increased cell proliferation in both cell lines. Fibroblasts were more susceptible with significant proliferation peaks on days 2 & 6, and days 1 & 4 for keratinocytes. No relevant changes in gene expression were detected for E2F1 and PCNA nor for TP53. Annexin-V/7-AAD-staining of fibroblasts revealed a small, yet insignificant reduction of apoptosis induction for ZnGluc and ZnSO(4). ZnGluc and ZnSO(4 )(0.1%) achieved high microbial reductions (4-5 log(10) reductions) against tested pathogens. ZnGluc and ZnSO(4) showed relevant pro-proliferative and antimicrobial, as well as tendential anti-apoptotic features in a simulated nutrient-deficient microenvironment in vitro. This further validates a potential benefit of local zinc treatment in deficient wound microenvironments.