S.Seiser¹ , D. Cerbu¹ , A. Gallhofer¹ , J. Matiasek² & A. Elbe‑Bürger^1*

Ewa A. BURIAN¹, Lubna SABAH¹, Klaus KIRKETERP-MØLLER¹, Glenn GUNDERSEN² and Magnus S. ÅGREN^1,3,4

¹Department of Dermatology and Copenhagen Wound Healing Center, Bispebjerg Hospital, University of Copenhagen, Copenhagen, Denmark, ²SoftOx Solutions AS, Oslo, Norway, ³Digestive Disease Center, Bispebjerg Hospital, University of Copenhagen, and ⁴Department of Clinical Medicine, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark

Jana Jandova ^a,b , Jeremy Snell ^a,b , Anh Hua ^a,b , Sally Dickinson ^b , Jocelyn Fimbres ^a,b , Georg T. Wondrak ^a,b,*

^aDepartment of Pharmacology and Toxicology, College of Pharmacy and UA Cancer Center, University of Arizona, Tucson, AZ, USA

^bUA Cancer Center, University of Arizona, Tucson, AZ, USA

ABSTRACT

Hypochlorous acid (HOCl) is the active oxidizing principle underlying drinking water disinfection, also delivered by numerous skin disinfectants and released by standard swimming pool chemicals used on a global scale, a topic of particular relevance in the context of the ongoing COVID-19 pandemic. However, the cutaneous consequences of human exposure to HOCl remain largely unknown, posing a major public health concern. Here, for the first time, we have profiled the HOCl-induced stress response in reconstructed human epidermis and SKH-1 hairless mouse skin. In addition, we have investigated the molecular consequences of solar simulated ultraviolet (UV) radiation and HOCl combinations, a procedure mimicking co-exposure experienced for example by recreational swimmers exposed to both HOCl (pool disinfectant) and UV (solar radiation). First, gene expression elicited by acute topical HOCl exposure was profiled in organotypic human reconstructed epidermis. Next, co-exposure studies (combining topical HOCl and UV) performed in SKH-1 hairless mouse skin revealed that the HOCl-induced cutaneous stress response blocks redox and inflammatory gene expression elicited by subsequent acute UV exposure (Nos2, Ptgs2, Hmox1, Srxn1), a finding consistent with emerging clinical evidence in support of a therapeutic role of topical HOCl formulations for the suppression of inflammatory skin conditions (e.g. atopic dermatitis, psoriasis). Likewise, in AP-1 transgenic SKH-1 luciferase-reporter mice, topical HOCl suppressed UV-induced inflammatory signaling assessed by bioluminescent imaging and gene expression analysis. In the SKH-1 high-risk mouse model of UV-induced human keratinocytic skin cancer, topical HOCl blocked tumorigenic progression and inflammatory gene expression (Ptgs2, Il19, Tlr4), confirmed by immunohisto-chemical analysis including 3-chloro-tyrosine-epitopes. These data illuminate the molecular consequences of HOCl-exposure in cutaneous organotypic and murine models assessing inflammatory gene expression and modulation of UV-induced carcinogenesis. If translatable to human skin these observations provide novel insights on molecular consequences of chlorination stress relevant to environmental exposure and therapeutic.

Keywords:

Pathway-focused gene expression profiling Hypochlorous acid Skin cancer Solar UV radiation Inflammation Oxidative stress

Julian-Dario Rembe^{1 *†} , Lioba Huelsboemer^2† , Isabell Plattfaut² , Manuela Besser² and Ewa K. Stuermer³

¹Department of Vascular and Endovascular Surgery, University Hospital Düsseldorf, Heinrich-Heine-University of Düsseldorf, Düsseldorf, Germany, ²Chair for Translational Wound Research, Centre for Biomedical Education and Research, Witten/Herdecke University, Witten, Germany, ³ Department of Vascular Medicine, University Heart and Vascular Center Hamburg, Translational Wound Research, University Medical Centre Hamburg-Eppendorf, Hamburg, Germany

Biofilms pose a relevant factor for wound healing impairment in chronic wounds. With 78% of all chronic wounds being affected by biofilms, research in this area is of high priority, especially since data for evidence-based selection of appropriate antimicrobials and antiseptics is scarce. Therefore, the objective of this study was to evaluate the anti-biofilm efficacy of commercially available hypochlorous wound irrigation solutions compared to established antimicrobials. Using an innovative complex in-vitro human plasma biofilm model (hpBIOM), quantitative reduction of Pseudomonas aeruginosa, Staphylococcus aureus, and Methicillin-resistant S. aureus (MRSA) biofilms by three hypochlorous irrigation solutions [two <0.08% and one 0.2% sodium hypochlorite (NaClO)] was compared to a 0.04% polyhexanide (PHMB) irrigation solution and 0.1% octenidine-dihydrochloride/phenoxyethanol (OCT/PE). Efficacy was compared to a non-challenged planktonic approach, as well as with increased substance volume over a prolonged exposure (up to 72 h). Qualitative visualization of biofilms was performed by scanning electron microscopy (SEM). Both reference agents (OCT/PE and PHMB) induced significant biofilm reductions within 72 h, whereby high volume OCT/PE even managed complete eradication of P. aeruginosa and MRSA biofilms after 72 h. The tested hypochlorous wound irrigation solutions achieved no relevant penetration and eradication of biofilms despite increased volume and exposure. Only 0.2% NaClO managed a low reduction under prolonged exposure. The results demonstrate that low-dosed hypochlorous wound irrigation solutions are significantly less effective than PHMB-based irrigation solution and OCT/PE, thus unsuitable for biofilm eradication on their own. The used complex hpBIOM thereby mimics the highly challenging clinical wound micro-environment, providing a more profound base for future clinical translation.

Keywords: wound infection, biofilm, antimicrobial, antiseptic, hypochlorous acid, human plasma biofilm model, wound irrigation, chronic wound