The Antiseptic Octenidine Inhibits Langerhans Cell Activation and Modulates Cytokine Expression upon Superficial Wounding with Tape Stripping

Nikolić, Nenad; Kienzl, Philip; Tajpara, Pooja; Vierhapper, Martin; Matiasek, Johannes; Elbe-Bürger, Adelheid

doi:https://doi.org/10.1155/2019/5143635

Journal of Immunology Research

On this page

Abstract Introduction Materials and Methods Results Discussion Data Availability Disclosure Conflicts of Interest Acknowledgments References Copyright Related Articles

Research Article | Open Access

Volume 2019 | Article ID 5143635 | https://doi.org/10.1155/2019/5143635

The Antiseptic Octenidine Inhibits Langerhans Cell Activation and Modulates Cytokine Expression upon Superficial Wounding with Tape Stripping

Nenad Nikolić,¹Philip Kienzl,¹Pooja Tajpara,¹Martin Vierhapper,²Johannes Matiasek,³and Adelheid Elbe-Bürger¹

Academic Editor: Xiao-Feng Yang

Received31 Jul 2018

Revised03 Dec 2018

Accepted14 Jan 2019

Published03 Mar 2019

Abstract

Ideal agents for the topical treatment of skin wounds should have antimicrobial efficacy without negative influence on wound healing. Octenidine (OCT) has become a widely used antiseptic in professional wound care, but its influence on several components of the wound healing process remains unclear. In the present study, we have used a superficial wound model using tape stripping on human full-thickness skin ex vivo to investigate the influence of OCT on epidermal Langerhans cells (LCs) and cytokine secretion pattern of skin cells during wound healing in a model without disruption of the normal skin structure. Histological and immunofluorescence studies showed that OCT neither altered human skin architecture nor the viability of skin cells upon 48 hours of culture in unwounded or wounded skin. The epidermis of explants and LCs remained morphologically intact throughout the whole culture period upon OCT treatment. OCT inhibited the upregulation of the maturation marker CD83 on LCs and prevented their emigration in wounded skin. Furthermore, OCT reduced both pro- and anti-inflammatory mediators (IL-8, IL-33, and IL-10), while angiogenesis and growth factor mediators (VEGF and TGF-β1) remained unchanged in skin explant cultures. Our data provide novel insights into the host response to OCT in the biologically relevant environment of viable human (wounded) skin.

1. Introduction

The skin is one of the body’s largest interfaces and is exposed to the outer environment, functioning as a physical barrier to protect against the invasion of pathogens. In addition to mechanical defense, T cells and immature CD1a⁺CD207⁺ Langerhans cells (LCs) reside in the epidermis and participate in immunosurveillance. LCs are a specialized subset of dendritic cells (DCs) that play an essential role in sensing pathogenic microorganisms and tissue damage to initiate immune responses and maintain skin homeostasis [1–5].

In inflamed or injured skin, inflammatory signals produced by various cell types can promote LC activation and migration towards regional lymph nodes, where they elicit primary immune responses. During migration, LCs mature and upregulate the maturation marker CD83 and other molecules associated with antigen presentation [6]. LCs also play a crucial immunologic role in wound healing. Upon acute wounding, it is most important that the skin barrier function is restored as fast as possible. This is achieved by a complex wound healing process which involves four sequentially overlapping phases: hemostasis, inflammation, proliferation, and remodeling, resulting in the formation of a scar [7–10]. This well-coordinated sequence of events is regulated by a variety of cells. Immediately following wounding, DCs [11, 12], keratinocytes [13, 14], T cells [15], and mast cells [16] are activated, subsequently releasing signaling molecules to recruit other immune cells [17]. In particular, LCs represent an important immune cellular component during the initial stages of acute wound healing [18–20] and higher numbers of LCs have been shown in healing compared to nonhealing diabetic foot ulcers implying their involvement in chronic wounds as well [20]. While our understanding of LC involvement during wound healing has somewhat advanced in recent years, little is known about the influence of topically applied antiseptics on LC physiology in wound healing. In that context, a widely used molecule in modern wound care is octenidine (OCT). Compared to other antiseptics (e.g., chlorhexidine, polyhexanide, and PVP-iodine), OCT is highly effective within a short contact time at low concentrations, with a broad spectrum of antimicrobial activity against (even multidrug resistant) bacteria [21–24]. Furthermore, OCT is stable at pH 1.6-12.2, which is crucial in wound care due to pH change in the wounding process [25]. In addition, it is the only antiseptic which remains active locally for up to 48 hours and is not associated with systemic side effects [26, 27]. OCT is widely used in Europe for skin, mucous membrane, and wound antisepsis [25, 28] as well as for patient decolonization in various clinical settings [29–31], and resistances have not been reported [32]. Even though very few cases of irritant and/or allergic contact dermatitis have been reported when superficial skin infections were treated with OCT [33, 34], it is generally considered to be a safe and efficient antiseptic [25].

Results from animal studies [35–37] and clinical data [26, 38–41] have shown that besides its high antimicrobial effects, OCT may also have a positive influence on wound healing, including significant improved scar quality after abdominoplasty [42]. The treatment of chronic venous leg ulcers with OCT was associated with higher healing rates [39] and appears to have positive influence in skin transplantation in patients with impaired wound healing [38].

As animal wound repair can never be a direct and true reflection of human wound healing and its clinical challenges, it is essential to use human skin as the basis of a wound healing model because the pathology and physiology of healing is identical to that found in patients. Furthermore, there are increasing restrictions in Europe for using animals for testing properties of compounds and creams. Therefore, in line with the 3Rs (reduction, refinement, and replacement of animal models), we have used a superficial wound model using tape stripping on human full-thickness skin ex vivo as it is the simplest partial thickness injury of the skin involving removal of the stratum corneum leaving the epidermal compartment generally intact, thus allowing to study the effects of OCT on epidermal LCs which play an essential immunologic role during wound healing. Further, each phase of wound healing can be characterized by the secretion of cytokines, chemokines, and growth factors which were also analyzed.

2. Materials and Methods

2.1. Skin Specimens and Treatment/Culture Procedures

Skin was collected from anonymous healthy participants (aged 20-55 years) undergoing routinely performed body contouring surgeries and processed within 1-3 hours. No morphological or histological pathologies of the skin were observed. The study was approved by the ethics committee of the Medical University of Vienna and conducted according to the Declaration of Helsinki principles. Written informed consent from the participants was obtained.

To generate a superficial wound, the stratum corneum was removed using a standardized tape-stripping method as reported previously by our group [43]. For this, D102-squame standard self-adhesive discs (CuDerm Corporation, USA) were applied with a constant pressure for 10 seconds. Fifty consecutive tape strips were made on the identical spot by the same performer to reduce variability. The efficient removal of the stratum corneum was tested by immunohistochemical staining of punch biopsies (mm) taken from wounded (=tape stripped) skin. In parallel, skin biopsies were cultured at the air-liquid interphase in triplicates per group in DMEM complete medium (supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin (Gibco, Austria)) for 24 to 48 hours without treatment and application of 50 μl control gel (Normlgel®, 0.9% w/w sodium chloride, Mölnlycke Health Care, Sweden) or 50 μl OCT (octenilin® wound gel, 0.05% OCT, Schülke & Mayr GmbH, Germany) onto the epidermal side of each biopsy. Supernatants were collected at selected time points and frozen at -80°C for further analysis.

2.2. Histology and Epidermal Sheet Preparation

From skin explants, one-third was embedded in the optimal cutting temperature compound (Tissue-plus, Scigen Scientific Inc., USA), snap frozen in liquid nitrogen, and stored at -80°C until further processing and another third was fixed in 7.5% formaldehyde overnight and embedded in paraffin. Five μm sections were cut and stained with hematoxylin and eosin to assess tissue morphology. From the last third of the biopsies, skin was incubated on 3.8% ammonium thiocyanate solution for 1 hour at 37°C (Carl Roth GmbH + Co. KG, Germany). Subsequently, the epidermis was separated from the underlying dermis, washed twice for 5 minutes with PBS, fixed with ice-cold acetone (Merck, USA) for 10 minutes, and stored at -80°C until further processing.

2.3. Skin Cell Apoptosis

Skin cryosections (5 μm) were stained with an activation-specific anti-caspase 3 polyclonal rabbit antibody (Cell Signaling, USA) and visualized with Alexa Fluor 546 goat anti-rabbit. As a positive control, normal human skin was exposed to UVB (280–320 nm) and similarly analyzed [44].

2.4. Immunofluorescence

Staining of acetone-fixed epidermal sheets was performed with the following primary antibodies: CD1a (mIgG2b; BioLegend, USA), CD83 (BD Biosciences, USA), CD207 (Sigma-Aldrich, USA), and HLA-DR-Alexa88 (mIgG2; BioLegend, USA). Primary antibodies were incubated overnight at 4°C and respective isotype controls were performed. After a washing step, species- and isotype-specific secondary Abs goat anti-mouse Alexa Fluor488 and goat anti-rabbit Alexa Fluor546 (both Life Technologies, USA) were added when needed and incubated for 1 hour at room temperature. Sheets were mounted with 20 μl mounting media containing DAPI (Vector Laboratories Inc., USA). Images were taken using the AX70 microscope with the imaging software MetaMorph version 7.8.6.0 (Olympus, Germany).

2.5. Immunohistochemistry

Acetone-fixed epidermal sheets were washed for 5 minutes with wash buffer followed by blocking of endogenous peroxidase activity for 5 minutes in methanol containing 0.03% hydrogen peroxide. Subsequently, sheets were incubated with an antibody directed against CD207 (Leica Biosystems, Germany) for 1 hour at room temperature, followed by a secondary antibody for 20 minutes at room temperature. Sheets were then incubated in streptavidin peroxidase solution for 20 minutes, and staining was visualized with amino-ethyl-carbazole (all Dako). Finally, sheets were mounted with Aquatex (Merck Millipore, USA) and examined.

2.6. Quantification of Cells in Skin Sections and Epidermal Sheets

Immunoreactive cells were counted in 6 images/section (Figure 1(c)) or epidermal sheets (Figures 2(b) and 2(d)) from a total of 6 different sections or epidermal sheets/donor from all experimental groups in 6 (Figure 1) or 7 (Figures 2(b) and 2(d)) different donors using ImageJ (1.51j, Wayne Rasband, National Institutes of Health, USA). Data are presented as mean±standard deviation (SD) of all measurements.

(a)

(b)

(c)

(a)

(b)

(c)

(d)

(e)

(f)

2.7. ELISA

96-well plates were coated with the appropriate capture antibodies: IL-8 (M801; Thermo Fisher Scientific, USA) and IL-10 (BioLegend, USA) overnight at 4°C and IL-33, VEGF, TGF-β1 (all R&D, USA) overnight at room temperature. On the next day, plates were washed with wash buffer (PBS-Tween 0.05%), incubated with blocking buffer (IL-8: 4% BSA in PBS-Tween 0.05%) or reagent diluent (IL-33, VEGF: 1% BSA in PBS) or block buffer (TGF-β1: 5% Tween-PBS) or assay diluent (IL-10) for 1 hour at room temperature. Standards and samples were applied to plates and incubated for either 1 hour (IL-8) or 2 hours (IL-33, VEGF, TGF-β1, and IL-10). The activation of latent TGF-β1 in supernatants was assessed by adding 1 N HCl for 10 minutes and stopped with 1.2 N NaOH/0.5 M HEPES. Next, the respective detection antibodies were incubated for either 1 hour (IL-8 and IL-10) or 2 hours (IL-33, VEGF, and TGF-β1) and subsequently incubated with a streptavidin-HRP for 20-30 minutes. TMB substrate solution (Thermo Fisher Scientific) was added and incubated for 20 minutes (IL-8, IL-33, VEGF and TGF-β1) or 30 minutes (IL-10) in dark at room temperature. After adding 0.18 M H₂SO₄ (IL-8) and 2 N H₂SO₄ (IL-33, VEGF, TGF-β1, and IL-10) to stop the reaction, the optical densities at 450 nm were measured using Multiskan™ FC Microplate Photometer (Thermo Fisher Scientific).

2.8. Statistical Analysis

Data was analyzed using GraphPad Prism 5 (GraphPad Software, USA). Unpaired -test was used for comparing means. The results were considered significant with values smaller than 0.05.

3. Results

3.1. OCT Neither Alters Skin Anatomy Nor Enhances Apoptosis in Skin Cells upon Wounding

To test whether the removal of the stratum corneum may influence the penetration capacity of topically applied OCT and consequently affect morphological and behavioral changes of skin cells when compared with unwounded OCT-treated skin, a human full-thickness skin ex vivo culture model was employed. We comparatively assessed unwounded with wounded human skin explants after culture without or with topical application of OCT or control gel. Compared with unwounded human skin, OCT did not cause obvious changes in the skin structure in wounded skin within 48 hours (Figure 1(a)). Thus, OCT does not alter the human skin architecture and preserves the structure of the epidermis and dermis. Next, we analyzed whether OCT induces apoptosis of skin cells in cultured wounded skin. Similar to untreated normal skin, we found no caspase 3⁺ cells in wounded skin before culture (data not shown), whereas caspase 3 activation was generally detected in some epidermal cells of all three groups and was most pronounced upon application of the control gel (Figure 1(b)). Quantitative analysis revealed significantly higher numbers of caspase 3⁺ cells in cryosections derived from control gel treatment compared to OCT and untreated groups when assessed 48 hours upon culture (Figure 1(c)). These observations suggest an increased apoptosis due to the wounding procedure itself and subsequent application of control gel rather than OCT treatment.

3.2. OCT Preserves LC Morphology and Prevents their Emigration and Maturation upon Wounding

Once human skin is excised and subsequently cultured, epidermal LCs get activated and start to emigrate from the epidermis [6]. This nonantigen-mediated reduction of the LC density in the epidermis is visible in normal human skin specimens upon culture and was used as a baseline against which LC changes (frequency and mobilization) in skin samples in response to topical application of OCT or control gel. Analysis of freshly isolated, unwounded, and untreated epidermal sheets before culture that were stained with an antibody directed against CD207 revealed a network of highly dendritic LCs (data not shown) as previously reported [3]. LCs also exhibited many dendrites in untreated as well as OCT or control gel-treated unwounded skin after 48 hours of culture as evidenced by staining of epidermal sheets with antibodies directed against CD207, CD1a, and HLA-DR (Figures 2(a), 2(e), and 2(f); upper panels). Of note, we found a trend toward slightly less LCs in general and fewer dendrites per LCs in particular compared to freshly isolated unwounded skin (data not shown). No significant changes in LC density became apparent in untreated unwounded skin compared to unwounded skin upon topical application of OCT or control gel (Figure 2(b)). However, when the skin was wounded, LCs underwent distinct changes after 48 hours of culture in all groups. Many cells appeared round, some cells had only short surface protrusions or formed one or two single dendrites (Figures 2(c), 2(e), and 2(f); lower panels and insets). Unexpectedly, significantly more LCs were present with a better preservation of the dendritic morphology in the OCT group compared to the control group (Figures 2(c), 2(e), and (f), lower panels and insets). We next investigated whether this observation may correlate with an inhibition of LC maturation. In freshly isolated, unwounded skin, CD207⁺ LCs did not express CD83 (Figure 3(a)). However, after wounding and culture for 48 hours, many CD207⁺CD83⁺ LCs were found in epidermal sheets from untreated and control gel-treated skin, while only some double-positive LCs were found in OCT-treated epidermis (Figure 3(a)). Subsequent enumeration revealed significantly lower numbers of CD207⁺CD83⁺ LCs in epidermal sheets of OCT-treated wounded skin compared to control gel (Figure 3(b)). Our observation that OCT prevents the emigration and maturation of LCs in wounded but not unwounded skin suggests that a potentially higher concentration of OCT in wounded skin may regulate inflammatory cytokines/factors/receptors related with LC maturation and migration as well as/or cytokines and factors that are crucial in balancing/resolving inflammatory responses in our skin model.

(a)

(b)

3.3. OCT Significantly Inhibits the Secretion of IL-8, IL-33, and IL-10 but Not VEGF and TGF-β1

Upon wounding, keratinocytes act as immunomodulators, managing inflammation via a rigorously coordinated network of inflammatory cascades, triggered by keratinocyte-receptor communication with the surroundings in a paracrine and autocrine manner. Among several cytokines, IL-1β and TNF-α are the principal cytokines involved in inflammation-induced LC migration. However, OCT treatment of tape-stripped skin compared to control skin revealed no significant regulation of these cytokines at the mRNA level (data not shown), implying no involvement in the LC behavior in our model. The interplay among proinflammatory and anti-inflammatory cytokines and growth factors and angiogenesis factors determines the inflammatory response. To unravel whether OCT alters their secretion, supernatants from untreated, control gel, and OCT-treated explant cultures with unwounded and wounded skin were analyzed. We found significantly lower IL-8 levels in supernatants of unwounded and wounded OCT-treated cultures compared to controls throughout the observation period of 48 hours (Figure 4(a)). Similarly, when the skin was left unwounded, there was a trend to lower IL-33 levels in OCT-treated skin explants compared to controls, which became even more apparent and statistically significant 48 hours after wounding (Figure 4(b)). Significantly lower levels of IL-10 were measured in supernatants with OCT-treated skin cultures compared to controls upon 48 hours after wounding (Figure 4(c)). Our results clearly show that OCT has either direct or indirect anti-inflammatory properties. We next investigated, whether VEGF, one of the key regulators of angiogenesis, is affected by OCT. We found very low VEGF levels in supernatants of all three groups after 24 hours of culture, which increased at 48 hours, however, with no significant difference between the various treatments groups (Figure 4(d)). Similarly, the production of TGF-β1, which represents one of the most important growth factors with regard to pathological scar formation during wound healing, was not significantly affected upon OCT treatment after 48 hours of culture (Figure 4(e)). Thus, VEGF and TGF-β1 were not altered by the topical application of OCT.

(a)

(b)

(c)

(d)

(e)

4. Discussion

The application of topical antiseptics and antibiotics represents the first strategy of preventing and treating wound infection. However, during the last decades, the unrestrictive use of antibiotics has led to the development of multidrug-resistant pathogens [45], while antiseptics are less likely to cause resistance because of their unspecific mode of action [28]. Data obtained in animal [35–37] and clinical [26, 38–42] studies led to the hypothesis that besides its high antimicrobial effects, OCT may also positively influence wound healing processes including better scar quality.

Ex vivo skin culture models have been used previously to investigate the function of the skin immune system [46, 47]. We provide further work investigating skin viability and morphology as well as following LC behavioral changes upon application of OCT. No obvious changes in skin morphology due to its application on wounded skin was detected, when the most upper layer of the skin, the stratum corneum, was removed to mimic a mild wound process. Even though this skin model is the most advanced to date, we are aware of its limitations since both lymph and blood vasculatures are absent, which are essential for trafficking of immune cells during tissue repair and skin disease.

Several reports have described the properties of LCs in skin organ culture [48–50] as they conceivably represent the closest laboratory model attainable to the in vivo environment with regard of fidelity to physiology as well as biological complexity, even though tissue viability in general vanishes from the time of excision [51]. When healthy human skin is excised and cultured, like in our experiments, LCs start to migrate from the epidermis due to mechanical trauma, triggering a degree of inflammation, orchestrated by skin cytokines, thus mimicking the first part of sensitization and innate immunity. When following LC behavior in all groups of unwounded skin, LCs only partially retracted their dendrites over the whole observation period which was most obvious when viewed in epidermal sheets at the end of the culture period. In contrast, when epidermal sheets from wounded skin were inspected, LCs in all groups displayed a “rounded” morphology already at 48 hours of culture. Previous studies demonstrated comparable changes in LCs after intradermal vaccination, showing a “rounded” morphology and lower LC numbers after 72 hours of culture [52]. Our observations about morphologic changes of LCs in wounded skin in all experimental groups correspond to previous findings [53]. A slightly more pronounced reduction in dendrites per LC was observed in control groups when compared with OCT treatment at 48 hours in wounded skin, implying that OCT may preserve the LC morphology. In line with this, LC numbers in OCT-treated wounded skin were higher compared to untreated skin or control gel. Intriguingly, higher LC numbers in the wounded OCT-treated skin also correlated with the failure of an upregulation of the maturation marker CD83 on LC when compared with the control groups implying that LCs do not undergo a maturation process. These results showed that OCT prevents LC migration to the dermis and inhibits their maturation indicating that OCT may have some influence on signals usually favoring their emigration. To address this hypothesis, potential changes in the production of cytokines and other factors were analyzed in skin explant culture supernatants. Evaluation of the proinflammatory cytokine IL-8 revealed slightly higher levels in wounded skin compared with unwounded skin at 48 hours of culture, indicating that IL-8 secretion is upregulated in wounded skin. Surprisingly, significantly lower IL-8 concentrations were identified in OCT-treated skin cultures compared to controls at all time points. It has been reported that the inhibition of mast cell activation and degranulation led to the downregulation of IL-1β and IL-8 in wounds, which influenced the healing response, characterized by the reduction in wound scar width and improved collagen fiber organization [54]. Previous experiments also revealed increased IL-8 levels in fibroblasts from keloid scars compared with normal human fibroblasts. This highlights a possible role of IL-8 in activation in keloid scars and leukocyte recruitment [55]. Furthermore, it has been shown that the equilibrium in inflammation regulated by low expression of proinflammatory cytokines like IL-8 is crucial in preventing the scar formation in the fetus [56]. Therefore, downregulation of IL-8 secretion by skin cells represents a key point in wound care which might explain the OCT-mediated improved outcome in scar quality as seen in clinical settings.

When analyzing other inflammatory-related cytokines, we identified that OCT largely inhibited IL-33 secretion at all time points in wounded and unwounded skin. Our findings that OCT has anti-inflammatory capacities are in line with results from previous research studies, showing that OCT prevented TNF-α secretion [57], a cytokine involved in inflammation, apoptosis, and immune response [58, 59]. In addition, it has been observed that OCT led to a faster decay of wound inflammation in vivo without occlusive cover in pigs, where redness and swelling of the wound was absent after 4 days of treatment [60]. OCT was also highly effective in the treatment of facial acne lesions [61].

A previous study investigating the correlation between IL-8 and VEGF secretion reported a possible mechanism by which IL-8 and other inflammatory mediators may promote the expression of VEGF in endothelial cells [62]. We identified similar VEGF production but significant inhibition of IL-8 secretion in OCT-treated skin. These data imply that OCT does not affect VEGF secretion but might dampen the inflammatory immune response. IL-10, one of the most important anti-inflammatory cytokines besides TGF-β and IL-35 [63], and also a regulatory cytokine with important functions in the control of inflammation and immune-mediated tissue damage [64], was significantly blocked after OCT treatment. Previous results have shown an increased level of IL-10 in wounded skin of mice together with high levels of TNF-α and more inflammation expressed by increased levels of macrophages and mast cells in wounds suggesting a controlled inflammatory process that favored successful wound healing [65]. TGF-β1, a promising target for the modulation of the cutaneous scarring response during wound healing that influences angiogenesis, inflammatory response, reepithelialization, and extracellular matrix remodeling and deposition, was not significantly altered after OCT treatment. These data imply that OCT does not affect TGF-β1 and VEGF secretion. In conclusion, we have shown that the cytokine secretion pattern of skin cells in wounded and unwounded skin upon topical OCT treatment appears to be rather similarly regulated with regard to the investigated cytokines. However, the observation that the maturation marker CD83 is not upregulated and that LCs do not emigrate from wounded skin but only in unwounded skin strongly suggests that OCT also affects other, possibly yet unexplored, cytokines/factors/receptors in wounded skin and remains to be further explored in a future study.

A reliable skin model, which recapitulates all features of human wound repair, is essential for the clinical and mechanical investigation of human cutaneous wound healing. Although providing highly relevant and promising data in a first stage, we are aware that our currently used ex vivo wound model with sequential tape stripping on human full-thickness skin has limitations, especially as it bears no relevance to deeper wound pathology. Thus, we now aim to investigate the influence of OCT in optimized, clinically even more relevant wound models (e.g., suction blister and biopsy punch) to follow molecular changes during the wound healing process in more detail for a prolonged treatment period.

In conclusion, our data not only provide novel insights into the host response to OCT within the viable human (wounded) skin but also suggest, in addition to its known antimicrobial activity, that a modulation of mediator expression might positively contribute to its wound healing influence resulting in better scar quality.

Data Availability

The data used to support the findings of this study are available from the corresponding author upon request.

Disclosure

Pooja Tajpara’s present affiliation is Department of Medicine, Solna, Center for Molecular Medicine (CMM), Karolinska Institute, 171 76 Stockholm, Sweden. Martin Vierhapper’s present affiliation is Department of Plastic and Reconstructive Surgery, Wilhelminenspital, Montleartstraße 37, Vienna, Austria. Parts of the data were presented as a poster at the Jahrestagung der Österreichischen Gesellschaft für Dermatologie und Venerologie (ÖGDV). Salzburg, Austria, Nov. 30–Dec. 2, 2017.

Conflicts of Interest

The authors declare no conflict of interest.

Acknowledgments

This work was supported by the Austrian Science Fund (FWF; DK W1248-B30) and the Medical Scientific Fund of the Mayor of the City of Vienna (18045). The authors would like to thank Christopher Schuster and René Reitermaier for helpful discussions, as well as Ana Rakita, Ksenija Prpa, and Nada-Katarina Kašić for technical assistance. We thank Erwin Tschachler and Michael Mildner with their teams for providing access to scientific devices and discussions.

References

M. Merad, F. Ginhoux, and M. Collin, “Origin, homeostasis and function of Langerhans cells and other langerin-expressing dendritic cells,” Nature Reviews Immunology, vol. 8, no. 12, pp. 935–947, 2008.
View at: Publisher Site | Google Scholar
T. Ouchi, A. Kubo, M. Yokouchi et al., “Langerhans cell antigen capture through tight junctions confers preemptive immunity in experimental staphylococcal scalded skin syndrome,” The Journal of Experimental Medicine, vol. 208, no. 13, pp. 2607–2613, 2011.
View at: Publisher Site | Google Scholar
M. Chopin and S. L. Nutt, “Establishing and maintaining the Langerhans cell network,” Seminars in Cell & Developmental Biology, vol. 41, pp. 23–29, 2015.
View at: Publisher Site | Google Scholar
T. Hieronymus, M. Zenke, J. H. Baek, and K. Sere, “The clash of Langerhans cell homeostasis in skin: should I stay or should I go?” Seminars in Cell & Developmental Biology, vol. 41, pp. 30–38, 2015.
View at: Publisher Site | Google Scholar
M. Collin and P. Milne, “Langerhans cell origin and regulation,” Current Opinion in Hematology, vol. 23, no. 1, pp. 28–35, 2016.
View at: Publisher Site | Google Scholar
N. Romani, B. E. Clausen, and P. Stoitzner, “Langerhans cells and more: langerin-expressing dendritic cell subsets in the skin,” Immunological Reviews, vol. 234, no. 1, pp. 120–141, 2010.
View at: Publisher Site | Google Scholar
P. Martin, “Wound healing--aiming for perfect skin regeneration,” Science, vol. 276, no. 5309, pp. 75–81, 1997.
View at: Publisher Site | Google Scholar
P. Martin and S. J. Leibovich, “Inflammatory cells during wound repair: the good, the bad and the ugly,” Trends in Cell Biology, vol. 15, no. 11, pp. 599–607, 2005.
View at: Publisher Site | Google Scholar
G. C. Gurtner, S. Werner, Y. Barrandon, and M. T. Longaker, “Wound repair and regeneration,” Nature, vol. 453, no. 7193, pp. 314–321, 2008.
View at: Publisher Site | Google Scholar
L. J. van den Broek, G. C. Limandjaja, F. B. Niessen, and S. Gibbs, “Human hypertrophic and keloid scar models: principles, limitations and future challenges from a tissue engineering perspective,” Experimental Dermatology, vol. 23, no. 6, pp. 382–386, 2014.
View at: Publisher Site | Google Scholar
J. Gregorio, S. Meller, C. Conrad et al., “Plasmacytoid dendritic cells sense skin injury and promote wound healing through type I interferons,” The Journal of Experimental Medicine, vol. 207, no. 13, pp. 2921–2930, 2010.
View at: Publisher Site | Google Scholar
Z. Han, Y. Chen, Y. Zhang et al., “MiR-21/PTEN axis promotes skin wound healing by dendritic cells enhancement,” Journal of Cellular Biochemistry, vol. 118, no. 10, pp. 3511–3519, 2017.
View at: Publisher Site | Google Scholar
S. Werner, T. Krieg, and H. Smola, “Keratinocyte-fibroblast interactions in wound healing,” The Journal of Investigative Dermatology, vol. 127, no. 5, pp. 998–1008, 2007.
View at: Publisher Site | Google Scholar
S. W. Spiekstra, M. Breetveld, T. Rustemeyer, R. J. Scheper, and S. Gibbs, “Wound-healing factors secreted by epidermal keratinocytes and dermal fibroblasts in skin substitutes,” Wound Repair and Regeneration, vol. 15, no. 5, pp. 708–717, 2007.
View at: Publisher Site | Google Scholar
A. Toulon, L. Breton, K. R. Taylor et al., “A role for human skin-resident T cells in wound healing,” The Journal of Experimental Medicine, vol. 206, no. 4, pp. 743–750, 2009.
View at: Publisher Site | Google Scholar
C. Noli and A. Miolo, “The mast cell in wound healing,” Veterinary Dermatology, vol. 12, no. 6, pp. 303–313, 2001.
View at: Publisher Site | Google Scholar
N. Strbo, N. Yin, and O. Stojadinovic, “Innate and adaptive immune responses in wound epithelialization,” Advances in Wound Care, vol. 3, no. 7, pp. 492–501, 2014.
View at: Publisher Site | Google Scholar
I. Juhasz, M. Simon Jr., M. Herlyn, and J. Hunyadi, “Repopulation of Langerhans cells during wound healing in an experimental human skin/SCID mouse model,” Immunology Letters, vol. 52, no. 2-3, pp. 125–128, 1996.
View at: Publisher Site | Google Scholar
T. Helfman, J. W. Streilein, W. H. Eaglstein, and P. M. Mertz, “Studies on the repopulation of Langerhans cells in partial-thickness wounds. Air exposed and occlusively dressed,” Archives of Dermatology, vol. 129, no. 5, pp. 592–595, 1993.
View at: Publisher Site | Google Scholar
O. Stojadinovic, N. Yin, J. Lehmann, I. Pastar, R. S. Kirsner, and M. Tomic-Canic, “Increased number of Langerhans cells in the epidermis of diabetic foot ulcers correlates with healing outcome,” Immunologic Research, vol. 57, no. 1-3, pp. 222–228, 2013.
View at: Publisher Site | Google Scholar
T. Koburger, N. O. Hubner, M. Braun, J. Siebert, and A. Kramer, “Standardized comparison of antiseptic efficacy of triclosan, PVP-iodine, octenidine dihydrochloride, polyhexanide and chlorhexidine digluconate,” The Journal of Antimicrobial Chemotherapy, vol. 65, no. 8, pp. 1712–1719, 2010.
View at: Publisher Site | Google Scholar
A. M. Slee and J. R. O'Connor, “In vitro antiplaque activity of octenidine dihydrochloride (WIN 41464-2) against preformed plaques of selected oral plaque-forming microorganisms,” Antimicrobial Agents and Chemotherapy, vol. 23, no. 3, pp. 379–384, 1983.
View at: Publisher Site | Google Scholar
T. Conceicao, H. de Lencastre, and M. Aires-de-Sousa, “Efficacy of octenidine against antibiotic-resistant Staphylococcus aureus epidemic clones,” The Journal of Antimicrobial Chemotherapy, vol. 71, no. 10, pp. 2991–2994, 2016.
View at: Publisher Site | Google Scholar
R. Alvarez-Marin, M. Aires-de-Sousa, P. Nordmann, N. Kieffer, and L. Poirel, “Antimicrobial activity of octenidine against multidrug-resistant Gram-negative pathogens,” European Journal of Clinical Microbiology & Infectious Diseases, vol. 36, no. 12, pp. 2379–2383, 2017.
View at: Publisher Site | Google Scholar
O. Assadian, “Octenidine dihydrochloride: chemical characteristics and antimicrobial properties,” Journal of Wound Care, vol. 25, no. Sup3, pp. S3–S6, 2016.
View at: Publisher Site | Google Scholar
W. Vanscheidt, K. Harding, L. Teot, and J. Siebert, “Effectiveness and tissue compatibility of a 12-week treatment of chronic venous leg ulcers with an octenidine based antiseptic - a randomized, double-blind controlled study,” International Wound Journal, vol. 9, no. 3, pp. 316–323, 2012.
View at: Publisher Site | Google Scholar
G. Muller and A. Kramer, “Biocompatibility index of antiseptic agents by parallel assessment of antimicrobial activity and cellular cytotoxicity,” The Journal of Antimicrobial Chemotherapy, vol. 61, no. 6, pp. 1281–1287, 2008.
View at: Publisher Site | Google Scholar
N. O. Hubner, J. Siebert, and A. Kramer, “Octenidine dihydrochloride, a modern antiseptic for skin, mucous membranes and wounds,” Skin Pharmacology and Physiology, vol. 23, no. 5, pp. 244–258, 2010.
View at: Publisher Site | Google Scholar
G. Pichler, C. Pux, R. Babeluk et al., “MRSA prevalence rates detected in a tertiary care hospital in Austria and successful treatment of MRSA positive patients applying a decontamination regime with octenidine,” European Journal of Clinical Microbiology & Infectious Diseases, vol. 37, no. 1, pp. 21–27, 2018.
View at: Publisher Site | Google Scholar
P. Gastmeier, K. P. Kampf, M. Behnke, C. Geffers, and F. Schwab, “An observational study of the universal use of octenidine to decrease nosocomial bloodstream infections and MDR organisms,” The Journal of Antimicrobial Chemotherapy, vol. 71, no. 9, pp. 2569–2576, 2016.
View at: Publisher Site | Google Scholar
U. Rohr, C. Mueller, M. Wilhelm, G. Muhr, and S. Gatermann, “Methicillin-resistant Staphylococcus aureus whole-body decolonization among hospitalized patients with variable site colonization by using mupirocin in combination with octenidine dihydrochloride,” The Journal of Hospital Infection, vol. 54, no. 4, pp. 305–309, 2003.
View at: Publisher Site | Google Scholar
Z. Al-Doori, P. Goroncy-Bermes, C. G. Gemmell, and D. Morrison, “Low-level exposure of MRSA to octenidine dihydrochloride does not select for resistance,” The Journal of Antimicrobial Chemotherapy, vol. 59, no. 6, pp. 1280-1281, 2007.
View at: Publisher Site | Google Scholar
T. Calow, K. Oberle, L. Bruckner-Tuderman, T. Jakob, and H. Schumann, “Contact dermatitis due to use of Octenisept in wound care,” Journal der Deutschen Dermatologischen Gesellschaft, vol. 7, no. 9, pp. 759–765, 2009.
View at: Publisher Site | Google Scholar
J. M. Lachapelle, “A comparison of the irritant and allergenic properties of antiseptics,” European Journal of Dermatology, vol. 24, no. 1, pp. 3–9, 2014.
View at: Publisher Site | Google Scholar
S. Langer, M. Sedigh Salakdeh, O. Goertz, H. U. Steinau, L. Steinstraesser, and H. H. Homann, “The impact of topical antiseptics on skin microcirculation,” European Journal of Medical Research, vol. 9, no. 9, pp. 449–454, 2004.
View at: Google Scholar
O. Goertz, T. Hirsch, A. Ring et al., “Influence of antiseptics on microcirculation after neuronal and receptor blockade,” International Wound Journal, vol. 8, no. 4, pp. 365–373, 2011.
View at: Publisher Site | Google Scholar
A. Kramer, B. Roth, G. Muller, P. Rudolph, and N. Klocker, “Influence of the antiseptic agents polyhexanide and octenidine on FL cells and on healing of experimental superficial aseptic wounds in piglets. A double-blind, randomised, stratified, controlled, parallel-group study,” Skin Pharmacology and Physiology, vol. 17, no. 3, pp. 141–146, 2004.
View at: Publisher Site | Google Scholar
J. Matiasek, G. Djedovic, L. Unger et al., “Outcomes for split-thickness skin transplantation in high-risk patients using octenidine,” Journal of Wound Care, vol. 24, no. Sup6, pp. S8–S12, 2015.
View at: Publisher Site | Google Scholar
G. Hammerle and R. Strohal, “Efficacy and cost-effectiveness of octenidine wound gel in the treatment of chronic venous leg ulcers in comparison to modern wound dressings,” International Wound Journal, vol. 13, no. 2, pp. 182–188, 2016.
View at: Publisher Site | Google Scholar
W. Eisenbeiß, G. Amtsberg, P. Hinz et al., “Prospective, double-blinded, randomised controlled trial assessing the effect of an octenidine-based hydrogel on bacterial colonisation and epithelialization of skin graft wounds in burn patients,” International Journal of Burns and Trauma, vol. 2, pp. 71–79, 2012.
View at: Google Scholar
J. Matiasek, G. Djedovic, M. Mattesich et al., “The combined use of NPWT and instillation using an octenidine based wound rinsing solution: a case study,” Journal of Wound Care, vol. 23, no. 11, pp. 590–596, 2014.
View at: Publisher Site | Google Scholar
J. Matiasek, P. Kienzl, L. W. Unger, C. Grill, R. Koller, and B. R. Turk, “An intra-individual surgical wound comparison shows that octenidine-based hydrogel wound dressing ameliorates scar appearance following abdominoplasty,” International Wound Journal, vol. 15, no. 6, pp. 914–920, 2018.
View at: Publisher Site | Google Scholar
P. Tajpara, C. Schuster, E. Schön et al., “Epicutaneous administration of the pattern recognition receptor agonist polyinosinic-polycytidylic acid activates the MDA5/MAVS pathway in Langerhans cells,” The FASEB Journal, vol. 32, no. 8, pp. 4132–4144, 2018.
View at: Publisher Site | Google Scholar
M. Mildner, J. Jin, L. Eckhart et al., “Knockdown of filaggrin impairs diffusion barrier function and increases UV sensitivity in a human skin model,” The Journal of Investigative Dermatology, vol. 130, no. 9, pp. 2286–2294, 2010.
View at: Publisher Site | Google Scholar
T. Hirsch, A. Koerber, F. Jacobsen et al., “Evaluation of toxic side effects of clinically used skin antiseptics in vitro,” The Journal of Surgical Research, vol. 164, no. 2, pp. 344–350, 2010.
View at: Publisher Site | Google Scholar
T. D. de Gruijl, O. J. A. E. Ophorst, J. Goudsmit et al., “Intradermal delivery of adenoviral type-35 vectors leads to high efficiency transduction of mature, CD8⁺ T cell-stimulating skin-emigrated dendritic cells,” Journal of Immunology, vol. 177, no. 4, pp. 2208–2215, 2006.
View at: Publisher Site | Google Scholar
C. L. Lehe, J. J. L. Jacobs, C. M. Hua, P. Courtellemont, G. R. Elliott, and P. K. Das, “Subtoxic concentrations of allergenic haptens induce LC migration and maturation in a human organotypic skin explant culture model: a novel method for identifying potential contact allergens,” Experimental Dermatology, vol. 15, no. 6, pp. 421–431, 2006.
View at: Publisher Site | Google Scholar
C. P. Larsen, R. M. Steinman, M. Witmer-Pack, D. F. Hankins, P. J. Morris, and J. M. Austyn, “Migration and maturation of Langerhans cells in skin transplants and explants,” Journal of Experimental Medicine, vol. 172, no. 5, pp. 1483–1493, 1990.
View at: Publisher Site | Google Scholar
A. Rambukkana, J. D. Bos, D. Irik, W. J. Menko, M. L. Kapsenberg, and P. K. Das, “In situ behavior of human Langerhans cells in skin organ culture,” Laboratory Investigation, vol. 73, no. 4, pp. 521–531, 1995.
View at: Google Scholar
M. Pearton, S. M. Kang, J. M. Song et al., “Changes in human Langerhans cells following intradermal injection of influenza virus-like particle vaccines,” PLoS One, vol. 5, no. 8, article e12410, 2010.
View at: Publisher Site | Google Scholar
Y. Tomita, M. Nihira, Y. Ohno, and S. Sato, “Ultrastructural changes during in situ early postmortem autolysis in kidney, pancreas, liver, heart and skeletal muscle of rats,” Legal Medicine, vol. 6, no. 1, pp. 25–31, 2004.
View at: Publisher Site | Google Scholar
K. W. Ng, M. Pearton, S. Coulman et al., “Development of an ex vivo human skin model for intradermal vaccination: tissue viability and Langerhans cell behaviour,” Vaccine, vol. 27, no. 43, pp. 5948–5955, 2009.
View at: Publisher Site | Google Scholar
S. Holzmann, C. H. Tripp, M. Schmuth et al., “A model system using tape stripping for characterization of Langerhans cell-precursors in vivo,” The Journal of Investigative Dermatology, vol. 122, no. 5, pp. 1165–1174, 2004.
View at: Publisher Site | Google Scholar
L. Chen, M. E. Schrementi, M. J. Ranzer, T. A. Wilgus, and L. A. DiPietro, “Blockade of mast cell activation reduces cutaneous scar formation,” PLoS One, vol. 9, no. 1, article e85226, 2014.
View at: Publisher Site | Google Scholar
C. P. Lim, T. T. Phan, I. J. Lim, and X. Cao, “Cytokine profiling and Stat3 phosphorylation in epithelial-mesenchymal interactions between keloid keratinocytes and fibroblasts,” The Journal of Investigative Dermatology, vol. 129, no. 4, pp. 851–861, 2009.
View at: Publisher Site | Google Scholar
C. Zgheib, J. Xu, and K. W. Liechty, “Targeting inflammatory cytokines and extracellular matrix composition to promote wound regeneration,” Advances in Wound Care, vol. 3, no. 4, pp. 344–355, 2014.
View at: Publisher Site | Google Scholar
H. Menke, M. Pelzer, T. Raff, J. Siebert, and G. Germann, “A new local antiseptic for topical treatment in patients with deep burns - results of the follow-up of local and systemic effects after application,” Aktuelle Traumatologie, vol. 31, no. 5, pp. 211–215, 2001.
View at: Publisher Site | Google Scholar
R. M. Locksley, N. Killeen, and M. J. Lenardo, “The TNF and TNF receptor superfamilies: integrating mammalian biology,” Cell, vol. 104, no. 4, pp. 487–501, 2001.
View at: Publisher Site | Google Scholar
T. Banno, A. Gazel, and M. Blumenberg, “Effects of tumor necrosis factor-α (TNF α) in epidermal keratinocytes revealed using global transcriptional profiling,” The Journal of Biological Chemistry, vol. 279, no. 31, pp. 32633–32642, 2004.
View at: Publisher Site | Google Scholar
J. Stahl, M. Braun, J. Siebert, and M. Kietzmann, “The effect of a combination of 0.1% octenidine dihydrochloride and 2% 2-phenoxyethanol (octenisept®) on wound healing in pigs in vivo and its in vitro percutaneous permeation through intact and barrier disrupted porcine skin,” International Wound Journal, vol. 7, no. 1, pp. 62–69, 2010.
View at: Publisher Site | Google Scholar
S. Mayr-Kanhauser, B. Kranke, and W. Aberer, “Efficacy of octenidine dihydrochloride and 2-phenoxyethanol in the topical treatment of inflammatory acne,” Acta Dermatovenerologica Alpina, Pannonica, et Adriatica, vol. 17, no. 3, pp. 139–143, 2008.
View at: Google Scholar
D. Martin, R. Galisteo, and J. S. Gutkind, “CXCL8/IL8 stimulates vascular endothelial growth factor (VEGF) expression and the autocrine activation of VEGFR2 in endothelial cells by activating NFκB through the CBM (Carma3/Bcl10/Malt1) complex,” The Journal of Biological Chemistry, vol. 284, no. 10, pp. 6038–6042, 2009.
View at: Publisher Site | Google Scholar
R. Sabat, G. Grütz, K. Warszawska et al., “Biology of interleukin 10,” Cytokine & Growth Factor Reviews, vol. 21, no. 5, pp. 331–344, 2010.
View at: Publisher Site | Google Scholar
W. H. Peranteau, L. Zhang, N. Muvarak et al., “IL-10 overexpression decreases inflammatory mediators and promotes regenerative healing in an adult model of scar formation,” The Journal of Investigative Dermatology, vol. 128, no. 7, pp. 1852–1860, 2008.
View at: Publisher Site | Google Scholar
M. C. C. Canesso, A. T. Vieira, T. B. R. Castro et al., “Skin wound healing is accelerated and scarless in the absence of commensal microbiota,” Journal of Immunology, vol. 193, no. 10, pp. 5171–5180, 2014.
View at: Publisher Site | Google Scholar

Copyright

Copyright © 2019 Nenad Nikolić et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

PDF Download Citation

Download other formats

Order printed copies

Views

4970

Downloads

1476

Citations