A novel murine model of pyoderma gangrenosum reveals that inflammatory skin-gut crosstalk is mediated by IL-1ß-primed neutrophils.

Pyoderma gangrenosum (PG) is a debilitating skin condition often accompanied by inflammatory bowel disease (IBD). Strikingly, ~40% of patients that present with PG have underlying IBD, suggesting shared but unknown mechanisms of pathogenesis. Impeding the development of effective treatments for PG is the absence of an animal model that exhibits features of both skin and gut manifestations. This study describes the development of the first experimental drug-induced mouse model of PG with concomitant intestinal inflammation. Topical application of pyrimidine synthesis inhibitors on wounded mouse skin generates skin ulcers enriched in neutrophil extracellular traps (NETs) as well as pro-inflammatory cellular and soluble mediators mimicking human PG. The mice also develop spontaneous intestinal inflammation demonstrated by histologic damage. Further investigations revealed increased circulating low density IL-1ß primed neutrophils that undergo enhanced NETosis at inflamed tissue sites supported by an increase in circulatory citrullinated histone 3, a marker of aberrant NET formation. Granulocyte depletion dampens the intestinal inflammation in this model, further supporting the notion that granulocytes contribute to the skin-gut crosstalk in PG mice. We anticipate that this novel murine PG model will enable researchers to probe common disease mechanisms and identify more effective targets for treatment for PG patients with IBD.

Abnormal thrombosis and neutrophil activation increase hospital-acquired sacral pressure injuries and morbidity in COVID-19 patients.

Hospitalized patients have an increased risk of developing hospital-acquired sacral pressure injury (HASPI). However, it is unknown whether SARS-CoV-2 infection affects HASPI development. To explore the role of SARS-CoV-2 infection in HASPI development, we conducted a single institution, multi-hospital, retrospective study of all patients hospitalized for ≥5 days from March 1, 2020 to December 31, 2020. Patient demographics, hospitalization information, ulcer characteristics, and 30-day-related morbidity were collected for all patients with HASPIs, and intact skin was collected from HASPI borders in a patient subset. We determined the incidence, disease course, and short-term morbidity of HASPIs in COVID-19(+) patients, and characterized the skin histopathology and tissue gene signatures associated with HASPIs in COVID-19 disease. COVID-19(+) patients had a 63% increased HASPI incidence rate, HASPIs of more severe ulcer stage (OR 2.0, p<0.001), and HASPIs more likely to require debridement (OR 3.1, p=0.04) compared to COVID-19(-) patients. Furthermore, COVID-19(+) patients with HASPIs had 2.2x increased odds of a more severe hospitalization course compared to COVID-19(+) patients without HASPIs. HASPI skin histology from COVID-19(+) patients predominantly showed thrombotic vasculopathy, with the number of thrombosed vessels being significantly greater than HASPIs from COVID-19(-) patients. Transcriptional signatures of a COVID-19(+) sample subset were enriched for innate immune responses, thrombosis, and neutrophil activation genes. Overall, our results suggest that immunologic dysregulation secondary to SARS-CoV-2 infection, including neutrophil dysfunction and abnormal thrombosis, may play a pathogenic role in development of HASPIs in patients with severe COVID-19.

Development of a reproducible porcine model of infected burn wounds.

Severe burns are traumatic and physically debilitating injuries with a high rate of mortality. Bacterial infections often complicate burn injuries, which presents unique challenges for wound management and improved patient outcomes. Currently, pigs are used as the gold standard of pre-clinical models to study infected skin wounds due to the similarity between porcine and human skin in terms of structure and immunological response. However, utilizing this large animal model for wound infection studies can be technically challenging and create issues with data reproducibility. We present a detailed protocol for a porcine model of infected burn wounds based on our experience in creating and evaluating full thickness burn wounds infected with Staphylococcus aureus on six pigs. Wound healing kinetics and bacterial clearance were measured over a period of 27 d in this model. Enumerated are steps to achieve standardized wound creation, bacterial inoculation, and dressing techniques. Systematic evaluation of wound healing and bacterial colonization of the wound bed is also described. Finally, advice on animal housing considerations, efficient bacterial plating procedures, and overcoming common technical challenges is provided. This protocol aims to provide investigators with a step-by-step guide to execute a technically challenging porcine wound infection model in a reproducible manner. Accordingly, this would allow for the design and evaluation of more effective burn infection therapies leading to better strategies for patient care.

Thread Size and Polymer Composition of 3D Printed and Electrospun Wound Dressings Affect Wound Healing Outcomes in an Excisional Wound Rat Model.

Thread size and polymer composition are critical properties to consider for achieving a positive healing outcome with a wound dressing. Three-dimensional (3D) printed scaffolds and electrospun mats both offer distinct advantages as replaceable wound dressings. This research aims to determine if the thread size and polymer compositions of the scaffolds affect skin wound healing outcomes, an aspect that has not been adequately explored. Using a modular polymer platform, four polyester direct-write 3D printed scaffolds and electrospun mats were fabricated into wound dressings. The dressings were applied to splinted, full thickness skin wounds in an excisional wound rat model and evaluated against control wounds to which no dressing was applied. Wound closure rates and reduction of the wound bed width were not affected by the thread size or polymer composition. However, epidermal thickness was larger in wounds treated with electrospun dressings and was slightly affected by the polymer composition. Two of the four tested polymer compositions lead to delayed reorganization of granulation tissues. Moreover, enhanced angiogenesis was seen in wounds treated with 3D printed dressings compared to those treated with electrospun dressings. The results from this study can be used to inform the choice of dressing architecture and polymer compositions to achieve positive wound healing outcomes.

Amorphous silicon dioxide nanoparticles modulate immune responses in a model of allergic contact dermatitis.

Amorphous silicon dioxide nanoparticles (SiNPs) are ubiquitous, and they are currently found in cosmetics, drugs, and foods. Biomedical research is also focused on using these nanoparticles as drug delivery and bio-sensing platforms. Due to the high potential for skin exposure to SiNPs, research into the effect of topical exposure on both healthy and inflammatory skin models is warranted. While we observe only minimal effects of SiNPs on healthy mouse skin, there is an immunomodulatory effect of these NPs in a model of allergic contact dermatitis. The effect appears to be mediated partly by keratinocytes and results in decreases in epidermal hyperplasia, inflammatory cytokine release, immune cell infiltration, and a subsequent reduction in skin swelling. Additional research is required to further our mechanistic understanding and to validate the extent of this immunomodulatory effect in human subjects in order to assess the potential prophylactic use of SiNPs for treating allergic skin conditions.

Pyrimidine synthesis inhibition enhances cutaneous defenses against antibiotic resistant bacteria through activation of NOD2 signaling.

Multidrug-resistant bacterial strains are a rapidly emerging healthcare threat; therefore it is critical to develop new therapies to combat these organisms. Prior antibacterial strategies directly target pathogen growth or viability. Host-directed strategies to increase antimicrobial defenses may be an effective alternative to antibiotics and reduce development of resistant strains. In this study, we demonstrated the efficacy of a pyrimidine synthesis inhibitor, N-phosphonacetyl-L-aspartate (PALA), to enhance clearance of methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa, and Acinetobacter baumannii strains by primary human dermal fibroblasts in vitro. PALA did not have a direct bactericidal effect, but enhanced cellular secretion of the antimicrobial peptides human ß-defensin 2 (HBD2) and HBD3 from fibroblasts. When tested in porcine and human skin explant models, a topical PALA formulation was efficacious to enhance MRSA, P. aeruginosa, and A. baumannii clearance. Topical PALA treatment of human skin explants also resulted in increased HBD2 and cathelicidin (LL-37) production. The antimicrobial actions of PALA required expression of nucleotide-binding, oligomerization domain 2 (NOD2), receptor-interacting serine/threonine-protein kinase 2 (RIP2), and carbamoyl phosphatase synthase II/aspartate transcarbamylase/dihydroorotase (CAD). Our results indicate that PALA may be a new option to combat multidrug-resistant bacterial infections of the skin through enhancement of an integral pathway of the cutaneous innate immune defense system.

Immunomodulatory Effects of Nanoparticles on Skin Allergy.

In recent years there has been considerable effort to understand the interaction of nanomaterials with the skin. In this study we use an in vivo mouse model of allergic contact dermatitis to investigate how nanoparticles (NPs) may alter allergic responses in skin. We investigate a variety of NPs that vary in size, charge and composition. Results show that small (<200 nm) negative and neutral charged NPs exhibit an immunosuppressive effect but that positively charged NPs do not. Confocal imaging suggests positively charged NPs may penetrate skin to a lesser extent and thereby are less able interact with and alter the local immune responses. Interestingly, negatively charged silica (20 nm) NPs suppress allergic response to two chemically distinct sensitizers; 1-fluoro-2, 4-dinitrobenzene and 2-deoxyurushiol. Skin wiping and NP application time studies suggest that the immunomodulatory mechanism is not due solely to the blocking of sensitizer adduct formation in skin. Results suggest that NPs modulate early immune events that impact mast cell degranulation. Our study shows for the first time the potential to modulate the elicitation phase of the allergic response which depends on the NP charge and composition. These finding can be used to inform the design topical therapeutics to mitigate allergic responses in skin.

In vivo quantification of quantum dot systemic transport in C57BL/6 hairless mice following skin application post-ultraviolet radiation.

BACKGROUND: Previous work has demonstrated size, surface charge and skin barrier dependent penetration of nanoparticles into the viable layers of mouse skin. The goal of this work was to characterize the tissue distribution and mechanism of transport of nanoparticles beyond skin, with and without Ultraviolet Radiation (UVR) induced skin barrier disruption. Atomic absorption spectroscopy (AAS), flow cytometry and confocal microscopy were used to examine the effect of UVR dose (180 and 360 mJ/cm2 UVB) on the skin penetration and systemic distribution of quantum dot (QD) nanoparticles topically applied at different time-points post UVR using a hairless C57BL/6 mouse model. RESULTS: Results indicate that QDs can penetrate mouse skin, regardless of UVR exposure, as evidenced by the increased cadmium in the local lymph nodes of all QD treated mice. The average % recovery for all treatment groups was 69.68% with ~66.84% of the applied dose recovered from the skin (both epicutaneous and intracutaneous). An average of 0.024% of the applied dose was recovered from the lymph nodes across various treatment groups. When QDs are applied 4 days post UV irradiation, at the peak of the skin barrier defect and LC migration to the local lymph node, there is an increased cellular presence of QD in the lymph node; however, AAS analysis of local lymph nodes display no difference in cadmium levels due to UVR treatment. CONCLUSIONS: Our data suggests that Langerhans cells (LCs) can engulf QDs in skin, but transport to the lymph node may occur by both cellular (dendritic and macrophage) and non-cellular mechanisms. It is interesting that these specific nanoparticles were retained in skin similarly regardless of UVR barrier disruption, but the observed skin immune cell interaction with nanoparticles suggest a potential for immunomodulation, which we are currently examining in a murine model of skin allergy.

Impact of Cosmetic Lotions on Nanoparticle Penetration through ex vivo C57BL/6 Hairless Mouse and Human Skin: A Comparison Study.

Understanding the interactions of nanoparticles (NPs) with skin is important from a consumer and occupational health and safety perspective, as well as for the design of effective NP-based transdermal therapeutics. Despite intense efforts to elucidate the conditions that permit NP penetration, there remains a lack of translatable results from animal models to human skin. The objectives of this study are to investigate the impact of common skin lotions on NP penetration and to quantify penetration differences of quantum dot (QD) NPs between freshly excised human and mouse skin. QDs were mixed in 7 different vehicles, including 5 commercial skin lotions. These were topically applied to skin using two exposure methods; a petri dish protocol and a Franz diffusion cell protocol. QD presence in the skin was quantified using Confocal Laser Scanning Microscopy. Results show that the commercial vehicles can significantly impact QD penetration in both mouse and human skin. Lotions that contain alpha hydroxyl acids (AHA) facilitated NP penetration. Lower QD signal was observed in skin studied using a Franz cell. Freshly excised human skin was also studied immediately after the sub-cutaneous fat removal process, then after 24 hours rest ex vivo. Resting human skin 24 hours prior to QD exposure significantly reduced epidermal presence. This study exemplifies how application vehicles, skin processing and the exposure protocol can affect QD penetration results and the conclusions that maybe drawn between skin models.

Understanding engineered nanomaterial skin interactions and the modulatory effects of ultraviolet radiation skin exposure.

The study of engineered nanomaterials for the development of technological applications, nanomedicine, and nano-enabled consumer products is an ever-expanding discipline as is the concern over the impact of nanotechnology on human environmental health and safety. In this review, we discuss the current state of understanding of nanomaterial skin interactions with a specific emphasis on the effects of ultraviolet radiation (UVR) skin exposure. Skin is the largest organ of the body and is typically exposed to UVR on a daily basis. This necessitates the need to understand how UVR skin exposure can influence nanomaterial skin penetration, alter nanomaterial systemic trafficking, toxicity, and skin immune function. We explore the unique dichotomy that UVR has on inducing both deleterious and therapeutic effects in skin. The subject matter covered in this review is broadly informative and will raise awareness of potential increased risks from nanomaterial skin exposure associated with specific occupational and life style choices. The UVR-induced immunosuppressive response in skin raises intriguing questions that motivate future research directions in the nanotoxicology and nanomedicine fields.

Quantification of quantum dot murine skin penetration with UVR barrier impairment.

Ultraviolet radiation (UVR) skin exposure is a common exogenous insult that can alter skin barrier and immune functions. With the growing presence of nanoparticles (NPs) in consumer goods and technological applications the potential for NPs to contact UVR-exposed skin is increasing. Therefore it is important to understand the effect of UVR on NP skin penetration and the potential for systemic translocation. Previous studies qualitatively showed that UVR skin exposure can increase the penetration of NPs below the stratum corneum. In this work, an in vivo mouse model was used to quantitatively examine the skin penetration of carboxylated (CdSe/ZnS, core/shell) quantum dots (QDs) through intact and UVR barrier-disrupted murine skin by organ Cd mass analysis. Transepidermal water loss was used to measure the magnitude of the skin barrier defect as a function of UVR dose and time post-UVR exposure. QDs were applied to mice 3-4 days post-UVR exposure at the peak of the skin barrier disruption. Our results reveal unexpected trends that suggest these negative-charged QDs can penetrate barrier intact skin and that penetration and systemic transport depends on the QD application time post-UVR exposure. The effect of UVR on skin-resident dendritic cells and their role in the systemic translocation of these QDs are described. Our results suggest that NP skin penetration and translocation may depend on the specific barrier insult and the inflammatory status of the skin.