Optimal microneedle length for hair regrowth in hair cycle arrest (alopecia X) in six dogs.

BACKGROUND: Hair cycle arrest (HCA) is a chronic alopecic disorder in dogs. Clinical responses vary and are often insufficient. Microneedling (MN) has been used as a successful treatment for HCA in dogs; ideal protocols have not yet been established. OBJECTIVES: The objective of the study was to compare the efficacy and safety of three needle lengths for MN in dogs with HCA. ANIMALS: Six unrelated client-owned dogs, including five Pomeranians and one mixed-breed dog, diagnosed with HCA. MATERIALS AND METHODS: Individual alopecic sites were divided into three sections. For each section, different lengths of needles (1, 2 and 3 mm) were used. Efficacy and safety were evaluated at 1, 3 and 6 months. Treated sections were monitored for 20 months. RESULTS: Three months after treatment with 3 mm needles, all sections showed hair regrowth. There was no hair regrowth in two of six sections treated with 2 mm needles, and four of six sections did not show a response to treatment with 1 mm needles. Two dogs developed transient pruritus. Five of six dogs had recurrent hair loss between 5 and 16 months of follow-up. CONCLUSION AND CLINICAL RELEVANCE: Microneedling using longer needles stimulated better hair regrowth in dogs with HCA. Alopecia relapsed in most dogs and minor pruritus occurred in some dogs.

Application of cold atmospheric microwave plasma as an adjunct therapy for wound healing in dogs and cats.

BACKGROUND: Cold atmospheric plasma is a novel innovative approach for wound care, and it is currently underrepresented in veterinary medicine. OBJECTIVES: To investigate the efficacy and safety of using cold atmospheric microwave plasma (CAMP) as an adjunct therapy for wound healing in dogs and cats. METHODS: Wound healing outcomes were retrospectively analyzed using clinical records of client-owned dogs and cats who were first managed through standard wound care alone (pre-CAMP period) and subsequently via CAMP therapy (CAMP period). The degree of wound healing was estimated based on wound size and a modified wound scoring system. RESULTS: Of the 27 acute and chronic wounds included in the analysis, 81.48% showed complete healing after the administration of CAMP as an adjunct therapy to standard care. Most wounds achieved complete healing in < 5 weeks. Compared with the pre-CAMP period, the rate of wound healing significantly increased every week in the CAMP period in terms of in wound size (first week, p < 0.001; second week, p = 0.012; third week, p < 0.001) and wound score (first week, p < 0.001; second week, p < 0.001; third week, p = 0.001). No adverse events were noted except for mild discomfort and transient erythema. CONCLUSIONS: CAMP is a well-tolerated therapeutic option with immense potential to support the treatment of wounds of diverse etiology in small animal practice. Further research is warranted to establish specific criteria for CAMP treatment according to wound characteristics.

Effect of cold atmospheric microwave plasma (CAMP) on wound healing in canine keratinocytes.

Cutaneous wound healing is a biological process that occurs upon skin injury and involves different mechanisms to repair tissue damage. Improper healing or prolonged curation period of wound lesions may induce unpleasant complications. Cold atmospheric microwave plasma (CAMP) is an upcoming medical therapeutic option for skin infection and wound treatment. However, the molecular mechanisms of CAMP-mediated canine wound healing are not well characterized. Wound-healing activity was examined to elucidate the biological effects and molecular mechanisms of CAMP. Canine keratinocytes (CPEKs) were treated using CAMP, and their wound-healing activities were evaluated. The molecular mechanisms of that effect were examined, based on RNA-Seq analysis data, and verified using immunoblotting and polymerase chain reaction. It was found that the CAMP-treated cells exhibited a significant increase in cell migration evaluated by scratch assay in human keratinocytes (HaCaT) and canine keratinocytes (CPEK). Additionally, CAMP-treated CPEK cells showed a significant positive effect on cell invasion. The RNA-Seq data revealed that CAMP alters different genes and pathways in CPEK cells. Gene expression involved in the cell cycle, cell proliferation, angiogenesis, cell adhesion, and wound healing was upregulated in CAMP-treated cells compared with gas-activated media used as a control. The Hippo pathway was also analyzed, and the protein and mRNA levels of YAP were significantly increased in CAMP-treated cells. CAMP-treated CPEK cells indicated the downregulation of E-cadherin and upregulation of vimentin, Snail, and Slug at transcription and translation levels, contributing to a favorable effect on cell migration. Our findings suggested that CAMP treatment provided beneficial effects on the curative wound process through the induction of genes involved in wound healing, promotion of EMT, and increase in the molecular targets in the Hippo signaling pathway.

The number of mitochondrial DNA mutations as a genetic feature for hair cycle arrest (alopecia X) in Pomeranian dogs.

BACKGROUND: Hair cycle arrest (HCA) is a noninflammatory alopecic disease affecting various dog breeds, particularly Pomeranian dogs. This disease is probably a hereditary disorder considering the strong breed predisposition. Despite efforts to identify the pathogenesis of this disease, an underlying specific cause is unknown. OBJECTIVE: To identify candidate gene mutations for HCA in Pomeranian dogs. ANIMALS: Four Pomeranian dogs diagnosed with HCA and four unaffected Pomeranian dogs. MATERIALS AND METHODS: Whole blood was used for DNA extraction. Whole-genome sequencing (WGS) was performed, and variants were analysed using a Genome Analysis Toolkit (GATK) and SnpEff. All reads were aligned to the reference genome, Dog10K_Boxer_Tasha. Sanger sequencing was performed to define the complex mutations. RESULTS: A total of 113 variants of mitochondrial DNA were found to be effective gene mutations in the eight dogs. The affected dogs showed significantly increased effective mutations (average 57 variants) compared with unaffected dogs (average eight variants; p < 0.05). There was no significant difference in the number of chromosomal DNA mutations between the two groups. CONCLUSION AND CLINICAL IMPORTANCE: We suggest that an increased number of mitochondrial gene mutations are features for HCA in Pomeranian dogs.

Evaluation of cold atmospheric microwave plasma on skin physiological parameters and tolerability in dogs.

BACKGROUND: Cold atmospheric microwave plasma (CAMP) is a promising therapeutic option for treating skin infections and wounds. Changes in biophysical skin parameters and the tolerability in dogs after applying CAMP is unknown. OBJECTIVE: This study aimed to evaluate the in vivo effects of CAMP on skin biophysical parameters [hydration, transepidermal water loss (TEWL) and surface temperature] and tolerability in dogs. ANIMALS: Twenty client-owned dogs with normal skin. MATERIALS AND METHODS: Cold atmospheric microwave plasma treatment was performed for 30 s and 1, 2 and 4 min, respectively, at different sites of normal canine skin in the inguinal area. Hydration, TEWL and surface temperature were measured five, three and three times, respectively, before and after CAMP application. After treatment, pain and adverse effects were evaluated using a modified Melbourne Pain Scale and the modified short form Glasgow Composite Measure Pain Scale (modified CMPS-SF). RESULTS: Transepidermal water loss values significantly decreased with 4 min of treatment, and hydration decreased significantly with 2 min of treatment. Temperature increased significantly with increasing treatment time. For other parameters, no significant changes were observed. No significant pain response or adverse effects were observed in most dogs, aside from mild erythema in the treatment area after 4 min. CONCLUSION AND CLINICAL SIGNIFICANCE: Cold atmospheric microwave plasma treatment was well-tolerated and did not significantly change canine skin biophysical parameters. CAMP achieves basic recommendations for safe use and is a potential therapeutic option for various skin diseases in dogs.

Contexte - Le CAMP (Cold Atmospheric Microwave Plasma) est une option thérapeutique prometteuse pour le traitement des infections cutanées et des plaies. Les modifications des paramètres biophysiques de la peau et la tolérance chez les chiens après l'application de CAMP sont inconnues. Objectif - Cette étude visait à évaluer les effets in vivo du CAMP sur les paramètres biophysiques de la peau [hydratation, perte d'eau transépidermique (TEWL) et température de surface] et la tolérance chez le chien. Animaux - Vingt chiens de propriétaires à peau normale. Matériels et méthodes - Le traitement CAMP a été effectué pendant 30 s et 1, 2 et 4 min, respectivement, sur différents sites de peau canine normale dans la région inguinale. L'hydratation, la TEWL et la température de surface ont été mesurées cinq, trois et trois fois, respectivement, avant et après l'application de CAMP. Après le traitement, la douleur et les effets indésirables ont été évalués à l'aide d'une échelle de douleur de Melbourne modifiée et de la forme courte modifiée de l'échelle de mesure de la douleur composite de Glasgow (CMPS-SF modifiée). Résultats - Les valeurs de TEWL ont diminué de manière significative après 4 minutes de traitement et l'hydratation a diminué de manière significative après 2 minutes de traitement. La température a augmenté de manière significative avec l'augmentation du temps de traitement. Pour les autres paramètres, aucun changement significatif n'a été observé. Aucune réponse significative à la douleur ni aucun effet indésirable n'ont été observés chez la plupart des chiens, à l'exception d'un léger érythème dans la zone de traitement après 4 minutes. Conclusion et signification clinique - Le traitement CAMP a été bien toléré et n'a pas modifié de manière significative les paramètres biophysiques de la peau canine. CAMP répond aux recommandations de base pour une utilisation sûre et constitue une option thérapeutique potentielle pour diverses maladies de la peau chez les chiens.

Introducción- el plasma de microondas atmosférico frío (CAMP) es una opción terapéutica prometedora para el tratamiento de infecciones y heridas de la piel. Se desconocen los cambios en los parámetros biofísicos de la piel y la tolerabilidad en perros después de aplicar CAMP. Objetivo- este estudio tuvo como objetivo evaluar los efectos in vivo de CAMP en los parámetros biofísicos de la piel [hidratación, pérdida de agua transepidérmica (TEWL) y temperatura superficial] y la tolerabilidad en perros. Animales - Veinte perros de propietarios particulares con piel normal. Materiales y métodos - El tratamiento CAMP se realizó durante 30 s y 1, 2 y 4 min, respectivamente, en diferentes sitios de piel canina normal en el área inguinal. La hidratación, el TEWL y la temperatura superficial se midieron cinco, tres y tres veces, respectivamente, antes y después de la aplicación de CAMP. Después del tratamiento, el dolor y los efectos adversos se evaluaron mediante una escala de dolor de Melbourne modificada y la escala de dolor de medida compuesta de Glasgow de forma abreviada modificada (CMPS-SF modificada). Resultados- los valores de TEWL disminuyeron significativamente con 4 min de tratamiento y la hidratación disminuyó significativamente con 2 min de tratamiento. La temperatura aumentó significativamente con el aumento del tiempo de tratamiento. Para otros parámetros no se observaron cambios significativos. En la mayoría de los perros no se observaron reacciones significativas de dolor ni efectos adversos, aparte de un leve eritema en el área de tratamiento después de 4 min. Conclusión y significado clínico- el tratamiento con CAMP fue bien tolerado y no cambió significativamente los parámetros biofísicos de la piel canina. CAMP obtuvo recomendaciones básicas para un uso seguro y es una opción terapéutica potencial para diversas enfermedades de la piel en perros.

Contexto - O plasma frio atmosférico de micro-ondas (CAMP) é uma opção terapêutica promissora para o tratamento de infecções cutâneas e feridas. Não se sabe a respeito das alterações nos parâmetros biofísicos da pele e a tolerabilidade de cães após a aplicação de CAMP. Objetivo - Este estudo tem como objetivo avaliar os efeitos in vivo de CAMP nos parâmetros biofísicos da pele [hidratação, perda de água transepidérmica (TEWL) e temperatura da superfície] e a tolerabilidade em cães. Materiais e métodos - O tratamento com CAMP foi realizado por 30s e 1, 2 e 4 min, respectivamente, em diferentes locais da pele canina normal na região inguinal. Hidratação, TEWL e temperatura da superfície foram medidas cinco, três e três vezes, respectivamente, antes e após a aplicação do CAMP. Após o tratamento, a dor e os efeitos adversos foram avaliados usando uma escala de dor de Melbourne modificada e a escala de medida composta de dor de Glasgow modificada (CMPS-SF modificada). Resultados - Os valores de TEWL reduziram significativamente com o tratamento de 4 min, e a hidratação reduziu significativamente com dois minutos de tratamento. A temperatura aumentou significativamente com o aumento do tempo de tratamento. Não foram observadas alterações significativas para outros parâmetros. Não se observou uma resposta de dor significativa ou efeitos adversos na maioria dos cães, além de eritema leve na área tratada após 4 min. Conclusão e significância clínica - O tratamento com CAMP foi bem tolerado e não alterou significativamente os parâmetros biofísicos da pele canina. CAMP requer recomendações básicas de segurança na sua utilização e é uma opção terapêutica potencial para várias dermatopatias em cães.

In vitro antifungal activity of cold atmospheric microwave plasma and synergistic activity against Malassezia pachydermatis when combined with chlorhexidine gluconate.

BACKGROUND: The antifungal efficacy of cold atmospheric microwave plasma (CAMP) against Malassezia pachydermatis has not been to be evaluated. OBJECTIVE: To examine the antifungal effects of CAMP against M. pachydermatis and its synergistic effects with chlorhexidine gluconate (CHX). METHODS: A M. pachydermatis isolate was collected from a dog with otitis externa and Malassezia dermatitis at the Seoul National University Veterinary Medical Teaching Hospital. The antifungal effect was determined by applying CAMP to a M. pachydermatis isolate that was incubated for 3 days at 37°C. After 1, 2, 3 and 5 min of application, the efficacy of the plasma treatment was determined according to the number of colony forming units (CFUs). A mixture consisting of inoculum and CHX was applied to evaluate the synergistic effect of the plasma treatment in the same way. RESULTS: The application of CAMP showed significant antifungal effects against M. pachydermatis. The antifungal effect of CAMP was enhanced by an increased exposure time and output power. The application of CAMP with 0.02% and 0.2% CHX resulted in lower survival rates against M. pachydermatis when compared with its sole application at 1 or 2 min. CONCLUSIONS: The study findings demonstrate that CAMP has a potential as a new antifungal option for M. pachydermatis and has synergistic antifungal effects with CHX in vitro. Clinical applications for CAMP are necessary to assess the antifungal efficacy for patients.

In vitro antibacterial and antibiofilm effects of cold atmospheric microwave plasma against Pseudomonas aeruginosa causing canine skin and ear infections.

BACKGROUND: Pseudomonas aeruginosa is an opportunist pathogen that causes purulent inflammation in the skin and in the ears of dogs. Among the various virulence factors of P. aeruginosa, biofilms have been reported to result in antibiotic resistance, leading to therapeutic limitations. Cold atmospheric microwave plasma (CAMP) is known to have a high antimicrobial effect, which causes physical cell wall rupture and DNA damage. HYPOTHESIS/OBJECTIVES: The objective of this study was to evaluate the antibacterial and antibiofilm effects of CAMP against planktonic bacteria and the biofilm of P. aeruginosa. METHODS AND MATERIALS: The antibacterial effect of CAMP against P. aeruginosa ATCC10145 and clinical isolates (n = 30) was evaluated using the colony count method. We also assessed the effect of CAMP on biofilm of P. aeruginosa ATCC strain by the colony count method, water-soluble tetrazolium salt (WST) assay and confocal laser scanning microscopy (CLSM). RESULTS: The complete eradication of P. aeruginosa (ATCC strain and clinical isolates) was achieved within 120 s at 50 W, and clinical isolates required 60 s shorter than the ATCC strain for complete eradication at 50 W. We also confirmed the time-dependent bactericidal effect of CAMP at 50 W against ATCC strain biofilm. CONCLUSIONS AND CLINICAL IMPORTANCE: CAMP was effective against both planktonic bacteria and biofilm formation of P. aeruginosa. However, further studies on in vivo efficacy and safety in canine skin and ears are necessary to fully validate its clinical application.

Successful Treatment of Mucocutaneous Lupus Erythematosus in a Dog with Prednisolone, Mycophenolate Mofetil and Tacrolimus.

A 6-year-old, intact male miniature Pinscher dog had erosive lesions on perilabial, peripenial and perianal mucocutaneous areas, which were exacerbated by ulcerations, crusts, with pain while defecating and urinating. The lesions were symmetrical, and no systemic signs were observed. Histopathological evaluation showed parakeratotic hyperkeratosis, ulceration and cell-rich lymphoplasmacytic interface dermatitis with basal keratinocyte apoptosis. Immunohistochemistry revealed strong reaction in the dermoepidermal junction against goat-canine IgG and mild-to-moderate reaction against goat-canine IgA, IgM and C3. Based on these findings, the dog was diagnosed with mucocutaneous lupus erythematosus (MCLE). Oral prednisolone 1 mg/kg twice daily, mycophenolate mofetil (MMF) 18.3 mg/kg twice daily and 0.1% tacrolimus ointment were prescribed as initial treatment. The lesions showed remarkable improvement within 4 weeks, but the dog exhibited polyuria, polydipsia and hepatomegaly with high dosage of prednisolone. Hence, the dosage of prednisolone was gradually tapered for 9 weeks and discontinued, but MMF and tacrolimus were continued. No new lesion or associated side effect was observed while reducing the MMF dose to 10 mg/kg twice daily and with continuous use of tacrolimus ointment after steroid discontinuation. In conclusion, this case report emphasizes the usefulness of MMF and tacrolimus as steroid-sparing agents in the treatment of dogs with MCLE. To the best of our knowledge, this is the first case report of MCLE that was successfully managed long-term with MMF and tacrolimus.