Ear Skin Research Articles

Metabolipidomic changes induced by dermal nickel penetration determined in an ex vivo porcine ear skin model.

Nickel is one of humans' most prevalent triggers of allergic contact dermatitis. However, the underlying mechanisms of this allergy still need to be fully understood. One aspect that has yet to be explored is the direct impact of common metal allergens on the skin's metabolites and lipids composition. Our study employed matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) to analyze spatially resolved metabolic alterations induced by nickel exposure. Cross-sections of ex vivo porcine ear skin exposed to increasing nickel (II) ion concentrations (17-167 μg/cm2) were measured with an AP-SMALDI5 AF ion source coupled to Q Exactive HF Orbitrap mass spectrometer. Additionally, the penetration of nickel ions into the skin was observed through its pink complexation with dimethylglyoxime under light microscopy. For nickel ion concentrations up to 84 μg/cm2, most nickel ions were stopped within the stratum corneum, while only a very small proportion of nickel ions penetrated the viable epidermis and dermis. Stratum corneum locations with high nickel ion concentrations showed a decrease in arginine and ceramides. Furthermore, several phosphatidylcholine and sphingomyelin species were found to be downregulated in the viable epidermis and dermis due to the nickel exposure. Nickel penetrates at a trace level into the viable skin and induces severe metabolomic and lipidomic changes in the stratum corneum, epidermis, and dermis, indicating a change in the skin (barrier) function. These findings contribute to a deeper understanding of nickel-induced skin allergies and provide a solid foundation for further research.

Bay 11-7082, an NF-κB Inhibitor, Prevents Post-Inflammatory Hyperpigmentation Through Inhibition of Inflammation and Melanogenesis.

Post-inflammatory hyperpigmentation (PIH) is a very common disorder of cutaneous hyperpigmentation, which poses a persistent management challenge in the fields of dermatology and esthetics. This study was designed to explore the anti-melanogenic and anti-inflammatory effects of Bay 11-7082, an NF-κB inhibitor, using small-molecule screening, to determine its potential application for PIH prevention. The molecular mechanisms were investigated invitro and exvivo in epidermis-humanized mice using melanin content, RT-PCR, and immunoblotting. Bay 11-7082 suppressed proinflammatory cytokines and ameliorated 2,4-dinitrofluorobenzene (DNFB)-induced contact dermatitis on day 15. The suppression of melanin synthesis by Bay 11-7082 was attributed to the reduction of MITF, which was induced by extracellular signal-regulated kinase activation. Bay 11-7082 reduced epidermal melanin accumulation in UVB-stimulated exvivo human epidermis as well as in the ear and tail skin of K14-stem cell factor (SCF) transgenic mice. Topical administration of Bay 11-7082 improved PIH on day 35 in the post-DNFB dorsal skin of K14-SCF transgenic mice. In conclusion, Bay 11-7082 can be considered a promising candidate for the development of a preventive topical agent for PIH.

Quantifying skin permeation of a novel metformin lotion using modified hydrophilic interaction liquid chromatography.

Aim: This study aimed to quantify the permeation of metformin (Met) lotion through pig ear skin using high-performance liquid chromatography, specifically hydrophilic interaction liquid chromatography (HILIC), to separate Met from biological contaminants and effectively measure its permeation through skin similar to human skin.Materials & methods: A Franz cell permeation assay was used to assess the permeation kinetics of 6% Met lotion through pig ear skin. Samples were collected at various time points and prepared for high-performance liquid chromatography analysis by removing large biological contaminants. The permeated Met was quantified by monitoring its retention time (RT) at 9min using HILIC, with an acidic, polar mobile phase and a normal-phase column.Results: A distinct Met peak with a RT of approximately 9min was observed in the 6% Met lotion, which was absent in the permeation samples from the 0% Met lotion. This peak (RT 9min) was distinct from the 'biological-contaminants' peaks at RT 2-3min and increased linearly over time, reaching 36.8% of the total applied Met at 24h.Conclusion: These findings demonstrate that the HILIC method effectively separates Met from biological components in pig ear skin, allowing accurate quantification of Met despite the presence of skin lipids and proteins.

Swellable Biopolymer Composite Microneedle Patch for Pain-Free Collection of Interstitial Fluid to Analyze Multiple Biomolecules.

Sampling of interstitial fluid (ISF) using microneedle (MN) patch offers a pain-free minimally invasive alternative to syringe needle-based blood sample collection. However, there is a challenge in the development of MN patch that provides swelling behavior with sufficient mechanical strength for skin penetration. Here, we report fabrication of MN patch made of biopolymer composite containing iota-carrageenan, gelatin, and polyethylene glycol. Calcium chloride was used as a crosslinker to improve mechanical strength. MN patch was characterized for integrity, swelling behavior, mechanical strength, aspiration of fluid from agarose gel, and the excised porcine ear skin. An array of 361 MNs was able to aspirate 36 ± 5 and 14 ± 1 μL fluid after application in agarose gel matrix and the ex vivo porcine skin model, respectively. MN patch applied in vivo rat model for 30 min resulted in the collection of ISF containing 267 ± 128 mg/dL, 24 ± 13 mg/dL, and 0.6 ± 0.4 mIU/mL of glucose, uric acid and thyroid stimulating hormone (TSH), respectively. The concentration of glucose, uric acid, and TSH in rat blood was found to be 199 ± 47 mg/dL, 8.4 ± 6 mg/dL, and 1.1 ± 0.6 mIU/mL at the same time. Furthermore, MN patch applied on the forearm of 10 healthy human volunteers for 30 min was able to aspirate 32 ± 14 μL of ISF. The concentration of glucose, uric acid, and TSH determined from ISF samples of human volunteers was 64 ± 25 mg/dL, 4.2 ± 4.1 mg/dL, and 0.16 ± 0.08 mIU/mL, respectively. The visual analogue scale (VAS) pain score after MN application was lower compared with hypodermic syringe needle insertion. Taken together, biopolymer composite-based swellable MN patch can be developed for collection of ISF for simultaneous determination of multiple biomolecules in a minimally invasive manner.

Application of validated size-exclusion chromatography method for physicochemical characterization of topical gel formulation of deferoxamine conjugated with PEGylated carbon nanoparticles

The focus of current research work was to develop and validate size-exclusion chromatography method and develop and evaluate gel formulation of deferoxamine conjugated with PEGylated carbon nanoparticles (DEF-PEG-CNP) for topical delivery. Size-exclusion chromatography-based method was validated as per ICH guidelines. Effect of Carbopol® 974P and Transcutol® on the nanoparticles’ permeation was studied by 3-level full factorial design of experiment. Gel formulations were characterized for viscosity, cohesive and adhesive force by texture analyzer, and drug permeation through pig ear and human skin. The analytical method was specific as no interference from solvent or excipients were observed and met preset criteria of validation with limit of quantification of 0.24 ± 0.00 μg/mL. The nanoparticles permeation, steady state flux, and retained drug were statistically (p < 0.05) affected by Carbopol® 974P and Transcutol® percentage in the gel formulations. The permeation, steady state flux, and retained nanoparticles from the gel formulations varied from 23.2 ± 2.5 % to 70.9 ± 113.3 %, 0.8 ± 0.3 to 6.6 ± 2.1 μg/cm2.h, and 5.6 ± 0.3 to 38.8 ± 8.8 µg/g, respectively. Permeation of the nanoparticles was 1.9 folds higher in pig skin compared to human skin. Immunofluorescence detected successful permeation of DEF-PEG-CNP particles into skin. In conclusion, the analytical method can quantify the nanoparticles from the gel formulation without interference, and gel formulation of the nanoparticles can permeate across the skin.

3D Printed Microneedles for the Transdermal Delivery of NAD+ Precursor: Toward Personalization of Skin Delivery.

3D printing of microneedles (μNDs) for transdermal therapy has the potential to enable patient personalization based on the target disease, site of application, and dosage requirements. To convert this concept to reality, it is necessary that the 3D printing technology can deliver high resolution, an affordable cost, and large print volumes. With the introduction of benchtop 4K and 8K 3D printers, it is now possible to manufacture medical devices like μNDs at sufficient resolution and low cost. In this research, we systematically optimized the 3D printing design parameters such as resin viscosity, print angle, layer height, and curing time to generate customizable μNDs. We have also developed an innovative 3D coating microtank device to optimize the coating method. We have applied this to the development of novel μNDs to deliver an established NAD+ precursor molecule, nicotinamide mononucleotide (NMN). A methacrylate-based polymer photoresin (eSun resin) was diluted with methanol to adjust the resin viscosity. The 3D print layer height of 25 μm yielded a smooth surface, thus reducing edge-ridge mismatches. Printing μNDs at 90° to the print platform yielded 84.28 ± 2.158% (n = 5) of the input height thus increasing the tip sharpness (48.52 ± 10.43 μm, n = 5). The formulation containing fluorescein (model molecule), sucrose (viscosity modifier), and Tween-20 (surface tension modifier) was coated on the μNDs using the custom designed microtank setup, and the amount deposited was determined fluorescently. The dye-coated μND arrays inserted into human skin (in vitro) showed a fluorescence signal at a depth of 150 μm (n = 3) into the skin. After optimization of the 3D printing parameters and coating protocol using fluorescein, NMN was coated onto the μNDs, and its diffusion was assessed in full-thickness human skin in vitro using a Franz diffusion setup. Approximately 189 ± 34.5 μg (5× dipped coated μNDs) of NMN permeated through the skin and 41.2 ± 7.53 μg was left in the skin after 24 h. Multiphoton microscopy imaging of NMN-coated μND treated mouse ear skin ex vivo demonstrated significantly (p < 0.05) increased free-unbound NADPH and reduced fluorescence lifetime of NADPH, both of which are indicative of cellular metabolic rates. Our study demonstrates that low-cost benchtop 3D printers can be used to print high-fidelity μNDs with the ability to rapidly coat and release NMN which consequently caused changes in intracellular NAD+ levels.

Identification of Reference Gene for Quantitative Gene Expression in Early-Term and Late-Term Cultured Canine Fibroblasts Derived from Ear Skin.

Fibroblasts are cells that reside within the fibrous or loose connective tissues of most mammalian organs. For research purposes, fibroblasts are often subjected to long-term culture under defined conditions, during which their properties can significantly change. It is essential to understand and document these changes to obtain reliable outcomes. For the quantification of specific gene expressions, the most reliable and widely used technique is quantitative real-time polymerase chain reaction (qRT-PCR). Here, we assessed the impact of a reference gene's stability on a qRT-PCR analysis of long-term cultured canine skin fibroblasts. After successfully isolating the fibroblasts from canine skin tissues, they were cultured and evaluated for proliferation and β-galactosidase activity at different passage numbers. With extended culture, the fibroblasts showed a long doubling time and elevated β-galactosidase activity. Using three widely used algorithms, geNorm, Normfinder, and Bestkeeper, we identified HPRT1, YWHAZ, and GUSB as the most stable reference genes for both early- and late-passage fibroblasts. Conventional reference genes such as GAPDH were found to be less stable than those genes. The normalization of Vimentin by the stable genes showed statistical differences, whereas normalization by an unstable gene did not. Collectively, this study indicates that using stable reference genes is essential for accurately and reliably measuring gene expression in both early- and late-passage fibroblasts. These findings provide valuable insights into internal controls for gene expression studies and are expected to be utilized for analyzing gene expression patterns in molecular biology research.

Chondroitin Sulfate for Cartilage Regeneration, Administered Topically Using a Nanostructured Formulation.

In the pharmaceutical sector, solid lipid nanoparticles (SLN) are vital for drug delivery incorporating a lipid core. Chondroitin sulfate (CHON) is crucial for cartilage health. It is often used in osteoarthritis (OA) treatment. Due to conflicting results from clinical trials on CHON's efficacy in OA treatment, there has been a shift toward exploring effective topical systems utilizing nanotechnology. This study aimed to optimize a solid lipid nanoparticle formulation aiming to enhance CHON permeation for OA therapy. A 3 × 3 × 2 Design of these experiments determined the ideal parameters: a CHON concentration of 0.4 mg/mL, operating at 20,000 rpm speed, and processing for 10 min for SLN production. Transmission electron microscopy analysis confirmed the nanoparticles' spherical morphology, ensuring crucial uniformity for efficient drug delivery. Cell viability assessments showed no significant cytotoxicity within the tested parameters, indicating a safe profile for potential clinical application. The cell internalization assay indicates successful internalization at 1.5 h and 24 h post-treatment. Biopharmaceutical studies supported SLNs, indicating them to be effective CHON carriers through the skin, showcasing improved skin permeation and CHON retention compared to conventional methods. In summary, this study successfully optimized SLN formulation for efficient CHON transport through pig ear skin with no cellular toxicity, highlighting SLNs' potential as promising carriers to enhance CHON delivery in OA treatment and advance nanotechnology-based therapeutic strategies in pharmaceutical formulations.

Physiologically-based pharmacokinetic model of in vitro porcine ear skin permeation for drug delivery research.

In silico techniques, such as physiologically based pharmacokinetic modeling (PBKP), are recently gaining importance. Computational methods in drug discovery and development and the generic drugs industry enhance research effectiveness by saving time and money and avoiding ethical issues. One key advantage is the ability to conduct toxicology studies without risking harm to living beings. This study aimed to repurpose the multi-phase multi-layer mechanistic dermal absorption (MPML MechDermA) PBPK model for simulation permeation through porcine ear skin under in vitro conditions. The work was divided into four steps: (1) the development of a pig ear skin model based on a previously collected dataset; (2) testing the model's ability to discriminate permeation between pig ear, human abdomen, and human back skin; (3) development of a caffeine permeation model; and (4) testing the caffeine model's performance against in vitro generated data sourced from the scientific literature. Data from 31 manuscripts were used for the development of the pig skin model. Based on these data, values specific to pig skin were found for 22 parameters of the MPML MechDermA model. The model was able to discriminate permeation between pig and human skin. A caffeine model was developed and used to simulate seven experiments identified in the literature. The model's performance was assessed by comparing simulated to observed results. Based on a visual check, all simulations were considered acceptable, whereas three out of seven experiments met the twofold difference criterion. The variability of the experimental data was considered the biggest challenge for reliable model assessment.

Sustained release microneedle patch for pronounced systemic delivery of doxazosin mesylate

Introduction: Microneedle patch is one of the fascinating drug delivery approaches that offers low invasiveness and a painless physical application to enhance the delivery of micro and macro-molecules into the skin. Methods: Variable contents of chitosan and polyvinyl alcohol were used for the development of doxazosin mesylate containing sustained release microneedle patches via solvent casting technique. The prepared patches were evaluated for microscopic evaluation, mechanical strength, drug loading (%) and Fourier transform infrared spectroscopy (FTIR) etc. The skin penetration study was performed by using pig ear skin and results were captured through confocal microscopy. Ex-vivo release study and pharmacokinetic evaluation were also performed. Results: Sharp needle tips with a height of 600µm and a base of 200µm were confirmed through microscopic examination. Optimized formulation (SRF-6) exhibited loading of 92.11% doxazosin mesylate with appreciable strength up to 1.94N force. Ex-vivo release studies revealed 87.24% release within 48 hours. Moreover, the pharmacokinetic parameters in case of optimized patch formulation (SRF-6) were markedly improved i.e. MRT (19.46 h), AUC (57.12 μg.h /mL), Cmax (2.16 µg /mL), tmax (10.10 h) and t1/2 (6.32 h) as compared to commercially available tablet. Biocompatibility of the developed patches was validated from skin irritation studies. Conclusion: Results confirmed the successful fabrication of microneedle patch having sufficient strength and effective penetration ability into the skin to ensure controlled release of incorporated drug for the intended duration. It can be employed as an efficient carrier system for other therapeutics those are prone to bioavailability issues due to first pass effect after their oral administration.

Hydrophobic binary mixtures containing amphotericin B as lipophilic solutions for the treatment of cutaneous leishmaniasis

Cutaneous leishmaniasis, caused by Leishmania parasites, requires treatments with fewer side effects than those currently available. The development of a topical solution based on amphotericin B (AmB) was pursued. The considerable interest in deep eutectic solvents (DESs) and their remarkable advantages inspired the search for a suitable hydrophobic excipient. Various mixtures based on commonly used hydrogen bond donors (HBDs) and acceptors (HBAs) for DES preparations were explored. Initial physical and in-vitro screenings showed the potential of quaternary phosphonium salt-based mixtures. Through thermal analysis, it was determined that most of these mixtures did not exhibit eutectic behavior. X-ray scattering studies revealed a sponge-like nanoscale structure. The most promising formulation, based on a combination of trihexyl(tetradecyl)phosphonium chloride and 1-oleoyl-rac-glycerol, showed no deleterious effects through histological evaluation. AmB was fully solubilized at concentrations between 0.5 and 0.8 mg·mL−1, depending on the formulation. The monomeric state of AmB was observed by circular dichroism. In-vitro irritation tests demonstrated acceptable viability for AmB-based formulations up to 0.5 mg·mL−1. Additionally, an ex-vivo penetration study on pig ear skin revealed no transcutaneous passage, confirming AmB retention in healthy, unaffected skin.

Local keratinocyte-nociceptor interactions enhance obesity-mediated small fiber neuropathy via NGF-TrkA-PI3K signaling axis.

The pathology of diabetic small fiber neuropathy, characterized by neuropathic pain and axon degeneration, develops locally within the skin during the stages of obesity and pre-diabetes. However, the initiation and progression of morphological and functional abnormalities in skin sensory nerves remains elusive. To address this, we utilized ear skin from mice with diet-induced obesity (DIO), the mouse models for obesity and pre-type 2 diabetes. We evaluated pain-associated wiping behavior and conducted ex vivo live Ca2+ imaging of the DIO ear skin to detect sensory hypersensitivity. Our findings reveal sensory hypersensitivity in skin nociceptive axons followed by axon degeneration. Further mechanistic analysis identified keratinocytes as a major source of nerve growth factor (NGF) in DIO skin, which locally sensitizes nociceptors through NGF-mediated signaling. Indeed, the local inactivation of NGF and its receptor TrkA-mediated downstream signaling, including the phosphoinositide 3-kinases (PI3K) pathway, suppresses sensory hypersensitivity in DIO skin. Thus, targeting these local interactions between keratinocytes and nociceptors offers a therapeutic strategy for managing neuropathic pain, avoiding the adverse effects associated with systemic interventions.

Daily Supplementation with Bifidobacterium longum KACC91563 Alleviates Allergic Contact Dermatitis in an Animal Model.

Allergic contact dermatitis (ACD) is the most common chronic inflammatory skin disease (or immune-mediated disease), causing disruption to our psychological condition and life quality. In this study, the therapeutic properties of probiotic Bifidobacterium longum (B. longum) was investigated by using an ACD-induced animal model. For ACD induction, BALB/c mice ear and dorsal skin were sensitized with 240 µL of 1% (w/v) 2,4-dinitrochlorobenzene (DNCB) twice (3-day intervals). After a week of the first induction, the mice were re-sensitized by painting on their dorsal skin and ear with 0.4% (w/v) DNCB for a further three times (once per week). Before the ACD induction of 2 weeks and throughout the trial period, the BALB/c mice were supplemented daily with 1 mL of 1.0 × 109 CFU or 5.0 × 109 CFU B. longum using an intragastric gavage method. The ACD-induced mice without B. longum supplementation were used as a control. Results show that B. longum supplementation significantly alleviated ACD symptoms (e.g., ear swelling, epidermal damage) and immune response (e.g., reduced immune cell recruitment, serum IgE level, and cytokine production). The therapeutic efficiency of B. longum increased as the supplementation dose increased. Thus, daily supplementation with 5.0 × 109 CFU probiotic B. longum could be an effective method for the prevention and treatment of ACD.

LPS exacerbates TRPV4-mediated itch through the intracellular TLR4-PI3K signalling.

Pruritus is often accompanied with bacterial infections, but the underlying mechanism is not fully understood. Although previous studies revealed that lipopolysaccharides (LPS) could directly activate TRPV4 channel and TRPV4 is involved in the generation of both acute itch and chronic itch, whether and how LPS affects TRPV4-mediated itch sensation remains unclear. Here, we showed that LPS-mediated TRPV4 sensitization exacerbated GSK101-induced scratching behaviour in mice. Moreover, this effect was compromised in TLR4-knockout mice, suggesting LPS acted through a TLR4-dependent mechanism. Mechanistically, LPS enhanced GSK101-evoked calcium influx in mouse ear skin cells and HEK293T cells transfected with TRPV4. Further, LPS sensitized TRPV4 channel through the intracellular TLR4-PI3K-AKT signalling. In summary, our study found a modulatory role of LPS in TRPV4 function and highlighted the TLR4-TRPV4 interaction in itch signal amplification.

Delivery of small hydrophilic molecules across the stratum corneum: Identification of model systems and parameters to study topical delivery of free amino acids

Free amino acids (FAAs) constitute the largest component (∼40 %) of the so-called natural moisturizing factors of the skin. Their level declines in dry skin conditions and one strategy to overcome this problem may involve the topical delivery of FAAs through appropriate strategy. The objective of the present study was therefore to identify alternative skin models and study the corneocyte-water partition coefficients (KCOR/W) and permeation coefficient (KP) of 18 FAAs. The KCOR/W was studied using standard protocols and the permeation studies were conducted using Franz diffusion cell. The results indicate that the FAAs have high partitioning behavior to the corneocytes. The KCOR/W values of the human COR and that of pig ear skin were better correlated with each other than that of keratin isolated from chicken feathers. The presence of lipid in the stratum corneum (SC), initial concentration of the FAAs, and permeation enhancers affect the KCOR/W. The FAAs have low permeation into the SC which suggests the need for permeation enhancers in designing dosage form containing these compounds. Even though the investigated mathematical models show good prediction of the Kp values, better prediction could be obtained by considering factors such as the possible entrapment of the FAAs by the CORs.

Effect of gelling agents on properties of coumestrol-loaded nanoemulsions – Formulation, skin retention/permeation, antioxidant and skin photoprotective activities

Coumestrol is an isoflavonoid that exhibits antioxidant activity. In this study, hydrogels containing coumestrol-loaded nanoemulsions were designed to protect the skin against UVA/UVB light. The systems investigated were coumestrol-loaded nanoemulsions obtained by spontaneous emulsification procedure formulated in semisolid hydrogels composed of ionic (Carbopol 940®), cationic (chitosan) or non-ionic (hydroxyethylcellulose) gelling materials. Transmission electron microscopy images of formulations showed oil droplets exhibiting 200–300 nm. Hydrogels displayed non-Newtonian shear-thinning behavior. Coumestrol content in formulations remained close to 100 % (1 mg/mL) over time (up to 90 days). Regardless of the gelling material used, a lowering of coumestrol release rate was observed, in comparison with the release from nanoemulsion. The permeation and retention of coumestrol in porcine ear skin was evaluated using Franz-type diffusion cells. A higher retention of coumestrol in the epidermis was noticed (20 μg/g) from Carbopol 940-based formulation after 8 h of kinetics. No histological evidence of tissue damage was observed. The nanoemulsion containing coumestrol thickened with Carbopol 940® was selected for further studies. The formulation exhibited antioxidant potential as estimated by TRAP and TAR assays. The skin photoprotective effect was evidenced by means of TBARS, protein carbonylation, and protein thiol content experiments. Overall, our research suggests that the formulation is promising to protect skin against oxidative damage from UVA/UVB light.

Next-generation Sequencing of MicroRNA in Acquired Middle Ear Cholesteatoma.

The pathophysiology of cholesteatoma is not precisely understood, and research on the associated microRNAs (miRNAs) is also deficient. We demonstrated the expression of miRNA in normal skin and middle ear cholesteatoma by next-generation sequencing (NGS) technology. The profiles of miRNA and relevant molecular interaction pathways were investigated. Case-control experimental study. Middle ear cholesteatoma and post-auricular skin tissue specimens were collected from 13 adult patients. Total RNA was extracted, and miRNA expression profiles were analyzed by NGS technology. Functional gene classification to predict target genes and relevant biological pathways was performed using DIANA-microT-CDS and the Kyoto Encyclopedia Gene and Genome database (KEGG) pathways. The expression of 2588 miRNAs from middle ear cholesteatoma and skin tissue samples was analyzed. The expression of 76 upregulated and 128 downregulated miRNAs was identified in the cholesteatoma samples compared to normal skin (FC ≥2 and p < 0.05). Ninety-nine differentially expressed miRNAs (FC ≥4 and p < 0.05) were used to explore the biological pathways involved in the etiopathogenesis of cholesteatoma. The most predicted pathway in cholesteatoma in the upregulated miRNA group was the ErbB signaling pathway and it was extracellular matrix (ECM)-receptor interaction in the downregulated miRNA group. This was the first study investigating small miRNAs in human acquired cholesteatoma using NGS technique. We were able to identify new miRNAs and pathways related to cholesteatoma. The results of this study are expected to be helpful in revealing new pathophysiologies of cholesteatoma. N/A Laryngoscope, 134:4374-4382, 2024.

Genomic regions, candidate genes, and pleiotropic variants associated with physiological and anatomical indicators of heat stress response in lactating sows

BackgroundHeat stress (HS) poses significant threats to the sustainability of livestock production. Genetically improving heat tolerance could enhance animal welfare and minimize production losses during HS events. Measuring phenotypic indicators of HS response and understanding their genetic background are crucial steps to optimize breeding schemes for improved climatic resilience. The identification of genomic regions and candidate genes influencing the traits of interest, including variants with pleiotropic effects, enables the refinement of genotyping panels used to perform genomic prediction of breeding values and contributes to unraveling the biological mechanisms influencing heat stress response. Therefore, the main objectives of this study were to identify genomic regions, candidate genes, and potential pleiotropic variants significantly associated with indicators of HS response in lactating sows using imputed whole-genome sequence (WGS) data. Phenotypic records for 18 traits and genomic information from 1,645 lactating sows were available for the study. The genotypes from the PorcineSNP50K panel containing 50,703 single nucleotide polymorphisms (SNPs) were imputed to WGS and after quality control, 1,622 animals and 7,065,922 SNPs were included in the analyses.ResultsA total of 1,388 unique SNPs located on sixteen chromosomes were found to be associated with 11 traits. Twenty gene ontology terms and 11 biological pathways were shown to be associated with variability in ear skin temperature, shoulder skin temperature, rump skin temperature, tail skin temperature, respiration rate, panting score, vaginal temperature automatically measured every 10 min, vaginal temperature measured at 0800 h, hair density score, body condition score, and ear area. Seven, five, six, two, seven, 15, and 14 genes with potential pleiotropic effects were identified for indicators of skin temperature, vaginal temperature, animal temperature, respiration rate, thermoregulatory traits, anatomical traits, and all traits, respectively.ConclusionsPhysiological and anatomical indicators of HS response in lactating sows are heritable but highly polygenic. The candidate genes found are associated with important gene ontology terms and biological pathways related to heat shock protein activities, immune response, and cellular oxidative stress. Many of the candidate genes with pleiotropic effects are involved in catalytic activities to reduce cell damage from oxidative stress and cellular mechanisms related to immune response.

Anatomical Vascular Differences and Leishmania-Induced Vascular Morphological Changes Are Associated with a High Parasite Load in the Skin of Dogs Infected with Leishmania infantum.

Canine visceral leishmaniasis (CVL), caused by the protozoan Leishmania infantum, affects several organs, including the skin. Dogs are considered the major domestic reservoir animals for leishmaniasis, and through their highly parasitized skin, they can serve as a source of infection for sandfly vectors. Therefore, studies of the skin parasite-host relationship can contribute to the understanding of the infectious dissemination processes of parasites in the dermis and help to identify targets for diagnosis and treatment. Thus, the aim of this study was to evaluate the association of anatomical vascular differences and Leishmania-induced vascular morphological changes with clinical signs and parasite load by analyzing the ear and abdominal skin from dogs naturally infected with L. infantum. Paired samples of ear and abdominal skin from L. infantum-positive dogs (n = 26) were submitted for histological and immunohistochemistry analyses. The ear skin samples showed a more intense and more diffusely distributed granulomatous inflammatory reaction, a higher number and larger diameter of blood vessels, increased parasite load, higher expression of VEGF+ (vascular endothelial growth factor) and MAC 387+ (calprotectin) recently infiltrating cells, and more intense collagen disruption compared to the abdominal skin samples. Intracellular amastigotes were observed in blood vessels and inside endothelial cells and were diffusely distributed throughout the dermis in the ear skin samples. The NOS2/MAC387+ cell ratio was lower in the ear skin samples than in those of the abdomen, suggesting that in the ear dermis, the inflammatory infiltrate was less capable of producing NO and thereby control the parasite load. Together, these findings indicate how parasites and immune cells are distributed in the skin and suggest an important role for dermal vascularization in cellular influx and thereby in parasite dissemination through the skin of naturally infected dogs.

Assessing children's outdoor thermal comfort with facial expression recognition: An efficient approach using machine learning

In this study, children's physiological indices and facial expressions during activities with distinct intensities in outdoor open spaces were real-timely measured. Correspondingly, meteorological measurements and questionnaire surveys were conducted to explore change laws of children's physiological feedback, facial expressions, and subjective perception. Then, a predictive model of children's outdoor thermal sensations was built with the coupling relations of physiological indices and facial expressions. Results showed that the skin temperature on the face tended to increase as thermal stress increased but gradually declined with increasing activity intensity. Mean ear skin temperature (Tear) was the most sensitive, followed by mean cheek temperature (Tcheek). When conducting moderate and vigorous intensity activities, the heart rate (HR) increased significantly within 5 min of exercise and then stabilized. The HR increase in vigorous intensity activities was greater than that during moderate intensity activities. With an increase in thermal stress and activity intensity, the proportion of negative emotions (NegEmo) in facial expressions gradually increased. Sadness and disgust expressions both increased significantly, followed by fear and anger. Thus, Tear and NegEmo expressed facially were important indices to predict children's outdoor thermal sensation. Finally, the predictive model for children's outdoor thermal sensation based on facial expressions, Tear, and HR allowed for non-invasive data collection was built integrating machine learning, and its accuracy reached as high as 97.1 %. Our results propose a faster feedback method for children's thermal perception based on facial expression recognition, which can real-timely monitor thermal comfort for children, ensuring their outdoor thermal health and safety.