Angelica polysaccharide attenuates LPS-induced inflammation response of primary dairy cow claw dermal cells via NF-κB and MAPK signaling pathways.

BACKGROUND: Laminitis, an inflammation of the claw laminae, is one of the major causes of bovine lameness, which can lead to enormous economic losses and animal welfare problems in dairy farms. Angelica polysaccharide (AP) is proved to possess anti-inflammatory properties. But the role of AP on inflammatory response of the claw dermal cells has not been reported. The aim of this study was to investigate the anti-inflammatory effects of AP on lipopolysaccharide (LPS)-induced primary claw dermal cells of dairy cow and clarify the potential mechanisms. In the current research, the primary claw dermal cells were exposed to gradient concentrations of AP (10, 50, 100 µg/mL) in the presence of 10 µg/mL LPS. The levels of cytokines and nitric oxide (NO) were detected with ELISA and Griess colorimetric method. The mRNA expressions of TLR4, MyD88 and chemokines were measured with qPCR. The activation of NF-κB and MAPK signaling pathways was detected with western blotting. RESULTS: The results indicated that AP reduced the production of inflammatory mediators (TNF-α, IL-1ß, IL-6 and NO), downregulated the mRNA expression of TLR4, MyD88 and some pro-inflammatory chemokines (CCL2, CCL20, CXCL2, CXCL8, CXCL10), and suppressed the NF-κB and MAPK signaling pathways evidenced by inhibition of the phosphorylation of IκBα, p65 and ERK, JNK, p38. CONCLUSIONS: Our results demonstrated that AP may exert its anti-inflammatory effects on claw dermal cells of dairy cow by regulating the NF-κB and MAPK signaling pathways.

Effect of Penetration Enhancers on Toenail Delivery of Efinaconazole from Hydroalcoholic Preparations.

The incorporation of permeation enhancers in topical preparations has been recognized as a simple and valuable approach to improve the penetration of antifungal agents into toenails. In this study, to improve the toenail delivery of efinaconazole (EFN), a triazole derivative for onychomycosis treatment, topical solutions containing different penetration enhancers were designed, and the permeation profiles were evaluated using bovine hoof models. In an in vitro permeation study in a Franz diffusion cell, hydroalcoholic solutions (HSs) containing lipophilic enhancers, particularly prepared with propylene glycol dicaprylocaprate (Labrafac PG), had 41% higher penetration than the HS base. Moreover, the combination of hydroxypropyl-ß-cyclodextrin with Labrafac PG further facilitated the penetration of EFN across the hoof membrane. In addition, this novel topical solution prepared with both lipophilic and hydrophilic enhancers was physicochemically stable, with no drug degradation under ambient conditions (25 °C, for 10 months). Therefore, this HS system can be a promising tool for enhancing the toenail permeability and therapeutic efficacy of EFN.

A novel technique to evaluate nail softening effects of different urea formulations.

Urea has been incorporated into several topical ungual formulations to hydrate and soften the nail plate. In this study, we employed various characterization techniques (visual observation, scanning electron microscopy, measurement of thickness, transonychial water loss, nail electrical resistance, and mechanical study) to investigate the effect of urea concentration on the hydration of bovine hoof membranes - an in vitro model of infected human nails. We obtained inconsistent results in the thickness, transonychial water loss, nail electrical resistance, and scanning electron microscopy studies. In the mechanical study using a modified Texture Analyzer method, we reported an inverse and linear correlation between urea concentrations in the formulations and the force required to puncture the treated membrane (R2 = 0.9582, n ≥ 8). As the urea concentration decreased from 4x to 2x, 1x, and 0x % w/w, the puncture force increased significantly from 0.47 ± 0.07 to 0.77 ± 0.07, 0.91 ± 0.09, and 1.33 ± 0.26 N, respectively (p < 0.05). Thus, urea provided a positive softening effect on the membranes and the puncture force could indicate the urea level in topical formulations. In this study, we provided a novel, efficient, and reliable tool to evaluate the hydration level and physical properties of bovine hoof membranes.

Potential of Naftifine Application for Transungual Delivery.

Naftifine is used to treat fungal skin infections as it inhibits dermatophytes, which are the cause of onychomycosis. However, naftifine's ability to permeate the human nail barrier has not been investigated, thus, the antimycotic potential is not clearly established. This work aims to evaluate the effect of penetration enhancing factors on the accumulation of naftifine hydrochloride through human nail clippings. Naftifine polymeric nail lacquers with Eudragit RL100 were developed as a suitable delivery system. Low penetration of naftifine into nail has been determined as less than 10% of applied drug dose accumulated in the nail layers. Incorporation of thioglycolic acid into formulations resulted in increased accumulation of antifungal agent in the nail layers by 100% compared with a control group. Salicylic acid did not effect naftifine accumulation in the human nail. The permeation of naftifine through the nail increased by threefold when the thioglycolic acid-containing formulation was applied and the nail was pretreated with a fractional CO2 laser. Structural changes of the nail barrier, induced by fractional CO2 laser, were visualized by microscopy. The results suggest, that naftifine nail penetration could be significantly increased when physical and chemical enhancing factors are applied.

Ex vivo nail infection as an effective preclinical method for screening of new topical antifungals.

Onychomycosis are fungal nail infections comprising of about 50% of onychopathies and are commonly caused by dermatophytes. The treatment of this dermatomycosis requires a long period of time and is associated with high rates of recurrence. In view of the need to evaluate the antifungal performance of promising preclinical compounds, we developed, in this study, a practical and accessibleex vivo model for establishing a Trichophyton rubrum onychomycosis framework using porcine hooves. This model has as its main advantage the similar structural and three-dimensional characteristics that the porcine hooves have with the human nail. The proposed model allowed to evaluate the antifungal activity of a new antifungal compound and a reference drug (terbinafine), both already incorporated into a nail lacquer for topical use. Treatments with compound 3-selenocyanate-indole (Se4a) and with terbinafine incorporated into this nail lacquer completely inhibited fungal growth, corresponding to the profile of in vitro activity observed against T. rubrum. This study concludes that the ex vivo porcine hoof model is an effective alternative method for preclinical screening of drugs or new topical compounds developed to combat onychomycosis. Further studies are needed to compare the permeability of porcine hooves with human nails permeability.

Efficacy of non-antibiotic treatment options for digital dermatitis on an organic dairy farm.

The objective of this study was to evaluate the efficacy of two non-antibiotic treatment options for digital dermatitis (DD) on an organic certified dairy farm. A randomized clinical trial was conducted using 70 multiparous Holstein cows with an early DD lesion at a USDA certified organic dairy farm in Northern Colorado, USA. Cows were enrolled in the study based on the presence of early DD lesions (scores M1 and M2) and randomly assigned to one of three treatments: (1) topical application of copper sulfate and iodine (CUI); (2) topical application of honey and iodine (HOI); and (3) control subject to no treatment (CON). Cows were evaluated at enrolment and on days 3, 12, 28, and 120 post treatment for pain and lesion size and received a locomotion and a lesion score. Cure was defined as the transition from active to non-active stages (M1/M2 to M0 or M4). The formulations had variable effects on the treatment of DD. The cure rate was numerically higher for CUI on all follow up days. The proportion of cows experiencing pain on d3 after treatment was greater in CON, followed by HOI and CUI. However, this proportion increased in HOI during the follow up period. The CUI group had a greater reduction in lesion size and larger lesions persisted in HOI. Non-antibiotic treatment formulations were partially effective in the treatment of DD in organic dairy cows: The two non-antibiotic formulations resulted in an earlier transition to mature lesions compared with the control group. The CUI combination was the most effective treatment in reducing lesion size, pain, and lameness in affected cows. However, this combination had short-term efficacy, which did not persist throughout the duration of the study. The HOI combination produced only transient reduction in lesion size.

Ciclopirox and Efinaconazole Transungual Permeation, Antifungal Activity, and Proficiency To Induce Resistance in Trichophyton rubrum.

Onychomycosis is a nail fungal infection, mostly caused by dermatophytes. The treatment efficacy is impaired by difficulties of reaching effective drug levels at the site of infection; frequent relapses occur after cessation of antifungal therapy. The aim of the study was to compare two commercial products containing ciclopirox or efinaconazole for antimycotic activity and antifungal drug resistance. A study of permeation and penetration through bovine hoof membranes, as a nail model, was performed to evaluate the antimycotic activity of permeates against clinical isolates of selected fungi, and the frequency of spontaneous in vitroTrichophyton rubrum-resistant strains was assessed by broth microdilution assays. The results suggest that ciclopirox creates a depot in the nail, leading to a gradual release of the drug over time with action on both the nail plate and bed. Conversely, efinaconazole, mildly interacting with nail keratin, mainly exerts its antifungal activity in the nail bed. However, in the case of T. rubrum, the antifungal activities of the drugs in the nail plate seem comparable. Finally, efinaconazole showed a potential for induction of resistance in T. rubrum, which may limit its efficacy over time. Ciclopirox did not show any potential to induce resistance in T. rubrum and appears endowed with a more complete activity than efinaconazole in the management of onychomycosis as the nail keratin is a substrate for the growth of fungal cells, and the availability of drug in large concentration just in the nail bed may not be sufficient to guarantee the complete eradication of pathogens.

Effects of silymarin on p65 NF-κB, p38 MAPK and CYP450 in LPS-induced hoof dermal inflammatory cells of dairy cows.

BACKGROUND: Laminitis is considered as one of the most important causes of hoof lameness in dairy cows, which can lead to enormous economic losses. However, the etiology and pathogenesis of laminitis have not been clarified yet. Besides, it is of great significant to find alternative herbs for the prevention and treatment of dairy hooves to avoid the antibiotic abuse. In this study, the primary hoof dermal cells of dairy cows were isolated, the inflammatory model was induced by LPS, and treated with silymarin to find whether silymarin has protective effect on the inflammatory dermal cells. The viability of dermal cells, the levels of IL-1ß and TNF-α, the degree of p65 NF-κB and p38 MAPK phosphorylation, the expressions of CYP3A4 and CYP1A1 were measured. RESULTS: Hoof dermal cells of dairy cows were successfully isolated and cultured by tissue adherent culture method. Certain concentrations of LPS can increase the levels of IL-1ß and TNF-α, promote the phosphorylation of p65 NF-κB and p38 MAPK, and inhibit the mRNA expressions of CYP3A4 and CYP1A1. The optimal concentration for LPS to establish a hoof dermal cells inflammatory model was 10 µg/mL. Certain concentrations of silymarin can markedly decrease the secretions of IL-1ß and TNF-α, inhibit the phosphorylation of p65 NF-κB and p38 MAPK, and promote the mRNA expressions of CYP3A4 and CYP1A1 in LPS-induced dermal inflammatory model. CONCLUSIONS: LPS can be used for inducing the hoof dermal cells inflammatory model of dairy cows. Silymarin has protective effects on the LPS-induced inflammatory model.

Use of topical local anesthetics to control pain during treatment of hoof lesions in dairy cows.

Hoof pathologies in dairy cows have a major effect on both production and animal welfare. Trimming of excess or diseased hoof tissue is essential for the treatment of many of these conditions. Trimming hoof lesions can cause severe pain, resulting in adverse behavioral responses with risk for animal and human safety. Interventions are usually carried out by nonveterinary technicians in the absence of pain management training. Pain control during trimming is not only an ethical obligation but also allows for better manipulation and more meticulous treatment. The aim of this study was to test the efficacy of Tri-Solfen (Bayer Australia Ltd., Pymble, NSW, Australia), a combination of local anesthetics in a topical gel form, containing lidocaine, bupivacaine, adrenaline, and cetrimide, for the treatment of pain associated with trimming of hoof lesions. Sixty-two Holstein-Frisian cows were selected for trimming at the drying-off period and were visually scored for lameness before entering the chute. After diagnosis of the hoof lesion but before deep trimming was initiated, each animal was randomly distributed to 2 groups: C, usual trimming with no pain control, and T, trimming with a local anesthetic formulation being applied immediately after live corium was exposed. During curative trimming, behavior observation was conducted by 2 observers blind to treatment. In 27 cows, algometry measurements were performed before and after the procedure to assess animal reaction to pressure. Lameness scoring was again performed as the cow left the chute. Nonparametric tests and ANOVA were performed. Results showed that use of the topical anesthetic formulation significantly reduced reaction to trimming and lameness score after trimming when compared with nontreated animals. Algometry values showed increased pressure threshold after application of topical anesthetics. This study suggests that the use of topical local anesthesia with lidocaine and bupivacaine helps reduce pain associated with corrective trimming of severe hoof lesions, enhancing animal welfare and potentially ensuring safety of trimmers.

Ex vivo effects of insulin on the structural integrity of equine digital lamellae.

BACKGROUND: Laminitis has a considerable impact on the equine industry. Endocrinopathic laminitis is the most common form and affected horses often have hyperinsulinaemia due to an underlying metabolic disorder. OBJECTIVES: The aim of this study was to determine if insulin weakens the structural integrity of digital lamellae and to develop an ex vivo model for the study of hyperinsulinaemia-induced lamellar failure. STUDY DESIGN: Ex vivo experiment. METHODS: Biomechanical testing was used to assess the structural integrity of lamellar explants exposed to either medium alone (control) or medium supplemented with insulin. Lamellar explants comprised of hoof wall, lamellar tissue and distal phalanx were harvested from four adult horses with no evidence of inflammatory disease or pre-existing disease of the digit. Following an equilibration period, explants were incubated in medium or medium supplemented with insulin (2.5 µg/ml) for 8 h prior to biomechanical testing to obtain load (N), stress (MPa), elongation to failure (mm), and Young's modulus (MPa) for each explant. Significant differences were assessed using a mixed linear model with horses as a random factor and control or insulin-treated group as a fixed factor. RESULTS: Lamellar explants incubated in medium supplemented with insulin failed at significantly lower load (P = 0.0001) and lower stress (P = 0.001) and had greater elongation to failure (P = 0.02). MAIN LIMITATIONS: In addition to the ex vivo nature of the study, location-dependent variability in explant structural integrity and variable diffusion of nutrients due to explant size may have been limitations. However, the study design attempted to account for these limitations through random assignment of explants to treatment groups independent of location and by evaluating stress to failure. CONCLUSIONS: Insulin weakens the structural integrity of equine lamellar explants and an ex vivo model for evaluation of hyperinsulinaemia-induced lamellar failure was established. The summary is available in Spanish - see Supporting Information.

Influence of dietary zinc on the claw and interdigital skin of sheep.

Claw diseases like interdigital dermatitis and footrot threaten sheep health and are major welfare issues. Several studies mainly done in cattle suggested that zinc (Zn) supplementation may improve claw integrity. However, Zn supplements may differ markedly regarding Zn bioavailability. Zn bound to single amino acids has been shown to be more bioavailable than inorganic Zn sources. The aim of this study was to determine the effect of different Zn supplements on the integrity of the claw and interdigital skin of healthy sheep. At weaning 30 Merino lambs were randomly allocated to three different dietary treatments which were provided through the pelleted concentrates as follows: 1) no supplemental Zn (Zn0); 2) addition of 40 mg/kg Zn as Zn sulphate (ZnS); 3) addition of 40 mg/kg organic Zn as Zn amino acid complex (CZn). Barley straw and pelleted concentrates were given ad-libitum. The calculated Zn concentration of the total diet (roughage and concentrate) without supplemental Zn (Zn0) was 38 mg Zn/kg DM. The concentrates were formulated to meet the nutritional requirements for growing lambs and contained 207 g/kg DM crude protein and 12.4 MJ/kg DM metabolizable energy. After 8 weeks the lambs were slaughtered and the following specimens were collected: blood serum, liver, sole and coronary band of the claw, and interdigital skin. Serum and tissue Zn and copper (Cu) concentrations and claw hardness were determined. Routine pathohistology and electron microscopy were conducted. Franz diffusion cell system and Ki-67 immunostaining were used to determine the permeability of the interdigital skin and the keratinocyte proliferation in the basal layer of sole horn, coronary band and interdigital skin, respectively. The concentrations of Zn and Cu in serum and liver tissue as well as the Zn concentration in claw horn were not affected by dietary treatment. Zn0 lambs showed higher (p < 0.05) Cu concentrations in claw horn compared to both Zn supplemented groups. Routine pathohistology as well as electron microscopy did not show significant morphological differences between the three groups. Franz diffusion cell system proved to be a suitable method examining the interdigital skin permeability, but the group differences in this study were not significant. Dietary treatment did not affect keratinocyte proliferation in the coronary band. In the sole keratinocyte proliferation was significantly higher (p < 0.05) in the Zn0 group compared to CZn with ZnS being intermediate. Keratinocyte proliferation in the interdigital skin was significantly higher (p < 0.05) in the CZn group compared to the Zn0 with ZnS being intermediate. The results of the current experiment indicate that serum and tissue Zn concentrations and horn hardness are not affected by adding a moderate amount of Zn sulphate or Zn amino acid complex to a basal diet. However, supplemental Zn amino acid complex seems to affect keratinocyte proliferation of interdigital skin and sole horn of lambs. Effects on skin permeability should be retested using a higher number of animals prospectively.

Comparison of effects of routine topical treatments in the milking parlor on digital dermatitis lesions.

Digital dermatitis (DD), an infectious bacterial disease affecting the feet of dairy cattle, can cause lameness and decrease milk production, fertility, and animal welfare. Current DD treatment typically involves routine hoof trimming and topical antibiotics. Several nonantibiotic commercial topical products are used for controlling DD lesions; however, there is limited or no evidence regarding their effectiveness. The objectives of this study were to evaluate 2 commercially available topical applications on their ability to (1) clinically cure active DD lesions to nonactive lesions and (2) prevent recurrence of active DD lesions. Ten farms were visited weekly. In the milking parlor, the hind feet of lactating cattle were cleaned and scored (M-stage scoring system). Cattle with DD lesions at the first visit were randomly allocated to 1 of 4 treatment groups: positive control (tetracycline solution), HealMax (AgroChem Inc., Saratoga Springs, NY), HoofSol (Diamond Hoof Care Ltd., Intracare BV, Veghel, the Netherlands), and a negative control (saline). All products were applied to lesions using a spray bottle. Tetracycline, HealMax, and HoofSol had a higher probability of clinical cure for active lesions compared with saline 1 wk after the first treatment (wk 1), with 69, 52, and 79% clinical cure of active lesions, respectively, compared with 34% with saline. At wk 7, the probability of clinical cure for active lesions was 10, 33, 31, and 45% of lesions treated weekly with saline, tetracycline, HealMax, and HoofSol, respectively (no difference among treatments). The substantial clinical cure with saline highlighted the potential importance of cleaning feet. In wk 1, treatment with saline, tetracycline, HealMax, and HoofSol resulted in a probability of recurrence of active DD lesions of 9, 11, 11, and 8%, respectively, with no product being superior to saline. After 7 wk, the probability of recurrence of active lesions was 5, 7, 6, and 6% for saline, tetracycline, HealMax, and HoofSol respectively, with no difference among groups in wk 7. These results provide alternatives to antibiotics for treatment of DD lesions and highlight the potential importance of cleaning feet in the milking parlor.

Evaluation of the promoting effect of soluble cyclodextrins in drug nail penetration.

Soluble derivatives of ß-cyclodextrin (CD) have a high capacity to solubilise hydrophobic molecules and to interact with proteins and membrane component. As consequence CD derivatives shows a significant activity as drug absorption enhancers through different delivery routes, such as the oral, nasal, ocular or topical route. In this paper, the effect of two CD derivatives -methyl-ß-cyclodextrin (MBCD) and hydroxypropyl-ß-cyclodextrin (HPB)- on the structure and permeability of the nail plate has been studied using the drug model ciclopirox olamine. Results shows that MBCD and HPB interacting with the nail plate components, modifying their microporous structure and swelling characteristics. The ability of the cyclodextrins to interact with aromatic amino acids and to stabilise and unfold protein structures could be the most likely mechanisms responsible of the nail microstructure modifications. Aditionally CD allows to increase the soluble dose of ciclopirox olamine in aqueous lacquers made with poloxamer and N-acetylcysteine via the formation of high solubility complexes with the drug. Finally the studies of diffusion and penetration obtained using bovine hoof model confirm the enhancing effect of the cyclodextrins on the penetration and accumulation of the drug in the nail structure. Results shows the great potential of the CD for the elaboration of aqueous based nail lacquers containing hidrofobic drugs.

Influence of a Combination of Chemical Enhancers and Iontophoresis on In Vitro Transungual Permeation of Nystatin.

To promote transungual permeation of nystatin (NYST), molecule with high molecular weight, no water-soluble, amphoteric by iontophoresis. The synergic effect of the combination of cetylpyridinium chloride, CPC, or polyoxyethylene (20) sorbitan monooleate, TW80, and iontophoresis was investigated. In vitro permeation experiments were carried out through bovine hoof slices using vertical diffusion cells. A low current density (0.2 mA/cm2) was applied by introducing Ag/AgCl electrodes in the donor (anode) and receptor (cathode) chambers. The donor phase consisted of a solution, a suspension, or gel-type vehicles containing NYST and surfactants in pH 5.6 HEPES buffer. The addition of CPC to NYST suspension (SOSP) produced a fivefold increase on the permeability of the bovine hoof membrane to the drug. The application of anodal iontophoresis further improved NYST flux. Conversely, NYST transungual permeation was not influenced by TW80 either in the passive diffusion or iontophoretic flux. Furthermore, the iontophoretic treatment does not appear to induce irreversible alterations to the hoof bovine membranes. The present work demonstrated the efficacy of iontophoresis as a treatment for different nail pathologies with large molecules very slightly soluble in water without irreversibly affecting the nail structure. A synergistic effect between CPC and iontophoresis was observed.

Terbinafine hydrochloride nail lacquer for the management of onychomycosis: formulation, characterization and in vitro evaluation.

AIM: The present investigation's intention was to develop an optimized nail lacquer (NL) for the management of onychomycosis. MATERIALS & METHODS: The NL was optimized statistically adopting 32 full factorial design having different polymer ratios and solvent ratios. The formulations were assessed for drug permeation drying time and peak adhesive strength of the film. Characterization was done using techniques including attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR), scanning electron microscopy (SEM), differential scanning calorimetry (DSC), x-ray diffraction (XRD), etc. RESULTS & CONCLUSION: The formulation that had 1:1 polymer ratio and 80:20 solvent ratio was chosen as the optimized formulation. In vitro permeation studies showed better penetration (∼3.25-fold) as well as retention (∼11-fold) of the optimized NL formulation in the animal hoof as compared with the commercial formulation. The findings of in vitro and ex vivo studies elucidated the potential of the optimized formulation. [Formula: see text].

Voriconazole-loaded nanostructured lipid carriers (NLC) for drug delivery in deeper regions of the nail plate.

Voriconazole-loaded nanostructured lipid carriers (VOR-NLC) were developed and drug penetration evaluated in porcine hooves in vitro. Synergistic effect of urea (Ur), selected among other known chemical enhancers according to hoof hydration potential, was also evaluated. VOR-NLC presented a high encapsulation efficiency (74.52±2.13%), approximate mean diameter of 230nm and were positively charged (+27.32±2.74mV). Stability studies indicated they were stable under refrigeration (4±2°C) for up to 150days. SEM images revealed hooves treated with VOR-NLC and VOR-NLC-Ur suffered a disturbance on the surface depicting high roughness and porosity. Permeation data showed a substantial VOR amount retained in superficial hooves sections independent of the formulation used (2.42±0.26; 2.52±0.36 and 2.41±0.60µg/cm2 for unloaded VOR, VOR-NLC and VOR-NLC-Ur, respectively, p>0.05). Still, successive extractions, revealed the amount of VOR retained in deeper regions was significantly higher when VOR-NLC or VOR-NLC-Ur was used (0.17±0.04, 0.47±0.14 and 0.36±0.07µg/cm2 for unloaded VOR, VOR-NLC and VOR-NLC-Ur, respectively, p<0.05). Such results indicate NLC are promising formulations for the management of onychomycosis. Further studies in diseased nail plates are necessary.

A Review of the Design and Management of Footbaths for Dairy Cattle.

This article summarizes current footbath practices, questions the mechanism by which footbaths function, and reviews the available scientific literature testing footbaths in the field. Copper sulfate appears the most efficacious agent to include in a footbath program, but disposal concerns should limit the frequency of its use. Other agents such as formaldehyde have some merit when used with care. Use of water alone in a flush bath appears to have minimal impact. Footbaths should be used as infrequently as possible to achieve lameness prevention goals for the herd.

Development of ciclopirox nail lacquer with enhanced permeation and retention.

Onychomycosis is a prevailing disease caused by fungal infection of nails that mostly affects athletes and the elderly. Ciclopirox is approved by the US Food and Drug Administration for the topical treatment of onychomycosis. However, the desired penetration of ciclopirox into the nail bed has not been achieved via topical application for efficient treatment. Therefore, the main aim of this study was to enhance ciclopirox permeation and retention in nail by the development of a new nail lacquer formulation. We screened the effects of different solvents, alkalizing agents, and permeation enhancers on the permeation of bovine hooves by ciclopirox and its retention in human nail clippings. The results suggest that isopropyl alcohol, potassium hydroxide, and urea as the solvent, alkalizing agent, and permeation enhancer, respectively, improved the permeation of the ciclopirox nail lacquer formulation the most with high flux rates. Comparison of the final formulation and marketed product revealed enhanced retention of ciclopirox from our developed formulation in human nail clippings. Therefore, our newly developed nail lacquer may be a potentially effective formulation for the treatment of onychomycosis in humans.

Fumonisin B1 (FB1) Induces Lamellar Separation and Alters Sphingolipid Metabolism of In Vitro Cultured Hoof Explants.

One of the most important hoof diseases is laminitis. Yet, the pathology of laminitis is not fully understood. Different bacterial toxins, e.g. endotoxins or exotoxins, seem to play an important role. Additionally, ingestion of mycotoxins, toxic secondary metabolites of fungi, might contribute to the onset of laminitis. In this respect, fumonsins are of special interest since horses are regarded as species most susceptible to this group of mycotoxins. The aim of our study was to investigate the influence of fumonisin B1 (FB1) on primary isolated epidermal and dermal hoof cells, as well as on the lamellar tissue integrity and sphingolipid metabolism of hoof explants in vitro. There was no effect of FB1 at any concentration on dermal or epidermal cells. However, FB1 significantly reduced the separation force of explants after 24 h of incubation. The Sa/So ratio was significantly increased in supernatants of explants incubated with FB1 (2.5-10 µg/mL) after 24 h. Observed effects on Sa/So ratio were linked to significantly increased sphinganine concentrations. Our study showed that FB1 impairs the sphingolipid metabolism of explants and reduces lamellar integrity at non-cytotoxic concentrations. FB1 might, therefore, affect hoof health. Further in vitro and in vivo studies are necessary to elucidate the effects of FB1 on the equine hoof in more detail.

Evaluation of topical antifungal products in an in vitro onychomycosis model.

Many topical commercial products are currently available for the treatment of onychomycosis. However, limited data are available concerning their antifungal activity. Using an in vitro onychomycosis model, the daily application of seven nail formulations was compared to the antifungal reference drug amorolfine (Loceryl(®) ) and evaluated for inhibitory activity against Trichophyton mentagrophytes using an agar diffusion test. Of all commercial nail formulations, only Excilor(®) and Nailner(®) demonstrated inhibitory activity, which was much lower compared to the daily application of Loceryl(®) . However, Excilor(®) showed similar efficacy compared to the conventional weekly application of Loceryl(®) . These results suggest a role for organic acids in the antifungal effect of Excilor(®) (acetic acid, ethyl lactate) and Nailner(®) (lactic acid, citric acid, ethyl lactate) as all tested formulations without organic acids were inactive.