Effect of L-4-Thiazolylalanine (Protinol™) on skin barrier strength and skin protection.

OBJECTIVES: Skin barrier properties are critical for maintaining epidermal water content, protecting from environmental factors and providing the first line of defense against pathogens. In this study, we investigated the non-proteinogenic amino acid L-4-Thiazolylalanine (L4) as a potential active ingredient in skin protection and barrier strength. METHODS: L4 on wound healing, anti-inflammatory and anti-oxidant properties were evaluated using monolayers and 3D skin equivalents. The transepithelial electrical resistance (TEER) value was used in vitro as a strong indicator of barrier strength and integrity. Clinical L4 efficacy was assessed for the evaluation of the skin barrier integrity and soothing benefits. RESULTS: In vitro treatments of L4 show beneficial effects in wound closure mechanism, and we demonstrate that L4 anti-oxidant benefits with markedly increased HSP70 and decreased reactive oxygen species production induced by UVs exposure. Barrier strength and integrity were significantly improved by L4, confirmed clinically by an increase in 12R-lipoxygenase enzymatic activity in the stratum corneum. In addition, soothing benefits of L4 have been shown clinically with the decrease in redness after methyl nicotinate application on the inner arm and the significant reduction of the erythema and the skin desquamation on the scalp. CONCLUSION: L4 delivered multiple skin benefits by strengthening the skin barrier, accelerating the skin repair process as well as soothing the skin and the scalp with anti-inflammaging effects. The observed efficacy validates L4 as a desirable skincare ingredient for topical treatment.

OBJECTIFS: La barrière cutanée est primordiale pour le maintien d'un épiderme hydraté, sa protection contre les facteurs environnementaux et pour conférer une première ligne de défense contre les pathogènes. Dans cette étude, nous nous intéressons à un aminoacide non-proteinogénique, L-4-Thiazolylalanine (L4) comme potentiel actif cosmétique et évaluons ses effets sur la protection de la peau et le renforcement de la barrière cutanée. MÉTHODES: Les propriétés cicatrisantes, anti-inflammatoires et antioxydantes de L4 ont été déterminées par culture cellulaire et sur modèles de peau 3D. La mesure de la résistance électrique transépithéliale a été utilisée in vitro comme indicateur de la résistance de la barrière de la peau. Des études cliniques ont été réalisées pour évaluer L4 sur ses capacités apaisantes et son impact sur l'intégrité de la barrière cutanée. RÉSULTATS: In vitro, nous déterminons qu'un traitement L4 a des effets bénéfiques dans le processus de cicatrisation mais aussi des propriétés antioxydantes, induisant une augmentation de HSP70 ainsi qu'une diminution de la production d'espèces réactives a l'oxygène induite par rayons UV. Un renforcement de la barrière cutanée et une amélioration de son intégrité sont observés après un traitement L4 et confirme cliniquement par une augmentation de l'activité enzymatique de la 12R-lipoxygenase dans le stratum corneum. De plus, les effets protecteurs de L4 ont été confirmes cliniquement avec une rougeur cutanée diminuée après application de nicotinate de methyle sur l'avant-bras interne ainsi qu'une réduction significative d'érythème et de desquamation cutanée du cuir chevelu. CONCLUSION: L4 offre de multiples bénéfices pour la peau en renforçant la barrière cutanée, accélérant le processus de cicatrisation et offrant des actions anti-inflammatoires apaisantes pour la peau et le cuir chevelu. Cela fait de L4 un ingrédient cosmétique intéressant pour une application topic.

Surgical techniques used in the management of intra-arachnoid diverticula in dogs across four referral centres and their immediate outcome.

OBJECTIVES: To report the surgical techniques being used to treat single focal spinal intra-arachnoid diverticula in dogs, their complications and immediate postoperative outcomes. MATERIALS AND METHODS: Retrospective multi-centre case series across four referral centres. RESULTS: Fifty-seven dogs were included in the study. The most common type of surgery was durectomy (28 dogs) followed by marsupialisation (11 dogs), durotomy alone (seven dogs), shunt placement (six dogs) and stabilisation (five dogs). A higher proportion of intra-arachnoid shunt dogs became unable to walk in the immediate postoperative period (24 hours postsurgery) (4/6, 66%) compared to all dogs five of 57, 9% (2/7 durotomy alone, 3/28 durectomy alone). Of the nine dogs with immediate postoperative deterioration, seven had improved, walking without assistance, by 3 to 5 weeks postoperatively. CLINICAL SIGNIFICANCE: This study does not identify an influence of surgical technique on short-term outcome. Dogs with a thoracolumbar intra-arachnoid diverticulum that undergo a shunt placement are likely to deteriorate neurologically in the immediate 24-hour postoperative period but appear to improve by 3 to 5 weeks after surgery. Further work is required to evaluate whether one surgical technique is superior for preventing or reducing long-term relapse.

Comparison of nerve conduction parameters in canine studies using recording needle and surface electrodes.

Surface electrodes have been used in electromyography and nerve conduction studies in human and veterinary medicine, but comparisons have not been made between surface and needle electrode recordings in dogs. Our aim in this method comparison study was to determine whether surface electrodes captured larger compound motor action potentials (CMAP) than needle electrodes. Tibial nerve CMAP from 25 dogs with normal limb function was acquired using both surface and needle recording electrodes; the stimulus was elicited with monopolar concentric needles. Paired Wilcoxon signed rank test (if data was not normally distributed) or a paired two tailed t-test was used if data were normally distributed; significance was set at P<0.05. Mean CMAP amplitude (P=0.009), area (P=0.045) and latency (P=0.02) recorded with needle electrodes were larger compared with surface recording. CMAP duration was not significantly longer when recorded with surface electrodes (P=0.898). Needle electrode recordings are suitable for canine studies, although surface electrodes could also be considered. Low CMAP amplitudes recorded with surface electrodes should be verified with needle electrodes.

Same-day surgery may reduce the risk of losing pain perception in dogs with thoracolumbar disc extrusion.

OBJECTIVES: To compare the proportions of dogs with thoracolumbar disc extrusion that lose pelvic limb pain perception if surgery is performed on the day of admission or delayed overnight. To describe the outcome of those dogs that deteriorate to lose pain perception. MATERIALS AND METHODS: Retrospective, single centre study on 273 client-owned dogs with thoracolumbar disc extrusion and intact pain perception, but inability to walk unaided on their pelvic limbs. Dogs were subdivided into two groups: early surgery (spinal decompression between their examination at day of admission and the following morning), and delayed surgery (did not undergo surgery between admission and the following morning). The proportion of dogs that lost pelvic limb pain perception overnight was compared between the early and delayed surgery groups. RESULTS: Seven of 151 dogs in the early surgery group lost pain perception overnight compared to 15 of 122 in the delayed surgery group (Fisher's exact test, P = 0.025). Number-needed-to-treat analysis suggested that 14 dogs (95% confidence interval: 7-106 dogs) need early surgery to prevent one losing pain perception overnight. Five of the seven dogs that lost pain perception in the early surgery group recovered pain perception by 3 weeks post-operatively, compared to eight of 14 in the delayed group. CLINICAL SIGNIFICANCE: This study suggests that an overnight delay before spinal decompression increases the risk of clinically meaningful deterioration in dogs unable to walk following thoracolumbar disc extrusion.

Anti-inflammatory activity of parthenolide-depleted Feverfew (Tanacetum parthenium).

Extracts of Tanacetum parthenium (L.) Sch. Bip., a plant known under the common name "Feverfew", contains the sesquiterpene lactone parthenolide, a potent skin sensitizer. To eliminate the risk of skin sensitization from Feverfew, we developed a parthenolide-depleted extract of Feverfew (PD-Feverfew) and determined its effectiveness as an anti-inflammatory agent. We confirmed that PD-Feverfew was sufficiently depleted of parthenolide since PD-Feverfew did not inhibit TNF-alpha induced-NF-kappaB activity unlike parthenolide containing whole Feverfew. PD-Feverfew directly inhibited the activity of pro-inflammatory enzymes 5-lipoxygenase, phosphodiesterase-3 and phosphodiesterase-4. PD-Feverfew inhibited the release of pro-inflammatory mediators nitric oxide, PGE(2) and TNF-alpha from macrophages and TNF-alpha, IL-2, IFN-gamma and IL-4 from human peripheral blood mononuclear cells. Additionally, PD-Feverfew inhibited TPA-induced release of PGE(2) from human skin equivalents. In vivo, PD-Feverfew inhibited oxazolone-induced dermatitis, and was more potent than whole Feverfew in reducing TPA-induced dermatitis. Finally the efficacy of PD-Feverfew was confirmed clinically by a reduction in erythema in a methyl nicotinate-induced vasodilation model. In conclusion, our results indicate that PD-Feverfew extracts have potent anti-inflammatory activity suggesting that this botanical would be efficacious in relieving inflammation without inducing immune sensitization.

Risk factors for early post-operative neurological deterioration in dogs undergoing a cervical dorsal laminectomy or hemilaminectomy: 100 cases (2002-2014).

Early post-operative neurological deterioration is a well-known complication following dorsal cervical laminectomies and hemilaminectomies in dogs. This study aimed to evaluate potential risk factors for early post-operative neurological deterioration following these surgical procedures. Medical records of 100 dogs that had undergone a cervical dorsal laminectomy or hemilaminectomy between 2002 and 2014 were assessed retrospectively. Assessed variables included signalment, bodyweight, duration of clinical signs, neurological status before surgery, diagnosis, surgical site, type and extent of surgery and duration of procedure. Outcome measures were neurological status immediately following surgery and duration of hospitalisation. Univariate statistical analysis was performed to identify variables to be included in a multivariate model. Diagnoses included osseous associated cervical spondylomyelopathy (OACSM; n = 41), acute intervertebral disk extrusion (IVDE; 31), meningioma (11), spinal arachnoid diverticulum (10) and vertebral arch anomalies (7). Overall 54% (95% CI 45.25-64.75) of dogs were neurologically worse 48 h post-operatively. Multivariate statistical analysis identified four factors significantly related to early post-operative neurological outcome. Diagnoses of OACSM or meningioma were considered the strongest variables to predict early post-operative neurological deterioration, followed by higher (more severely affected) neurological grade before surgery and longer surgery time. This information can aid in the management of expectations of clinical staff and owners with dogs undergoing these surgical procedures.

An alternative approach to depigmentation by soybean extracts via inhibition of the PAR-2 pathway.

The protease-activated receptor 2, expressed on keratinocytes but not on melanocytes, has been ascribed functional importance in the regulation of pigmentation by phagocytosis of melanosomes. Inhibition of protease-activated receptor 2 activation by synthetic serine protease inhibitors requires keratinocyte-melanocyte contact and results in depigmentation of the dark skinned Yucatan swine, suggesting a new class of depigmenting mechanism and agents. We therefore examined natural agents that could exert their effect via the protease-activated receptor 2 pathway. Here we show that soymilk and the soybean-derived serine protease inhibitors soybean trypsin inhibitor and Bowman-Birk inhibitor inhibit protease-activated receptor 2 cleavage, affect cytoskeletal and cell surface organization, and reduce keratinocyte phagocytosis. The depigmenting activity of these agents and their capability to prevent ultraviolet-induced pigmentation are demonstrated both in vitro and in vivo. These results imply that inhibition of the protease-activated receptor 2 pathway by soymilk may be used as a natural alternative to skin lightening.

Modulation of microphthalmia-associated transcription factor gene expression alters skin pigmentation.

The microphthalmia-associated transcription factor is implicated in melanocyte development and in the regulation of melanogenesis. Microphthalmia-associated transcription factor is thought to bind to the M-box promoter elements of tyrosinase, tyrosinase-related protein-1 and dopachrome tautomerase/tyrosinase-related protein-2 and transactivate these genes, resulting in increased pigmentation. Using a luciferase reporter construct driven by the microphthalmia-associated transcription factor promoter, we identified agents that modulate microphthalmia-associated transcription factor promoter activity. Changes in endogenous microphthalmia-associated transcription factor expression levels upon treatment with these agents were confirmed using northern and western blots, and their pigmentary modulating activities were demonstrated. Ultraviolet B irradiation and traditional Chinese medicine-1, a natural extract used in traditional Chinese medicine, upregulated microphthalmia-associated transcription factor gene expression and enhanced tyrosinase activity in vitro. Dihydrolipoic acid, lipoic acid, and resveratrol reduced microphthalmia-associated transcription factor and tyrosinase promoter activities. These agents also inhibited the forskolin- and ultraviolet B-stimulated promoter activities of these genes and significantly reduced tyrosinase activity in melanocyte cultures, resulting in depigmentation. Overexpressed microphthalmia-associated transcription factor was capable of rescuing the repressive effects of these compounds on the cotransfected tyrosinase promoter. Dark-skinned Yucatan swine treated with these agents showed visible skin lightening, which was confirmed histologically, whereas ultraviolet B-induced tanning of light-skinned swine was inhibited using these agents. Our findings suggest that modulation of microphthalmia-associated transcription factor expression can alter skin pigmentation and further confirm the central role of microphthalmia-associated transcription factor in melanogenesis.

Evaluation of topical retinoids for cutaneous pharmacological activity in Yucatan microswine.

The pharmacological effects of retinoids on skin have been studied primarily in test systems using small animals, such as mice and rabbits. Because of potentially significant differences in skin permeation and metabolism between small animals and humans, we have used Yucatan microswine as an alternative model for testing topical retinoids. Microswine skin resembles human skin, functionally and anatomically, more closely than most other species. In these studies, microswine skin was treated topically with retinoids for 5 consecutive days per week for 5 weeks. We found microswine epidermis to be functionally responsive to retinoids in that it undergoes hyperplasia and shows an increase in transepidermal water loss (TEWL). All-trans-retinoic acid, and its analogs, 13-cis-retinoic acid, 4-hydroxy-retinoic acid and 4-oxo-retinoic acid all caused epidermal thickening and increased TEWL. The three analogs were less potent than all-trans-retinoic acid. A synthetic retinoid, TTNPB, potently induced epidermal hyperplasia and increased TEWL, but a close structural analog, m-carboxy-TTNPB, which is also inactive on nuclear retinoic acid receptors, was without effects on microswine epidermis. These findings show that microswine are useful for evaluating the cutaneous effects of topical retinoids. This model could be of value in identifying retinoids with potential clinical activity.

Establishing a minimal erythema concentration of methyl nicotinate for optimum evaluation of anti-inflammatories.

Topical administration of chemicals such as methyl nicotinate that induce erythema have been employed to measure the effectiveness of formulations containing anti-inflammatory agents. Prior studies have utilized a single concentration of methyl nicotinate, between 36.5 and 100 mM, for all test subjects in evaluations of topical formulations. However, individuals have different thresholds of response to methyl nicotinate; thus, a single concentration may not be appropriate for all individuals and could result in the apparent lack of anti-inflammatory activity of the formulation being evaluated. In the current study, we evaluated the use of a minimal erythema concentration (MEC) of methyl nicotinate, defined as the lowest concentration that produces a complete and even erythema at the test site, compared with a 36.5-mM concentration of methyl nicotinate. Hydroalcoholic gels containing the nonsteroidal anti-inflammatory drug ibuprofen were compared with placebo. Diffuse reflectance spectroscopy was employed to measure differences in cutaneous inflammatory response between the control (placebo)-treated group and the ibuprofen-treated group. When chemical erythema was induced using an MEC of methyl nicotinate, greater reductions in erythema were seen in ibuprofen-treated sites compared with sites treated with a 36.5-mM concentration of methyl nicotinate. In conclusion, for an accurate assessment method of erythema induced by methyl nicotinate, consideration should be given to determining the extent of response of an erythema-producing agent on an individual basis. An MEC of methyl nicotinate should be determined and employed for each individual to obtain more consistent and reliable efficacy results of anti-inflammatory activity.

Androgen-induced delay of hair growth in the golden Syrian hamster.

The golden Syrian hamster flank organ has been used to study the stimulatory effect of androgens on sebaceous glands and hair. Androgens cause the sebaceous glands and hair follicles in this organ to grow. We have made the novel observation that exogenously administered androgen, testosterone propionate (TP), suppresses hair growth in the area surrounding the flank organ. When given in a time-release (systemic) subcutaneous dosage form (pellet), 25 mg TP inhibited the regrowth of clipped hair in peri-flank organ skin for up to 21 days; however, by 28 days hair grew back to the same extent as in controls. The peak serum level of testosterone in TP-treated animals occurred at 14 days, and declined thereafter. When two separate TP pellets (25 mg/pellet) were administered 14 days apart in order to maintain high serum levels for 28 days, the amount of hair regrowth after 35 days was identical to animals receiving a single TP pellet or placebo. This suggests that the systemic level of testosterone was not the only factor in hair regulation. Hair growing within the flank organ appeared to be unaffected by TP administration. In the golden Syrian hamster, androgen, as in humans, can exert stimulatory and inhibitory effects on hair growth depending on the body site. We conclude that this animal model could serve as a useful system to investigate the mechanisms responsible for the opposing effects of androgen on hair growth.