A blackberry-dill extract combination synergistically increases skin elasticity.

BACKGROUND: The loss of structural elastin due to intrinsic and extrinsic ageing results in the skin's inability to stretch and recoil (decrease in elasticity) and manifests as loss of skin firmness and sagging. While other extracellular matrix (ECM) components such as collagen and hyaluronic acid are continually synthesized and assembled through life, elastic fibres are not. Elastic fibre assembly and functionality require fibre cross-linking, induced by the lysyl oxidase-like (LOXL) enzymes, which sharply decrease during ageing. OBJECTIVE: To evaluate the enhanced elastogenic effect of a blackberry-dill extract combination, which was hypothesized to induce elastin fibre component synthesis, fibre cross-linking and reduce elastin fibre degradation. METHODS: The blackberry and the dill extracts were tested separately and in combination to confirm single ingredient bioactivity and synergistic benefits. Human skin explants, dermal fibroblasts, elastase assays, ELISAs, quantitative real-time PCRs and spectrofluorometer measurements were used. Moreover, a double-blinded, placebo-controlled clinical study was carried out to assess skin elasticity using Cutometer and histologically from biopsies. RESULTS: The blackberry extract induced elastin gene expression, elastin promoter activity and inhibited elastic fibre degradation by matrix metalloproteinases (MMPs) 9 and 12. The dill extract induced elastin, collagen and LOXL1 gene expression, resulting in enhanced fibre cross-linking in human skin explants. Clinically, the blackberry and dill combination treatment displayed synergistic pro-elasticity activity as compared to each ingredient alone and placebo. CONCLUSION: Taken together, these results demonstrated the two multimodal plant-based extracts complemented each other in terms of bioactivity and resulted in a synergistic elastogenesis induction.

CONTEXTE: la perte de l'élastine structurelle causée par un vieillissement intrinsèque et extrinsèque provoque l'incapacité de la peau à s'étirer et à rebondir (diminution de l'élasticité) et se manifeste comme une perte de fermeté et un relâchement de la peau. Alors que d'autres composants de la matrice extracellulaire (MEC), tels que le collagène et l'acide hyaluronique sont continuellement synthétisés et assemblés tout au long de la vie, les fibres élastiques ne le sont pas. L'assemblage et la fonctionnalité des fibres élastiques nécessitent une réticulation des fibres, causée par les enzymes de type lysyle oxydase (LOXL), qui diminuent fortement au cours du vieillissement. OBJECTIF: évaluer l'effet élastogène amélioré d'une combinaison d'extrait de mûre et d'aneth, qui était supposée induire la synthèse des composants des fibres d'élastine, la réticulation des fibres et réduire la dégradation des fibres d'élastine. MÉTHODES: les extraits de mûre et d'aneth ont été testés séparément et ensemble pour confirmer la bioactivité d'un seul ingrédient et les avantages synergiques. Des explants de peau humaine, des fibroblastes cutanés, des dosages d'élastase, des ELISA, des analyses PCR quantitatives en temps réel et des mesures de spectrofluorimètre ont été utilisés. De plus, une étude clinique en double aveugle, contrôlée par placebo, a été réalisée pour évaluer l'élasticité de la peau à l'aide du cutomètre et histologiquement à partir de biopsies. RÉSULTATS: l'extrait de mûre a induit l'expression génique de l'élastine, l'activité de promoteur de l'élastine et a inhibé la dégradation des fibres élastiques par des métalloprotéinases matricielles (MPM) 9 et 12.L'extrait d'aneth a causé l'expression génique de l'élastine, du collagène et du gène LOXL1, entraînant une amélioration de la réticulation des fibres dans les explants de peau humaine. Cliniquement, le traitement par une combinaison de mûre et d'aneth a montré une activité de pro-élasticité synergique par rapport à chaque ingrédient seul et au placebo. CONCLUSION: ensemble, ces résultats ont démontré que les deux extraits de plantes multimodales se complètent en termes de bioactivité et ont entraîné une induction synergique de l'élastogenèse.

4-Hexyl-1,3-phenylenediol, a nuclear factor-κB inhibitor, improves photodamaged skin and clinical signs of ageing in a double-blinded, randomized controlled trial.

BACKGROUND: The nuclear factor-κB (NF-κB) pathway is a key mediator of inflammation; however, few studies have examined the direct effects of NF-κB inhibition on the skin. OBJECTIVES: To investigate NF-κB activity in cultured human fibroblasts and to investigate the effects of 4-hexyl-1,3-phenylenediol (an NF-κB inhibitor) on elastin and collagen gene expression in vitro and on the clinical appearance of photodamaged skin. METHODS: The amount and activity of NF-κB in human fibroblasts obtained from donors (17-78 years old) was measured after transfection with a NF-κB reporter and a luciferase promoter system. The expression of extracellular matrix (ECM) genes was determined using quantitative polymerase chain reaction. Women with moderate skin photodamage were randomized to daily treatment with a topical lotion containing 4-hexyl-1,3-phenylenediol (n = 30) or vehicle (n = 29) for 8 weeks, with clinical assessments at baseline and weeks 2, 4 and 8. RESULTS: Fibroblasts obtained from donors older than 50 years had higher NF-κB activity compared with cells from younger donors; inhibition of the NF-κB pathway with 4-hexyl-1,3-phenylenediol enhanced the expression of ECM genes. In women, treatment for 8 weeks with 4-hexyl-1,3-phenylenediol significantly improved crow's feet fine lines, cheek wrinkles, age spots, mottled pigmentation and radiance compared with both the vehicle and baseline. Furthermore, treatment with 4-hexyl-1,3-phenylenediol resulted in a twofold greater clinical improvement in overall photodamage compared with the vehicle group. CONCLUSIONS: Inhibition of the proinflammatory NF-κB pathway resulted in increased expression of ECM proteins in vitro and significant clinical improvement in photodamaged skin.

Twenty-four hour exposure to prostaglandin downregulates prostanoid receptor binding but does not alter PGE(2)-mediated sensitization of rat sensory neurons.

Although the tissue levels of prostaglandins are elevated for a relatively long period during injury or inflammation, few studies have been performed to assess the effects of prolonged prostaglandin exposure on receptor binding and activity in sensory neurons. Consequently, we examined whether unilateral inflammation or a 24 h exposure to prostaglandin E2 (PGE2) altered binding of this prostanoid in spinal cord tissue or in isolated sensory neurons, respectively. To assess functional changes in EP receptors, we also examined PGE2-induced cAMP production and the prostanoid-mediated augmentation of substance P release from isolated sensory neurons after acute and 24 h pretreatment with PGE2. Injection of complete Freund's adjuvant into the hindpaw decreased binding of PGE2 in ipsilateral, but not contralateral dorsal spinal cord 24 h after injection. This decrease in Bmax was blocked by administration of intrathecal ketorolac (10 nmol/microl/h) for 24 h prior to and throughout the period of inflammation, suggesting that the inflammation-induced decrease in binding is dependent on prostaglandin synthesis. In an analogous manner, treating sensory neurons grown in culture with 1 microM PGE2 for 24 h decreased [3H]-PGE2 binding by approximately 50% without altering binding affinity. Exposing neuronal cultures to 1 microM PGE2 for 24 h also reduced, but did not abolish the ability of the prostanoid to increase the production of cAMP. This treatment, however, did not significantly alter the ability of PGE2 to augment the evoked release of immunoreactive substance P from sensory neurons. These results demonstrate that under conditions that significantly downregulate PGE2 binding, sensory neurons are still capable of maintaining PGE2-mediated sensitization.

Prostaglandin E(2)-mediated sensitization of rat sensory neurons is not altered by nerve growth factor.

To ascertain whether chronic exposure to nerve growth factor (NGF) alters the responsiveness of sensory neurons to prostaglandin E(2) (PGE(2)), sensory neurons taken from adult rats were grown in culture in the presence or absence of NGF for 7 days. Neurons then were exposed to PGE(2) and release of immunoreactive calcitonin gene-related peptide (iCGRP) and production of immunoreactive cAMP (icAMP) were examined. Growing neurons in the presence of 250 ng/ml NGF increased the content and the release of iCGRP from sensory neurons. Independent of NGF treatment, exposure to 100 nM PGE(2) augmented capsaicin- or potassium-stimulated release of iCGRP by 1. 5-fold compared with cells not exposed to PGE(2). In a similar manner, NGF treatment did not alter the ability of PGE(2) to increase the content of icAMP. These data suggest that prostaglandin-induced sensitization of sensory neurons is not influenced by NGF.

Age dependent changes in serotonin and dopamine receptors in Aplysia californica.

Age related changes in dopaminergic and serotonergic receptors were examined in Aplysia californica. In this study dopamine (DA) and serotonin (5-HT) receptor levels were examined for animals belonging to 4-, 5-, 6-, 8-, 9- and 12-month age groups. Receptors analysis was performed using radio-labeled d-[3H] lysergic acid diethylamide (LSD) as the specific ligand. Specific binding for 5-HT was found to be significantly greater than that for DA in the young (4-month post-hatch) animals. The total DA and 5-HT receptor levels changed significantly with age. Dopamine levels increased from 5.34 fmol/mg of protein at 4 months to 19.11 fmol/mg at 12 months. Serotonin receptor levels increased from 7.35 fmol/mg at 4 months to 20.45 at 12 months.

Differential levels of elastin fibers and TGF-ß signaling in the skin of Caucasians and African Americans.

BACKGROUND: While skin color is the most notable difference among ethnic skins the current knowledge on skin physiological and aging properties are based mainly on Caucasian skin studies. OBJECTIVE: To evaluate histological differences in elastin fiber network and differential responsiveness to TGF-ß in skin of Caucasians and African Americans. METHODS: These studies were undertaken using human skin biopsies, primary dermal fibroblasts, Western blot analyses, immunofluorescence microscopy, cDNA array and quantitative real-time PCR. RESULTS: In Caucasian subjects, tropoelastin expression and elastin fibers in photoprotected skin was substantially less than in age-matched African American skin. Expression of tropoelastin in photoexposed skin of African American was similar to their photoprotected skin, suggesting that photoexposure did not affect elastin fibers in African American skin to the same extent as Caucasian skin. An elevated level of TGF-ß1 present in media from dermal fibroblasts derived from African American skins correlated well with the higher levels of TGF-ß mRNA in African American skins analyzed by cDNA array. Treatment with TGF-ß1 resulted in a considerably higher induction of elastin mRNA in dermal fibroblasts from African Americans than from Caucasian fibroblasts, indicative of enhanced TGF-ß signaling in African American skins. Furthermore, UVA exposure decreased levels of elastin mRNA in Caucasian fibroblasts compared to African Americans fibroblasts. CONCLUSION: These results suggest that there are ethnic differences in the elastin fiber network and in TGF-ß signaling in African American and Caucasian skin, and that African American have less UV dependent loss of elastin than Caucasian which may contribute to the different perceived aging phenotypes.

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.

Prostaglandin receptor subtypes, EP3C and EP4, mediate the prostaglandin E2-induced cAMP production and sensitization of sensory neurons.

Although a number of prostaglandin E(2) (PGE(2)) receptor subtypes have been cloned, limited studies have been performed to elucidate subtypes that subserve specific actions of this eicosanoid, in part because of a paucity of selective receptor antagonists. Using reverse transcription-polymerase chain reaction (PCR) and antisense oligonucleotides, we examined which prostaglandin E(2) receptor (EP receptor) subtypes are expressed in sensory neurons and which mediate the PGE(2)-induced increase in cAMP production and augmentation of peptide release. Reverse transcription-PCR of cDNA isolated from rat sensory neurons grown in culture revealed PCR products for the EP1, EP2, EP3C, and EP4 receptor subtypes but not the EP3A or EP3B. Preexposing neuronal cultures for 48 h to antisense oligonucleotides of EP3C and EP4 mRNA diminished expression of the respective receptors by approximately 80%, abolished the PGE(2)-stimulated production of cAMP, and blocked the ability of PGE(2) to augment release of immunoreactive substance P and calcitonin gene-related peptide. Pretreating with individual antisense against the EP2, EP3C, or EP4 receptors or combinations of missense oligonucleotides had no effect on PGE(2)-induced activity. Treatment with antisense to EP3C and EP4 receptor subtypes did not alter the ability of forskolin to increase cAMP or enhance peptide release. These results demonstrate that sensory neurons are capable of expressing multiple EP receptor subtypes but that only the EP3C and EP4 receptors mediate PGE(2)-induced sensitization of sensory neurons.

The platelet-activating factor receptor protects epidermal cells from tumor necrosis factor (TNF) alpha and TNF-related apoptosis-inducing ligand-induced apoptosis through an NF-kappa B-dependent process.

A number of chemical mediators can induce human keratinocytes and epidermal-derived carcinomas to undergo apoptosis, or programmed cell death. Recent evidence suggests pro-inflammatory cytokines, such as interleukin-1 beta or transforming growth factor alpha, protects carcinomas from numerous pro-apoptotic stimuli. Platelet-activating factor (1-alkyl-2-acetyl-3-glycerophosphocholine; PAF) is a lipid mediator with pro-inflammatory effects on numerous cell types. Although PAF can be metabolized to other bioactive lipids, the majority of PAF effects occur through activation of a G protein-coupled receptor. Using a model system created by retroviral transduction of the PAF receptor (PAF-R) into the PAF-R-negative human epidermal cell line KB and the PAF-R-expressing keratinocyte cell line HaCaT, we now demonstrate that activation of the epidermal PAF-R results in protection from apoptosis induced by tumor necrosis factor (TNF) alpha or TNF-related apoptosis-inducing ligand. The PAF-mediated protection was inhibited by PAF-R antagonists, and protection did not occur in PAF-R-negative KB cells. Additionally, we show protection from TNFalpha- or TRAIL-induced apoptosis by PAF-R activation is dependent on the transcription factor nuclear factor (NF)-kappa B, because PAF-R activation-induced NF-kappa B and epidermal cells transduced with a super-repressor form of inhibitor kappa B were not protected by the PAF-R. These studies provide a mechanism whereby the epidermal PAF-R, and possibly other G protein-coupled receptors, can exert anti-apoptotic effects through an NF-kappa B-dependent process.

Isoprostanes, novel eicosanoids that produce nociception and sensitize rat sensory neurons.

Isoprostanes are a novel class of eicosanoids primarily formed by peroxidation of arachidonic acid. Because of their potential as inflammatory and/or hyperalgesic agents whose formation is largely independent of cyclooxygenases, we examined whether 8-iso prostaglandin E(2) (8-iso PGE(2)) or 8-iso prostaglandin F(2alpha) (8-iso PGF(2alpha)) reduces mechanical and thermal withdrawal threshold in rats, and whether they sensitize rat sensory neurons. Injection of 1 microg of 8-iso PGE(2) (in 2.5 microl) into the hindpaw of rats significantly reduced mechanical and thermal withdrawal thresholds, whereas 1 microg of 8-iso PGF(2alpha) elicited a transient decrease in only the mechanical withdrawal threshold. Both isoprostanes enhanced the firing of C-nociceptors in a concentration-dependent manner when injected into peripheral receptive fields. Exposing sensory neurons grown in culture to 1 microM 8-iso PGE(2) or 8-iso PGF(2alpha) augmented the number of action potentials elicited by a ramp of depolarizing current. In contrast, 8-iso PGE(2) but not 8-iso PGF(2alpha) enhanced the release of substance P- and calcitonin gene-related peptide-like immunoreactivity from isolated sensory neurons. Ten micromolar 8-iso PGE(2) stimulated peptide release directly, whereas treatment with 1 microM 8-iso PGE(2) augmented the release evoked by either bradykinin or capsaicin. Pretreating neuronal cultures with the nonsteroidal anti-inflammatory drug ketorolac did not alter the sensitizing action of 8-iso PGE(2) on peptide release, suggesting that this action of the isoprostane was not secondary to the production of prostaglandins via the cyclooxygenase pathway. These data support the notion that isoprostanes are an important class of inflammatory mediators that augment nociception.