'Speed-ageing' of human skin in serum-free organ culture ex vivo: An instructive novel assay for preclinical human skin ageing research demonstrates senolytic effects of caffeine and 2,5-dimethylpyrazine.

Preclinical human skin ageing research has been limited by the paucity of instructive and clinically relevant models. In this pilot study, we report that healthy human skin of different age groups undergoes extremely accelerated ageing within only 3 days, if organ-cultured in a defined serum-free medium. Quantitative (immuno-)histomorphometry documented this unexpected ex vivo phenotype on the basis of ageing-associated biomarkers: the epidermis showed significantly reduced rete ridges and keratinocyte proliferation, sirtuin-1, MTCO1 and collagen 17a1 protein levels; this contrasted with significantly increased expression of the DNA-damage marker, γH2A.X. In the dermis, collagen 1 and 3 and hyaluronic acid content were significantly reduced compared to Day 0 skin. qRT-PCR of whole skin RNA extracts also showed up-regulated mRNA levels of several (inflamm-) ageing biomarkers (MMP-1, -2, -3, -9; IL6, IL8, CXCL10 and CDKN1). Caffeine, a methylxanthine with recognized anti-ageing properties, counteracted the dermal collagen 1 and 3 reduction, the epidermal accumulation of γH2A.X, and the up-regulation of CXCL10, IL6, IL8, MMP2 and CDKN1. Finally, we present novel anti-ageing effects of topical 2,5-dimethylpyrazine, a natural pheromone TRPM5 ion channel activator. Thus, this instructive, clinically relevant "speed-ageing" assay provides a simple, but powerful new research tool for dissecting skin ageing and rejuvenation, and is well-suited to identify novel anti-ageing actives directly in the human target organ.

Fluoxetine exerts anti-inflammatory effects on human epidermal keratinocytes and suppresses their endothelin release.

Fluoxetine is a safe antidepressant with remarkable anti-inflammatory actions; therefore, we aimed to investigate its effects on immortalized (HaCaT) as well as primary human epidermal keratinocytes in a polyinosinic-polycytidylic acid (p(I:C))-induced inflammatory model. We found that a non-cytotoxic concentration (MTT-assay, CyQUANT-assay) of fluoxetine significantly suppressed p(I:C)-induced expression and release of several pro-inflammatory cytokines (Q-PCR, cytokine array, ELISA), and it decreased the release of the itch mediator endothelins (ELISA). These effects were not mediated by the inhibition of the NF-κB or p38 MAPK pathways (western blot), or by the suppression of the p(I:C)-induced elevation of mitochondrial ROS production (MitoSOX Red labeling). Instead, unbiased activity profiling revealed that they were most likely mediated via the inhibition of the phosphoinositide 3-kinase (PI3K) pathway. Importantly, the PI3K-inhibitor GDC0941 fully mimicked the effects of fluoxetine (Q-PCR, ELISA). Although fluoxetine was able to occupy the binding site of GDC0941 (in silico molecular docking), and exerted direct inhibitory effect on PI3K (cell-free PI3K activity assay), it exhibited much lower potency and efficacy as compared to GDC0941. Finally, RNA-Seq analysis revealed that fluoxetine deeply influenced the transcriptional alterations induced by p(I:C)-treatment, and exerted an overall anti-inflammatory activity. Collectively, our findings demonstrate that fluoxetine exerts potent anti-inflammatory effects, and suppresses the release of the endogenous itch mediator endothelins in human keratinocytes, most likely via interfering with the PI3K pathway. Thus, clinical studies are encouraged to explore whether the currently reported beneficial effects translate in vivo following its topical administration in inflammatory and pruritic dermatoses.

Mechanical epilation exerts complex biological effects on human hair follicles and perifollicular skin: An ex vivo study approach.

OBJECTIVE: Electrical epilation of unwanted hair is a widely used hair removal method, but it is largely unknown how this affects the biology of human hair follicles (HF) and perifollicular skin. Here, we have begun to explore how mechanical epilation changes selected key biological read-out parameters ex vivo within and around the pilosebaceous unit. METHODS: Human full-thickness scalp skin samples were epilated ex vivo using an electro-mechanical device, organ-cultured for up to 6 days in serum-free, supplemented medium, and assessed at different time points by quantitative (immuno-)histomorphometry for selected relevant read-out parameters in epilated and sham-epilated control samples. RESULTS: Epilation removed most of the hair shafts, often together with fragments of the outer and inner root sheath and hair matrix. This was associated with persistent focal thinning of the HF basal membrane, decreased melanin content of the residual HF epithelium, and increased HF keratinocyte apoptosis, including in the bulge, yet without affecting the number of cytokeratin 15+ HF epithelial stem cells. Sebocyte apoptosis in the peripheral zone was increased, albeit without visibly altering sebum production. Epilation transiently perturbed HF immune privilege, and increased the expression of ICAM-1 in the bulge and bulb mesenchyme, and the number of perifollicular MHC class II+ cells as well as mast cells around the distal epithelium and promoted mast cell degranulation around the suprabulbar and bulbar area. Moreover, compared to controls, several key players of neurogenic skin inflammation, itch, and/or thermosensation (TRPV1, TRPA1, NGF, and NKR1) were differentially expressed in post-epilation skin. CONCLUSION: These data generated in denervated, organ-cultured human scalp skin demonstrate that epilation-induced mechanical HF trauma elicits surprisingly complex biological responses. These may contribute to the delayed re-growth of thinner and lighter hair shafts post-epilation and temporary post-epilation discomfort. Our findings also provide pointers regarding the development of topically applicable agents that minimize undesirable sequelae of epilation.

OBJECTIF: L'épilation électrique des poils indésirables est une méthode d'épilation largement utilisée, mais on ne connaît pas l'ampleur de son effet sur la biologie des follicules pileux humains (FP) et de la peau périfolliculaire. Dans cette étude, nous avons commencé à explorer comment l'épilation mécanique modifie certains paramètres de mesures biologiques clés ex vivo à l'intérieur et autour de l'unité pilo­sébacée. MÉTHODES: Des échantillons de peau du cuir chevelu humain de pleine épaisseur ont été épilés ex vivo à l'aide d'un dispositif électromécanique, cultivés biologiquement pendant un maximum de 6 jours dans un milieu complet sans sérum, et évalués à différents moments par (immuno­)histomorphométrie quantitative pour certains paramètres de mesures pertinents dans des échantillons avec épilation et des échantillons témoins avec épilation simulée. RÉSULTATS: L'épilation a enlevé la plupart des poils, souvent avec des fragments de la gaine de la racine externe et de la matrice pileuse. Cela a été associé à un amincissement focal persistant de la membrane basale du FP, à une diminution de la teneur en mélanine de l'épithélium résiduel du FP et à une augmentation de l'apoptose des kératinocytes du FP, y compris dans la surface arrondie, mais sans affecter le nombre de cellules souches épithéliales du FP positives pour la cytokératine 15. L'apoptose des sébocytes de la zone périphérique était augmentée, sans pour autant altérer visiblement la production de sébum. L'épilation a temporairement perturbé l'immunoprivilège du FP et a augmenté l'expression de l'ICAM­1 dans la surface arrondie et le mésenchyme du bulbe, ainsi que le nombre de cellules périfolliculaires du CMH de classe II et des mastocytes autour de l'épithélium distal, et a favorisé la dégranulation des mastocytes autour de la zone supra­bulbaire et bulbaire. En outre, par rapport aux échantillons témoins, plusieurs acteurs clés de l'inflammation neurogène cutanée, de la démangeaison et/ou de la thermosensation (TRPV1, TRPA1, NGF et NKR1) ont été exprimés de manière différentielle dans la peau après l'épilation. CONCLUSION: Ces données générées dans la peau du cuir chevelu humain dénervée et cultivée biologiquement démontrent que le traumatisme du FP induit par l'épilation mécanique provoque des réponses biologiques étonnamment complexes. Celles­ci peuvent contribuer à retarder la repousse des poils plus fins et plus clairs après l'épilation, et à provoquer une gêne temporaire après l'épilation. Nos résultats fournissent également des pistes concernant le développement d'agents applicables par voie topique qui minimisent les séquelles indésirables de l'épilation.

Analogies and Relations between Non-Additive Entropy Formulas and Gintropy.

We explore formal similarities and mathematical transformation formulas between general trace-form entropies and the Gini index, originally used in quantifying income and wealth inequalities. We utilize the notion of gintropy introduced in our earlier works as a certain property of the Lorenz curve drawn in the map of the tail-integrated cumulative population and wealth fractions. In particular, we rediscover Tsallis' q-entropy formula related to the Pareto distribution. As a novel result, we express the traditional entropy in terms of gintropy and reconstruct further non-additive formulas. A dynamical model calculation of the evolution of Gini index is also presented.

N,N-Dimethylglycine Sodium Salt Exerts Marked Anti-Inflammatory Effects in Various Dermatitis Models and Activates Human Epidermal Keratinocytes by Increasing Proliferation, Migration, and Growth Factor Release.

N,N-dimethylglycine (DMG) is a naturally occurring compound being widely used as an oral supplement to improve growth and physical performance. Thus far, its effects on human skin have not been described in the literature. For the first time, we show that N,N-dimethylglycine sodium salt (DMG-Na) promoted the proliferation of cultured human epidermal HaCaT keratinocytes. Even at high doses, DMG-Na did not compromise the cellular viability of these cells. In a scratch wound-closure assay, DMG-Na augmented the rate of wound closure, demonstrating that it promotes keratinocyte migration. Further, DMG-Na treatment of the cells resulted in the upregulation of the synthesis and release of specific growth factors. Intriguingly, DMG-Na also exerted robust anti-inflammatory and antioxidant effects, as assessed in three different models of human keratinocytes, mimicking microbial and allergic contact dermatitis as well as psoriasis and UVB irradiation-induced solar dermatitis. These results identify DMG-Na as a highly promising novel active compound to promote epidermal proliferation, regeneration, and repair, and to exert protective functions. Further preclinical and clinical studies are under investigation to prove the seminal impact of topically applied DMG-Na on relevant conditions of the skin and its appendages.

Non-Additive Entropy Formulas: Motivation and Derivations.

Entropy is a great tool in thermodynamics and statistical physics [...].

The role of protein kinase C isoenzymes in the pathogenesis of human autoimmune diseases.

The physiological role of protein kinase C (PKC) enzymes in the immune system is presented briefly. From earlier publications of others data were collected how the defects of one/two isoenzymes of PKC system suggested their involvement in the pathogenesis of human autoimmune diseases. Our observations on the defects of seven PKC isoenzymes in the peripheral blood mononuclear cells (PBMC) demonstrate that these molecular impairments are not prerequisits of the pathogenesis of systemic lupus erythematosus (SLE), mixed connective tissue disease and Sjögren's syndrome. However, these defects can modulate the disease activity and symptoms especially in SLE by several pathways. The role of PKC system in other forms of autoimmune diseases is also very small. It was of note that we detected decreased expression of PKC isoenzymes in PBMC of a European white family with an X-linked genetic background showing seasonal undulations in the lupus patient and also in her healthy mother.

Transient receptor potential vanilloid 3 expression is increased in non-lesional skin of atopic dermatitis patients.

TRPV3 (transient receptor potential vanilloid 3) is a pro-inflammatory ion channel mostly expressed by keratinocytes of the human skin. Previous studies have shown that the expression of TRPV3 is markedly upregulated in the lesional epidermis of atopic dermatitis (AD) patients suggesting a potential pathogenetic role of the ion channel in the disease. In the current study, we aimed at defining the molecular and functional expression of TRPV3 in non-lesional skin of AD patients as previous studies implicated that healthy-appearing skin in AD is markedly distinct from normal skin with respect to terminal differentiation and certain immune function abnormalities. By using multiple, complementary immunolabelling and RT-qPCR technologies on full-thickness and epidermal shave biopsy samples from AD patients (lesional, non-lesional) and healthy volunteers, we provide the first evidence that the expression of TRPV3 is markedly upregulated in non-lesional human AD epidermis, similar to lesional AD samples. Of further importance, by using the patch-clamp method on cultured healthy and non-lesional AD keratinocytes, we also show that this upregulation is functional as determined by the significantly augmented TRPV3-specific ion current (induced by agonists) on cultured non-lesional AD keratinocytes when compared to healthy ones.

Acne: Transient Arrest in the Homeostatic Host-Microbiota Dialog?

We propose that acne vulgaris represents a naturally developing, transient inflammatory interaction of adolescent facial skin with its new microbial/chemical milieu (Cutibacterium acnes, sebum), replacing a state of previous childhood skin homeostasis. This concept might explain why acne is characterized by strong regional and age specificity, prevalent occurrence, and resolution.

The Effect of Femtosecond Laser Irradiation and Plasmon Field on the Degree of Conversion of a UDMA-TEGDMA Copolymer Nanocomposite Doped with Gold Nanorods.

In this work, the effects of femtosecond laser irradiation and doping with plasmonic gold nanorods on the degree of conversion (DC) of a urethane dimethacrylate (UDMA)-triethylene glycol dimethacrylate (TEGDMA) nanocomposite were investigated. The UDMA-TEGDMA photopolymer was prepared in a 3:1 weight ratio and doped with dodecanethiol- (DDT) capped gold nanorods of 25 × 75 or 25 × 85 nm nominal diameter and length. It was found that the presence of the gold nanorods alone (without direct plasmonic excitation) can increase the DC of the photopolymer by 6-15%. This increase was found to be similar to what could be achieved with a control heat treatment of 30 min at 180 °C. It was also shown that femtosecond laser impulses (795 nm, 5 mJ pulse energy, 50 fs pulse length, 2.83 Jcm-2 fluence), applied after the photopolymerization under a standard dental curing lamp, can cause a 2-7% increase in the DC of undoped samples, even after thermal pre-treatment. The best DC values (12-15% increase) were obtained with combined nanorod doping and subsequent laser irradiation close to the plasmon resonance peak of the nanorods (760-800 nm), which proves that the excited plasmon field can directly facilitate double bond breakage (without thermoplasmonic effects due to the short pulse length) and increase the crosslink density independently from the initial photopolymerization process.

Non-Additive Entropy Composition Rules Connected with Finite Heat-Bath Effects.

Mathematical generalizations of the additive Boltzmann-Gibbs-Shannon entropy formula have been numerous since the 1960s. In this paper we seek an interpretation of the Rényi and Tsallis q-entropy formulas single parameter in terms of physical properties of a finite capacity heat-bath and fluctuations of temperature. Ideal gases of non-interacting particles are used as a demonstrating example.

Kaniadakis Entropy Leads to Particle-Hole Symmetric Distribution.

We discuss generalized exponentials, whose inverse functions are at the core of generalized entropy formulas, with respect to particle-hole (KMS) symmetry. The latter is fundamental in field theory; so, possible statistical generalizations of the Boltzmann formula-based thermal field theory have to take this property into account. We demonstrate that Kaniadakis' approach is KMS ready and discuss possible further generalizations.

f-Gintropy: An Entropic Distance Ranking Based on the Gini Index.

We consider an entropic distance analog quantity based on the density of the Gini index in the Lorenz map, i.e., gintropy. Such a quantity might be used for pairwise mapping and ranking between various countries and regions based on income and wealth inequality. Its generalization to f-gintropy, using a function of the income or wealth value, distinguishes between regional inequalities more sensitively than the original construction.

Human epithelial stem cell survival within their niche requires "tonic" cannabinoid receptor 1-signalling-Lessons from the hair follicle.

The endocannabinoid system (ECS) regulates multiple aspects of human epithelial physiology, including inhibition/stimulation of keratinocyte proliferation/apoptosis, respectively. Yet, how the ECS impacts on human adult epithelial stem cell (eSC) functions remains unknown. Scalp hair follicles (HFs) offer a clinically relevant, prototypic model system for studying this directly within the native human stem cell niche. Here, we show in organ-cultured human HFs that, unexpectedly, selective activation of cannabinoid receptor-1 (CB1)-mediated signalling via the MAPK (MEK/Erk 1/2) and Akt pathways significantly increases the number and proliferation of cytokeratin CK15+ or CK19+ human HF bulge eSCs in situ, and enhances CK15 promoter activity in situ. In striking contrast, CB1-stimulation promotes apoptosis in the differentiated progeny of these eSCs (CK6+ HF keratinocytes). Instead, intrafollicular CB1 gene knockdown or CB1 antagonist treatment significantly reduces human HF eSCs numbers and stimulates their apoptosis, while CB1 knockout mice exhibit a reduced bulge eSCs pool in vivo. This identifies "tonic" CB1 signalling as a required survival stimulus for adult human HF eSCs within their niche. This novel concept must be taken into account whenever the human ECS is targeted therapeutically.

Anandamide Concentration-Dependently Modulates Toll-Like Receptor 3 Agonism or UVB-Induced Inflammatory Response of Human Corneal Epithelial Cells.

Photodamage-induced and viral keratitis could benefit from treatment with novel nonsteroid anti-inflammatory agents. Therefore, we determined whether human corneal epithelial cells (HCECs) express members of the endocannabinoid system (ECS), and examined how the endocannabinoid anandamide (AEA, N-arachidonoyl ethanolamine) influences the Toll-like receptor 3 (TLR3) agonism- or UVB irradiation-induced inflammatory response of these cells. Other than confirming the presence of cannabinoid receptors, we show that endocannabinoid synthesizing and catabolizing enzymes are also expressed in HCECs in vitro, as well as in the epithelial layer of the human cornea in situ, proving that they are one possible source of endocannabinoids. p(I:C) and UVB irradiation was effective in promoting the transcription and secretion of inflammatory cytokines. Surprisingly, when applied alone in 100 nM and 10 µM, AEA also resulted in increased pro-inflammatory cytokine production. Importantly, AEA further increased levels of these cytokines in the UVB model, whereas its lower concentration partially prevented the transcriptional effect of p(I:C), while not decreasing the p(I:C)-induced cytokine release. HCECs express the enzymatic machinery required to produce endocannabinoids both in vitro and in situ. Moreover, our data show that, despite earlier reports about the anti-inflammatory potential of AEA in murine cornea, its effects on the immune phenotype of human corneal epithelium may be more complex and context dependent.

Transient Dynamics in the Random Growth and Reset Model.

A mean-field type model with random growth and reset terms is considered. The stationary distributions resulting from the corresponding master equation are relatively easy to obtain; however, for practical applications one also needs to know the convergence to stationarity. The present work contributes to this direction, studying the transient dynamics in the discrete version of the model by two different approaches. The first method is based on mathematical induction by the recursive integration of the coupled differential equations for the discrete states. The second method transforms the coupled ordinary differential equation system into a partial differential equation for the generating function. We derive analytical results for some important, practically interesting cases and discuss the obtained results for the transient dynamics.

Mitochondrial energy metabolism is negatively regulated by cannabinoid receptor 1 in intact human epidermis.

Epidermal energy metabolism is relevant to skin physiology, ageing and photodamage. While selected hormones stimulate epidermal keratinocyte mitochondrial activity, its negative regulation remains unknown. In several cell types, cannabinoid receptor 1 (CB1 ) is expressed both on the cell membrane (cmCB1 ) and on the mitochondrial outer membrane (mtCB1 ), where its stimulation directly suppresses mitochondrial functions. In the current pilot study, we investigated if CB1 is a negative regulator of human epidermal energy metabolism under physiological conditions. Using organ-cultured full-thickness human skin specimens of healthy individuals, we showed that antagonizing the homeostatic CB1 signalling by the administration of the CB1 inverse agonist AM251 increased respiratory chain complex I and II/IV activity. The effect was CB1 -dependent, since the CB1 -selective agonist arachidonyl-2'-chloroethylamide could prevent the effect. Moreover, the phenomenon was also reproduced by siRNA-mediated down-regulation of CB1 . As revealed by the unaltered expression of several relevant markers (TFAM, VDAC1, MTCO1 and NDUFS4), modulation of CB1 signalling had no effect on the epidermal mitochondrial mass. Next, by using immunoelectron microscopy, we found that human epidermal keratinocytes express both cmCB1 and mtCB1 . Finally, by using equipotent extracellularly restricted (hemopressin) as well as cell-permeable (AM251) inverse agonists, we found that mitochondrial activity is most likely exclusively regulated by mtCB1 . Thus, our data identify mtCB1 as a novel negative regulator of keratinocyte mitochondrial activity in intact human epidermis, and raise the question, whether topical therapeutic interventions capable of selectively activating mtCB1 can reduce excessive mitochondrial ROS production resulting from dysregulated mitochondrial activity during skin ageing or photodamage.

Synthesis and antiproliferative activity of 6-naphthylpterocarpans.

The Heck-oxyarylation of racemic 2-(1-naphthyl)- and 2-(2-naphthyl)-2H-chromene derivatives were carried out resulting diastereoselectively in (6S*,6aR*,11aR*)-6-(1-naphthyl)- and 6-(2-naphthyl)-pterocarpans as major products and bridged (6R*,12R*)-6,12-methanodibenzo[d,g][1,3]dioxocine derivatives as minor products. Antiproliferative activity of two 6-naphthylpterocarpans was identified by MTT assay against A2780 and WM35 human cancer cell lines with low micromolar IC50 values. The measured 0.80 and 3.51 µM IC50 values of the (6S*,6aR*,11aR*)-6-(1-naphthyl)pterocarpan derivative with 8,9-methylenedioxy substitution represent the best activities in the pterocarpan family. Enantiomers of the pterocarpan and dioxocine derivatives and their chiral 2-naphthylchroman-4-one and 2-naphthyl-2H-chromene precursors were separated by HPLC using chiral stationary phase. HPLC-ECD spectra were recorded and absolute configuration and low-energy solution conformations were determined by TDDFT-ECD calculations. Characteristic ECD transitions of the separated enantiomers were correlated with their absolute configuration.

Small Extracellular Vesicles Isolated from Serum May Serve as Signal-Enhancers for the Monitoring of CNS Tumors.

Liquid biopsy-based methods to test biomarkers (e.g., serum proteins and extracellular vesicles) may help to monitor brain tumors. In this proteomics-based study, we aimed to identify a characteristic protein fingerprint associated with central nervous system (CNS) tumors. Overall, 96 human serum samples were obtained from four patient groups, namely glioblastoma multiforme (GBM), non-small-cell lung cancer brain metastasis (BM), meningioma (M) and lumbar disc hernia patients (CTRL). After the isolation and characterization of small extracellular vesicles (sEVs) by nanoparticle tracking analysis (NTA) and atomic force microscopy (AFM), liquid chromatography -mass spectrometry (LC-MS) was performed on two different sample types (whole serum and serum sEVs). Statistical analyses (ratio, Cohen's d, receiver operating characteristic; ROC) were carried out to compare patient groups. To recognize differences between the two sample types, pairwise comparisons (Welch's test) and ingenuity pathway analysis (IPA) were performed. According to our knowledge, this is the first study that compares the proteome of whole serum and serum-derived sEVs. From the 311 proteins identified, 10 whole serum proteins and 17 sEV proteins showed the highest intergroup differences. Sixty-five proteins were significantly enriched in sEV samples, while 129 proteins were significantly depleted compared to whole serum. Based on principal component analysis (PCA) analyses, sEVs are more suitable to discriminate between the patient groups. Our results support that sEVs have greater potential to monitor CNS tumors, than whole serum.

Gintropy: Gini Index Based Generalization of Entropy.

Entropy is being used in physics, mathematics, informatics and in related areas to describe equilibration, dissipation, maximal probability states and optimal compression of information. The Gini index, on the other hand, is an established measure for social and economical inequalities in a society. In this paper, we explore the mathematical similarities and connections in these two quantities and introduce a new measure that is capable of connecting these two at an interesting analogy level. This supports the idea that a generalization of the Gibbs-Boltzmann-Shannon entropy, based on a transformation of the Lorenz curve, can properly serve in quantifying different aspects of complexity in socio- and econo-physics.