Impaired incorporation of fibronectin into the extracellular matrix during aging exacerbates the senescent state of dermal cells.

Fibronectin (Fn) is a ubiquitous extracellular matrix (ECM) glycoprotein that acts as an ECM scaffold organizer and is essential in many biological functions, including tissue repair, differentiation or cancer dissemination. Evidence suggests that the amount of Fn changes during aging. However, how these changes influence the aging process remains unclear. This study aims to understand Fn influence on cell aging. First, we assess the relative level of Fn abundance in both different biopsies of skin donors and replicative senescence cellular model. In skin biopsies, we observed that Fn level decreases with aging in the reticular dermis, while its expression remains relatively stable in the papillary dermis, likely to sustain the dermis-epidermis junction. During replicative senescence, in BJ skin fibroblasts, while intracellular Fn increases, we found that secretion and Fn fibrils formation are less effective. Reduced Fn fibrils leads to disorganization of the ECM. This could be explained by the expression of different Fn isoforms observed in the secretome of senescent cells. Surprisingly, the knockdown of Fn delays the onset of senescence while cultivating cells onto a Fn-coated support promotes it. Taken together, these new insights on the role of Fn during aging may emerge new therapeutic strategies on aged-related diseases.

Proteomic and secretomic comparison of young and aged dermal fibroblasts highlights cytoskeleton as a key component during aging.

Crucial for skin homeostasis, synthesis and degradation of extracellular matrix components are orchestrated by dermal fibroblasts. During aging, alterations of component expression, such as collagens and enzymes, lead to reduction of the mechanical cutaneous tension and defects of skin wound healing. The aim of this study was to better understand the molecular alterations underwent by fibroblasts during aging by comparing secretomic and proteomic signatures of fibroblasts from young (<35years) and aged (>55years) skin donors, in quiescence or TGF-stimulated conditions, using HLPC/MS. The comparison of the secretome from young and aged fibroblasts revealed that 16 proteins in resting condition, and 11 proteins after a 24h-lasting TGF-ß1-treatment, were expressed in significant different ways between the two cell groups (fold change>2, p-value <0.05), with a 77% decrease in the number of secreted proteins in aged cells. Proteome comparison between young and aged fibroblasts identified a significant change of 63 proteins in resting condition, and 73 proteins in TGF-ß1-stimulated condition, with a 67% increase in the number of proteins in aged fibroblasts. The majority of the differentially-expressed molecules belongs to the cytoskeleton-associated proteins and aging was characterized by an increase in Coronin 1C (CORO1C), and Filamin B (FLNB) expression in fibroblasts together with a decrease in Cofilin (CFL1), and Actin alpha cardiac muscle 1 (ACTC1) detection in aged cells, these proteins being involved in actin-filament polymerization and sharing co-activity in cell motility. Our present data reinforce knowledge about an age-related alteration in the synthesis of major proteins linked to the migratory and contractile functions of dermal human fibroblasts.

The impact of air pollution on the facial skin of Caucasian women using real-life pollutant exposure measurements.

BACKGROUND: To date, studies examining the effect of air pollution on skin characteristics have relied on regional pollution estimates obtained from fixed monitoring sites. Hence, there remains a need to characterize the impact of air pollution in vivo in real-time conditions. We conducted an initial investigation under real-life conditions, with the purpose of characterizing the in vivo impact of various pollutants on the facial skin condition of women living in Paris over a 6-month period. MATERIALS AND METHODS: A smartphone application linked to the Breezometer platform was used to collect participants' individual exposures to pollutants through the recovery of global positioning system (GPS) data over a 6-month period. Daily exposure to fine particulate matter (PM 2.5 µm and PM 10 µm), pollen, and air quality was measured. Facial skin color, roughness, pore, hydration, elasticity, and wrinkle measurements were taken at the end of the 6-month period. Participants' cumulated pollutant exposure over 6 months was calculated. Data were stratified into two groups (lower vs. higher pollutant exposure) for each pollutant. RESULTS: 156 women (20-60 years-old) were recruited, with 124 women completing the study. Higher PM 2.5 µm exposure was associated with altered skin color and increased roughness under the eye. Higher PM 10 µm exposure with increased wrinkles and roughness under the eye, increased pore appearance, and decreased skin hydration. Exposure to poorer air quality was linked with increased forehead wrinkles and decreased skin elasticity, while higher pollen exposure increased skin roughness and crow's feet. CONCLUSION: This study suggests a potential correlation between air pollution and facial skin in real-life conditions. Prolonged exposure to PM, gases, and pollen may be linked to clinical signs of skin ageing. This study highlights the importance of longer monitoring over time in real conditions to characterize the effect of pollution on the skin.

Effects of psychological stress on the emission of volatile organic compounds from the skin.

Thirty-five women were included in a clinical study to characterize the volatile organic compounds (VOCs) emitted by the skin during exposure to psychological stress. An original silicon-based polymeric phase was used for VOC sampling on the forehead before and after stress induction. Cognitive stress was induced using specialized software that included a chronometer for semantic and arithmetic tasks. Assessment of stress was monitored using a State-trait anxiety inventory questionnaire, analysis of participants' verbal expressions and clinical measurements. Identification and relative quantification of VOCs were performed by gas chromatography-mass spectrometry. Stress induction was validated by a significant increase in state-anxiety as indicated by the questionnaire, modifications in electrodermal activity measurements and the expression of stress verbatims. In parallel, a sebum production increase and a skin pH decrease were observed. A total of 198 VOCs with different potential sources were identified. They were categorized in 5 groups: probable cosmetic composition, VOCs produced by the body or its microbiota, environmental origin, and dietary intake. In our qualitative statistical approach, three VOCs were found to be correlated with stress induction and 14 compounds showed significance in the paired Wilcoxon test. Fatty-acyls derived from lipids were predominantly identified as well as ethylbenzenes.

Road-traffic-related air pollution contributes to skin barrier alteration and growth defect of sensory neurons.

The effects of air pollution on health are gaining increasing research interest with limited data on skin alterations available. It was suggested that air pollution is a trigger factor for sensitive skin (SS). However, this data was based on surveys with a lack of experimental data. SS is related to altered skin nerve endings and cutaneous neurogenic inflammation. TTe present study was to assess the in vitro effect of particulate matter (PM) on epidermis and nerve ending homeostasis. PM samples were collected according to a validated protocol. Reconstructed human epidermis (RHE, Episkin®) was exposed to PM and subsequently the supernatants were transferred to a culture of PC12 cells differentiated into sensory neurons (SN). Cell viability, axonal growth and neuropeptide-release were measured. The modulation of the expression of different inflammatory, keratinocytes differentiation and neurites growth markers was assessed. PM samples contained a high proportion of particles with a size below 1 µm and a complex chemical composition. Transcriptomic and immunohistochemical analyses revealed that PM altered keratinocytes terminal differentiation and induced an inflammatory response. While viability and functionality of the SN were not modified, their outgrowth was significantly decreased after incubation with PM-exposed Episkin® supernatants. This was closely related to the modification of nerve growth factor/semaphorin 3A balance. This study showed that air pollutants have negative effects on keratinocytes and sensory nerve endings including inflammatory responses. These effects are probably involved in the SS pathophysiology and might be involved in inflammatory skin disorders.

In a Preliminary Study on Human Subjects, a Cosmetic Cream Containing a Harungana madagascariensis Plant Extract Induces Similar Anti-Aging Effects to a Retinol-Containing Cream.

Retinoids are the most effective anti-aging ingredients. Yet, their use can result in adverse reactions. Even bakuchiol, a natural functional analog, can lead to contact dermatitis. We previously showed that a Harungana madagascariensis (Lam. ex Poir.) plant extract (HME) presents retinol-like properties in vitro. Therefore, we performed a preliminary analysis of the anti-aging potential of a cream containing HME on 46 subjects. The participants applied a HME cream on half of their face and one forearm. The effects induced were compared to those produced by a retinol cream applied to the contralateral side. Clinical evaluations indicate that the two creams rapidly (28 days) improve wrinkles underneath the eyes, ptosis, color homogeneity, smoothness, plumpness, firmness, and elasticity of the facial skin. The improvement of crow's feet is only significant after 56 days. For all clinical signs, the effects of both creams are indistinguishable. Instrumental measurements of silicon replica from the eye contour region indicate that the wrinkle surface reduction is already noticeable after 28 days with the HME and retinol cream, while it takes 56 days to have a significant depth decrease. Only the retinol cream improves wrinkle length after 56 days. Ultrasound assessment of forearm skin revealed that the HME cream improves superficial dermis density as early as 28 days with further improvement at day 56, timepoint at which improvement is at the limit of significance with the retinol cream. These preliminary results indicate that HME has similar in vivo functional properties to retinol for reducing the severity of aging signs. Future works, including a bona fide clinical study, are needed to confirm these findings.

Endothelial DLL4 Is an Adipose Depot-Specific Fasting Sensor Regulating Fatty Acid Fluxes.

BACKGROUND: Adaptation of fat depots to change in fuel availability is critical for metabolic flexibility and cardiometabolic health. The mechanisms responsible for fat depot-specific lipid sensing and shuttling remain elusive. Adipose tissue microvascular endothelial cells (AT-EC) regulates bidirectional fatty acid fluxes depending on fed or fasted state. How AT-EC sense and adapt to metabolic changes according to AT location remains to be established. METHODS: We combined transcriptional analysis of native human AT-EC together with in vitro approaches in primary human AT-EC and in vivo and ex vivo studies of mice under fed and fasted conditions. RESULTS: Transcriptional large-scale analysis of human AT-EC isolated from gluteofemoral and abdominal subcutaneous AT revealed that the endothelium exhibits a fat depot-specific signature associated with lipid handling and Notch signaling enrichment. We uncovered a functional link between metabolic status and endothelial DLL4 (delta-like canonical notch ligand 4), which decreases with fasting. DLL4 regulates fatty acid uptake through nontranscriptional modulation of macropinocytosis-dependent long chain fatty acid uptake. Importantly, the changes in DLL4 expression, in response to energy transition state, is impaired under obesogenic conditions, an early alteration coinciding with a defect in systemic fatty acid fluxes adaptation and a resistance to weight loss. CONCLUSIONS: DLL4 is a major actor in the adaptive mechanisms of AT-EC to regulate lipid fluxes. It likely contributes to fat depot-dependent metabolism in response to energy transition states. AT-EC alteration with obesity may favor metabolic inflexibility and the development of cardiometabolic disorders.

Modeling of global skin aging indices among Caucasian and Asian women.

BACKGROUND: The skin aging process is defined as the gradual degradation of several skin properties such as firmness, color, or the appearance of wrinkles. These properties can be assessed by trained experts, who perform an overall evaluation of the entire face. OBJECTIVE: The objective of this paper is the construction of two Global Skin Aging Indices specifically designed to model the overall skin aging process of Caucasian and Asian women. METHODS: Two hundred forty Asian women and 129 Caucasian women aged between 20 and 60 years old are recruited. Parameters related to wrinkles, sagging, elasticity, and skin tone are measured (clinically or instrumentally). The global skin aging index is defined as the normalized projection on the first principal component of a principal component analysis of the skin measurements. Then, linear regressions are performed between the indices and age of both panels. RESULTS: The first principal component carries around 50% of the initial variance for both indices. Both Global Skin Aging Indices statistically correlate with age (R2 ≥ 0.7, p-value < 0.05). An equation linking the indices with age is computed. CONCLUSION: The proposed indices are good indicators of the overall aging process for Caucasian and Asian women. They offer new approaches to assess antiaging product efficacy.

A Harungana madagascariensis extract with retinol-like properties: Gene upregulations and protein expressions in human fibroblasts and skin explants.

OBJECTIVE: Because they limit, even reverse, age-induced skin alterations, retinoids became a staple in cosmetology. However, their use can result in undesired secondary effects and there is a demand for natural sources of compounds with retinoid-like effects. A preliminary screening identified a Harungana madagascariensis plant extract (HME) as possibly inducing genes stimulated by retinol. We analysed its effect on gene and protein expression, comparing it to retinoids. METHODS: Gene expression was analysed by real-time qPCR on RNA from isolated fibroblasts subjected to retinol or the plant extract for 6, 48 or 96 h. Skin markers were quantified in fibroblasts cultured with retinol or extract containing medium, and UV-aged skin explants subjected to topical applications of creams containing retinol, retinaldehyde or HME. RESULTS: Real-time qPCR shows that the extract induced all RARs and RXRs, even RXRγ that was not induced by retinol. Eighty-eight per cent of the 25 early retinoid reaction genes induced by a concentration of retinol are induced by the extract. In fibroblasts, only the extract increased collagen III labelling, while collagen I and fibronectin labelling are increased by retinol and the extract, with higher levels for the extract. When topically applied to UV-aged skin explants, only the cream containing the HME led to increased labelling of CRABP1 in the epidermis. CRABP2 and Ki67 are induced by all three creams and no effect was detected on RXRs. In the dermisthe extract containing cream increased CRABP2, total collagen, procollagen I and collagen I while creams with retinol or retinaldehyde only affected some of these proteins. CONCLUSIONS: The HME induces an overall retinol-like gene induction profile in isolated fibroblasts and retinoid-like stimulation of protein synthesis in both isolated fibroblasts and photoaged skin explants.

OBJECTIFS: Limitant, voire inversant les altérations cutanées induites par l'âge, les rétinoïdes sont devenus incontournables en cosmétologie. Cependant, leur application topique peut entraîner des effets secondaires indésirables et il existe une demande pour des composés naturels ayant des effets similaires à ceux des rétinoïdes. Un screening préliminaire nous avait permis d'identifier un extrait de la plante Harungana madagascariensis (HME) comme pouvant induire des gènes stimulés par le rétinol. Nous avons donc analysé son effet sur l'expression de gènes et de protéines induits par les rétinoïdes et comparé les résultats à ceux obtenus en présence de rétinoïdes. MÉTHODES: L'expression de gènes a été analysée par qPCR en temps réel réalisée sur l'ARN de fibroblastes isolés soumis au rétinol ou à l'extrait végétal pendant 6, 48 ou 96 heures. Différentes protéines cutanées ont été quantifiés dans des fibroblastes cultivés en présence de rétinol ou d'un milieu contenant l'extrait. Des quantifications ont également été faites sur des explants de peau vieillie par les UV et soumis à des applications topiques de crèmes contenant du rétinol, du rétinaldéhyde ou le HME. RESULTATS: La qPCR en temps réel montre que l'extrait induit tous les gènes RARs et RXRs, même RXRγ qui n'était pas induit par le rétinol. Quatre-vingt-huit pour cent des 25 gènes impliqués dans la réaction précoce aux rétinoïdes induits par une concentration de rétinol ont été induits par l'extrait. Dans les fibroblastes, seul l'extrait a augmenté le marquage du collagène III, tandis que le marquage du collagène I et de la fibronectine a été augmenté par le rétinol et l'extrait, avec des niveaux plus élevés pour l'extrait. En application topique sur des explants de peau vieillie par les UV, seule la crème contenant le HME a entraîné une augmentation du marquage de CRABP1 dans l'épiderme. CRABP2 et Ki67 ont été induits par les trois crèmes et aucun effet n'a été détecté sur les RXRs. Dans le derme, la crème contenant l'extrait a augmenté CRABP2, le collagène total, le procollagène I et le collagène I, tandis que les crèmes contenant du rétinol ou du rétinaldéhyde n'ont affecté que certaines de ces protéines. CONCLUSIONS: Chez les fibroblastes isolés, le HME induit un profil d'induction génique globalement similaire à celui du rétinol. Chez les fibroblastes isolés et des explants de peau photo-vieillie, il entraine une stimulation de la synthèse protéique similaire à celle des rétinoïdes.

In elderly Caucasian women, younger facial perceived age correlates with better forearm skin microcirculation reactivity.

BACKGROUND: Visual and molecular changes occurring upon aging are rather well characterized. Still, aging signs show great significant inter-individual variations, and little is known concerning the link between perceived age and cutaneous microcirculation. MATERIALS AND METHODS: To investigate this point, we recruited Caucasian women in their mid-50's to mid-70's and subsampled women looking older or younger than their age. We studied their facial skin color, as well as their microvascular reactivity to local heating assessed in the forearm skin. We also used skin biopsies from some of these women for gene expression or immunohistochemical analysis. RESULTS: Clinical and instrumental analysis of skin color revealed that subjects who look 5 years younger differ only by a higher glowing complexion. Our most striking result is that subjects looking 5 years younger than their age present a higher microcirculation reactivity in forearm skin. Transcriptome comparison of skin samples from women looking older or younger than their age revealed 123 annotated transcripts differentially expressed, among which MYL9 relates to microcirculation. MYL9 is downregulated in the group of women looking younger than their real age. Microscopy shows that the labeling of MYL9 and CD31 are altered and heterogeneous with age, as is the morphology of microvessels. CONCLUSION: Therefore, assessing generalized vascular reactivity in non-photo-exposed skin to focus on the intrinsic aging allows subtle discrimination of perceived age within elderly healthy subjects.

Visible characteristics and structural modifications relating to enlarged facial pores.

BACKGROUND: Many factors relate to enlargement and visibility of facial pores. However, how enlargement relates to modification of the skin structure is still poorly analysed. Therefore, our aim was to characterise the visible features and the micro-environment of facial pores. MATERIALS AND METHODS: We recruited 160 women, half of them with normal-size facial pores, the other half presenting enlarged pores. We analysed their right cheek using standardised photography, in vivo reflectance microscopy and optical coherence tomography. RESULTS: If the area of pores is indeed the most differentiating visible characteristic, enlarged facial pores are also denser and more contrasted. At the structural level, enlarged facial pores present a more regular organisation of keratinocytes, an irregular dermo-epidermal junction and a higher attenuation of collagen between the surface and deeper layers. Still, the most relevant alterations of enlarged facial pores are the presence of bright tubular structures surrounding hair follicles and enlarged hair shafts. CONCLUSION: These observations suggest that alterations of the stroma could result in altered skin tensions, leading to the dilation of the infundibulum and enlarged hair shaft. The modified tension would also explain the more regular organisation of the keratinocytes and the irregular dermo-epidermal junction.

Influence of the environmental relative humidity on the inflammatory response of skin model after exposure to various environmental pollutants.

The skin is an essential barrier, protecting the body against the environment and its numerous pollutants. Several environmental pollutants are known to affect the skin, inducing premature aging through mechanisms including oxidative stress, inflammation, and impairment of skin functions. Even climate conditions can impact the skin. Therefore, using a Reconstructed Human Epidermis (RHE), we tested the effect of two samples of fine particulate matters (PM0.3-2.5 - one metals-rich sample and the other organic compounds-rich), two Volatile Organic Compounds mixtures (VOCs - from a solvent-based paint and a water-based paint) and Tobacco Smoke (TS). All pollutants affected cellular functionality, but to a lesser extent for the water-based paint VOC. This effect was enhanced when RHE were preconditioned for 2 h by a semi-dry airflow (45% relative humidity) before pollutants application, compared to preconditioning by a humid airflow (90% relative humidity). In the absence of preconditioning, IL-1α, IL-6, IL-8, and RANTES were almost systematically induced by pollutants. When RHE were preconditioned by a semi-dry or humid airflow before being subjected to pollutants, the increase of IL-1α, IL-8, and RANTES falls into two groups. Similarly to RHE not treated with pollutants, RHE treated with VOCs after preconditioning by a semi-dry airflow showed increased IL-1α, IL-8, and RANTES release. On the contrary, RHE treated with PM or TS after preconditioning by a semi-dry airflow show a lower increase in IL-1α, IL-8, and RANTES compared to preconditioning by a humid airflow. The effect of real environmental relative humidity conditions of the air, combined with acute exposure to various environmental pollutants, seemed to relate mainly to structural changes of the skin, determining the outcome of the inflammatory response depending on the physicochemical characteristics of pollutants.

Longevity strategies in response to light in the reef coral Stylophora pistillata.

Aging is a multifactorial process that results in progressive loss of regenerative capacity and tissue function while simultaneously favoring the development of a large array of age-related diseases. Evidence suggests that the accumulation of senescent cells in tissue promotes both normal and pathological aging. Oxic stress is a key driver of cellular senescence. Because symbiotic long-lived reef corals experience daily hyperoxic and hypoxic transitions, we hypothesized that these long-lived animals have developed specific longevity strategies in response to light. We analyzed transcriptome variation in the reef coral Stylophora pistillata during the day-night cycle and revealed a signature of the FoxO longevity pathway. We confirmed this pathway by immunofluorescence using antibodies against coral FoxO to demonstrate its nuclear translocation. Through qPCR analysis of nycthemeral variations of candidate genes under different light regimens, we found that, among genes that were specifically up- or downregulated upon exposure to light, human orthologs of two "light-up" genes (HEY1 and LONF3) exhibited anti-senescence properties in primary human fibroblasts. Therefore, these genes are interesting candidates for counteracting skin aging. We propose a large screen for other light-up genes and an investigation of the biological response of reef corals to light (e.g., metabolic switching) to elucidate these processes and identify effective interventions for promoting healthy aging in humans.

Coupling of melanocyte signaling and mechanics by caveolae is required for human skin pigmentation.

Tissue homeostasis requires regulation of cell-cell communication, which relies on signaling molecules and cell contacts. In skin epidermis, keratinocytes secrete factors transduced by melanocytes into signaling cues promoting their pigmentation and dendrite outgrowth, while melanocytes transfer melanin pigments to keratinocytes to convey skin photoprotection. How epidermal cells integrate these functions remains poorly characterized. Here, we show that caveolae are asymmetrically distributed in melanocytes and particularly abundant at the melanocyte-keratinocyte interface in epidermis. Caveolae in melanocytes are modulated by ultraviolet radiations and keratinocytes-released factors, like miRNAs. Preventing caveolae formation in melanocytes increases melanin pigment synthesis through upregulation of cAMP signaling and decreases cell protrusions, cell-cell contacts, pigment transfer and epidermis pigmentation. Altogether, we identify that caveolae serve as molecular hubs that couple signaling outputs from keratinocytes to mechanical plasticity of pigment cells. The coordination of intercellular communication and contacts by caveolae is thus crucial to skin pigmentation and tissue homeostasis.

Age-Induced Reduction in Human Lipolysis: A Potential Role for Adipocyte Noradrenaline Degradation.

Gao et al. report that the observed reduction in adipose lipolysis with age in women could be explained by an upregulation of the catecholamine-degradation pathway in subcutaneous adipocytes. However, in contrast to findings in mice, these pathways are enriched in adipocytes and not in immune cells, suggesting species-specific differences in aging mechanisms.

Prospective analyses of white adipose tissue gene expression in relation to long-term body weight changes.

BACKGROUND: Transcriptome analysis of abdominal subcutaneous white adipose tissue (sWAT) has identified important obesity-associated disturbances. However, the relation between sWAT transcriptome and long-term future changes in body weight remains elusive. OBJECTIVE: To investigate sWAT transcriptome signatures before and after long-term weight changes and assess their predictive value for body weight changes. DESIGN: A total of 56 women were followed longitudinally and subdivided into weight-stable (WS, n = 25), weight-gaining (WG, n = 14) and weight-losing (WL, n = 17) groups between baseline and follow-up (13 ± 1 years). The fasting sWAT transcriptome was analyzed by gene microarray at baseline and follow-up. Key genes associated with weight changes were validated using quantitative real-time PCR. RESULTS: In total 285 transcripts exhibited difference (FDR < 30%) in expression fold change over time between WL and WS women. WL women displayed decreased pro-inflammatory (NLRP3) but increased insulin-response gene (FASN and GLUT4) expression over time. In comparison, 461 transcripts displayed difference in expression fold change over time between WG and WS women (P < 0.05). Genes involved in autophagic processes (CDK5, SQSTM1 and FBXL2) were generally upregulated in WG women. At baseline, 307 and 302 transcripts were differentially expressed (FDR < 30%) in WL and WG women, respectively, when independently compared against WS women. Baseline expression of adipogenic and lipogenic genes (PPARG, IRS2 and HACD2) was lower, while pro-fibrotic (COL6A1) was higher, in WL than WS women; whereas protein processing genes were lower expressed in WG than in WS women. CONCLUSION: In adult women, long-term body weight change associates with altered sWAT transcriptome. Expression of genes associated with inflammation, insulin response, adipogenesis and lipogenesis are linked to weight loss. However, other pathways such as autophagy not only associate but also predict future weight gain suggesting that intrinsic factors in sWAT impact tissue expansion.

An in vitro model to evaluate the impact of environmental fine particles (PM0.3-2.5) on skin damage.

Exposure to airborne particulate matter (PM) has significant effects on human health mainly leading to cardio-respiratory diseases. However very few data are available regarding the impact of PM on the skin, so to better understand the impact of fine particle (PM0.3-2.5) on both inflammatory response and epidermal structure, we exposed a reconstructed human epidermis (RHE) to several doses of PM collected in Cotonou (Benin, West Africa). After 24 h of exposure, inflammatory response, histological observations, and gene expression related to oxidative stress, antioxidant defense and structural damages were determined. No PM-linked changes in tissue morphology or membrane integrity were observable. PM was however cytotoxic in a dose dependent manner. An inflammatory response appeared as shown by the increase in IL-1α and IL-8 cytokine productions. PM also induced oxidative stress, leading to an increase in 4-HNE immunostaining and to the up-regulation of HMOX1, MT1G and MT1E. Finally, PM had a negative impact on fundamental skin functions such as tissue anchorage, cell differentiation, cornification / skin desquamation and apoptosis. Our data show that airborne fine particles have an adverse effect on skin integrity, most probably leading to accelerated ageing.

Non-senescent keratinocytes organize in plasma membrane submicrometric lipid domains enriched in sphingomyelin and involved in re-epithelialization.

Membrane lipid raft model has long been debated, but recently the concept of lipid submicrometric domains has emerged to characterize larger (micrometric) and more stable lipid membrane domains. Such domains organize signaling platforms involved in normal or pathological conditions. In this study, adhering human keratinocytes were investigated for their ability to organize such specialized lipid domains. Successful fluorescent probing of lipid domains, by either inserting exogenous sphingomyelin (BODIPY-SM) or using detoxified fragments of lysenin and theta toxins fused to mCherry, allowed specific, sensitive and quantitative detection of sphingomyelin and cholesterol and demonstrated for the first time submicrometric organization of lipid domains in living keratinocytes. Potential functionality of such domains was additionally assessed during replicative senescence, notably through gradual disappearance of SM-rich domains in senescent keratinocytes. Indeed, SM-rich domains were found critical to preserve keratinocyte migration before senescence, because sphingomyelin or cholesterol depletion in keratinocytes significantly alters lipid domains and reduce migration ability.

Repeated short climatic change affects the epidermal differentiation program and leads to matrix remodeling in a human organotypic skin model.

Human skin is subject to frequent changes in ambient temperature and humidity and needs to cope with these environmental modifications. To decipher the molecular response of human skin to repeated climatic change, a versatile model of skin equivalent subject to "hot-wet" (40°C, 80% relative humidity [RH]) or "cold-dry" (10°C, 40% RH) climatic stress repeated daily was used. To obtain an exhaustive view of the molecular mechanisms elicited by climatic change, large-scale gene expression DNA microarray analysis was performed and modulated function was determined by bioinformatic annotation. This analysis revealed several functions, including epidermal differentiation and extracellular matrix, impacted by repeated variations in climatic conditions. Some of these molecular changes were confirmed by histological examination and protein expression. Both treatments (hot-wet and cold-dry) reduced the expression of genes encoding collagens, laminin, and proteoglycans, suggesting a profound remodeling of the extracellular matrix. Strong induction of the entire family of late cornified envelope genes after cold-dry exposure, confirmed at protein level, was also observed. These changes correlated with an increase in epidermal differentiation markers such as corneodesmosin and a thickening of the stratum corneum, indicating possible implementation of defense mechanisms against dehydration. This study for the first time reveals the complex pattern of molecular response allowing adaption of human skin to repeated change in its climatic environment.

In vitro model adapted to the study of skin ageing induced by air pollution.

More than a barrier against environmental agents, skin reflects individual health and is a visible sign of ageing with the progressive loss of skin integrity. In order to evaluate the consequences of an environmental complex mixture, with tobacco smoke (TS) as model, on cellular and morphological changes, a 3D skin model was used. Morphologically, tissue integrity was intact after one TS-exposure while the superficial layers were drastically reduced after two TS-exposures. However, TS modified epidermal organisation at the molecular level after just one exposure. A decrease in loricrin protein staining was showed in the epidermis, while production of inflammatory cytokines (IL-8, IL-1α, IL-18) and metalloproteinase (MMP-1, MMP-3) were stimulated. Oxidative stress was also illustrated with an increase in 4-HNE protein staining. Moreover, terminal differentiation, cell-cell junction and anchorage gene expression was down-regulated in our model after one TS-exposure. In conclusion, tobacco smoke impacted the fundamental functions of skin, namely tissue anchorage, cornification and skin desquamation. Oxidative stress resulted in skin ageing. The tissue was even reactive with the inflammatory pathways, after one TS-exposure. The 3D-RHE model is appropriate for evaluating the impact of environmental pollutants on skin ageing.