CCN proteins as potential actionable targets in scleroderma

Systemic sclerosis (SSc) is a complex autoimmune connective tissue disease combining inflammatory, vasculopathic and fibrotic manifestations. Skin features, which give their name to the disease and are considered as diagnostic as well as prognostic markers, have not been thoroughly investigated in terms of therapeutic targets. CCN proteins (CYR61/CCN1, CTGF/CCN2, NOV/CCN3 and WISP1â 2â 3 as CCN4â 5â 6) are a family of secreted matricellular proteins implicated in major cellular processes such as cell growth, migration, differentiation. They have already been implicated in key pathophysiological processes of SSc, namely fibrosis, vasculopathy and inflammation. In this review, we discuss the possible implication of CCN proteins in SSc pathogenesis, with a special focus on skin features, and identify the potential actionable CCN targets.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/147777/1/exd13806.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/147777/2/exd13806_am.pd

Xeroderma pigmentosum: clues to understanding cancer initiation

AbstractXeroderma pigmentosum (XP) type C is a rare autosomal recessive disorder that occurs because of inactivation of the xeroderma pigmentosum group C (XPC) protein, which is an important DNA damage recognition protein involved in DNA nucleotide excision repair (NER). This defect, which prevents removal of a wide array of direct and indirect DNA lesions, is associated with a decrease in catalase activity. As a novel photoprotective approach, lentivirus-mediated catalase overexpression in XPC human keratinocytes results in a marked decrease in sunburn cell formation, caspase-3 activation, and p53 accumulation following UVB irradiation. While not correcting the gene defect, indirect gene therapy using antioxidant enzymes may be helpful in limiting photosensitivity in XP type C, as well as in other monogenic/polygenic photosensitive disorders characterized by reactive oxygen species (ROS) accumulation. Hypoxia-inducible factor-1 (HIF-1), a major transcription factor sensitive to oxygen levels, responds to various stress factors. As a common stressor of skin, UVB induces a biphasic HIF-1a variation through ROS generation in keratinocytes. HIF-1a has an important regulator effect on the expression of XPC protein and other NER genes, indicating indirect regulation of NER by ROS. The intrinsic genomic instability arising in XP type C provides a good opportunity to investigate the complex molecular mechanisms underlying the Warburg effect (the shift of mito-chondrial metabolism towards glycolysis). Overall, the monogenic disorder XP type C is a powerful tool for studying photoprotection and cancer

Rôle des mélanocytes dans l'unité épidermique de mélanisation reconstruite ex-vivo après une irradiation UV aiguë

Influence of melanocytes in skin pigmentation is well documented, however its photoprotective role is controvert. Role of melanocytes have been investigated on reconstructed epidermis with 100 % of keratinocytes or 95 % of keratinocytes and 5 % of melanocytes. In a first time, effect of an acute UVB dose has been studied on both reconstructs, next we have investigated UVA and UVA+B effects on these epidermis. Following irradiation, presence of melanocytes in reconstructed epidermis protects against apoptosis without protects significantly against DNA damage formation (CPD, 6-4PP) and protects against UV-induced unbalance of the SOD/catalase ratio (antioxidants enzymes). On the contrary, presence of melanocytes in reconstructed epidermis amplifies lipids and proteins oxidations but seems to protect against DNA oxidations. Melanocytes differ from keratinocytes by their melanin content and their more important concentration in polyunsaturated fatty acids. To evaluate what is the part of melanin and the part of polyunsaturated fatty acids in epidermal UV responses, reconstructed epidermis with keratinocytes have been supplemented with polyunsaturated fatty acid. This study indicates that polyunsaturated fatty acids are responsible for lipids and proteins oxidations and that melanin protect against DNA oxidation induced by lipoperoxidation. All these studies demonstrate that, model of reconstructed epidermis and epidermis in-vivo have the same behaviour following UV irradiation. In a last time, sunscreens and antioxidants have been tested on reconstructed epidermis and have demonstrated that model of reconstructed epidermis is suitable for photoprotective molecules screening.Le rôle du mélanocyte dans la pigmentation de la peau n'est plus a démontré, par contre son rôle photoprotectif est controversé. Le rôle du mélanocyte a été étudié en comparant des épidermes reconstruits avec 100 % de kératinocytes et des épidermes reconstruits avec 95 % de kératinocytes et 5 % de mélanocytes. Dans un premier temps, l'effet d'une irradiation UVB aiguë a été étudié sur ces deux types de reconstructions, puis l'étude a été élargie aux effets des UVA et des UVA+B. Ces études ont permis de montrer, qu'après irradiation, la présence de mélanocytes au sein de l'épiderme reconstruit prévient l'apoptose sans pour autant protéger de façon significative de la formation des lésions directes de l'ADN (CPD et 6-4PP) et permet le maintien du rapport SOD/Catalase (principales enzymes antioxydantes). Par contre, la présence de mélanocytes au sein de l'épiderme amplifie les oxydations lipidiques et protéiques UV-induites mais semble prévenir l'oxydation de l'ADN. Les mélanocytes possèdent en plus de la mélanine, une plus grande concentration en acides gras polyinsaturés membranaires que les kératinocytes. Afin d'estimer quelle est la part de la mélanine et quelle est celle des acides gras polyinsaturés mélanocytaires dans les réponses UV-induites, des épidermes reconstruits avec des kératinocytes ont été suppléméntés avec des acides gras polyinsaturés. Cette étude a permis de mettre en évidence que ce sont les acides gras qui induisent l'amplification des oxydations lipidiques et protéiques alors que la mélanine protège l'ADN de l'oxydation induite par la lipoperoxydation. Ces différentes études ont également permis de montrer que face aux UV, l'épiderme reconstruit se comporte de façon similaire à l'épiderme normal in-vivo. Dans un dernier temps, des crèmes solaires et des antioxydants systémiques ont été testés et ont permis de confirmer que le modèle d'épiderme reconstruit est tout à fait adapté au test de molécules photoprotectrices

Les molécules d'adhésion CCN3 et DDR1 au cours du vitiligo

Le vitiligo généralisé est une leucodermie acquise qui touche 0,5 à 1% de la population mondiale et qui résulte d une perte progressive des mélanocytes.Le mécanisme à l origine de la perte des mélanocytes au cours du vitiligo reste obscur et la destruction des mélanocytes n a jamais été observée.De nombreuses hypothèses ont été avancées pour expliquer la disparition des mélanocytes : une susceptibilité génétique, l auto-immunité, la théorie neurale et la théorie oxydative.Notre équipe a développé une théorie intégrée qui reprend ces différents mécanismes. Cette théorie considère que le vitiligo est lié au détachement des mélanocytes et à leur élimination à travers l épiderme ou mélanocytorrhagie. Dans cette théorie, un défaut d adhésion des mélanocytes est le facteur prédisposant au vitiligo.L interaction des mélanocytes avec les kératinocytes environnants et avec la membrane basale est médiée par les intégrines et les cadhérines. L expression de l intégrine et de la E-cadhérine n est pas modifiée dans le vitiligo généralisé.En 2006, Fukunaga-Kalabis et al. montrent que l attachement des mélanocytes à la membrane basale est en partie dû à DDR1 qui est sous le contrôle de la protéine CCN3. Ils ont observé que l inhibition de CCN3 induit le détachement des mélanocytes.Récemment, des variants génétiques de DDR1 ont été observés chez des patients d origine ethnique différente atteints de vitiligo. Nous avons décidé d étudier d une part l expression de CCN3 au niveau de la peau lésionnelle et non lésionnelle de patients atteints de vitiligo et d autre part l impact de l inhibition de CCN3 et de DDR1 au niveau de mélanocytes utilisés pour des reconstructions épidermiques. Nos résultats in vitro et in vivo suggèrent que CCN3 est impliqué dans la physiopathologie du vitiligo.Common generalized vitiligo is an acquired hypopigmentation which is found in 0, 5-1% of individuals world-wide and which results in progressive loss of melanocytes.The mechanism underlying the elimination of melanocytes in vitiligo remains unclear and melanocyte destruction has never been clearly demonstrated in non segmental vitiligo. Various hypotheses have been put forward to explain the disappearance of melanocytes in vitiligo: genetic susceptibility, autoimmunity, neural and impared redox status. We previously proposed a new theory that integrates those pathomechanisms. This theory considers vitiligo as a disease caused by the chronic detachment and transepidermal loss of melanocytes named melanocytorrhagy. In this theory, the defective adhesion of melanocytes is the predisposing factor.Interactions between melanocytes and the basement membrane are mediated by integrins and interactions between melanocytes and keratinocytes are mediated by cadherins in association with b-catenin. But integrin expression is not affected in NSV and a normal expression of E-cadherin in lesional and non lesional vitiligo skin is observed.In 2006, Fukunaga-Kalabis et al. reported that attachment of melanocytes to basal lamina is in part due to DDR1, which is under control of CCN3. They have observed that inhibition of CCN3 induces the detachment of melanocytes.Recently, DDR1 genetics variants have been associated with vitiligo in patients of different ethnic origin. We have decided to study in parallel the expression of CCN3 and DDR1 in lesional and non lesional skin of vitiligo patients and the impact of inhibition of CCN3 and DDR1 in melanocytes on their behaviour in reconstructed epidermis.In conclusion, our in vivo and in vitro data suggest that CCN3 is implicated in vitiligo etiology.BORDEAUX2-Bib. électronique (335229905) / SudocSudocFranceF

Transport of hydrocortisone in targeted layers of the skin by multi-lamellar liposomes

Hydrocortisone (HyC), a hydrophobic pharmaceutical active, was encapsulated in multi-lamellar liposomes (MLLs) composed of P100, a mixture of phospholipids, and Tween (R) 80. Three different HyC-loaded formulations were designed to target the stratum corneum, the living epidermis and the hypodermis. The impact of encapsulation on their size, elasticity and zeta potential, the three key factors controlling MLLs skin penetration, was studied. Raman mapping of phospholipids and HyC allowed the localisation of both components inside an artificial skin, Strat-M (R), demonstrating the efficiency of the targeting. Percutaneous permeation profiles through excised human skin were performed over 48 h, supporting results on artificial skin. Their modelling revealed that HyC encapsulated in MLLs, designed to target the stratum corneum and living epidermis, exhibited a non-Fickian diffusion process. In contrast, a Fickian diffusion was found for HyC administered in solution, in a pharmaceutical cream and in transdermal MLLs. These results allowed us to propose a mechanism of interaction between HyC-containing MLLs and the skin

Prediction of the penetration depth of multi-lamellar liposomes in artificial skin. Application to the vectorization of corticosteroid in human skin

Our previous work showed that the size, elasticity and charge of multi-lamellar liposomes (MLLs) could not be considered separately to predict the fate of MLLs in the skin [1]. Based on this study, we developed several MLLs formulations containing a corticosteroid, betamethasone 17-valerate (B17) to transport the drug into the stratum corneum, living epidermis, dermis or through the skin. MLLs encapsulation efficiency was found to exceed 74 +/- 3 % in all cases. In addition, we showed that MLLs protected the corticosteroid from thermal degradation. Comparing the penetration depth of all MLLs within artificial skin measured by Raman imaging, we established an equation for its determination, given the MLLs elasticity and size. This equation was verified experimentally on human explants: quantification of B17 in each skin layer, as well as its transdermal passage by ultra-high performance liquid chromatography, confirmed that B17 was predominantly and significantly transported in the desired layer. Eventually, we showed the benefits in using B17-loaded MLLs instead of a B17-containing pharmaceutical cream in terms of B17 penetration and thermal degradation

Journal of Drug Delivery Science and Technology

Trans-Resveratrol (RV) was encapsulated in multi-lamellar liposomes (MLLs) composed of a mixture of phosphatidylcholine (P100), Tween 80 (T80) and water. The P100-to-T80 ratio as well as their water content were chosen based on a previous study to promote transdermal transport of RV. The effect of RV on the size, elasticity, and charge of MLLs was evaluated as well as the effect of encapsulation on the apparent solubility of RV. The diffusion of RV and MLLs in artificial skin, namely Strat-M™, was monitored by confocal Raman imaging, and compared with that of a RV solution and empty MLLs, while the transdermal passage rate was measured by UV–vis spectrophotometry on both artificial and excised human skin. RV was found to remain localized in the outer layer of the skin with less than 3% passing through it over a 24-h period in both skin types. Encapsulation in MLLs drastically increased its transdermal passage: 73 ± 10% after 3 h of incubation on excised human skin, and 10% on Strat-M™ after 9 h. Whereas RV in its free form underwent cis isomerization, MLLs protected it up to 9h before undergoing chemical degradation after 24h

The Relation Between Constitutional Skin Color and Photosensitivity Estimated from UV-Induced Erythema and Pigmentation Dose-Response Curves

In 54 healthy volunteers we assessed predictors of sensitivity to ultraviolet (UV) light, including Fitzpatrick's sun reactive skin types and constitutional skin color, and compared these with one another and with responses of the skin to UV irradiation, as determined experimentally by a minimal erythema dose (MED), a minimal melanogenic dose (MMD), and dose-response curves for UV-induced erythema and pigmentation. For these studies, a xenon arc solar simulator was used as the source of UV irradiation, and a chromameter interfaced with a computer for objective measurement of UV-induced erythema and pigmentation was employed. The skin type did not correspond well to the constitutional skin color, as measured by a chromameter prior to UV irradiation. Within each skin type, there were large ranges of MED and MMD values and great variability in the shapes of the dose-response curves. Constitutional skin color was also not a good predictor of the measured MED and MMD values but did appear to correlate with the steepness of the dose-response curves for erythema and for pigmentation. From these studies, we propose that objectively measured constitutional skin color is a better predictor of UV responses of the skin than skin type and that steepness of dose-response curves for erythema is a better measure of the response of the skin to UV irradiation than is a MED measurement

Is Hyperpigmentation in Systemic Sclerosis a Perivascular Dermal Tattoo?

International audienc

Tspan8 Drives Melanoma Dermal Invasion by Promoting ProMMP-9 Activation and Basement Membrane Proteolysis in a Keratinocyte-Dependent Manner

International audienceMelanoma is the most aggressive skin cancer with an extremely challenging therapy. The dermal-epidermal junction (DEJ) degradation and subsequent dermal invasion are the earliest steps of melanoma dissemination, but the mechanisms remain elusive. We previously identified Tspan8 as a key actor in melanoma invasiveness. Here, we investigated Tspan8 mechanisms of action during dermal invasion, using a validated skin-reconstruct-model that recapitulates melanoma dermal penetration through an authentic DEJ. We demonstrate that Tspan8 is sufficient to induce melanoma cells' translocation to the dermis. Mechanistically, Tspan8+ melanoma cells cooperate with surrounding keratinocytes within the epidermis to promote keratinocyte-originated proMMP-9 activation process, collagen IV degradation and dermal colonization. This concurs with elevated active MMP-3 and low TIMP-1 levels, known to promote MMP-9 activity. Finally, a specific Tspan8-antibody reduces proMMP-9 activation and dermal invasion. Overall, our results provide new insights into the role of keratinocytes in melanoma dermal colonization through a cooperative mechanism never reported before, and establish for the first time the pro-invasive role of a tetraspanin family member in a cell non-autonomous manner. This work also displays solid arguments for the use of Tspan8-blocking antibodies to impede early melanoma spreading and therefore metastasis