The Région Languedoc Roussillon is the umbrella organisation for an interconnected and integrated project on active and healthy ageing (AHA). It covers the 3 pillars of the European Innovation Partnership on Active and Healthy Ageing (EIP on AHA): (A) Prevention and health promotion, (B) Care and cure, (C) and (D) Active and independent living of elderly people. All sub-activities (poly-pharmacy, falls prevention initiative, prevention of frailty, chronic respiratory diseases, chronic diseases with multimorbidities, chronic infectious diseases, active and independent living and disability) have been included in MACVIA-LR which has a strong political commitment and involves all stakeholders (public, private, patients, policy makers) including CARSAT-LR and the Eurobiomed cluster. It is a Reference Site of the EIP on AHA. The framework of MACVIA-LR has the vision that the prevention and management of chronic diseases is essential for the promotion of AHA and for the reduction of handicap. The main objectives of MACVIA-LR are: (i) to develop innovative solutions for a network of Living labs in order to reduce avoidable hospitalisations and loss of autonomy while improving quality of life, (ii) to disseminate the innovation. The three years of MACVIA-LR activities are reported in this paper.

The Region Languedoc-Roussillon is the umbrella organisation for an interconnected and integrated project on AHA covering the 3 pillars of the European Innovation Partnership on Active and Healthy Ageing. All sub-activities (A1 : electronic pharmaceutical file, A2: falls prevention initiative, A3: frailty, B3: chronic respiratory diseases, chronic diseases with comorbidities, oral health and hepatitis virus C chronic infection, C2 and 04 active and independent living and handicap) are included in MACVIA-LR that has a strong political commitment and includes all stakeholders (public, private, patients, policy makers). It is one of the Reference Sites of the European Innovation Partnership on Active and Healthy Ageing built around chronic diseases, ageing and handicap. The framework of MACVIA-LR has the vision that the prevention and management of CDs is essential for AHA promotion and for the reduction of handicap. The main objective of MACVIA-LR is to develop innovative solutions for a network of Living Labs in order to improve the care of patients affected by CDs in the Languedoc-Roussillon area and to disseminate the innovation. (C) 2014 Published by Elsevier Masson SAS.

Espeletiinae are plants which grow above 3000 m of altitude in the Northern Andes and kaurenic acid was extracted from the leaves of Coespeletia moritziana. This compound has shown a wide range of biological activities, including cytotoxicity which is efficient in cancer therapy. The percutaneous penetration of this compound was measured in vitro using Franz cells. At appropriate intervals for up to 24 h, diffusion samples were analyzed using HPLC. At the end of the test period, the amount of kaurenic acid was determined in each compartment and approximately 10% of kaurenic acid had been absorbed and was found in the skin layers.

Solid lipid microspheres (SLM), lipid-in-water formulations made from oil-and-wax mixtures, were studied concerning feasibility. SLMs were then loaded with a benzophenone-3, water insoluble UVAB-filter intended for dermal application. Microspheres were prepared by dispersion with homogenisers and investigated by polarizing micrography and scanning electron micrography. For the selected formulations, investigations on percutaneous penetration of B-3 capacity were performed “in vitro” using Franz cells. Microspheres, 5–50 μm in size, and a spherical shape were obtained from several mixtures. B-3 was added and the loading capacity of this drug in the SLM was obtained for a maximum of 5% when the lipophilic phase was 18%. The lipophilic mixture with non-ionic surfactants in the selected formulation of lipid microspheres has a favorable effect on size. The selected formulation is also cosmetically adapted and it is composed of physiological and biodegradable lipids. B-3 was released and penetrated into skin more quickly and in greater quantity than in SLM form, from vehicles containing free B-3. This work has shown that SLM is an excellent carrier for lipophilic sunscreens like B-3 in order to decrease the release and penetration rate of this UV absorber compared with B-3 in oily solution.

Nanoemulsions are biphasic systems with droplet size in the nanometric scale (20–200nm). Oil/water nanoemulsions are promising colloidal drug carrier systems for diverse therapeutic applications as topical, intravenous, oral and ocular [1]. They are non-equilibrium systems, a high energy input generally from mechanical devices or from the chemical potential of the components, is required for their formation [2]. In this work we prepared an oil/water nanoemulsion with the ethanolic extract obtained from flowers of Calceolaria chelidonioides 5% with droplet size of 45,46±3,64nm and high stability using a classical macroemulsion protocol by mixing two formulation concepts, the H.L.B. and the phase diagram, to choose the best non-ionic surfactant match. Calceolaria chelidonioides is an original Brazilian plant belonging to the Scrophulariaceae family. This plant has been studied in our group and we were able to identify its antimicrobial and antioxidant properties [3]. We evaluate this nanoemulsion skin hydration power in vivo with female volunteers between (20–40 years old). The measurement of the skin humidity was carried out by a Corneometer® CM820 using the capacitance method, before and 10, 20, 30, 45 minutes after the administration. Two moisturizing comercial products well known by its properties, Nivea Soft® from Nivea® and Ictyane® from Ducray® were used as control, and the formulated nanoemulsion without the plant active was evaluated. The C. chelidonioides' nanoemulsion showed some interesting skin hydration actitvity higher than Nivea Soft® cream, especially after 45 minutes, showing an increase of 25% in skin hydration. However, the most important hydration power was obtained with the nanoemulsion base which showed 30% of increase after 45 minutes.

Calceolaria chelidonioides is an original Brazilian plant belonging to the Scrophulariaceae family, which is used in the Brazilian folk medicine for the treatment of several kinds of cancer. Its cytotoxicity, photoxicity and genotoxicity potential were evaluated in different methodologies in vitro using the flowers ethanol extract. The cytotoxicity and phototoxicity were evaluated by the neutral red dye assay using keratinocyte human cells (NCTC 2544). For the phototoxicity evaluation the cell culture containing the compounding test was submitted to UVA radiation (345nm) during 15 minutes. The assays showed the cell viability after the treatment with the extract and its metabolites formed by the UV radiation. The genotoxicity potential was evaluated by two different methods, both of them suggested in the „Genotoxicity: a standard battery for genotoxicity testing of pharmaceuticals“ guide [1]. The first one was the Comet assay [2,3] using keratinocyte human cells incubated for 1 hour with the extract, with and without metabolic activation using the S9 mix. The Comet assay is able to detect different kinds of DNA fragmentations caused by the genotoxic agents. The cells which DNA were damaged show an image comet like with a „head“ and a „tail“ that elongates proportionally to the DNA damages. The second method was the Ames' test [4] which is capable to detect compounds with carcinogenic and mutagenic properties using mutants Salmonella typhimurium strains to detect base substitution and frame shift point mutations. These tests were also evaluated with and without metabolic activation. The C. calceolaria flowers ethanol extract or even its metabolites didn't show any kind of toxicity in all tested models.

Langerhans cells (LC) are key mediators of contact allergenicity in the skin. However, no in vitro methods exist which are based on the activation process of LC to predict the sensitization potential of chemicals. In this study, we have evaluated the performances of MUTZ-3, a cytokine-dependent human monocytic cell line, in its response to sensitizers. First, we compared undifferentiated MUTZ-3 cells with several standard human cells such as THP-1, KG-1, HL-60, K-562, and U-937 in their response to the strong sensitizer DNCB and the irritant SDS by monitoring the expression levels of HLA-DR, CD54, and CD86 by flow cytometry. Only MUTZ-3 and THP-1 cells show a strong and specific response to sensitizer, while other cell lines showed very variable responses. Then, we tested MUTZ-3 cells against a wider panel of sensitizers and irritants on a broader spectrum of cell surface markers (HLA-DR, CD40, CD54, CD80, CD86, B7-H1, B7-H2, B7-DC). Of these markers, CD86 proved to be the most reliable since it detected all sensitizers, including benzocaine, a classical false negative in local lymph node assay (LLNA) but not irritants. We confirmed the MUTZ-3 response to DNCB by real-time PCR analysis. Taken together, our data suggest that undifferentiated MUTZ-3 cells may represent a valuable in vitro model for the screening of potential sensitizers.

Caffeic acid, chlorogenic acid and oraposide, a natural glycoside, are phenyl-propanoid compounds. These natural products have been reported to have antioxidant activities such as the scavenging of superoxide anions and hydroxyl radicals. These compounds could be used in the dermocosmetic field to protect the skin from oxidative stress induced by UV radiation. To this end, the permeation of caffeic acid, chlorogenic acid, and oraposide, through pig-ear skin was evaluated in vitro. The percutaneous permeation of these three compounds through pig skin was measured and compared using Franz diffusion cells. At appropriate intervals, up to 72h, diffusion samples were analyzed using an HPLC assay. After 48h of drug contact the permeation was also evaluated with a fluorescent microscope on vertical microtomed pig skin sections. In this study on excised pig skin, the flux value was found to be equal to 0.32 and 0.48μgcm−2h−1 for caffeic and chlorogenic acids, respectively; for oraposide the levels were below the limit of detection and the flux was not evaluated. These results were corroborated by fluorescent microscopy. Caffeic and chlorogenic acids were found in all skin sections, and these might represent a systemic activity, whereas oraposide remained in the upper superficial layer of the skin. This latter phenomenon seems to be interesting for dermocosmetic applications.

Glutathione-S-transferase Pi1 (GSTP1) and multidrug resistance protein 1 (MRP1) are overexpressed in melanoma, a skin cancer notoriously resistant to all current modalities of cancer therapy. To investigate the involvement of these detoxifying enzymes in the drug resistance of melanoma, an inducible (Tet-On™ system) antisense (AS) RNA strategy was used to specifically inhibit GSTP1 expression in A375 cells, a human melanoma cell line expressing high levels of GSTP1 and MRP1. Stable transfectant clones were established and analysed for GSTP1 inhibition by AS RNA. The clone A375-ASPi1, presenting a specific 40% inhibition of GSTP1 expression in the presence of doxycycline, was selected. Lowering the GSTP1 level significantly increased (about 3.3-fold) the sensitivity of A375-ASPi1 cells to etoposide. Inhibitors of glutathione synthesis (BSO), GSTs (curcumin, ethacrynic acid), and also of MRPs (MK571, sulphinpyrazone) improved the sensitising effect of GSTP1 AS RNA. All these inhibitors had stronger sensitising effects in control cells expressing high GSTP1 level (A375-ASPi1 cells in the absence of doxycycline). In conclusion, GSTP1 can act in a combined fashion with MRP1 to protect melanoma cells from toxic effects of etoposide.

Le but de l’étude est de vérifier si les monteurs de pneus sont exposés au plomb des masselottes utilisées pour équilibrer les roues des automobiles. Nous avons comparé 36 monteurs et 37 témoins. Les prélèvements sanguins ont permis d’analyser l’hémogramme, la plombémie, la protoporphyrine-Zinc et l’activité de l’ALA-D. Monteurs et témoins ont rempli un questionnaire pour comparer les expositions extraprofessionnelles éventuelles et préciser l’activité professionnelle dans le cas des monteurs. Dans le cadre de l’étude, nous n’avons pas décelé de différence liée à l’habitat ou à l’eau consommée. Il n’y a pas non plus de différence d’antécédents d’exposition au plomb, d’onychophagie ni d’activités extraprofessionnelles pouvant exposer au plomb. Par contre, nous avons constaté que les monteurs de l’étude étaient plus jeunes, fumaient davantage et avaient les mains plus abîmées que celles des témoins. Nous n’avons pas mis en évidence de différences au niveau des résultats hématologiques et toxicologiques (plombémie, PPZ et ALA-D) entre les monteurs et les témoins. Tant chez les monteurs que chez les témoins, la plombémie augmente avec l’âge. La suppression de cette influence permet de mettre en évidence une différence très significative de la moyenne de la plombémie entre monteurs et témoins. L’augmentation de la moyenne de la plombémie (non corrigée par l’âge) entre monteurs et témoins est décelable et significative à partir de 10 pneus/jour. De même, entre les monteurs qui équilibrent plus ou moins de 10 pneus/jour. Le port des gants diminue significativement la plombémie des monteurs par rapport à ceux qui n’en mettent pas. Même s’il s’agit de très faibles doses, le port des gants doit être encouragé en raison de l’exposition quotidienne et du caractère cumulatif du plomb dans l’organisme. Mais c’est seulement la finesse de l’étude qui a permis de mettre en évidence une différence significative entre monteurs et témoins. Elle ne justifie cependant pas d’envisager une modification de la réglementation en ce domaine. This study evaluates tyre-fitters’ exposure to lead from the inertia blocks used to equilibrate car wheels. A study was carried out on 36 fitters and 37 controls. Blood samples were used to determine hemogram, blood lead level, ZPP and ALAD activity. Moreover, participants filled in a questionnaire so as to investigate possible hobby exposure and provide details about the mechanics’ professional activities. Within the limits of this study, no differences are shown between the two groups according to accomodation, water drinking habits, anterior lead exposure, onychophagia or lead exposure hobbies. We nevertheless observed in our study that the mechanics are younger, heavier smokers and have rougher hands than controls. Between mechanics and controls, the haematological or toxicological results (lead, ZPP, ALAD) showed no differences. However we noticed, among mechanics and controls, that blood lead level increases with age. When the age influence is cancelled, a highly significant difference then appears for blood lead level average between mechanics and controls. Moreover an increase in blood lead level average, without correction for age, between mechanics and controls, is significant for tyre-fitters fitting 10 or more tyres a day. The difference is also significant between mechanics who equilibrate more than 10 tyres a day versus those who set less than 10 tyres a day. Glove wearing significantly decreases the mechanics’ blood lead results. Therefore even for very low exposure workers must be encouraged to wear gloves in order to prevent the cumulative effects of lead in the human body. But it's only the acuteness of the study which allowed to show a significant difference between fitters and controls. This difference does not justify a modification of the regulations in this field.

Previous studies have shown that glutathione S-transferases (GSTs) can operate in synergy with efflux transporters, multi-drug resistance proteins (MRPs), to confer resistance to several carcinogens, mutagens and anticancer drugs. To address the poorly documented role of the GSTM1 in cancer chemoresistance, we used CAL1 human melanoma cells expressing no endogenous GSTM1 and a high level of MRP1. Cells were transfected with an expression vector containing the GSTM1 cDNA, and different clones were selected expressing different levels of GSTM1 (RT-PCR, Western blot, and enzyme activity). Cells overexpressing GSTM1 displayed a 3- to 4-fold increase in resistance to anticancer drugs vincristine (VCR) and chlorambucil (CHB) in proliferation, cytotoxic, and clonogenic survival assays. Inhibitors of MRP1 (sulfinpyrazone, verapamil) and GST (dicumarol, curcumin) completely reversed the GSTM1-associated resistance to VCR, indicating that a MRP efflux function is necessary to potentiate GSTM1-mediated resistance to VCR. Conversely, MRP1 inhibitors had no effect on the sensitivity to CHB. Using immunofluorescence assay, GSTM1 was also shown to protect microtubule network integrity from VCR-induced inhibition of microtubule polymerization. In conclusion, these results show that GSTM1 alone is involved in melanoma resistance to CHB, whereas it can act in synergy with MRP1 to protect cells from toxic effects of VCR.

Fotemustine is a third generation chloroethylnitrosourea that has demonstrated significant antitumoral effects in malignant melanoma. However, its use is somewhat limited by its toxic side effects and chemoresistance caused by direct repair of O6-alkyl groups by the enzyme O6-methylguanine DNA-methyltransferase (MGMT). The aim of this work was to determine to what extent the expression of MGMT influences cytotoxicity, DNA damage, and apoptosis induced by new nitrososulfamide analogs of fotemustine (compounds 4 and 8), which have previously demonstrated interesting antiproliferative properties. We carried out complementary strategies that consisted of MGMT cDNA transfection in CAL77 Mer- melanoma cells and of MGMT inhibition with O6-benzylguanine (BG) in A375 Mer+ melanoma cells. MGMT-transfected cells were 7 to 9 times less sensitive to fotemustine than parent cells, whereas no difference between the transfected and parent cells was observed for nitrososulfamide analogs. The cytotoxicity of these analogs vis à vis a MGMT-proficient A375 melanoma cell line was approximately 3 times greater than that of fotemustine. Coincubation of these cells with O6-benzylguanine significantly increased the cytotoxicity of fotemustine and compound 8, whereas BG had little effect on the cytotoxicity of compound 4. Furthermore, DNA fragmentation determined by a comet assay was greater with nitrososulfamide analogs than with fotemustine. O6-benzylguanine increased DNA fragmentation for fotemustine and compound 8, but not for compound 4, which induced comets with a typical apoptotic appearance. The ability of this compound to induce apoptosis in the absence of BG was confirmed by a specific enzyme-linked immunosorbent assay apoptotic assay using a single-stranded DNA monoclonal antibody.

Liverbeads, cryopreserved hepatocytes entrapped within an alginate matrix, were examined for their relevance in the comet assay. It was estimated by their capacity to activate the indirectly acting mutagens, cyclophosphamide (CP), benzo[a]pyrene (BP), dimethylbenzanthracene (DMBA) and 2-acetylaminofluorene (2-AAF), into DNA reactive metabolites. The comet assay performed in alkaline condition is a sensitive method for detecting strand breaks at the level of individual cells and allows use of quiescent cells. Experimental conditions as treatment time, cell density, beads dissociation and viability were investigated. Significant statistical positive results assessed by the tail extent moment (TEM) were observed with both human and rat Liverbeads after 12h duration incubation compared to metabolic non-competent cells, HeLa S3. Due to the maintenance of specific functions assessed by the observed capacity to metabolize xenobiotics, Liverbeads represent a suitable tool system, easy to handle, for the detection of promutagens using the comet assay.

The study described in this paper constitutes a practical assay system to evaluate in vivo drug penetration using two complementary non-invasive methods. An electrical capacitance test was first applied to the skin on the forearm to evaluate the hydration of the skin, and check the integrity of the stratum corneum. In the first step, the percentage absorption was measured using an occlusive and difference method; following benzophenone-3 application any residual formulation was washed off and the amount removed analyzed. In the second step, the tape stripping method-a useful procedure for selectively removing the skin's outermost layer, the stratum corneum, and measuring the stratum corneum adsorption-was performed. Under these conditions the human skin permeation of this UV-filter over four hours was near to 35% of the applied dose with the occlusive method. The amount of topically applied benzophenone-3 found in the stratum corneum after 30 min exposure using the stripping procedure was evaluated at 4% to the applied dose.

BACKGROUND:Ultraviolet (UV) B-induced effects on the skin immune system have been extensively investigated, but little is known regarding the immunological changes induced by UVA exposure of human skin. Recent data assessing the protection afforded by sunscreens against photoimmunosuppression stress the need for broad-spectrum sunscreens with an adequate UVA protection.OBJECTIVES:The purpose of this study was first to determine the changes observed in epidermal Langerhans cells (ELC) density and epidermal antigen-presenting cell (APC) activity after exposure of human skin to UVAI (340-400 nm) radiation, and secondly to assess the immune protection afforded in vivo by a sunscreen formulation containing a long wavelength UVA filter with a low UVA protection factor (UVA-PF = 3).METHODS:Epidermal cell (EC) suspensions were prepared from skin biopsies 3 days after exposure to a single dose of UVAI (either 30 or 60 J cm(-2)).RESULTS:Flow-cytometric analysis of EC suspensions revealed that exposure to 60 J cm(-2) UVAI resulted in a decreased number of ELC without infiltration of CD36+ DR+ CD1a- antigen-presenting macrophages into the epidermis, and a significant reduction of HLA-DR expression on viable ELC. In vivo exposure to both 30 and 60 J cm(-2) resulted in a decreased allogeneic CD4+ T-cell proliferation induced by UVAI-irradiated ECs. The sunscreen application partially prevented (57 +/- 9%) the decrease in epidermal allogeneic APC activity induced by 60 J cm(-2) UVAI.CONCLUSIONS:In vivo UVAI exposure of human skin results in a decreased number of ELC and in a downregulation of epidermal APC activity. This last effect is partially prevented by prior application of a sunscreen with a low UVAI-PF value. These results indicate that increasing the absorption of UV filters for long UVA wavelengths may lead to an improved immune protection.

The relationship between uridine phosphorylase (UP) expression level in cancer cells and the tumour sensitivity to fluoropyrimidines is unclear. In this study, we found that UP overexpression by gene transfer, and the subsequent efficient metabolic activation of 5-fluorouracil (5-FU) by the ribonucleotide pathway, does not increase the fluoropyrimidine sensitivity of MCF-7 human cancer cells. © 2001 Cancer Research Campaign http://www.bjcancer.com

Background. Contact dermatitis to henna are rarely reported. Most of the lime, they are related to additives, especially scented oils or paraphenylene diamine. This latter substance is added to shorten the time of application and to enhance the darkness of the color. We report one case of an allergic contact dermatitis to henna skin tattoo, probably related to a paraphenylene diamine derivative. Case report. A 25-year-old woman developed an allergic contact dermatitis at the site of application of two henna skin tattoos on her right arm. She never dyed her hair and wore dark clothing without problems. Epicutaneous tests were positive for henna paste, paraphenylene diamine, N-isopropyl-N'phenyiparaphenylene diamine and five chemical dyes, probably related to cross-reactions between paraphenylene diamine and these dyes. Epicutaneous test with henna powder was negative, confirming the absence of allergy to henna. Fluorescence analysis detected the presence of a primary amine in henna paste but high performance liquid chromatography analysis did not reveal the presence of paraphenylene diamine or any suspect dye. An another allergen, related compound of paraphenylene diamine, was probably present but we failed to identify it with precision. Discussion. Labile henna skin tattoos are in fashion at the moment in western European countries. They can be a secrete source of paraphenylene diamine, or one of its related compounds, sensitization.