Arlette Baillet-Guffroy

Age-dependent changes in stratum corneum barrier function

The Stratum Corneum (SC) barrier function mainly depends on the SC structure at the tissue level, its composition, and the organization of intercellular lipidic cement at the molecular level. The goal of this study was to assess the age‐dependent changes of the SC barrier function and the associated physiological parameters.

Molecular interactions of penetration enhancers within ceramides organization: A FTIR approach

The barrier function of the skin is related to the unique composition of the stratum corneum (SC) lipids and their complex structural arrangement. The high content of ceramides would seem to be ideally suited for the formation of ordered impermeable membrane. Skin penetration enhancers (PE) are molecules which reversibly remove the barrier resistance of the SC. Interactions with SC intercellular lipids are of crucial importance for the effectiveness of PE action. Their mode of action on the lipid bilayer may involve interactions at two sites, i.e., at or near the polar head groups of the lipid bilayer and/or between the hydrophobic tails of the bilayer. This paper discusses the local effect of four PE, among the most investigated, limonene, ethanol, oleic acid and DMSO. FTIR is used in this study to highlight the local effect of the PE on ceramides films. Lipophilic PE, i.e., oleic acid and limonene, both present a direct fluidizing action on the alkyl chains and an indirect action on the polar head groups resulting in a more spacing lipid packing. Hydrophilic PE, i.e., ethanol and DMSO, have no interaction on the lipid bilayer but show a complex action on the polar headgroup, weakening the H-bonds. Our most significant finding is that each PE we investigated interacted with the ceramide packing, depending of these structures. Such modifications contribute a share to interpretation, at the molecular level, of the decrease of skin barrier properties with PE described in published data.

Mobility of water molecules in the stratum corneum: effects of age and chronic exposure to the environment.

enhanced antimicrobial activity against P. acnes and inhibited the expression of proinflammatory cytokines, such as IL-8 and tumor necrosis factor-a. Our data showed that the T-zone had a low incidence of inflammatory acne lesions compare with U-zone. These data support the previous findings that hBD-2 levels indicate the presence of antiinflammatory lipids, which inhibit inflammation, therefore, only a few clinically inflammatory lesions occur (Georgel et al., 2005; Zouboulis et al., 2010). Previous studies demonstrated that FFAs enhance the innate antibacterial defenses of the skin by inducing the expression of AMP (Lee et al., 2008; Nakatsuji et al., 2010). Our findings also showed that considerable FFAs in the high-sebum-secreting area (T-zone) induce more abundant hBD-2 expression, and enhanced antimicrobial activity against P. acnes and inhibited the expression of proinflammatory cytokines. However, it needs to further evaluation for the expression of other AMP in each area, because other AMP may also have a role in regional difference of inflammatory acne lesions (Lee et al., 2008, 2009). There was a previous study that hBD-2 strikingly killed P. acnes at concentrations higher than 10 mM (Nakatsuji et al., 2010). Although we showed which amount of hBD-2 in sebocytes from T-zones able to influence the growth of P. acnes in vitro, it needs further in vivo study especially in acne lesions. In conclusion, we suggest that increased hBD-2 in the T-zone prevents the progress of comedones to papulopustular lesions via either an upregulation of antiinflammatory sebaceous lipid synthesis and/or an inhibitory activity of P. acnes proliferation. These data suggest a functional link between topographical variation of inflammatory acne lesions and innate immunity in acne pathogenesis. Further studies of the relationship between hBD-2 expression and P. acnes in acne lesion are warranted to gain the confirmation of the effects of hBD-2 on regional difference of inflammatory acne lesions.

Comparison of structure and organization of cutaneous lipids in a reconstructed skin model and human skin: spectroscopic imaging and chromatographic profiling

The use of animals for scientific research is increasingly restricted by legislation, increasing the demand for human skin models. These constructs present comparable bulk lipid content to human skin. However, their permeability is significantly higher, limiting their applicability as models of barrier function, although the molecular origins of this reduced barrier function remain unclear. This study analyses the stratum corneum (SC) of one such commercially available reconstructed skin model (RSM) compared with human SC by spectroscopic imaging and chromatographic profiling. Total lipid composition was compared by chromatographic analysis (HPLC). Raman spectroscopy was used to evaluate the conformational order, lateral packing and distribution of lipids in the surface and skin/RSM sections. Although HPLC indicates that all SC lipid classes are present, significant differences are observed in ceramide profiles. Raman imaging demonstrated that the RSM lipids are distributed in a non‐continuous matrix, providing a better understanding of the limited barrier function.

Effect of UVA or UVB irradiation on cutaneous lipids in films or in solution.

The barrier function of the skin is largely due to the stratum corneum which is essentially composed of lipids. Different external factors, such as UV irradiation, affect this skin layer and are responsible for a destabilization of the supramolecular organization of its constituted lipids. In this work, mass spectrometry and infrared spectroscopy are combined to study the correlation between the formation of oxidative compounds by UV irradiation and the lipid organization. Experiments were carried out on unsaturated lipids in film or solution form, exposed to UVA or UVB irradiation. UV exposure leads to the formation of oxygenated entities in the case of lipids with an unsaturated fatty acid moiety, resulting in a decrease in their packing which is greater when the lipids are in solution. The packing decrease is even greater following UVB irradiation.

Raman spectroscopy: in vivo quick response code of skin physiological status.

Abstract. Dermatologists need to combine different clinically relevant characteristics for a better understanding of skin health. These characteristics are usually measured by different techniques, and some of them are highly time consuming. Therefore, a predicting model based on Raman spectroscopy and partial least square (PLS) regression was developed as a rapid multiparametric method. The Raman spectra collected from the five uppermost micrometers of 11 healthy volunteers were fitted to different skin characteristics measured by independent appropriate methods (transepidermal water loss, hydration, pH, relative amount of ceramides, fatty acids, and cholesterol). For each parameter, the obtained PLS model presented correlation coefficients higher than R2=0.9. This model enables us to obtain all the aforementioned parameters directly from the unique Raman signature. In addition to that, in-depth Raman analyses down to 20 μm showed different balances between partially bound water and unbound water with depth. In parallel, the increase of depth was followed by an unfolding process of the proteins. The combinations of all these information led to a multiparametric investigation, which better characterizes the skin status. Raman signal can thus be used as a quick response code (QR code). This could help dermatologic diagnosis of physiological variations and presents a possible extension to pathological characterization.

The contribution of Raman spectroscopy to the analytical quality control of cytotoxic drugs in a hospital environment: Eliminating the exposure risks for staff members and their work environment

The purpose of the study was to perform a comparative analysis of the technical performance, respective costs and environmental effect of two invasive analytical methods (HPLC and UV/visible-FTIR) as compared to a new non-invasive analytical technique (Raman spectroscopy). Three pharmacotherapeutic models were used to compare the analytical performances of the three analytical techniques. Statistical inter-method correlation analysis was performed using non-parametric correlation rank tests. The studys economic component combined calculations relative to the depreciation of the equipment and the estimated cost of an AQC unit of work. In any case, analytical validation parameters of the three techniques were satisfactory, and strong correlations between the two spectroscopic techniques vs. HPLC were found. In addition, Raman spectroscopy was found to be superior as compared to the other techniques for numerous key criteria including a complete safety for operators and their occupational environment, a non-invasive procedure, no need for consumables, and a low operating cost. Finally, Raman spectroscopy appears superior for technical, economic and environmental objectives, as compared with the other invasive analytical methods.

Discrimination and quantification of two isomeric antineoplastic drugs by rapid and non-invasive analytical control using a handheld Raman spectrometer.

Raman spectroscopy is a rapid, non-destructive and non-invasive method that is a promising tool for real-time analytical control of drug concentrations. This study evaluated a handheld Raman device to discriminate and quantify two isomeric drugs used to treat cancer. Doxorubicin (DOXO) and epirubicin (EPIR) samples were analyzed at therapeutic concentrations from 0.1 to 2mg/mL (n=90) and 0.08-2mg/mL (n=90) by non-invasive measurements using a portable Raman spectrometer. The discrimination of these two molecules was demonstrated for all concentrations (n=180) by qualitative analysis using partial least square discriminant analysis (PLS-DA) with 100% classification accuracy, sensitivity and specificity and 0% error rate. For each molecule, quantitative analyses were performed using PLS regression. The validity of the model was evaluated using root mean square error of cross validation (RMSECV) and prediction (RMSEP) that furnished 0.05 and 0.02mg/mL for DOXO and 0.17 and 0.16mg/mL for EPIR after pretreatment optimization. Based on the accuracy profile, the linearity range was from 1.256 to 2.000mg/mL for DOXO (R2=0.9988) and from 0.553 to 2.000mg/Ml for EPIR (R2=0.9240) and repeatability (CV% max of 1.8% for DOXO and 3.2% for EPIR) and intermediate precision (CV% max of 2.8% for DOXO and 4.5% for EPIR) were both acceptable. Despite the narrow validated concentration range for quantitative analysis, this study shows the potential of a handheld Raman spectrometer coupled to chemometric approaches for real-time quantification of cytotoxic drugs, as well for discriminating between two drugs with similar UV absorption profiles. Finally, the use of a handheld spectrometer with the possibility of a direct measurement of substances in containers is a potentially valuable tool for combining patient safety with security of healthcare workers.

Examination of the effect of Stratum Corneum isolation process on the integrity of the barrier function: a confocal Raman spectroscopy study

Stratum Corneum (SC) is the most superficial layer of the epidermis. It plays the main barrier role against water loss and the aggression of external chemical and biological substances. Thermal treatment in warm purified water followed by trypsin incubation of excised human skin is a well‐established in vitro method for SC removal. Different protocols can be found in literature, but little is described about the effect of temperature and trypsin during isolation process on its barrier function.

Vibrational spectroscopy coupled to classical least square analysis, a new approach for determination of skin moisturizing agents' mechanisms.

Skin dryness is an omnipresent symptom in various types of skin disorders. Thereby, a large panel of skin care products is developed for therapeutic purposes. However, there is still a lack of non‐invasive methods to determine the mechanisms of action of moisturizers at the molecular level.