http://repub.eur.nl/res/aut/12592/ List of Publications en http://repub.eur.nl/static-eur/img/logo.png http://repub.eur.nl http://repub.eur.nl/res/pub/33247/ 2011-11-01T00:00:00Z Caspase-14 is a protease that is mainly expressed in suprabasal epidermal layers and activated during keratinocyte cornification. Caspase-14-deficient mice display reduced epidermal barrier function and increased sensitivity to UVB radiation. In these mice, profilaggrin, a protein with a pivotal role in skin barrier function, is processed correctly to its functional filaggrin (FLG) repeat unit, but proteolytic FLG fragments accumulate in the epidermis. In wild-type stratum corneum, FLG is degraded into free amino acids, some of which contribute to generation of the natural moisturizing factors (NMFs) that maintain epidermal hydration. We found that caspase-14 cleaves the FLG repeat unit and identified two caspase-14 cleavage sites. These results indicate that accumulation of FLG fragments in caspase-14-/-mice is due to a defect in the terminal FLG degradation pathway. Consequently, we show that the defective FLG degradation in caspase-14-deficient skin results in substantial reduction in the amount of NMFs, such as urocanic acid and pyrrolidone carboxylic acid. Taken together, we identified caspase-14 as a crucial protease in FLG catabolism. http://repub.eur.nl/res/pub/33267/ 2011-10-01T00:00:00Z http://repub.eur.nl/res/pub/20619/ 2010-09-01T00:00:00Z Background: Filaggrin (FLG) has a central role in the pathogenesis of atopic dermatitis (AD). FLG is a complex repetitive gene; highly population-specific mutations and multiple rare mutations make routine genotyping complex. Furthermore, the mechanistic pathways through which mutations in FLG predispose to AD are unclear. Objectives: We sought to determine whether specific Raman microspectroscopic natural moisturizing factor (NMF) signatures of the stratum corneum could be used as markers of FLG genotype in patients with moderate-to-severe AD. Methods: The composition and function of the stratum corneum in 132 well-characterized patients with moderate-to-severe AD were assessed by means of confocal Raman microspectroscopy and measurement of transepidermal water loss (TEWL). These parameters were compared with FLG genotype and clinical assessment. Results: Three subpopulations closely corresponding with FLG genotype were identified by using Raman spectroscopy. The Raman signature of NMF discriminated between FLG-associated AD and non-FLG-associated AD (area under the curve, 0.94; 95% CI, 0.91-0.99). In addition, within the subset of FLG-associated AD, NMF distinguished between patients with 1 versus 2 mutations. Five novel FLG mutations were found on rescreening outlying patients with Raman signatures suggestive of undetected mutations (R3418X, G1138X, S1040X, 10085delC, and L2933X). TEWL did not associate with FLG genotype subgroups. Conclusions: Raman spectroscopy permits rapid and highly accurate stratification of FLG-associated AD. FLG mutations do not influence TEWL within established moderate-to-severe AD. http://repub.eur.nl/res/pub/26923/ 2009-02-01T00:00:00Z As the possibilities in the treatment of cancer continue to evolve, its early detection and correct diagnosis are becoming increasingly important. Froum the early detection of cancer to the guidance of oncosurgical procedures new sensitive in vivo diagnostic tools are much needed. Many studies report the Raman spectroscopic detection of malignant and premalignanl tissues in different sites of the body with high sensitivities. The great appeal of this technique lies in its potential for in vivo clinical implementation. We present an overview of the in vitro and in vivo work on the oncological application of Raman spectroscopy and discuss its potential as a new tool in the clinico-oncological practice. Opportunities for integration of Raman spectroscopy in oncological cure and care as a real-time guidance tool during diagnostic (i.e. biopsy) and therapeutic (surgical resection) modalities as well as technical shortcomings are discussed from a clinicians point of view. Hut lire Rinnan application lor real-time tumor-border http://repub.eur.nl/res/pub/29246/ 2008-08-01T00:00:00Z http://repub.eur.nl/res/pub/37094/ 2007-03-01T00:00:00Z Resonance Raman spectra of oxygenated and deoxygenated functional erythrocytes recorded using 785 nm laser excitation are presented. The high-quality spectra show a mixture of enhanced A1g, A2g, B1g, B2g, Euand vinyl modes. The high sensitivity of the Raman system enabled spectra from four oxygenation and deoxygenation cycles to be recorded with only 18 mW of power at the sample over a 60-minute period. This low power prevented photo-/thermal degradation and negated protein denaturation leading to heme aggregation. The large database consisting of 210 spectra from the four cycles was analyzed with principal components analysis (PCA). The PC1 loadings plot provided exquisite detail on bands associated with the oxygenated and deoxygenated states. The enhancement of a band at 567 cm-1, observed in the spectra of oxygenated cells and the corresponding PC1 loadings plot, was assigned to the Fe-O2stretching mode, while a band appearing at 419 cm-1was assigned to the Fe-O-O bending mode based on previous studies. For deoxygenated cells, the enhancement of B1gmodes at 785 nm excitation is consistent with vibronic coupling between band III and the Soret transition. In the case of oxygenated cells, the enhancement of iron-axial out-of-plane modes and non-totally symmetric modes is consistent with enhancement into the y,z-polarized transition a1u(π)→ddz+O2(πg) centered at 785 nm. The enhancement of non-totally symmetric B1gmodes in oxygenated cells suggests vibronic coupling between band IV and the Soret band. This study provides new insights into the vibrational dynamics, electronic structure and resonant enhancement of heme moieties within functional erythrocytes at near-IR excitation wavelengths. http://repub.eur.nl/res/pub/10186/ 2003-01-01T00:00:00Z In vivo confocal Raman spectroscopy is a noninvasive optical method to obtain detailed information about the molecular composition of the skin with high spatial resolution. In vivo confocal scanning laser microscopy is an imaging modality that provides optical sections of the skin without physically dissecting the tissue. A combination of both techniques in a single instrument is described. This combination allows the skin morphology to be visualized and (subsurface) structures in the skin to be targeted for Raman measurements. Novel results are presented that show detailed in vivo concentration profiles of water and of natural moisturizing factor for the stratum corneum that are directly related to the skin architecture by in vivo cross-sectional images of the skin. Targeting of skin structures is demonstrated by recording in vivo Raman spectra of sweat ducts and sebaceous glands in situ. In vivo measurements on dermal capillaries yielded high-quality Raman spectra of blood in a completely noninvasive manner. From the results of this exploratory study we conclude that the technique presented has great potential for fundamental skin research, pharmacology (percutaneous transport), clinical dermatology, and cosmetic research, as well as for noninvasive analysis of blood analytes, including glucose. http://repub.eur.nl/res/pub/10883/ 2003-01-01T00:00:00Z In vivo confocal Raman spectroscopy is a noninvasive optical method to obtain detailed information about the molecular composition of the skin with high spatial resolution. In vivo confocal scanning laser microscopy is an imaging modality that provides optical sections of the skin without physically dissecting the tissue. A combination of both techniques in a single instrument is described. This combination allows the skin morphology to be visualized and (subsurface) structures in the skin to be targeted for Raman measurements. Novel results are presented that show detailed in vivo concentration profiles of water and of natural moisturizing factor for the stratum corneum that are directly related to the skin architecture by in vivo cross-sectional images of the skin. Targeting of skin structures is demonstrated by recording in vivo Raman spectra of sweat ducts and sebaceous glands in situ. In vivo measurements on dermal capillaries yielded high-quality Raman spectra of blood in a completely noninvasive manner. From the results of this exploratory study we conclude that the technique presented has great potential for fundamental skin research, pharmacology (percutaneous transport), clinical dermatology, and cosmetic research, as well as for noninvasive analysis of blood analytes, including glucose. http://repub.eur.nl/res/pub/10880/ 2002-01-01T00:00:00Z The stratum corneum (SC) barrier typically consists of layers of corneocytes embedded in a lipid continuum that regulates barrier function. The lipid domain containing ceramides, cholesterol, and free fatty acids provides the major pathway for most drugs permeating across SC (1). Penetration enhancers diminish the SC barrier function. The classic enhancer is dimethyl sulfoxide (DMSO) (2). Its mechanisms of action remain unclear, although DMSO disrupts lipid organisation and may displace protein-bound water (3). Here we use confocal Raman spectroscopy to probe molecular interactions between a finite (depleting) dose of DMSO and SC, as functions of depth and time, providing novel information about residence time and location of DMSO in human SC in vivo. http://repub.eur.nl/res/pub/10881/ 2001-03-20T00:00:00Z Confocal Raman spectroscopy is introduced as a noninvasive in vivo optical method to measure molecular concentration profiles in the skin. It is shown how it can be applied to determine the water concentration in the stratum corneum as a function of distance to the skin surface, with a depth resolution of 5 mum. The resulting in vivo concentration profiles are in qualitative and quantitative agreement with published data, obtained by in vitro X-ray microanalysis of skin samples. Semi-quantitative concentration profiles were determined for the major constituents of natural moisturizing factor (serine, glycine, pyrrolidone-5-carboxylic acid, arginine, ornithine, citrulline, alanine, histidine, urocanic acid) and for the sweat constituents lactate and urea. A detailed description is given of the signal analysis methodology that enables the extraction of this information from the skin Raman spectra. No other noninvasive in vivo method exists that enables an analysis of skin molecular composition as a function of distance to the skin surface with similar detail and spatial resolution. Therefore, it may be expected that in vivo confocal Raman spectroscopy will find many applications in basic and applied dermatologic research. http://repub.eur.nl/res/pub/10882/ 2000-01-01T00:00:00Z An automated confocal Raman microspectrometer for rapid measurement of molecular concentration profiles in the skin is described. It permits the successive collection of Raman spectra at a range of depths below the skin surface. The axial resolution of the confocal Raman microspectrometer is 5.1 +/- 0.2 mu m. The setup was applied to determine water concentration profiles of the stratum corneum and to determine changes therein as a result of hydration of the skin.