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BACKGROUND: A preference for type 2 immunity plays a central role in the pathogenesis of atopic dermatitis (AD). Dupilumab, a monoclonal antibody targeting the IL-4α receptor subunit, inhibits IL-4 and IL-13 signaling. These cytokines contribute significantly to IgE class switch recombination in B-cells, critical in atopic diseases. Recent studies indicate IgG+CD23hiIL-4RA+ memory B-cells (MBC2) as IgE-producing B-cell precursors, linked to total IgE serum levels in atopic patients. Total IgE serum levels decreased during dupilumab treatment in previous studies. OBJECTIVE: To assess the effects of dupilumab treatment in comparison to alternative therapies on the frequency of MBC2 and the correlation to total IgE levels in pediatric patients with AD. METHODS: Pediatric patients with AD, participating in an ongoing trial, underwent randomization into three treatment groups: dupilumab (n=12), cyclosporine (n=12), or topical treatment (n=12). Plasma and Peripheral Blood Mononuclear Cells (PBMCs) were collected at baseline (T0) and after 6 months (T6). Flow cytometry was employed for PBMC phenotyping, ELISA was utilized to assess total IgE levels in plasma. For detailed Methods, please see the Methods section in this article's Online Repository at www.jacionline.org RESULTS: Our findings revealed a significant reduction in MBC2 frequency and total IgE levels among patients treated with dupilumab. Additionally, a significant correlation was observed between MBC2s and total IgE levels CONCLUSION: Systemic blocking of the IL-4RA subunit leads to a decrease in circulating MBC2 cells and total IgE in pediatric AD patients. Our findings unveil a novel mechanism through which dupilumab exerts its influence on the atopic signature.
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Near-infrared (NIR) fluorescence imaging using exogenous fluorescent agents provides whole-field images in real-time to assist the surgeon in the excision of a tumor. Although the method has high sensitivity, the specificity can sometimes be lower than expected. Raman spectroscopy can detect tumors with high specificity. Therefore, a combination of both techniques can be advantageous. A complication that must be addressed is that the NIR spectral region is favored by both techniques for (in vivo) tissue analysis. When fluorescence and Raman emissions spectrally overlap, it becomes challenging or impossible to detect the Raman signal. In this paper, by avoiding this overlap, we describe a Raman spectroscopy setup capable of recording high-quality Raman spectra from tissue containing NIR exogenous fluorescent agents. We identify an optimal wavelength interval (900-915 nm) for Raman excitation, which avoids both excitation of fluorescent dyes and Raman signal self-absorption by the tissue. In this way, Raman spectroscopy can be combined with the currently most-used NIR fluorescent dyes. This combined novel setup could pave the way for clinical trials benefiting from both fluorescence imaging and Raman spectroscopy to avoid positive margins in cancer surgery.
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Corantes Fluorescentes , Neoplasias , Humanos , Corantes Fluorescentes/química , Análise Espectral Raman/métodos , Espectroscopia de Luz Próxima ao Infravermelho/métodos , Neoplasias/diagnóstico por imagem , Neoplasias/cirurgia , Imagem ÓpticaRESUMO
BACKGROUND: Carriers of loss-of-function mutations in the filaggrin gene (LoF FLG) have less natural moisturizing factor (NMF) in their stratum corneum (SC) and an increased risk of atopic dermatitis (AD). Natural moisturizing factor can be measured noninvasively by Raman spectroscopy. The use of Raman-derived NMF at birth to screen for FLG genotype could inform targeted AD prevention, but values in neonatal populations are largely unexplored. OBJECTIVE: To examine the associations between Raman-derived neonatal NMF measurements and FLG genotype. METHODS: Natural moisturizing factor was measured by Raman spectroscopy in the SC of the thenar eminence within 4 days of birth in 139 term neonates. Filaggrin genotyping was performed for 117 neonates (84%). RESULTS: The mean (SD) NMF was 0.37 (0.11) g/g protein, with values increasing across the first 3 days (day 1 vs 3: 0.29 [0.09] vs 0.43 [0.08, P < .001]). Twelve infants (10.3%) were carriers of LoF FLG, all heterozygous. Natural moisturizing factor was lower in LoF FLG carriers compared with wild-type (0.27 [0.08] vs 0.38 [0.11] g/g protein, P ≤ .001). Natural moisturizing factor had good discriminatory power for FLG genotype (area under the receiver operating curve [AUROC]: 0.79; 95% CI: 0.66, 0.91; P ≤ .001). This improved after correcting day 1 and 2 measurements to day 3 (AUROC: 0.83; 95% CI: 0.75, 0.92; P < .001). CONCLUSION: This study suggests that Raman-derived NMF measured in the early postnatal period may have the potential to classify by FLG genotype. The full translational value of this needs to be determined.
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Dermatite Atópica/genética , Genótipo , Mutação/genética , Proteínas S100/genética , Pele/patologia , Análise Espectral Raman/métodos , Eczema , Feminino , Proteínas Filagrinas , Predisposição Genética para Doença , Heterozigoto , Humanos , Higroscópicos/metabolismo , Lactente , Recém-Nascido , Masculino , Pele/metabolismoRESUMO
BACKGROUND: Clinical diagnosis of early melanoma (Breslow thickness less than 0.8 mm) is crucial to disease-free survival. However, it is subjective and can be exceedingly difficult, leading to missed melanomas, or unnecessary excision of benign pigmented skin lesions. An objective technique is needed to improve the diagnosis of early melanoma. METHODS: We have developed a method to improve diagnosis of (thin) melanoma, based on Raman spectroscopy. In an ex vivo study in a tertiary referral (pigmented lesions) centre, high-wavenumber Raman spectra were collected from 174 freshly excised melanocytic lesions suspicious for melanoma. Measurements were performed on multiple locations within the lesions. A diagnostic model was developed and validated on an independent data set of 96 lesions. RESULTS: Approximately 60% of the melanomas included in this study were melanomas in situ. The invasive melanomas had an average Breslow thickness of 0.89 mm. The diagnostic model correctly classified all melanomas (including in situ) with a specificity of 43.8%, and showed a potential improvement of the number needed to treat from 6.0 to 2.7, at a sensitivity of 100%. CONCLUSION: This work signifies an important step towards accurate and objective clinical diagnosis of melanoma and in particular melanoma with Breslow thickness <0.8 mm.
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Detecção Precoce de Câncer , Melanoma/diagnóstico , Neoplasias Cutâneas/diagnóstico , Análise Espectral Raman/métodos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Reprodutibilidade dos Testes , Adulto JovemRESUMO
Oncological applications of Raman spectroscopy have been contemplated, pursued, and developed at academic level for at least 25 years. Published studies aim to detect pre-malignant lesions, detect cancer in less invasive stages, reduce the number of unnecessary biopsies and guide surgery towards the complete removal of the tumour with adequate tumour resection margins. This review summarizes actual clinical needs in oncology that can be addressed by spontaneous Raman spectroscopy and it provides an overview over the results that have been published between 2007 and 2017. An analysis is made of the current status of translation of these results into clinical practice. Despite many promising results, most of the applications addressed in scientific studies are still far from clinical adoption and commercialization. The main hurdles are identified, which need to be overcome to ensure that in the near future we will see the first Raman spectroscopy-based solutions being used in routine oncologic diagnostic and surgical procedures.
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Neoplasias/diagnóstico por imagem , Neoplasias/cirurgia , Análise Espectral Raman , HumanosRESUMO
BACKGROUND/AIMS: Sensitive skin (SS), a frequently reported condition in the Western world, has been suggested to be underlined by an impaired skin barrier. The aim of this study was to investigate the skin barrier molecular composition in SS subjects using confocal Raman microspectroscopy (CRS), and to compare it with that of non-SS (NSS) individuals as well as atopic dermatitis (AD) and allergic rhinoconjunctivitis (AR) subjects, who frequently report SS. METHODS: Subjects with SS (n = 29), NSS (n = 30), AD (n = 11), and AR (n = 27) were included. Stratum corneum (SC) thickness, water, ceramides/fatty acids, and natural moisturizing factor (NMF) were measured by CRS along with transepidermal water loss and capacitance on the ventral forearm, thenar, and cheek. Sebum levels were additionally measured on the forearm and cheek. RESULTS: No differences between SS and NSS subjects were found regarding SC thickness, water, and NMF content, yet a trend towards lower ceramides/fatty acids was observed in the cheek. Compared to AD subjects, the SS group showed higher ceramides/fatty acid content in the forearm, whereas no differences emerged with AR. The correlation of macroscopic biophysical techniques and CRS was weak, yet CRS confirmed the well-known lower content of NMF and water, and thinner SC in subjects with filaggrin mutations. CONCLUSION: The skin barrier in SS is not impaired in terms of SC thickness, water, NMF, and ceramides/fatty acid content. The failure of biophysical techniques to follow alterations in the molecular composition of the skin barrier revealed by CRS emphasizes a strong need in sensitive and specific tools for in vivo skin barrier analysis.
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Pele/metabolismo , Adolescente , Adulto , Ceramidas/análise , Conjuntivite Alérgica/metabolismo , Dermatite Atópica/metabolismo , Ácidos Graxos/análise , Feminino , Proteínas Filagrinas , Glicerol/farmacocinética , Humanos , Masculino , Rinite Alérgica/metabolismo , Pele/química , Absorção Cutânea , Análise Espectral Raman , Adulto JovemRESUMO
Melanoma is a pigmented type of skin cancer, which has the highest mortality of all skin cancers. Because of the low clinical diagnostic accuracy for melanoma, an objective tool is needed to assist clinical assessment of skin lesions that are suspected of (early) melanoma. The aim of this study was to identify spectral differences in the CH region of HWVN (high-wavenumber) Raman spectra between melanoma and benign melanocytic lesions clinically suspected of melanoma. We used these spectral differences to explore preliminary classification models to distinguish melanoma from benign melanocytic lesions. Data from 82 freshly excised melanocytic lesions clinically suspected of melanoma were measured using an in-house built Raman spectrometer, which has been optimized for measurements on pigmented skin lesions (excitation wavelength 976 nm and a wavelength range of the Raman signal 1340-1540 nm). Clear spectral differences were observed between melanoma and benign melanocytic lesions. These differences can be assigned mainly to the symmetric CH2 stretching vibrations of lipids. Our results show that the Raman bands between 2840 and 2930 cm(-1) have increased intensity for melanoma when compared to benign melanocytic lesions, suggesting an increase in lipid content in melanoma. These results demonstrate that spectroscopic information in the CH-stretching region of HWVN Raman spectra can discriminate melanoma from benign melanocytic lesions that are often clinically misdiagnosed as melanoma and that Raman spectroscopy has the potential to provide an objective clinical tool to improve the clinical diagnostic accuracy of skin lesions suspected of melanoma.
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Melanoma/patologia , Neoplasias Cutâneas/patologia , Análise Espectral Raman , Área Sob a Curva , Análise Discriminante , Humanos , Melanócitos/química , Melanócitos/citologia , Melanoma/química , Análise de Componente Principal , Curva ROC , Neoplasias Cutâneas/químicaRESUMO
BACKGROUND: Penetration, autoxidation and N-acetylation of p-phenylenediamine (PPD) have been studied in vitro and ex vivo. However, a clear understanding of in vivo PPD penetration and the formation of PPD derivatives is lacking. OBJECTIVES: To obtain insights into the in vivo penetration, clearance and formation of PPD derivatives in human skin. METHODS: Patch test chambers containing PPD 1% pet. were applied to the forearms of two human volunteers, with increasing application times. Non-invasive Raman microspectroscopy was used for detection of PPD (derivatives) in skin at several follow-up times. RESULTS: Application of a PPD 1% pet. patch for 30 min resulted in substantial amounts of PPD in the stratum corneum of 90 mg PPD/g keratin. PPD contents were highest after three applications for 1 h each (330 mg PPD/g keratin), followed by single applications for 2 h 40 min, 2 h, and 23 h. The PPD half-time in the skin was 3 h. No spectral contributions of Bandrowski's base, monoacetyl-PPD and diacetyl-PPD were detected. CONCLUSIONS: We have gained insights into the in vivo penetration of PPD in human skin by using non-invasive Raman spectroscopy. Penetration into the skin was fast, and the PPD concentrations detected in the stratum corneum were high. PPD was detected in both the stratum corneum and the viable epidermis. Oxidized or acetylated PPD derivatives could not be detected.
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Corantes/metabolismo , Corantes/farmacocinética , Fenilenodiaminas/metabolismo , Fenilenodiaminas/farmacocinética , Absorção Cutânea , Pele/metabolismo , Acetilação , Adulto , Corantes/administração & dosagem , Feminino , Meia-Vida , Humanos , Masculino , Fenilenodiaminas/administração & dosagem , Pele/química , Análise Espectral RamanAssuntos
Melanoma , Neoplasias Cutâneas , Diagnóstico Diferencial , Humanos , Análise Espectral RamanRESUMO
An important feature of atopic eczema (AE) is a decreased skin barrier function. The stratum corneum (SC) lipids - comprised of ceramides (CERs), free fatty acids (FFAs) and cholesterol - fulfil a predominant role in the skin barrier function. In this clinical study, the carbon chain length distribution of SC lipids (FFAs and CERs) and their importance for the lipid organization and skin barrier function were examined in AE patients and compared with control subjects. A reduction in FFA chain length and an increase in unsaturated FFAs are observed in non-lesional and lesional SC of AE patients. The reduction in FFA chain length associates with a reduced CER chain length, suggesting a common synthetic pathway. The lipid chain length reduction correlates with a less dense lipid organization and a decreased skin barrier function. All changes are more pronounced in lesional SC compared with non-lesional skin. No association was observed between lipid properties and filaggrin mutations, an important predisposing factor for developing AE. The results of this study demonstrate an altered SC lipid composition and signify the importance of these changes (specifically regarding the CER and FFA chain lengths) for the impaired skin barrier function in AE. This provides insights into epidermal lipid metabolism as well as new opportunities for skin barrier repair.
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Dermatite Atópica/metabolismo , Ácidos Graxos não Esterificados/metabolismo , Pele/metabolismo , Adulto , Estudos de Casos e Controles , Ceramidas/química , Ceramidas/metabolismo , Colesterol/metabolismo , Dermatite Atópica/genética , Epiderme/metabolismo , Ácidos Graxos não Esterificados/química , Feminino , Proteínas Filagrinas , Humanos , Proteínas de Filamentos Intermediários/genética , Metabolismo dos Lipídeos , Masculino , Mutação , Espectroscopia de Infravermelho com Transformada de Fourier , Adulto JovemRESUMO
We have developed a method to determine the limit of detection (LoD) for quantitative measurement of exogenous analytes in the outer layer of the human skin by in vivo confocal Raman spectroscopy. The method is in accordance with the guidelines of the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use that have been adopted by regulatory authorities such as the American Food and Drug Administration and the European Medicines Agency. The method can be applied in silico so that the limit of detection can be assessed before starting a skin penetration study, for example, in areas of pharmaceutical formulation, pharmacokinetics, or toxicokinetics. This can significantly reduce the need for expensive and time-consuming feasibility studies. This paper describes the method to calculate this LoD as well as the experimental and methodological factors that can influence the calculation of the LoD.
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A hallmark of atopic eczema (AE) is skin barrier dysfunction. Lipids in the stratum corneum (SC), primarily ceramides, fatty acids, and cholesterol, are crucial for the barrier function, but their role in relation to AE is indistinct. Filaggrin is an epithelial barrier protein with a central role in the pathogenesis of AE. Nevertheless, the precise causes of AE-associated barrier dysfunction are largely unknown. In this study, a comprehensive analysis of ceramide composition and lipid organization in nonlesional SC of AE patients and control subjects was performed by means of mass spectrometry, infrared spectroscopy, and X-ray diffraction. In addition, the skin barrier and clinical state of the disease were examined. The level of ceramides with an extreme short chain length is drastically increased in SC of AE patients, which leads to an aberrant lipid organization and a decreased skin barrier function. Changes in SC lipid properties correlate with disease severity but are independent of filaggrin mutations. We demonstrate for the first time that changes in ceramide chain length and lipid organization are directly correlated with the skin barrier defects in nonlesional skin of AE patients. We envisage that these insights will provide a new therapeutic entry in therapy and prevention of AE.
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Ceramidas/metabolismo , Dermatite Atópica/metabolismo , Metabolismo dos Lipídeos , Adulto , Ceramidas/química , Dermatite Atópica/genética , Feminino , Proteínas Filagrinas , Genótipo , Humanos , Proteínas de Filamentos Intermediários/genética , Masculino , Estrutura Molecular , MutaçãoRESUMO
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.
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Dermatite Atópica/genética , Predisposição Genética para Doença , Proteínas de Filamentos Intermediários/genética , Fenótipo , Pele/metabolismo , Criança , Feminino , Proteínas Filagrinas , Genótipo , Humanos , Irlanda , Masculino , Mutação , Análise Espectral RamanRESUMO
The composition of topical and transdermal formulations is known to determine the rate and the extent of drug delivery to and through the skin. However, to date, the role of excipients in these formulations on skin delivery of actives has received little attention from scientists in the field. Monitoring skin absorption of both drug and vehicle may provide insights into the mechanism by which excipients promote permeation and may facilitate the design of effective and safer products. Previously, we have investigated the use of quantitative Confocal Raman Spectroscopy (CRS) to investigate the delivery of an active to the skin, and we also reported the first fully quantitative study that compared this method with the well-established in vitro permeation test (IVPT) model. To further explore the potential of quantitative CRS in assessing topical delivery, the present work investigated the effects of commonly used excipients on the percutaneous absorption of a model drug, ibuprofen (IBU). Permeation of IBU and selected solvents following finite dose applications to human skin was determined in vitro and in vivo by Franz diffusion studies and quantitative CRS, respectively. The solvents used were propylene glycol (PG), dipropylene glycol (DPG), tripropylene glycol (TPG), and polyethylene glycol 300 (PEG 300). Overall, the cumulative amounts of IBU that permeated at 24 h in vitro were similar for PG, DPG, and TPG (p > 0.05). These three vehicles outperformed PEG 300 (p < 0.05) in terms of drug delivery. Concerning the vehicles, the rank order for in vitro skin permeation was DPG ≥ PG > TPG, while PEG 300 did not permeate the skin. A linear relationship between maximum vehicle and IBU flux in vitro was found, with a correlation coefficient (R2) of 0.95. When comparing in vitro with in vivo data, a positive in vitro-in vivo (IVIV) correlation between the cumulative permeation of IBU in vitro and the total amount of IBU that penetrated the stratum corneum (SC) in vivo was observed, with a Pearson correlation coefficient (R2) of 0.90. A strong IVIV correlation, R2 = 0.82, was found following the linear regression of the cumulative number of solvents permeated in vitro and the corresponding skin uptake in vivo measured with CRS. This is the first study to correlate in vivo permeation of solvents measured by CRS with data obtained by in vitro diffusion studies. The IVIV correlations suggest that CRS is a powerful tool for profiling drug and vehicle delivery from dermal formulations. Future studies will examine additional excipients with varying physicochemical properties. Ultimately, these findings are expected to lead to new approaches for the design, evaluation, and optimization of formulations that target actives to and through the skin.
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The goal of head and neck oncological surgery is complete tumor resection with adequate resection margins while preserving acceptable function and appearance. For oral cavity squamous cell carcinoma (OCSCC), different studies showed that only 15%-26% of all resections are adequate. A major reason for the low number of adequate resections is the lack of information during surgery; the margin status is only available after the final histopathologic assessment, days after surgery. The surgeons and pathologists at the Erasmus MC University Medical Center in Rotterdam started the implementation of specimen-driven intraoperative assessment of resection margins (IOARM) in 2013, which became the standard of care in 2015. This method enables the surgeon to turn an inadequate resection into an adequate resection by performing an additional resection during the initial surgery. Intraoperative assessment is supported by a relocation method procedure that allows accurate identification of inadequate margins (found on the specimen) in the wound bed. The implementation of this protocol resulted in an improvement of adequate resections from 15%-40%. However, the specimen-driven IOARM is not widely adopted because grossing fresh tissue is counter-intuitive for pathologists. The fear exists that grossing fresh tissue will deteriorate the anatomical orientation, shape, and size of the specimen and therefore will affect the final histopathologic assessment. These possible negative effects are countered by the described protocol. Here, the protocol for specimen-driven IOARM is presented in detail, as performed at the institute.
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Neoplasias Bucais , Carcinoma de Células Escamosas/cirurgia , Humanos , Cuidados Intraoperatórios , Margens de Excisão , Neoplasias Bucais/cirurgiaRESUMO
OBJECTIVE: The depth of invasion (DOI) is considered an independent risk factor for occult lymph node metastasis in oral cavity squamous cell carcinoma (OCSCC). It is used to decide whether an elective neck dissection (END) is indicated in the case of a clinically negative neck for early stage carcinoma (pT1/pT2). However, there is no consensus on the cut-off value of the DOI for performing an END. The aim of this study was to determine a cut-off value for clinical decision making on END, by assessing the association of the DOI and the risk of occult lymph node metastasis in early OCSCC. METHODS: A retrospective cohort study was conducted at the Erasmus MC, University Medical Centre Rotterdam, The Netherlands. Patients surgically treated for pT1/pT2 OCSCC between 2006 and 2012 were included. For all cases, the DOI was measured according to the 8th edition of the American Joint Committee on Cancer guideline. Patient characteristics, tumor characteristics (pTN, differentiation grade, perineural invasion, and lymphovascular invasion), treatment modality (END or watchful waiting), and 5-year follow-up (local recurrence, regional recurrence, and distant metastasis) were obtained from patient files. RESULTS: A total of 222 patients were included, 117 pT1 and 105 pT2. Occult lymph node metastasis was found in 39 of the 166 patients who received END. Univariate logistic regression analysis showed DOI to be a significant predictor for occult lymph node metastasis (odds ratio (OR) = 1.3 per mm DOI; 95% CI: 1.1-1.5, p = 0.001). At a DOI of 4.3 mm the risk of occult lymph node metastasis was >20% (all subsites combined). CONCLUSION: The DOI is a significant predictor for occult lymph node metastasis in early stage oral carcinoma. A NPV of 81% was found at a DOI cut-off value of 4 mm. Therefore, an END should be performed if the DOI is >4 mm.
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Previously, we reported the use of Confocal Raman Spectroscopy (CRS) to investigate the topical delivery of actives and excipients. We have also correlated the results from CRS with findings from in vitro diffusion studies in human skin. However, until now CRS has only been used as a semi-quantitative method of determining the skin uptake of molecules, with results expressed as arbitrary units of signal intensity. Clearly, this posed challenges for using CRS to determine skin delivery and to assess the drug bioavailability and bioequivalence of topical formulations. In the present work, the permeation of niacinamide (NIA) from various formulations in human skin was studied in vitro using conventional Franz cells and in vivo using a quantitative CRS method under finite dose conditions. The selection of NIA was based on its wide use in pharmaceutical and personal care formulations for many years. This is the first fully quantitative study to compare these methods. The vehicles investigated were neat Transcutol® P (TC); binary combinations of propylene glycol (PG) with propylene glycol monolaurate (PGML); and ternary mixtures of PG, PGML, and isopropyl myristate (IPM). These solvents were selected to encompass a range of physicochemical properties. NIA permeation was evident from all formulations in vitro and in vivo. The vehicles PG:PGML and PG:PGML:IPM delivered comparable amounts across the skin in vitro at 24 h (100.3-106.7 µg/cm2, p > 0.05) that were significantly higher compared with those of TC (1.3 µg/cm2, p < 0.05). An excellent in vitro in vivo correlation (R2 = 0.98) was found following the linear regression of the cumulative amounts of NIA permeated in vitro and the amounts of NIA at 2 µm in the skin measured with CRS. A very good correlation between the cumulative permeation of NIA in vitro and the total amount of NIA that penetrated the stratum corneum (SC) per unit of surface area (µg/cm2) in vivo was also observed, with a Pearson correlation coefficient (R2) of 0.94. The findings support the use of CRS for the quantitative measurement of actives delivered to the skin in vivo. Future studies will focus on exploring the reproducibility and reliability of the method by investigating the delivery of different actives from a wider range of vehicles. Additionally, quantitative CRS will be evaluated further as a method for assessing the bioequivalence of topical formulations.
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The high-frequency electrical conductance of tape-stripped human skin in vivo can be used to evaluate the hydration profile of Stratum corneum (SC). Tape-stripping provides access to the underlying SC layers, and the conductance of these layers (as measured by the Skicon instrument) correlates well with their water content, as demonstrated by independent confocal Raman spectroscopic measurements. The correlation shows high inter-individual variance and is not linear over the full measurement range of the instrument, but is helpful to discriminate between dry, normal and highly hydrated SC. The depth profile of hydration in tape-stripped SC corresponds to the one in intact SC only if the barrier function of the skin is not impaired. Thus, conductometry of tape-stripped skin must be used in conjunction with a method that allows to estimate the barrier damage inflicted to SC during the tape-stripping procedure, for example, measurement of the trans-epidermal water loss. The methodology described here is simple, rapid and minimally invasive, and it employs commercially available instrumentation that is cheap, portable and easy to use. This approach is applicable to in vivo estimation of the SC hydration in studies in the areas of dermatology, skin care and transdermal drug delivery.