Blind source separation of molecular components of the human skin in vivo: non-negative matrix factorization of Raman microspectroscopy data.

Determination of the molecular composition of the skin is crucial for numerous tasks in medicine, pharmacology, dermatology and cosmetology. Confocal Raman microspectroscopy is a sensitive method for the evaluation of molecular depth profiles in the skin in vivo. Since the Raman spectra of most of the skin constituents significantly superimpose, a spectral decomposition by a set of predefined library components is usually performed to disentangle their contributions. However, the incorrect choice of the number and type of components or differences between the spectra of the basic components measured in vitro and in vivo can lead to incorrect results of the decomposition procedure. Here, we investigate an alternative data-driven approach based on a non-negative matrix factorization (NNMF) algorithm of depth-resolved Raman spectra of skin that does not require a priori information of spectral data for the analysis. Using the model and experimentally measured depth-resolved Raman spectra of the upper epidermis in vivo, we show that NNMF provides depth profiles of endogenous molecular components and exogenous agents penetrating through the upper epidermis for the spectra and concentration. Moreover, we demonstrate that this approach is capable of providing new information on the molecular profiles of the skin.

Stratum corneum occlusion induces water transformation towards lower bonding state: a molecular level in vivo study by confocal Raman microspectroscopy.

OBJECTIVE: It is conventionally understood that occlusive effects are the retention of excessive water in the stratum corneum (SC), the increase of SC thickness (swelling) and a decrease of the transepidermal water loss. However, the influence of occlusion on water binding properties in the SC is unknown. METHODS: The action of plant-derived jojoba and almond oils, as well as mineral-derived paraffin oil and petrolatum topically applied on human skin, is investigated in vivo using confocal Raman microspectroscopy. To understand the oils' influence on the SC on the molecular level, the depth-dependent hydrogen bonding states of water in the SC and their relationship to the conformation of keratin, concentration of natural moisturizing factor (NMF) molecules and lipid organization were investigated. RESULTS: A significant SC swelling was observed only in petrolatum-treated skin. The water concentration was increased in oil-treated skin in the intermediate SC region (40-70% SC depth). Meanwhile, the amount of free, weakly and tightly bound water increased, and strongly bound water decreased in the uppermost SC region (0-30% SC depth). The NMF concentration of oil-treated skin was significantly lower at 50-70% SC depth. The lateral organization of lipids in oil-treated skin was lower at 0-30% SC depth. The secondary structure of keratin was changed towards an increase of ß-sheet content in mineral-derived oil-treated skin and changed towards an increase of α-helix content in plant-derived oil-treated skin. CONCLUSION: The occlusive properties can be summarized as the increase of free water and the transformation of water from a more strongly to a more weakly hydrogen bonding state in the uppermost SC, although some oils cause insignificant changes of the SC thickness. The accompanied changes in the keratin conformation at the intermediate swelling region of the SC also emphasize the role of keratin in the SC's water-transporting system, that is the water in the SC transports intercellularly and intracellularly in the intermediate swelling region and only intercellularly in the uppermost non-swelling region. Bearing this in mind, almond, jojoba and paraffin oils, which are not occlusive from the conventional viewpoint, have an occlusion effect similar to petrolatum on the SC.

OBJECTIF: Il est généralement entendu que les effets occlusifs consistent en la rétention d'un excès d'eau dans la couche cornée (stratum corneum, SC), l'augmentation d'épaisseur de la SC (gonflement) et une diminution de la perte d'eau trans-épidermique. Cependant, l'influence de l'occlusion sur les propriétés de fixation de l'eau dans le SC est inconnue. MÉTHODES: L'action des huiles de jojoba et d'amande d'origine végétale, ainsi que des huiles de paraffine et de pétrolatum d'origine minérale appliquées topiquement sur la peau humaine est étudiée in vivo à l'aide de la microspectroscopie Raman confocale. Pour comprendre l'influence des huiles sur le SC au niveau moléculaire, on a étudié les états de liaison hydrogène de l'eau dans le SC en fonction de la profondeur et leur relation avec la conformation de la kératine, la concentration des molécules du facteur naturel d'hydratation (NMF) et l'organisation des lipides. RÉSULTATS: Un gonflement significatif de le SC n'a été observé que dans la peau traitée au pétrolatum. La concentration en eau a été augmentée dans la peau traitée au pétrolatum dans la région SC intermédiaire (40-70% de profondeur du SC). En meme temps, la quantité d'eau libre, faiblement et fortement liée augmentait, tandis que l'eau fortement liée diminuait dans la région SC supérieure (0-30% de profondeur du SC). La concentration en NMF de la peau traitée à l'huile était plus basse d´une manière significative à 50-70% de profondeur du SC. L'organisation latérale des lipides dans la peau huilée était plus basse à une profondeur du SC de 0 à 30 %. La structure secondaire de la kératine a été modifiée pour augmenter la teneur en feuillet-ß dans les peaux huilées d'origine minérale et pour augmenter la teneur en hélice α dans les peaux huilées d'origine végétale. CONCLUSION: Les propriétés occlusives peuvent être résumées comme l'augmentation de l'eau libre et la transformation de l'eau d'un état de liaison hydrogène plus fort à un état de liaison hydrogène plus faible dans le SC supérieure, bien que certaines huiles provoquent des changements insignifiants de l'épaisseur de la SC. Les modifications de la conformation de la kératine dans la zone de gonflement intermédiaire du SC soulignent également le rôle de la kératine dans le système de transport de l'eau du SC, c'est-à-dire que l'eau est transportée du SC de manière intercellulaire et intracellulaire dans la zone de gonflement intermédiaire et seulement de manière intercellulaire dans la zone non gonflée la plus élevée. En considérant cela, les huiles d'amande, de jojoba et de paraffine, qui ne sont pas occlusives du point de vue conventionnel, ont un effet d'occlusion similaire à celui du pétrolatum sur le SC.

Skin type differences in solar-simulated radiation-induced oxidative stress.

BACKGROUND: Human skin protects the body from external damage, pathogens and oxidative stress factors such as ultraviolet (UV) radiation. Excessive exposure to UV radiation can lead to increased production of free radicals and hence to skin damage such as inflammation, premature skin ageing and skin cancer. Besides UV, the visible and near infrared (NIR) regions are also a source of radical production. Half of all free radicals are induced by the visible + NIR region of the solar spectrum in people with skin types I-III, but data on the effects in people with skin types IV-VI are missing. OBJECTIVES: This in vivo pilot study addressed the distribution of radical production in skin types IV and V during irradiation in the UV, visible and NIR spectral regions, comparing the first results with those of skin type II. METHODS: The measurements were performed in vivo using L-band electron paramagnetic resonance spectroscopy and the spin probe PCA. RESULTS: In skin types IV-V most radicals were induced in the visible + NIR region, followed by the NIR and UV regions of the sun spectrum. Significantly (P ≤ 0·05) more radicals were induced in skin types IV-V than in type II during NIR irradiation, whereas skin types IV-V exhibited significantly less UV-induced radicals (P ≤ 0·01) than skin type II. CONCLUSIONS: All spectral regions (UV, visible and NIR) cause free radical formation in skin types II and IV-V. After 4 min of solar-simulated exposure (UV-NIR), the radical formation in skin types IV-V is 60% of that in skin type II. Therefore people with darker skin types also need solar protection.

Methods for Optical Skin Clearing in Molecular Optical Imaging in Dermatology.

This short review describes recent progress in using optical clearing (OC) technique in skin studies. Optical clearing is an efficient tool for enhancing the probing depth and data quality in multiphoton microscopy and Raman spectroscopy. Here, we discuss the main mechanisms of OC, its safety, advantages, and limitations. The data on the OC effect on the skin water content are presented. It was demonstrated that 70% glycerol and 100% OmnipaqueTM 300 reduce the water content in the skin. Both OC agents (OCAs) significantly affect the strongly bound and weakly bound water. However, OmnipaqueTM 300 causes considerably less skin dehydration than glycerol. In addition, the results of examination of the OC effect on autofluorescence in two-photon excitation and background fluorescence in Raman scattering at different skin depths are presented. It is shown that OmnipaqueTM 300 is a promising OCA due to its ability to reduce background fluorescence in the upper skin layers. The possibility of multimodal imaging combining optical methods and OC technique is discussed.

Label-Free Multiphoton Microscopy: The Origin of Fluorophores and Capabilities for Analyzing Biochemical Processes.

Multiphoton microscopy (MPM) is a method of molecular imaging and specifically of intravital imaging that is characterized by high spatial resolution in combination with a greater depth of penetration into the tissue. MPM is a multimodal method based on detection of nonlinear optical signals - multiphoton fluorescence and optical harmonics - and also allows imaging with the use of the parameters of fluorescence decay kinetics. This review describes and discusses photophysical processes within major reporter molecules used in MPM with endogenous contrasts and summarizes several modern experiments that illustrate the capabilities of label-free MPM for molecular imaging of biochemical processes in connective tissue and cells.

Multiphotonic staging of chronic wounds and evaluation of sterile, optical transparent bacterial nanocellulose covering: A diagnostic window into human skin.

BACKGROUND: Diagnostics of healing, infection, and inflammation in chronic wounds in comparison with physiological wound healing in acute wounds may help for therapy decisions toward individualized therapy management. With emerging new optical techniques the coupling of optical diagnostic devices with tissue provides a great challenge. Traditional coupling with cover slips is used since the early days of microscopy. In modern health care, hygienic covering of surfaces is necessary to avoid infections and cross-contaminations. METHODS: Measurements in chronic wounds were performed at three different areas including the center of the wound, the border of the wound and healthy skin as comparison area. For each measurement area, three vertical stacks were taken by MPT. Additionally, three different optical measuring procedures (MPT, OCT, CLSM) were used for the examination of BNC foil. Examinations of BNC foil were carried out at two different areas of healthy skin compared to a standard setup as control. RESULTS: The MPT evaluation revealed a distinct difference in the second harmonic generation-to-autofluorescence aging index of dermis (SAAID) behavior between the vertical stacks taken at central wound areas and wound margins as well as unaffected skin. Through BNC foil covers, MPT CLSM and OCT images were captured with good quantitative and qualitative results. CONCLUSIONS: Phases in chronic wounds could be matched with physiologically healing in acute wounds according to SAAID and MPT imaging. BNC provided an alternative covering for MPT, OCT, and CLSM with clear morphological images.

Kinetics and tissue repair process following fractional bipolar radiofrequency treatment.

INTRODUCTION: Fractionated radiofrequency (RF) tissue tightening is an alternative method to fractionated laser treatment of skin wrinkling, laxity and acne scars, with reduced risk of scarring or persistent pigmentation. The aim of this study was to evaluate and quantify the wound healing process after RF treatment. MATERIALS AND METHODS: 12 patients were treated with a 64-pin fractional bipolar RF device with 60 mJ/pin applied energy. Confocal laser scanning microscopy (CLSM) examination was performed on day 1, day 2, day 7 and day 14 after treatment. Clinical wound healing process was measured and expressed as a percentage. RESULTS: All patients developed erythema, mild edema and crusts at the treated areas. Two weeks after treatment clinical symptoms resolved. During ablation patients reported moderate pain. Directly after ablation microscopic ablation zones could be detected in CLSM. Measurement of MAZ at epidermis, dermo-epidermal junction and papilary dermis showed a constant diameter until two weeks after treatment. Re-epithelization of the MAZ could be detected already 1 week after treatment. However, 2 weeks after ablation the honeycomb pattern of the epidermis was not yet completely restored. DISCUSSION: Bipolar fractionated RF treatment demonstrates clinically a rapid wound healing response. The subepidermal remodelling process still ongoing after 14 days, showing new granulation tissue. Therefore, treatment intervals of at least 14 days should be recommended to allow completion of the remodelling process.

[Follicular penetration of nanocarriers is an important penetration pathway for topically applied drugs]. / Bedeutung des follikulären Penetrationswegs für den Wirkstofftransport mittels Nanocarriern.

BACKGROUND: The hair follicle represents a significant penetration route for topically applied substances. ISSUE: The percutaneous absorption of substances can be significantly increased and accelerated by the involvement of hair follicles. In addition, nanoparticles have the characteristic to penetrate deeply and effectively into the hair follicles. MATERIALS AND METHODS: An optimization of drug delivery for topically applied substances is possible if the nanoparticles act solely as a carrier to transport active ingredients into the hair follicle. Once the nanocarrier has penetrated into the hair follicle, the active substance must be released there. This can be triggered by various mechanisms. RESULTS: The released drug can thus pass into the living tissue surrounding the hair follicle independently. With the help of this innovative strategy, the bioavailability of topically applied substances can be significantly improved. CONCLUSION: The transport of active ingredients into the hair follicles with the help of particles and the release of active substances there is a very effective new method for transporting active substances through the skin barrier.

Non-invasive evaluation of human mucosal structures by multiphoton laser scanning tomography in vitro.

BACKGROUND: Mucous membranes may be affected by various diseases and the diagnostic accessibility is limited. Multiphoton laser tomography (MPT) is a useful tool for in vivo evaluation of superficial skin structures and might also be useful for the imaging of mucosa. OBJECTIVES: In order to investigate the suitability of MPT for the evaluation of mucous membranes, tissue samples of different donors and anatomical localizations have been imaged. METHODS: Human mucosa samples from the urinary bladder, palatine tonsil and ocular conjunctiva were investigated by MPT and subsequently compared with conventional histology. RESULTS: Horizontal images of the epithelium and the underlying connective tissue were obtained using the autofluorescence and second harmonics generation signals by MPT. The stratification of multilayered epithelium was consistently reproduced. Also inflammatory changes like lymphocytic infiltrates and widened intercellular spaces were imaged in some cases. CONCLUSION: Mucous tissue samples were obtained and MPT imaging was successfully performed. In the direct comparison with histology of the same tissue samples, the characteristic morphology of the epithelia was found. Further studies are necessary in order to compare images of healthy and diseased mucosal epithelium and the use of MPT for its evaluation in vivo.

Prevention of palmoplantar erythrodysesthesia in patients treated with pegylated liposomal doxorubicin (Caelyx®).

PURPOSE: Palmoplantar erythrodysesthesia (PPE) is one of the most frequent side effects during systemic treatment with pegylated liposomal doxorubicin (PLD, Caelyx®). PPE lesions show a range of symptoms, from numbness to painful erosions, and can have a major impact on the quality of life in affected patients. Previously, a possible pathomechanism of PPE was found in doxorubicin-treated patients based on radical formation in the skin. Here, a preventive strategy using a topically applied ointment with a high radical protection factor was investigated. METHODS: In this randomized placebo-controlled double-blind study the antioxidant-containing ointment was compared with a placebo ointment regarding PPE grade III occurrence, overall PPE grade I-III occurrence and PPE severity in PLD patients. The verum or placebo cream was topically applied for a period of 16 weeks, starting 3 days prior to the first cycle of chemotherapy. Clinical evaluations were carried out by a dermatologist prior to the first cycle of chemotherapy and every 4 weeks for the duration of 16 weeks. RESULTS: Thirty-two patients were enrolled in total, of which 17 (66%) completed the study. No PPE grade III was found in the verum group, while five out of seven patients (71%) had to be unblinded in the placebo arm due to PPE grade III (p = 0.003). General PPE occurrence of all grades was 60% under verum and 86% under placebo treatment. CONCLUSIONS: The preventive application of an antioxidant-containing ointment was shown to be significantly more effective in the prevention of PPE grade III compared to placebo treatment.

Fluorescence detection of protein content in house dust: the possible role of keratin.

We propose a fluorescence method for protein content assessment in fine house dust, which can be used as an indicator of the hygienic state of occupied rooms. The results of the measurements performed with 30 house dust samples, including ultrafiltration experiments, strongly suggest that the fluorescence emission of house dust extracts excited at 350 nm is mainly due to protein fragments, which are presumably keratin hydrolysates. This suggestion is supported by several facts: (i) Spectral band shapes for all the samples under investigation are close and correspond to that of keratin; (ii) fluorescence intensity correlates with the total protein content as provided by Lowry assay; (iii) treatment of the samples with proteinase K, which induces keratin hydrolysis, results in fluorescence enhancement without changing fluorescence band shape; and (iv) Raman spectra of keratin and fine house dust samples exhibit a very similar structure. Based on the obtained results and literature data, we propose a hypothesis that keratin is a major substrate for fluorescence species in fine house dust, which are responsible for emission at 350-nm excitation.

Examination of wound healing after curettage by multiphoton tomography of human skin in vivo.

BACKGROUND: The multiphoton tomography (MPT) has evolved into a useful tool for the non-invasive investigation of morphological and biophysical characteristics of human skin in vivo. Until now, changes of the skin have been evaluated mainly by using clinical and histological techniques. In this study, the progress of wound healing was investigated by MPT over 3 weeks with a final examination after 24 months. Especially, the collagen degradation, reepithelization and tissue formation were examined. As specific parameter for wound healing and its course the second-harmonic generation-to-autofluorescence aging index of dermis (SAAID) was used. METHODS: About 10 volunteers aged between 25 and 58 years were examined. Acute wounds were scanned with three Z-stacks taken per visit. The stacks were taken up to a depth of 225 µm at increments of 5 µm and a scan time for 3 seconds per scan. Subsequently, the SAAID was evaluated as an indicator for wound healing. Furthermore, single scans were taken for morphological investigations. RESULTS: The evaluation revealed a distinct difference in the SAAID behavior between the Z-stacks taken at each visit. Furthermore, the degradation of collagen and cells and their reappearance could be shown in the course of the visits. CONCLUSIONS: Clear differences in the curve behavior of the SAAID at every visit were shown in this study. The SAAID curves and morphological images could be correlated with findings of the clinical examination of different wound healing phases. Therefore, SAAID curves and morphological MPT imaging could provide a non-invasive tool for the determination of wound healing phases in patients in vivo.

Multiple spatially resolved reflection spectroscopy to monitor cutaneous carotenoids during supplementation of fruit and vegetable extracts in vivo.

BACKGROUND: A nutrition rich in fruit and vegetables and a healthy lifestyle become more and more important in the industrial countries to counteract oxidative stress and promote health. For many years, it has been possible to control human cutaneous carotenoids noninvasively by resonance Raman spectroscopic systems and by spatially resolved reflectance spectroscopy. METHODS: Ten volunteers took a commercially available fruit and vegetable extract daily for a time period of 5 weeks. A second group served as control group and did not take any supplements (10 volunteers). To monitor the status of the cutaneous carotenoids noninvasively, an optical sensor based on multiple spatially resolved reflectance spectroscopy was applied once a week. RESULTS: The study could demonstrate that the intake of the supplement significantly increase the cutaneous carotenoid values of the young adults by 50%. The control group without any supplementation showed also significantly increased values, ie, by 10%, which might be due to the fact that their lifestyle was controlled. CONCLUSION: The results illustrate that a biofeedback by measuring the skin carotenoids could improve the lifestyle of young adults and that a regular consumption of fruit and vegetables directly or as a drink can increase the concentration of cutaneous carotenoids significantly.

[New aspects in the development of sunscreening agents]. / Neue Aspekte bei der Entwicklung von Sonnenschutzmitteln.

Sunlight is indispensable for life. Skin aging is determined not only by genetic factors but also by the influence of solar radiation which in all its spectral regions induces free radicals in the human skin. Below a critical concentration, free radicals are important for signaling processes in the human body. Exceeding a critical threshold value (FRTV), however, free radicals lead to cellular damage which, in turn, can induce skin aging, immunosuppression and even skin cancer. Recently it could be shown that 50% of the free radicals induced in the human skin by solar radiation were caused by the visible and infrared spectral regions. Taking into consideration that people who use sunscreen stay considerably longer in the sun than unprotected people, their skin could be exposed to free radical formation in the visible and infrared spectral regions, which far exceeds the critical threshold. Filter substances like those used in sunscreens for skin protection in the ultraviolet region are not available for the visible and infrared spectral regions. Using the natural protection mechanisms of the human skin as examples, however, the protective effect of sunscreens can be considerably enhanced by the addition of scattering and reflective pigments, such as titanium dioxide, as well as of antioxidants which neutralize the free radicals. Currently the focus of sunscreen development is shifting from mere UV protection to protection covering the entire solar spectrum.

Optimization of the measurement procedure during multiphoton tomography of human skin in vivo.

BACKGROUND: The in vivo multiphoton tomography has evolved into a useful tool for the non-invasive investigation of morphological and biophysical characteristics of human skin. Until now, changes of skin have been evaluated mainly by clinical and histological techniques. The current study addresses the effects of a changed acquisition time for single scans in a Z-stack on the directly related qualitative and quantitative interpretability of the data. METHODS: A test area of the skin was used for scanning 12 Z-stacks of 10 volunteers aged between 25 and 34 years. The stacks were taken up to a depth of 220 µm at increments of 10 µm at four different times, 1, 3, 7, 13 s, per scan. Subsequently, the second harmonic generation (SHG)-to-autofluorescence aging index of dermis (SAAID) was evaluated at three different measuring depths, i.e. at the maximum of SHG as well as at depths of 60 and 150 µm. RESULTS: The evaluation did not reveal any significant differences in the SAAID behavior between the Z-stacks of each test area scanned at different acquisition times. However, the acquisition time of 1 s/frame increases the measurement stability without influencing the SAAID behavior. The resolution of subcellular structures decreases significantly at scan times ≤3 s, whereas the acquisition time from 7 to 13 s warrants a high image quality. CONCLUSION: The study has shown that there are no significant differences between the scan speeds per scan in a Z-stack and the resulting SAAID. Acquisition times of 7 s are suitable for the morphological evaluation whereas a further extension to 13 s does not result in any benefits. A scan time per image of 1 s is sufficient for the quantitative evaluation of SAAID thus substantially reducing the possible influence of movement artifacts.

Correlation of optical coherence tomography and histology in microcystic adnexal carcinoma: a case report.

BACKGROUND/AIMS: Herein, we report a case of microcystic adnexal carcinoma (MAC), which we correlated and evaluated by optical coherence tomography (OCT) and conventional H&E histology. METHODS: A commercially available OCT scanner was used for imaging. Several multi-slice images were obtained from the central portion of the lesion. Correlation of OCT findings with histology was performed retrospectively. RESULTS: Microcystic adnexal carcinoma showed characteristic criteria, which were divided into superficial and sub-epidermal findings. CONCLUSION: The use of OCT can visualize characteristic criteria of MAC, thus enabling prompt diagnosis before surgery.

Assessment of a scoring system for Basal Cell Carcinoma with multi-beam optical coherence tomography.

BACKGROUND: Multi-beam optical coherence tomography (OCT) is a novel method of non-invasive skin imaging allowing the evaluation of tissue at high level of lateral and axial resolution. It permits the horizontal and vertical evaluation of the extent of diseases. OBJECTIVE: Herein, we aimed to validate diagnosing basal cell carcinoma (BCC) by OCT using a newly developed scoring system ('Berlin Score'-BS). This was based on the predetermined criteria such as dark border underneath the tumour and ovoid structures. Their frequency and distribution in subtypes of BCC were evaluated. METHODS: The study was conducted in two phases, in which the experience of examiner differed. A total of 127 BCC and 50 other skin diseases were examined. In phase one, students performed the evaluation of skin lesions using the BS, while in phase two an expert performed the scoring in a different subset of patients. RESULTS: Application of BS by students revealed sensitivity and specificity of 92.8% (95% CI 85.4-96.8) and 24.1% (95% CI 11.0-43.4) when reaching the lower threshold BS≥8. The most common BCC subtypes were superficial (28.7%) and nodular (22.6%) BCC. Second phase was carried out to verify collected data by a dermatological specialist and expert in using OCT. Increased sensitivity and specificity for OCT amounted to 96.6% (95% CI 80.4-99.8) and 75.2% (95% CI 52.5-90.9). Thereby 88% of all diagnoses were correctly classified confirmed by histopathology. CONCLUSION: Multi-beam optical coherence tomography revealed to be a fast and promising device for assessing lesions by means of BS. Both students, who benefit from practice in handling OCT, and experts are able to perform this procedure. However, experience and training in the interpretation markedly increased sensitivity and specificity of the BS in our study. Moreover, redefinition and refining of the criteria seems necessary and may further increase the diagnostic value of OCT for NMSC.

Clinical use of cold atmospheric pressure argon plasma in chronic leg ulcers: A pilot study.

OBJECTIVE: In the age of multiresistant microbes and the increasing lack of efficient antibiotics, conventional antiseptics play a critical role in the prevention and therapy of wound infections. Recent studies have demonstrated the antiseptic effects of cold atmospheric pressure plasma (APP). In this pilot, study we investigate the overall suitability of one of the first APP sources for wound treatment focusing on its potential antimicrobial effects. METHOD: The wound closure rate and the bacterial colonisation of the wounds were investigated. Patients suffering from chronic leg ulcers were treated in a clinical controlled monocentric trial with either APP or octenidine (OCT). In patients who presented with more than one ulceration in different locations, one was treated with APP and the other one with OCT. Each group was treated three times a week over a period of two weeks. The antimicrobial efficacy was evaluated immediately after and following two weeks of treatment. RESULTS: Wounds treated with OCT showed a significantly higher microbial reduction (64%) compared to wounds treated with APP (47%) immediately after the treatment. Over two weeks of antiseptic treatment the bacterial density was reduced within the OCT group (-35%) compared to a slight increase in bacterial density in the APP-treated group (+12%). Clinically, there were no signs of delayed wound healing observed in either group and both treatments were well tolerated. CONCLUSION: The immediate antimicrobial effects of the APP prototype source were almost comparable to OCT without any signs of cytotoxicity. This pilot study is limited by current configurations of the plasma source, where the narrow plasma beam made it difficult to cover larger wound surface areas and in order to avoid untreated areas of the wound bed, smaller wounds were assigned to the APP-treatment group. This limits the significance of AAP-related effects on the wound healing dynamics, as smaller wounds tend to heal faster than larger wounds. However, clinical wound healing studies on a larger scale now seem justifiable. A more advanced plasma source prototype allowing the treatment of larger wounds will address APP's influence on healing dynamics, synergetic treatment with current antiseptics and effects on multiresistant bacteria.

Ultraviolet radiation and nanoparticle induced intracellular free radicals generation measured in human keratinocytes by electron paramagnetic resonance spectroscopy.

BACKGROUND: Several nanoparticle-based formulations used in cosmetics and dermatology are exposed to sunlight once applied to the skin. Therefore, it is important to study possible synergistic effects of nanoparticles and ultraviolet radiation. METHODS: Electron paramagnetic resonance spectroscopy (EPR) was used to detect intracellular free radicals induced by ultraviolet B (UVB) radiation and amorphous silica nanoparticle and to evaluate the influence of nanoparticle surface chemistry on particle cytotoxicity toward HaCaT cells. Uncoated titanium dioxide nanoparticles served as positive control. In addition, particle intracellular uptake, viability, and induction of interleukin-6 were measured. RESULTS: We found that photo-activated titanium dioxide particles induced a significant amount of intracellular free radicals. On the contrary, no intracellular free radicals were generated by the investigated silica nanoparticles in the dark as well as under UVB radiation. However, under UVB exposure, the non-functionalized silica nanoparticles altered the release of IL-6. At the same concentrations, the amino-functionalized silica nanoparticles had no influence on UVB-induced IL-6 release. CONCLUSION: EPR spectroscopy is a useful technique to measure nanoparticle-induced intracellular free radicals. Non-toxic concentrations of silica particles enhanced the toxicity of UVB radiation. This synergistic effect was not mediated by particle-generated free radicals and correlated with particle surface charge and intracellular distribution.

Gain or loss? Sunscreen efficiency after cosmetic pretreatment of the skin.

Sunscreens are a key pillar of the multimodal protection strategy against short- and long-term impacts of intermittent and continuous UV exposure. Hitherto, an unanswered part of current scientific discourse is the question whether a cosmetic pretreatment has an impact on distribution and adhesiveness of sunscreens on the skin and therefore affects UV protection. In order to evaluate the homogeneity of sunscreen filter distribution, water resistance as a parameter of adhesiveness and effective UV protection of sunscreens after a pretreatment with cream or lotion was investigated in 18 volunteers who were examined before and after swimming, using the established combination of the tape stripping procedure and UV/VIS spectroscopy. It was shown that a cosmetic skin pretreatment affects neither filter homogeneity nor effective UV protection prior to water contact. However, compared to nonpretreated skin, a considerable loss of water resistance is caused. Therefore, using a cream or lotion before application of sunscreens is not to be recommended.