Lessons learned in a decade: Medical-toxicological view of tattooing.

Tattooing has been part of the human culture for thousands of years, yet only in the past decades has it entered the mainstream of the society. With the rise in popularity, tattoos also gained attention among researchers, with the aim to better understand the health risks posed by their application. 'A medical-toxicological view of tattooing'-a work published in The Lancet almost a decade ago, resulted from the international collaboration of various experts in the field. Since then, much understanding has been achieved regarding adverse effects, treatment of complications, as well as their regulation for improving public health. Yet major knowledge gaps remain. This review article results from the Second International Conference on Tattoo Safety hosted by the German Federal Institute for Risk Assessment (BfR) and provides a glimpse from the medical-toxicological perspective, regulatory strategies and advances in the analysis of tattoo inks.

Photodynamic inactivation of different pathogenic bacteria on human skin using a novel photosensitizer hydrogel.

BACKGROUND: The colonization of skin with pathogenic, partially antibiotic-resistant bacteria is frequently a severe problem in dermatological therapies. For instance, skin colonization with Staphylococcus aureus is even a disease-promoting factor in atopic dermatitis. The photodynamic inactivation (PDI) of bacteria could be a new antibacterial procedure. Upon irradiation with visible light, a special photosensitizer exclusively generates singlet oxygen. This reactive oxygen species kills bacteria via oxidation independent of species or strain and their antibiotic resistance profile causing no bacterial resistance on its part. OBJECTIVE: To investigate the antibacterial potential of a photosensitizer, formulated in a new hydrogel, on human skin ex vivo. METHODS: The photochemical stability of the photosensitizer and its ability to generate singlet oxygen in the hydrogel was studied. Antimicrobial efficacy of this hydrogel was tested step by step, firstly on inanimate surfaces and then on human skin ex vivo against S. aureus and Pseudomonas aeruginosa using standard colony counting. NBTC staining and TUNEL assays were performed on skin biopsies to investigate potential necrosis and apoptosis effects in skin cells possibly caused by PDI. RESULTS: None of the hydrogel components affected the photochemical stability and the life time of singlet oxygen. On inanimate surfaces as well as on the human skin, the number of viable bacteria was reduced by up to 4.8 log10 being more effective than most other antibacterial topical agents. Histology and assays showed that PDI against bacteria on the skin surface caused no harmful effects on the underlying skin cells. CONCLUSION: Photodynamic inactivation hydrogel proved to be effective for decolonization of human skin including the potential to act against superficial skin infections. Being a water-based formulation, the hydrogel should be also suitable for the mucosa. The results of the present ex vivo study form a good basis for conducting clinical studies in vivo.

Photodynamic inactivation of Salmonella enterica and Listeria monocytogenes inoculated onto stainless steel or polyurethane surfaces.

The photodynamic inactivation (PDI) uses molecules (photosensitizers) that absorb visible light (385-450 nm) energy, transfer it to adjacent molecular oxygen and thereby generating the biocidal singlet oxygen and other reactive oxygen species in situ. Efficacy of PDI was tested against Listeria monocytogenes and Salmonella enterica in three ways. Firstly, by adding the photosensitizer to bacterial suspensions. Secondly, bacteria were placed on inanimate surfaces and then sprayed with a photosensitizer suspension. Thirdly, bacteria were placed on coated inanimate surfaces, where the photosensitizer was permanently fixed in this coating (antimicrobial coating, AMC). Experiments were performed without and with soiling (albumin, sheep erythrocytes). In suspension, PDI reduced the number of viable Listeria monocytogenes and Salmonella enterica by more than 6 Log CFU/mL within seconds of light exposure. Photosensitizer spray suspension reduced the bacterial burden on surfaces with up to about 6 Log CFU/mL (5 s light exposure). PDI, even in the presence of high soiling, achieved a reduction of up to 5.1 ± 1.2 Log CFU/mL. The AMC showed a bacterial reduction that decreased from 5.1 to 0.7 Log CFU/mL with increasing soiling. Depending on the soiling and the respective bacteria, the spray suspension or AMC achieved a bacterial reduction on the running conveyor belt demonstrator ranging from 2.9 to 5.3 or 0.5 to 4.5 Log CFU/mL, respectively. PDI used visible light, phenalene-1-one and curcumin photosensitizers, and oxygen from ambient air to reduce the bioburden on typical surfaces in food processing. The AMC acts slower than the spray suspension but enables a permanent, self-sanitizing effect.

Phototherapy: Theory and practice.

Despite the development of highly effective biologics for skin diseases such as psoriasis or atopic dermatitis, UVA and UVB therapy, alone or in combination, are still essential components of various guidelines. Phototherapy is not only a first-line treatment and highly effective for a number of skin diseases, but is also economical and has few side effects. The targeted use of UVA and UVB, if necessary, in combination with the photosensitizer psoralen in the context of PUVA therapy, enables the dermatologist to effectively treat a wide variety of skin diseases. Indications for phototherapy include epidermal diseases such as atopic dermatitis, psoriasis and vitiligo, as well as photodermatoses, mycosis fungoides, graft-versus-host disease and deep dermal diseases such as scleroderma. This article reviews the physical principles, molecular mechanisms, current treatment regimens, and individual indications for phototherapy and photochemotherapy.

Antimicrobial coatings for environmental surfaces in hospitals: a potential new pillar for prevention strategies in hygiene.

Recent reports provide evidence that contaminated healthcare environments represent major sources for the acquisition and transmission of pathogens. Antimicrobial coatings (AMC) may permanently and autonomously reduce the contamination of such environmental surfaces complementing standard hygiene procedures. This review provides an overview of the current status of AMC and the demands to enable a rational application of AMC in health care settings. Firstly, a suitable laboratory test norm is required that adequately quantifies the efficacy of AMC. In particular, the frequently used wet testing (e.g. ISO 22196) must be replaced by testing under realistic, dry surface conditions. Secondly, field studies should be mandatory to provide evidence for antimicrobial efficacy under real-life conditions. The antimicrobial efficacy should be correlated to the rate of nosocomial transmission at least. Thirdly, the respective AMC technology should not add additional bacterial resistance development induced by the biocidal agents and co- or cross-resistance with antibiotic substances. Lastly, the biocidal substances used in AMC should be safe for humans and the environment. These measures should help to achieve a broader acceptance for AMC in healthcare settings and beyond. Technologies like the photodynamic approach already fulfil most of these AMC requirements.

Tattoos - more than just colored skin? Searching for tattoo allergens.

During tattooing, a high amount of ink is injected into the skin. Tattoo inks contain numerous substances such as the coloring pigments, impurities, solvents, emulsifiers, and preservatives. Black amorphous carbon particles (carbon black), white titanium dioxide, azo or polycyclic pigments create all varieties of color shades in the visible spectrum. Some ingredients of tattoo inks might be hazardous and allergenic chemicals of unknown potential. In Germany, about 20 % of the general population is tattooed and related adverse reactions are increasingly reported. Since tattoo needles inevitably harm the skin, microorganisms can enter the wound and may cause infections. Non-allergic inflammatory reactions (for example cutaneous granuloma and pseudolymphoma) as well as allergic reactions may emerge during or after wound healing. Especially with allergies occurring after weeks, months or years, it remains difficult to identify the specific ingredient(s) that trigger the reaction. This review summarizes possible adverse effects related to tattooing with a focus on the development of tattoo-mediated allergies. To date, relevant allergens were only identified in rare cases. Here we present established methods and discuss current experimental approaches to identify culprit allergens in tattoo inks - via testing of the patient and in vitro approaches.

Biological effects of a new ultraviolet A1 prototype based on light-emitting diodes on the treatment of localized scleroderma.

Ultraviolet A1 (UVA1 ) phototherapy (spectral range 340-400 nm) is a well-established treatment option for various skin diseases such as localized scleroderma. Recent improvements of conventional UVA1 light sources (metal-halide or fluorescent lamps) have brought attention to a new light-emitting diode (LED) technology with remarkable advantages in handling and clinical routine. This study provides a preclinical histological and molecular evaluation of an LED-based UVA1 prototype with a narrower spectral range (360-400 nm) for treating localized scleroderma. Scleroderma mouse models and fibroblasts in vitro were exposed to LED-based UVA1 phototherapy or to irradiation with a commercially available metal-halide lamp emitting low-dose (20, 40 J/cm2 ), medium-dose (60 J/cm2 ) and high-dose (80, 100 J/cm2 ) UVA1 light. Both UVA1 light sources affected inflammatory genes (IL-1α and IL-6) and growth factors (TGFß-1 and TGFß-2). Increased collagen type 1 was reduced after UVA1 phototherapy. Matrix metalloproteinase-1 was more enhanced after a medium dose of LED-based UVA1 phototherapy than after conventional treatment. In vivo, dermal thickness and the amount of collagen were reduced after both treatment methods. Remarkably, myofibroblasts were more effectively reduced by a medium dose of LED-based UVA1 phototherapy. The study indicates that LED-based UVA1 phototherapy yields similar or even better results than conventional treatment. In terms of biosafety and patient comfort, LED-based UVA1 phototherapy offers clear advantages over conventional treatment because of the use of a narrower and less harmful UVA1 spectrum, less heat generation and shorter treatment times at the same irradiation intensity. Clinical studies are required to confirm these results in patients with localized scleroderma.

Distribution of nickel and chromium containing particles from tattoo needle wear in humans and its possible impact on allergic reactions.

BACKGROUND: Allergic reactions to tattoos are amongst the most common side effects occurring with this permanent deposition of pigments into the dermal skin layer. The characterization of such pigments and their distribution has been investigated in recent decades. The health impact of tattoo equipment on the extensive number of people with inked skin has been the focus of neither research nor medical diagnostics. Although tattoo needles contain high amounts of sensitizing elements like nickel (Ni) and chromium (Cr), their influence on metal deposition in skin has never been investigated. RESULTS: Here, we report the deposition of nano- and micrometer sized tattoo needle wear particles in human skin that translocate to lymph nodes. Usually tattoo needles contain nickel (6-8%) and chromium (15-20%) both of which prompt a high rate of sensitization in the general population. As verified in pig skin, wear significantly increased upon tattooing with the suspected abrasive titanium dioxide white when compared to carbon black pigment. Additionally, scanning electron microscopy of the tattoo needle revealed a high wear after tattooing with ink containing titanium dioxide. The investigation of a skin biopsy obtained from a nickel sensitized patient with type IV allergy toward a tattoo showed both wear particles and iron pigments contaminated with nickel. CONCLUSION: Previously, the virtually inevitable nickel contamination of iron pigments was suspected to be responsible for nickel-driven tattoo allergies. The evidence from our study clearly points to an additional entry of nickel to both skin and lymph nodes originating from tattoo needle wear with an as yet to be assessed impact on tattoo allergy formation and systemic sensitization.

A medical-toxicological view of tattooing.

Long perceived as a form of exotic self-expression in some social fringe groups, tattoos have left their maverick image behind and become mainstream, particularly for young people. Historically, tattoo-related health and safety regulations have focused on rules of hygiene and prevention of infections. Meanwhile, the increasing popularity of tattooing has led to the development of many new colours, allowing tattoos to be more spectacular than ever before. However, little is known about the toxicological risks of the ingredients used. For risk assessment, safe intradermal application of these pigments needs data for toxicity and biokinetics and increased knowledge about the removal of tattoos. Other concerns are the potential for phototoxicity, substance migration, and the possible metabolic conversion of tattoo ink ingredients into toxic substances. Similar considerations apply to cleavage products that are formed during laser-assisted tattoo removal. In this Review, we summarise the issues of concern, putting them into context, and provide perspectives for the assessment of the acute and chronic health effects associated with tattooing.

A novel set of symmetric methylene blue derivatives exhibits effective bacteria photokilling - a structure-response study.

This study focuses on the structure-response relationship of symmetrically substituted phenothiazinium dyes. Four hydrophilic derivatives with the ability of additional hydrogen bonding (, ) or additional electrostatic interaction (, ) were synthesized, photophysically characterized and compared to the parent compound methylene blue (MB, ) and a lipophilic derivative () without additional coordination sites. Derivative was most effective against Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli reaching a maximum photodynamic efficacy of >5log10 steps (≥99.999%) of bacteria killing in 10 minutes (5 µM, 30 J cm(-2)) without inherent dark toxicity after one single treatment with the incoherent light source PDT1200 (λmax = 660 nm, 50 mW cm(-2)). Interestingly, one derivative with two additional primary positive charges () showed selective killing of Escherichia coli (5 µM, 30 J cm(-2), 4log10 steps inactivation (≥99.99%)) and no antimicrobial effect on Staphylococcus aureus. This might allow the development of a new generation of photosensitizers with higher antimicrobial efficacy and selectivity for future applications.

Fast and effective inactivation of Bacillus atrophaeus endospores using light-activated derivatives of vitamin B2.

Highly resistant endospores may cause severe problems in medicine as well as in the food and packaging industries. We found that bacterial endospores can be inactivated quickly with reactive oxygen species (ROS) that were generated by a new generation of flavin photosensitizers. Flavins like the natural compound vitamin B2 are already known to produce ROS but they show a poor antimicrobial photodynamic killing efficacy due to the lack of positive charges. Therefore we synthesized new flavin photosensitizers that have one (FLASH-01a) or eight (FLASH-07a) positive charges and can hence attach to the negatively charged surface of endospores. In this study we used standardized Bacillus atrophaeus endospores (ATCC 9372) as a biological surrogate model for a proof-of-concept study of photodynamic inactivation experiments using FLASH-01a and FLASH-07a. After incubation of spores with different flavin concentrations, the flavin derivatives were excited with blue light at a light dose of 70 J cm(-2). The inactivation of spores was investigated either in suspension or after attachment to polyethylene terephthalate (PET) surfaces. Incubation of spores suspended in Millipore water with 4 mM FLASH-01a for 10 seconds and irradiation with blue light for 10 seconds caused a biologically relevant decrease of spore survival of 3.5 log10 orders. Using FLASH-07a under the same conditions we achieved a decrease of 4.4 log10 orders. Immobilized spores on PET surfaces were efficiently killed with 7.0 log10 orders using 8 mM FLASH-07a. The total treatment time (incubation + irradiation) was as short as 20 seconds. The results of this study show evidence that endospores can be fastly and effectively inactivated with new generations of flavin photosensitizers that may be useful for industrial or medical applications in the future.

Exposure of vitamins to UVB and UVA radiation generates singlet oxygen.

Deleterious effects of UV radiation in tissue are usually attributed to different mechanisms. Absorption of UVB radiation in cell constituents like DNA causes photochemical reactions. Absorption of UVA radiation in endogenous photosensitizers like vitamins generates singlet oxygen via photosensitized reactions. We investigated two further mechanisms that might be involved in UV mediated cell tissue damage. Firstly, UVB radiation and vitamins also generate singlet oxygen. Secondly, UVB radiation may change the chemical structure of vitamins that may change the role of such endogenous photosensitizers in UVA mediated mechanisms. Vitamins were irradiated in solution using monochromatic UVB (308 nm) or UVA (330, 355, or 370 nm) radiation. Singlet oxygen was directly detected and quantified by its luminescence at 1270 nm. All investigated molecules generated singlet oxygen with a quantum yield ranging from 0.007 (vitamin D3) to 0.64 (nicotinamide) independent of the excitation wavelength. Moreover, pre-irradiation of vitamins with UVB changed their absorption in the UVB and UVA spectral range. Subsequently, molecules such as vitamin E and vitamin K1, which normally exhibit no singlet oxygen generation in the UVA, now produce singlet oxygen when exposed to UVA at 355 nm. This interplay of different UV sources is inevitable when applying serial or parallel irradiation with UVA and UVB in experiments in vitro. These results should be of particular importance for parallel irradiation with UVA and UVB in vivo, e.g. when exposing the skin to solar radiation.

Singlet oxygen generation in porphyrin-doped polymeric surface coating enables antimicrobial effects on Staphylococcus aureus.

Surfaces can be coated with photosensitizer molecules, which generate singlet oxygen ((1)O2) when the surface is exposed to light. (1)O2 may diffuse from the coating and has the potential to kill microorganisms present on the surface. In the present study a derivative of the meso-tetraphenylporphyrin (TPP) was immobilized onto polyurethane (PU) after being sprayed and polymerized as a thin layer onto poly-methylmethacrylate (PMMA). PU is gas permeable and thus a sufficient amount of oxygen reaches the photosensitizer in this coating. The surface generation of (1)O2 and its diffusion were investigated by detecting its luminescence at 1270 nm and a tri-iodide assay. Antimicrobial photodynamic surface effects were tested on Staphylococcus aureus. The spectrally resolved detection of (1)O2 luminescence yielded a clear peak at 1275 nm. The time-resolved luminescence showed multi-exponential decay, revealing rise and decay times in the range of 5-2 × 10(2)µs. The photodynamic inactivation of S. aureus was monitored at different photosensitizer concentrations and radiant exposures of light. A photodynamic killing of >99.9% (>3log10-steps) was achieved within an irradiation time of 30 min. The photodynamic killing on the bioactive surface confirmed the antimicrobial effect of (1)O2 that was generated in the PU-coating and reached the bacteria by diffusion.

An Internet-based survey on characteristics of laser tattoo removal and associated side effects.

Tattoo removal by laser therapy is a frequently performed procedure in dermatological practices. Quality-switched ruby, alexandrite, or Nd:YAG lasers are the most suitable treatment devices. Although these techniques are regarded as safe, both temporary and permanent side effects might occur. Little has been published on the frequency of complications associated with laser tattoo removal. We performed an Internet survey in German-speaking countries on characteristics of laser tattoo removal and associated side effects. A total number of 157 questionnaires entered the final analysis. Motivations for laser tattoo removal were mainly considering the tattoo as youthful folly (29%), esthetic reasons (28%), and 6% indicated medical problems. One third of participants were unsatisfied with the result of laser tattoo removal, and a complete removal of the tattoo pigment was obtained in 38% only. Local transient side effects occurred in nearly all participants, but an important rate of slightly visible scars (24%) or even important scarring (8%) was reported. Every fourth participant described mild or intense tan when the laser treatment was performed, and the same number of people indicated UV exposure following laser therapy, which should normally be avoided in these circumstances. As reported in the literature, nearly half of the participants experienced hypopigmentation in the treated area. Our results show that from the patients' point of view there is an important rate of side effects occurring after laser tattoo removal. Appropriate pretreatment counseling with regard to realistic expectations, possible side effects, and the application of test spots is mandatory to ensure patient satisfaction. Laser treatment should be performed by appropriately trained personnel only.

Dirty hands: photodynamic killing of human pathogens like EHEC, MRSA and Candida within seconds.

Hand hygiene is one of the most important interventions for reducing transmission of nosocomial life-threatening microorganisms, like methicillin resistant Staphylococcus aureus (MRSA), enterohemorrhagic Escherichia coli (EHEC) or Candida albicans. All three pathogens have become a leading cause of infections in hospitals. Especially EHEC is causing severe diarrhoea and, in a small percentage of cases, haemolytic-uremic syndrome (HUS) as reported for E. coli 104:H4 in Germany 2011. We revealed the possibility to inactivate very fast and efficiently MRSA, EHEC and C. albicans using the photodynamic approach. MRSA, EHEC and C. albicans were incubated in vitro with different concentrations of TMPyP for 10 s and illuminated with visible light (50 mW cm(-2)) for 10 and 60 s. 1 µmol l(-1) of TMPyP and an applied radiant exposure of 0.5 J cm(-2) achieved a photodynamic killing of ≥99.9% of MRSA and EHEC. Incubation with higher concentrations (up to 100 µmol l(-1)) of TMPyP caused bacteria killing of >5 log(10) (≥99.999%) after illumination. Efficient Candida killing (≥99.999%) was achieved first at a higher light dose of 12 J cm(-2). Different rise and decay times of singlet oxygen luminescence signals could be detected in Candida cell suspensions for the first time, indicating different oxygen concentrations in the surrounding for the photosensitizer and singlet oxygen, respectively. This confirms that TMPyP is not only found in the water-dominated cell surrounding, but also within the C. albicans cells. Applying a water-ethanol solution of TMPyP on ex vivo porcine skin, fluorescence microscopy of histology showed that the photosensitizer was exclusively localized in the stratum corneum regardless of the incubation time. TMPyP exhibited a fast and very effective killing rate of life-threatening pathogens within a couple of seconds that encourages further testing in an in vivo setting. Being fast and effective, antimicrobial photodynamic applications might become acceptable as a tool for hand hygiene procedures and also in other skin areas.

Luminescence spectroscopy of singlet oxygen enables monitoring of oxygen consumption in biological systems consisting of fatty acids.

The interaction of singlet oxygen ((1)O2) generated in a photosensitized process with well-known reference photosensitizers Perinaphthenone (PN) and TMPyP is investigated in a model system consisting of fatty acids and the respective exogenous photosensitizer (PS) in solution by direct detection of the luminescence photons of (1)O2 at 1270 nm. Such a model system is a first approach to mimic the complex environment of (1)O2 in a biological cell which consists mainly of water, proteins, sugars and lipids. Firstly, the important issue of oxygen consumption is evaluated which has to be considered during luminescence detection of (1)O2. It is known that the luminescence signal of (1)O2 is dependent on the oxygen concentration of the environment. Cellular components such as lipids represent oxygen consumers due to peroxidation of their unsaturated double bonds. Secondly, the experimental conditions for this model system regarding oxygen consumption are optimized to estimate the rates and rate constants of the coupled system. Thirdly, the triplet decay of the PS can provide more precise information about the actual oxygen concentration close to the PS and can be used, therefore, as a more precise method to determine the oxygen concentration in more complex systems such as a biological cell. The aim is to get a better understanding of photosensitized reactions of (1)O2 with cellular components to further improve methodologies, in particular at a cellular level using luminescence spectroscopy. In conclusion, luminescence detection might be a helpful tool to monitor precisely and promptly changes in oxygen concentration in a complex environment.

Singlet oxygen-induced photodegradation of the polymers and dyes in optical sensing materials and the effect of stabilizers on these processes.

A comprehensive study of photodegradation processes in optical sensing materials caused by photosensitized singlet oxygen in different polymers is presented. The stabilities of the polymers are accessed in the oxygen consumption measurements performed with help of optical oxygen sensors. Polystyrene and poly(phenylsilesquioxane) are found to be the most stable among the polymers investigated, whereas poly(2,6-dimethyl-p-phenylene oxide) and particularly poly(methyl methacrylate) and their derivatives show the fastest oxygen consumption. The effect of the stabilizers (singlet oxygen quenchers) on the oxygen consumption rates, the photostability of the sensitizer, and the total photon emission (TPE) by singlet oxygen is studied. 1,4-Diazabicyclo[2.2.2]octane (DABCO) was found to significantly reduce both the TPE and the oxygen consumption rates, indicating its role as a physical quencher of singlet oxygen. The addition of DABCO also significantly improved the photostability of the sensitizer. The N-alkylated derivative of DABCO and DABCO covalently grafted to the polystyrene backbone are prepared in an attempt to overcome the volatility and water solubility of the quencher. These derivatives as well as other tertiary amines investigated were found to be inefficient as stabilizing agents, and some of them even negatively affected the oxygen consumption rates.

Adverse reactions after tattooing: review of the literature and comparison to results of a survey.

The number of tattooed people has substantially increased in the past years. Surveys in different countries reveal this to be up to 24% of the population. The number of reported adverse reactions after tattooing has also increased including infections, granulomatous and allergic reactions and tumors. However, the case reports do not reflect the frequency of adverse reactions. This review compares the medically documented adverse reactions published in 1991-2011 with the findings of a nation-wide survey that recently revealed the features and health problems associated with tattoos. To compare the data with the survey, the sex of patients was reported and the location and color of tattoos were evaluated. The results show clearly that colored tattoo inks are mainly responsible for adverse skin reactions and that tattoos on the extremities are involved most.