RESUMO
This work reveals a versatile new method to produce films with antimicrobial properties that can also bond materials together with robust tensile adhesive strength. Specifically, we demonstrate the formation of coatings by using a dielectric barrier discharge (DBD) plasma to convert a liquid small-molecule precursor, m-cresol, to a solid film via plasma-assisted on-surface polymerisation. The films are quite appealing from a sustainability perspective: they are produced using a low-energy process and from a molecule produced in abundance as a by-product of coal tar processing. This process consumes only 1.5 Wh of electricity to create a 1 cm2 film, which is much lower than other methods commonly used for film deposition, such as chemical vapour deposition (CVD). Plasma treatments were performed in plain air without the need for any carrier or precursor gas, with a variety of exposure durations. By varying the plasma parameters, it is possible to modify both the adhesive property of the film, which is at a maximum at a 1 min plasma exposure, and the antimicrobial property of the film against Escherichia coli, which is at a maximum at a 30 s exposure.
Assuntos
Adesivos , Anti-Infecciosos , Antibacterianos/química , Antibacterianos/farmacologia , Anti-Infecciosos/farmacologia , Cresóis , Escherichia coliRESUMO
Hexaminolevulinate (HAL) induced Protoporphyrin IX (PpIX) fluorescence is commonly used to differentiate cancer cells from normal cells in vivo, as for instance in blue light cystoscopy for bladder cancer diagnosis. A detailed approach is here provided to use this diagnostic principle ex vivo in an immunosensor device, towards enabling non-invasive cancer diagnostic from body fluids, such as urine. Several factors susceptible to affect the applicability of HAL-assisted diagnosis in body fluids were tested. These included the cell viability and its impact on PpIX fluorescence, the storage condition and shelf life of HAL premix reagent, light exposure (360-450 nm wavelengths) and its corresponding effect on both intensity and bleaching of the PpIX fluorescence as a function of the microscopy imaging conditions. There was no significant decrease in the viability of bladder cancer cells after 6 h at 4 °C (student's t-test: p > 0.05). The cellular PpIX fluorescence decreased in a time-dependent manner when cancer cells were kept at 4 °C for extended period of time, though this didn't significantly reduce the fluorescence intensity contrast between cancer and non-cancer cells kept in the same condition for 6 h. HAL premix reagent kept in long term storage at 4 °C induced stronger PpIX fluorescence than reagent kept in the - 20 °C freezer. The PpIX fluorescence was negatively affected by repeated light exposure but increased with illumination intensity and exposure time. Though this applied to both healthy and cancer cell lines, and therefore did not statistically improved the differentiation between cell types. This study revealed important experimental settings that need to be carefully considered to benefit from the analytical potential of HAL induced fluorescence when used in technologies for the diagnosis of cancer from body fluids.
Assuntos
Ácido Aminolevulínico/análogos & derivados , Técnicas Biossensoriais/métodos , Testes Imunológicos/métodos , Fármacos Fotossensibilizantes/química , Neoplasias da Bexiga Urinária/patologia , Ácido Aminolevulínico/química , Técnicas Biossensoriais/normas , Linhagem Celular Tumoral , Células Cultivadas , Humanos , Testes Imunológicos/normas , Biópsia Líquida/métodos , Biópsia Líquida/normas , Microfluídica/métodos , Microfluídica/normas , Protoporfirinas/metabolismo , Sensibilidade e Especificidade , Neoplasias da Bexiga Urinária/urina , Urotélio/metabolismo , Urotélio/patologiaRESUMO
Chronic wounds fail to heal and are accompanied by an ongoing infection. They cause suffering, shorten lifespans, and their prevalence is increasing. Unfortunately, the medical treatment of chronic wounds has remained unchanged for decades. A novel approach to break the biological vicious cycle is the long-lived radical (2,2,6,6-Tetramethylpiperidin-1-yl)oxyl (TEMPO). TEMPO can be plasma polymerised (TEMPOpp) into thin coatings that have antimicrobial properties. However, due to its radical nature, quenching causes it to lose effectiveness over time. Our aim in this study was to extend the shelf-life of TEMPOpp coatings using various storage conditions: Namely, room temperature (RT), room temperature & vacuum sealed (RTV), freezer temperature & vacuum sealed (FTV). We have analysed the coatings' quality via the surface analytical methods of X-Ray Photoelectron spectroscopy (XPS) and electron paramagnetic resonance (EPR); finding marked differences among the three storage conditions. Furthermore, we have compared the antimicrobial efficacy of the stored coatings against two major bacterial pathogens, Staphylococcus aureus and Staphylococcus epidermidis, commonly found in chronic wounds. We did so both qualitatively via live/dead staining, as well as quantitatively via (2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino) carbonyl]-2H-tetrazolium (XTT) viability assay for up to 15 weeks in 5 weeks increments. Taken all together, we demonstrate that samples stored under FTV conditions retain the highest antimicrobial activity after 15 weeks and that this finding correlates with the retained concentration of nitroxides.
RESUMO
The stable nitroxide radical TEMPO [(2,2,6,6-Tetramethylpiperidin-1-yl)oxyl] has a multitude of applications in fields ranging from energy storage to biomedical applications and many more. However, to date, the processes of incorporating nitroxide radicals into thin-film coatings are laborious and not cost-effective, which hinders their wider use in many applications. In contrast, the authors have recently demonstrated the facile method of plasma polymerization of TEMPO into thin-film coatings that retain the stable nitroxide radicals. In this work, we are using three types of mass spectroscopic methods (plasma-mass spectrometry, time of flight secondary ion mass spectrometry, and high-performance liquid chromatography-mass spectrometry) and electron spin resonance to track the fate of the TEMPO molecule from monomer flask through the plasma and inside the resulting coatings. The results of this study demonstrate that TEMPO is a versatile monomer that can be used across different plasma reactors and reliably retain the stable nitroxide radical in the resulting thin-film coatings if certain process conditions are observed, namely, higher process pressures and lower powers.
Assuntos
Óxidos N-Cíclicos/química , Óxidos de Nitrogênio/química , Gases em Plasma/química , Polimerização , Espectroscopia de Ressonância de Spin Eletrônica , Íons , Análise de Componente PrincipalRESUMO
Chronic wounds are affecting increasingly larger portions of the general population and their treatment has essentially remained unchanged for the past century. This lack of progress is due to the complex problem that chronic wounds are simultaneously infected and inflamed. Both aspects need to be addressed together to achieve a better healing outcome. Hence, we hereby demonstrate that the stable nitroxide radical (2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO) can be plasma polymerized into smooth coatings (TEMPOpp), as seen via atomic force microscopy, X-ray photoelectron spectroscopy and ellipsometry. Upon contact with water, these coatings leach nitroxides into aqueous supernatant, as measured via EPR. We then exploited the known cell-signalling qualities of TEMPO to change the cellular behaviour of bacteria and human cells that come into contact with the surfaces. Specifically, the TEMPOpp coatings not only suppressed biofilm formation of the opportunistic bacterium Staphylococcus epidermidis but also dispersed already formed biofilm in a dose-dependent manner; a crucial aspect in treating chronic wounds that contain bacterial biofilm. Thus the coatings' microbiological efficacy correlated with their thickness and the thickest coating was the most efficient. Furthermore, this dose-dependent effect was mirrored in significant cytokine reduction of activated THP-1 macrophages for the four cytokines TNF-α, IL-1ß, IL-6 and IP-10. At the same time, the THP-1 cells retained their ability to adhere and colonize the surfaces, as verified via SEM imaging. Thus, summarily, we have exploited the unique qualities of plasma polymerized TEMPO coatings in targeting both infection and inflammation simultaneously; demonstrating a novel alternative to how chronic wounds could be treated in the future.