Matrix Metalloproteinases in Asthma-Associated Airway Remodeling – Dr. Jekyll or Mr. Hyde ?

Matrix metalloproteinases (MMPs) are Zn2+-dependent endoproteases, which digest extracellular matrix (ECM) components and various non-ECM molecules. Main physiological role of MMPs concerns regulation of tissue remodeling and regeneration. The production and activity of MMPs are tightly supervised by multistage control mechanisms. These mechanisms include regulation of gene expression, and various post-transcriptional/post-translational modifications. However, without proper control MMPs reveal dual nature, similarly to character from the novella by R.L. Stevenson, “Strange Case of Dr Jekyll and Mr Hyde”. They become dangerous molecules, involved in cancer metastasis, or cardiovascular diseases

Label-free smartphone quantitation of bacteria by darkfield imaging of light scattering in fluoropolymer micro capillary film allows portable detection of bacteriophage lysis

Conventional methods for the detection and quantitation of bacteria are slow, laborious and require a laboratory. Microfluidic systems offer faster and portable testing, and smartphone cameras can record colorimetric or fluorometric bioassays, but this requires dye addition. Here, we demonstrate for the first time label-free smartphone detection of bacterial light scattering by darkfield microfluidic imaging to measure bacteria and bacteriophage lysis. A single LED and portable 3D printed imaging box allowed bacterial concentration and growth to be measured by direct imaging of bacterial light scattering. Bacteriophage lysis was detected within a 10-channel microfluidic device made from melt-extruded fluoropolymer micro capillary film, allowing rapid detection of host specificity. Elimination of unwanted reflections and optimising illumination angle are critical for successful darkfield bacterial imaging, with 15° giving maximal intensity. Bacterial sedimentation was directly observed within microfluidic devices, and detection sensitivity significantly increased by allowing bacteria to sediment for 30 min. With this simple, low-cost, 3D printed system bacterial concentrations down to an optical density of 0.1 could be measured corresponding to 8 × 104 colony forming units (CFU) per microdevice, approaching the sensitivity of conventional spectrophotometers. Bacteriophage lysis could be detected at a range of starting cell concentrations. With a low starting cell concentration, the increase in light scatter signal with incubation was prevented in the presence of bacteriophage. Conversely, with high starting cell concentration, the light scatter signal detected at the start was clearly eliminated when phage were added, indicating this simple system allows direct visualisation of bacteriophage eliminating light scattering by lysis

Mikrosolenoidowe przepływowe systemy bioanalityczne do oznaczania wybranych analitów istotnych w diagnostyce medycznej

Diagnostyka medyczna jest procesem rozpoznania chorób na podstawie stwierdzonych objawów przy pomocy wielu rodzajów badań. Jednym z obszarów diagnostyki medycznej jest analiza kliniczna. Wynika to ze znaczenia i wkładu wyników otrzymanych podczas badań laboratoryjnych w toku identyfikacji zespołów chorobowych. Znaczna większość podejmowanych decyzji medycznych jest oparta właśnie o jakościowe lub/i ilościowe dane analityczne. Od technik pomiarowych wprowadzanych na potrzeby analityki klinicznej wymagany jest szereg parametrów, które przyczynią się do coraz efektywniejszego oznaczania wielu, różnorodnych analitów w złożonych matrycach materiałów biologicznych. Dlatego też wysoka precyzja, powtarzalność i odtwarzalność oraz zmniejszenie kosztów i czasu przeprowadzanych analiz to główne parametry charakteryzujące nowoczesne układy pomiarowe. W dużym stopniu zostało to osiągnięte poprzez wprowadzenie zmechanizowania lub zautomatyzowania procedur analitycznych i zaimplementowania ich w systemach klinicznych. Jednym z trzech głównych typów analizatorów dostępnych w laboratoriach klinicznych, poza analizatorami wirówkowymi i dyskretnymi, są analizatory przepływowe. Koncepcja analiza przepływowej powstała w późnych latach pięćdziesiątych jako odpowiedź na problemy analizy klinicznej (do których głównie należy zaliczyć niewielką ilość przeprowadzanych oznaczeń oraz niską precyzję pomiarów) i od samego początku była prężnie rozwijana. Dowodem tego są powstałe na jej podstawie liczne techniki z wykorzystaniem coraz to wydajniejszych urządzeń do sterowania przepływem. Zalicza się do nich między innymi technika z zastosowaniem urządzeń mikrosolenoidowych (określana w literaturze anglojęzycznej jako MCFA – MultiCommutated Flow Analysis). Celem prezentowanych badań była konstrukcja zmechanizowanych układów analitycznych typu MCFA z detekcją optoelektroniczną typu PEDD (Paired Emitter Detector Diode) do oznaczania wybranych biomarkerów w płynach ustrojowych, istotnych z punktu widzenia diagnostyki medycznej, w tym: enzymów (fosfataza kwaśna i zasadowa), metabolitów (kreatynina), białka całkowitego oraz frakcji albuminowej. Dodatkowo, elastyczność i uniwersalność mikrosolenoidowych układów multikomutacyjnych pozwoliła na sprawne i efektywne analizowanie próbek o bardzo złożonej matrycy (mocz, surowica, płyn mózgowo-rdzeniowy). W pracy została przedstawiona budowa, zasada działania oraz charakterystyka opisywanych systemów wraz z potwierdzeniem eksperymentalnym ich przydatności analitycznej do zastosowania w analizie klinicznej.Medical diagnostics is the process of disease recognition based on the symptoms observed with many types of research. One of the areas of medical diagnostics is clinical analysis. The contribution and importance of the results obtained in clinical laboratories are significant in identification process of syndromes. The vast majority of medical decisions are based on the qualitative and / or quantitative analytical data. From analytical techniques, which are introduced for the clinical analysis purpose, a number of parameters is required to efficient determination of many analytes in complex biological matrices. Therefore, high precision, repeatability, reproducibility and reduction of costs and time of the analysis are the main parameters that characterize modern measuring systems. It has been achieved by the introduction of mechanization or automation of analytical procedures and implementation them in clinical systems. One of the three main types of analyzers available in clinical laboratories, besides centrifugal and discrete analyzers are flow analyzers. The concept of flow analysis was introduced in the late fifties as an answer to the problems of clinical analysis (which mainly include a small amount of performed determinations and low precision of measurements). From the very beginning, flow analysis has been rapidly developed. A number of techniques with the use of increasingly efficient flow control devices have been invented on the basis of flow analysis concept. One of them is MultiCommutated Flow Analysis (MCFA) - the technique using solenoid flow control devices. The aim of this study was to construct mechanized analytical MCFA systems with optoelectronic PEDD (Paired Emitter Detector Diode) detection for the determination of selected biomarkers in body fluids, significant from medical diagnostics point of view, including enzymes (acid phosphatase and alkaline), metabolites (creatinine ), total protein and albumin. In addition, the flexibility and versatility solenoid multicommutation systems allowed for efficient and effective sample analysis of very complex matrices (urine, serum, cerebrospinal fluid). In presented dissertation, design, principle of operation and characterization of described systems is presented with experimental confirmation of their analytical usefulness in clinical analysis

Electrochemical Properties of Lipid Membranes Self-Assembled from Bicelles

Supported lipid membranes are widely used platforms which serve as simplified models of cell membranes. Among numerous methods used for preparation of planar lipid films, self-assembly of bicelles appears to be promising strategy. Therefore, in this paper we have examined the mechanism of formation and the electrochemical properties of lipid films deposited onto thioglucose-modified gold electrodes from bicellar mixtures. It was found that adsorption of the bicelles occurs by replacement of interfacial water and it leads to formation of a double bilayer structure on the electrode surface. The resulting lipid assembly contains numerous defects and pinholes which affect the permeability of the membrane for ions and water. Significant improvement in morphology and electrochemical characteristics is achieved upon freeze–thaw treatment of the deposited membrane. The lipid assembly is rearranged to single bilayer configuration with locally occurring patches of the second bilayer, and the number of pinholes is substantially decreased. Electrochemical characterization of the lipid membrane after freeze–thaw treatment demonstrated that its permeability for ions and water is significantly reduced, which was manifested by the relatively high value of the membrane resistance

Multicommutated Flow Analysis System for Determination of Horseradish Peroxidase and Its Inhibitors

A fully mechanized multicommutated flow analysis (MCFA) system dedicated to determining horseradish peroxidase (HRP) activity was developed. Detection was conducted using a flow-through optoelectronic detector-constructed of paired LEDs operating according to the paired emitter-detector diode (PEDD) principle. The PEDD-MCFA system is dedicated to monitoring the enzyme-catalyzed oxidation of p-phenylenediamine (pPD) by a hydrogen peroxide. Under optimized conditions, the presented bioanalytical system was characterized by a linear response range (33.47–200 U/L) with a detection limit at 10.54 U/L HRP activity and 1.66 mV·L/U sensitivity, relatively high throughput (12 signals recordings per hour), and acceptable precision (RSD below 6%). Additionally, the utility of the developed PEDD-MCFA system for the determination of HRP inhibitors allowing the detection of selected thiols at micromolar levels, is demonstrated. The practical utility of the flow system was illustrated by the analysis of some dietary supplements containing L-cysteine, N-acetylcysteine, and L-glutathione

Tobacco Smoke Induces and Alters Immune Responses in the Lung Triggering Inflammation, Allergy, Asthma and Other Lung Diseases: A Mechanistic Review

Many studies have been undertaken to reveal how tobacco smoke skews immune responses contributing to the development of chronic obstructive pulmonary disease (COPD) and other lung diseases. Recently, environmental tobacco smoke (ETS) has been linked with asthma and allergic diseases in children. This review presents the most actual knowledge on exact molecular mechanisms responsible for the skewed inflammatory profile that aggravates inflammation, promotes infections, induces tissue damage, and may promote the development of allergy in individuals exposed to ETS. We demonstrate how the imbalance between oxidants and antioxidants resulting from exposure to tobacco smoke leads to oxidative stress, increased mucosal inflammation, and increased expression of inflammatory cytokines (such as interleukin (IL)-8, IL-6 and tumor necrosis factor α ([TNF]-α). Direct cellular effects of ETS on epithelial cells results in increased permeability, mucus overproduction, impaired mucociliary clearance, increased release of proinflammatory cytokines and chemokines, enhanced recruitment of macrophages and neutrophils and disturbed lymphocyte balance towards Th2. The plethora of presented phenomena fully justifies a restrictive policy aiming at limiting the domestic and public exposure to ETS