Marek Kawecki

In-vivo expression profiling of Pseudomonas aeruginosa infections reveals niche-specific and strain-independent transcriptional programs.

Pseudomonas aeruginosa is a threatening, opportunistic pathogen causing disease in immunocompromised individuals. The hallmark of P. aeruginosa virulence is its multi-factorial and combinatorial nature. It renders such bacteria infectious for many organisms and it is often resistant to antibiotics. To gain insights into the physiology of P. aeruginosa during infection, we assessed the transcriptional programs of three different P. aeruginosa strains directly after isolation from burn wounds of humans. We compared the programs to those of the same strains using two infection models: a plant model, which consisted of the infection of the midrib of lettuce leaves, and a murine tumor model, which was obtained by infection of mice with an induced tumor in the abdomen. All control conditions of P. aeruginosa cells growing in suspension and as a biofilm were added to the analysis. We found that these different P. aeruginosa strains express a pool of distinct genetic traits that are activated under particular infection conditions regardless of their genetic variability. The knowledge herein generated will advance our understanding of P. aeruginosa virulence and provide valuable cues for the definition of prospective targets to develop novel intervention strategies.

Poly[tri(ethylene glycol) ethyl ether methacrylate]-coated surfaces for controlled fibroblasts culturing.

Well-defined thermosensitive poly[tri(ethylene glycol) monoethyl ether methacrylate] (P(TEGMA-EE)) brushes were synthesized on a solid substrate by the surface-initiated atom transfer radical polymerization of TEGMA-EE. The polymerization reaction was initiated by 2-bromo-2-methylpropionate groups immobilized on the surface of the wafers. The changes in the surface composition, morphology, philicity, and thickness that occurred at each step of wafer functionalization confirmed that all surface modification procedures were successful. Both the successful modification of the surface and bonding of the P(TEGMA-EE) layer were confirmed by X-ray photoelectron spectroscopy (XPS) measurements. The thickness of the obtained P(TEGMA-EE) layers increased with increasing polymerization time. The increase of environmental temperature above the cloud point temperature of P(TEGMA-EE) caused the changes of surface philicity. A simultaneous decrease in the polymer layer thickness confirmed the thermosensitive properties of these P(TEGMA-EE) layers. The thermosensitive polymer surfaces obtained were evaluated for the growth and harvesting of human fibroblasts (basic skin cells). At 37 °C, seeded cells adhered to and spread well onto the P(TEGMA-EE)-coated surfaces. A confluent cell sheet was formed within 24 h of cell culture. Lowering the temperature to an optimal value of 17.5 °C (below the cloud point temperature of the polymer, TCP, in cell culture medium) led to the separation of the fibroblast sheet from the polymer layer. These promising results indicate that the surfaces produced may successfully be used as substrate for engineering of skin tissue, especially for delivering cell sheets in the treatment of burns and slow-healing wounds.

Laser in the treatment of hypertrophic burn scars

This prospective study looked at the outcome of laser (light amplification by stimulated emission of radiation) treatment for hypertrophic scarring. Derma™ K laser (a set of combined lasers erbium:yttrium aluminium garnet/carbon dioxide, qualified as a class IV laser) was used. Between 21 June 2000 and 19 November 2002, at the Siemianowice Burn Center, Poland, 592 interventions, using laser, were performed on N= 327 patients (220 women and 107 men, aged between 3 and 80 years). The majority of cases [N= 223 (68·9%)] were patients with post‐burn hypertrophic scars, and 104 cases (31·8%) had various types of hypertrophic scars. Evaluation took place using an adapted Vancouver Scar Scale and digital photographs as well as the patient’s opinion. It was noted that after laser treatment, satisfactory results were achieved in 72% of cases. The scars had become less red (192/327 scored no redness at the end of the study versus 92/327 upon initial), less raised (272/327 scored a flat scar versus 72/327 upon initial) and demonstrated an improved viscoelasticity (192/327 scored a soft skin versus 62/327 upon initial). Laser treatment did not improve contractures in post‐burn hypertrophic scars. Results were not confirmed using objective measurement tools, as these were not available to us.

The impact of the types of microorganisms isolated from blood and wounds on the results of treatment in burn patients with sepsis.

UNLABELLED Despite development of combustiology, infections continue to be the most important cause of death among patients with burns. Sepsis is the most severe clinical presentation of infection in patients after thermal injuries who require immediate treatment. Early diagnosis and proper treatment of sepsis are important in the clinical management that is often hampered for multiple reasons, e.g. impaired patient immunity, problems with microorganisms with multi-antibacterial drug resistance. The aim of the study was to assess effect of type of a microorganism isolated from blood and wound on results of treatment of sepsis in patients with burns. MATERIAL AND METHODS Effect of type of microorganisms isolated from blood and wound on the result of treatment of sepsis was studied in 338 patients hospitalized immediately after an injury in Centre for Burn Treatment in Siemianowice Śląskie in years 2003 - 2004 (at the age of 18 - 96 years, 66 women and 272 men). Clinical symptoms of generalized infection were found in all study subjects. The study group was divided into two subgroups: cured patients and patients who died of sepsis. The following parameters were assessed in both subgroups: type of microorganism isolated from blood, type of microorganism isolated from wound as well as occurrence of the same and different infections of blood and burn wound. RESULTS positive blood cultures were found in 165 patients (48.8%), 106 (64.2%) were cured, 59 (35.8%) died. The most commonly isolated microorganisms in cured patients were Gram(+) Staphylococcus epidermidis MRSE (19.81%) and Staphylococcus aureus MRSA (18.87%). Gram(-) intestinal rods were least commonly isolated from this group. The most commonly isolated microorganisms from blood of patients who were to die, included non-fermenting Gram(-) rods Acinetobacter baumannii (35.59%) and Pseudomonas aeruginosa (22.03%). Mixed bacterial flora was found in the blood of 22.03% patients. Among patients who were to die, the same microorganisms were found in the blood and in the wound in 32.2% of patients, while this rate was 17.92 in cured patients. The most commonly found bacteria in the blood and burn wound in the cured patients included Staphylococcus aureus MRSA (31.58%) and Staphylococcus aureus (21.05%). In the group of patients who were to die, the most common bacteria isolated simultaneously from the blood and burn wound included Acinetobacter baumannii (47.37%) and Pseudomonas aeruginosa (36.84%). CONCLUSIONS 1. The patients with thermal injuries are at higher risk of death in the event of sepsis caused by Gram(-) bacteria versus Gram(+) bacteria. 2. Infection of blood and burn wound caused by the same bacteria Pseudomonas aeruginosa and Acinetobacter baumanni increases the risk of death due to sepsis in patients with burns following thermal injuries.

A review of decellurization methods caused by an urgent need for quality control of cell-free extracellular matrix' scaffolds and their role in regenerative medicine: REVIEW OF DECELLURIZATION METHODS

The natural extracellular matrix (ECM),thanks to its specific properties (e.g., collagenous lattice, a reservoir of growth factors, ECM-cell anchoring areas, an optimal pH and CO2 ),ensures an optimal microenvironment for homeostatic and regenerative cell development. In the context of regenerative medicine, ECM is a lair for residual and infiltrative cells. The aim of the clinical usage of cell-free ECM scaffolds is the enhancement of tissue regeneration with possible minimization of an adverse host reaction on allogeneic or xenogeneic biomaterial. Thus, the objective of decellularization is to obtain acellular grafts characterized by optimal biological properties, such as a lack of remaining cellular elements (e.g., cell membrane phospholipids and proteins, nucleic acids, mitochondria), lack of immunogenicity, lack of calcification promotion and lack of cytotoxicity (e.g., in unrinsed detergents). Furthermore, cell-free ECM scaffolds should present the optimal mechanical and structural properties that may ensure the biocompatibility of the graft. The maintenance of the ultrastructure composition of the ECM is one of the most important goals of decellularization. All physical, chemical, and biological methods proposed (used separately or in combination to extract cells from tissues/organs) are not 100% effective in cell removal and always cause a disruption of the ECM texture, as well as a probable loss of important structure components. Although cell-free ECM scaffolds are generally classified as medical devices, there are no widely accepted or legally defined criteria for quality control/evaluation methods of obtained matrices. Such criteria must be provided. Some of them have been proposed in this manuscript.

Controlling the Crystallinity of Thermoresponsive Poly(2-oxazoline)-Based Nanolayers to Cell Adhesion and Detachment

Semicrystalline, thermoresponsive poly(2-isopropyl-2-oxazoline) (PIPOx) layers covalently bonded to glass or silica wafers were obtained via the surface-termination of the living polymer chains. Polymer solutions in acetonitrile were exposed to 50 °C for various time periods and were poured onto the functionalized solid wafers. Fibrillar crystallites formed in polymerization solutions settled down onto the wafers next to the amorphous polymer. The amount of crystallites adsorbed on thermoresponsive polymer layers depended on the annealing time of the PIPOx solution. The wettability of PIPOx layers decreased with the increasing amount of crystallites. The higher content of crystallites weakened the temperature response of the layer, as evidenced by the philicity and thickness measurements. Semicrystalline thermoresponsive PIPOx layers were used as biomaterials for human dermal fibroblasts (HDFs) culture and detachment. The presence of crystallites on the PIPOx layers promoted the proliferation of HDFs. Changes in the physicochemical properties of the layer, caused by the temperature response of the polymer, led to the change in the cells shape from a spindle-like to an ellipsoidal shape, which resulted in their detachment. A supporting membrane was used to assist the detachment of the cells from PIPOx biosurfaces and to prevent the rolling of the sheet.

Transfer of fibroblast sheets cultured on thermoresponsive dishes with membranes

In cell or tissue engineering, it is essential to develop a support for cell-to-cell adhesion, which leads to the generation of cell sheets connected by extracellular matrix. Such supports must be hydrophobic and should result in a detachable cell sheet. A thermoresponsive support that enables the cultured cell sheet to detach using only a change in temperature could be an interesting alternative in regenerative medicine. The aim of this study was to evaluate plates covered with thermoresponsive polymers as supports for the formation of fibroblast sheets and to develop a damage-free procedure for cell sheet transfer with the use of membranes as transfer tools. Human skin fibroblasts were seeded on supports coated with a thermoresponsive polymer: commercial UpCell™ dishes (NUNC™) coated with thermoresponsive poly(N-isopropylacrylamide) (PNIPAM) and dishes coated with thermoresponsive poly(tri(ethylene glycol) monoethyl ether methacrylate) (P(TEGMA-EE)). Confluent fibroblast sheets were effectively cultured and harvested from both commercial PNIPAM-coated dishes and laboratory P(TEGMA-EE)-coated dishes. To transfer a detached cell sheet, two membranes, Immobilon-P® and SUPRATHEL®, were examined. The use of SUPRATHEL for relocating the cell sheets opens a new possibility for the clinical treatment of wounds. This study established the background for implementing thermoresponsive supports for transplanting in vitro cultured fibroblasts.

The efficiency of burn wounds debridement with use of hydrosurgery--our experiences.

UNLABELLED The aim of the study was to present experience of doctors from the Center for Burns Treatment in Siemianowice Śląskie who use a Versajet system. MATERIAL AND METHODS Debridement with the use of a water jet was applied in 70 patients with IIb° /III° thermal burns in the period between 2009-2013. The corresponding evaluation involved duration of operation, locations, wound surfaces and technique of debridement after operation. Microbiological tests were also performed before and after debridement. RESULTS AND CONCLUSIONS In the paper, its authors draw attention to the short time of debridement. The application of a water jet allows precise debridement, particularly in hardly accessible places, preparation of wounds to be covered with autologous split thickness skin graft, protection of healthy tissues.

Literature Review Concerning Cell and Skin Substitute Cultures Obtained by Means of Tissue Engineering used in the Treatment of Burns

Center For The Treatment of Burns in Siemianowice Śląskie1 Dyrektor: dr n. med. M. nowak Technical-Humanistic Academy in Bielsko Biała. Faculty of Health Science. Faculty of Emergency Medicine2 Dziekan Wydziału: dr hab. M. Mikulska, prof. ATH Higher School of Applied Sciences in Ruda Śląska. Department of Health3 Dziekan Wydziału: doc. dr n. med. I. Caus-Woźniak Center of Polymer and Carbon Resources of the Polish Academy of Science in Zabrze4 Dyrektor: prof. dr hab. a. dworak Chair and Department of General, Molecular, and Genetic Biology, Silesian Medical University in Katowice5 Kierownik: prof. dr hab. A. L. Sieroń

Alterations in taste perception as a result of hyperbaric oxygen therapy.

The present study evaluates the effect of hyperbaric oxygen therapy on taste sensitivity, hedonic perception of taste, and food preferences. The studied groups included 197 people in total (79 in the study group; 118 in the control group). All patients from the study group were treated with hyperbaric oxygen therapy due to chronic non-healing wounds. The control group consisted of healthy people, who did not receive hyperbaric oxygen therapy. The taste intensity, recognition thresholds, and hedonic perception were examined using gustatory tests. The aqueous solutions of sucrose for sweet, sodium chloride for salty, citric acid for sour, quinine hydrochloride for bitter, and monosodium glutamate for umami taste were used. The participants fulfilled the questionnaire to examine pleasure derived from eating certain types of dishes. Gustatory tests and analyses of the pleasure derived from eating in the study group were carried out before the first exposure to hyperbaric oxygen and then at the end of therapy, after at least 25 sessions of treatment. In the control group, examination of perception of taste sensations was conducted only once. The results of comparing patients with non-healing wounds with healthy people are characterized by reduced taste sensitivity. After participation in hyperbaric oxygen therapy, the improvement in perception of taste sensations and changes in hedonic evaluation have occurred among patients with non-healing wounds. In terms of food preference, a decreased desire for eating sweet desserts, chocolate, and crisps was observed in those patients who received hyperbaric oxygen therapy.