Silver Nanoforms as a Therapeutic Agent for Killing Escherichia coli and Certain ESKAPE Pathogens.

The scope of this study included the preparation of silver nanoforms with high antimicrobial efficacy, low cost, and ease of application. The term 'silver nanoforms' refers to silver located on the amorphous or crystalline titanium dioxide (TiO2). Silver nanoforms may be used as an alternative to antibiotics in killing bacteria. Pure and silver-incorporated titanium (used as a carrier) was prepared using the sol-gel-modified method. Physical and chemical properties of the samples were described, and the antibacterial activity was indicated using the following strains of bacteria: Staphylococcus aureus, Klebsiella pneumoniae (ESKAPE pathogens), and Escherichia coli. The results have shown that the antibacterial activity of silver nanoforms with amorphous TiO2 is much better than that in the samples based on anatase (crystalline TiO2). The sensitivity of the tested bacteria to silver nanoforms depends on physical and chemical properties of the nanoforms and individual characteristics of the bacteria. For the first time, significant participation of amorphous TiO2 in antibacterial compounds has been described through this study.

Selection and electrophoretic characterization of Salmonella enterica subsp. enterica biocide variants resistant to antibiotics.

The proposed research outlines a serious common concern of Salmonella resistance to antimicrobials following prolonged exposure to the disinfectants (biocides). These phenotypes of bacteria could potentially result in hard to treat infections. Typical for avian sources, biocide sensitive S. enterica subsp. enterica serovars: Typhimurium, Enteritidis, Virchow and Zanzibar and their isogenic biocide-tolerant variants were studied in order to investigate bacteriostatic effect of two commercially available biocide formulations: potassium peroxymonosulfate (P) and dodecylamine based structure (triamine, D). We found that cultivating of the bacteria in the medium supplemented with a blend containing P did not influence their antibiotic susceptibility pattern. In contrast, tolerance of bacteria to D compound resulted in resistance to co-trimoxazole, cefotaxime and ciprofloxacin of which two cefotaxime and ciprofloxacin are used commonly for the treatment of invasive Salmonella infections in humans. The dependency between OMP patterns and the level of Salmonella survival in media containing the biocides was observed merely in serovar Typhimurium. In conclusion, these results suggest that Salmonella strains challenged by prolonged treatment with the disinfectants become resistant to antibiotics, however it depends on Salmonella serovar and the chemical used. This paper also highlights the loop-mediated isothermal amplification (LAMP) as a technique that offers great benefits to microbiological detecting of Salmonella species in the samples.

Osteoprotegerin, RANKL, and bone turnover in primary hyperparathyroidism: the effect of parathyroidectomy and treatment with alendronate.

Receptor activator of nuclear factor-κB (RANK), RANK ligand (RANKL), and its decoy receptor osteoprotegerin (OPG) play key roles in regulating bone turnover. The OPG/RANKL/RANK system is regulated by many hormones and cytokines, among which parathormone (PTH) seems to be one of the most important. Primary hyperparathyroidism (PHPT) with chronic excess of PTH and enhanced bone resorption provides an opportunity to observe the relationships between PTH, OPG and RANKL. From a group of 63 patients with PHPT, 29 underwent effective parathyroidectomy (PTX) and 33 were treated with alendronate. After one year, bone mineral density (BMD) improved in both groups, but the biochemical disturbances and PTH returned to normal only after PTX. The baseline serum concentrations of OPG and RANKL were higher in PHPT patients than in healthy controls, whilst the OPG/RANKL-F ratio was lower. The mean OPG concentration did not change after PTX, and slightly increased after alendronate treatment despite the unchanged PTH. Twelve months after treatment, RANKL slightly declined in both groups and the ratio of OPG/RANKL consistently increased. Serum OPG and RANKL did not correlate with PTH before or after PTX or alendronate treatment. In conclusion, bone resorption in PHPT is accompanied by a high serum concentration of OPG and RANKL as well as a low OPG/RANKL ratio. Both parathyroidectomy and treatment with alendronate diminish bone resorption, and correct the OPG/RANKL ratio in favor of OPG, although the mechanisms of their actions are different. Serum OPG concentration does not depend directly on PTH.

Human PEX1 is mutated in complementation group 1 of the peroxisome biogenesis disorders.

Human peroxisome biogenesis disorders (PBDs) are a group of genetically heterogeneous autosomal-recessive disease caused by mutations in PEX genes that encode peroxins, proteins required for peroxisome biogenesis. These lethal diseases include Zellweger syndrome (ZS), neonatal adrenoleukodystrophy (NALD) and infantile Refsum's disease (IRD), three phenotypes now thought to represent a continuum of clinical features that are most severe in ZS, milder in NALD and least severe in IRD2. At least eleven PBD complementation groups have been identified by somatic-cell hybridization analysis compared to the eighteen PEX complementation groups that have been found in yeast. We have cloned the human PEX1 gene encoding a 147-kD member of the AAA protein family (ATPases associated with diverse cellular activities), which is the putative orthologue of Saccharomyces cerevisiae Pex1p (ScPex1p). Human PEX1 has been identified by computer-based 'homology probing' using the ScPex1p sequence to screen databases of expressed sequence tags (dbEST) for human cDNA clones. Expression of PEX1 rescued the cells from the biogenesis defect in human fibroblasts of complementation group 1 (CG1), the largest PBD complementation group. We show that PEX1 is mutated in CG1 patients.

Type of desmin expression in cardiomyocytes - a good marker of heart failure development in idiopathic dilated cardiomyopathy.

AIM: The aim of this study was to determine whether remodelling of the desmin (DES) cytoskeleton affects myocardial function and whether it could be a useful marker of disease progression in patients with idiopathic dilated cardiomyopathy (IDCM). MATERIAL AND METHODS: Endomyocardial biopsy was performed in 195 IDCM patients, and five to six specimens were collected from the left ventricle. DES expression was evaluated using tissue immunostaining and Western blotting. The study population was assigned to four groups according to DES expression type: I, normal DES staining at Z-lines giving a regular pattern of cross-striation (n = 57); IIA, increased DES staining with a regular pattern of cross-striation (n = 40); IIB, increased DES staining with an irregular pattern of cross-striation and/or the presence of aggregates (n = 56); and III, decreased/lack of DES staining (n = 42). Fibrosis, cardiomyocyte hypertrophy and ultrastructure were assessed for the four types of DES expression. RESULTS: The pathological types of DES expression (IIB or III) were associated with pathological changes in mitochondria and the contractile apparatus. Cardiomyocyte diameter and level of fibrosis were both significantly affected. DES expression type correlated with NYHA class, left ventricular end-diastolic diameter, left ventricular ejection fraction and the level of N-terminal pro-brain natriuretic protein. CONCLUSION: The type of immunohistochemical DES expression correlated with the level of myocardial injury at the cellular and organ levels. This correlation was similar to that observed between DES expression and the well-established biochemical, echocardiographic and clinical parameters of heart failure (HF). DES expression type could be used as an important diagnostic feature of HF development.

Vitamin D and selected cytokine concentrations in postmenopausal women in relation to metabolic disorders and physical activity.

Obesity and metabolic disturbances constitute significant health problems in elderly women. Due to the multifactorial background of these disorders, assessing the interaction between risk factors remains a significant part of prevention and health promotion. Studies have illustrated a relationship between low physical activity and vitamin D deficiency with obesity and its complications. Furthermore, vitamin D affects the production of adipokines and the inflammatory response in adipose tissue. The aim of our study was to determine the association between selected adipokines, vitamin D concentrations, physical activity (PA), and visceral adiposity index (VAI) in postmenopausal women. The study sample consisted of 318 ethnically homogenous postmenopausal women aged 50-60. Both anthropometric measurements (BMI, WC, WHR) and biochemical measurements (TC, HDL, LDL, TG, AIP, glucose, insulin, IL-6, TNF-α, adiponectin, leptin) were made, and PA by IPAQ were recorded. Body mass index (BMI), waist-to-hip ratio (WHR), HOMA, atherogenic index of plasma (AIP), and VAI were calculated using the standard formulas. We observed a negative correlation between BMI, WC, insulin, HOMA, and PA. We determined that there is a negative association between leptin and vitamin 25(OH)D concentrations (P = 0.007) and a positive association with adiponectin (P = 0.014). The results of the multiple linear regression analysis indicate that vitamin D and HOMA are independent factors that significantly affect leptin and adiponectin levels, contrary to VAI.

Ultrastructural response of arcuate nucleus neurons to fasting in aged rats.

The arcuate nucleus of the hypothalamus (ARH) is involved in the control of energy homeostasis. Leptin - an adipocyte derived hormone - is known to act on the hypothalamic nuclei and thus to control body weight by food intake reduction. Oxidative stress is believed to be implicated in leptin signalling. However, its relevance for leptin-induced signal transduction within ARH remains unclear. The goal of the study was to investigate the effect of fasting on morphological alterations of the neuronal endoplasmic reticulum/Golgi network as well as on the expression of leptin receptors in the arcuate nucleus of aged rats. Male Wistar rats, aged 24 months, were fasted for 96 hours. The control animals were fed ad libitum. Membranous whorls in the ARH neurons were visualized using the electron microscopy technique. Leptin receptors in the membranes of ARH neurons were determined immunohistochemically (IHC), and soluble leptin receptors in the plasma as well as plasma isoprostanes were quantified immunochemically (ELISA). An intense formation of membranous whorls was observed, directly associated with the cisternae of the rough endoplasmic reticulum, as well as lamellar bodies. Interestingly, the whorls were often localized near a well-developed Golgi complex. Moreover, some Golgi complexes displayed an early stage of whorl formation. Groups of residual lipofuscin granules were found in the immediate proximity of the whorls. An increased immunoreactivity with neuronal leptin receptors suggests that hypersensitive neurons may still effectively respond to the fasting serum levels of leptin, mediating ultrastructural transformation of ARH neurons during short-term fasting. Having observed a significant accumulation of lipofuscin granules and a marked increase of total 8-isoprostane serum level in the fasting rats, we hypothesize that signal transduction within the neurons of ARH is dependent on oxidative stress phenomena.

"Extracorporeal Membrane Oxygenation for Greater Poland" Program: How to Save Lives and Develop Organ Donation?

The "ECMO for Greater Poland" program takes full advantage of the extracorporeal membrane oxygenation (ECMO) perfusion therapy opportunities to promote the health of the 3.5 million inhabitants in the region. The main implementation areas are treatment of patients with hypothermia; severe reversible respiratory failure (RRF); critical states resulting in heart failure, that is, cardiac arrest, cardiogenic shock, or acute intoxication; and promotion of the donor after circulatory death (DCD) strategy in selected organ donor cases, after unsuccessful life-saving treatment, to achieve organ recovery. This organizational model is complex and expensive, so we used advanced high-fidelity medical simulation tests to prepare for real-life experience. Over the course of 4 months we performed scenarios including "ECMO for DCD," "ECMO for extended cardiopulmonary resuscitation," "ECMO for RRF," and "ECMO in hypothermia." Soon after these simulations, Maastricht category II DCD procedures were performed involving real patients and resulting in 2 successful double kidney transplantations for the first time in Poland. One month later we treated 2 hypothermia patients (7 adult patients with heart failure and 5 patients with reversible respiratory failure) with ECMO for the first time in the region. Fortunately, we have discovered an important new role of medical simulation. It can be used not only for skills testing but also as a tool to create non-existing procedures and unavailable algorithms. The result of these program activities will promote the care and treatment of patients in critical condition with ECMO therapy as well as increase the potential organ pool from DCDs in the Greater Poland region of Poland.

Interaction of plasmenylcholine with free radicals in selected model systems.

Plasmalogens (Plg) - naturally occurring glycerophospholipids with the vinyl-ether group in the sn-1 position are generally viewed as physiological antioxidants. Although there are numerous examples of antioxidant action of plasmalogen in cell cultures and in experimental animals, this hypothesis is far from being satisfactorily proven due to substantial limitations of such studies. Thus, plasmalogen reactivity in cells results in the accumulation of toxic byproducts and the experimental design is usually too complicated to evaluate the protective function of solely one type of lipid molecular species. In this study, experiments were performed in homogenous and heterogeneous model systems consisting of solutions in organic solvents as well as micelles and liposomes containing pure synthetic plasmenylcholines. Under the experimental conditions used, chemical reactivity of plasmalogens could be attributed to specific fatty acid esterification pattern. This is important because the chemical reactivity cannot be separated from physico-chemical properties of the lipids. Time-dependent formation of phospholipid and cholesterol hydroperoxides were determined by iodometric assay and HPLC-EC. EPR oximetry and Clark electrode were employed to detect the accompanying changes in oxygen concentration. Oxidation of the studied lipids was monitored by standard colorimetric TBARS method as well as MALDI-TOF mass spectrometry. Our data indicate that the reactivity of sn-2 monounsaturated vinyl ether lipids in peroxyl radical-induced or iron-catalyzed peroxidation reactions is comparable with that of their diacyl analogs. In samples containing cholesterol and plasmalogens, oxidative processes lead to accumulation of the radical oxidation product of cholesterol. It can be concluded that the antioxidant action of plasmalogens takes place intramolecularly rather than intermolecularly and depends on the degree of unsaturation of esterified fatty acids. Thus, it is questionable if plasmalogens can really be viewed as "endogenous antioxidant", even though they may exhibit, under special conditions, protective effect.

Oxidation-Induced Increase In Photoreactivity of Bovine Retinal Lipid Extract.

The mammalian retina contains a high level of polyunsaturated fatty acids, including docosahexaenoic acid (22:6) (DHA), which are highly susceptible to oxidation. It has been shown that one of the products of DHA oxidation-carboxyethylpyrrole (CEP), generated in situ, causes modifications of retinal proteins and induces inflammation response in the outer retina. These contributing factors may play a role in the development of age-related macular degeneration (AMD). It is also possible that some of the lipid oxidation products are photoreactive, and upon irradiation with blue light may generate reactive oxygen species. Therefore, in this work we analysed oxidation-induced changes in photoreactivity of lipids extracted from bovine neural retinas. Lipid composition of bovine neural retinas closely resembles that of human retinas making the bovine tissue a convenient model for studying the photoreactivity and potential phototoxicity of oxidized human retinal lipids. Lipid composition of bovine neural retinas Folch' extracts (BRex) was determined by gas chromatography (GC) and liquid chromatography coupled to an electrospray ionization source-mass spectrometer (LC-ESI-MS) analysis. Liposomes prepared from BRex, equilibrated with air, were oxidized in the dark at 37 °C for up to 400 h. The photoreactivity of BRex at different stages of oxidation was studied by EPR-oximetry and EPR-spin trapping. Photogeneration of singlet oxygen (1O2, 1Δg) by BRex was measured using time-resolved detection of the characteristic phosphorescence at 1270 nm. To establish contribution of lipid components to the analysed photoreactivity of Folch' extract of bovine retinas, a mixture of selected synthetic lipids in percent by weight (w/w %) ratio resembling that of the BRex has been also studied. Folch's extraction of bovine neural retinas was very susceptible to oxidation despite the presence of powerful endogenous antioxidants such as α-tocopherol and zeaxanthin. Non-oxidized and oxidized BRex photogenerated singlet oxygen with moderate quantum yield. Blue-light induced generation of superoxide anion by Folch' extract of bovine neural retinas strongly depended on the oxidation time. The observed photoreactivity of the studied extract gradually increased during its in vitro oxidation.

A novel canine B-cell leukaemia cell line. Establishment, characterisation and sensitivity to chemotherapeutics.

We established a new B-cell leukaemia cell line CLB70 from a dog with chronic lymphocytic leukaemia. This cell line is positive for CD20, CD45, CD79a, MHC class II, IgG, IgM; weakly positive for CD21; and negative for CD3, CD4, CD5, CD8, CD14, CD34, CD117. PCR for antigen receptor gene rearrangement (PARR) analysis revealed a biclonal immunoglobulin heavy chain (IgH) gene rearrangement and negative result for TCRγ. Western blot analysis of anti- and pro-apoptotic proteins showed increased expression of Bcl-2, Mcl-1, NF-kB, and Ras, and decreased expression of p53. CLB70 cells grow rapidly in vitro and are tumourigenic in nude mice. The CLB70 line is highly sensitive to doxorubicin, less sensitive to etoposide and imatinib, and resistant to piroxicam, celecoxib and dexamethasone. Our results indicate that CLB70 cells are derived from mature B-cells and they may be a useful tool for the development of new therapeutic strategies for both dogs and humans.

Human testis expresses a specific poly(A)-binding protein.

In testis mRNA stability and translation initiation are extensively under the control of poly(A)-binding proteins (PABP). Here we have cloned a new human testis-specific PABP (PABP3) of 631 amino acids (70.1 kDa) with 92.5% identical residues to the ubiquitous PABP1. A northern blot of multiple human tissues hybridised with PABP3- and PABP1-specific oligonucleotide probes revealed two PABP3 mRNAs (2.1 and 2.5 kb) detected only in testis, whereas PABP1 mRNA (3.2 kb) was present in all tested tissues. In human adult testis, PABP3 mRNA expression was restricted to round spermatids, whereas PABP1 was expressed in these cells as well as in pachytene spermatocytes. PABP3-specific antibodies identified a protein of 70 kDa in human testis extracts. This protein binds poly(A) with a slightly lower affinity as compared to PABP1. The human PABP3 gene is intronless with a transcription start site 61 nt upstream from the initiation codon. A sequence of 256 bp upstream from the transcription start site drives the promoter activity of PABP3 and its tissue-specific expression. The expression of PABP3 might be a way to bypass PABP1 translational repression and to produce the amount of PABP needed for active mRNA translation in spermatids.

Immunophenotypic characterization of canine malignant lymphoma: a retrospective study of cases diagnosed in Poland Lower Silesia, over the period 2011-2013.

Lymphoma is the most frequently diagnosed cancer of the canine haematopoietic system. In this study, the flow cytometry and polymerase chain reaction (PCR) analysis were used to characterize a series of canine lymphomas in detail. The aim of this study was to determine the incidence of B- and T-cell high-grade lymphomas and their immunophenotypic characterization in Lower Silesia, Poland. The results show that the frequency of each type of lymphoma is 71% for B-cell and 17% for T-cell lymphomas. In two cases the PCR techniques confirmed the presence of simultaneous double gene rearrangements of the BCR and TCR receptors.

Ataxia telangiectasia heterozygotes and patients display increased fluidity and decrease in contents of sulfhydryl groups in red blood cell membranes.

In red blood cell membranes of ataxia telangiectasia mutated (ATM) homozygotes and heterozygotes, decreased values of the corrected fluorescence anisotropy and the anisotropy parameter were found, indicating increased fluidity and decreased microviscosity, respectively. These changes in membranes were accompanied by a decrease in SH-groups and an increase in malondialdehyde (MDA) contents. The content of MDA both in homozygotes and in heterozygotes exceeded roughly threefold the respective control values. Decreased content of GSH in red blood cells was found only in ATM homozygotes. The change most specific for the ATM gene appears to be the increase in fluidity, since only this parameter displays the proportionally greater changes in ATM homozygotes compared to ATM heterozygotes. The observations presented here may indicate that the ATM gene is expressed in precursors of red blood cells and deficiency of normal AT gene function may produce the changes which persist in circulating cells.

Carriers of ataxia-telangiectasia gene display additional protein fraction and changes in the environment of SH groups in erythrocyte membrane.

Additional protein fraction migrating slower than spectrin has been detected in erythrocyte membranes from an ataxia-telangiectasia (A-T) patient and from his mother (A-T heterozygote). In erythrocyte membranes labelled with maleimide spin label changes in signal of the weakly immobilized spin label as related to that of strongly immobilized one (w/s) were noted. In comparison to age-matched control groups the values of w/s were lower in A-T heterozygotes (ten persons) and higher in A-T homozygotes (four persons). In control persons the values of w/s increased with age, whereas in families with A-T no significant differences in this parameter were noted between children and parents. The presence of additional protein fraction in erythrocytes membranes of A-T patient and A-T heterozygote indicates that these phenotypes can be differentiated from the healthy control persons for the first time on the basis of changes detected in the erythrocytes. This change in erythrocyte membrane may explain the decrease in the w/s parameter of electron spin resonance in A-T heterozygotes. On the other hand increased values of w/s in A-T patients may be caused by disease process.

Human testis specifically expresses a homologue of the rodent T lymphocytes RT6 mRNA.

A human homologue of the rodent T cell mono ADP-ribosyl transferase RT6 mRNA was identified by a systematic analysis of human testis transcripts. This messenger encodes for a precursor protein of 367 aa (MW: 41.5 kDa) which exhibits a peptide signal, consensus domains for mono ADP-ribosyl transferase and a C-terminal part characteristic of glycophosphatidyl inositol anchored protein. This mRNA, transcribed from a gene localized in 4q13-q21, is not expressed in white blood cells but is specific for human testis in which it is likely to correspond to a new ADP-ribosyl transferase.

Fluctuations in somatosensory responsiveness and baseline firing rates of neurons in the lateral striatum of freely moving rats: effects of intranigral apomorphine.

Somatosensory responsiveness and baseline firing rates of 102 striatal neurons were studied in freely moving rats. For individual neurons, mean levels of responsiveness and baseline firing fluctuated unpredictably in direction and magnitude and independently of each other throughout an experiment. Following microinjections of apomorphine into the substantia nigra, which were used as a means of reducing nigral output activity, the magnitude of fluctuations in striatal somatosensory responsiveness significantly increased, while the magnitude of fluctuations in baseline firing was unaltered. The receptive zones of 54 neurons studied in control experiments remained stable, whereas receptive zones changed in 12 of 25 neurons studied after apomorphine microinjection. Normal nigrostriatal dopamine transmission appears to selectively restrict the magnitude of fluctuations in responsiveness of striatal neurons to corticostriatal synaptic input and may exert additional control over afferent projections from cutaneous receptive zones to these neurons.

Low constitutive activity of aryl hydrocarbon hydroxylase in phytohaemoagglutinin-stimulated peripheral blood mononuclear cells of children after treatment for acute lymphoblastic leukemia.

In six children in the first remission after treatment of acute lymphoblastic leukemia (ALL) the constitutive activity of aryl hydrocarbon (benzo-[alpha]pyrene) hydroxylase (AHH) (0.14 +/- 0.04 U, i.e. pmol of 3-OH-benzo[alpha] pyrene x 10(-6) cells x min-1) in phytohaemoagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PBMC) was significantly lower (P 0.05) than the constitutive activities in the parents of the children (0.25 +/- 0.09 U) and in the control adult group (0.26 +/- 0.09 U). In methylcholanthrene (MC)-induced activities of AHH no significant differences were observed between the above groups.