Search

The risk of a hemolytic reaction during the transfusion of ABO non‐identical PC is determined by the presence of natural anti‐A IgM antibodies, the titer of which may increase after infections. The aim of the study was to evaluate the titer of anti‐A isohemagglutinins in platelet concentrate (PC) obtained by apheresis from group O donors who experienced SARS‐CoV‐2 infection, and to compare the titer before and after infection. A retrospective single‐center analysis of 21 PC donors with a previous COVID‐19 history was performed. The results showed neither a statistically important increase in the anti‐A IgM antibody titers nor a significant correlation between the anti‐A IgM antibody level and anti‐SARS‐CoV‐2S1 antibody titer in the donors with an asymptomatic or mild COVID‐19. Further population‐based studies on anti‐A titers are necessary for a comprehensive assessment of this phenomenon.

The treatment of cancer patients with immune checkpoint inhibitors (ICI) (anti-CTLA-4, anti-PD-1, anti-PD-L1, combined therapy anti-PD-1/PD-L1 with anti-CTLA-4) has without doubt been a significant breakthrough in the field of oncology in recent years and constitutes a major step forward as a novel type of immunotherapy in the treatment of cancer. ICIs have contributed to a significant improvement in the outcome of treatment and prognosis of patients with different types of malignancy. With the expansion of the use of ICIs, it is expected that caregivers will face new challenges, namely, they will have to manage the adverse side effects associated with the use of these drugs. New treatment options pose new challenges not only for oncologists but also for specialists in other clinical fields, including general practitioners (GPs). They also endorse the need for taking a holistic approach to the patient, which is a principle widely recognized in oncology and especially relevant in the case of the expanding use of ICIs, which may give rise to a wide variety of organ complications resulting from treatment. Knowledge and awareness of the spectrum of immune-related adverse events (irAEs) will allow doctors to qualify patients for treatment more appropriately, prevent complications, correctly recognize, and ultimately treat them. Additionally, patients with more non-specific symptoms would be expected, in the first instance, to consult their general practitioners, as complications may appear even after the termination of treatment and do not always proceed in line with disease progression. Dealing with any iatrogenic complications, will not only be the remit of oncologists but because of the likelihood that specific organs may be affected, is likely to extend also to specialists in various fields of internal medicine. These specialists, e.g., endocrinologists, dermatologists, pulmonologists, and gastroenterologists, are likely to receive referrals for patients suffering from specific types of adverse events or will be asked to provide care in cases requiring hospitalization of patients with complications in their field of expertise. In view of these considerations, we believe that there is an urgent need for multidisciplinary teamwork in the treatment of cancer patients undergoing immunotherapy and suffering the consequent adverse reactions to treatment.

Despite intensive research, the physiology of the serological response to SARS-CoV-2 infection and its dynamics during the recovery period remain incompletely understood. Regulation of the immune response seems all the more important as it plays a role in both virus clearance during infection and the potential development of long-term resistance to reinfection. A case of convalescent plasma donor is described in whom was observed a prolonged enhanced immune response to infection in the form of a persistently high level of anti-Spike IgG despite subsequent plasma donations. The presented donor experienced COVID-19 interstitial pneumonia, requiring pharmacotherapy in a hospital setting (therapy involving azithromycin, chloroquine and protease inhibitors), which allowed him to achieve remission. The described donor donated plasma nine times during convalescence, each time showing a satisfactory level of anti-Spike IgG. The presented case highlights the multifactorial regulation of the serological response in the course of SARS-CoV-2 infection, which may include the long-term effect of pharmacotherapy, especially in the field of antiretroviral drugs. While the authors are not yet able to clearly define the importance of antiretroviral therapy in regulating the humoral response in COVID-19 patients, it is supposed it may be important in the subsequent antibody production.

Transfusion of blood and blood components is one of the widely used medical procedures. The responsibility for provision of blood and blood components lies with Polish blood transfusion centers (CKiK) substantively supervised by the Institute of Hematology and Transfusion Medicine. Hospital based blood banks, hospital wards, immunohematology laboratories are supervised by CKiK. Every year approximately 600 thousand people donate blood, more than 75% of which are regular donors. The annual number of donations is maintained at a constant level of about 1.3 million. The aim of this position paper is to present the current status, achievements and advancement in collection, preparation and testing procedures and methods used to obtain the best possible quality blood components dedicated for clinical use. The aim is also to show the structure of blood transfusion service in Poland. Collaboration of all organizational units involved in transfusion medicine is crucial for providing high quality health care for patients. Polish transfusiologists and blood transfusion officers have largely contributed to transfusion science with numerous publications and significant research work. The implementation of novel methods and ongoing research positions Polish blood transfusion service among those of highly developed countries.

Despite the increasing number of patients suffering from tick-borne encephalitis (TBE), Lyme disease, and their co-infection, the mechanisms of the development of these diseases and their effects on the human body are still unknown. Therefore, the aim of this study was to evaluate the changes in the proteomic profile of human plasma induced by the development of TBE and to compare it with changes in TBE patients co-infected with other tick-borne pathogens. The results obtained by proteomic analysis using a nanoLC-Q Exactive HF mass spectrometer showed that the most highly elevated groups of proteins in the plasma of TBE patients with co-infection were involved in the pro-inflammatory response and protein degradation, while the antioxidant proteins and factors responsible for protein biosynthesis were mainly downregulated. These results were accompanied by enhanced GSH- and 4-HNE-protein adducts formation, observed in TBE and co-infected patients at a higher level than in the case of patients with only TBE. In conclusion, the differences in the proteomic profiles between patients with TBE and co-infected patients indicate that these diseases are significantly diverse and, consequently, require different treatment, which is particularly important for further research, including the development of novel diagnostics tools.

Anemia is a common feature in about 40% of patients at the moment of cancer diagnosis and in more than half of patients on anticancer therapy. Therapeutic alternatives in cancer patients with anemia include substitution of lacking agents, red blood cell transfusions, and erythropoiesis-stimulating agents (ESAs). The advantages of red blood cell transfusions are rapid increase of hemoglobin concentration and effectiveness independent of the cause of anemia. However, several adverse reactions may occur after blood component transfusion. ESAs act through stimulation of erythropoietin receptors. Use of ESAs reduces the need for red blood cell transfusions, decreases the risk of post-transfusion adverse reactions, and improves the quality of life of cancer patients with chemotherapy-induced anemia. In accordance with registered indications, ESA may be administered in non-myeloid cancer patients with chemotherapy-induced anemia. Thromboembolic events and arterial hypertension are known risks of ESA treatment. If ESAs are used in accordance with currently approved indications and are not administered when hemoglobin (Hb) concentration is 12 g/dL or above, there is no observed unfavorable effect on survival or thromboembolic risk. The administration of RBC transfusions without delay is justified in patients with Hb under 7–8 g/dL and/or severe anemia-related symptoms (even at higher Hb levels) and the need for immediate Hb and symptom improvement. The goal of ESA treatment is maintenance of the lowest hemoglobin concentration needed to avoid red blood cell transfusion. ESAs may be used in patients with symptomatic chemotherapy-induced anemia and Hb concentration at 10 g/dL or below. There is no indication for ESAs in patients who are not receiving chemotherapy or who are receiving radiotherapy.

Osteochondral defects of the knee are common in orthopaedic patients. They are challenging to treat, especially in young, highly demanding patients who do not qualify for arthroplasty. Among the many possibilities to treat osteochondral lesions presented so far, none is ideal. Because of the poor healing potential of cartilage, treatment outcomes significantly worsen with larger lesions. The treatment of large defects usually requires expensive solutions, sometimes including second-stage surgery. Using mesenchymal stem cell transplantation and cancellous bone autografts, the technique presented here for osteochondral lesion reconstruction can be effectively used to treat large osteochondral lesions in a single-stage procedure.

Although IgG1 and IgG3 have been shown to be the dominant subclasses in the acute phase of SARS-CoV-2 infection, little is known about the distribution of IgG subclasses during the recovery phase of COVID-19. The aim of the study was to analyze the profile of IgG subclasses in COVID-19 convalescent plasma donors.A total of 36 convalescent plasma donors were included in the analysis. IgG and IgG subclass levels were measured using a nephelometric assay in plasma samples obtained directly from the plasma container.Although there was no significant difference in the concentration of IgG subclasses between the study and control groups, the contribution of IgG1 to the total IgG pool between the study and control groups was statistically significant (p = 0.0478). In addition, there was a discrepancy between the total IgG and IgG sum values in the study group, exceeding 15 % in 19,4 % of samples (n = 7), while in the control group no samples with a sum/ total IgG difference 15 % were observed.The selective affinity of the IgG1 subclass for the polyclonal anti-IgG reagent may interfere with the determination of total IgG and should be considered when interpreting the results of enzyme immunoassays DATA AVAILABILITY: The data that support the findings of this study are available on request from the corresponding author.

Autohemotherapy with ozonated blood is used in the treatment of a broad spectrum of clinical disorders. Ozone demonstrates strong oxidizing properties and causes damage to cell membranes. The impact of whole-blood ozonation on the release of microparticles from blood and endothelial cells and the concentration of selected markers in the hemostatic system (APTT, PT, D-dimer, fibrinogen) were investigated. Venous blood, obtained from 19 healthy men, was split into four equal parts and treated with air, 15 µg/mL ozone, or 30 µg/mL ozone, or left untreated. The number and types of microparticles released were determined using flow cytometry on the basis of surface antigen expression: erythrocyte-derived microparticles (CD235+), platelet-derived microparticles (CD42+), leukocyte-derived microparticles (CD45+), and endothelial-derived microparticles (CD144+). The study is the first to demonstrate that ozone induces a statistically significant increase in the number of microparticles derived from blood and endothelial cells. Although statistically significant, the changes in some coagulation factors were somewhat mild and did not exceed normal values.

This study investigated the estrogen-like effects and mechanism of action most commonly used parabens: methyl- (MeP), ethyl- (EtP), propyl- (PrP) and butylparaben (BuP) in human neutrophils. Neutrophils were isolated from 50 blood donors, pre-incubated with antagonists of estrogen receptor α (ERα), ERβ and G-protein coupled estrogen receptor 1 (GPER), then incubated with MeP, EtP, PrP, BuP and 17β-estradiol (E2; 10 nM). Cytotoxic effect was evaluated by MTT test. Neutrophils apoptosis, necrosis and NETs formation were assessed in flow cytometry and confocal microscopy. The ability of the neutrophils for chemotaxis, phagocytosis, NADPH oxidase activity and generation of superoxide anion was assessed in Boyden's chamber, Park's method with latex, the NBT test, and reduction of cytochrome C, respectively. The total nitric oxide concentration was measured in neutrophils supernatants by the Griess reaction. The expression of cathepsin G, neutrophil elastase, proteinase 3, ERα, ERβ and GPER was assessed in Western blot method. In our research, parabens did not cause a cytotoxic effect on human neutrophils nor affect their lifespan. Parabens exposure did not change neutrophils functions (chemotaxis, phagocytosis, NETs formation and oxygen-dependent killing mechanism) and expression of estrogen receptors. Our results suggest that parabens do not cause estrogen receptor-mediated neutrophils-related effects at concentrations measured in the plasma of individuals using products preserved with parabens.

Patient: Female, 63-year-old Final Diagnosis: COVID-19 Symptoms: Cough • fever Medication: — Clinical Procedure: PCR • plasma exchange • serology Specialty: Infectious Diseases Objective: Unusual clinical course Background: There is no evidence-based treatment for coronavirus disease 2019 (COVID-19). We report the case of a 63-year-old woman with SARS-CoV-2 infection who developed severe COVID-19 pneumonia and was treated with convalescent plasma. Case Report: A 63-year-old woman who presented with severe and prolonged course of COVID-19 disease (fever up to 39.4°C, persistent cough, and dyspnea) received a convalescent plasma transfusion, which led to complete recovery. The diagnosis was confirmed by RT-PCR testing using the CFX96 Real-Time System (Bio-Rad, USA) from nasopharyngeal swabs. In laboratory tests, an increase in acute-phase parameters was observed. Chest computed tomography (CT) showed abnormalities typical for COVID-19. On days 9 and 11 of the disease, she received the convalescent plasma prepared from a single plasmapheresis donation from a male donor. This male donor was qualified as a convalescent plasma donor according to Polish guidelines, which are compliant with European guidelines. He donated plasma at the Regional Centre for Transfusion Medicine in Białystok, Poland. The therapy with convalescent plasma led to clinical improvement and normalization of inflammatory parameters. Conclusions: This report presents a case of severe COVID-19 pneumonia in a 63-year-old woman who was given supportive treatment with convalescent plasma. Ongoing clinical trials will determine whether convalescent plasma therapy is an effective treatment for SARS-CoV-2 infection.

BackgroundIn the present study, we aimed to investigate selected functions of human neutrophils exposed to bisphenol A (BPA) under in vitro conditions. As BPA is classified among xenoestrogens, we compared its action and effects with those of 17β-estradiol (E2).MethodsChemotaxis of neutrophils was examined using the Boyden chamber. Their phagocytosis and nicotinamide adenine dinucleotide phosphate hydrogen (NADPH) oxidase activity were assessed via Park’s method with latex beads and Park’s test with nitroblue tetrazolium. To assess the total concentration of nitric oxide (NO), the Griess reaction was utilized. Flow cytometry was used to assess the expression of cluster of differentiation (CD) antigens. The formation of neutrophil extracellular traps (NETs) was analyzed using a microscope (IN Cell Analyzer 2200 system). Expression of the investigated proteins was determined using Western blot.ResultsThe analysis of results obtained for both sexes demonstrated that after exposure to BPA, the chemotactic capacity of neutrophils was reduced. In the presence of BPA, the phagocytic activity was found to be elevated in the cells obtained from women and reduced in the cells from men. Following exposure to BPA, the percentage of neutrophils with CD14 and CD284 (TLR4) expression, as well as the percentage of cells forming NETs, was increased in the cells from both sexes. The stimulatory role of BPA and E2 in the activation of NADPH oxidase was observed only in female cells. On the other hand, no influence of E2 on the expression of CD14 and CD284, chemotaxis, phagocytosis, and the amount of NET-positive neutrophils was found for both sexes. The study further showed that BPA intensified NO production and iNOS expression in the cells of both sexes. In addition, intensified expression of all tested PI3K-Akt pathway proteins was observed in male neutrophils.ConclusionsThe study demonstrated the influence of BPA on neutrophil functions associated with locomotion and pathogen elimination, which in turn may disturb the immune response of these cells in both women and men. Analysis of the obtained data showed that the effect of this xenoestrogen on the human neutrophils was more pronounced than E2.

Hypercoagulable state and neoangiogenesis are common phenomena associated with malignancy. Cancer patients have increased levels of circulating endothelium-derived microparticles (EMPs), which have been hypothesized to be involved in numerous pathophysiological processes. Hemostasis and angiogenesis are also activated in colorectal cancer (CRC) patients. The study aimed to investigate potential influence of chemotherapy on EMPs, thrombin anti-thrombin complex (TAT) and vascular endothelial growth factor (VEGF) levels in CRC patients undergoing chemotherapy. The study group consisted of 18 CRC patients: 8 stage III colon cancer (CC) and 10 stage IV rectal cancer (RC) patients. EMPs, TAT and VEGF levels were assessed before chemotherapy and after the third course. Results were compared with 10 healthy subjects. EMP concentration was measured by flow cytometry, while TAT and VEGF concentrations were assayed employing ELISA. Compared to the control group, CC and RC patients had significantly higher levels of tissue factor (TF)-bearing and non-TF-bearing EMPs before and after three courses of chemotherapy. VEGF concentrations in CRC patients were higher than in the control groups and increased following chemotherapy. TAT levels were elevated in CRC patients before chemotherapy compared to healthy subjects and significantly increased after the third course of chemotherapy. No significant correlation was found either between EMP and TAT levels, or between EMP concentrations and VEGF levels in the study group. CRC patients have increased EMPs, and TAT as well as VEGF levels tend to increase during chemotherapy.

Background: The study objective was to assess the impact of a xenoestrogen—bisphenol A (BPA) (at the environmental concentration and 100-fold increased dose)—on selected functions of neutrophils, as well as to compare the effect of this compound and 17-β-estradiol (E2) (at the physiological concentration) on the studied functions of these cells in women and men.Methods and Results: Analysis of the results obtained for both sexes demonstrated a reduction in the chemotactic potential of neutrophils after exposure to BPA. In the presence of BPA, the phagocytic activity, as measured via the SCORE index, was found to be elevated in the neutrophils from women and reduced in those isolated for men. Exposure to BPA resulted in an increased percentage of neutrophils expressing CD14 and CD284 (TLR4), as well as an increased percentage of NET-forming cells, in the case of both sexes. In addition, a reduced capacity to release NETs was observed in cells that were preincubated with xenoestrogen and then stimulated with LPS. The stimulating role of BPA and 17-β-estradiol in the activation of NADPH oxidase, evaluated using NBT stimulation test, was evident only in the neutrophils from women. No influence of E2 could be observed on the expression of CD14 and CD284 by neutrophils, their chemotactic potential and phagocytic activity, and the amount of NETs for both sexes. The study further showed that BPA intensified the NO production and iNOS expression in the neutrophils from both sexes. In addition, an increase in expression was found for all the tested proteins of the PI3K/AKT pathway for men.Conclusions: The conducted study demonstrated that BPA influenced the functions of neutrophils associated with locomotion and pathogen elimination, which may disturb the response of these cells in both women and men. Furthermore, neutrophils isolated from women were found to be more susceptible to the effect of BPA in terms of oxygen-dependent killing, compared to those obtained from men. The variable results of the tests conducted in the presence of BPA indicated the stronger effect of xenoestrogen on human neutrophils compared to that of 17-β-estradiol.

Objectives: The growing incidence of multidrug-resistant (MDR) bacteria is an inexorable and fatal challenge in modern medicine. Colistin is a cationic polypeptide considered a “last-resort” antimicrobial for treating infections caused by MDR Gram-negative bacterial pathogens. Plasmid-borne mcr colistin resistance emerged recently, and could potentially lead to essentially untreatable infections, particularly in hospital and veterinary (livestock farming) settings. In this study, we sought to establish the molecular basis of colistin-resistance in six extraintestinal Escherichia coli strains. Methods: Molecular investigation of colistin-resistance was performed in six extraintestinal E. coli strains isolated from patients hospitalized in Medical University Hospital, Bialystok, Poland. Complete structures of bacterial chromosomes and plasmids were recovered with use of both short- and long-read sequencing technologies and Unicycler hybrid assembly. Moreover, an electrotransformation assay was performed in order to confirm IncX4 plasmid influence on colistin-resistance phenotype in clinical E. coli strains. Results: Here we report on the emergence of six mcr-1.1-producing extraintestinal E. coli isolates with a number of virulence factors. Mobile pEtN transferase-encoding gene, mcr-1.1, has been proved to be encoded within a type IV secretion system (T4SS)-containing 33.3 kbp IncX4 plasmid pMUB-MCR, next to the PAP2-like membrane-associated lipid phosphatase gene. Conclusion: IncX4 mcr-containing plasmids are reported as increasingly disseminated among E. coli isolates, making it an “epidemic” plasmid, responsible for (i) dissemination of colistin-resistance determinants between different E. coli clones, and (ii) circulation between environmental, industrial, and clinical settings. Great effort needs to be taken to avoid further dissemination of plasmid-mediated colistin resistance among clinically relevant Gram-negative bacterial pathogens.

Parabens, including the most common methylparaben (MeP), are popular preservatives, which possess estrogenic activity. The aims of this study were to assess the impact of MeP on estrogen receptors (ERs) and/or NF-κB-dependent generation of IL-8 and production of nitric oxide (NO), and also to verify the hypothesis about the crosstalk of ERs with NF-κB in xenoestrogen-exposed neutrophils. Human neutrophils were incubated for 20-h with MeP (0.06 μM) and/or ER antagonist (1 μM) and/or NF-κB inhibitor (100 μM). After the isolation of cell lysates and cytoplasmic and nuclear fraction, the expression of ERα, ERβ, p-IKKα/β, p65 NF-κB, and inducible nitric oxide synthase (iNOS) was measured by Western blot analysis, The concentration of NO was evaluated by Griess reaction, and that of IL-8 was measured by ELISA. The results showed that MeP modulated the expression of ERα, but not ERβ. Exposure to paraben activated iKKα/β-dependent NF-κB pathway, but translocation of p65 NF-κB into the cell nucleus was inhibited by ERs. MeP also decreased the iNOS-dependent production of NO, but did not influence the secretion of IL-8 by neutrophils. The study indicates that MeP may affect the functioning of human neutrophils by modulating intracellular signal transduction pathways, including ERs and NF-κB pathway.

Curcumin (CUR) is a natural compound that exhibits anti-inflammatory, anti-bacterial, and other biological properties. However, its application as an effective drug is problematic due to its poor oral bioavailability, solubility in water, and poor absorption from the gastrointestinal tract. The aim of this work is to synthesize monocarbonyl analogs of CUR based on the 9-methyl-9-azabicyclo[3.2.1]nonan-3-one (pseudopelletierine, granatanone) scaffold to improve its bioavailability. Granatane is a homologue of tropane, whose structure is present in numerous naturally occurring alkaloids, e.g., l-cocaine and l-scopolamine. In this study, ten new pseudopelletierine-derived monocarbonyl analogs of CUR were successfully synthesized and characterized by spectral methods and X-ray crystallography. Additionally, in vitro test of the cytotoxicity and anti-inflammatory properties of the synthesized compounds were performed.

In our previous study, we introduced the platelet endothelial cell adhesion molecule 1 (PECAM-1)/thrombus ratio, which is a parameter indicating the proportion of PECAM-1 in laser-induced thrombi in mice. Because PECAM-1 is an antithrombotic molecule, the higher the PECAM-1/thrombus ratio, the less activated the platelets. In this study, we used an extracorporeal model of thrombosis (flow chamber model) to verify its usefulness in the assessment of the PECAM-1/thrombus ratio in animal and human studies. Using the lipopolysaccharide (LPS)-induced inflammation model, we also evaluated whether the PECAM-1/thrombus ratio determined in the flow chamber (without endothelium) differed from that calculated in laser-induced thrombosis (with endothelium). We observed that acetylsalicylic acid (ASA) decreased the area of the thrombus while increasing the PECAM-1/thrombus ratio in healthy mice and humans in a dose-dependent manner. In LPS-treated mice, the PECAM-1/thrombus ratio decreased as the dose of ASA increased in both thrombosis models, but the direction of change in the thrombus area was inconsistent. Our study demonstrates that the PECAM-1/thrombus ratio can more accurately describe the platelet activation status than commonly used parameters such as the thrombus area, and, hence, it can be used in both human and animal studies.

Microparticles (MP-microparticles) – cell membrane vesicles 0.1–1 μm diameter, released in response to activation or apoptosis, both in physiological and pathological conditions. They reveal a wide spectrum of biological activities, express cell surface antigens characteristic for cells of their origin. In this article reviewed quantitative and qualitative methods for detection of microparticles, presents pre-analytical conditions as an potential source of variability in the analysis of MP. In conclusion there are several methods for detection of microparticles but they are not standarized. Methods of microparticles detection need to be standarized to be clinically relevant.

Neutrophils (PMN) play a key role in eliciting congenital immune response. These cells are equipped with specific receptors that are located on the surface of their cell membrane. These receptors produce various signals which in turn help in the effective functioning of PMN. The activity of these cells may be modified by factors of endo- and exogenous origin, including xenoestrogens such as bisphenol A (BPA). The aim of this study was to evaluate the effect of BPA on the expression of CD11c, CD14, CD15, CD16, CD62L and CD284 compounds on the surface of neutrophils in women and men. The study material included PMN isolated from the whole blood. The cells were incubated in the presence of BPA and/or LPS. Flow cytometry technique was used to evaluate the expression of CD antigens. Studies of these receptors indicate that BPA, at a concentration corresponding to the serum level of this compound in healthy subjects as well as at higher doses, induces changes in the immunophenotype of PMN, which may lead to immunity disorders associated with the dysfunction of these cells. Moreover, the observed effects of xenoestrogen on the expression of CD11c, CD14, CD15, CD16, CD62L and CD284 differentiation markers on these cells are sex-independent.

Background The growing incidence of MDR Gram-negative bacteria is a rapidly emerging challenge in modern medicine. Objectives We sought to establish the role of intrinsic drug-resistance regulators in combination with specific genetic mutations in 11 Enterobacter cloacae isolates obtained from a single patient within a 7 week period. Methods The molecular characterization of eight carbapenem-resistant and three carbapenem-susceptible E. cloacae ST89 isolates included expression-level analysis and WGS. Quantitative PCR included: (i) chromosomal cephalosporinase gene (ampC); (ii) membrane permeability factor genes, e.g. ompF, ompC, acrA, acrB and tolC; and (iii) intrinsic regulatory genes, e.g. ramA, ampR, rob, marA and soxS, which confer reductions in antibiotic susceptibility. Results In this study we describe the influence of the alterations in membrane permeability (ompF and ompC levels), intrinsic regulatory genes (ramA, marA, soxS) and intrinsic chromosomal cephalosporinase AmpC on reductions in carbapenem susceptibility of E. cloacae clinical isolates. Interestingly, only the first isolate possessed the acquired VIM-4 carbapenemase, which has been lost in subsequent isolates. The remaining XDR E. cloacae ST89 isolates presented complex carbapenem-resistance pathways, which included perturbations in permeability of bacterial membranes mediated by overexpression of ramA, encoding an AraC/XylS global regulator. Moreover, susceptible isolates differed significantly from other isolates in terms of marA down-regulation and soxS up-regulation. Conclusions Molecular mechanisms of resistance among carbapenem-resistant E. cloacae included production of acquired VIM-4 carbapenemase, significant alterations in membrane permeability due to increased expression of ramA, encoding an AraC/XylS global regulator, and the overproduction of chromosomal AmpC cephalosporinase.

Background/Aim: Endothelial microparticles (EMP) are small vesicles which are released from the endothelium and contribute to blood coagulation activation in various clinical settings. The aim of this study was to examine whether EMP influence blood coagulation activation in cancer patients during radiotherapy/ radiochemotherapy (RT/RCT). Materials and Methods: Sixteen head and neck cancer (HNC) patients undergoing RT/RCT and 10 controls were examined. EMP and thrombin-antithrombin complex (TAT) were measured by flow cytometry and enzyme-linked immunosorbent assay (ELISA), respectively. Tissue factor-positive EMP (TF(+)EMP) were defined as CD31(+)/CD142(+)/CD42b(–). Results: TF(+)EMP were significantly elevated in HNC patients before RT/RCT (T(0)) (1299±1154/μl), one day after RT/RCT (T(1d)) (1257±603/μl) and 3 months after RT/RCT (T(3m)) (1289±372/μl) compared to controls (688±647/μl). TF(+)EMP levels at T(0)/T(1d) and T(0), as well as at T(1d) and T(3m) were not significantly different. TAT levels at T(0) and T(1d) did not differ significantly but at T(3m) were significantly lower compared to T(0) and T(1d). TF(+)EMP and TAT concentrations were not significantly correlated at T(0) (r=0.058; p=0.828), T(1d )(r=0.373, p=0.154) and T(3m) (r=–0.302, p=0.204). Conclusion: TF(+)EMP may not contribute to hemostatic abnormalities in HNC patients.

Highlights • Pathogen reduction can be an effective method of reducing emerging infectious agents. • Assessment of the effectiveness of pathogen reduction against SARS-CoV-2 is required. • The timing of pathogen reduction should be adapted to the actual plasma demand. • Pathogen reduction performed post thawing may ensure safe plasma for transfusion. • Pathogen reduction techniques can solve the issue of plasma from one-time-only donors., In the field of transfusion medicine, many pathogen reduction techniques (PRTs) are currently available, including those based on photochemical (PI) and photodynamic inactivation (PDI). This is particularly important in the face of emerging viral pathogens that may pose a threat to blood recipients, as in the case of the COVID-19 pandemic. However, PRTs have some limitations, primarily related to their adverse effects on coagulation factors, which should be considered before their intended use. A comprehensive search of PubMed, Wiley Online Library and Science Direct databases was conducted to identify original papers. As a result, ten studies evaluating fresh plasma and frozen-thawed plasma treated with different PI/ PDI methods and evaluating concentrations of coagulation factors and natural anticoagulants both before and after photochemical treatment were included in the review. The use of PI and PDI is associated with a significant decrease in the activity of all analysed coagulation factors, while the recovery of natural anticoagulants remains at a satisfactory level, variable for individual inactivation methods. In addition, the published evidence reviewed above does not unequivocally favour the implementation of PI/PDI either before freezing or after thawing as plasma products obtained with these two approaches seem to satisfy the existing quality criteria. Based on current evidence, if implemented responsibly and in accordance with the current guidelines, both PI and PDI can ensure satisfactory plasma quality and improve its safety.

Attributable mortality due to transfusion-associated graft-versus-host disease (TA-GVHD) in contrary to the reaction occurring after transplantations has been estimated to be more than 95%. This type of reaction may occur not only in immunocompromised patients but also in patients without immune deficits. Because there is currently no available treatment for TA-GVHD and very high mortality the goal has been to reduce the likelihood of occurrence in recipients at risk of TA-GVHD. The removal of all lymphocytes from blood components is very difficult and highly impracticable. Much easier way to neutralize lymphocytes presents irreversible injury of their genetic material. It can be achieved by using ionizing irradiation (gamma- or X-ray) or pathogen reduction technologies (PRT). PRT use ultraviolet light with or without photosensitizers. Since PRT cause damage to nucleic acids, they also have potential to inactivate lymphocytes T, making them unable to proliferate, engraft and cause TA-GVHD. Transmission of infections, particularly those not routinely tested in blood components, presents still significant problem in spite of constant improvement of methods applied for donor qualification, blood testing and preparation techniques. PRT have a goal to minimize the risk of transfusion-related pathogen transmission. PRT are equally effective in TA-GVHD prophylaxis as gamma irradiation and according to current guidelines may be applied alternatively. However PRT have advantage over gamma irradiation because they additionally decrease the risk of pathogen transmission via platelet concentrate transfusion and limit proinflammatory cytokine production as well as lymphocyte activation in blood components.

BACKGROUND In radiofrequency identification (RFID) systems used in labeling of blood components, blood cells are subjected to the direct influence of electromagnetic waves throughout the storage period. The aim of this study was to prove the safety of storage of platelet concentrates (PCs) in containers labeled with RFID tags. STUDY DESIGN AND METHODS Ten pooled PCs obtained from 12 buffy coats each suspended in additive solution were divided into three separate containers that were assigned to three groups: control, PCs labeled with ultrahigh frequency (UHF) range tags and exposed to 915-MHz radio waves, and PCs labeled with high-frequency (HF) range tags and exposed to 13.56-MHz radio waves. PCs were stored at 20 to 24°C for 7 days. In vitro tests of platelet (PLT) function were performed on the first, fifth, and seventh days of storage. RESULTS There were no significant differences in pH; hypotonic shock resistance; surface expression of CD62P, CD42a, or CD63; release of PLT-derived microparticles; PLT aggregation; and number of PLTs between PCs stored at a constant exposure to radio waves of two different frequencies and the control group on the first, fifth, and seventh days of storage. CONCLUSION The results of the study indicate no impact of electromagnetic radiation generated in HF and UHF RFID systems and constant contact with the tags on the quality of stored PCs.

Background: Endothelial microparticles (EMPs) released from activated or apoptotic endothelial cells may play a role in coagulation and thrombus formation. However, there is insufficient evidence regarding the impact of EMPs on angiogenesis in patients with cancer. Materials and Methods: Sixteen patients with head and neck cancer (HNC) undergoing radiotherapy/radiochemotherapy (RT/RCT) and 10 healthy controls were studied. Serum EMPs were counted by flow cytometry, and vascular endothelial growth factor (VEGF) was measured by enzyme-linked immunosorbent assay (ELISA). Results: The mean EMP level was significantly higher in patients with HNC before RT/RCT (1,601±1,479 EMP/μl) compared to the control group (782±698 EMP/μl). The number of EMPs was not notably increased after RT/RCT (1,629±769 EMP/μl). There was no significant correlation between the plasma EMP number and concentration of VEGF before (r=0.131; p=0.625), 1 day after (r=−0.042, p=0.874), nor 3 months after RT/RCT (r=0.454, p=0.076). Conclusion: Released EMPs may not influence promotion of neovascularization in patients with HNC.

Background and objectives: Radio frequency identification (RFID) tags have advantages over bar codes in blood management system. Ultra-high frequency (UHF) RFID technology (820-960 MHz) in many ways has more advantages than high frequency (HF) RFID (13.56 MHz). The aim of our study was the evaluation of the effects of UHF RFID in comparison to HF RFID radiation on quality markers of RBC and on the oxidoreductive balance in RBC stored in containers labelled with RFID tags. Materials and methods: Ten RBC units were split into three components, one control unit-marked with bar code and two test units labeled with RFID tags - one operating on UHF and the second on HF radio waves. All units were stored at 2-6°C for 35 days. The test groups were exposed to radio waves continuously during storage. The quality parameters as well as concentration of malondialdehyde (MDA), and activities of superoxide dismutase, glutathione peroxidase and glutathione reductase were measured. Results: The degree of hemolysis and the concentration of K+ were statistically significant higher on 35th day of storage in RBC labelled with UHF RFID tags compare to control and HF RFID groups. On the 35th day of storage the concentration of MDA was statistically significant higher and the activities of oxidoreductive enzymes were statistically significant lower in the UHF group compare to control and HF groups. Conclusion: UHF RFID tags affect oxidoreductive balance in RBC and may lead to eryptosis. The approval of UHF RFID system for blood components needs further studies.

Background. Acute lymphoblastic leukemia (ALL) is a malignant disease of lymphoid progenitor cells. ALL chemotherapy is associated with numerous side effects including neutropenia that is routinely prevented by the administration of growth factors such as granulocyte colony-stimulating factor (G-CSF). To date, the effects of G-CSF treatment on the level of mobilization of different stem and progenitor cells in ALL patients subjected to clinically effective chemotherapy have not been fully elucidated. Therefore, in this study we aimed to assess the effect of administration of G-CSF to ALL patients on mobilization of other than hematopoietic stem cell (HSCs) subsets, namely, very small embryonic-like stem cells (VSELs), endothelial progenitor cells (EPCs), and different monocyte subsets. Methods. We used multicolor flow cytometry to quantitate numbers of CD34+ cells, hematopoietic stem cells (HSCs), VSELs, EPCs, and different monocyte subsets in the peripheral blood of ALL patients and normal age-matched blood donors. Results. We showed that ALL patients following chemotherapy, when compared to healthy donors, presented with significantly lower numbers of CD34+ cells, HSCs, VSELs, and CD14+ monocytes, but not EPCs. Moreover, we found that G-CSF administration induced effective mobilization of all the abovementioned progenitor and stem cell subsets with high regenerative and proangiogenic potential. Conclusion. These findings contribute to better understanding the beneficial clinical effect of G-CSF administration in ALL patients following successful chemotherapy.

Background Oxidoreductive imbalance is a major cause of excessive haemolysis in in vitro conditions. Leucocytes and blood platelets present in red blood cell concentrates (RBCs) are one of the sources of free radicals, which have a significant effect on the status of stored erythrocytes. The study objective was to assess the effect of leucoreduction on the intensity of lipid peroxidation and the activity of antioxidant barrier enzymes in RBC. Study Design and Methods Red blood cell concentrates units obtained from 10 whole-blood units were split into two equal units, one of which was leucoreduced on the day of donation. Both units were stored for 35 days. The following markers of oxidoreductive balance were measured on day 0 (donation day) and on storage days 7, 14, 21 and 35: concentration of malondialdehyde (MDA) and activities of antioxidant barrier components, that is superoxide dismutase, glutathione peroxidase and glutathione reductase. Results Lipid peroxidation in leucodepleted units (LRBC) was slower than that in non-leucodepleted ones. The analysis of LRBC revealed statistically significant decrease in concentrations of MDA. The activities of superoxide dismutase, glutathione peroxidase and glutathione reductase were higher throughout the storage period as compared to non-leucoreduced RBC. Statistically significant differences between RBC and LRBC units were noted throughout the storage in the activity of lactate dehydrogenase, and concentrations of K+ ions and free haemoglobin. Conclusions Leucoreduction of RBC before storage helps to preserve the activity of antioxidant barrier enzymes in stored RBCs and significantly improves the quality of stored red blood cell components.

The lack of evidence for the tissue-factor dependent activation of the coagulation system and the release of thrombin on one hand, and a decreased concentration of factor XII after short term air, saturated air and heliox exposures, as well as an increased concentration of the plasmin-antiplasmin complex (PAP) after short dives indicate that diving and decompression possibly affect fibrinolysis. The aim of our research was to verify the assumption that diving and decompression activate the system of fibrinolysis and the clarification of the pathomechanism of this activation. The study involved 50 healthy volunteers who were subjected to short-term, air hyperbaric exposures at 400 kPa and 700 kPa, which correspond to 30m and 60m dives. Decompression was applied in accordance with Naval tables of decompression. Before hyperbaric exposition and after decompression the following factors were determined: activity of factor XII, concentration and activity of t-PA, concentration and activity of PAI-1, concentration of alpha2- antiplasmin, concentration of PAP, concentration of neutrophil elastase. The following observations have been made: a statistically significant increase in the factor XII activity, increase in the PAP complex concentration and a simultaneous significant decline in the α2-AP activity. No measurable t-PA activity or significant changes in t-PA concentration have been observed. In addition, a statistically significant decline in both the activity and concentration of PAI-1 has been observed, which was more pronounced after the expositions that corresponded to 60 m dives. The concentrations of granulocyte elastase did not differ significantly before and after decompression. Conclusions: People qualified for diving should have the following risk factors examined: risk factors of increased fibrynolytic activity - haemostasis abnormalities that increase the risk of haemorrhage, possibility of parietal blood clots/thrombi.

Purpose Epstein–Barr virus is a common human pathogen which infects the great majority of population worldwide. A striking proliferation of CD8 + T cells is an immune response to EBV invasion of B lymphocytes during infectious mononucleosis. The aim of the study was to analyze frequencies of CD28 + CD95 − , CD28 + CD95 + , CD28 − CD95 + T cell subsets putative naive ( T N ), central ( T CM ) and effector memory ( T EM ) T cells in children with infectious mononucleosis. Material/methods Multiparameter flow cytometric analysis of CD4 + and CD8 + T cell subsets was performed in 19 children with acute infectious mononucleosis. Results The CD4 + /CD8 + ratio was found to be decreased (0.53) in children with infectious mononucleosis. Median T N , T CM , T EM frequencies were estimated to be 3.7, 4.5, 15.1% of CD8 + and 23, 59.3, 5.5% of CD4 + T cells, respectively. In the present study we demonstrated negative correlations between CD8 + CD28 + CD95 + and CD8 + CD28 − CD95 + T cells and both VCA IgM antibody titers and disease duration. However, no such correlation was found when subset of CD4 + T cells or CD8 + CD28 + CD95 − cells was compared. Conclusions We conclude that there is a rapid decrease in the number of memory CD8 + T cells in early acute stage of infectious mononucleosis.

Ozonated autohemotherapy (O3-AHT) is a medical approach during which blood obtained from the patient is ozonated and injected back into the body. Despite an increasing number of evidence that O3-AHT is safe, this type of therapy remains controversial. To extend knowledge about the changes in blood evoked by O3-AHT, LC-MS- and GC-MS-based metabolic fingerprinting was used to compare plasma samples obtained from blood before and after the treatment with potentially therapeutic concentrations of ozone. The procedure was performed in PVC bags utilized for blood storage to study also possible interactions between ozone and plastic. By use of GC-MS, an increase in lactic acid and pyruvic acid was observed, which indicated an increased rate of glycolysis. With LC-MS, changes in plasma antioxidants were observed. Moreover, concentrations of lipid oxidation products (LOP) and lysophospholipids were increased after ozone treatment. This is the first report of increased LOPs metabolites after ozonation of blood. Seven metabolites detected by LC-QTOF-MS only in ozonated samples could be considered as novel biomarkers of oxidative stress. Several plasticizers have been detected by both techniques in blood stored in PVC bags. PVC is known to be an ozone resistant material, but ozonation of blood in PVC bags stimulates leaching of plasticizers into the blood.

The aim of the study was to assess the activity of AP-1 family proteins, e.g. Fra-1, Fra-2, JunB, JunD, and FosB, engaged in the regulation of inducible nitric oxide synthase (iNOS) expression and the production of NO by neutrophils (PMN) exposed to N-nitrosodimethylamine (NDMA) xenobiotic. Isolated human PMN were incubated in the presence of NDMA. iNOS mRNA expression was then analyzed using Northern blot and the expression of other proteins in the cytoplasmic and nuclear fractions were assessed using Western blot. The obtained results indicate that NDMA increased iNOS mRNA and protein expression in human PMN. Furthermore, it increased the expression of Fra-1, Fra-2, JunB, and JunD in the cytoplasmic fraction, and FosB expression in the fractions of analyzed cells. As a consequence of inhibiting p38 pathway and JNK, reduced iNOS expression and NO production was noted in PMN exposed to NDMA. Inhibition of the p38 pathway resulted in reduced expression of all analyzed proteins in the cytoplasmic fraction of PMN exposed to NDMA. Furthermore, increased Fra-2 expression and reduced FosB expression were found in the nuclear fraction of those cells. Inhibiting ERK5 pathway resulted in increased JunB expression in both fractions of the analyzed cells. Therefore, no changes in the expression of analyzed proteins in the presence of NDMA were observed in PMN pre-incubated with JNK pathway inhibitor. In conclusion, the results here indicate a role of Fra-1, Fra-2, JunB, JunD, and FosB transcription factors in the regulation of iNOS expression and NO production by human neutrophils exposed to NDMA.

International Congress of Clinical Transfusiology, Polish National Transfusion Service September 24th 2010, Hotel Ossa, Ossa, Poland This symposiumwas a part of the International Congress of Clinical Transfusiology, which took place in September 2010 in Ossa, Poland. An overview of the Mirasol Pathogen Reduction Technology (PRT) and the results from its use in research and clinical settings were presented. Among presenters were research scientists and doctors who used PRT treated blood components in the research laboratories and routinely for the transfusion support of patients.

Background Bacterial contamination of platelet concentrates (PCs) still remains a significant problem in transfusion with potential important clinical consequences, including death. The International Society of Blood Transfusion Working Party on Transfusion-Transmitted Infectious Diseases, Subgroup on Bacteria, organised an international study on Transfusion-Relevant Bacteria References to be used as a tool for development, validation and comparison of both bacterial screening and pathogen reduction methods. Material and Methods Four Bacteria References (Staphylococcus epidermidis PEI-B-06, Streptococcus pyogenes PEI-B-20, Klebsiella pneumoniae PEI-B-08 and Escherichia coli PEI-B-19) were selected regarding their ability to proliferate to high counts in PCs and distributed anonymised to 14 laboratories in 10 countries for identification, enumeration and bacterial proliferation in PCs after low spiking (0·3 and 0·03 CFU/ml), to simulate contamination occurring during blood donation. Results Bacteria References were correctly identified in 98% of all 52 identifications. S. pyogenes and E. coli grew in PCs in 11 out of 12 laboratories, and K. pneumoniae and S. epidermidis replicated in all participating laboratories. The results of bacterial counts were very consistent between laboratories: the 95% confidence intervals were for S. epidermidis: 1·19–1·32 × 107 CFU/ml, S. pyogenes: 0·58–0·69 × 107 CFU/ml, K. pneumoniae: 18·71–20·26 × 107 CFU/ml and E. coli: 1·78–2·10 × 107 CFU/ml. Conclusion The study was undertaken as a proof of principle with the aim to demonstrate (i) the quality, stability and suitability of the bacterial strains for low-titre spiking of blood components, (ii) the property of donor-independent proliferation in PCs, and (iii) their suitability for worldwide shipping of deep frozen, blinded pathogenic bacteria. These aims were successfully fulfilled. The WHO Expert Committee Biological Standardisation has approved the adoption of these four bacteria strains as the first Repository for Transfusion-Relevant Bacteria Reference Strains and, additionally, endorsed as a project the addition of six further bacteria strain preparations suitable for control of platelet contamination as the next step of enlargement of the repository.

Ratajczak-Wrona W, Jablonska E, Garley M, Jablonski J, Radziwon P. Effect of N-nitrosodimethylamine on inducible nitric oxide synthase expression and production of nitric oxide by neutrophils and mononuclear cells: the role of JNK signalling pathway, APMIS 2011; 119: 431–41. In neutrophils (PMN) and mononuclear cells (PBMC), one of the enzymes responsible for nitric oxide (NO) synthesis is inducible nitric oxide synthase (iNOS). Changes in iNOS expression result from various signalling pathways including the mitogen-activated protein kinase (MAPK) pathway activated by endogenic and exogenic factors. N-nitrosodimethylamine (NDMA) is a xenobiotic with widespread occurrence in human environment and has an effect on cells of the immune system. The aim of this study was to determine the effect of NDMA on iNOS expression and NO production by human PMN and PBMC cells in the light of superoxide anion production by PMN cells. Moreover, the role of JNK and p38 pathways in NO production with involvement of iNOS was studied. Additionally, the function of JNK pathway in generation of superoxide anion was determined. Moreover, nitrotyrosine expression was studied in PMN and PBMC cells in the presence of NDMA. This work shows that NDMA increases iNOS expression and NO production by PMN and PBMC cells. In addition, elevated expression of phospho-JNK and phospho-p38, which are markers of JNK and p38 MAPK pathways activation, were observed. Lower iNOS expression and NO production by neutrophils exposed to extended action of NDMA were observed after application of inhibitor of JNK and p38 pathways. Lower phospho-p38 expression in PMN stimulated by NDMA was found as a result of arresting JNK pathway, whereas, application of inhibitor of p38 pathway resulted in enhanced phospho-JNK expression in PMN and PBMC cells. Increased ability to release superoxide anion by NDMA-stimulated PMN cells was observed. This ability was reduced after the application of inhibitor of JNK pathway. In PMN and PBMC cells exposed to NDMA, an increased expression of nitrotyrosine, which is dependant on JNK and p38 pathways that are activated by this particular xenobiotic, was observed. Generally, increased induction of iNOS related to elevated production of NO by PMN and PBMC cells exposed to NDMA may result in dysfunction of regulation of immunity responses controlled by this molecule in various conditions. Increased expression of nitrotyrosine in PMN and PBMC cells exposed to NDMA may affect their functions in an auto- and/or a paracrine way. Mutual interactions of JNK and p38 MAPK during the induction of iNOS expression in cells exposed to NDMA indicate complex mechanism of induction of iNOS synthase.

More and more is known about the role of venous wall abnormalities and valvular incompetence in the development of chronic venous disorders (CVD). Unfortunately detailed mechanisms of CVD pathophysiology are not well understood. Recent studies focus on involvement of the inflammatory process in the structural remodeling of venous valves and venous wall. The aim of this study is to investigate and to document the memory T cells homeostasis in CVD patients. In this study we present lymphocytic changes in blood from varicose veins in terms of total CD4+ and CD8+ T cells and their particular subsets of memory T cells: TN, TCM and TEM. Results suggest that immunological memory may be involved in the CVD development.

Purpose Short-term and saturated simulated dives followed by decompression with air, cause a decrease in platelet count and increased activation of fibrinolysis. The aim of this study was to determine whether short-term dives with trimix as a breathing mixture induce the activation of platelets, and/or fibrinolysis. Material and Methods 30 male divers were subjected to short-term hyperbaric exposures to 0.7 MPa. Thirty divers used air and then the same divers used trimix as a breathing mixture. Results: The mean platelet count dropped significantly after decompression only in the group breathing air. The number of CD62P positive platelets and the amount of platelet-derived micro particles were statistically significant higher after decompression in both exposures. The number of CD61 positive platelets increased significantly only in the group breathing air. We observed a significant decrease of factor XII and fibrinogen concentrations after decompression only in the group breathing air. A significant increase in the concentration of plasmin-antiplasmin complex in both groups was detected. Conclusions Short-term hyperbaric exposure and decompression performed according to current safety standards activates platelets and the fibrinolytic system. Trimix protects divers from a reduction in the amount of platelets, fibrinogen and factor XII in the course of these exposures.