The role of shear forces in primary and secondary nucleation of amyloid fibrils.

Shear forces affect self-assembly processes ranging from crystallization to fiber formation. Here, the effect of mild agitation on amyloid fibril formation was explored for four peptides and investigated in detail for A[Formula: see text]42, which is associated with Alzheimer's disease. To gain mechanistic insights into the effect of mild agitation, nonseeded and seeded aggregation reactions were set up at various peptide concentrations with and without an inhibitor. First, an effect on fibril fragmentation was excluded by comparing the monomer-concentration dependence of aggregation kinetics under idle and agitated conditions. Second, using a secondary nucleation inhibitor, Brichos, the agitation effect on primary nucleation was decoupled from secondary nucleation. Third, an effect on secondary nucleation was established in the absence of inhibitor. Fourth, an effect on elongation was excluded by comparing the seeding potency of fibrils formed under idle or agitated conditions. We find that both primary and secondary nucleation steps are accelerated by gentle agitation. The increased shear forces facilitate both the detachment of newly formed aggregates from catalytic surfaces and the rate at which molecules are transported in the bulk solution to encounter nucleation sites on the fibril and other surfaces. Ultrastructural evidence obtained with cryogenic transmission electron microscopy and free-flow electrophoresis in microfluidics devices imply that agitation speeds up the detachment of nucleated species from the fibril surface. Our findings shed light on the aggregation mechanism and the role of detachment for efficient secondary nucleation. The results inform on how to modulate the relative importance of different microscopic steps in drug discovery and investigations.

Aß Oligomer Dissociation Is Catalyzed by Fibril Surfaces.

Oligomeric assemblies consisting of only a few protein subunits are key species in the cytotoxicity of neurodegenerative disorders, such as Alzheimer's and Parkinson's diseases. Their lifetime in solution and abundance, governed by the balance of their sources and sinks, are thus important determinants of disease. While significant advances have been made in elucidating the processes that govern oligomer production, the mechanisms behind their dissociation are still poorly understood. Here, we use chemical kinetic modeling to determine the fate of oligomers formed in vitro and discuss the implications for their abundance in vivo. We discover that oligomeric species formed predominantly on fibril surfaces, a broad class which includes the bulk of oligomers formed by the key Alzheimer's disease-associated Aß peptides, also dissociate overwhelmingly on fibril surfaces, not in solution as had previously been assumed. We monitor this "secondary nucleation in reverse" by measuring the dissociation of Aß42 oligomers in the presence and absence of fibrils via two distinct experimental methods. Our findings imply that drugs that bind fibril surfaces to inhibit oligomer formation may also inhibit their dissociation, with important implications for rational design of therapeutic strategies for Alzheimer's and other amyloid diseases.

Discovery of potent inhibitors of α-synuclein aggregation using structure-based iterative learning.

Machine learning methods hold the promise to reduce the costs and the failure rates of conventional drug discovery pipelines. This issue is especially pressing for neurodegenerative diseases, where the development of disease-modifying drugs has been particularly challenging. To address this problem, we describe here a machine learning approach to identify small molecule inhibitors of α-synuclein aggregation, a process implicated in Parkinson's disease and other synucleinopathies. Because the proliferation of α-synuclein aggregates takes place through autocatalytic secondary nucleation, we aim to identify compounds that bind the catalytic sites on the surface of the aggregates. To achieve this goal, we use structure-based machine learning in an iterative manner to first identify and then progressively optimize secondary nucleation inhibitors. Our results demonstrate that this approach leads to the facile identification of compounds two orders of magnitude more potent than previously reported ones.

Design of amyloidogenic peptide traps.

Segments of proteins with high ß-strand propensity can self-associate to form amyloid fibrils implicated in many diseases. We describe a general approach to bind such segments in ß-strand and ß-hairpin conformations using de novo designed scaffolds that contain deep peptide-binding clefts. The designs bind their cognate peptides in vitro with nanomolar affinities. The crystal structure of a designed protein-peptide complex is close to the design model, and NMR characterization reveals how the peptide-binding cleft is protected in the apo state. We use the approach to design binders to the amyloid-forming proteins transthyretin, tau, serum amyloid A1 and amyloid ß1-42 (Aß42). The Aß binders block the assembly of Aß fibrils as effectively as the most potent of the clinically tested antibodies to date and protect cells from toxic Aß42 species.

Improving community-based COPD care in general practice in Poland - a cluster randomized controlled trial.

INTRODUCTION AND OBJECTIVE: Chronic obstructive pulmonary disease (COPD) is the third leading cause of death worldwide. The aim of the study was to evaluate the impact of intervention on exacerbations of COPD in elderly patients compared to those receiving usual care. MATERIAL AND METHODS: A 12 month, multicentre, three-arm, pragmatic, cluster randomized controlled trial was performed (CRCT). The 97 largest PHC clinics with at least 46 COPD registered patients in the Lódz Province, in central Poland. In total, 27,534 COPD patients aged 65 and over were identified from the National Health Fund (NFZ) electronic health records. A checklist of selected, recommended COPD interventions sent to GPs once or twice by post and shown on their desk in their clinics, in the intervention arms. RESULTS: A primary outcome was the difference in exacerbations or deaths between the 3 arms at 12 months. The amounts of specific short- and long-acting drugs purchased by patients were also assessed as secondary outcomes. Only 0.44% (122 of 27 534) COPD patients demonstrated exacerbations after the one-year study period. No statistically significant associations were found between interventions and exacerbations (p=0.1568, Chi-Square) or deaths (p=0.8128, Chi-Square) at 12 months. CONCLUSIONS: As this study coincided with the pandemic period, the results should be interpreted with care. The intervention had no association with exacerbations. Future research on interventions aimed at improving chronic illness care are needed.

The Hsc70 disaggregation machinery removes monomer units directly from α-synuclein fibril ends.

Molecular chaperones contribute to the maintenance of cellular protein homoeostasis through assisting de novo protein folding and preventing amyloid formation. Chaperones of the Hsp70 family can further disaggregate otherwise irreversible aggregate species such as α-synuclein fibrils, which accumulate in Parkinson's disease. However, the mechanisms and kinetics of this key functionality are only partially understood. Here, we combine microfluidic measurements with chemical kinetics to study α-synuclein disaggregation. We show that Hsc70 together with its co-chaperones DnaJB1 and Apg2 can completely reverse α-synuclein aggregation back to its soluble monomeric state. This reaction proceeds through first-order kinetics where monomer units are removed directly from the fibril ends with little contribution from intermediate fibril fragmentation steps. These findings extend our mechanistic understanding of the role of chaperones in the suppression of amyloid proliferation and in aggregate clearance, and inform on possibilities and limitations of this strategy in the development of therapeutics against synucleinopathies.

A checklist-based method for improving COPD care for the elderly in general practice: study protocol for a cluster randomized controlled trial using electronic health records.

BACKGROUND: The third most frequent chronic condition, and the fourth most common cause of death, in Poland is chronic obstructive pulmonary disease (COPD). The diagnosis and treatment of COPD is the responsibility of the general practitioner (GP); the GP also serves as gatekeeper, referring patients to the other levels of public health care system when necessary. Undertreatment of COPD can result in a greater frequency of exacerbations and hospitalizations. Elderly patients require special attention due to the increased prevalence of COPD and systemic comorbidities. However, both the occurrence of exacerbations and the quality of life of the patients may be improved by developing and implementing guidelines for practice and ensuring their adherence. This proposal concerns the development of a checklist-based educational program to assist general practitioners in managing COPD patients. METHODS: No less than eighty-four general clinics in the Lodz region, Poland (28 clusters in each of three study arms), will be identified, randomized, and included in the trial. The trial will be based on anonymized data in electronic health records within the national public health care system. The educational intervention program will consist of GPs in two intervention arms being provided with a COPD management checklist: those in the first intervention arm with receive the checklist once at the beginning, while those in the second with receive it twice. The third (control) arm receives standard care without the checklist. The study used the International Code of Diseases (ICD)-10 for COPD. The primary aim is to determine the effect of interventions delivered to general practitioners (GPs) in primary health care. These interventions are aimed at decreasing the hospitalization of elderly patients with medical code J-44 (COPD) as the main reason for hospital admission. DISCUSSION: The results of this trial will be directly applicable to primary care in Poland and add new data to the growing body of evidence regarding interventions aimed at improving chronic illness care. TRIAL REGISTRATION: This trial has been registered with the Clinical Trials Protocol Registration System. Please see in ClinicalTrial.gov identifier (NCT Number): NCT04301505 . Registered on 10 March 2020.

Anchor Effect in Polymerization Kinetics: Case of Monofunctionalized POSS.

The effect of the anchoring group on the detailed polymerization kinetics was investigated using monomethacryloxy-heptaisobutyl POSS (1M-POSS). This compound was copolymerized with lauryl methacrylate (LM) as the base monomer, at various molar ratios. The process was initiated photochemically. The polymerization kinetics were followed by photo-DSC and photorheology while the polymers were characterized by nuclear magnetic resonance (NMR), gel permeation chromatography (GPC), transmission electron microscopy (TEM), and differential scanning calorimetry (DSC). For comparison, a methacrylate containing the branched siloxy-silane group (TSM) was also studied. It was found that the modifiers with a bulky substituent have a dual effect on the termination process: (i) At low concentrations, they increase the molecular mobility by increasing the free volume fraction, which leads to an acceleration of the termination and slows the polymerization; while (ii) at higher concentrations, they retard molecular motions due to the "anchor effect" that suppresses the termination, leading to acceleration of the polymerization. The anchor effect can also be considered from a different point of view: The possibility of anchoring a monomer with a long substituent (LM) around the POSS cage, which can further enhance propagation. These conclusions were derived based on kinetic results, determination of polymerization rate coefficients, and copolymer analysis.

Origins Left, Right, and Centre: Increasing the Number of Initiation Sites in the Escherichia coli Chromosome.

The bacterium Escherichia coli contains a single circular chromosome with a defined architecture. DNA replication initiates at a single origin called oriC. Two replication forks are assembled and proceed in opposite directions until they fuse in a specialised zone opposite the origin. This termination area is flanked by polar replication fork pause sites that allow forks to enter, but not to leave. Thus, the chromosome is divided into two replichores, each replicated by a single replication fork. Recently, we analysed the replication parameters in E. coli cells, in which an ectopic origin termed oriZ was integrated in the right-hand replichore. Two major obstacles to replication were identified: (1) head-on replication⁻transcription conflicts at highly transcribed rrn operons, and (2) the replication fork trap. Here, we describe replication parameters in cells with ectopic origins, termed oriX and oriY, integrated into the left-hand replichore, and a triple origin construct with oriX integrated in the left-hand and oriZ in the right-hand replichore. Our data again highlight both replication⁻transcription conflicts and the replication fork trap as important obstacles to DNA replication, and we describe a number of spontaneous large genomic rearrangements which successfully alleviate some of the problems arising from having an additional origin in an ectopic location. However, our data reveal additional factors that impact efficient chromosome duplication, highlighting the complexity of chromosomal architecture.

Tissue and serum mRNA profile of MMPs-2/9 as a potential novel biomarker for the most individual approach in infantile hemangiomas and cancer disease.

Propranolol is a widely-known beta-blocker approved for treating infantile hemangiomas (IH). The mechanisms behind the spectacular IH involution after propranolol treatment remain unclear. Recently, there is strong evidence of overexpression of numerous angiogenic factors in IH tissues, and it is reported that propranolol influences their pathways. However, a number of MMPs studies is highly limited. Here, for the first time, we propose a comprehensive approach by analyzing the expression levels of metalloproteinases-2/9 (MMPs-2/9) and tissue metalloproteinase inhibitor-2 (TIMP-2) in vivo on both, molecular and immunohistochemical levels, and in both, IH tissues and in the serum of IH patients, and relates the obtained results to the tumor's biology and systemic propranolol treatment. MATERIAL AND METHODS: MMPs-2/9 and TIMP-2 were analyzed in 71 IH tissue samples using immunohistochemistry and real-time PCR, and in 50 serum samples of IH patients by ELISA. RESULTS: Significantly lower MMPs-2/9 and higher TIMP-2 levels were observed in IH tissues on the mRNA level as well as lower serum MMP-2 concentration among the treated individuals. CONCLUSION: MMPs-2/9 and TIMP-2 are both involved in the biology of IH and the propranolol pathways enabling their antiangiogenic properties. The most reliable method of IH examination appears to be direct MMPs-2/9 mRNA evaluation in tumor tissue; and MMP-2 evaluation in patients' serum is a valuable complement to it. Tissue and serum mRNA MMPs assessment may represent a suitable novel biomarker identifying tumor progression and involution processes with potential clinical impact in IH as well as in cancer disease.

Dataset for acrylate/silica nanoparticles formulations and photocured composites: Viscosity, filler dispersion and bulk Poisson׳s ratio.

UV-curable polymer composites are of importance in industry, biomedical applications, scientific fields, and daily life. Outstanding physical properties of polymer composites were achieved with nanoparticles as filler, primarily in enhancing mechanical strength or barrier properties. Structure-property relationships of the resulting nanocomposites are dictated by the polymer-filler molecular architecture, i.e. interactions between polymer matrix and filler, and high surface area to volume ratio of the filler particles. Among monomers, acrylates and methacrylates attracted wide attention due to their ease of polymerization and excellent physicochemical and mechanical properties of the derived polymers. We prepared and photopolymerized two series of formulations containing hydrophobized silica nanofiller (Aerosil R7200) dispersed in 2-hydroxyethyl acrylate (HEA) or polyethylene glycol diacrylate (PEGDA) monomers. We compared selected physical properties of the formulations, both before and after photocuring; specifically the viscosity of formulations and dispersion of the filler in the polymer matrices. Additionally, we estimated the bulk Poisson׳s ratio of the investigated nanocomposites. This article contains data related to the research article entitled "Nanoscale Young׳s modulus and surface morphology in photocurable polyacrylate/nanosilica composites" (Gojzewski et al., 2017) [1].

Molecular and immunohistochemical expression of apoptotic proteins Bax, Bcl-2 and Caspase 3 in infantile hemangioma tissues as an effect of propranolol treatment.

BACKGROUND: Infantile hemangiomas (IHs) are the most common benign tumors of childhood. They are characterized by a unique clinical course with two phases, proliferation and involution, which are followed by regression. The therapy of infantile hemangiomas was revolutionized in 2008 by the introduction of propranolol, however, the mechanism of its influence on hemangiomas remains unclear. METHODS: The study included 71 patients with IHs, 27 of whom were treated with propranolol while the remaining 44 were used as a comparative group. The expression of Bcl-2, Bax and Caspase3 was determined with immunohistochemistry and mRNA of Bax, Bcl-2 and Caspase3 were assessed with the use of RT-PCR. RESULTS: Both methods revealed a statistically significant decrease in Bcl-2 expression and an increase in Bax in IHs tissues after propranolol treatment. CONCLUSIONS: The results obtained for Bax and Bcl-2 proteins may indicate a link between the effect of propranolol and apoptosis. Higher Bax and lower Bcl-2 expression in the propranolol treated group indicates a strong pro- apoptotic action countering any anti-apoptotic activity; apoptosis was indicted in IH tissue as a potential result of propranolol treatment, with potential clinical impact in other tumors.

Serum and tissue profile of VEGF and its receptors VGFR1/R2 in children with infantile hemangiomas on systemic propranolol treatment.

UNLABELLED: In the last few years propranolol has revolutionized infantile hemangioma therapy. This nonselective ß bloker has been proven to be safe and effective but the molecular bases of its actions remain unclear. One of debated theories holds that propranolol may inhibit angiogenesis and induce apoptosis. To investigate this claim, this study aims to analyze the serum and tissue profiles of VEGF and VEGRR1/2 in patients treated with propranolol. MATERIALS AND METHODS: To assess the expression if VEGF and VEGRR1/2 we used three independent methods. First we analyzed serum VEGF levels in 50 children with IH before and 3 months after the therapy using ELISA test (I.). Then we used immunohistochemistry to evaluate tissue expression of VEGF and VEGFR1/2 in IH treated (n=27) and not treated (n=45) with propranolol (II.). Finally we assessed mRNA of VEGF and VEGFR1/2 in the same patients as in part II (III.). RESULTS: (I) There was no distinct decrease of VEGF level in children with IH after propranolol treatment. (II) We found no significant difference in VEGFR1 and VEGFR2 expression in hemangiomas from the study and control group. The expression of VEGF was even higher than before therapy. (III) VEGF and VEGFR1 mRNA expression was significantly lower in IH tissue after propranolol treatment compared to those without treatment. VEGFR2 demonstrated no differences in expression between the two groups. CONCLUSIONS: The obtained results show distinct discrepancies between in vitro and clinical studies as well as among different methods used for analyzing the same phenomenon. Only VEGF and VEGFR1 expression in mRNA studies may prove the proposed theory of antiangiogenic properties of propranolol. Other results do not confirm it and remain inconsistent with the fantastic clinical response to this medication.

Selected Factors of Innate Immunity in Healthy Individuals with S. aureus Nasal Carriage.

Nasal carriage of Staphylococcus aureus represents a well-defined factor of risk involving community and hospital-acquired infections. Recently a significance of several host factors has been pointed out and, in particular, of immune determinants in nasal S. aureus colonization. Therefore, this study aimed at analysis of manifestation involving manifestation in the nasal secretions of important components of the host innate immunity - human beta-defensin-2 (HBD-2), lysozyme (Ly), and interferon-gamma (IFN-γ) in healthy individuals and in persons with persistent carriage of S. aureus. The studies were conducted in two groups of healthy volunteers, encompassing non-carriers (group 1) or persistent carriers of S. aureus (group 2). Elisa assays were employed to evaluate levels of HBD-2, Ly, and IFN-γ in nasal secretions of the examined donors. In S. aureus carriers a significant variability of HBD-2 levels was detected, corresponding to, respectively, the high (averaging at 1.46 ng/ml) and the low (averaging at 0.13 ng/ml) secretory response of the defensin. The level of Ly in S. aureus carriers averaged at 1.46 µg/ml and it manifested no significant difference as compared to that noted in non-carriers. In turn, concentrations of IFN-γ in nasal secretions in the group of carriers of S. aureus amounted on the average to 81.7 pg/ml and they were 1.3-fold higher that in the group of non-carriers. The obtained results allow to conclude that IFN-γ secretion by the nasal cavity-colonizing S. aureus remains quantitatively insufficient to eliminate the pathogen. Nevertheless, a significant increase in levels of this host factor may be important for restriction of the staphylococcal colonization and protection against development of an invasive infection. In turn, the role of HBD-2 and Ly in inactivation of the colonizing S. aureus remains doubtful.

Evaluation of potential prognostic value of Bmi-1 gene product and selected markers of proliferation (Ki-67) and apoptosis (p53) in the neuroblastoma group of tumors.

INTRODUCTION: Cancer in children is a very important issue in pediatrics. The least satisfactory treatment outcome occurs among patients with clinically advanced neuroblastomas. Despite much research, the biology of this tumor still remains unclear, and new prognostic factors are sought. The Bmi-1 gene product is a currently highly investigated protein which belongs to the Polycomb group (PcG) and has been identified as a regulator of primary neural crest cells. It is believed that Bmi­1 and N-myc act together and are both involved in the pathogenesis of neuroblastoma. The aim of the study was to assess the potential prognostic value of Bmi-1 protein and its relations with mechanisms of proliferation and apoptosis in the neuroblastoma group of tumors. MATERIAL/METHODS: 29 formalin-fixed and paraffin-embedded neuroblastoma tissue sections were examined using mouse monoclonal antibodies anti-Bmi-1, anti-p53 and anti-Ki-67 according to the manufacturer's instructions. RESULTS: There were found statistically significant correlations between Bmi-1 expression and tumor histology and age of patients. CONCLUSIONS: Bmi-1 seems to be a promising marker in the neuroblastoma group of tumors whose expression correlates with widely accepted prognostic parameters. The pattern of BMI-1 expression may indicate that the examined protein is also involved in maturation processes in tumor tissue.

Prognostic significance of MCM 2 and Ki-67 in neuroblastic tumors in children.

INTRODUCTION: Neuroblastic tumors can be characterized by three features: spontaneous regression, maturation and aggressive proliferation. The most common and routinely used method of assessing tumor cell proliferation is to determine the Ki-67 index in the tumor tissue. Despite numerous studies, neuroblastoma biology is not fully understood, which makes treatment results unsatisfactory. MCM 2 is a potential prognostic factor in the neuroblastoma group. MATERIAL/METHODS: The study is based on retrospective analysis of 35 patients treated for neuroblastic tumors in the Department of Pediatric Surgery and Oncology of the Medical University of Lodz, during the period 2001-2011. The material comprised tissues of 16 tumors excised during the operation and 19 biopsy specimens. Immunohistochemical examinations were performed with immunoperoxidase using mouse monoclonal anti-MCM 2 and anti-Ki-67 antibodies. RESULTS: We observed that MCM 2 expression ranged from 2% to 98% and the Ki-67 index ranged from 0 to 95%. There was a statistically significant correlation between expression of MCM 2 and the value of the Ki-67 index and a correlation close to statistical significance between expression of MCM 2 and unfavorable histopathology. There was no statistical relationship between expression of MCM 2 and age over 1 year and N-myc amplification. DISCUSSION: The presented research shows that MCM 2 may have prognostic significance in neuroblastic pediatric tumors and as a potential prognostic factor could be the starting point of new individualized therapy.

The Role of Ultrasound Imaging of Callus Formation in the Treatment of Long Bone Fractures in Children.

BACKGROUND: In the process of diagnosis and treatment of fractures, an X-ray study is typically performed. In modern medicine very important is the development of new diagnostic methods without adverse effects on the body. One of such techniques is ultrasound imaging. It has a high value in imaging most areas of the body, including the musculoskeletal system. Reports on the use of ultrasound in the evaluation of the callus are rare and this could be a method equivalent to or even better than standard radiographs. The aim of the study was to analyze the correlation of ultrasound with radiographs in imaging of callus formation after fractures of long bones in children and to analyze the correlation of vascular resistance index (RI) and the degree of vascularization of the callus with a subjective radiological assessment of the bone union quality. MATERIAL/METHODS: The prospective study was planned to qualify 50 children treated for long bones fractures of the arm, forearm, thigh and lower leg. Ultrasound diagnosis was carried out using a Philips iU22 camera equipped with a linear probe with 17-5-MHz resolution and MSK Superficial program. During ultrasound examination measurements of the callus were performed. Using the Power Doppler callus vascularity was visualized and vascular resistance index (RI) was measured. The same measurements were made within the corresponding area of the healthy limb. The results obtained by ultrasound were compared with radiograph measurements and with the subjective assessment of the callus quality. RESULTS: Preliminary results were developed on a group of 24 patients, where 28 fractured bones and 28 corresponding healthy bones were examined. Fifteen boys and 9 girls participated in the study. The average age at injury was, respectively, 11 and 9 years. In both groups fractures without displacement were the most frequent. A similar frequency was observed in fractures requiring reposition and subperiosteal fractures. In contrast, fractures with a slight displacement of the fragments, were 3 times more common in girls. Statistical analysis of the measurements of length and width of the callus demonstrated that the differences between results obtained in the ultrasound in comparison with X-rays were not statistically significant. Moreover, preliminary results showed a significantly higher degree of vascularization of the callus than of the healthy periosteum. CONCLUSIONS: Preliminary results indicate the high efficacy of ultrasound in the evaluation of callus formation after fractures of long bones in children and the possibility of its alternative use to X-ray examinations.

Evaluation of antimicrobial resistance of Helicobacter pylori in the last 15 years in West Poland.

Increasing resistance to drugs represents a serious problem in treatment of infections with Helicobacter pylori, providing cause of frequent therapeutic failures. Present study aimed at analysis of changes in resistance of H. pylori to antibiotics in West Poland within the recent 15 years. 108 strains of H. pylori were analysed, isolated from gastric mucosa of adult patients. Group 1 involved 66 strains isolated in years of 1998/1999. Group 2 comprised 42 isolates obtained in years of 2013/2014. Susceptibility to amoxicillin (AMX), clarithromycin (CL), tetracycline (TC) and metronidazole (MTZ) was determined by E-test (AB Biodisc). All strains on both studied groups were susceptible to AMX. In group 1 all strains proved to be susceptible to TC, while 9% and 36% of tested strains were resistant to CL and MTZ, respectively. By contrast, in group 2, 31% and 83% of strains were resistant to CL and MTZ, respectively. In parallel, 14% strains were found to be resistant to TC (according to EUCAST interpretations). In West Poland, within recent 15 years a dramatic increase was noted in H. pylori strains resistant to metronidazole. In parallel, a significant increase was noted in proportion of strains resistant to clarithromycin.

[Evaluation of COX-2 protein expression in melanocytic nevi in children]. / Ocena odczynów z COX-2 w znamionach melanocytarnych u dzieci.

UNLABELLED: Melanocytic skin tumors can be divided into benign nevi and malignant which take the form of melanoma. Melanocytic nevi are common in both adults and children. It is widely acknowledged that they are one of the risk factors of the formation of melanoma. However, melanoma among children is rare. In addition, differentiation of benign and malignant melanocytic tumors is often challenging. Recent studies suggest that COX-2 protein maybe useful in excluding malignant transformation of melanocytic lesion. The aim of the study was to evaluate the occurrence and differences of reactions with COX-2 in groups of nevi in children and melanoma adults. MATERIAL AND METHODS: The study included 75 common nevi and 43 atypical nevi incised in children and 15 cases of melanoma removed in adults. Paraffin blocks were used to make a preparations with routine hematoxylin and eosin staining (H + E) and immunohistochemistry. The results were statistically analyzed. RESULTS: Positive reactions were observed in both melanocytic nevi and melanoma. Differences between benign and malignant melanocytic tumors were statistically significant. Differences within melanocytic nevi were not statistically significant. CONCLUSIONS: The reactions of COX-2 are present in all nevi and do not allow to differentiate between their various types. COX-2 has potential utility in the differentiation of benign and malignant melanocytic tumors.

Minichromosome maintenance 2 (MCM2) is a new prognostic proliferative marker in Wilms tumour.

OBJECTIVE: We examined expression of minichromosome maintenance 2 (MCM2) by immunohistochemistry in nephrectomy specimens of children with nephroblastoma treated according to the Society International d'Oncologie Pediatrique (SIOP) scheme to determine its potential prognostic significance. MATERIAL AND METHODS: 18 children with nephroblastoma, 9 females and 9 males, 2 months to 7 years of age, treated in the Department of Oncology and Paediatric Surgery, Medical University of Lodz, during the period 1994-2006 were analysed. Children were treated by neoadjuvant chemotherapy and subsequent nephrectomy according to SIOP protocols -93 and 2001 and followed up from 2 to 11 years. The tumour stage and classification in nephrectomy specimens were established according to the revised 2001 SIOP working classification of renal tumours of childhood. RESULTS: In low risk nephroblastoma MCM2 expression was low, ranging from 0% in two cases of completely necrotic nephroblastoma to 5% in one child with cystic partially differentiated nephroblastoma. In mesoblastic nephroma, which is a distinct type of neoplasm with a low malignant potential and the most common congenital renal neoplasm, MCM2 expression was variable ranging from 2-5% in 2 children with stage I disease to 70% in one child with stage III disease In intermediate risk nephroblastoma MCM2 expression was low (10%) in one case of regressive type nephroblastoma and stage II disease and intermediate to high in children with epithelial type nephroblastoma, ranging from 40-50% in one case with stage I disease to 70% and 70-100% in 2 children with stage I and stage IV disease, respectively. In high risk nephroblastoma (7 children with nephroblastoma blastemal type) MCM2 expression was intermediate to high, ranging from 40 to 90%. MCM2 expression tends to be lower in children with less advanced stage of disease (stage II) and higher in more advanced disease (stage III and IV). Two children with blastemal type and high (> 60% MCM2) died of disease within 2-4 years from diagnosis and one child was lost to follow-up. Both children who died were older 8.5 yo M and 7 yo M and presented with advanced disease stage IV or III with anaplasia. CONCLUSION: MCM2 is a promising prognostic factor in WT treated according to the SIOP scheme.