Serum neurofilament light for detecting disease activity in individual patients in multiple sclerosis: A 48-week prospective single-center study.

BACKGROUND: Serum neurofilament light (sNfL) reflects neuroaxonal damage and is now used as an outcome in treatment trials of relapsing-remitting multiple sclerosis (RRMS). However, the diagnostic properties of sNfL for monitoring disease activity in individual patients warrant further investigations. METHOD: Patients with suspected relapse and/or contrast-enhancing lesions (CELs) were consecutively included and performed magnetic resonance imaging (MRI) of the brain at baseline and weeks 28 and 48. Serum was obtained at baseline and 2, 4, 8, 16, 24, and 48 weeks. Neurofilament light concentration was measured using Single molecule array technology. RESULTS: We included 44 patients, 40 with RRMS and 4 with clinically isolated syndrome. The median sNfL level peaked at 2 weeks post-baseline (14.6 ng/L, interquartile range (IQR); 9.3-31.6) and reached nadir at 48 weeks (9.1 ng/L, IQR; 5.5-15.0), equivalent to the median sNfL of controls (9.1 ng/L, IQR; 7.4-12). A baseline Z-score of more than 1.1 (area under the curve; 0.78, p < 0.0001) had a sensitivity of 81% and specificity of 70% to detect disease activity. CONCLUSION: One out of five patients with relapse and/or CELs did not change significantly in post-baseline sNfL levels. The utility of repeated sNfL measurements to monitor disease activity is complementary rather than a substitute for clinical and MRI measures.

Measles outbreaks in 2017-2019 - molecular surveillance started in the Czech Republic.

OBJECTIVE: Between 2017 and 2019, measles virus spread globally, causing a large measles epidemic that suddenly ended in 2020. Measles outbreaks also occurred in the Czech Republic (CR) as part of the global public health problem. In the recent alarming epidemiological situation, molecular surveillance is becoming increasingly important as it plays a vital role in the identification of imported cases and in the monitoring of virus transmission. Molecular surveillance makes it possible to obtain evidence of the discontinuation of the endemic spread and is indispensable for the verification of measles elimination. The study aim is to find out whether any of measles virus genotypes circulated in the CR for more than 12 months in order to either confirm or refute the endemic spread of measles virus in the country in relation to the recent loss of the measles elimination status. Another aim is to assess the current laboratory diagnosis from the perspective of recent measles outbreaks and the obligation to refer samples for confirmation and genotyping. MATERIAL AND METHODS: In total, 243 positive nasopharyngeal swabs collected from outbreak patients from all over the CR in 2018 and 2019 were analysed by molecular methods. The most variable part of the measles virus genome, the nucleoprotein gene (N-450), was sequenced according to the WHO protocol. The sequence analysis was performed by Sanger method using the Applied Biosystems 3 500 sequencer, and sequence data were analysed by the bioinformatics programe Geneious. RESULTS: In the CR, only two genotypes were found in measles outbreaks in 2018-2019, eight variants of the dominant D8 and six B3 variants, while genotype A was detected in eight samples. The dominant genotype of 2017 (D8, 4283) was identified for the first time in the CR in January 2018. Four months later, it was replaced by genotype D8, 4683, occurring in the CR from March 2018 to June 2019. This genotype was identified in 170 of 243 samples (70%). There was a 3-month window between the first and the second detection of this genotype, which does not imply that in the meantime the virus did not circulate in the population. The analysis of seven samples from 2017 conducted by the collaborating Regional Reference Laboratory at the Robert Koch Institute (RRL RKI) in Berlin assigned five samples from Ostrava to genotype B3 and detected two variants of genotype D8 (Praha, Liberec). Laboratory diagnosis was facilitated by a higher proportion of clinical specimens available for direct detection of the virus, which increased from 18% in 2017 to 43% in 2019. Samples were referred to the National Reference Laboratory (NRL) in Prague for sequencing in accordance with the set legal rules. Between 2018 - 2019, laboratories sent 424 samples. Two hundred and forty-three samples (60%) were successfully sequenced, while the sequencing of the remaining samples failed due to low viral load. CONCLUSIONS: Measles virus sequencing was introduced in the Czech Republic as a necessary part of molecular surveillance, and almost 60% of positive samples were analysed. The sequencing analysis confirmed the endemic spread of measles virus, with genotype D8, 4683 MVs/GirSomnath.IND/42.16 found to circulate in the CR for 16 months between 2018 and 2019. Laboratory diagnosis is recently focusing more on direct detection of the virus, which along with genotyping extended to include another part of the genome will improve molecular surveillance.

Monitoring influenza virus oseltamivir resistance - our experience to date.

OBJECTIVE: The monitoring of influenza virus resistance is a routine part of influenza virus surveillance conducted by the National Reference Laboratory for Influenza and Non-Influenza Respiratory Viral Diseases (NRL/INI) at the National Institute of Public Health (NIPH). The aim is to detect neuraminidase inhibitor (oseltamivir) resistance in patients diagnosed with influenza. MATERIAL AND METHODS: A total of 326 influenza virus isolates from tissue culture were included in the study. They were obtained from inpatient and outpatient nasopharyngeal swabs which were referred to the NRL/INI during the seasons 2013/2014 to 2019/2020 and turned out to be RTPCR (reverse transcription polymerase chain reaction) positive for RNA (ribonucleic acid) of influenza virus A or B. The MDCK (Madin-Darby canine kidney) tissue culture cells were used for virus isolation from nasopharyngeal swabs. Oseltamivir resistance was tested using the NA-Star Influenza Neuraminidase Inhibitor Resistance Detection Kit (Applied Biosystems, Foster City, CA). RESULTS: Nine of 326 positive specimens were oseltamivir resistant. Resistant strains showed IC50 values 100 times as high on average as those in oseltamivir sensitive strains. CONCLUSIONS: Monitoring influenza virus resistance is helpful in controlling reasonable prescription of antivirals and thus becomes an integral part of influenza virus surveillance. Antiviral resistance monitoring is necessary not only in hospitalized patients on antivirals but also in symptomatically treated outpatients as the detection of antiviral drug resistant strains in the latter group can suggest the emergence and/or spread of antiviral drug resistance in the population.

Natalizumab, rituximab and fingolimod as escalation therapy in multiple sclerosis.

BACKGROUND AND PURPOSE: Breakthrough disease on first-line injectables in relapsing-remitting multiple sclerosis (RRMS) is a common clinical situation where comparative studies between different escalation therapies are lacking. The aim of this study was to compare the efficacy, safety and medication persistence of natalizumab (NTZ), rituximab (RTX) and fingolimod (FGL) as escalation therapy in RRMS. METHODS: Patients switching from interferon or glatiramer acetate to NTZ, RTX or FGL due to breakthrough disease were identified through the Swedish multiple sclerosis (MS) registry at four large MS centers in this retrospective observational study. Data were collected from the MS registry and medical charts. Hazard ratios (HRs) for relapses, adverse events and drug discontinuation with 95% confidence interval (CI) were calculated using multivariable confounder-adjusted Cox proportional hazard models. RESULTS: A total of 241 patients were included. The annualized relapse rates were 0.02 for NTZ, 0.03 for RTX and 0.07 for FGL. Compared with NTZ, the adjusted HR for relapse was 1.0 (95% CI, 0.2-5.6) for RTX and 3.4 (95% CI, 1.3-9.2) for FGL. The annualized drug discontinuation rates were 0.15, 0.01 and 0.15 for NTZ, RTX and FGL, respectively. The adjusted HR for drug discontinuation was 0.05 (95% CI, 0.01-0.38) for RTX and 1.0 (95% CI, 0.6-1.7) for FGL vs. NTZ. CONCLUSIONS: In patients with RRMS on interferon/glatiramer acetate with breakthrough disease, switching to NTZ or RTX was associated with less disease activity compared with FGL. RTX displayed superior medication persistence compared with both NTZ and FGL.

Current state of bioanalytical chromatography in clinical analysis.

Chromatographic methods have become popular in clinical analysis in both routine and research laboratories. The purpose of this review article is to provide an overview of the current state of chromatographic methods, i.e., high-performance liquid chromatography (HPLC), gas chromatography (GC), and supercritical fluid chromatography (SFC), in clinical analysis. The aspects related to method sensitivity, selectivity, analysis time, and throughput have been discussed in detail. Adequate solutions to improve these features have also been presented. HPLC is the most widely used method among the chromatographic methods, whereas GC is dedicated to several specific applications, and SFC is used only marginally certainly due to its only recent comeback to the analytical scene. Based on the literature search, the application fields in clinical analysis are divided into the following groups: drugs, hormones, drugs of abuse, metabolomics, lipidomics, volatile organic compounds, biomarkers and endogenous compounds, proteomics, multi-analyte approches, and others. The important features of these applications have been emphasized.

Diffusion tensor imaging in multiple sclerosis at different final outcomes.

OBJECTIVES: Methods to evaluate the relative contributions of demyelination vs axonal degeneration over the long-term course of MS are urgently needed. We used magnetic resonance diffusion tensor imaging (DTI) to estimate degrees of demyelination and axonal degeneration in the corpus callosum (CC) in cases of MS with different final outcomes. MATERIALS AND METHODS: We determined DTI measures mean diffusivity (MD), fractional anisotropy (FA), and axial (AD) and radial (RD) diffusivities in the CC of 31 MS patients, of whom 13 presented a secondary progressive course, 11 a non-progressive course, and seven a monophasic course. The study participants were survivors from an incidence cohort of 254 attack-onset MS patients with 50 years of longitudinal follow-up. As reference, we included five healthy individuals without significant morbidity. RESULTS: In patients with secondary progression, compared to all other groups, the corpus callosum showed increased RD and reduced FA, but no change in AD. None of the parameters exhibited differences among non-progressive and monophasic course groups and controls. CONCLUSION: Increased RD was observed in secondary progressive MS, indicating significant myelin loss. Normal RD values observed in the clinically isolated syndrome and non-progressive groups confirm their benign nature. AD was not a characterizing parameter for long-term outcome. Demyelination revealed by increased RD is a distinguishing trait for secondary progression.

[Enterovirus sequencing as a new approach to the laboratory diagnosis for clinical and epidemiological purposes]. / Sekvenace enteroviru jako nový prístup k laboratorní diagnostice pro klinické i epidemiologické úcely.

OBJECTIVE: Introducing enterovirus sequencing as an advanced approach to classify the viruses isolated according to the novel nomenclature and to characterize isolates in detail. MATERIAL AND METHODS: Seventy-five specimens collected from 64 patients in two hospitals, Liberec Regional Hospital, and Plzen University Hospital, were analyzed. The study patients' age ranged from four to 54 years, with a median of 15 years in males and 16 years in females. In most patients, the reasons for admission were intense headache, fever, vomiting, tiredness, meningeal symptoms, intestinal symptoms (in two patients), and skin symptoms (in one patient). The specimens collected were rectal and throat swabs, cerebrospinal fluid (CSF) and stool specimens. Molecular detection and typing were performed using the RT-PCR method. A segment of the 5´non-coding RNA was selected for typing. Specimens were amplified using single-step PCR with external primers and with the same primers extended to include M13 sequences (Generi-Biotech). The LASERGENE software (DIASTAR) was used in sequence editing, alignment, and quality check. The sequences obtained were checked against the central GenBank sequence database using the BLAST algorithm. RESULTS: The identification of the study isolates resulted in 61 ECHO viruses 30, three coxsackie viruses B1, one coxsackie virus B3, one coxsackie virus A9, one enterovirus 86, one enterovirus 71, Two ECHO viruses 13/coxsackie virus B5, one ECHO virus 7/30/coxsackie virus B4, one coxsackie virus B4/enterovirus B, one enterovirus 87/ECHO virus 30/enterovirus B, and one ECHO virus 3. All viruses isolated, except enterovirus 71 classified into group A, were of group B. CONCLUSION: The enteroviruses were identified unambigously, although the sequencing only targeted a short, conserved segment that showed considerable variability. The sequencing was an effective alternative to enterovirus identification by the neutralisation test and allowed for detailed characterization of the isolates. The predominance of ECHO 30 as the cause of aseptic meningitis is in accordance with the literature data.

The decrease in number and change in phenotype of mucosal-associated invariant T cells in the elderly and differences in men and women of reproductive age.

Mucosal-associated invariant T (MAIT) cells are innate-like T cells comprising up to 10% of the peripheral blood T cells in humans. During ontogeny, MAIT cells can first be detected in the cord blood in low amounts, but rise steadily after birth. In this population-based study, we show that their counts continue to increase, reaching maximal levels (4.5% of CD3(+) cells, 65 cells/µl) in the third and fourth decenniums. At this age, the amounts of MAIT cells exhibit the highest interindividual variability. The values then dramatically decline; subjects 80 years old and older have on average 10 times less MAIT cells, both absolutely and as a percentage among CD3(+) T cells, than subjects in fertile age. The senescence of MAIT cells is associated with decreased CD8/double negative (DN) ratio. Finally, we observed significantly higher amounts of MAIT cells in women of reproductive age than in men of the same age. Our data suggest that further studies aimed at elucidating a role of MAIT cells in human pathologies must recruit age- and gender-matched controls.

Clinically isolated syndromes with no further disease activity suggestive of multiple sclerosis at the age of population life expectancy.

The proportion of patients with clinically isolated syndrome (CIS) reported to convert to clinically definite multiple sclerosis varied between 30 and 75%. We studied the lifetime probability of remaining in the "CIS only" condition. The study was based on the longitudinally followed Gothenburg 1950-1964 incidence cohort (n = 306). Survival analysis revealed that 17.8% of 236 attack onset patients remained "CIS only". Patients with afferent (optic and sensory) symptoms had a better prognosis with approximately 30% of these patients remaining "CIS only". Patients who had experienced no relapse during the first 25 years remained "CIS only" for the subsequent 25 years of follow-up.

Monitoring of methylation changes in 9p21 region in patients with myelodysplastic syndromes and acute myeloid leukemia.

Epigenetic de novo methylation of CpG islands is an important event in malignant transformation. Two genes are frequently methylated: cyclin-dependent kinase inhibitor 2B (CDKN2B) and cyclin-dependent kinase inhibitor 2A (CDKN2A). In our study methylation of these genes was studied in 63 patients with myelodysplastic syndromes (MDS), 2 with myelodysplastic/myeloproliferative neoplasms (MDS/MPN) and 13 with acute myeloid leukemia (AML). Five patients were monitored during 5-azacytidine treatment. Twenty-six healthy donors were tested in a control group. Methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) method with all associated techniques was used for detection. Aberrant methylation was present in the CDKN2A gene in 38% and in the CDKN2B gene in 77% of the patients in MDS group. The level of methylation was higher in the group of AML patients - 77% in CDKN2A gene and 100% in CDKN2B gene. In MDS patients, an aberrant methylation was associated with a tendency to disease progression towards more advanced forms according to the World Health Organization (WHO) classification and the International Prognostic Scoring System (IPSS). Significant differences in methylation level were observed between early and advanced forms of MDS in CDKN2B gene (P value < 0.05) but not for CDKN2A gene. The trend of methylation in patients treated with azacitidine was analyzed in CDKN2B gene and correlated with the course of the disease. Increased methylation was connected with disease progression. We concluded that the methylation level of CDKN2B gene might be used as a marker of leukemic transformation in MDS. Our study indicates the role of hypermethylation as an important event in the progression of MDS to AML.

Use of ultra high performance liquid chromatography-tandem mass spectrometry to demonstrate decreased serum statin levels after extracorporeal LDL-cholesterol elimination.

BACKGROUND: Using our statin analysis method, it was possible to uncover a significant drop in statin levels (atorvastatin, simvastatin, and metabolites) after extracorporeal LDL-cholesterol elimination (EE) in severe familial hypercholesterolemia (FH). The purpose of this work was to identify the mechanism underlying this drop and its clinical significance as well as to propose measures to optimize a pharmacotherapeutical regimen that can prevent the loss of statins. METHODS: Ultra High Performance Liquid Chromatography (UHPLC) connected to the triple quadrupole MS/MS system was used. Patients. A group of long-term treated patients (3-12 years of treatment) with severe FH (12 patients) and treated regularly by LDL-apheresis (immunoadsorption) or haemorheopheresis (cascade filtration) were included in this study. RESULTS: After EE, the level of statins and their metabolites decreased (atorvastatin before/after LDL-apheresis: 8.83/3.46 nmol/l; before/after haemorheopheresis: 37.02/18.94 nmol/l). A specific loss was found (concentration of atorvastatin for LDL-apheresis/haemorheopheresis: 0.28/3.04 nmol/l in washing fluids; 11.07 nmol/l in filters). To prevent substantial loss of statin concentrations, a pharmacotherapeutic regimen with a longer time interval between the dose of statins and EE is recommended (15 hours). CONCLUSIONS: A specific loss of statins was found in adsorbent columns and filters. The decrease can be prevented by the suggested dosage scheme.

Incidence of second malignancies during treatment of chronic myeloid leukemia with tyrosine kinase inhibitors in the Czech Republic and Slovakia.

Tyrosine kinase inhibitors (TKI) have completely changed the prognosis of patients with Ph+ chronic myeloid leukemia (CML). The occurrence of a second malignancy (SM) in CML patients successfully treated with TKI may significantly affect their prognosis. In a retrospective study of 1,038 patients with CML treated at 10 centers in the Czech Republic and Slovakia between 2000 and 2009, SM was detected in 35 (3.37%) patients after TKI therapy was initiated. The median intervals from the diagnosis of CML and from the start of TKI therapy to the diagnosis of SM were 58 months (range 2 - 214) and 32 months (range 1 - 102), respectively. The observed age-standardized incidence of SM after the start of TKI therapy was 8.95 / 1,000 person-years. Comparison of the incidence of SM in CML patients with population data was performed only for patients from the Czech Republic. The age-standardized incidence rate of all malignant tumors except non-melanoma skin cancers was 6.76 (95% CI: 6.74; 6.78) / 1,000 person-years in 2000 - 2007 while the incidence rate of SM in 708 CML patients from the Czech Republic treated with TKI was 9.84 (95% CI: 6.20; 13.48) / 1,000 person-years, i.e. 1.5-fold higher, although the difference was statistically insignificant. The distribution of SM types in CML patients treated with TKI was similar to that in the age-standardized general Czech population. The median overall survival (OS) of patients treated with TKI who also developed SM (57 months) was shorter than the OS of patients treated with TKI but not suffering from SM (median OS not reached, log rank test p < 0.001. Prospective long-term population-based studies in CML patients treated with TKI as first-line therapy are needed to determine the relationship of SM to KTI therapy.

[Diagnosis and treatment of BCR/ABL-negative myeloproliferative diseases--principles and rationale of CZEMP recommendations]. / Diagnostika a lécba BCR/ABL-negativních myeloproliferativních onemocnení--principy a výchdiska doporuceni CZEMP.

In 2009, the recommendations of the Czech Collaborative Group for Ph- Myeloproliferative Diseases (CZEMP) for diagnosis and treatment of BCR/ABL-negative myeloproliferative diseases (MPD), i.e., essential thrombocythemia (ET), polycythaemia vera (PV) and primary myelofibrosis (PMF) were updated and extended. The present article gives the rationale of the recommendations in full detail. The CZEMP diagnostic criteria for ET and PMF are based on histopathological (HP) findings, which must unconditionally be in line with the given clinical and laboratory characteristics of ET or of a certain stage of PMF, respectively. The platelet count is not decisive for diagnosis. In cases lacking an adequately taken and read HP finding, the Polycythemia Vera Study Group (PVSG) criteria are recommended. The diagnosis of typical PV is based on demonstration of the V617F mutation of the JAK2 gene along with a significant increase of red cell parameters. If these are close to borderline, the demonstration of increased total red cell mass (RCM) is required. In atypical cases lacking polyglobulia or elevated RCM, the HP picture of PV (in accordance with WHO description) plus JAK2 V617F mutation is satisfactory for diagnosis, or, in cases lacking JAK2 V617F mutation, the HP picture of PV along with polyglobulia (or increased RCM) is sufficient. The treatment principles of ET and other MPDs with thrombocythemia (MPD-T; i.e., the early stages of PMF and PV) are identical. The patients are stratified by their thrombotic risk (preceding thrombosis, another thrombophilic state, jAK2 mutation), presence of disease symptoms (mainly microcirculatory), platelet count and age. Only patients up to 65 years lacking the above mentioned risks with a platelet count < 1000 x 10(9)/l are considered as low-risk and do not demand cytoreducing therapy. The others are high-risk ones and have an indication for thromboreduction. In patients older than 65 years, the potentially leukemogenic drug hydroxyurea (HU) may be used. In the younger ones, the choice is between anagrelide (ANG) or interferon-alpha (IFN). In high-risk patients, the treatment goal is to maintain platelet counts below 400, and in low-risk ones, below 600 x 10(9)/l. In PV, polycythemia itself is another thrombotic risk factor. The condition is treated by bloodletting or erythrocytaphereses. If hematocrit levels < or =45 are not achieved, cytoreductive therapy using HU in patients over 65 years, or IFN in younger individuals is required. All patients with thrombocythemia in PV are high-risk and have an indication for cytoreduction. Acetylsalicylic acid is given to all patients with MPD-T with platelets < 1000 x 10(9)/l (at higher counts, hemorrhage is imminent), and to all individuals with PV, unless contraindication is present. In case of platelet count normalization, it may be withdrawn in cases of low-risk ET or PMF, not in JAK2+ PV. The treatment of advanced stages of PMF is symptomatic, with substitution of blood derivatives being the basis. The only curative treatment is allogeneic stem cell transplantation, which should not be indicated too early seeing to its risks, but also not too late--we must not allow transition into acute leukemia, which is heralded by blasts in the blood picture. The indication is the presence of any of the following criteria: values of hemoglobin < 10 g/dl, WBC < 4 x 10(9)/l and platelets < 100 x 10(9)/l, any percentage of blasts or > or = 10% immature granulocytes in the differential picture, >1 erythroblast per 100 cells--all at repeated examinations within at least a 2-month interval, and in addition, rapid progression of hepato-/splenomegaly, presence of general symptoms of the disease, portal hypertension and extensive swellings.

Dance Intervention Impact on Brain Plasticity: A Randomized 6-Month fMRI Study in Non-expert Older Adults.

Background: Dance is a complex activity combining physical exercise with cognitive, social, and artistic stimulation. Objectives: We aimed to assess the effects of dance intervention (DI) on intra and inter-network resting-state functional connectivity (rs-FC) and its association to cognitive changes in a group of non-demented elderly participants. Methods: Participants were randomly assigned into two groups: DI and life as usual (LAU). Six-month-long DI consisted of supervised 60 min lessons three times per week. Resting-state fMRI data were processed using independent component analysis to evaluate the intra and inter-network connectivity of large-scale brain networks. Interaction between group (DI, LAU) and visit (baseline, follow-up) was assessed using ANOVA, and DI-induced changes in rs-FC were correlated with cognitive outcomes. Results: Data were analyzed in 68 participants (DI; n = 36 and LAU; n = 32). A significant behavioral effect was found in the attention domain, with Z scores increasing in the DI group and decreasing in the LAU group (p = 0.017). The DI as compared to LAU led to a significant rs-FC increase of the default mode network (DMN) and specific inter-network pairings, including insulo-opercular and right frontoparietal/frontoparietal control networks (p = 0.019 and p = 0.023), visual and language/DMN networks (p = 0.012 and p = 0.015), and cerebellar and visual/language networks (p = 0.015 and p = 0.003). The crosstalk of the insulo-opercular and right frontoparietal networks were associated with attention/executive domain Z-scores (R = 0.401, p = 0.015, and R = 0.412, p = 0.012). Conclusion: The DI led to intervention-specific complex brain plasticity changes that were of cognitive relevance.

Dasatinib in imatinib-resistant or -intolerant CML patients: data from the clinical practice of 6 hematological centers in the Czech Republic.

Dasatinib is effective second line treatment for patients with chronic myeloid leukemia (CML) resistant or intolerant to imatinib. We report here the first experiences with dasatinib therapy in 71 CML patients resistant or intolerant to imatinib from the real clinical practice of 6 hematological centers in the Czech Republic. Dose 100 mg daily and 70 mg twice daily was administered to patients with chronic phase (CP) and advanced phases (AP) CML. In chronic phase (n=46), complete hematological reponse (CHR) was achieved in 97%, major cytogenetic reponse (MCgR) in 77% and complete cytogenetic response (CCgR) in 67%. Major molecular reponse (MMR) was achieved in 19/31 patients in median of 10 months. In advanced phase (n=25), CHR was attained in 77%, MCgR in 39%, CCgR in 33% and MMR in 2/18 patients. Eleven different baseline mutations were followed up in 15 patients. Dasatinib eliminated mutations in most of the patients, but 3 patients acquired a new one. Novel mutations were detected under dasatinib therapy in 2 patients. Dasatinib was well tolerated, cytopenias were common and was managed by dose modification. The estimated progression free survival (PFS) at 12 months was 97+/-3% in CP and 62+/-21% in AP. The median time to treatment failure was 605 days in AP while it was not reached in CP patients. Our clinical experiences, described here, confirmed that dasatinib is associated with high response rates especially in imatinib resistant or intolerant CML patients in chronic phase.

NFL and CXCL13 may reveal disease activity in clinically and radiologically stable MS.

BACKGROUND: Cerebrospinal fluid (CSF) levels of neurofilament light (NFL), a biomarker of axonal damage, and CXCL13, a chemokine involved in B-cell regulation, are both associated with disease activity in multiple sclerosis (MS). OBJECTIVE: To explore the potential of NFL and CXCL13 to detect residual disease activity in patients with no signs of clinical or ongoing radiological activity and to study the clinical relevance of such activity. METHODS: NFL and CXCL13 concentrations were determined with ELISA in CSF obtained from 90 relapsing-remitting (RR) MS and 47 Progressive (Pr) MS (including primary and secondary PrMS) at baseline and after 12 months of follow-up. The patients were assessed at baseline, before initiating or switching disease modifying therapy (DMT) and again after 12 and 27 months of follow-up. RESULTS: All patients with ongoing disease activity (relapse or contrast-enhancing lesions on MRI) had increased NFL or CXCL13. The proportion of RRMS and PrMS patients without ongoing disease activity with elevation of either NFL or CXCL13 (residual disease activity) was 39% and 50%, respectively, and both were increased in 11% and 16%, respectively. The treatment with DMTs decreased the proportion with residual disease activity in both RRMS and PrMS significantly. We could not show any significant association between residual disease activity and clinical or MRI measures at 12 or 27 months of follow-up. CONCLUSIONS: Although most of this real-world study population had been treated with second-line DMTs and achieved clinical and radiological stability, a significant proportion of patients still displayed increased CSF levels of both NFL and CXCL13, indicating residual disease activity. Thus, these markers seemed considerably more sensitive to disease activity than clinical and MRI measures. However, the long-term clinical significance of such activity remains to be determined.

[Pre-operative care for cardiac surgery patients with cold antibody disorder, cryoglobulinaemia and cryofibrinogenemia]. / Péce a pacienty s nemocí chladovych protilátek, kryoglobulinemií a kryofibrinogenemií pred kardiochirurgickými zákroky.

We present an example of a patient with confirmed cold agglutinin disease who underwent cardiac surgery in hypothermia to illustrate a known fact that, when exposed to cold, cold agglutinins induce haemolysis of erythrocytes and that cryoglobulins and cryofibrinogens may, upon exposition to cold during a surgery under hypothermia, precipitate or gelify and thus increase plasma viscosity and damage microcirculation. Detailed immunological and haematological investigations in all patients awaiting cardiac surgery with a risk of developing hypothermia is not advantageous considering the low number of patients with clinical and laboratory signs of cold agglutinin disease, autoimmune haemolytic anaemia or paroxysmal cold haemoglobinuria and considering that these investigations, in addition, might not detect cryoglobulinaemia and cryofibrinogenemia. Identification of in-risk patients from the warning signs in the medical history, physical or basal laboratory testing who would subsequently undergo confirmatory investigations to verify the presence of these entities and define them accurately might be a potential solution to this clinical issue. Cardiac surgery strategy and peri-operative care should be tailored to the results of these investigations. Well-structured, practiced and functional cooperation between clinicians and laboratory personnel is a prerequisite for success in these circumstances.

[The results of patients with essentials thrombocythemia and other myeloproliferation-related thrombocythemia--a report of patients treated with Thromboreductin]. / Výsledky lécby nemocných s esentiální trombocytemií a dalsími myeloproliferacemi provázenými trombocytemií--zpráva z registru pacientu lécených Thromboreductinem.

The registry of patients treated with Thromboreductin (anagrelide) in the Czech Republic contains data concerning patients that have been treated using this drug since 2004. As of June 2009, the total number of patients was 549. The current analysis focused mainly on evaluation of anagrelide dosage needed to achieve a complete response in high-risk patients: reduction in platelet count to below 400 x 10(9)/l, which was also considered as reaching the therapeutic goal. The outcomes of the registry confirm that although anagrelide (Thromboreductin) is a very effective platelet-reducing agent, the administration of which is related to a low incidence of adverse effects and complications, the therapeutic goal is not achieved in all cases and or despite a quick treatment response, the therapeutic goal is achieved more slowly.

Comparison of a novel ultra-performance liquid chromatographic method for determination of retinol and alpha-tocopherol in human serum with conventional HPLC using monolithic and particulate columns.

Retinol and alpha-tocopherol are biologically active compounds often monitored in blood samples because of their evident importance in human metabolism. In this study a novel ultra-performance liquid chromatographic (UPLC) method used for determination of both vitamins in human serum has been compared with conventional HPLC with particulate and monolithic C(18) columns. In UPLC a sub-two-micron particle-hybrid C(18) stationary phase was used for separation, in contrast with a five-micron-particle packed column and a monolithic column with a highly porous structure. Methanol, at flow rates of 0.48, 1.5, and 2.5 mL min(-1), respectively, was used as mobile phase for isocratic elution of the compounds in the three methods. Detection was performed at 325 nm and 290 nm, the absorption maxima of retinol and alpha-tocopherol, respectively. Analysis time, sensitivity, mobile-phase consumption, validation data, and cost were critically compared for these different chromatographic systems. Although cost and mobile-phase consumption seem to make UPLC the method of choice, use of the monolithic column resulted in almost the same separation and performance with a slightly shorter analysis time. These methods are alternatives and, in routine laboratory practice, more economical means of analysis of large numbers of biological samples than use of a traditional particulate column.

HPLC determination of chlorhexidine gluconate and p-chloroaniline in topical ointment.

A novel fast isocratic reversed-phase HPLC method for simultaneous determination of chlorhexidine and its degradation product p-chloroaniline was developed. Zorbax SB Phenyl column (75 mm x 4.6 mm, 3.5 microm) was used for the separation. Mobile phase composed of acetonitrile and buffer solution of 0.08 M sodium phosphate monobasic containing 5 ml of triethylamine (0.5%) and adjust with 85% phosphoric acid to pH 3.0 in ratio 35:65 (v/v) pumped isocratically at flow rate 0.6 ml min(-1) was used. UV detection was performed at 239 nm, the total analysis time was about 10 min. The method is suitable for practical routine analysis of topical ointment in the quality control laboratory.