[Change of secondhand smoke levels in a public hospital in Budapest after implementation of anti-smoking policy in 2011]. / Dohányfüst-koncentráció változása egy budapesti közkórházban a nemdohányzók védelmét célzó 2011-es törvényt követoen.

INTRODUCTION: Our previous 2009 study demonstrated high secondhand smoke levels throughout a public hospital in Budapest. AIM: To compare changes in indoor air pollution level between 2009 and 2012, before and after the Hungarian anti-smoking policy legislation adopted in 2011. METHODS: TSI SidePak AM510 Personal Aerosol Monitor was used. RESULTS: In-patient care department PM2.5 levels declined by 92% from 87.7 µg/m3 to 6.9 µg/m3. Non-patient care area PM2.5 level increased by 67% from 64.8 µg/m3 to 108.0 µg/m3. The increase was driven entirely by a large increase in the level in public toilets. Excluding these, there was a 83% drop in PM2.5 in non-patient care areas from 64.8 µg/m3 to 11.1 µg/m3. CONCLUSIONS: PM2.5 decreased significantly due to the 2011 law. However, smoking still occurred in the hospital, albeit in less frequently visited areas. A stricter enforcement of this beneficial law is needed to reach a comprehensive smoke-free hospital environment.

Effects of adalimumab treatment on vascular disease associated with early rheumatoid arthritis.

BACKGROUND: Increased cardiovascular morbidity has become a leading cause of mortality in rheumatoid arthritis (RA). Tumor necrosis factor-alpha (TNFa) inhibitors may influence flow-mediated vasodilation (FMD) of the brachial artery, common carotid intima-media thickness (ccIMT) and arterial stiffness indicated by pulse-wave velocity (PWV) in RA. OBJECTIVES: To assess the effects of adalimumab treatment on FMD, ccIMT and PWV in early RA. METHODS: Eight RA patients with a disease duration < or =1 year received 40 mg adalimumab subcutaneously every 2 weeks. Ultrasound was used to assess brachial FMD and ccIMT. PWV was determined by arteriograph. These parameters were correlated with C-reactive protein, vonWillebrand factor (vWF), immunoglobulin M (IgM)-rheumatoid factor (RF), anti-CCP levels and 28-joint disease activity score (DAS28). RESULTS: Adalimumab therapy successfully ameliorated arthritis as it decreased CRP levels (P = 0.04) and DAS28 (P < 0.0001). Endothelial function (FMD) improved in comparison to baseline (P < 0.05). ccIMT decreased after 24 weeks, indicating a mean 11.9% significant improvement (P = 0.002). Adalimumab relieved arterial stiffness (PWV) after 24 weeks. Although plasma vWF levels decreased only non-significantly after 12 weeks of treatment, an inverse correlation was found between FMD and vWF (R = -0.643, P = 0.007). FMD also inversely correlated with CRP (R = -0.596, P= 0.015). CRP and vWF also correlated with each other (R = 0.598, P = 0.014). PWV and ccIMT showed a positive correlation (R = 0.735, P = 0.038). CONCLUSIONS: Treatment with adalimumab exerted favorable effects on disease activity and endothelial dysfunction. It also ameliorated carotid atherosclerosis and arterial stiffness in patients with early RA. Early adalimumab therapy may have an important role in the prevention and management of vascular comorbidity in RA.

The immunoregulatory role of vitamins A, D and E in patients with primary Sjogren's syndrome.

OBJECTIVE: The aim of the present study was to investigate the immunomodulating role of fat-soluble vitamins in 25 patients with primary SS (pSS) and 15 healthy individuals. METHODS: Plasma levels of vitamins A, D and E were determined by HPLC. Peripheral NK, NK T cells, T-cell subsets, B cells, IL-10 producing Tr1 cells, CD4(+)CD25(+) Treg cells and Th17 were determined by flow cytometry. Various Th1- and Th2-soluble cytokines were assessed by ELISA, whereas intracytoplasmic cytokines (IFN-gamma, IL-4, -10 and -17) were measured by flow cytometry. Correlation was assessed between vitamin levels and immunological and clinical parameters. RESULTS: Vitamin A levels did not differ between patients and controls, yet in patients with extraglandular manifestations (EGMs) a significant decrease in vitamin A levels was apparent compared with pSS patients without EGMs (P = 0.005). Vitamin E levels were increased in patients compared with controls (P = 0.004), whereas vitamin D levels were similar in pSS and control subjects. In patients, vitamin A showed a positive correlation with both NK cell (P = 0.038) and Th17 cell (P = 0.025), and a negative correlation with Schirmer's test values (P = 0.035). Positive correlation was found between vitamin E and NK cells (P = 0.043), Th1 cells (P = 0.049) and the Th1/Th2 ratio (P = 0.043). In the control group, we found correlation between vitamin E and serum IL-10 levels (P = 0.003). CONCLUSIONS: Our data suggest that fat-soluble vitamins may be important in immunoregulatory processes in patients with pSS.

A coagulation factor becomes useful in the study of acute leukemias: studies with blood coagulation factor XIII.

The intracellular form of the coagulation factor XIII has previously been identified by immunomorphological techniques using polyclonal antibodies. In these studies, only the A subunit (FXIII-A) was detectable in megakaryocytes/platelets and in monocytes/macrophages. We developed several novel monoclonal antibody clones directed to both subunits (FXIII-A and FXIII-B) and investigated their appearance in normal and leukemic cells. By using 3- and 4-color flow cytometry FXIII expression was investigated in normal peripheral blood and bone marrow samples and in acute myeloblastic (AML) and lymphoblastic (ALL) leukemia cases. Samples were studied by Western blotting and confocal laser scanning microscopy. With a previously published ELISA assay applying two monoclonal antibodies directed to different epitopes in FXIII-A, we were able to measure the intracytoplasmic content of FXIII-A in normal cells and leukemic blasts. FXIII-A was detectable in normal peripheral blood monocytes and in large quantities in platelets, but both cell types were negative for FXIII-B. There was no surface staining for FXIII-A, it only appeared intracellularly. In samples derived from patients with AML M4 and M5, FXIII-A sensitively identified blast cells. Although normal lymphocytes do not express FXIII-A, 40% of ALL cases showed significant FXIII-A expression as determined by flow cytometry. FXIII-A positivity of lymphoblasts was verified by Western blotting, ELISA, and confocal laser scanning microscopy cytometry. These data provide evidence that FXIII-A is a sufficiently sensitive marker in differentiating myeloblasts and monoblasts and is suitable for identifying leukemia-associated phenotypes in ALL.

[Web-based laboratory service]. / "Webalapú" laboratóriumi szolgáltatás.

UNLABELLED: The internet has revolutionized not only the clinical laboratory operation but has also made laboratory service more user friendly. With its help the laboratory can provide precise and up to date information about the services and laboratory tests to offer, furthermore can provide insight into the different operating procedures. AIM: To provide an overview of the laboratory services in an easily accessible manner, promote efficient information transfer to the clinicians, familiarize the test repertoire in different classifications, provide information on tests requiring special sampling and transportation, create a search engine for tests performed, provide in a downloadable format the different order forms and the specifications of the newly introduced tests, provide links to websites relevant to the field, familiarize with the departments educational and research activities. RESULTS: With the creation of our own website we have provided clinicians and students with a practical, easily accessible, user friendly tool that provides details of our services and activities, furthermore the number of preanalytical queries has also reduced significantly. CONCLUSIONS: Unfortunately, in the present Hungarian healthcare system few users utilize adequately the possibilities offered by the internet, however, departments like ours and others are making efforts to change this tendency.

Expression of Coagulation Factor XIII Subunit A Correlates with Outcome in Childhood Acute Lymphoblastic Leukemia.

Previously we identified B-cell lineage leukemic lymphoblasts as a new expression site for subunit A of blood coagulation factor XIII (FXIII-A). On the basis of FXIII-A expression, various subgroups of B-cell precursor acute lymphoblastic leukemia (BCP-ALL) can be identified. Fifty-five children with BCP-ALL were included in the study. Bone marrow samples were obtained by aspiration and the presence of FXIII-A was detected by flow cytometry. G-banding and fluorescent in situ hybridization was performed according to standard procedures. The 10-year event-free survival (EFS) and overall survival (OS) rate of FXIII-A-positive and FXIII-A-negative patients showed significant differences (EFS: 84% vs. 61%, respectively; p = 0.031; OS: 89% vs. 61%; p = 0.008). Of all the parameters examined, there was correlation only between FXIII-A expression and 'B-other' genetic subgroup. Further multivariate Cox regression analysis of FXIII-subtype and genetic group or 'B-other' subgroup identified the FXIII-A negative characteristic as an independent factor associated with poor outcome in BCP-ALL. We found an excellent correlation between long-term survival and the FXIII-A-positive phenotype of BCP lymphoblasts at presentation. The results presented seem to be convincing enough to suggest a possible role for FXIII-A expression in the prognostic grouping of childhood BCP-ALL patients.

Expression of coagulation factor XIII subunit A in acute promyelocytic leukemia.

Leukemic cells often express markers, which are not characteristic of their particular cell lineage. In this study, we identified the "A" subunit of coagulation factor XIII (FXIII-A) in leukemic promyelocytes in de novo AML M3 cases. The cytoplasmic presence of factor XIII-A has previously been shown only in platelets/megakaryocytes and monocytes/macrophages. Furthermore, more recently we described the presence of FXIII-A in leukemic lymphoblasts. We studied 14 patients with this rare type of acute leukemia in a period of 4 years and investigated their bone marrow samples by 3-color flow cytometry upon diagnosis, mainly focusing on FXIII-A expression of leukemic cells. We detected FXIII-A also by ELISA, Western-blot, and confocal laser scanning microscopy. This was a homogenous group of AML M3 patients with translocation t(15;17)(q22;q21) detected by fluorescence in situ hybridization (FISH). In 10 out of 14 samples, FXIII-A was detectable by flow cytometry and was coexpressed with markers characteristic for leukemic promyleocytes (CD45dim/CD13+/CD33+/CD117+/cyMPO+ and HLA-DR-/CD34-/CD14-/CD15-). Staining for the markers GPIIb and GPIX were negative, and FXIII-A was identified in the cytoplasm of the cells by confocal microscopy in a relatively high quantity, as measured by ELISA. By Western blot analysis we could identify FXIII-A in the native 82 kDa form and in cleaved forms corresponding to cleavage products observed when purified FXIII-A was treated by human neutrophil elastase. This novel expression site of FXIII-A in AML M3 can be considered as a leukemia associated immunophenotype and may have pathophysiological significance.

Classical and atypical neuroblastoma--case reports.

Neuroblastoma is the most common extracranial solid tumor in early childhood with a pluripotent neural cell origin. The disease often infiltrates the bone marrow, and these cases are candidates for flow cytometric diagnosis and disease follow-up, more so in atypical cases that are diagnostically challenging by other methods. Here, we report two neuroblastoma cases, one with characteristic neuroblastoma cells forming Homer-Wright rosettes found in the bone marrow sample by histological examination. This case showed the classical immunophenotype with CD81/CD56/CD117 positive and CD45 negative labeling and 80% bone marrow infiltration. Minimal residual disease measurement was performed in a regular fashion, and flow cytometry results showed good correlation with other disease markers. Analyzing 300,000 events provided a sensitivity level of 0.01%. The second case showed atypical cell morphology on histological examination, after which flow cytometric analysis was initiated. Atypical cells displayed CD81 and bright CD56 but CD117 negative immunophenotype, with a 28% bone marrow infiltration. In most cases the primary tumor can be identified, but in this particular case only bone marrow involvement could be detected using flow cytometry, as such making it a powerful tool for diagnosis and disease monitoring.

Vitamin D insufficiency in a large MCTD population.

OBJECTIVES: The aim of the present study was to evaluate the vitamin D status in patients with mixed connective tissue disease (MCTD) and to determine which clinical symptoms, laboratory parameters and endothelial cell markers are associated with low vitamin D levels. METHODS: 125 female MCTD patients and 48 age- and sex-matched healthy controls were enrolled in the study. The clinical symptoms, autoantibodies (anti-U1-RNP, anti-cardiolipin - anti-CL and anti-endothelial cell antibody - AECA), serum cytokines (IFN-γ, IL-6, IL-12, IL-23, IL-17 and IL-10), soluble endothelial cell markers (endothelin, thrombomodulin - TM, and von Willebrand factor antigen - vWFAg) and serum lipids (total cholesterol, triglyceride, LDL-C, HDL-C, apolipoprotein A1, and apolipoprotein B) were investigated for an association with vitamin D levels by univariate and multivariate statistical analyses. RESULTS: The mean vitamin D levels were significantly lower in MCTD patients, as compared with the control group (26.16±13.50ng/ml vs. 34.92±9.64ng/ml; p<0.001). In laboratory parameters, vitamin D levels were inversely associated with serum IL-6 (p<0.001), IL-23 (p=0.011), IL-10 (p=0.033) cytokine levels, TM (p=0.001) and endothelin (p=0.033) levels. Low vitamin D levels were also significantly associated with carotid artery intima media thickness (p<0.001), fibrinogen (p=0.010), total cholesterol (p=0.042) and ApoA1 (p=0.004) levels. Among the clinical symptoms, the cardiovascular involvement showed an inverse correlation with vitamin D status in MCTD (p<0.001). CONCLUSIONS: The prevalence of vitamin D insufficiency is high in patients with MCTD. We assume that vitamin D insufficiency along with inflammatory parameters and lipid abnormalities may provoke cardiovascular events.

Alfacalcidol treatment restores derailed immune-regulation in patients with undifferentiated connective tissue disease.

Vitamin D deficiency may contribute to pathological changes in the number and function of CD4+ T helper cell subsets (CD4+Th1, CD4+Th17, CD4+CD25(bright)Foxp3-natural regulatory T cells-nTreg) in patients with undifferentiated connective tissue disease (UCTD). The aim of the present study was to evaluate, whether alfacalcidol could restore immune-regulatory changes in patients with UCTD. We assessed the optimal dose of alfacalcidol that could normalize the elevated levels of IFN-γ expressed by the CD4+Th1 cells and the IL-17 expressed by Th17 cells. Furthermore alfacalcidol decreased the Th1 and Th17 related cytokine levels, repaired the nTreg/Th7 balance, and restored the functional activity of nTreg cells. Twenty one UCTD patients with Vitamin D deficiency (<30 ng/ml) were administered with three different daily doses of alfacalcidol. Seven patients were supplemented with 0.5 µg/day, 7 patients with 1.0 µg/day, and 7 patients with 1.5 µg/day alfacalcidol treatment during 5 weeks. Our results indicated that 1.0 µg/day alfacalcidol during 5 weeks was the optimal therapeutic regime to increase the vitamin D levels, repair the nTreg/Th17 balance and raise the capacity of nTreg cells to suppress the proliferation of autologous CD4+CD25- cells. 1.5 µg daily dose alfacalcidol was not more effective than the 1.0 µg/day treatment. In this study we described that vitamin D deficiency can contribute to the complex immune-regulatory abnormalities in patients with UCTD and vitamin D substitution therapy can improve the fine balance of pro- and anti-inflammatory processes in the disease.

Vitamin D deficiency in undifferentiated connective tissue disease.

INTRODUCTION: Both experimental and clinical data provide evidence that vitamin D is one of those important environmental factors that can increase the prevalence of certain autoimmune diseases such as systemic lupus erythematosus, rheumatoid arthritis, insulin-dependent diabetes mellitus, and inflammatory bowel disease. The aim of the present study was to investigate the prevalence of vitamin D insufficiency in patients with undifferentiated connective tissue disease (UCTD). METHODS: Plasma 25(OH)D3 levels in 161 UCTD patients were measured in both summer and winter periods. Autoantibody profiles (antinuclear antibody, anti-U1-ribonucleoprotein, anti-SSA, anti-SSB, anti-Jo1, anti-Scl70, anti-double-stranded DNA, anti-centromere, anti-cardiolipin, rheumatoid factor, and anti-cyclic citrullinated peptide) and clinical symptoms of the patients were assessed. RESULTS: Plasma levels of 25(OH)D3 in UCTD patients were significantly lower compared with controls in both summer and winter periods (UCTD summer: 33 +/- 13.4 ng/mL versus control: 39.9 +/- 11.7 ng/mL, P = 0.01; UCTD winter: 27.8 +/- 12.48 ng/mL versus control: 37.8 +/- 12.3 ng/mL, P = 0.0001). The presence of dermatological symptoms (photosensitivity, erythema, and chronic discoid rash) and pleuritis was associated with low levels of vitamin D. During the average follow-up period of 2.3 years, 35 out of 161 patients (21.7%) with UCTD further developed into well-established connective tissue disease (CTD). Patients who progressed into CTDs had lower vitamin D levels than those who remained in the UCTD stage (vitamin D levels: CTD: 14.7 +/- 6.45 ng/mL versus UCTD: 33.0 +/- 13.4 ng/mL, P = 0.0001). CONCLUSIONS: In patients with UCTD, a seasonal variance in levels of 25(OH)D3 was identified and showed that these levels were significantly lower than in controls during the corresponding seasons. Our results suggest that vitamin D deficiency in UCTD patients may play a role in the subsequent progression into well-defined CTDs.

Age dependence of serum beta-N-acetylhexosaminidase (NAG) activity.

Serum N-acetyl-beta-D-glucosaminidase (NAG; EC 3.2.1.30) is a hexosaminidase and may be a predictor of vascular injury, e.g., in infant respiratory distress syndrome, pneumonia, broncho-pulmonary dysplasia and necrotizing enterocolitis. To estimate the new diagnostic prospects we have modified our urinary NAG assay. In this sensitive colorimetric micro-assay, VRA-GlcNAc was used as a substrate. In the present study the age dependence of serum NAG activity was investigated in newborn babies, infants (1-24 months), children (2-18 years) and adults (19-80 years). Serum NAG activity was found to be age-dependent; it is higher in early childhood (11-59 U/l) but decreases to a constant value at the age of 1-2 years. After the age of 2 years it is similar to adults' NAG (10-30 U/l). In pediatrics age-matched reference ranges must be taken into consideration.