Ceruloplasmin expression in renal cell carcinoma correlates with higher-grade and shortened survival.

Aim: We aimed to explore ceruloplasmin (CP) expression in clear cell renal cell carcinoma (ccRCC). Materials & methods: CP was analyzed in biofluid samples of 63 ccRCC patients, divided into three grading groups, and immunohistochemically, in 308 ccRCC. Results: Significant differences of mean plasma and urine CP levels in different grading groups were found. CP immunoreactivity was significantly linked to high-grade disease. Log rank tests showed a significant shorter overall survival rate in CP-positive cases (all p < 0.05). Conclusion: CP protein levels in biofluid samples confirmed differential CP expressions, depending on nuclear grade in ccRCC as previously seen in RNA expression analysis. CP expression was linked to high-grade disease and reduced survival rate in RCC.

Transient Howell-Jolly-body-like cytoplasmic inclusions in neutrophils after severe intoxication with alpha-lipoic acid.

After intravenous supplementation of an unintentionally high dose of the antioxidant alpha-lipoic acid (ALA), a 53-year-old female complained of myalgia, chills and nausea, and showed signs of haemorrhagic diathesis. The laboratory findings were excessive hyperferritinemia, leukoerythroblastosis, severe thrombocytopenia, elevated liver enzymes and impaired coagulation. The toxicological tests resulted in an ALA serum concentration of 10 280 µg/L. The peripheral blood film of the patient showed some neutrophil dysplasia with unusual small dark-blue stained round cytoplasmic inclusions resembling 'Howell-Jolly-body-like' (HJBL) cytoplasmic inclusions, aptly named due to the morphologic similarity to their erythrocytic counterparts. Such HJBL inclusions are occasionally associated with acquired immunodeficiency, or immunosuppressive or cytostatic treatment. An association with ALA intoxication has not been described before. There are only a few reports on unintentional, harmful and lethal intoxications with ALA. The underlying molecular background of its toxicity on liver function or haematopoiesis is not yet known in detail, but ALA seems to interact with enzyme functions, e.g. with mitochondrial enzyme-complexes, possibly due to its pro-oxidant potential at high doses.

Inhibition of agonist-induced platelet aggregation by magnesium sulfate warrants its use as an alternative in vitro anticoagulant in pseudothrombocytopenia.

MgSO4 is effective in preventing spontaneous in vitro platelet agglutination in anticoagulant-induced pseudothrombocytopenia (PTCP). In order to learn more about its potential as an in vitro anticoagulant, platelets from MgSO4-anticoagulated blood were stimulated by several differentially-acting agonists (ADP, ARA, TRAP, epinephrine, collagen and ristocetin). Platelet aggregation in blood samples from 11 and 17 volunteers was measured by light-transmission aggregometry (LTA) according to Born and impedance aggregometry (MultiplateTM), respectively. Agonist-induced platelet aggregation was markedly lower in MgSO4-anticoagulated samples when compared with citrate-anticoagulated samples (decrease of 95.75% (ristocetin), 69.02% (collagen) and 75.73% (epinephrine)) or hirudin-anticoagulated samples (decrease of 85.99% (ADP), 80.98% (ARA), 77.24% (ristocetin), 54.37% (collagen) and 50.14% (TRAP)). The anti-aggregatory effect of MgSO4 is dose-dependent and readily detectable at a concentration of 7.5 mmol/l. Analysis of the agonist signaling pathways suggest that MgSO4 interferes with the final step of platelet aggregation, namely the intracellular mobilization of Ca2+.

Magnesium Sulfate as an Alternative In Vitro Anticoagulant for the Measurement of Platelet Parameters?

OBJECTIVES: There are conflicting reports on the reliable measurement of platelet count and mean platelet volume (MPV) using EDTA or citrate. The anticoagulant properties of magnesium sulfate (MgSO4) are known from the literature. The aim of this study was to evaluate MgSO4 as an in vitro anticoagulant for platelet count, MPV, platelet distribution width, and platelet activation. METHODS: Whole blood from volunteers was anticoagulated by EDTA, citrate, or MgSO4 Platelets were counted by the XE 5000 (Sysmex, Norderstedt, Germany) impedance and fluorescence optical technique. RESULTS: The mean impedance platelet counts were 227.7, 197.0, and 201.1 × 10(9)/L in EDTA-, citrate-, or MgSO4-anticoagulated blood, respectively. The counts were 4.7% higher (EDTA) after 3 hours of storage but 4% lower in citrate-anticoagulated blood. The counts in magnesium samples remained stable. The MPV was 10.4 fL (EDTA), 9.5 fL (citrate), and 9.3 fL (MgSO4). EDTA samples showed cell swelling within the first 3 hours. This was lower in citrate and only marginal in magnesium samples. High activation of platelets was observed only in EDTA samples. CONCLUSIONS: Magnesium anticoagulation might be advantageous for more reliable MPV measurements. Although platelet count is underestimated when the impedance method is used, the platelet count reveals similar results when measured by the fluorescent optical method.

Performance of the XN-2000 WPC channel-flagging to differentiate reactive and neoplastic leukocytosis.

BACKGROUND: The distinction between reactive and neoplastic leukocytes, especially atypical lymphocytes suspected to be reactive or neoplastic, is a particular challenge in automated hematological cell differentiation. The aim of the study was to evaluate the performance of the XN analyzer supplemented with the WPC channel for differentiating between reactive and neoplastic leukocytosis. METHODS: Blood samples of 253 patients with viral infections, lymphoma or leukemia were analyzed by the Sysmex XN-2000 analyzer equipped with the WPC channel. The results were compared to routine leukocyte differentiation using the routine Sysmex XE-2100 analyzer and automated digital microscopy (DM96). The combined information from standard morphology, immune phenotyping and clinical diagnosis served as a reference. RESULTS: The XN WPC channel demonstrated an excellent performance for differentiating neoplastic (AUC=0.933) and reactive leukocytosis (AUC=0.900) as compared to morphological smear examination (AUC=0.949 and AUC=0.968, respectively) or to the differentiation results of our routine hematology analyzer (AUC=0.630 and AUC=0.635, respectively). CONCLUSIONS: Our data show that the combined WDF/WPC of the Sysmex XN-Series analyzer is advantageous in the automated differentiation of neoplastic and reactive leukocytosis, thus supporting the correct diagnostic decision in the daily laboratory routine.

Measurement of Platelet Counts and Volume Using Magnesium Sulfate as an Anticoagulant: Comparison of Impedance and Light-Scatter Technology.

OBJECTIVES: Magnesium sulfate (MgSO 4 ) was recently reported as an alternative in vitro anticoagulant in pseudo-thrombocytopenia. Its suitability as an anticoagulant for the determination of reliable platelet parameters is the subject of this study. METHODS: Platelet count and mean platelet volume were measured in blood samples anticoagulated with EDTA and MgSO 4 and compared. The platelet parameters were determined by impedance (XE 5000 [Sysmex, Norderstedt, Germany]; DxH 800 [Beckman-Coulter, Krefeld, Germany]) and laser light-scatter technology (Advia 120 [Siemens Healthcare Diagnostics, Eschborn, Germany]). RESULTS: MgSO 4 anticoagulation underestimated platelet counts compared with EDTA. Mean platelet volume (MPV) in magnesium-anticoagulated blood was lower when measured by impedance but higher when light-scatter technology was used. Storage of the differently anticoagulated blood led to differently lower platelet counts after 24 hours, independent of the anticoagulant. In EDTA blood, the mean platelet volume increased moderately when measured by impedance but markedly when measured by laser light scatter. In MgSO 4 -anticoagulated blood, the MPV increase was negligible. CONCLUSIONS: Impedance technology and magnesium anticoagulation might be advantageous for standardizing MPV measurements, although the mean platelet count is slightly underestimated by both technologies.

Exclusion of thrombocytopenia as a contraindication for invasive radiofrequency ablation in a patient with paroxysmal atrial fibrillation by using magnesium anticoagulation instead of EDTA: another case of anticoagulant-induced pseudo-thrombocytopenia.

Thrombocytopenia might be an exclusion criterion for invasive radiofrequency catheter ablation; therefore it is necessary to differentiate between pseudo-thrombocytopenia and a low platelet count due to other etiologies.A 69-year-old female presented to the cardiology department with recurrent atrial fibrillation that was resistant to conventional drug treatment. The initial laboratory findings were within the normal ranges, except for low platelet counts that occurred without a specific bleeding history. The reason for thrombocytopenia was anticoagulant-induced in vitro aggregation of platelets in the presence of EDTA as well as in citrated blood samples. As recently communicated, magnesium anticoagulated blood samples prevent platelet aggregation in individuals with anticoagulant-associated pseudo-thrombocytopenia. Although its aggregation-inhibiting effect is known from previous clinical observations, magnesium sulphate has not been introduced as an anticoagulant in analytical medicine.Based on our observations, blood anticoagulated with magnesium sulphate is recommended to verify low routine platelet counts before final clinical decisions are made.

Effective estimation of correct platelet counts in pseudothrombocytopenia using an alternative anticoagulant based on magnesium salt.

Pseudothrombocytopenia remains a challenge in the haematological laboratory. The pre-analytical problem that platelets tend to easily aggregate in vitro, giving rise to lower platelet counts, has been known since ethylenediamine-tetra acetic acid EDTA and automated platelet counting procedures were introduced in the haematological laboratory. Different approaches to avoid the time and temperature dependent in vitro aggregation of platelets in the presence of EDTA were tested, but none of them proved optimal for routine purposes. Patients with unexpectedly low platelet counts or flagged for suspected aggregates, were selected and smears were examined for platelet aggregates. In these cases patients were asked to consent to the drawing of an additional sample of blood anti-coagulated with a magnesium additive. Magnesium was used in the beginning of the last century as anticoagulant for microscopic platelet counts. Using this approach, we documented 44 patients with pseudothrombocytopenia. In all cases, platelet counts were markedly higher in samples anti-coagulated with the magnesium containing anticoagulant when compared to EDTA-anticoagulated blood samples. We conclude that in patients with known or suspected pseudothrombocytopenia the magnesium-anticoagulant blood samples may be recommended for platelet counting.

Role of lipid apheresis in changing times.

During the last decades, LDL-apheresis was established as an extracorporeal treatment option for patients with severe heterozygous or homozygous familial hypercholesterolemia (FH) that is resistant to conventional treatment strategies such as diet, drugs, and changes in lifestyle. Nearly half a century ago, the first LDL-apheresis treatment was performed by plasma exchange in a child with homozygous FH. At the beginning of the 1970s, the clinical advantage of regular extracorporeal LDL-elimination was demonstrated in siblings suffering from homozygous FH. These findings encouraged researchers especially from Germany and Japan to develop extracorporeal devices to selectively eliminate LDL-cholesterol in the 1980s. Although the selectivity of the currently available LDL-apheresis devices is different, the efficacy of LDL-elimination during a single treatment is rather similar and ranges between 55 and 65 % of the pretreatment LDL plasma concentration.In the 1990s, the patients regularly treated by extracorporeal LDL-elimination, diet, and drugs were included in regression studies assessed by angiography. It was shown that the combined treatment with LDL-apheresis, diet, and drugs resulted in less progression of coronary lesions than drugs and/or diet alone. However, although a tendency was evident, results did not reach criteria for significance. During the last decade, apheresis registries were established to collect data on efficiency, safety, and clinical outcome of regular long-term LDL-apheresis. The evaluation of registry data will certainly permit further insights in the therapeutic benefit of this expensive and time-consuming therapeutic approach. Furthermore, the future of LDL-apheresis will depend upon the availability of highly efficient new drugs and molecular genetic approaches such as RNA silencing of the apoB gene, whereas the liver transplantation and gene therapy of the LDL-receptor deficiency will not replace LDL-apheresis in severe familial hypercholesterolemia in the near future.

Health care consequences of demographic changes in Mecklenburg-West Pomerania: projected case numbers for age-related diseases up to the year 2020, based on the Study of Health in Pomerania (SHIP).

BACKGROUND: The population in the German federal state of Mecklenburg-West Pomerania is growing older. A resulting rise in age-related diseases will likely lead to a greater need for medical care, even though the population as a whole is declining. The predicted number of patients affected by these diseases varies from one district to another because of local differences in demographic trends. METHODS: Case numbers were forecasted on the basis of representative data on the morbidity from chronic diseases, which were derived from the Study of Health in Pomerania (SHIP), the conjoint cancer registry of the East German federal states (GKR), and a study on dementia morbidity. These data were combined with demographic prognoses for Mecklenburg-West Pomerania and its rural and urban districts up to the year 2020. RESULTS: The largest increases in case numbers are predicted for dementia (+91.1%), myocardial infarction (+28.3%), diabetes mellitus (+21.4%), and incident colon carcinoma (+31.0%; all figures are expressed in relation to the year 2005 as a baseline). The predicted changes in case numbers vary widely from one district to another. CONCLUSION: All of the German federal states located in the former East Germany are likely to experience similar developments to those predicted for Mecklenburg-West Pomerania, as will many rural areas of the former West Germany, in which a demographic transition is already evident. Because of the predicted rise in the number of patients, new health care concepts will have to be rapidly developed, implemented, and evaluated in order to ensure that comprehensive medical care will be delivered where it is needed.

The European Register of Specialists in Clinical Chemistry and Laboratory Medicine: guide to the Register, version 3-2010.

In 1997, the European Communities Confederation of Clinical Chemistry and Laboratory Medicine (EC4) set up a Register for European Specialists in Clinical Chemistry and Laboratory Medicine. The operation of the Register is undertaken by a Register Commission (EC4RC). During the last 12 years, more than 2200 specialists in Clinical Chemistry and Laboratory Medicine have joined the Register. In 2007, EC4 merged with the Forum of European Societies of Clinical Chemistry and Laboratory Medicine (FESCC) to form the European Federation of Clinical Chemistry and Laboratory Medicine (EFCC). Two previous Guides to the Register have been published, one in 1997 and another in 2003. The third version of the Guide is presented in this article and is based on the experience gained and development of the profession since the last revision. Registration is valid for 5 years and the procedure and criteria for re-registration are presented as an Appendix at the end of the article.

The European Register of Specialists in Clinical Chemistry and Laboratory Medicine: Code of Conduct, Version 2--2008.

In 1997, the European Communities Confederation of Clinical Chemistry and Laboratory Medicine (EC4) set up a Register for European Specialists in Clinical Chemistry and Laboratory Medicine. The operation of the Register is undertaken by a Register Commission (EC4RC). During the last 10 years, more than 2000 specialists in Clinical Chemistry and Laboratory Medicine have joined the Register. In 2007, EC4 merged with the Federation of European Societies of Clinical Chemistry and Laboratory Medicine (FESCC) to form the European Federation of Clinical Chemistry and Laboratory Medicine (EFCC). A Code of Conduct was adopted in 2003 and a revised and updated version, taking account particularly of the guidelines of the Conseil Européen des Professions Libérales (CEPLIS) of which EFCC is a member, is presented in this article. The revised version was approved by the EC4 Register Commission and by the EFCC Executive Board in Paris on 6 November, 2008.

Renal Insufficiency Following Radiocontrast Exposure Trial (REINFORCE): a randomized comparison of sodium bicarbonate versus sodium chloride hydration for the prevention of contrast-induced nephropathy.

INTRODUCTION: For the prevention of contrast-induced nephropathy (CIN) after coronary angiography, hydration by 0.9% sodium chloride solution and N-acetylcysteine is currently recommended. However, it is unclear whether volume supplementation with sodium bicarbonate is better than with sodium chloride when used in conjunction with nonionic, low-osmolar iopamidol. The aim of this study was to analyze and compare the effects of sodium bicarbonate and sodium chloride on renal function in 145 patients exposed to nonionic iso-osmolar contrast medium iodixanol in a randomized study. PATIENTS AND METHODS: Renal Insufficiency Following Radiocontrast Exposure is a prospective, randomized, single-center, double-blinded trial of 145 patients (age 72.6+/-6.7 years) with elevated baseline serum creatinine levels (mean 132.6+/-29.3 micromol/l). Eligible patients were randomized to either a 154 mEq/l infusion of sodium bicarbonate (n=71, group I) or sodium chloride 0.9% solution (n=74, group II). The primary endpoint was serum creatinine elevation beyond 25% or 44 micromol/l on the first or second day following exposure to the contrast medium. Serum creatinine, serum cystatin C, plasma viscosity, urinary enzymes alanine aminopeptidase and N-acetyl-beta-D-glucosaminidase, and alpha1-microglobulin were measured at baseline and on days 1 and 2 after contrast medium administration. RESULTS: An overall proportion of five CIN (3.4%) was observed with equal distribution among the groups (4.2% in sodium bicarbonate group vs. 2.7% in sodium chloride group; P=0.614). Parameters of renal function demonstrated no differences between the two hydration regimens on day 1 after angiography; even on day 2 most parameters were similar in groups I and II. CONCLUSION: Renal Insufficiency Following Radiocontrast Exposure demonstrates a homogeneously low rate of CIN after exposure to nonionic, iso-osmolar iodixanol regardless of the use of either bicarbonate sodium or sodium chloride solution for volume supplementation. Low-toxicity contrast media and any hydration may offset potential antioxidant effects of sodium bicarbonate.

Hydrogen peroxide-induced activation of defense mechanisms against oxidative stress in rat pancreatic acinar AR42J cells.

Oxidative stress has been implicated in the pathogenesis of acute pancreatitis. Generally, cells respond to oxidative stress with adaptive changes in gene expression aimed at preventing cellular damage and increasing their survival. However, the overall extent of these genetic changes remains poorly defined. This issue was, therefore, examined in the current study. Following exposure of rat pancreatic AR42J cells to 0.08 mM hydrogen peroxide (H(2)O(2)), a concentration failing to induce necrotic cell death, the expression of 96 stress-related genes was monitored by cDNA microarray analysis. H(2)O(2) provoked a time-dependent reorientation of 54 genes. In particular, at 6 and 24 h, 27 and 11 genes were induced, whereas 10 and 6 genes were suppressed, respectively, showing that the degree of change was stronger at the early time point, and that the number of up-regulated genes was obviously larger than the number of down-regulated genes. Reverse transcription-PCR for selected genes confirmed the gene expression pattern. Many of the differentially up-regulated genes can be related to the antioxidant enzymatic defense system, to cell cycle arrest, to repair and/or replacement of damaged DNA, to repair of damaged protein, and to activation of the NF-kappaB pathway. The results suggest that AR42J cells respond to sublethal oxidative stress with transient transcriptional activation of multiple defense mechanisms that may be an indication for a complex adaptation process. An understanding of the cellular stress responses may lead to new insights into the pathogenesis of oxidative stress-related diseases including acute pancreatitis.

Platelet and plasma BDNF in lower respiratory tract infections of the adult.

Enhanced bronchial responsiveness during and following lower respiratory tract infections is a major clinical problem, but its pathogenesis is poorly understood. Brain-derived neurotrophic factor (BDNF), which can be released by platelets and leukocytes, has been identified as a mediator of bronchial hyperresponsiveness. It is unknown whether the release of BDNF is altered during lower respiratory tract infections of the adult. In this clinical pilot study, 16 patients (35-80 years old) with the diagnosis of an acute bacterial lower respiratory tract infection and elevated serum concentrations of c-reactive protein (>100 microg/ml) and procalcitonin (>0.1 ng/ml) were examined on admission to the hospital and 1 week after antibiotic treatment. Sixteen age- and sex-matched controls were examined in the same time period. BDNF concentrations in serum and platelets, but not in plasma, were markedly reduced in patients on the day of admission (median <25% of the controls). Analysis of the platelet marker serotonin (5-HT) suggested that the decrease of platelet BDNF is part of a non-specific release of platelet-derived mediators in this condition. Clinical improvement was accompanied by a restoration of serum and platelet BDNF concentrations which returned to control levels after 1 week of treatment. Cell culture experiments revealed that bacterial lipopolysaccharide (LPS) enhanced the release of BDNF by peripheral blood mononuclear cells of the patients at both time points. In conclusion, these data suggest that lower respiratory tract infections might be associated with an augmented release of BDNF by platelets and mononuclear cells.

Improvement of early vascular changes and cardiovascular risk factors in obese children after a six-month exercise program.

OBJECTIVES: The present study aimed to assess the effect of a 6-month exercise program in obese children on flow-mediated vasodilation (FMD) carotid intima-media thickness (IMT) and cardiovascular risk factors (RF). BACKGROUND: Childhood obesity contributes to adult obesity and subsequent cardiovascular disease. Physical inactivity is a major RF for obesity, endothelial dysfunction, and elevated carotid IMT, culminating in early atherosclerotic disease. METHODS: Sixty-seven obese subjects (age 14.7 +/- 2.2 years) were randomly assigned to 6 months' exercise or non-exercise protocol. We examined the influence of exercises (1 h, 3 times/week) on FMD, IMT, and cardiovascular risk profile. RESULTS: Compared with lean control subjects, obese children demonstrated at baseline significantly impaired FMD (4.09 +/- 1.76% vs. 10.65 +/- 1.95%, p < 0.001), increased IMT (0.48 +/- 0.08 mm vs. 0.37 +/- 0.05 mm, p < 0.001), and a number of obesity-related cardiovascular RF. Significant improvements were observed in the exercise group for IMT (0.44 +/- 0.08 mm, p = 0.012, -6.3%) and FMD (7.71 +/- 2.53%, p < 0.001, +127%). This improvement correlated with reduced RF, such as body mass index standard deviation scores, body fat mass, waist/hip ratio, ambulatory systolic blood pressure, fasting insulin, triglycerides, low-density lipoprotein/high-density lipoprotein ratio, and low-degree inflammation (C-reactive protein, fibrinogen). CONCLUSIONS: The present study documented increased IMT, impaired endothelial function, and various elevated cardiovascular RF in young obese subjects. Regular exercise over 6 months restores endothelial function and improves carotid IMT associated with an improved cardiovascular risk profile in obese children.

Impaired flow-mediated vasodilation, carotid artery intima-media thickening, and elevated endothelial plasma markers in obese children: the impact of cardiovascular risk factors.

OBJECTIVES: Childhood obesity contributes to the development of adult obesity and subsequent cardiovascular disease. The present study aimed to assess vascular status (flow-mediated vasodilation [FMD], intima-media thickness [IMT]) and to analyze plasma surrogate endothelial markers (von Willebrand factor [vWf], E-selectin, and thrombomodulin) in obese children as compared with controls. Associations between early morphologic and functional vascular changes, surrogate soluble markers of early atherosclerosis, and the cardiovascular risk profile were determined. METHODS: We examined 32 obese children versus 20 control subjects. All of the children underwent identical screening, comprehensive risk factor assessment, and measurements of E-selectin, vWf, thrombomodulin, FMD, and IMT. RESULTS: Compared with controls, obese children demonstrated significantly impaired FMD and increased IMT. Concentrations of soluble E-selectin and thrombomodulin were significantly elevated in obese children, whereas vWf showed no significant differences between obese children and controls. FMD, IMT, E-selectin, and thrombomodulin were significantly associated with various risk factors, including the extent of obesity, arterial hypertension, fibrinogen, C-reactive protein, and low physical fitness. CONCLUSIONS: The present study documented increased IMT, impaired endothelial function, and elevated plasma markers of endothelial activation and injury in obese children. Morbid obesity, arterial hypertension, subclinical inflammation, and low physical fitness formed a risk profile associated with the risk of early atherosclerosis in these children. Sonographic assessment of vascular status and the estimation of soluble endothelial plasma markers, combined with comprehensive risk factor screening, may form a rationale to identify high-risk children susceptible to early atherosclerotic disease and to monitor vascular changes during follow-up studies and therapeutic measures.

Thrombin/thrombin receptor (PAR-1)-mediated induction of IL-8 and VEGF expression in prostate cancer cells.

Interleukin 8 (IL-8) and vascular endothelial growth factor (VEGF) are two cytokines promoting prostate tumor growth and angiogenesis. The main coagulation protease thrombin may modulate the malignant phenotype of prostate cancer cells via its cellular receptor(s). We aimed to investigate the effects of thrombin on IL-8 and VEGF expression in DU 145 prostate cancer cells. Thrombin induced the expression and secretion of IL-8 and VEGF, with more pronounced effects on IL-8. Target-specific siRNA-induced protease-activated receptor 1 (PAR-1) knockdown completely neutralized thrombin-enhanced cytokine secretion, demonstrating the essential role of PAR-1. Inhibitors of either extracellular signal-regulating kinase (ERK) or phosphatidylinositol 3-kinase (PI3K) partly reversed the thrombin-induced cytokine expression, suggesting that both ERK and PI3K kinase pathways may be involved in IL-8 and VEGF expression. The results suggest that the thrombin/PAR-1 system upregulates cytokines in prostate cancer cells which in turn may contribute to the progression of prostate cancer.