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Factor V Leiden (FVL) mutation and prothrombin G20210A mutation are common hereditary risk factors for venous thrombosis. In the current study, 40 patients (mean age +/- standard deviation, 35 +/- 11 years) and 764 healthy control subjects (mean age +/- standard deviation, 37 +/- 14 years) were enrolled to assess the potential role of these mutations in the manifestation of thrombotic microangiopathies. Compared with controls, neither the heterozygous FVL mutation (7.5% vs 8.5%; P = 1) nor the heterozygous prothrombin mutation (2.5% vs 2.8%; P = 1) was more prevalent in the patients. The findings do not support a significant role of FVL and prothrombin mutations as risk factors for the manifestation of thrombotic microangiopathies. Thus, screening for these mutations does not allow the identification of individuals at increased risk for these rare thrombotic disorders.

In this study, we assessed the potential role of the TT genotype of the gene of the methylenetetrahydrofolate reductase for the manifestation of thrombotic microangiopathies, enrolling 40 affected patients (mean age [+/- standard deviation] 35 +/- 11 years). As a result, the methylenetetrahydrofolate reductase 677TT genotype was more prevalent in patients with thrombotic microangiopathies compared with controls (adjusted odds ratio = 2.58, 95% confidence interval = 1.2-5.7, P = .018), particularly in those suffering from the hemolytic uremic syndrome. A hemolytic more severe clinical course of thrombotic microangiopathies in carriers of the methylenetetrahydrofolate reductase 677TT genotype was not observed. In summary, our findings suggest a significant influence of the methylenetetrahydrofolate reductase genotype on the manifestation of thrombotic microangiopathies. The 677 TT genotype of this polymorphism appears to be a risk factor for manifestation of these rare thrombotic disorders, possibly explained by endothelial activation and increased oxidative stress.

A 37-year-old female patient with systemic mastocytosis who was admitted with severe unexplained bleeding symptoms is studied. Laboratory procedures established the diagnosis of a patient-derived-heparin-like anticoagulant as a very rare hemostatic abnormality predisposing to bleeding. The patient died from refractory disease despite therapy with protamine, initiation of chemotherapy, and supportive measures. The case illustrates the clinical presentation and diagnosis of heparin-like anticoagulants. Etiology, pathophysiology, and therapeutic options are discussed.

Background: Gender-related differences in incidence of arterial thrombosis have been a focus of interest for years. The platelet integrin alphaIIbbeta3 is primarily responsible for the interaction between platelets and fibrinogen and consecutive thrombus growth. In this study, we evaluated platelet adhesion onto immobilized fibrinogen under venous and arterial flow conditions in men and women., Methods: Platelets in whole anticoagulated blood were labelled with the fluorescence dye Mepacrine and perfused through the rectangular flow chamber over glass cover slips coated with fibrinogen (shear rates of 50 s-1, 500 s-1 and 1500 s-1). A fluorescence laser-scan microscope was used for visualisation and quantification of platelet adhesion at 15 seconds, 1 and 5 minutes after the start of perfusion., Results: During perfusion, the platelet adhesion linearly increased in regard to exposition time and shear rate. After five minutes of perfusion the platelet adhesion onto immobilized fibrinogen showed no significant gender related difference, neither at 50 s-1 nor at 500 s-1 and 1500 s-1 (p > 0.05), respectively. No significant difference in platelet adhesion onto immobilized fibrinogen, in regard to the menopausal status, was either observed (p > 0.05)., Conclusion: In our in vitro experimental system, hormonal differences between men and women did not influence platelet adhesion onto immobilized fibrinogen, neither under venous nor under arterial rheological conditions.

Thromboangiitis obliterans or Buerger's disease is an episodic and segmental inflammatory and thrombotic process of the medium and small arteries of the lower extremities. Even though the disease was described 90 years ago, the etiopathogenesis is still under consideration. Afflicted patients are mostly young male cigarette smokers without signs of atherosclerosis or other risk factors for peripheral arterial occlusive disease. This indicates that hereditary thrombophilic factors could play a role in the etiopathogenesis. Recently, increasing evidence shows that platelet receptor polymorphisms (HPA-1 polymorphism of beta3 subunit of alphaIIbbeta3 and 807 C/T polymorphism alpha2beta1) are associated with early onset of arterial thrombosis (myocardial infarction, stroke). This case-control study was designed to assess whether the 807 C/T polymorphism or the HPA-1 polymorphism is involved in the pathogenesis of Buerger's disease or has any influence on the clinical course of Buerger's disease. Eighteen patients with Buerger's disease and 81 (sex and age matched) healthy control subjects (mean age 44 +/- 10 vs 45 +/- 8 years, respectively) were genotyped for platelet receptor HPA-1 and GPIa 807 C/T polymorphism. The gene frequency of HPA-1 and GPIa 807 C/T polymorphisms was identical in both groups. Prevalence of hetero- and homozygous carriers of the HPA-1b allel (1a1b and 1b1b genotype) as well as the prevalence of the 807 C/T and 807 T/T carriers did not differ significantly between the two groups, p >0.05. The grade of clinical disease manifestation as well as disease progression did not reveal any significant relationship with HPA-1 and 807 C/T polymorphisms. A relationship between the age at onset of the disease and HPA-1 polymorphism was not found. Otherwise analysis of the GPIa 807 C/T platelet receptor polymorphism showed that the average age of patients who are carriers of the T allele at early onset of disease was 32 +/- 6 years (range 27-48 years) compared to 42 +/- 6 years (range 34-53 years) of the C/C carriers (p <0.05). This indicates that the GPIa 807 C/T polymorphism does not represent a risk factor for Buerger's disease itself, but could be associated with premature onset of this disorder in predisposed individuals.

Background: Platelet adhesion and subsequent thrombus formation on a subendothelial matrix at the site of vascular damage play a crucial role in the arrest of posttraumatic bleeding but also in different pathological thrombotic events, such as acute coronary syndrome and stroke. Recently published studies have clearly demonstrated that platelet integri alphaIIbbeta3 is intimately involved in the occlusive thrombus formation at the site of endothelial damage. Therefore, any genetic variation in the expression of this receptor may lead to an excessive bleeding or excessive thrombus formation. In this study, we evaluated the influence of HPA-1 polymorphism of integrin alphaIIbbeta3 on platelet adhesion onto immobilized fibrinogen using an in vitro system simulating blood flow., Methods: Platelets in anticoagulated whole blood [49 healthy previously genotyped blood donors) were labelled with fluorescence dye and perfused through a rectangular flow chamber (shear rates of 50 s(-1), 500 s(-1) and 1500 s(-1)). A fluorescence laser-scan microscope was used for visualisation and quantification of platelet adhesion at 15 sec, 1 and 5 minutes after start of perfusion., Results: During perfusion, the platelet adhesion linearly increased with regard to exposition time and shear rate. Perfusion of blood preincubated with Abciximab over fibrinogen-coated cover-slips showed reduced platelet adherence (absolute fluorescence: 168 +/- 35 U vs. 53000 +/- 19000 at control experiments, p < 0.05), as well as by perfusion over BSA-coated glass coverslips. Platelet with HPA-1a/1a genotype exhibited initial better adhesion but they also exhibited higher detachment under arterial flow conditions compared to the HPA-1b/1b platelets. Analysis of stable adhesion rate indicate that the platelets carrying the HPA-1b/1b genotype have a higher reactivity threshold for initial interaction with fibrinogen but under the higher shear rate (in regard to time of perfusion) also realize more stable bonds with fibrinogen than platelets with the HPA-1a/1a genotype., Conclusion: Our data support the contention that genetically determined variants of platelet integrins alphaIIbbeta3 could play a role in arterial thrombogenesis and thus confirm the hypothesis derived from epidemiological studies.

Background: Adhesion of platelets onto immobilized fibrinogen is of importance in initiation and development of thrombosis. According to a recent increase in evidence of a multiple biological property of antithrombin, we evaluated the influence of antithrombin on platelet adhesion onto immobilized fibrinogen using an in-vitro flow system., Methods: Platelets in anticoagulated whole blood (29 healthy blood donors) were labelled with fluorescence dye and perfused through a rectangular flow chamber (shear rates of 13 s-1 to 1500 s-1). Platelet adhesion onto fibrinogen-coated slips was assessed using a fluorescence laser-scan microscope and compared to the plasma antithrombin activity. Additionally the effect of supraphysiological AT supplementation on platelets adhesion rate was evaluated., Results: Within a first minute of perfusion, an inverse correlation between platelet adhesion and plasma antithrombin were observed at 13 s-1 and 50 s-1 (r = -0.48 and r = -0.7, p < 0.05, respectively). Significant differences in platelet adhesion related to low (92 +/- 3.3%) and high (117 +/- 4.1%) antithrombin activity (1786 +/- 516 U vs. 823 +/- 331 U, p < 0.05) at low flow rate (13 s-1, within first minute) have been found. An in-vitro supplementation of whole blood with antithrombin increased the antithrombin activity up to 280% and platelet adhesion rate reached about 65% related to the adhesion rate in a non-supplemented blood (1.25 +/- 0.17 vs. 1.95 +/- 0.4 p = 0.008, respectively)., Conclusion: It appears that antithrombin in a low flow system suppresses platelet adhesion onto immobilized fibrinogen independently from its antithrombin activity. A supraphysiological substitution of blood with antithrombin significantly reduces platelet adhesion rate. This inhibitory effect might be of clinical relevance.

Background: Hyperhomocysteinemia is an important and independent risk factor for vascular disease. About 35% of patients with stroke and 47% of patients with peripheral arterial disease have elevated plasma homocysteine (HCY) concentrations. The relationship between plasma HCY and the methylentetrahydrofolate reductase (MTHFR) C677T polymorphism is still unclear, especially in regard to screening/diagnostic power., Methods: This case-control study was performed on 96 patients, who underwent surgery due to asymptomatic or symptomatic high grade stenosis of the internal carotid artery (ICA), and 96 healthy age and sex-matched, controls. Plasma HCY concentration was determined using a commercial kit for fully automated analysis (AxSYM, Abbott). The C677T polymorphism of the MTHFR-gene was assessed by PCR., Results: The mean plasma HCY concentration was significantly higher in the group with stenosis of ICA compared to the controls, 12.43 +/- 6.96 microM and 10.16 +/- 3.16 microM, respectively, (p < 0.05). An HCY plasma concentration of 1.5 SD above the mean value of the control group, was defined as cut-off for a pathological versus physiological plasma concentration. The sensitivity and specificity of HCY was 0.27 and 0.94, respectively. The positive predictive value was 0.82. There was no significant difference in the frequency of the MTHFR 677 CT and TT genotype between patients and controls (47% vs. 47% and 8.3% vs. 11.4%, respectively). Carriers of the T-allele (CT and TT genotypes) have significantly higher plasma HCY concentrations than CC patients, 14.1 +/- 7.6 microM and 10.29 +/- 5.2 microM, respectively, p < 0.05. Sensitivity and specificity of the MTHFR C677T polymorphism (T-allele) were 0.56 and 0.40, respectively. The positive predictive value was 0.48. There was no significant difference in plasma HCY or genotype frequency of the MTHFR C677T polymorphism between asymptomatic and symptomatic patients., Conclusion: Our study shows that in a population with a given pretest disease probability of 50%, the determination of plasma HCY concentration, with a positive predictive value of 0.82, is more suitable for screening of patients at risk than analysis of the MTHFR C677T polymorphism.

Background: Current debates are focused on inflammatory processes in atherosclerotic lesions as a possible pathomechanism for destabilization and thrombembolism. In this prospective study the role of systemic and local infection in patients with high-grade internal carotid artery stenosis (ICA) was evaluated., Patients and Methods: Serum antibody titers of 109 consecutive patients, who underwent surgery for ICA stenosis (asymptomatic n = 40, symptomatic n = 69) were prospectively measured for Chlamydia pneumoniae (Cpn) (IgA and IgG), Herpes simplex virus (HSV) (IgG, IgM) and Cytomegalovirus (CMV) (IgG, IgM) respectively. 53 carotis plaques of this group (asymptomatic n = 17, symptomatic n = 36) could be analyzed by polymerase chain reaction (PCR) for Cpn-, HSV- and CMV-DNA presence., Results: Seropositivity was found in 61,5% for Cpn, 91,7% for HSV and 72,5% CMV respectively. No significant relation was found between symptomatic and asymptomatic patients as well as no difference was seen for presence of IgA antibodies against Cpn comparing both groups. Plaque-PCR revealed Cpn in 7 cases (13,2%), HSV in 2 cases (3,8%) and no CMV had been detected. Again, no significant relationship was found concerning symptomatic and asymptomatic patients. All 9 PCR-positive plaques displayed lesions of "complicated atherosclerosis" as central fibrous necrosis and calcification or plaque bleeding and surface thrombosis., Conclusions: Our results do not support the hypothesis that systemic Cpn, HSV or CMV- infection or evidence of Cpn-, HSV- or CMV-DNA in carotid plaques causes plaque destabilization and cerebral thromboembolism. Plaque infection could only be observed in cases with advanced atherosclerosis.

Homocysteine may have deleterious effects on the cardiovascular system. It has been hypothesized that these effects may be brought about by a decrease in the adenosine concentration via the S-adenosylhomocysteine hydrolase reaction. A requirement for this causal relationship is proof of a reduction in vascular adenosine concentration during conditions of elevated homocysteine concentrations. In the present communication we summarize published data obtained during systematic variation of the arterial homocysteine concentration. Most of the results reported show that an increase in homocysteine concentration to 100 microM is associated with a 20-50% decrease in vascular adenosine concentration and an increase in tissue S-adenosylhomocysteine level. Homocysteine effects on the adenosine concentration seem to be more pronounced under conditions of impaired oxygenation. Further experiments, in particular on organs and tissue that release high amounts of homocysteine, i.e., the liver, are warranted to study the potential effects of homocysteine on vascular and tissue adenosine concentrations and consequent effects on organ function. The evidence obtained may be relevant for future assessment of risk indicators in conjunction with homocysteine pathogenicity, which might potentially be extended to measurements of adenosine or S-adenosylhomocysteine levels.

5, 10-Methylenetetrahydrofolate Reductase (MTHFR) is one of the key enzymes in the metabolism of homocysteine, where it catalyses its remethylation. The autosomal recessive bp 677 C --> T mutation in the MTHFR gene leads to the substitution of valine for alanine. Individuals who are homozygous for this C677T mutation exhibit a decreased specific activity and increased thermolability of this enzyme. This leads to increased plasma levels of homocysteine, which is a known risk factor for atherosclerosis and various manifestations of the atherosclerotic disease. The aim of this study was to find out the distribution and frequency of this mutation in the general Croatian population. A group of 228 volunteers (175 males and 53 females) has been analyzed for the MTHFR polymorphism, which revealed the following distribution: 105 (46.05%) individuals were without mutation (C/C), 102 (44.74%) were heterozygous (C/T) and 21 (9.21%) homozygous (T/T). These findings are within the results of studies on other European populations.

Aim: Ferritin acts as an iron scavenger and thereby may reduce iron catalysed oxygen radical production during reperfusion injury. We tested the hypothesis that the myocardial ferritin concentration is enhanced during ischaemia in proportion to the blood flow reduction., Methods: In 10 anaesthetized, open chest Beagle dogs (six controls and four with 60 min coronary occlusion) regional myocardial blood flow (RMBF) was measured with the tracer microsphere technique and ferritin was determined in samples with an average mass of 125 mg (124-256 samples per heart)., Results: Under physiological conditions heart rate was 88 +/- 12 bpm, mean aortic pressure 98 +/- 8 mmHg, and RMBF 0.99 +/- 0.33 mL min-1 g-1. Data did not differ between experimental groups, P > 0.05. In the control group regional myocardial ferritin concentration averaged 11.76 +/- 3.54 ng mg-1 protein and exhibited a significant blood flow independent heterogeneity (CV(biol) = 0.27). However, between low and high flow areas (relative flow <0.5 and >1.5 times the average RMBF, respectively) no significant difference in ferritin was found, P > 0.05. In four experiments, in which regional blood flow was reduced by 40% to 0.60 +/- 0.23 mL min-1 g-1, regional ferritin content was significantly higher as compared with the control group 27.95 +/- 6.16 vs. 11.76 +/- 3.54 ng mg-1 protein, respectively. An inverse relationship was observed between ferritin and RMBF, r = -0.61, P < 0.001. Thus, a reduction of RMBF of >80% was associated with a 2.75-fold increase of the average ferritin content. Between subepicardium and subendocardium no significant difference in ferritin content was observed, neither in the control group nor in the group with induced ischaemia. Regions with control low and high flow responded similarly to the coronary constriction with regard to the local ferritin concentration: 27.88 +/- 15.22 vs. 30.10 +/- 14.91 ng mg-1, P > 0.05, respectively. A data analysis using Baye's theorem indicated that sensitivities were 0.28 and 0.94 for average flow reductions of 5 and 93%. In additional in vitro measurements (ischaemic incubation at 37 degrees C) myocardial ferritin content increased almost linearly within the first 60 min of incubation and thereafter remained unchanged., Conclusions: (1). Local physiological ferritin content in myocardium is heterogeneous and unrelated to control myocardial blood flow. (2). Ischaemia results in an enhanced ferritin content in relation to the degree of ischaemia. (3). The increase of myocardial ferritin requires a severe degree of ischaemia.

Unlabelled: Local myocardial blood flow measurements are of great importance in experimental and clinical settings. However, a lack of ideal markers is evident. Adenosine is suggested to be a potential candidate because of its high uptake and rapid intracellular sequestration. We specifically tested the hypothesis that the local deposition density of labeled adenosine within the heart reflects local myocardial blood flow., Methods: Tracer microspheres, the recognized standard for local blood flow measurements, were injected and compared with simultaneously injected labeled adenosine ((3)H/(14)C) in tracer concentration into the left atrium of anesthetized Beagle dogs (n = 7). Myocardial deposition densities were assessed through beta-scintillation and gamma-counting measurements in samples (100-128 per heart) of an average wet mass of 487 +/- 54 mg. To challenge local myocardial blood flow distribution, alprostadil was infused into the left circumflex artery in 3 experiments. In 2 other experiments, erythro-9-hydroxy-nonyl-adenine (EHNA) was infused to inhibit degradation of injected adenosine to inosine., Results: Tracer adenosine and microspheres did not exert significant local or systemic hemodynamic effects. Both were almost completely extracted from blood within 2 min and locally retained in the tissue. Deposition densities of tracer microspheres and labeled adenosine correlated closely in each experiment, independently of the respective protocol (control, EHNA, or alprostadil), over a wide range of local myocardial blood flows (0.23-12.9 mL min(-1) g(-1)). The mean correlation coefficient (n = 293) was r = 0.93 (r(2) = 0.86; P < 0.0001), indicating that the deposition density of (3)H-adenosine could explain local blood flow as measured with the tracer microsphere technique with 86% probability., Conclusion: Adenosine appears to be a reliable marker of local blood flow in dog myocardium.

It is well established that myocardial blood flow is heterogeneous on the local level. During recent years comprehensive studies have been undertaken to assess the relation between myocardial metabolism and spatial blood flow heterogeneity. Based on the type of measurements two major groups of studies have been performed: enzyme activity and tissue metabolite level assessments. Enzyme activity measurements have provided only limited insight into the coupling of local metabolism and flow. This is probably due to the fact that, in addition to estimated Vmax values, local substrate affinity (Km values) and substrate concentrations affect the metabolite fluxes. However, the latter two variables remain normally unknown. In contrast, valuable insight has been obtained concerning flow-metabolism matching from tissue metabolite measurements, especially when connected with mathematical model analyses. The latter permitted the calculation of metabolic flux rates (e.g., production of oxidation water, citric acid cycle flux, glucose uptake, fatty acid uptake) or the translation of the metabolic indexes into physiologically meaningful local metabolite concentrations (e.g., free cytosolic adenosine). The bottom line of the studies reported to date is that the broad range of myocardial flows observed under resting control conditions correlates with local metabolism possibly affected by spatial differences in adrenergic stimulation. Thus, high flow samples exhibit a higher oxidative metabolism than low flow samples. As a result the flow threshold below which local myocardial ischemia ensues is higher in control high flow samples. The importance of these findings with respect to local flow-metabolism matching is underlined by the finding that the probability of developing an infarction following ischemia/reperfusion is related to the functional state of the myocardium under control conditions, i.e., the local level of flow-metabolism matching.

Background: Homocysteine (HCY) was recently established as an independent risk factor for atherosclerosis. The prevalence of an increased homocysteine plasma concentration was reported to be up to 6-fold higher in patients with different locations of arterial occlusive diseases., Aim: This study evaluated critically whether the total HCY plasma concentration can be used as a screening marker for peripheral arterial disease in the general population., Methods: Study subjects were 40 patients (51.8 +/- 7.5 years) with symptomatic lower limb peripheral arterial disease (PAD) (stage II) and 40 healthy volunteers (45.6 +/- 6.8 years, P< 0.05 vs PAD). The percentage of women in both groups was 30%. The plasma HCY concentration was determined by using derivatization techniques and subsequent fluorescence high-performance liquid chromatography., Results: Total plasma HCY concentration was significantly higher in the PAD group than in controls (14.90 +/- 5.78 microM vs 11.32 +/- 2.95 microM, respectively, P< 0.001). Also, the coefficient of variation of plasma HCY in PAD was significantly higher than that in the control group, 0.38 vs 0.25 (P< 0.001), respectively, reflecting greater interindividual differences. In addition to a PAD-specific effect, the plasma HCY concentration was also dependent on gender and age (both P< 0.05). Sensitivity and specificity of HCY as a marker of PAD were 0.3 and 0.95, respectively. Positive and negative predictive values were 0.85 and 0.42, respectively., Conclusions: From these data it is concluded that HCY metabolism may have an influence on the development of PAD in one-third of all patients with PAD, and that total plasma HCY concentration may not be suited as a screening test for PAD in the general population but rather serves as a monitoring marker in certain risk groups.

Objectives: The myocardium of the left ventricle exhibits spatial heterogeneity of blood flow under physiological conditions. This study was designed to investigate, whether oxygen supply is jeopardized in low flow areas (blood flow < 50% of mean) under physiological conditions and whether areas of high flow (> 150% of mean) exhibit perfusion in excess of demand ("luxury perfusion")., Methods: The study was performed in anesthetized and ventilated beagle dogs. Local blood flow was reduced by mechanically narrowing of the r. circumflexus of the left coronary artery; myocardial blood flow was measured by the tracer-microsphere technique, free concentrations cellular adenosine by the SAH-technique, regional metabolism of substrates by the desoxyglucose-technique., Results: Low flow areas exhibited normal oxygenation of the myocardium, while in high flow areas no luxury perfusion could be demonstrated., Conclusion: Myocardial blood flow and metabolism demonstrate significant spatial heterogeneity. There appears to be no absolute threshold of blood flow, where regional myocardial ischemia develops. Probably biochemical evidence of myocardial ischemia is determined by a local ratio of oxygen supply and demand.

The concentration of heat-shock proteins of 70 kD (HSP70) in heart tissue has been shown to increase during transient myocardial ischaemia and to persist during several hours of reperfusion. In this study the relationship between the local myocardial HSP70 concentration and blood flow was addressed for control physiological conditions and acute myocardial ischaemia. A specific aim of this study was to address the question of whether low flow areas under control physiological conditions have undergone a transient ischaemia during the preceding hours and thus may be in a state of hibernation or stunning. In 12 anaesthetized, open-chest beagle dogs (6 control and 6 with 60-min coronary artery stenosis) heart rate, mean aortic pressure, mean arterial partial pressure of O2 and partial pressure of CO2 averaged 85+/-16 beats/min, 94+/-14 mmHg, 102+/-17 mmHg and 39+/-6 mmHg, respectively. Regional HSP70 and myocardial blood flow (RMBF) were measured using an HSP70-enzyme-linked immunosorbent assay and the tracer microsphere technique, respectively, in samples of 250 mg wet mass. In the control group the mean RMBF was 1.06+/-0.59 ml.min-1.g-1 and the local HSP70 concentration was 7.08+/-1.03 microg/mg cytosolic protein. Myocardial HSP70 showed a blood flow-independent regional biological heterogeneity, equivalent to a coefficient of variation of 0.31. Local HSP70 concentrations did not differ (P>0.05) between control low and high flow samples, 6.16+/-1.0 vs 6.08+/-0.75 microg/mg cytosolic protein, respectively. However, after 60 min of coronary artery occlusion the local HSP70 concentration increased from 7.08 +/-1.03 to 13.43+/-3.19 microg/mg cytosolic protein (P<0. 001). There was a significant inverse relationship between the percent reduction of local blood flow and HSP70 (r=-0.56, P<0.001). From these results it is concluded that: (1) low flow samples under control physiological conditions are unlikely to be in a state of hibernation or stunning since their HSP70 concentration is normal and (2) the increase in the local HSP70 concentration during myocardial ischaemia reflects the degree of impairment of O2 delivery.

Background: Left ventricular myocardial blood flow is spatially heterogeneous. The hypothesis we tested was whether myocardial areas with a steady-state flow <0.5 times mean flow are underperfused and areas with flow > 1.5 times mean flow are overperfused., Methods and Results: In anesthetized beagle dogs (n=10), the relationship between local blood flow versus S-adenosylhomocysteine (SAH) concentration, a measure of the free intracellular adenosine concentration, and lactate, a measure of the myocardial NADH/NAD+ ratio, were determined under control conditions and after coronary constriction. Control local myocardial blood flow was 0.99+/-0.46 mL x min(-1) x g(-1), with a coefficient of variation of 0.36+/-0.12 (n=256 per heart; sample wet mass, 125+/-30 mg). Tissue concentrations of SAH (3.4+/-2.5 nmol/g) and lactate (1.88+/-0.80 micromol/g) were not elevated in low-flow samples. However, after coronary artery constriction, poststenotic blood flow decreased from 1.00+/-0.27 to 0.49+/-0.22 mL x min(-1) x g(-1) (P<0.04), with significant correlation between local SAH and flow (r=-0.59) and lactate and flow (r = -0.50). Although nearly all samples from control high-flow regions showed increased SAH concentrations if relative flow after stenosis was <1.0, control low-flow samples frequently displayed low SAH concentrations. The percent reduction in flow determined the changes in the local SAH and lactate concentration, independent of the local control blood flow., Conclusions: When the coronary inflow is unrestricted, the oxygen supply to control low-flow regions meets metabolic demand. Flow to control high-flow regions reflects a higher local demand rather than overperfusion. Thus, blood flow heterogeneity most likely reflects differences in aerobic metabolism.

Myocardial ischemia can be described as an imbalance of energy demand and energy supply. Even during ischemia, energy metabolism is predominantly aerobic. Only during the most severe underperfusion (residual flow less than 5%) anaerobic metabolism exceeds aerobic metabolism quantitatively. ATP synthesis and ATP metabolism are in a steady state during myocardial ischemia, albeit on a reduced level. A locally low blood flow rate may exist under physiological conditions without the presence of ischemia. The basis for the underlying flow heterogeneity is functional. While experimental data have been widely obtained during acute ischemia, metabolic rates have scarcely been determined during conditions of chronic myocardial ischemia. First experimental studies, however, show that uptake of fluoro-deoxyglucose is enhanced for hours after induction of myocardial stunning and even for months during myocardial hibernation.

The S-adenosylhomocysteine (SAH) technique allows the estimation of the free cytosolic adenosine concentration using the kinetic properties of the enzyme SAH-hydrolase (adenosine+homocysteine reversible SAH+H2O). Besides the cytosolic adenosine concentration, the local SAH signal may also depend on the local homocysteine availability, the continuous production of SAH from S-adenosylmethionine (SAM-->SAH+CH3) and the activity of the enzyme SAH-hydrolase. These variables were studied with high spatial resolution (sample dry mass 25 mg) in left ventricular myocardium from 26 anesthetized open-chest dogs in which heart rate averaged 86 +/- 14 beats/min and mean aortic pressure 96 +/- 17 mmHg. Homocysteine infusion (48 mg/kg i.v.) increased the normal plasma homocysteine concentration from 5.0 +/- 0.8 to 586 +/- 40 microM after 30 min when the average tissue concentration was 94% of the plasma concentration and similar in low and high flow areas (flow range 0.04 to 1.91 ml/min/g). Local SAH content was 1.18 +/- 0.48 nmol/g under control conditions and increased to 4.33 +/- 0.59 nmol/g within 60 min following competitive blockage of the SAH-hydrolase by adenosine dialdehyde (10 mumol/kg i.v.). This increase of the SAH content was slightly more in high than in low-flow areas (P < 0.01). Regional SAH-hydrolase activity (9.0 +/- 0.5 nmol/min/g) was comparable in high and low flow areas. All three variables exhibited an observed variability which was larger than the methodical variability suggesting significant spatial heterogeneity in the myocardium. A regrouping analysis indicated that between four and five samples taken from distant sites should be averaged to obtain a robust estimate of the above metabolic parameters. Reconciling the measurements with a mathematical model of cardiac adenosine metabolism and fitting of the measured SAH tissue levels gave an estimate of 72 pmol/min/g for the mean transmethylation rate. Estimates of the cytosolic adenosine concentration of cardiomyocytes and endothelial cells under control physiological conditions were 24 and 7 microM, respectively. Thus, the present measurements provide a basis for the quantitative assessment of the local cytosolic adenosine concentration in relation to blood flow.

The spatial heterogeneity of myocardial perfusion and metabolism was studied in 11 anaesthetized dogs under resting conditions. In each heart local myocardial blood flow was assessed using the tracer microsphere technique in 256 samples (mean mass: 83.1 mg) taken from the left anterior ventricular wall. In the same samples, the following biochemical parameters were determined: accumulation of [3H]-deoxyglucose (a measure of glucose uptake), free cytosolic adenosine (S-adenosylhomocysteine accumulation technique, a measure of tissue oxygenation and a possible mediator of blood flow regulation), and the specific activities of oxidative (citrate synthase, cytochrome-c-oxidase) and glycolytic (hexokinase, phosphoglycerate kinase) enzymes. Capillary density and mitochondrial and myofibril volume densities were determined by morphometry. Myocardial perfusion in each sample (average 0.77 ml min-1 g-1) varied between 0.1 and 2.5 times the mean (coefficient of variation 0.30+/-0.02). [3H]-deoxyglucose was deposited locally in proportion to perfusion. Samples showing low flow (<0.2 ml min-1 g-1) did not exhibit increased levels of cytosolic adenosine. The specific activities of the oxidative and glycolytic enzymes, however, were uniformly distributed between low and high flow areas. Furthermore, capillary density and mitochondrial and myofibril densities were similar in high and low flow regions. The results show firstly that local glucose metabolism in the heart occurs in proportion to local blood flow, suggesting that high flow regions have a higher than average metabolic rate. Secondly, regions of low flow are not compromized by critical oxygenation and most likely have a lower than average oxygen demand and finally, the homogeneous distribution of oxidative and glycolytic enzymes, as well as the homogeneous myocardial ultrastructure, suggest that areas with high and low blood flow under resting conditions may increase their metabolic rate to similar levels when required.

The aim of the study was to determine the diagnostic value of carcinoembryonic antigen (CEA) and ferritine in malignant and tuberculous non-bloody pleural effusion. The etiology of diseases was determined by cytologic, histologic and microbiologic methods. CEA concentration above 5 ng/ml and ferritine concentration above 200 ng/ml were considered to be positive. There was significant difference in the value of CEA measured in malignant and in tuberculous pleural effusion (P < 0.005) as well as in the sera (P < 0.01) of these two groups. There was no correlation between concentration of CEA and ferritine in malignant pleural effusion. Ratio between CEA and ferritine in effusions and sera was of no help in discriminating malignant from tuberculous effusions. No correlation between examined markers and physical status of patients was observed. The sensitivity and specificity of CEA assay in malignant pleural effusion was 65% and 90%, respectively, and for ferritine 67% and 80%, respectively. A high correlation was observed between the CEA concentration in malignant pleural effusion and sera patients (r = 0.95). Combined sensitivity and specificity of CEA and ferritine was 65.9% and 85%. Bayes theorem was used to calculate the positive predictive values for CEA and ferritine, which were 53% and 37%, respectively. Results obtained in the study show the relatively good diagnostic potential of CEA.

To provide an objective measure of the hazard smoking parents represent to their children's health, continue concentration in urine was measured by the colorimetric method using barbituric acid (DBA). A total of 205 children, aged 10-12, were examined. The results of laboratory tests were correlated with the data collected by interview. A significant difference in the average value of cotinine concentration was demonstrated between the children whose parents did not smoke (3.2 mumol/L) and those whose one parent smoked (5.8 mumol/L). An even larger concentration was recorded when both parents smoked (7.8 mumol/L). The largest cotinine concentration was determined in the urine of children--passive smokers whose both parents smoked and who did not have a room of their own (9.2 mumol/L). The difference in cotinine concentration between girls and boys was not statistically significant.