Düzgün Özatli

Circulating and Local Bone Marrow Renin - angiotensin System in Leukemic Hematopoiesis: Preliminary Evidences

Abstract Until last the decade, the renin-angiotensin system (RAS) was considered as a circulating endocrine system. It is now known that there are local RASs in many tissues. It has also recently been hypothesized that there exists a local bone marrow (BM) RAS with paracrine/autocrine pathobiological functions. The aim of this study was to detect BM and peripheral blood levels of the essential RAS components in normal and leukemic hematopoiesis. Concentrations of renin and angiotensin-converting enzyme (ACE) were assayed in BM aspirates and in simultaneously drawn peripheral blood samples of 16 pre-chemotherapy leukemic and 10 post-treatment megaloblastic anemia patients with normal blood counts, as controls. In the leukemia group, the ACE concentration was found to be significantly higher in the BM (38±6.2 U/l) than in the peripheral blood (29.5±5.3 U/l), (p=0.029). In the leukemia group, although the BM renin concentration was higher than the peripheral blood levels (21.3±8.3 vs. 18.6±6.2 U/l), this difference was not statistically significant (p=0.196). In the control group, mean BM renin levels were insignificantly lower than in the peripheral blood (8.6±3 vs. 12.1±4.6 pg/ml) (p=0.059). In the leukemia group, serum ACE levels positively correlated with BM and peripheral blood blast percentages (p<0.05). Serum LDH level (p<0.01), BM blast (p<0.05) and peripheral blast percentages (p<0.01) were inversely correlated with serum potassium in the leukemia group. The results of this study can be considered as the preliminary evidence supporting the hypothesis of the presence of a local BM RAS. Further, molecular biologic and immunohistochemical studies are needed to shed light on this important subject. A better understanding of the interrelationships of RAS and hematopoiesis will bring new insights into the pathobiology and even novel therapies for such neoplastic diseases.

Rational use of the PFA-100 device for screening of platelet function disorders and von Willebrand disease.

Two hundred and five patients referred for evaluation of platelet functions and 126 healthy controls were tested with the PFA-100 instrument. A cut-off value of 150 s for collagen/epinephrine (CEPI) closure time (CT) produced most acceptable sensitivity (90%), specificity (85.2%), and positive (82.6%) and negative (91.6%) predictivity values for screening of platelet function disorders and von Willebrand disease (vWD). All patients with vWD and Glanzmann thrombasthenia could be detected by PFA-100. Both CEPI and collagen/adenosine diphosphate (CADP) CTs were elevated in all of these cases. Sensitivity of the device was 81.6% for patients with platelet secretion defects. CADP CT was normal in 63.9% of the patients in this subgroup. Specificity (47%) and positive predictivity (57%) of the instrument were diminished in patients with low hemoglobin concentrations. Depending on the results, an algorithm was developed for screening of platelet function disorders and vWD with PFA-100.

Evaluation of the Haemostatic System during Ketoacidotic Deterioration of Diabetes mellitus

Clinical observations have indicated the frequent development of thrombotic complications during diabetic ketoacidosis (DKA). This study aimed to examine whether haemostatic changes that could lead to a thrombotic tendency occur during ketoacidosis. Plasma levels of in vivo haemostatic markers reflecting activation degrees of the coagulation system, fibrinolytic system, platelets and endothelium were assayed in 34 patients with DKA, both at diagnosis and 1 week after recovery. We found coagulation system and platelet activation and endothelial injury/activation in the patients at diagnosis of DKA. Although significant improvements were observed after recovery, only platelet activity was completely normalized. Fibrinolytic activity was also increased, both at diagnosis and after recovery, compared to the control group. However, although coagulation activity was prominently increased at diagnosis compared to the recovery period, there was no change in fibrinolytic activity in the same periods; on the contrary, the fibrinolytic capacity of the endothelium was diminished at diagnosis of DKA compared to the recovery period, suggesting the presence of relative hypofibrinolysis during DKA. Indications for a role of hyperglycaemia in the emergence of haemostatic disturbances during DKA were observed.

Unchanged global fibrinolytic capacity despite increased factor VIIa activity in Behçet's disease: evidence of a prethrombotic state

Abstract.The aim of this study was to evaluate coagulation and fibrinolytic systems in Behçets disease (BD). Accordingly, various parameters of the coagulation and fibrinolytic systems were investigated in 39 patients with BD and 31 age- and sex-matched healthy volunteers as the control group. Seven of these patients with BD had histories of thrombotic complication. Three were found to have decreased protein S activity, and one patient had diminished protein C activity. Each of those patients had experienced a thromboembolic event. Activated protein C resistance was present in two patients, one of whom had had a thromboembolic episode. Activated FVII (FVIIa), fibrinogen, and cholesterol levels were significantly higher in patients with BD than in the control group. In the patient group, plasma FVIIa level was inversely correlated with age. Plasma global fibrinolytic capacity (GFC) did not differ between the patients and control group. No statistically significant difference was found in the GFC and FVIIa levels between patients with and without histories of thrombosis. Although the coagulation system was activated in vivo in patients with BD, there was no reactive activation in the fibrinolytic system to counteract the activated coagulation system. These findings suggest a relative hypofibrinolytic state in BD.

Successful treatment of rheumatoid arthritis is associated with a reduction in serum sE-selectin and thrombomodulin level

The aim of this study was to investigate the changes in serum levels of endothelial cell injury markers, soluble (s) E-selectin and thrombomodulin (TM), in patients with rheumatoid arthritis (RA) before and after antirheumatic drug treatment and to assess the relationship between these changes and clinical responses to the drug treatment. Eleven patients with RA having active arthritis and 12 healthy volunteers were enrolled in the study. They were monitored by clinical and laboratory parameters while receiving a combination of methotrexate, hydroxychloroquine and sulphasalazine. Pre- and post-treatment clinical and laboratory parameters, including sE-selectin and sTM levels, were measured. The ages of the patients were comparable with those of the control groups. Significant improvements were detected in erythrocyte sedimentation rate, C-reactive protein, hemoglobin, morning stiffness, patients’ global assessment, physicians’ global assessment, number of tender joints and number of swollen joints improved at the end of the therapy (for each parameter p<0.05). Significant improvements were detected in clinical and laboratory parameters. In the patient group there were significant decreases in the levels of sTM and sE-selectin after treatment (p<0.05). The patient group had significantly higher sTM and sE-selectin levels than the control group at the beginning of the study (p<0.01), but the difference returned to normal after the treatment (p>0.05). The sE-selectin and sTM levels significantly correlated with each other, and also with clinical and laboratory findings. Combination treatment successfully treated RA patients. sE-selectin and sTM levels probably reflect disease activity and can be helpful in monitoring disease status and response to therapy.

Anemias in Chronic Liver Diseases

Anemia is a frequently observed manifestation during the clinical course of chronic liver disease. In this study, we retrospectively reviewed the hospital files of 500 chronic liver disease patients and assessed the frequency, etiology and morphology of anemia in 50 patients who fulfilled the criteria to be included in the study. The mean age of the patients was 48 ± 16 years and male/female ratio was 1.4/1. The mean hemoglobin value was 9.54 ± 2.03 g/dl. The mean MCV was 82.9 ± 10.52 fl. Iron deficiency anemia, defined as absent bone marrow iron stores, was the most common anemia present in 50% of patients. Classical laboratory criteria used in the diagnosis of iron deficiency anemia (MCV < 80 fl, ferritin < 10 ng/ml) could not be applicable to all of the patients with iron deficiency anemia and hepatic disorders. Hemolytic anemia due to hypersplenism was the second most common anemia (24%) followed by anemias, namely anemia due to gastrointestinal hemorrhage (22%), anemia of chronic disease (8%), β-thalassemia major (8%), folate deficiency (6%), vitamin B12 deficiency (4%), macrocytic anemia (2%), aplastic anemia (2%) and immune hemolytic anemia (2%). Twenty-eight percent of the patients had more than a single cause of anemia. Morphologically, microcytic anemia was the most common seen in 46% of the patients followed by normocytic (42%) and macrocytic anemia (12%). As patients do not always present with classical laboratory findings and may have more than a cause of anemia, a complex diagnostic approach should be considered in anemic patients with hepatic disorders.

Circulating Thrombopoietin in Clonal Versus Reactive Thrombocytosis

Introduction: The aim of this study is to assess circulating thrombopoietin concentrations in patients with both clonal and reactive thrombocytosis (RT), which are two distinct categories of extreme platelet production circumstances. Investigation of the thrombopoietin levels in clonal versus reactive thrombocytosis may help us to understand the interactions of this key regulatory cytokine and the conditions in which abnormally increased platelet formation exist. Materials and methods: Thrombopoietin levels were measured in patients with platelet counts greater than 500 × 103 μl−1 The study population consisted of 21 patients with RT (13 with iron deficiency anemia, and 8 with rheumatoid arthritis), 24 patients with clonal thrombocytosis (six with essential thrombocytosis, three with myelofibrosis, eight with chronic myelogenous leukemia, and seven with polycythemia vera (PV)) and 16 healthy subjects were used as controls. Results: The median plasma thrombopoietin concentration was 100.5 pg ml−1 in patients with RT, 467pg ml−1 in patients with clonal thrombocytosis and 62.65pgml−1 in the control group. The thrombopoietin concentration was found to be higher in the patients with primary thrombocytosis when compared to the control group (p = 0.001), as well as in patients with RT (p = 0.002). However, there was no statistically significant difference between the patients with RT and the control group (p = 0.14). There was no correlation between thrombopoietin levels and the platelet counts in patients with clonal thrombocytosis, including essential thrombo- cythemia (ET). Conclusion: Increased levels of thrombopoietin were found in patients with clonal thrombocytosis versus patients with RT and control subjects as well. Defective clearance of thrombopoietin by megakaryocytes and platelets due to a reduced number of thrombopoietin receptors may be the causative mechanism behind this. These results indicate that plasma thrombopoietin levels may be helpful in distinguishing between clonal and reactive thrombocytosis.

Cytokines, endothelium, and adhesive molecules in pathologic thrombopoiesis.

Clonal thrombocytosis (CT) associated with myeloproliferative disorders (MPD) is believed to be secondary to autonomous unregulated platelet production. Secondary or reactive thrombocytosis (RT) can be observed in a number of clinical circumstances and may be related to persistent production of some thrombopoietic factors acting on megakaryocytes (MK). The goal of this study is to assess the serum concentrations of these cytokines in control subjects and patients with MPD associated with thrombocythemia, RT, and autoimmune thrombocytopenic purpura (ATP). Eleven patients with MPD, five with chronic myeloid leukemia (CML), three with polycythemia vera (PCV), two with essential thrombocythemia (ET), one with myelofibrosis, 15 with RT, eight with ATP, and 12 healthy volunteers were enrolled in the study. Serum interleukin (IL)-1β, IL-6, tumor necrosis factor-alpha (TNF), fibronectin, intracellular adhesion molecule-1 (ICAM-1), and thrombomodulin (TM) were measured in these groups. Interleukin-1β, IL-6, and TNF levels were high in patients with RT and ATP, suggesting that these cytokines act on early uncommitted progenitors, promoting commitment along the MK lineage and leading to thrombocytosis or compensation for thrombocytopenia. TM was significantly increased in patients with MPD compared to all other groups, probably indicating the presence of subclinical endothelial damage. Fibronectin levels were high in MPD and RT patients. This finding can be secondary to high platelet turnover in these patients. We found that ICAM-1 levels were high in patients with clonal thrombocytosis. ICAM-1 can be one of the factors initiating the events ultimately leading to clonal thrombocytosis. Thrombocythemia associated with MPD is an autonomous phenomenon not regulated by cytokines.

Bone Marrow and Peripheral Blood C-kit Ligand Concentrations in Patients with Thrombocytosis and Thrombocytopenia

Abstract C-kit ligand (stem cell factor, SCF) is a hematopoietic growth factor with diverse effects. It has stimulatory effects on megakaryocytopoiesis acting in synergism with interleukin-3 (IL-3), thrombopoietin (TPO) and granulocyte-macrophage colony stimulating factor (GM-CSF). The relationship between SCF and megakaryocytopoiesis, especially the correlations between blood and bone marrow SCF levels have been not clearly established in the literature. We therefore, investigated peripheral and bone marrow SCF levels in patients with thrombocytosis and thrombocytopenia. Subjects were divided into three groups: those with (i) thrombocytopenia, (ii) thrombocytosis and (iii) healthy adults as controls. When the three groups were compared, the mean peripheral blood SCF level of the thrombocytosis group (2149±197) was significantly higher than the thrombocytopenia (1586±178) and normal control groups (1371±68; p<0.05) and the bone marrow SCF level was higher (2694±267) than the thrombocytopenia group (1700±182; p<0.05). In the correlation analysis, considering all the groups together the bone marrow and peripheral blood SCF concentrations were positively and significantly correlated (p<0.01; r=0.93). Correlations between platelet number and both bone marrow SCF concentration (p<0.01; r=0.51) and peripheral blood concentrations (p<0.01; r=0.40) were also shown. Our results indicate that SCF is operative in the pathological megakaryopoiesis of clonal origin and reactive thrombocytosis both in the local bone marrow microenvironment and the peripheral circulating blood. We feel that further studies on the platelet-SCF relationship and SCF levels in different disease states are required.

Thrombosis and hemorrhage in chronic myeloproliferative disorders.

Bleeding and thrombosis are frequent complications in myeloproliferative disorders (MPD) and are associated with a high morbidity and mortality (1,2). Increased platelet counts, qualitative abnormalities of MPD platelets, elevated hematocrit levels, and patient’s age are regarded as risk factors for hemostatic complications commonly observed in MPD (1-3). The aim of this study was to determine the incidence of thrombohemorrhagic complications in MPD and to assess whether there are any correlations between these risk factors and the complications. Accordingly, we retrospectively reviewed the clinical course of 143 patients with chronic MPD admitted to our hospital during the last 5 years. The study group consisted of 80 patients with chronic myeloid leukemia (CML), 26 patients with idiopathic myelofibrosis (MF), 24 patients with polycythemia rubra