Elevated levels of serum vascular endothelial growth factor in patients with polycythaemia vera

Angiogenesis seems to be a prominent event of myeloproliferative diseases. There are few reported data on angiogenesis and the significance of its stimulator, the vascular endothelial growth factor (VEGF), in polycythaemia vera (PV). We report our observation of elevated serum VEGF levels in patients suffering from PV. Twenty patients with PV and 20 age-matched healthy subjects were enrolled. VEGF levels were measured by a quantitative sandwich enzyme immunoassay. Serum VEGF levels in PV were found to be very significantly higher than in healthy individuals (569.7 +/- 101.2 vs. 164.7 +/- 32.8 pg/ml, p = 0.001). We found no correlation between VEGF and haemoglobin, platelet or leucocyte counts in the patient group. Different therapeutic regimens had no influence on VEGF levels. However, in the control group, we observed a positive correlation between VEGF levels and platelet counts (r = 0.52, p = 0.02). Platelet counts did not differ between patients and healthy subjects. We also evaluated platelet-poor plasma VEGF levels in 10 patients and in all healthy individuals. We found very low levels of VEGF, approximately zero in most cases, in both groups and there was obviously no difference between the two groups. Our results indicate that VEGF is overproduced in PV. However, follow-up studies are needed to verify the role of this factor.     

Homocysteine levels in polycythaemia vera and essential thrombocythaemia

Patients with polycythaemia vera (PV) or essential thrombocythaemia (ET) have an increased risk of arterial and venous thromboembolic complications. Since hyperhomocysteinaemia (HHC) is a risk factor for vascular disease, we investigated the frequency of HHC in these disorders and analysed a possible association of elevated plasma homocysteine levels with vascular complications. In the cohort of 134 patients from Vienna (69 female, 65 male, median age 65.5 years, range 21-91 years) with PV (n = 74) or ET (n = 60), plasma homocysteine levels were significantly higher compared to 134 healthy controls. Median homocysteine level was 12.3 micromol/l (range 3.5-48.4 micromol/l) in patients with PV or ET and 8.9 micromol/l (range 4.8-30.5 micromol/l) in normal controls (P < 0. 0001). In addition to the 134 patients from Vienna, 48 patients (28 female, 20 male; median age 66.5 years, range 24-82) from Vicenza with PV (n = 25) or ET (n = 23) were included to evaluate the impact of HHC on the risk of thrombosis. Of 59 patients with HHC (44 from Vienna and 15 from Vicenza) 18 (31%) had a history of arterial and 10 (17%) of venous thrombosis. Of 123 patients with normal homocysteine levels, 30 (24%) had arterial and 16 (13%) had venous thromboses. The difference between the two groups was statistically not significant. Even though mild to moderate HHC occurred in a larger number of patients with PV or ET and thrombosis, it can presently not be regarded as an additional risk factor for thrombosis.     

Elevated soluble urokinase plasminogen activator receptor in plasma from patients with idiopathic myelofibrosis or polycythaemia vera

Extracellular proteolytic enzymes of the urokinase-type plasminogen activator (uPA) system and the family of metalloproteases play a crucial role in the matrix degradation and tissue remodelling processes characteristic of malignant disorders. The receptor for urokinase plasminogen activator (uPAR) serves to localise and intensify the action of uPA and is expressed on the surface of malignant as well as tumour stromal cells including fibroblasts. A soluble form of uPAR (suPAR) cleaved from its glycosyl-phosphatidylinositol anchor is detected in plasma from healthy individuals and increased levels of suPAR have been found in advanced malignancy, suggesting that suPAR may be a marker of extensive tissue remodelling. In an attempt to clarify whether suPAR might be a marker for bone marrow tissue remodelling we measured plasma suPAR levels in a patient cohort comprising 17 with myelofibrosis (MF), 17 with polycythaemia vera (PV), 15 with essential thrombocythaemia (ET), one with a transitional myeloproliferative disorder evolving from PV and 30 controls. Compared with controls suPAR levels were significantly higher in the patients (P < 0.0001) (median 3.35 vs. 2.32 microg L(-1)). Moreover, in subgroup analyses including patients with MF, PV, and ET, respectively, suPAR levels differed significantly with the highest levels found in patients with MF and PV (MF vs. PV vs. ET; P = 0.0003). When comparing suPAR levels of the individual patient subgroups with controls, only suPAR levels of PV and MF patients were significantly increased (P < 0.0001). Sixty-five percent of patients with PV and MF (22/34) had suPAR plasma values that were above the mean +2 standard deviations (SD) of controls. The concentration of suPAR was significantly correlated to plasma lactate dehydrogenase, thrombomodulin, and complex of tPA:PAI-1 in the patients. There was no difference between patients and controls when comparing plasma uPA levels. Increased plasma suPAR levels in patients with chronic myeloproliferative disorders may reflect increased uPAR production in the bone marrow, leading to enhanced bone marrow remodelling.     

The transcobalamins in polycythaemia vera.

The unsaturated B12 binding capacity (UBBC) of the serum and the binding capacity of each of the 3 vitamin B12 binders--the transcobalamins (TC) I, II and III were determined in 21 patients with polycythaemia vera (PV) during the course of the disease and following treatment, using the recently described charged cellulose filter technique. High serum UBBC due to elevated serum TCIII was found in all patients. TCI was moderately elevated in patients who had leucocytosis with a shift to the left. The changes in serum TCIII and UBBC correlated with the activity of the disease. Chemotherapy resulted in a decrease in TCIII and UBBC. The decrease in TCIII and UBBC folowing chemotherapy may be observed before a decrease in the haematocrit and the leucocyte count occurs. Activation of the disease may be assessed by the elevation of TCIII and UBBC. The onset of acute myeloblastic crisis in 1 patient was associated with a decrease in TCIII and TCI levels and a rise in serum TCII. The determination of TCIII and UBBC may be helpful in differentiating true from secondary polycythaemia.     

Histamine metabolism in polycythaemia vera.

Histamine metabolism was studied in 35 patients with polycythaemia vera (PV) at different stages of their disease and compared with controls and patients with secondary polycythaemia. In addition to blood and urinary histamine the main urinary metabolites of histamine, methylhistamine (MeHi) and 1-methyl-4-imidazoleacetic acid (MeImAA) were measured. In patients with active PV the excretion of MeHi and MeImAA was significantly higher than in controls and secondary polycythaemia, indicating an increased histamine formation. The MeImAA excretion was correlated to the blood histamine level, the degree of blood basophilia, the total white blood count and the spleen volume. The blood histamine level was significantly higher in PV patients compared with controls and secondary polycythaemia. No patients with secondary polycythaemia had an increased blood histamine level. With the bio-assay technique used in this study the urinary excretion of histamine in the PV patients was within the normal range. There was no correlation between the increased histamine formation and "histamine-related symptoms". Pruritus and duodenal ulcer occurred with a similar frequency in patients with and without increased MeImAA excretion. The similarity between the disturbance of the histamine metabolism in PV and that found by other authors in chronic myeloid leukaemia is pointed out.     

Red cell mass,spleen size and plasma erythropoietin in polycythaemia vera and apparent polycythaemia

Most frequently, an elevated packed cell volume (PCV) value arouses the suspicion of a polycythaemic state. The aim of the present work was to assess a few readily available variables which could help the clinician to differentiate between polycythaemia vera (PV) and apparent polycythaemia (AP). During a 5-year period, 31 consecutive newly diagnosed patients with PV were identified, and during a 4-year period 38 consecutive subjects were considered to be afflicted with AP. In each subject: (i) the red cell mass (RCM) and plasma volume were measured, (ii) the spleen size was assessed using gamma-camera imaging, and (iii) the plasma erythropoietin (EPO) concentration was determined. The diagnosis of PV was based upon recently proposed criteria. By definition, all PV patients had absolute erythrocytosis, i.e. the RCM was greater than 25% above the mean normal predicted value for the individual. There was no statistical difference between the plasma volumes for PV and AP patients. However, the mean measured/predicted plasma volume for subjects with AP was significantly lower than the mean for PV. The means for spleen scan areas (posterior and left lateral projections) for AP patients were identical to the mean reference values for our laboratory. As compared to AP, in PV the spleen scan areas were significantly increased, and the lateral spleen scan area was significantly larger than the posterior area. It was also shown that, in contrast to AP, both spleen scan areas were significantly larger in male than in female PV patients. All PV patients had plasma EPO concentrations below the lower reference limit, and in 68% of the patients undetectable EPO concentrations were present. Most AP patients (84%) had EPO values within the reference range; 8% had slightly subnormal, but not undetectable, plasma EPO levels.     

Elevated levels of circulating platelet aggregates and beta-thromboglobulin in scleroderma

Levels of circulating platelet aggregates and plasma beta-thromboglobulin reflect the degree of platelet activation in vascular injury and repair. Both values were evaluated in 38 patients with scleroderma and 18 control subjects matched for age, sex, and race. Circulating platelet aggregates (expressed as percentage of total platelet count) were 7.2 +/- 5.5% (mean +/- SD) in the control group and 31.2 +/- 13% in the scleroderma group (p less than 0.0005). Beta-thromboglobulin levels in the control groups were 20 +/- 10 ng/mL and 72.7 +/- 50 ng/mL in the group with scleroderma (p less than 0.0005). A positive correlation was found between the two values (r = 0.6, p less than 0.0005). Significant reductions in circulating platelet aggregates and beta-thromboglobulin levels were achieved in 10 patients by dipyridamole and aspirin therapy. These results show in-vivo activation of platelets in scleroderma with release of platelet granule constituents. Antiplatelet therapy in adequate doses returned both values to normal; however, its long-term effect on scleroderma is not yet known.     

Elevated plasma levels of beta-thromboglobulin and platelet factor 4 in patients with rheumatic disorders and cutaneous vasculitis

The efficacy of plasma levels of β-thromboglubulin (β-TG) and platelet factor 4 (PF4) as markers of the presence and activity of vasculiditic processes in rheumatic diseases were evaluated, first by serial measurement of their levels in a patient with rheumatoid arthritis and a chronic leg ulcer in the course of treatment, and second in 11 patients with rheumatoid arthritis without cutaneous vasculitis, and in nine patients with a variety of rheumatic diseases with cutaneous vasculitis. In the former, plasma levels of β-TG and PF4 were elevated and slowly reduced in parallel with healing, raised again after relapse, and normalized after disappearance of the leg ulcer. In the latter, both plasma levels were elevated in all of the nine patients with cutaneous vascular lesions and in one of the 11 patients rheumatoid arthritis without skin lesions. Levels of β-TG and PF4 may be useful to estimate the presence of vascular lesions in rheumatic disorders. Received: 5 November 2001 / Accepted: 18 June 2002 Correspondence and offprint requests to: Dr Toshihide Yamamoto, Kishiwada Tokushukai Hospital, 4-22-38 Isonokami-cho, Kishiwada, Osaka 596-0001, Japan. Tel: 81-724-38-8781; Fax: 81-724-37-7395; E-mail: ikyoku@kishiwada.tokushukai.or.jp     

Haemostatic defects in polycythaemia vera

Summary 24 patients with polycythaemia vera were studied as regards platelet function, blood coagulation and fibrinolysis. Platelet investigation showed increased red cell fall-out in the clot retraction test, abnormal results in the platelet adhesiveness and defective platelet aggregation with adrenalin, ADP, collagen and thrombin in frequency decreasing in that order. In a small number of cases the ADP release from platelets and the platelet factor-3 availability tests were abnormal while the bleeding time was normal in all cases. Blood coagulation study revealed various mild abnormalities, namely the screening tests in some cases were slightly prolonged, the mean values of factors V and X as well as the antithrombin-III activity were decreased while the factor-VIII level scattered widely. Fibrinolytic study showed normal plasma fibrinolytic activity but the plasminogen was slightly decreased and the fibrinogen/fibrin degradation products were elevated compared to normal. There was no clear correlation between abnormalities of platelets, coagulation or fibrinolysis tests and bleeding manifestations.

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Zusammenfassung 24 Patienten mit Polycythaemia vera wurden untersucht, und zwar die Funktion der Blutplättchen, die Blutkoagulation und die Fibrinolyse. Untersuchungen der Blutplättchen zeigten einen vermehrten Ausfall der roten Blutkörperchen in dem Gerinnungsretraktionstest, abnorme Ergebnisse der Plättchen-Adhäsivität und Defekte der Plättchenaggregation durch Adrenalin, ADP, Kollagen und Thrombin in abnehmender Häufigkeit in der Reihenfolge der Reagenzien. In einer kleinen Zahl der Fälle waren die ADP-Befreiung von den Plättchen und der Plättchen-Faktor-3-Labilitätstest abnorm, während die Blutungszeit in allen Fällen normal war. Die Untersuchung der Blutkoagulation zeigte geringe Abweichungen, und zwar waren die Screening-Tests in manchen Fällen leicht verlängert, die Durchschnittswerte des Faktors V und X und die Antithrombin-III-Aktivität waren vermindert, während die Werte des Faktors VIII sehr weit voneinander abwichen. Die Untersuchung der Fibrinolyse zeigte eine normale fibrinolytische Aktivität des Plasmas, aber das Plasminogen war leicht vermindert und die Fibrinogenspaltprodukte waren erhöht im Vergleich zu den normalen Fällen. Es wurde kein klarer Zusammenhang zwischen der Abnormalität der Thrombozyten, der Koagulation und der Fibrinolyse im Vergleich zu den Blutungserscheinungen festgestellt.

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Abbreviations AT-III Antithrombin-III - BT Bleeding time - CR Clot retraction - DIC Disseminated intravascular coagulation - f. c. Final concentration - FDP Fibrinogen degradation products - PCV Packed cell volume (haematocrit) - PF-3a Platelet factor-3 availability - PAd Platelet adhesiveness value - PPP Platelet poor plasma - PRP Platelet rich plasma - PT Prothrombin time - PTTK Partial thromboplastin time with kaolin - PV Polycythaemia vera - RT Reptilase time - TT Thrombin time     

Elevated levels of plasma and urine beta-thromboglobulin or thromboxane-B2 as markers of real platelet hyperactivation in diabetic nephropathy.

Serum creatinine, immunoreactive serum and urine beta 2-microglobulin, plasma and urine thromboglobulin, plasma thromboxane-B2 levels and daily protein excretion were determined in 61 insulin-treated diabetic patients, comparing the different patient groups (complication free, nephropathy without and with azotaemia) with control subjects. In the groups of diabetic patients, plasma and urine beta-thromboglobulin (BTG) and plasma thromboxane-B2 levels were higher than in the controls. There was a significant positive correlation between urine BTG and beta 2-microglobulin in the group without complication, and between the plasma BTG and beta 2-microglobulin, and plasma BTG and thromboxane levels in the diabetic group with azotaemia. In contrast to some previous assumptions, the increased level of plasma BTG reflects a real platelet hyperactivation in patients with diabetic nephropathy. At the same time, urine BTG also increases. Determination of urine BTG is more simple with less possibility of methodological error.     

Elevated soluble MUC1 levels and decreased anti-MUC1 antibody levels in patients with multiple myeloma

Soluble MUC1 (sMUC1) levels are elevated in many MUC1(+) cancers. We and others have shown that MUC1 is expressed on multiple myeloma (MM) plasma cells and B cells. In this study, we measured sMUC1 levels in bone marrow (BM) plasma from 71 MM patients and 21 healthy donors (HDs), and in peripheral blood (PB) plasma from 42 MM patients and 13 HDs using an immunoassay that detects the CA27.29 epitope of MUC1. sMUC1 levels were found to be significantly greater (mean 31.76 U/mL, range 5.69 to 142.48 U/mL) in MM patient BM plasma versus HD BM plasma (mean 9.68 U/mL, range 0.65 to 39.83 U/mL) (P <. 001). Importantly, BM plasma sMUC1 levels were related to tumor burden because sMUC1 levels were significantly higher for MM patients with active disease (34.62 U/mL, range 5.69 to 142.48 U/mL) versus MM patients with minimal residual disease (16.16 U/mL, range 5.7 to 56.68 U/mL) (P =.0026). sMUC1 levels were also elevated in the PB plasma of MM patients (32.79 U/mL, range 4.15 to 148.84 U/mL) versus HDs (18.47 U/mL, range 8.84 to 42.49) (P =.0052). Lastly, circulating immunglobulin M (IgM) and IgG antibodies to MUC1 were measured in 114 MM patients and 31 HDs, because natural antibodies to MUC1 have been detected in patients with other MUC1-bearing malignancies. These studies demonstrated lower levels of circulating IgM (P <.001) and IgG (P =.078) antibodies to MUC1 in MM patients compared with HDs. Our data therefore show that in MM patients, sMUC1 levels are elevated and correlate with disease burden, whereas anti-MUC1 antibody levels are decreased.