Induction of functional inducible nitric oxide synthase in monocytes of patients with congestive heart failure. Link with tumour necrosis factor-alpha.

AIMS: We studied the induction of monocytic inducible nitric oxide synthase expression and the tumour necrosis factor-alpha system in patients with congestive heart failure. METHODS AND RESULTS: Forty-three congestive heart failure patients and 15 healthy subjects were studied. Antigenic tumour necrosis factor-alpha and its soluble receptors, measured by ELISA, were increased in chronic heart failure and the increase was related to the clinical severity of the syndrome (tumour necrosis factor-alpha from 8.2+/-5.2 in NYHA class II to 18.2+/-7.2 in class III and 26.9+/-13.2 pg. ml(-1)in class IV, P<0.0001 classes III and IV vs class II; soluble tumour necrosis factor receptor I from 1.0+/-0.2 in class II to 2.3+/-1.1 in class III and 5.5+/-3.2 ng. ml(-1)in class IV, P<0.0001 classes III and IV vs class II; soluble tumour necrosis factor receptor II from 2.7+/-0.7 in class II to 4.9+/-1.9 in class III and 8.4+/-5.0 ng. ml(-1)in class IV, P<0.002 classes III and IV vs class II). Monocytic inducible nitric oxide synthase assessed by Western blot, was expressed only in congestive heart failure patients (13 out of 43). The association among monocytic inducible nitric oxide synthase expression, tumour necrosis factor-alpha system activation, neurohormones and other clinical parameters was studied. The univariate logistic regression showed that inducible nitric oxide synthase expression was strictly associated with NYHA class (P<0.05), antigenic tumour necrosis factor-alpha (P<0.01) and its soluble receptors (P<0.05). The multivariate analysis showed that antigenic tumour necrosis factor-alpha was the only predictor for monocytic inducible nitric oxide synthase expression (P<0.05, RR=2.75, CI 1. 34-5.43). CONCLUSIONS: Inducible nitric oxide synthase is expressed in circulating monocytes of patients with severe congestive heart failure. This phenomenon is linked to the activation of the tumour necrosis factor-alpha system.

[Activation and role of the tumor necrosis factor-alpha in congestive heart failure]. / Attivazione e ruolo del fattore di necrosi tumorale alfa nello scompenso cardiaco congestizio.

Recently, an activation of the immune system has been demonstrated in congestive heart failure (CHF). Aim of this study was to evaluate the effects of CHF on the activation of alpha tumor necrosis factor (TNF-alpha), a pleiotropic cytokine. Since the soluble forms of the TNF membrane receptors, sTNF-RI and sTNF-RII, have been shown to modulate TNF-alpha biological activity, we determined antigenic TNF-alpha, bioactive TNF-alpha, sTNF-RI and sTNF-RII in 52 patients with varying degrees of CHF (NYHA functional class II, III, IV). The etiology of CHF was coronary artery disease in 51% of the patients, idiopathic dilated cardiomyopathy in 38% and valvular disease in 11%. All patients were treated with ACE-inhibitors, digoxin and inotropic agents. Antigenic TNF-alpha was significantly increased in NYHA functional class IV patients (from 12.1 +/- 7.6 to 38.5 +/- 12.4 pg/ml, p < 0.001) whereas cytotoxic activity was always under the detection limit of the assay (100 pg/ml). Soluble TNF receptors were significantly elevated in NYHA functional class IV patients: sTNF-RI increased from 1.27 +/- 0.48 to 4.54 +/- 2.11 ng/ml (p < 0.001) and sTNF-RII from 2.25 +/- 0.55 to 7.78 +/- 2.13 ng/ml (p < 0.001). The possible modulation of TNF-alpha biological activity by the soluble receptors was investigated by means of spiking experiments after addition of 625 pg/ml human recombinant TNF-alpha to each serum sample. The biological activity of the added TNF-alpha was significantly inhibited by the high levels of soluble receptors present in the sera of NYHA functional class IV patients (from 625 to 249 +/- 176 pg/ml, p < 0.001). The results show that TNF-alpha and its soluble receptors are activated in severe CHF. The high concentration of soluble TNF receptors circulating in CHF patients are likely to play a protective role against TNF-alpha biological activity.

Oxidative stress during reperfusion of human hearts: potential sources of oxygen free radicals.

OBJECTIVE: The aim was to examine the role of neutrophil activation in the genesis of oxidative stress during the early phases of reperfusion after ischaemia in patients subjected to aortocoronary bypass grafting. METHODS: Ten selected patients were studied. All had normal ejection fraction and normal left ventricular end diastolic pressures before operation. Each patient required at least three grafts, so that the duration of aortic crossclamping exceeded 30 min, the minimum ischaemic period required to detect oxidative stress upon reperfusion. Oxidative stress was assessed by measuring the formation and release of oxidised glutathione (GSSG) in the coronary sinus 1 min before and 3 min after the start of the cardiopulmonary bypass, and then 1, 5, 10, and 20 min after removal of the aortic clamp, and again 5 and 10 min after the end of the cardiopulmonary bypass. The arterial-coronary sinus difference for neutrophils, elastase-alpha 1 protease complex (elastase), and creatine phosphokinase was also monitored at the same intervals. RESULTS: Before clamping GSSG was undetectable in arterial and coronary sinus blood. There was no significant arterial-coronary sinus difference for neutrophils or elastase [53(SEM 66) cell.ml-1 and 1.10(2.49) micrograms.litre-1, respectively[. Five minutes after re-establishment of coronary blood flow, there was both a release of GSSG into the coronary sinus [arterial-coronary sinus difference: 11(2.6) nmol.dl-1] and an accumulation of neutrophils in the heart [arterial-coronary sinus difference: 262(33), P < 0.01 cell.ml-1], whereas no elastase release from the heart was measured [arterial-coronary sinus difference 7.6(4.46) microgram.litre-1, NS]. The arterial levels of elastase increased progressively during the operation from 48(5) microgram.litre-1 (preclamping) to 405(62) microgram.litre-1, P < 0.01 (end of the cardiopulmonary bypass). CONCLUSIONS: These data indicate that, in man, neutrophils do accumulate in the myocardium during early reperfusion. However, they are not activated when oxidative stress occurs. It is unlikely that the neutrophil localisation in the heart has pathological significance in the production of oxygen free radicals during early reperfusion. Free radical accumulation in the coronary vessels may contribute to disorders of coronary flow associated with reperfusion.

The technicon H6000 automated hematology analyzer in the diagnosis and classification of the myelodysplastic syndromes.

The Technicon H6000 hematology blood counter is a fully automated analyzer which provides, in addition to conventional hematological blood values (i.e. RBC, PLT, etc.), a differential count of WBCs based on a combination of cytochemistry (peroxidase content) and cell volume analysis. Because of these characteristics, the H6000 quantitizes, as part of the full differential count, the number and percentage of "large unstained cells" (LUCs), i.e. large peroxidase-negative circulating elements with a cell volume above a predetermined threshold established for the lymphocytes of normal subjects. We evaluated the H6000 printouts (scattergrams) of 29 patients with myelodysplastic syndromes and of 12 other patients with transient cytopenias. The most important and constant diagnostic features for myelodysplastic syndromes were the increased proportion of LUCs and, in some cases, the high monocyte count, which are both automatically provided by the instrument.

Cefotaxime in the cerebrospinal fluid and serum in patients with purulent meningitis.

Cefotaxime concentrations in the cerebrospinal fluid and serum were determined in patients with purulent meningitis by means of a simple, rapid and reproducible method in agar medium. The CSF concentrations of cefotaxime fluctuated around 4 mg/l. The pharmacokinetics of the antibiotic in relation to the integrity of the blood-brain barrier was studied by means of an assay of the albumin and IgG present in the cerebrospinal fluid and in the serum.

[Plasma cell acid phosphatase in the differential diagnosis of monoclonal gammopathies]. / La fosfatasi acida plasmacellulare nella diagnostica differenziale delle gammopatie monoclonali.

Acid phosphatase activity was studied cytochemically in bone marrow plasma cells of 32 multiple myelomas, 45 non-myelomatous monoclonal gammopathies and 20 normal subjects. We have found significant differences among these three groups (P less than 0.001). The usefulness of this cytochemical reaction for the study of monoclonal gammopathies is discussed.

[Laboratory diagnosis of acute monocytic leukaemias]. / Diagnostica di laboratorio delle leucemie monocitiche acute.

The haemathological features of nine patients with acute monocytic leukaemia are presented. The peripheral blood and bone marrow showed differentiated and apparently undifferentiated blasts. It was necessary to type these blast cells using cytochemical reaction for specific and unspecific esterases. It is very important to differentiate this variety of acute leukaemia from the other non limphoblastic leukaemias for different prognosis and therapeutic assays.

[Laboratory diagnosis of idiopathic myelofibrosis. Review of the literature and presentation of 15 cases]. / Diagnostica di laboratorio della mielofibrosi idiopatica. Rassegna della letteratura e presentazione di quindici casi.

We report on 15 patients affected by myelofibrosis. The main clinical aspects were hepatosplenomegaly, anemia and leukocytosis. Immature cells and anisopoikilocytosis of the erythrocytes were frequently found in the peripheral blood. At bone biopsy, various aspects of panhyperplasia, increase in the reticular fibers, myeloid fibrosis and osteosclerosis were observed. Leukocyte alkaline phosphatase was frequently elevated. The clinical and laboratory course of the disease, causes of death and the possible etiopathogenetic aspects of this disease are considered and discussed.

[The Pelger-Huët leukocyte nuclear anomaly: a morphologic, cytochemical, cytogenetic and ultrastructural study of 39 cases (authors' transl)]. / L'anomalia nucleare leucocitaria di Pelger-Huët: studio morfologico, citochimico, citogenetico ed ultrastrutturale di 39 casi.

Thirtynine of nuclear leukocyte Pelger-Huët anomaly occurring in two Italian families are presented and discussed. The study of the genealogical trees has confirmed that the anomaly is a familial and congenital form, with autosomal dominant inheritance. The incidence of the anomaly in our popolation is 1 to 4,500. In the heterozygous form the anomaly is characterized by decreased segmentation of the nucleus, distinctive nuclear forms and pyknosis of the nuclear chromatin out of proportion of the nuclear area. The results of morphologic, cytochemical, ultrastructural and cytogenetic studies are reported.

[Diagnostic cytochemistry of acute leukemias (author's transl)]. / Diagnostica citochimica di laboratorio delle leucemie acute.

We have described the principal haematological features of 35 cases of acute leukemias, showing the usefulness of cytochemical reactions for the purpose of a right diagnosis and prognosis. The PAS, Sudan Black B and Peroxidase reactions have been very important in the differential diagnosis between lymphoblastic and myeloblastic acute leukemias. The specific and non specific esterases reactions have been useful in the monocytic leukemias diagnosis.