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Objective Estimate the effect of lifestyle adjustment activities in patients with metabolic syndrome treated by prescribed balneotherapy. Methods Observational pilot cohort study with 12-month follow-up after multidimensional lifestyle training (physical, dietary, educational) during 3-week standard stay in the spa town of Eugenie-les-Bains. Results Of 145 eligible patients, 97 were included; 63 were followed and analysable. At inclusion all had ≥ 3 National cholesterol education program-Adult treatment panel III (NCEP-ATPIII) criteria defining metabolic syndrome, 76.2% were female, mean age was 61.2 years. At the end of follow-up (median:10.4 months, Inter-Quartile Range: [6.7;11.4]), 48 of these 63 patients (76.2%) no longer had metabolic syndrome (95%CI [65.7;86.7]). These 48 patients without metabolic syndrome at the end of follow-up represented 49.5% of the 97 included (95%CI [39.5;59.4]). Conclusions Future studies of lifestyle interventions taking advantage of the spa environment can be expected to find least one third of patients free of metabolic syndrome at the end of 12-month follow-up in the intervention group.

Long-term metabolic changes in vein tissue of the limbs and feet have been reported in the course of chronic venous insufficiency (CVI). Low fibrinolytic potential was first demonstrated using fibrin-embedded vein rings (Todd's biopsy), then in blood via increased euglobulin lysis time. This defect is especially pronounced when blood is taken from foot veins of patients with grades 5 and 6 of CVI or skin lesions. Haemorrheological disorders such as increased blood viscosity and red cell aggregation, related mainly to high fibrinogen levels, have also been shown in patients with CVI. Such disorders, particularly observed in blood from foot veins, play an important role in the interaction between the blood cell components and the vessel wall. More recently, the margination and adhesion of white cells to the endothelium and the subsequent activation of white cells have been assumed. Adhesion molecules such as ICAM-1, VCAM-1 and P-selectin have been identified surrounding skin ulceration. There is also evidence that markers of neutrophil degranulation reach high levels in blood taken from foot veins after exposure to experimental venous hypertension. White cell activation results in elevated plasma levels of L-selectin, increased production of oxygen free radicals, pseudopods, CD11b integrin, platelet-monocyte aggregates and abnormal lymphocyte L-selectin. Increased nitric oxide, presumably due to monocyte activation, as well as increased urine haemosiderin related to skin microvessel leakage have also been reported. VCAM-1 and metalloproteinase MMP9 activity have been stated as sensitive markers in relation to endothelium activation and white cell trapping, which follows hypoxia and/or alterations in wall shear stress, mainly in skin venulae and in the region of valves.

Background: Some forms of chemoresistance in leukemia may start from failure of tumour cells to successfully undergo apoptosis and Bcl-2 may play a role in this defect. Therefore, we evaluated the Bcl-2 content and synthesis in relation with the apoptotic potential in leukemic cell lines after anthracycline treatment. Methods: U937, HL60, and K562 cells and their drug resistant (DR) variants were treated with varying concentrations of Idarubicin (IDA). Apoptosis was evaluated by fluorescence microscopy after acridine orange staining. Bcl-2 and Bax content were evaluated either by flow cytometry after indirect immunolabelling or by Western blot. Results: High Bcl-2 contents were not related to a poor ability to undergo apoptosis in U937, HL60, K562 and their DR variants. IDA induced a concentration-dependent increase in Bcl-2 content in all cell lines as long as they do not perform apoptosis. Enhanced Bcl-2 expression was inhibited by cycloheximide, actinomycin D, or antisense oligonucleotide directed against bcl-2 mRNA. Bcl-2 expression was also increased in the resistant U937 variant after serum deprivation or C2-ceramide treatment. The synthesis of Bcl-2 led to an increased Bcl-2/Bax ratio solely in the cells with an apoptosis-resistance phenotype. Conclusions: These data suggest that exposure to IDA induces Bcl-2 expression in leukemic cell lines, and that this mechanism could contribute to apoptosis resistance and participate in the acquisition of chemoresistance. They also confirm that the evolution of the Bcl-2/Bax ratio reflects apoptotic ability better than the steady state level of Bcl-2 expression. Cytometry 36:140–149, 1999. © 1999 Wiley-Liss, Inc.

Expression of the multidrug resistance (MDR) phenotype is responsible for chemotherapy failure in numerous cancers. This phenotype is generally due to the expression of the mdr1 gene-encoded P-gp. Modulation of P-gp activity by chemotherapy has limited possibilities because of toxicity and poor specificity. In contrast, specific transcription blockage of the mdr1 gene can be obtained by oligonucleotides forming a triple helix structure at the DNA level. We used here immunofluorescence and both flow cytometry and image analysis to evaluate surface and total P-gp content in K562 MDR cells. The mdr1 mRNA content was measured by RT-PCR. We confirm the capacity of a 27-mer oligodeoxynucleotide, targeted to an mdr1 DNA fragment, to cause a 10-fold decrease in mdr1 mRNA level. However, this specific genetic inhibition was functionally limited because cellular growth was not modified in a cytotoxic environment. We found that total P-gp content was reduced in resistant cells treated with the mdr1-targeted oligonucleotide, while it remained in high levels on the cell surface, suggesting the existence of a large cytoplasmic pool of P-gp (approximately 50% of the total cellular P-gp). Moreover, when cycloheximide was used for 72 h to suppress protein synthesis, surface P-gp expression showed no decrease, whereas total P-gp was considerably lowered. A rapid 35% decrease in surface P-gp level was reached when resistant cells were treated for 24 h with brefeldin A, an inhibitor of intracellular protein trafficking. Simultaneously, the total P-gp level remained stable, thus indicating a probable accumulation of cytoplasmic P-gp, in agreement with the interruption of protein migration. We propose that the cytoplasmic P-gp pool could be a storage pool consumed for maintaining a steady-state level of surface P-gp. Cytometry could be a useful tool to study such a mechanism of P-gp trafficking and cellular distribution, which could explain the difficulties encountered in achieving stable and rapid effects of MDR reversal with oligonucleotides.

An increasing number of methods are being described to detect apoptotic cells. However, attempts to detect apoptotic cells in clinical samples are rarely successful. A hypothesis is that apoptotic cells are cleared from the circulation by phagocytosis before they become detectable by conventional morphological or cytometric methods. Using LR73 adhering cells as phagocytes in a model ofin vitrophagocytosis, we found that phagocytosis of daunorubicin (DNR)-treated U937, HL60, or K562 leukemia cell lines occurred prior to phosphatidylserine externalization, DNA hydrolysis, chromatin condensation, nuclear fragmentation, or mitochondrial potential alteration. Moreover DNR-treated K562 cells were eliminated by phagocytes while apoptosis was never observed by any of the above methods. By contrast, using a fluorometric batch analysis assay to detect caspase activity in ceramide- or DNR-treated cells (fluorogenic substrate for caspase), we found that caspase activity increased in apoptosis-committed cells before they were detected by flow cytometry or recognized by phagocytes. Similarly a caspase activity increase was detected in circulating mononuclear cells of leukemic patients 15 h after the beginning of anthracyclin treatment. We suggest that recent findings on enzymatic events (caspase activation) occurring in the early events of apoptosis must now allow the development of new markers for apoptosis, irrespective of the morphological features or internucleosomal fragmentation which are late events in apoptosis.

A flow cytometry method has been introduced into the routine investigation of whole bone marrow samples following red blood cell lysis on the basis of a primary CD45/side scatter (SSC) gating procedure. Blast cells were first identified by CD45/SSC gating in 74 cases of acute myeloid leukemia (AML) and the results were compared to a conventional FSC/SSC gating procedure and to MGG-staining smears. The percentages of blast cells in these samples as defined by the morphological analysis of MGG smears correlated better with the values determined by CD45/SSC gating (r = 0.94) than with the blast cell counts recorded with FSC/SSC gating (r = 0.76). These findings were not surprising because while CD45 expression was regularly lower on leukemic blasts than on normal lymphoid and monocytic cells, the FCS/SSC characteristics of these populations were overlapping. In 53 samples, the blast cell populations were also analyzed with a panel of FITC-conjugated monoclonal antibodies that were utilized in double labeling with CD45-PE. We show that the CD45/SSC gating procedure improved phenotypic determination of the blast cells in three ways: (1) by discriminating between leukemic blast cells and residual normal cells; (2) by excluding normal cells from the phenotypic analysis of leukemic blast cells; and (3) by identifying blast cell heterogeneity in many cases of leukemia on the basis of different CD45 display. Moreover, this immunophenotyping procedure on whole bone marrow samples also allowed an efficient discrimination between the various cell lineages and facilitated the analysis of leukemic blasts present in low proportions.

Membrane thrombomodulin (TM) is a very efficient natural anti-thrombin glycoprotein with anticoagulant properties expressed on endothelial cell surface. Circulating plasmatic thrombomodulin (TMp) detected by enzyme immunoassay in plasma is considered as a cell marker of endothelial injury. The TMp levels are increased in many conditions (diabetes mellitus, atheromatous disease...). In cases of collagen vascular diseases, where vascular endothelium damage is suspected, TMp is increased particularly in systemic lupus erythematosus (SLE) and systemic sclerosis (SSc). It is noteworthy that the TMp level is correlated with disease activity. Since TMp is a non specific marker of endothelial damage, it may be of interest as a useful marker for the supervision of these diseases. Further studies are needed on larger series. TMp level change during spontaneous evolution or under treatment will help determine wether TMp is a predictor and prognostic marker of these systemic diseases.

Circulating levels of the adhesion molecule P-selectin (CD62P) are increased in the plasma of patients with atherosclerosis, but its relationship to the anatomical location of symptomatic disease or extent of symptomatic disease is unknown. The influence of the risk factors for atherosclerosis on soluble P-selectin is also unclear. To clarify these questions we analysed plasma samples from 170 patients with symptomatic peripheral vascular disease and 119 asymptomatic controls who were, as a group, age- and sex-matched. Soluble P-selectin (ELISA) was increased in 83 patients with symptomatic disease of the iliac and/or femoral arteries alone (P < 0.05, ANOVA) but not in 37 patients with symptomatic carotid artery disease alone compared with controls. Soluble P-selectin was equally raised in 120 patients with disease at one arterial site and in 50 patients with disease at two or more arterial sites (both P < 0.05) compared with controls. Smoking and atherosclerosis were both independent predictors of raised soluble P-selectin. We conclude that increased soluble P-selectin may have value as a marker of peripheral vascular disease of the iliac and/or femoral arteries in group comparisons only, as the poor discrimination and wide variation of data make comparisons at the individual level difficult.

Resume Nous avons mis au point une methode d'evaluation des alarmes de la formule leucocytaire fournies par les automates de numeration formule sanguine (NFS). Ces alarmes previennent l'operateur de la presence de cellules anormales dont la validation doit etre effectuee par l'examen au microscope. On peut les diviser en deux familles : (1) les alarmes quantitatives qui sont dites positives au dela d'un seuil defini par l'operateur ; (2) les alarmes qualitatives, qui interviennent selon deux modalites : presence ou absence. Nous montrons qu'il est possible de determiner le seuil optimal des alarmes quantitatives au moyen de la methode du “likelihood ratio” (LR) et de definir la capacite des alarmes qualitatives a identifier des types particuliers de cellules anormales au moyen de la valeur predictive d'une resultat positif (PV + pour predictive value). Nous avons teste ces methodes sur automate de NFS : le Cobas Argos 5 Diff (ABX, Montpellier, Groupe Hoffmann La Roche). Pour 1600 echantillons sanguins tous preleves dans le service des maladies du sang, la NFS a ete realisee sur l'Argos et la formule leucocytaire manuelle faite sur 200 elements. Des anomalies de la formule leucocytaire sur lame (myelemie, blastes, lymphocytes atypiques, cellules hyperbasophiles, erythroblastes et tricholeucocytes) ont ete retrouvees dans 597 echantillons. La methode du LR nous a permis de determiner le meilleur seuil de detection des alarmes quantitatives -lymphocutes atypiques (LYA) et grandes cellules immatures (GCI)-. Le calcul de la PV+ nous a permis de definir la capacite des alarmes qualitatives a determiner la presence de cellules anormales. La presence de combinaisons particulieres d'alarmes a pu etre associee avec les metamyelocytes matures (“band cells”) et les cellules blastiques de leucemies aigues lymphoblastiques (LAL) et myeloides (LAM). Les valeurs de sensitibilite, specificite et efficacite de l'Argos sont respectivement 92,1, 53,9, et 68,2. En conclusion, les methodes developpees dans ce travail nous ont permis de metrre en evidence la bonne capacite de l'Argos a detecter les anomalies des leucocytes sanguins et nous semblent de facon plus generale permettre une meilleur validation des alarmes donnees par les differents automates de NFS.

A variety of disorders of erythrocyte, platelet, and polymorphonuclear leukocyte (PMN) functions have been described in diabetes. The phospholipid composition of erythrocyte, platelet, and PMN membranes from controls and from type I and II diabetics was investigated in this study. Phospholipids were determined by densitometry using the molybdenum blue reagent. In diabetics, the relative abundance of phosphatidylethanolamine (PE) increased in all cell types studied, whereas those of sphingomyelin (Sph) and phosphatidylcholine (PC) were decreased in platelets and PMN. The percentage of phosphatidylserine (PS) was reduced in erythrocytes but increased in platelets. The level of Sph in PMN was significantly lower in type I than in type II diabetics. Moreover, the longer the duration of diabetes and the poorer the metabolic control, the greater the decrease in Sph. Rheological parameters, which reflect the behavior of red blood cells (RBC), were correlated with the alteration in PE/PS ratio in these cells.

SummaryPoor anticoagulant response to APC is conveniently screened by a commercially available functional test (Coatest® APC Resistance) allowing identification of APC-resistant patients. These patients may then be genotyped with respect to factor V, the Arg - > Gin mutation being the principle cause of APC resistance. However, determination of phenotype generally precedes that of genotype, and the need for an “abnormality threshold” prompted a study of inter-batch variations and the clinical conditions associated with an altered APC response. The response to APC was assessed twice in plasma from 111 patients using two of four successive kit batches. A modest but significant inter-batch variability was observed. At the same time, we also tested 130 patients with retinal venous occlusion (RVO), 28 patients with glaucoma and 24 normal volunteers. The APCaPTT/aPTT ratio was found to be lower in the presence of elevated thrombin-antithrombin complexes (r = 0.167, p

A flow cytometric method to detect apoptotic cells is described. This method is based on the detection of differences in chromatin condensation with Hoechst 33342 as a probe and the detection of dead cells with propidium iodide as a probe for membrane damage. By this method it was possible to detect, in the same sample and at the same time, intact cells, cells undergoing apoptosis, and dead cells resulting from apoptotic and/or necrotic processes. The method was successfully applied to the detection of apoptotic cells in two human cell models: cultured polymorphonuclear cells and the U937 cell line treated with antitumoral drugs. Staining specificity for apoptotic cells was controlled by cell sorting of the presumed apoptotic population, followed by morphologic examination or DNA analysis of the sorted populations. The usefulness of such a method is discussed in terms of applications in the analysis of heterogeneous clinical samples, populations with low DNA degradation during apoptosis, and cell cycle position of the apoptotic cells.

Cytarabine (Ara-C) is currently used in the treatment of adult acute myeloid leukemia (AML). To predict the results of induction chemotherapy, it could be useful to detect leukemic cells that are resistant to Ara-C in patients with AML. Using a bromodeoxyuridine/DNA (BrdUrd/DNA) staining method in flow cytometry (FCM), we have developed a cell resistance index to Ara-C (RI). The technique has been applied to 121 bone marrow (BM) samples from patients with de novo AML treated by a regimen containing Ara-C and daunorubicin (DNR). Ninety-seven patients achieved a complete remission (CR), and 24 patients did not and were considered drug-resistant (DR). The BM cells collected at diagnosis were cultured for 48 hours and underwent BrdUrd/DNA analysis. Among 25 patients with no or very low proliferative activity (

The antibody Ki67 is currently used to evaluate the proliferative fraction of solid tumors and some hematological malignancies. We have used phytohemagglutinin (PHA)-stimulated peripheral blood lymphocytes as a model to study the entry of quiescent cells into cell cycle and to follow their progress to the next cycle. Flow cytometric analysis of lymphocyte samples stained with the antibody Ki67 and a DNA marker has allowed us to follow the expression of Ki67 antigen (Ki67 Ag) as a function of the position of the cells in the cell cycle. The use of drugs blocking the stimulated lymphocytes in different phases of the cell cycle permitted us to demonstrate that Ki67 Ag expression started from the beginning of the first S phase. The level of Ki67 Ag increased during S phase until mitosis, when its expression was maximal. After division, the cells in G1 phase showed a decrease in Ki67 Ag expression (possibly corresponding to degradation) until they reentered S phase, when the level of Ki67 Ag increased again. The results confirm that the expression of Ki67 Ag is related to the proliferative state of the cells and suggest that it may be used to determine the proliferative cell fraction in hematopoietic tissues.

Hyperhomocysteinaemia is a risk factor for premature atherosclerosis and venous thromboembolic disease. Supplementation with folic acid and vitamin B6 has been shown to decrease plasma homocysteine but data fail to assess an effect on the progression of vascular disease. We measured plasma homocysteine and two markers of endothelial injury (plasma soluble thrombomodulin and von Willebrand factor) at baseline and after 3 months of treatment with folic acid and vitamin B6. After this treatment there was a significant decrease in fasting soluble thrombomodulin (−15 ng/ml, 95%CI 5–22.2). Von Willebrand factor was significantly raised after methionine load at baseline but did not significantly rise after supplementation.

Many clinical studies have evaluated the clinical efficiency of ticlopidine in vascular pathology and in few of these studies the plasma Fg level was determined as a biological parameter and not a primary end point. All these studies are heterogeneous because plasma Fg concentration have been measured using various methods, patients were included at various time after the acute event and were followed over a period of 1 to 24 months of treatment with ticlopidine, patients suffered from various pathology: peripheral arterial disease, cerebrovascular disease, diabetes or polycythemia vera. Despite the heterogeneity of these prospective studies, a general trend indicates a decrease in the plasma Fg levels by 10 to 25% with ticlopidine compared to placebo. This decrease in circulating Fg was associated with clinical improvement, and, when studied, hemorheological modifications (decreases in whole blood and plasma viscosities). The mechanism of ticlopidine action is discussed.

The antigen Ki-67 Ag, regarded as a marker for proliferating cells, was identified as a protein(s) (pKi-67) which can exist free or associated with DNA as evidenced by DNA digestion of cells before or after immunolabeling with Ki-67. The dual nature of this antigen was also supported by reconstitution of Ki-67 Ag from purified DNA and nuclear proteins extracted from the K562 cell line. The immunoreactivity of the resulting complexes was examined in solution using Ki-67 and MIB-1 antibodies. The interaction between Ki-67 or MIB-1 antibodies and pKi-67 was enhanced in the presence of undegraded ds DNA, indicating that ds DNA modulates the conformation of pKi-67 and that the altered conformation of pKi-67 is more reactive than the pure protein to both Ki-67 and MIB-1 antibodies.

The plasma thrombomodulin (TM) level depends on the integrity of the endothelium and the clearance of the molecule. In several different pathological conditions, plasma TM levels increase with damage to the endothelium. We studied plasma TM levels in patients with various localizations of atheromatous arterial disease who had normal serum creatinine levels. Two groups of patients had a single symptomatic localization, which was either peripheral occlusive arterial disease (POAD) or ischemic heart disease (IHD) and a third group of patients had multiple symptomatic localizations (polyvascular). We compared the plasma TM levels with the plasma levels of other specific markers of endothelial cell activation such as: prostacyclin (PGI2), tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor (PAI-1). Plasma TM levels were significantly increased in all three individual groups and when all patients were considered (total patients), as compared with normal controls. When all patients were considered, there was a significant positive correlation between plasma TM levels and t-PA and between plasma TM levels and PGI2. A significant positive correlation was also found between the plasma TM levels and PAI-1 for patients with POAD. Thus, our findings suggest that an increased influx of TM into the plasma may be caused by endothelial cell damage in patients with atheromatous arterial disease. However in our study, the plasma TM levels obtained were similar for all three types of atheromatous arterial disease. Though plasma thrombomodulin is a marker of endothelial cell injury, it cannot be of a clinical interest until its levels are related to the extend of the atheromatous lesions. Moreover, we show that plasma TM is not independent of the usual endothelial cell activation markers.

The cytotoxicity of ara-C derives from an inhibition of DNA synthesis after incorporation of ara-CTP into DNA. The rate of DNA synthesis can be determined from the amount of bromodeoxy-uridine (BrdUrd) incorporated into cells after a short exposure to BrdUrd. We developed a computer program to quantify the inhibition of the rate of DNA synthesis by analysis of the distribution of BrdUrd/ DNA. Inhibition was evaluated in ara-C-sensitive and resistant cells after incubation with different doses of ara-C. An index of resistance to ara-C (RI) was expressed as the ratio of the amount of BrdUrd incorporated into S phase cells incubated with ara-C to that incorporated in the absence of ara-C. In the ara-C-sensitive and resistant HL60 cells, a linear relationship between RI and log ara-C concentration was observed. Small numbers of slightly resistant cells in mixtures of ara-C-sensitive and resistant cells could be determined using this method, making it suitable for clinical use to test the resistance of leukemic cells to ara-C. © 1992 Wiley-Liss, Inc.

Anthracyclines (ANT) are used in the treatment of leukemia and other cancers. These drugs have been shown to intercalate between the strands of DNA. In the present study, we show that the amount of ANT intercalated into DNA can be determined by measuring the fluorescence resonance energy transfer (FRET) between Hoechst 33342 (H33342) and ANT bound to DNA. The transfer efficiency was found to depend on the amount of disposable ANT but was independent of the amount of H33342 bound to DNA over a wide range of H33342 concentrations. The method was adapted for flow cytometric measurement of FRET in whole living cells and was used to evaluate the degree of intercalation of daunorubicin (DAU) and idarubicine (IDA) into DAU-sensitive and DAU-resistant leukemic cell lines. ANT intercalation into DNA was affected by factors which modify the intracytoplasmic concentration of ANT, and it was shown that the action of ANT and the resistance to ANT could not be attributed solely to the intercalative effect of the drugs. The method has advantages over previously described methods and represents a useful complementary tool in studies on the mode of action of ANT and the mechanisms of chemoresistance.

In this issue of Clinical Hemorheology and Microcirculation Norbert Nemeth publishes a well-written analysis of the often strong attraction to arts, and especially to musical arts, of people of medical vocation and possible reasons therefore [1]. The quest to comprehend the human body and soul, to detect our own vital signs and signs of psychosomatic processes materializes especially in the music, the art with the most complex effects on human sensory reception. Some vital signs like appearance, dynamic changes and the seemingly endless movement of blood and blood circulation are used by artists as expression of life and death. The author beautifully describes the analogy between progress and flow of music with the running of blood, the speeding or slowing tempo and even the fatal arrest of circulation. He reminds A.L. Copley who linked scientific work and hemorheology with arts, nicely demonstrated in his design of the Fahraeus Medal [2]. A description of “music in medicine” with the known and scientifically approved effects on biological parameters of circulation and on the nervous system is given as well as a description of “medicine in music”. In a few divisions, physiological and pathological phenomena of circulation and blood, diseases or care taking medical persons are portrayed describing circulation, stream, rhythm, bleeding, and appearance of blood in the more or less brilliant compositions of musical arts. It may be unusual, but we decided that this manuscript should published in this journal because it is worthwhile considering the authors attitude to become one with an artwork can mean a momentary liberation for the mind grown wary of everyday limitations, duties, responsibilities, decisions and questions, contributing to a more balanced life and aiding medical activity.

Hyperchomocysteinaemia is a vascular risk factor. However few data are available as regards the efficacy of vitamin supplementation on the course of hyperhomocysteinaemia-related vascular disease. Since homocystein induces vascular endothelium cell dysfunction in vitro, we investigated the effect of vitamin supplementation on plasma soluble thrombomodulin (sTM) concentration, a marker of endothelium injury. The diagnosis of hyperhomocysteinaemia was assessed by oral L-methionin load. Homocysteinaemia was measured before and 6 hours after the load. The sTM concentration was measured while fasting before and 3-months after supplementation with 5 mg folic acid and 250 mg vitamin B6. The difference between fasting and post-load homocystein concentration decreased from 43.9+/14.9 mol/I before supplementation to 28.1+/-9.4 mol/I after a 3-monthslong supplementation. Then a 37% decrease was obtained in the post load rise in homocysteinaemia. 14 from the 17 patients (82%) no longer had detectable hyperhomocysteinaemia after the 3-months supplementation. The concentration of sTM decreased from 43.6+/19.8 ng/ml to 30.7+/I I ng/ml (p=0.005). After supplementation for 3 months with folic acid and vitamin B6, a significant decrease was obtained in sTM. This may reflect a decrease in injury of the vascular endothelium, sTM might be a useful marker to assess the efficacy of supplementation regiments on hyperhomocysteinaemia-related vascular disease.

Variations of plasma thrombomodulin levels, which is a putative marker of endothelial damage, were measured in 23 cases of systemic lupus erythematosus (without antiphospholipids and/or lupus anticoagulants) and in 7 dermatopolymyositis. This parameter was significantly increased in dermatopolymyositis suggesting the involvement of ischemia in endothelium injury. No change in plasma thrombomodulin levels was observed in patients with systemic lupus erythematosus.

Plasma thrombomodulin (TMp) is proned as a marker of endothelial cell domage. Significantly elevated levels were found in patients with atherosclerosis. Furthermore a positive correlation was pointed out between TMp and prostacyclin (PG12) and tissue-type plasminogen activator (t-PA) which are markers of endothelial cells activation.

Changes in the fluorescence of factors IX and II in the presence of calcium at procoagulant phospholipid interfaces suggest a conformational change of the proteins upon binding. Shifts of transition temperature of phosphatidylcholine—phosphatidylserine mixtures towards those of the pure lecithin component when calcium and factors are added suggest that lateral phase separation does exist. The higher resonance-energy transfer efficiency with charged pyrene-labelled phospholipids leads to the conclusion that within the membrane, factor II and IX binding sites are domains of phosphatidylserine. Calcium-independent phospholipid binding, selective for phosphatidylserine, is well documented and is proposed to be mediated by electrostatic forces between the arginine and lysine residues of the proteins and the negative charges of the phosphatidylserine. The binding of heparin or the synthetic anticoagulant polymer PSSO 2 —Glu with antithrombin or thrombin induces different fluorescence changes in these proteins which traduce local changes in the tryptophan residues of the proteins upon binding. The interaction of heparin with various mixtures of thrombin and antithrombin indicates that heparin does not modify the structure of the preformed thrombin—antithrombin complex, as far as can be seen by spectrofluorimetry.

Une vingtaine de te´trahydro-1,2,3,4 quinazolines substitue´es en 3 originales onte´te´synthe´tise´es. L'e´tude in vitro de leur activite´antiagre´gante plaquettaire vis-a`-vis de diffe´rents inducteurs de l'agre´gation plaquettaire (ADP—collage`ne—acide arachidonique) ae´te´re´alise´e, ainsi que la mesure de leur pouvoir inhibiteur de la libe´ration des constituants plaquettaires et de la synthe`se du throm☐ane A2. Toutes ces mole´cules posse`dent un pouvoir inhibiteur qui, compare´a`celui que manifeste l'aspirine dans les meˆmes conditions, est du meˆme ordre de grandeur ou supe´rieur pour certaines d'entre elles lorsque l'agre´gation est induite par l'ADP.

Rabbits given a hypercholesterolemic diet (500 mg/day) for 6 months and then maintained for another 6 months on a normal diet were found to have developed fibrous lipidic lesions in the aorta. Although circulating platelet levels in these animals were normal there was a reduction in mean megakaryocyte ploidy. The high concentrations of megakaryoblasts in all the sedimentation fractions collected by the ‘STAPUT’ system suggested an increase in megakaryocyte turnover with activation of committed stem cells. In addition, other defects in maturation of megakaryocytes were observed, such as abnormalities in the demarcation membrane system and granule number. These data reveal that defects in megakaryocyte maturation and turnover may occur during the process of reparative fibrosis of the arterial tree following a period of moderate hypercholesterolemic diet in the rabbit.

The in vitro antiaggregating action of paracetamol and ten of its derivatives was studied by observing their action on collagen, adenosine-5 diphosphate (ADP) and arachidonic acid-induced platelet aggregation. It is established that in vitro activity appears not only with paracetamol but also with its positional isomers and its isosteric derivatives; this involves the replacement of oxygen by sulfur and that of the NH group by oxygen. Paracetamol and its derivatives also have an inhibiting effect on the serotonin release and the thromboxane synthesis of collagen-stimulated platelets.