PADGEM (GMP140) is a component of Weibel-Palade bodies of human endothelial cells

PADGEM protein (PADGEM), also known as GMP140, is a platelet alpha- granule membrane protein that is translocated to the external membrane after platelet activation. Although the biosynthesis of this protein was originally thought to be confined to megakaryocytes, the synthesis of PADGEM in endothelial cells was recently demonstrated (McEver et al: Blood 70:1974a, 1987). We now describe the subcellular localization of this protein in endothelial cells. Immunofluorescence staining of permeabilized human umbilical vein endothelial cells with KC4, a well characterized monoclonal antibody to PADGEM, showed positively stained elongated structures similar in distribution and shape to Weibel-Palade bodies. Their identity as Weibel-Palade bodies was confirmed by double label immunofluorescence using KC4 and a polyclonal antiserum to von Willebrand factor (vWf), a protein known to be specifically stored in these organelles. All Weibel-Palade bodies were found to contain PADGEM. In contrast to strong perinuclear staining produced with anti- vWf antibodies, no significant perinuclear staining was obtained with KC4, indicating that relatively little PADGEM is present in the endoplasmic reticulum and in the Golgi apparatus. In endothelial cells treated with secretagogues that stimulate vWf release the elongated structures positive for PADGEM disappeared, further identifying these structures as Weibel-Palade bodies. This observation extends the parallels between Weibel-Palade bodies and alpha-granules and suggests a possible functional association between vWf and PADGEM.     

Simultaneous immunoaffinity purification of O6-methyl, O6-ethyl-, O6-propyl- and O6-butylguanine and their analysis by gas chromatography/mass spectrometry

A sensitive and specific method has been developed for the simultaneousanalysis of different O⁶-alkylguanines. The cross-reactivityof two different antibodies raised against O⁶-methylguanosineand O⁶-butylguanosine for a series of O⁶-alkylguanines was exploitedfor the immunoaffinity purification of biological samples beforequantitative analysis by gas chromatography/mass spectrometry.The method can be applied to the detection of O⁶-alkylguaninesin DNA and appears to be useful for studying chemical carcinogenmechanisms in animals and possibly for the detection of humanexposure to alkylating agents.     

Multiple endocrine cell types in thyroid medullary carcinoma

Summary 10 cases of thyroid medullary carcinoma (TMC) have been studied ultrastructurally and histochemically. Well differentiated calcitonin-producing C cells were present in all tumours, being prevalent in 9 cases. 5-Hydroxytryptamine (5HT) storing cells were found in two cases, somatostatin immunoreactive cells in at least 5 cases and ACTH-immunoreactive cells in 4 cases. Ultrastructurally, at least 3 types of apparently non-C cells were observed. Type 1 cells with large, poorly osmiophilic granules resembling those of gastroenteropancreatic D cells, were present in 6 cases; they appeared to correlate well with somatostatin immunoreactive cells. Type 2 cells with large osmiophilic granules were found in 5 cases; they resembled ACTH-MSH cells of the human pituitary and may correspond to the ACTH-immunoreactive cells of light microscopy. Type 3 cells with small granules and an unknown function were found in 6 cases, always in scarce number. It is concluded that TMC, although mainly made up of C cells, usually contains large proportions of other endocrine cell types.Supported in part by grant N. 75.00630.04 from the Italian National Research Council (C.N.R.). P.F. is a fellow of the Fondazione Anna Villa Rusconi, Varese     

Expression of gamma interferon on circulating lymphocytes in viral infections.

Peripheral blood lymphocytes from a total of 111 patients and 40 healthy individuals were studied for gamma interferon (IFN-gamma) expression on their surfaces by indirect immunofluorescence assay and flow cytometry, with a new anti-IFN-gamma monoclonal antibody (IGMB-14) as a specific reagent. Of 64 patients with proven acute viral infections, 59 had a significantly higher percentage of lymphocytes expressing IFN-gamma on their membranes than healthy individuals did. On the other hand, only 3 (8.9%) of 34 patients with proven bacterial infections had an increased percentage of IFN-gamma-expressing lymphocytes. None of the eight patients with other infections and none of the five with systemic lupus erythematosus showed an increased percentage of IFN-gamma-positive lymphocytes. The percentage of IFN-gamma-expressing lymphocytes during a viral infection was found to be related to different stages of the disease. Finally, some applications of this rapid IFN-gamma assay method in viral diseases are discussed.     

A new approach to the diagnosis of central venous catheter sepsis

One hundred forty-four cancer patients harboring a central venous catheter (CVC) were prospectively investigated to assess the relationship between hub culture, clinical assessment of sepsis before removal, and CVC sepsis. In 22 patients, the CVC was removed because of clinical assessment of catheter sepsis expressed by the staff prior to the removal. For each CVC removal, peripheral blood (qualitative method), hub, and CVC tip (quantitative method) cultures were performed. Clinical sepsis (disappearance of fever after CVC removal) was observed in 13 patients, microbiologic "sepsis" (identification of the same microorganisms on the CVC tip and in the peripheral blood) in seven patients, and clinical and/or microbiologic sepsis in 16 patients. Staphylococcus epidermidis was the microorganism most frequently identified. Hub culture was negative in 48% and positive for a low number and a high number of colonies in 35% and 17%, respectively. The predictive value of hub culture was 96% when testing negative and 8% and 37% (p = 3 x 10(-3)) when testing positive for a low and a high number of colonies, respectively. Predictive values of clinical assessment were 55% if positive and 97% if negative. Combining hub cultures and clinical assessment, the risk of sepsis varied from 2% with both evaluations negative to 89% in the case of positive clinical assessment associated with positive high-count hub. Inasmuch as the CVCs used have a disposable hub, it is possible to have an accurate diagnosis of CVC sepsis without removing the CVC.     

Nicotinamide and insulin secretion in normal subjects

Summary Nicotinamide has been given both before and after clinical onset of Type 1 (insulin-dependent) diabetes mellitus in an attempt to prolong beta-cell survival. Nicotinic acid, structurally similar to nicotinamide, induces insulin resistance and increases insulin secretion in healthy individuals. It is not known if nicotinamide has similar effects. Since insulin secretion, as measured by the acute insulin response to intravenous glucose, is used to predict diabetes and to monitor therapy, the effects of nicotinamide must be established before trials in individuals at high risk of progression to Type 1 diabetes can be interpreted. Intravenous tolerance tests were performed according to the ICARUS standard protocol in 10 healthy, adult subjects (age 32±5.7 years) before and after 14 days of treatment with nicotinamide 25 mg · kg^–1 · day^–1. The acute insulin response after nicotinamide did not differ from the control study, whether measured as the incremental 0–10 min insulin area (278±142 vs 298±130mU · l^–1 · 10 min^–1) or as the 1±3 min insulin level (78±39 vs 81±44 mU/l). The late insulin response was equally unaffected, as were basal insulin (5.2±1.6 vs 5.6±2.1 mU/l) and glucose (5.0±0.4 vs 4.9±0.2 mmol/l) levels and glucose disposal rates (1.98±0.88 vs 2.04±0.68%/min). Nicotinamide does not affect insulin secretion and glucose kinetics in normal subjects, confirming its suitability for trials designed to delay or prevent the onset of Type 1 diabetes.     

Thrombopoietic properties of 5-methoxytryptamine plus melatonin versus melatonin alone in the treatment of cancer-related thrombocytopenia

Recent studies have shown that the hematopoietic system is under neuroendocrine control. In particular, thrombopoiesis has been proven to be stimulated by melatonin, and the pineal indole has been shown to be effective in the treatment of thrombocytopenia resulting from different causes. At present, however, there are no data concerning the possible thrombopoietic activity of pineal indoles other than melatonin. The present study was carried out to evaluate the effect of a concomitant administration of the pineal indole 5-methoxytryptamine in patients with cancer-related thrombocytopenia who did not respond to melatonin alone. The present study included 30 patients, who were randomized to receive melatonin alone (20 mg/day orally in the evening) or melatonin plus 5-methoxytryptamine (1 mg/day orally in the early afternoon). A normalization of platelet count was achieved in 5/14 (36%) patients treated with melatonin plus 5-methoxytryptamine and in none of the patients treated with melatonin alone (P < 0.05). Moreover, mean platelet number significantly increased only in the patients treated with melatonin plus 5-methoxytryptamine. This preliminary clinical study would suggest that 5-methoxytryptamine, a pineal indole, may also exert thrombopoietic activity. Further studies, however, will be required to establish whether 5-methoxytryptamine may play a direct thrombopoietic activity, or whether it may act by improving melatonin's efficacy.     

Non-O blood type is the commonest genetic risk factor for VTE: results from a meta-analysis of the literature

It is well known that the ABO blood group exerts a major influence on hemostasis, as O blood group individuals have lower von Willebrand factor and factor VIII levels than non-O blood group subjects. To evaluate the possible clinical implication of the different ABO blood groups on the risk of developing venous thromboembolism (VTE), we conducted a meta-analysis of the existing literature. After an electronic search strategy using Medline and Embase and a manual review of abstract books of the International Society on Thrombosis and Haemostasis and of reference lists of all retrieved articles, we included in the systematic review 38 studies with 10,305 VTE cases. The prevalence of non-O blood group was significantly higher in VTE patients compared with controls with a resulting pooled odds ratio (OR) of 2.09 (95% confidence interval [CI], 1.83, 2.38; p < 0.00001). Similar findings were obtained when the genotypes A1O/BO/A2B (OR 1.73, 95% CI, 1.47, 2.05; p < 0.00001) and A1B/A1A1/BB (OR 1.87, 95% CI, 1.84, 2.44; p < 0.00001) were analyzed. The maximum VTE risk was observed in non-O-factor V Leiden patients (OR 7.60, 95% CI, 3.21, 17.99), while for G20210A prothrombin mutation it was not possible to perform a pooled analysis due to a paucity of published studies. Finally, the association between non-O blood group and VTE was weaker when provoked VTE cases were considered (OR 1.33, 95% CI, 1.18, 1.50), while it was substantially unchanged when unprovoked VTE cases were analyzed (OR 1.88, 95% CI, 1.42, 2.50). In conclusion, considering its prevalence, non-O blood group is a candidate to be one of the most important genetic risk factors for venous thrombosis.