Inhibition of adhesive and signaling functions of the platelet GPIb-V-IX complex by a cell penetrating GPIbα peptide

BACKGROUND: Interaction between the platelet glycoprotein (GP)Ib-V-IX complex and von Willebrand factor (VWF) is critical for initiating platelet-vessel wall contacts, particularly under high shear conditions. This interaction also plays an important role in initiating platelet activation through the generation of intracellular signals resulting in platelet shape change and integrin alpha(IIb)beta3 activation. OBJECTIVE: A cell-penetrating peptide strategy was used to study the role of the intracellular domain of the GPIbalpha subunit in VWF/GPIb-V-IX-dependent adhesion and activation. METHODS: Peptides of 11-13 amino acids, covering the 557-610 region, were coupled to a nine-arginine permeating tag (R9) and the effects of their cell entry on VWF-dependent responses were analyzed. RESULTS: The R9alpha557 peptide corresponding to the 557-569 segment reduced platelet agglutination in response to VWF, while the other peptides had no effect. The decreased platelet agglutination appeared to be an indirect consequence of adenosine diphosphate release as a normal response was restored by apyrase or a P2Y1 receptor antagonist. A more direct effect of R9alpha557 on GPIb VWF-dependent functions was observed in adhesion studies on a VWF matrix, where it decreased platelet adhesion and profoundly inhibited filopodia formation. In addition, cell adhesion was reduced and shape change absent when Chinese hamster ovary cells expressing the GPIb-IX complex were incubated with R9alpha557. CONCLUSION: This study performed in intact platelets suggests a functional role of the 557-569 domain of GPIbalpha in controlling VWF-dependent adhesion and signaling.     

Effect of enprostil on the gastroduodenal mucosa of healthy volunteers

Enprostil is a synthetic dehydro-prostaglandin E2 with gastroduodenal ulcer-healing and mucosal-protective properties. One hundred and three healthy volunteers were randomized to receive capsules of enprostil 35 micrograms b.d. (the clinically recommended dose), enprostil 70 micrograms b.d., or placebo b.d. All underwent endoscopic assessment of the gastroduodenal mucosa, scored using a 0-4 scale, at baseline and on Days 3, 7, 14, 21 and 28 of dosing. Mean and median maximum scores demonstrated a dose response, and the mean maximum scores were statistically significantly higher for both enprostil groups on each endoscopy day when compared with placebo. The majority of enprostil-treated subjects had petechial haemorrhages. The proportion of volunteers with small white-based mucosal breaks (erosions) was significantly higher for the fundus in the enprostil 70-microgram group on Days 21 and 28 when compared with placebo, but there were no significant differences between treatment groups for any area on the other study days. The 70-microgram dose was associated with significantly more gastrointestinal adverse events than the 35-microgram dose, which was similar to placebo. There were no significant differences between groups for large white-based mucosal breaks (ulcers). We conclude that oral enprostil produced gastric mucosal petechial haemorrhages, primarily in the fundus of the stomach. Gastric mucosal petechial haemorrhages are probably without clinical significance because they are very common in the general population (10-15%) and do not progress to erosions and ulcers.     

Ventricular Parasystole: A Chronobiologic Study

Previous works have reported circadian rhythms for several cardiovascular parameters. A chronobiologic rhythm is characterized by: mesor (a rhythm-determined average), amplitude (half difference between the highest and lowest values), and acrophase (timing of high point in degrees and/or in hours) along with 95% confidence limits. We performed 24-hour ECG Holler monitoring in seven patients (mean age, 50.6 years) with ventricular parasystole (VP) in order to determine whether the chronotropic activity of parasystolic foci has a circadian rhythm similar to that of the sinus node. For each Holter recording parasystolic rates (PRs) and heart rates (HRs) were calculated every hour. Furthermore, a mean hourly PR and a mean hourly HR were calculated from the hourly PRs and HRs of the patients. The statistic chronobiologic analysis was done by single and mean cosinor methods. Correlation between mean hourly PR and HR was evaluated by Pearson's V coefficient. A statistically significant rhythm (P < 0.05) was found for the single and mean rhythms both of HR and PR. In our patients, HR had acrophase at 1.27 P.M., mesor at 73.28 beats/min, and amplitude at 9.53 beats/min, whereas PR had acrophase at 1.42 P.M., mesor al 38.31 beats/min, and amplitude at 3.64 beats/ min. Chronobiological data and the high direct correlation between mean hourly HRs and mean hourly PRs (r = 0.96, P < 0.001) indicate a similar circadian variability of the chronotropic activity of sinus nodes and parasystolic foci. Although several hypotheses can be made, responsiveness of parasystolic foci to circadian variations of the autonomic nervous system tone (sympathetic and/or vagal) and/or circulating substances (particularly catecholamines) seems the more probable one for explaining our findings.     

Multigenetic lesions in infant acute leukaemias: correlations with ALL-1 gene status

The large majority of childhood B-precursor cell acute lymphoblastic leukaemia cases present IgH and TCRδ gene rearrangements. These rearrangements have been widely used as specific markers for monitoring minimal residual disease. However, their prognostic value still remains unclear. In order to determine whether IgH and TCRδ gene rearrangements have any influence on relapse and event-free survival (EFS), we analysed the clinical impact of these genetic characteristics in 51 B-precursor acute lymphoblastic leukaemia patients. 46/51 patients (90.2%) showed IgH gene rearrangements by Southern blot and/or polymerase chain reaction (PCR) analysis. No statistically significant associations were found between IgH gene rearrangement pattern and age, sex, WBC count, immunophenotype, risk factor, relapse or EFS. 27/41 patients (66%) showed Vδ₂Dδ₃ recombination by Southern blot and/or PCR analysis. At a median follow-up of 53 months the estimated 5-year EFS probability was 78 ± 3% for the whole group. The EFS probability among patients with a Vδ₂Dδ₃ recombination pattern in the TCRδ locus was 90 ± 3%, whereas for patients without Vδ₂Dδ₃ recombination was 39 ± 13% ( P  < 0.005).
IgH rearrangement patterns do not appear to influence relapse or EFS probability. However, TCRδ gene rearrangement patterns have a relevant impact on the relapse rate and the EFS probability. Patients with Vδ₂Dδ₃ recombination have better clinical outcome than patients without this recombination, independent of any other prognostic factors.