Serum thrombopoietin levels in patients with aplastic anaemia

Endogenous serum thrombopoietin (TPO) levels were measured in 31 patients with aplastic anaemia (AA) using an enzyme immunoassay with a sensitivity of 20 pg/ml. The median platelet count for all AA patients was 30 ± 29 × 10⁹/l (range 5–102) compared with a median of 284 ± 59 × 10⁹/l (range 148–538) for normal controls. Serum TPO levels were significantly elevated in all patients compared with normals (1706 ± 1114.2, range 375–5000 v 78 ± 54, range 16.5–312.9, P  < 0.0001). There was no correlation between serum TPO levels and the degree of thrombocytopenia in AA patients, but TPO levels were significantly higher in patients who were platelet transfusion dependent than in patients who were transfusion independent ( P  < 0.01). There was a trend for higher TPO levels in patients with severe AA compared with non-severe AA patients. Clinical trials of TPO and a related truncated, pegylated molecule, megakaryocyte growth and development factor (PEG-rHuMGDF), are awaited to determine whether treatment with these drugs will result in increased platelet counts in patients with AA.     

Balloon coarctation angioplasty in adolescents and adults: early and intermediate results.

Twenty-three adolescent and adult patients with native coarctation of the aorta underwent balloon dilatation. Dissection of the aorta developed in one patient. Data were collected on the remaining 22 patients. They ranged in age from 15 to 55 years (mean 23 +/- 9.2 years). Invasive measurement of the peak systolic gradient (PSG) and biplane angiography were performed before and immediately after angioplasty and at follow-up 4 to 48 months (mean 15 months) later. PSG before dilatation was 37 to 100 mm Hg (mean 66.9 +/- 19.9 mm Hg) and decreased to 0 to 30 mm Hg (mean 9.1 +/- 11 mm Hg) immediately after dilatation (p less than 0.001). Restenosis occurred in two patients 6 months after dilatation, and one patient had an incomplete dilatation. These three patients underwent successful redilatation and remained improved 12 to 19 months later. There was no significant change in gradient at repeat catheterization in the remaining 20 patients. PSG was 0 to 20 mm Hg (mean 5.8 +/- 7.2 mm Hg). Angiography showed that a small aneurysm developed in one patient immediately after dilatation and in another 6 months later. Eleven patients were restudied more than once, and no change in gradient or size of the aneurysm was noted at mean follow-up 25 months after dilatation. This study demonstrated that balloon angioplasty is an effective method of treating adolescent and adult patient with native coarctation of the aorta. However, because of the uncertain natural history of aneurysm after dilatation, this procedure should be considered investigational until much longer follow-up times are available.     

The effects of systemic immunization of pulmonary clearance of Pseudomonas aeruginosa

Systemic immunization with gram-negative organisms enhances the subsequent pulmonary clearance of these organisms. We studied the early time course of this phenomenon and related it to the time of appearance of polymorphonuclear leukocytes (PMN) and anti-Pseudomonas antibody in bronchoalveolar lavage (BAL). Mice were immunized intraperitoneally twice, separated by 1 wk, with 10(8) formalin-treated Pseudomonas aeruginosa. Two weeks later, they received an intrabronchial inoculum of 2.9 X 10(6) or 4.6 X 10(7) Pseudomonas organisms. Two, 4, and 6 h later, clearance and total PMN and anti-Pseudomonas antibody in the BAL were assessed. Clearance was enhanced in immunized mice at the lower inoculum. At the higher inoculum, bacteria were growing in lungs of both groups, although they were inhibited in immunized mice. Total PMN in the BAL increased progressively in both groups of mice, but net recruitment was diminished with the high inoculum. There were significant differences in the PMN in the BAL between control and immunized mice with high inoculum. Anti-Pseudomonas IgG first appeared in the BAL at 2 h, anti-Pseudomonas IgM at 6 h. These data suggest that anti-Pseudomonas IgG is an effective early pulmonary opsonin. Further, with high inoculums, immunization may aid pulmonary defenses by diminishing the magnitude of the decrement of PMN in the lung.     

Key role of diacylglycerol-mediated 12-lipoxygenase product formation in angiotensin II-induced aldosterone synthesis

We have shown earlier that the 12-lipoxygenase product of arachidonic acid (AA), 12-hydroxyeicosatetraenoic acid (12-HETE), plays an important role in mediating angiotensin II (AII)-induced aldosterone secretion (J. Clin. Invest. (1987) 80, 1763). In the present study, we have evaluated whether diacylglycerol (DG) is the source of arachidonic acid giving rise to this 12-HETE. Treatment of rat adrenal glomerulosa cells with a DG lipase inhibitor, RHC 80267, which prevents conversion of DG to AA and HETEs, blocked AII-induced aldosterone and 12-HETE formation. In contrast, a DG kinase inhibitor, R59022, which prevents conversion of DG to phosphatidic acid, potentiated AII-induced aldosterone and 12-HETE formation. These two inhibitors block DG metabolism which would be expected to lead to increased DG levels and protein kinase C activity and AII-induced steroidogenesis. However, only R59022 potentiated AII action while RHC 80267 was inhibitory. This suggests that conversion of DG to AA and 12-HETE is important for AII action. Further proof for this was obtained by measuring [3H]AA-labeled DG levels. The combination of the inhibitors significantly potentiated AII-induced DG formation even though this same combination was inhibitory on AII-induced aldosterone and 12-HETE. Thus, the inhibitory effect of RHC 80267 is due to blockade of AA release and not of DG formation. These results suggest that DG plays a dual role in AII action, both as an activator of protein kinase C and as a source of AA for 12-HETE formation.     

Early somatic mosaicism is a rare cause of long-QT syndrome

Somatic mosaicism, the occurrence and propagation of genetic variation in cell lineages after fertilization, is increasingly recognized to play a causal role in a variety of human diseases. We investigated the case of life-threatening arrhythmia in a 10-day-old infant with long QT syndrome (LQTS). Rapid genome sequencing suggested a variant in the sodium channel Na_V1.5 encoded by SCN5A, {"type":"entrez-nucleotide","attrs":{"text":"NM_000335","term_id":"124518660","term_text":"NM_000335"}}NM_000335:c.5284G > T predicting p.(V1762L), but read depth was insufficient to be diagnostic. Exome sequencing of the trio confirmed read ratios inconsistent with Mendelian inheritance only in the proband. Genotyping of single circulating leukocytes demonstrated the mutation in the genomes of 8% of patient cells, and RNA sequencing of cardiac tissue from the infant confirmed the expression of the mutant allele at mosaic ratios. Heterologous expression of the mutant channel revealed significantly delayed sodium current with a dominant negative effect. To investigate the mechanism by which mosaicism might cause arrhythmia, we built a finite element simulation model incorporating Purkinje fiber activation. This model confirmed the pathogenic consequences of cardiac cellular mosaicism and, under the presenting conditions of this case, recapitulated 2:1 AV block and arrhythmia. To investigate the extent to which mosaicism might explain undiagnosed arrhythmia, we studied 7,500 affected probands undergoing commercial gene-panel testing. Four individuals with pathogenic variants arising from early somatic mutation events were found. Here we establish cardiac mosaicism as a causal mechanism for LQTS and present methods by which the general phenomenon, likely to be relevant for all genetic diseases, can be detected through single-cell analysis and next-generation sequencing.There is growing recognition that somatic mosaicism, i.e., genetic variation within an individual that arises from errors in DNA replication during early development, may play a role in a variety of human diseases other than cancer (1). However, the extent to which cellular heterogeneity contributes to disease is minimally understood. One report suggests that 6.5% of de novo mutations presumed to be germline in origin may instead have arisen from postzygotic mosaic mutation events (2), and recent genetic investigations directly interrogating diseased tissues in brain malformations, breast cancer, and atrial fibrillation have revealed postzygotic causal mutations absent from germline DNA (3–6). Pathogenic mosaic structural variation is also detectable in children with neurodevelopmental disorders (7). However, a consequential category of genetic variation has not been surveyed systematically in clinical or research studies of other human diseases.The pathophysiological basis of long-QT syndrome (LQTS) is prolongation of cardiac ventricular repolarization by acquired factors such as drug exposure or genetic variation in the proteins controlling transmembrane ion-concentration gradients (8, 9); parental gonadal mosaicism is an infrequently described phenomenon in LQTS (10–12). Knowledge of the molecular subtyping of disease in LQTS has provided a foundation for genotype-specific risk stratification and treatment strategies (8, 9, 13, 14). However, nearly 30% of probands remain undiagnosed using standard commercial gene-panel testing, suggesting there is unrecognized genetic variation (genic, regulatory, or otherwise) yet to be associated with disease (15).In its most severe form, LQTS may occur in the neonatal period with bradycardia and functional 2:1 AV block that occur secondary to a severely prolonged ventricular refractory period greater than the short R–R interval characteristic of a normal heart rate in infancy (16). Patients of all ages presenting with LQTS are predisposed to torsades de pointes (TdP), a life-threatening cardiac arrhythmia, with patients presenting in infancy showing particularly poor outcomes (13, 16–18). In this study, we applied rapid-turnaround whole-genome sequencing (WGS) on day of life (DOL) 3 in a premature infant with perinatal LQTS and life-threatening arrhythmia and investigated the contribution of a discovered mosaic variant to abnormal cardiac electrophysiology at the molecular and tissue levels. Additionally we surveyed 7,500 individuals already tested for genetic arrhythmias, allowing us to estimate the prevalence of mosaicism in an unbiased population sample.     

Complete genome sequence of Vibrio fischeri: a symbiotic bacterium with pathogenic congeners

Vibrio fischeri belongs to the Vibrionaceae, a large family of marine gamma-proteobacteria that includes several dozen species known to engage in a diversity of beneficial or pathogenic interactions with animal tissue. Among the small number of pathogenic Vibrio species that cause human diseases are Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus, the only members of the Vibrionaceae that have had their genome sequences reported. Nonpathogenic members of the genus Vibrio, including a number of beneficial symbionts, make up the majority of the Vibrionaceae, but none of these species has been similarly examined. Here we report the genome sequence of V. fischeri ES114, which enters into a mutualistic symbiosis in the light organ of the bobtail squid, Euprymna scolopes. Analysis of this sequence has revealed surprising parallels with V. cholerae and other pathogens.     

Pain control in orthodontics using a micropulse vibration device: A randomized clinical trial

Objective:To investigate the relationship between a micropulse vibration device and pain perception during orthodontic treatment.Materials and Methods:This study was a parallel group, randomized clinical trial. A total of 58 patients meeting eligibility criteria were assigned using block allocation to one of two groups: an experimental group using the vibration device or a control group (n  =  29 for each group). Patients used the device for 20 minutes daily. Patients rated pain intensity on a visual analog scale at appropriate intervals during the weeks after the separator or archwire appointment. Data were analyzed using repeated measures analysis of variance at α  =  .05.Results:During the 4-month test period, significant differences between the micropulse vibration device group and the control group for overall pain (P  =  .002) and biting pain (P  =  .003) were identified. The authors observed that perceived pain was highest at the beginning of the month, following archwire adjustment.Conclusion:The micropulse vibration device significantly lowered the pain scores for overall pain and biting pain during the 4-month study period.     

Eicosanoids and control of mesangial cell contraction

Contraction of glomerular mesangial cells is stimulated in vitro by the vasoconstrictor metabolite of arachidonic acid, thromboxane A2. To establish the role of mesangial prostaglandin (PG) synthesis in the modulation of contractile responses, we studied the effects of the stable thromboxane A2/endoperoxide analogue U-46619 on cultured rat mesangial cells preincubated with 1) four structurally unrelated, nonsteroidal anti-inflammatory drugs, indomethacin, acetylsalicylic acid, meclofenamate, and piroxicam, to inhibit the synthesis of PGE2, the major mesangial metabolite of arachidonic acid; 2) exogenous PGE2 and the stable analogue of PGI2, iloprost; and 3) indomethacin in the presence of exogenous PGE2. Computer-assisted image analysis microscopy demonstrated enhancement of spontaneous and agonist-induced contraction by nonsteroidal anti-inflammatory drugs in individual cells grown on a glass substrate, from 37.2 +/- 7.3% to a maximum of 75.5 +/- 6.4% of the cells with piroxicam, at 1 microM U-46619. PGE2 and iloprost dose-dependently inhibited U-46619-induced contraction, to 5.0 +/- 2.8% and 12.5 +/- 4.7% of the cells, respectively, at 1 microM U-46619. PGE2 also completely reversed the effects of indomethacin. Both PGE2 and iloprost dose-dependently stimulated intracellular cyclic AMP (cAMP) accumulation during 3-minute incubations, an effect that was blocked by the inhibitor of adenylate cyclase, 2',5'-dideoxyadenosine. The latter reversed the inhibitory action of PGE2, enhancing spontaneous and agonist-induced contractility, thus indicating a modulatory role of cAMP. We conclude that endogenous arachidonate metabolism regulates mesangial cell contraction through elevation of intracellular cAMP.