Loss-of-function mutations in the EGF-CFC gene CFC1 are associated with human left-right laterality defects.

All vertebrates display a characteristic asymmetry of internal organs with the cardiac apex, stomach and spleen towards the left, and the liver and gall bladder on the right. Left-right (L-R) axis abnormalities or laterality defects are common in humans (1 in 8,500 live births). Several genes (such as Nodal, Ebaf and Pitx2) have been implicated in L-R organ positioning in model organisms. In humans, relatively few genes have been associated with a small percentage of human situs defects. These include ZIC3 (ref. 5), LEFTB (formerly LEFTY2; ref. 6) and ACVR2B (encoding activin receptor IIB; ref. 7). The EGF-CFC genes, mouse Cfc1 (encoding the Cryptic protein; ref. 9) and zebrafish one-eyed pinhead (oep; refs 10, 11) are essential for the establishment of the L-R axis. EGF-CFC proteins act as co-factors for Nodal-related signals, which have also been implicated in L-R axis development. Here we identify loss-of-function mutations in human CFC1 (encoding the CRYPTIC protein) in patients with heterotaxic phenotypes (randomized organ positioning). The mutant proteins have aberrant cellular localization in transfected cells and are functionally defective in a zebrafish oep-mutant rescue assay. Our findings indicate that the essential role of EGF-CFC genes and Nodal signalling in left-right axis formation is conserved from fish to humans. Moreover, our results support a role for environmental and/or genetic modifiers in determining the ultimate phenotype in humans.

Erythropoiesis-stimulating agents in oncology: a study-level meta-analysis of survival and other safety outcomes.

BACKGROUND: Cancer patients often develop the potentially debilitating condition of anaemia. Numerous controlled studies indicate that erythropoiesis-stimulating agents (ESAs) can raise haemoglobin levels and reduce transfusion requirements in anaemic cancer patients receiving chemotherapy. To evaluate recent safety concerns regarding ESAs, we carried out a meta-analysis of controlled ESA oncology trials to examine whether ESA use affects survival, disease progression and risk of venous-thromboembolic events. METHODS: This meta-analysis included studies from the 2006 Cochrane meta-analysis, studies published/updated since the 2006 Cochrane report, and unpublished trial data from Amgen and Centocor Ortho Biotech. The 60 studies analysed (15 323 patients) were conducted in the settings of chemotherapy/radiochemotherapy, radiotherapy only treatment or anaemia of cancer. Data were summarised using odds ratios (ORs) with 95% confidence intervals (CIs). RESULTS: Results indicated that ESA use did not significantly affect mortality (60 studies: OR=1.06; 95% CI: 0.97-1.15) or disease progression (26 studies: OR=1.01; 95% CI: 0.90-1.14), but increased the risk for venous-thromoboembolic events (44 studies: OR=1.48; 95% CI: 1.28-1.72). CONCLUSION: Though this meta-analysis showed no significant effect of ESAs on survival or disease progression, prospectively designed, future randomised clinical trials will further examine the safety and efficacy of ESAs when used according to the revised labelling information.

The role of acyl-CoA: cholesterol acyltransferase in the metabolism of free cholesterol to cholesteryl esters or bile acids in primary cultures of rat hepatocytes.

Sandoz compound 58-035 has been shown to inhibit acyl-CoA: cholesterol acyltransferase activity in a variety of cell types. We have shown that it does not inhibit rat liver microsomal cholesterol 7 alpha-hydroxylase, the rate-limiting enzyme of bile-acid synthesis, but it does inhibit acyl-CoA: cholesterol acyltransferase in both the microsomal fraction and in rat hepatocyte monolayers. To test the role of acyl-CoA: cholesterol acyltransferase in these cells, monolayers were incubated over 5 h in the presence and absence of 58-035 and in the presence of increasing amounts of mevalonic acid to provide a source of cholesterol. The addition of mevalonic acid increased the secretion of bile acids by the cells, and this was further increased by the addition of 58-035. The secretion of cholesteryl esters was conversely inhibited by the addition of 58-035. The results help define the role of acyl-CoA: cholesterol acyltransferase in determining the fate of intracellular cholesterol.

The effects of 6-azacholest-4-en-3 beta-ol-7-one, an inhibitor of cholesterol 7 alpha-hydroxylase, on cholesterol metabolism and bile acid synthesis in primary cultures of rat hepatocytes.

6-Azacholest-4-en-3 beta-ol-7-one (azacholesterol) was shown to be a specific inhibitor of cholesterol 7 alpha-hydroxylase. It inhibited cholesterol hydroxylation by a rat liver microsomal preparation with non-competitive kinetics and a Ki of 4 microM. No evidence was found for a time-dependent inhibition of activity. Azacholesterol did not inhibit acyl-CoA: cholesterol acyltransferase or 3-hydroxy-3-methylglutaryl coenzyme A reductase in rat liver microsomal preparations, or cholesterol esterification and synthesis in primary cultures of rat hepatocytes. The synthesis of bile acids was inhibited by azacholesterol in these cells in a dose-dependent way. When bile acid synthesis was inhibited by azacholesterol, newly-synthesized cholesterol from exogenous mevalonate was secreted by the hepatocyte cultures into the cell culture medium in several-fold excess over control incubations. No changes in the secretion of cholesteryl ester occurred in the presence of azacholesterol. This observation suggests that newly synthesised cholesterol that has entered the substrate pool for hydroxylation is no longer accessible to the substrate pool for esterification. This is further evidence for the compartmentation of cholesterol metabolism in the hepatocyte.

Vitamin D receptor polymorphisms are associated with altered prognosis in patients with malignant melanoma.

Calcitriol [1,25(OH)2D3], the hormonal derivative of vitamin D3, is an antiproliferative and prodifferentiation factor for several cell types, including cultured melanocytes and malignant melanoma (MM) cells. Several polymorphisms of the vitamin D receptor (VDR) gene have been described including a FokI RFLP in exon 2, BsmI, and ApaI polymorphisms in intron 8 and an adjacent TaqI RFLP in exon 9. Alterations in vitamin D/1,25(OH)2D3 levels and polymorphisms of the VDR have been shown to be associated with several systemic malignancies. We hypothesize that polymorphism in this gene may be associated with altered susceptibility and outcome in patients with MM. A hospital-based case-control study, using 316 MM cases and 108 controls, was used to assess associations with MM susceptibility. Breslow thickness, the most important single prognostic factor in MM, was used as the outcome measure. Polymorphisms at the FokI and TaqI restriction sites were determined using PCR-based methods. Polymorphism at the FokI, but not TaqI, RFLP was associated with an altered risk of MM (P = 0.014). More importantly, variant alleles were associated with increased Breslow thickness. Thus, homozygosity for variant alleles at both RFLP (ttff genotype combination) was significantly associated with thicker tumors. (> or = 3.5 mm; P = 0.001; odds ratio = 31.5). Thus, polymorphisms of the VDR gene, which would be expected to result in impaired function, are associated with susceptibility and prognosis in MM. These data suggest that 1,25(OH)2D3, the ligand of the VDR, may have a protective influence in MM, as has been proposed for other malignancies.

Nitroprusside selectively reduces ventricular preload in the stage 21 chick embryo.

OBJECTIVE: Embryonic cardiovascular function is dynamically regulated at the tissue level. Nitric oxide (NO) regulates vascular tone and influences cardiovascular function in neonatal and mature circulations. However, the role of NO in regulating embryonic cardiovascular function is undefined. We hypothesized that NO released from nitroprusside alters embryonic vascular tone with secondary effects on ventricular function. METHODS: We measured ventricular pressure and calculated ventricular volume from area using ellipsoid geometry for 180 s after suffusion of 0.0, 0.1, 1.0, or 2.5 micrograms of nitroprusside in 10 microliters of KHB buffer, or after acute venous hemorrhage in stage 21 chick embryos (n > or = 8 per group). We plotted pressure-volume loops and analyzed standard parameters of cardiovascular function by ANOVA, regression analysis, and ANCOVA. RESULTS: Following nitroprusside, heart rate was unchanged, end-diastolic volume (EDV), stroke volume (SV), and end-systolic pressure decreased (P < 0.05 at 180 s). For 1.0 microgram of nitroprusside, EDV decreased by 27 +/- 6%, SV decreased by 36 +/- 6%, and end-systolic pressure decreased by 28 +/- 3%. The EDV vs. SV relationship was unchanged with the exception of the 2.5 micrograms nitroprusside dose. Arterial elastance was unchanged with the exception of the 2.5 micrograms nitroprusside dose. The EDV vs. SV relationship and arterial elastance during preload reduction suggest that ventricular afterload did not decrease following NO. CONCLUSIONS: In contrast to the mature circulation, NO reduced preload without decreasing ventricular afterload. Thus, vasoactive agents may have unique roles in the regulation of cardiovascular structure and function during embryogenesis.

Evidence that adenosine 3',5'-monophosphate mediates hormonal stimulation of prostaglandin production in cultured mouse parietal bones.

The present investigation was undertaken to examine the possible role of cAMP in PTH-stimulated prostaglandin (PG) production in organ cultures of neonatal mouse parietal bones. Cultures were treated with PTH, forskolin, isobutylmethylxanthine (IBMX), and 8-bromo-cAMP (8BrcAMP). We found that similar concentrations of PTH stimulate cAMP formation and increase PG production in this culture system. Forskolin, a direct activator of adenylate cyclase, was also a potent stimulator of cAMP and PG production. The effect was dose dependent, with a maximum at 10(-5) M. The time courses for PTH- and forskolin-stimulated PG production were similar, and there was a close and similar correlation between cAMP production at 15 min and PGE2 production at 6 h for both agents. An increase in PG production was also observed when IBMX, which elevates cAMP levels in cells by inhibiting cAMP phosphodiesterase, or the cAMP analog 8BrcAMP was added to the cultures. In addition, IBMX enhanced the PGE2 responses to PTH, forskolin, and 8BrcAMP. These findings indicate that stimulation of PG production by PTH may be mediated by cAMP.

Left-right axis malformations associated with mutations in ACVR2B, the gene for human activin receptor type IIB.

Targeted disruption of the mouse activin receptor type IIB gene (Acvr2b) results in abnormal left-right (LR) axis development among Acvr2b-/- homozygotes [Oh and Li, 1997: Genes Dev 11:1812-1826]. The resulting malformations include atrial and ventricular septal defects, right-sided morphology of the left atrium and left lung, and spleen hypoplasia. Based on these results, we hypothesized that mutations in the type IIB activin receptor gene are associated with some cases of LR axis malformations in humans. We report here characterization of the ACVR2B genomic structure, analysis of ACVR2B splice variants, and screening for ACVR2B mutations among 112 sporadic and 14 familial cases of LR axis malformations. Two missense substitutions have been identified, one of which appears in two unrelated individuals. Neither of these nucleotide changes has been found in 200 control chromosomes. We conclude that ACVR2B mutations are present only rarely among human LR axis malformation cases.

Mesoridazine -- a pharmacodynamic and pharmacokinetic profile.

Available data on the clinical effectiveness and side effects of mesoridazine are reviewed, and an attempt is made to relate clinical efficacy to phenothiazine pharmacokinetics. Both controlled and open-label clinical studies have attested to the efficacy of mesoridazine in schizophrenia, as well as certain other psychiatric disorders. Clinical observations of the effectiveness of mesoridazine in patients refractory to treatment with thioridazine and other neuroleptics may be related to its slow rate of inactivation and to the relatively large proportion of free mesoridazine that is available for penetration to the target sites in the brain.

Analysis of skull anthropometric measurements in patients with neurofibromatosis type-1.

RATIONALE AND OBJECTIVES: The authors studied selected anthropometric measurements of plain postero-anterior and lateral skull roentgenograms to ascertain whether these were useful in distinguishing patients with clinically probable neurofibromatosis type-1 from controls. METHODS: A retrospective review of medical records of patients for whom skull roentgenograms were available was conducted. Patients were assigned to one of three groups: definite neurofibromatosis type-1 (DNF), probable neurofibromatosis type-1 (PNF), and controls. A blinded analysis of 29 measurements, 9 qualitative assessments, and 3 area/volume calculations was performed. RESULTS: There were 58 patients (29 controls, 14 DNF, and 15 PNF). The majority (75%) of all predetermined landmarks could be ascertained in 43 of these subjects. After age and gender were held constant, analysis of covariance showed that both DNF and PNF subjects could be distinguished from controls, but not from each other when comparing the mean: sella turcica height (P < .001), sella turcica depth (P < .005), skull width (P < .001), skull length (P < .002), skull height (P < .003), and skull volume (P < .0001). CONCLUSIONS: Anthropometric analysis of skull roentgenograms coupled with results of clinical examination improves the ability to distinguish between patients with DNF and PNF from controls.

The genetics of left-right development and heterotaxia.

Looping of the primitive heart tube is one of the earliest and most crucial steps in cardiac morphogenesis. Cardiac looping is dependent on normal left-right development, and defects in left-right development result in both heterotaxia and complex congenital heart disease. Single gene defects result in the wide spectrum of heterotaxy phenotypes, and conversely, different gene defects result in similar heterotaxy phenotypes. Elucidation of the molecular-genetic mechanisms of left-right development will greatly increase our understanding of the etiology of this complex group of congenital heart defects.

Acute ventricular hemorrhage in adults with hydrocephalus managed by corpus callosotomy and fenestration of the septum pellucidum. Report of three cases.

Three patients with hypertension-induced basal ganglia or thalamic hemorrhage and ventricular rupture underwent corpus callosotomy and fenestration of the septum pellucidum. A patient with a left thalamic hemorrhage underwent surgery on an emergency basis and made a complete physical recovery, although she retained mild psychomotor deficits. Another patient with a large right basal ganglia hemorrhage who also underwent surgery on an emergency basis retained a spastic left hemiparesis without evident psychomotor deficits. The third patient with a left thalamic and basal ganglia hemorrhage, who was initially awake and then lapsed into stupor days later, underwent surgery, but did not recover consciousness. Hydrocephalus was reversed and effectively controlled in all three patients without having to perform a shunt placement procedure.

Dogger Bank Itch and cyclosporin.

Two cases of Dogger Bank Itch responding to oral cyclosporin (Neoral) are reported. The first, a 42-year-old shell-fisherman, presented with eczema on his face, forearms and hands. Oral prednisolone was ineffective and in subsequent years systemic steroids and topical clobetasol propionate allowed him to continue his work. In the shellfishing season of 1996, cyclosporin (Neoral) was introduced and the eczema was subsequently satisfactorily controlled. The second fisherman, aged 46, presented with an irritant eczema of the hands and forearms. Initial treatment with emollients and potent topical steroids had little effect. A subsequent course of cyclosporin provided good control of the eczema.

EEG "maturational lag" profiles: follow-up analyses.

Nineteen children who had EEG diffuse slow frequency profiles suggesting a maturational lag were retested 1 to 2 years later receiving both EEG and psychometric assessments. They were compared to 18 children who also had been tested previously but did not have the slow frequency profile. Children with the maturational lag profile displayed varying degrees of EEG developmental change upon retest; i.e., some showed the expected developmental change while others remained the same. Within this group, there were significant relationships between the degree of EEG change and psychometric scores. Those that demonstrated the most EEG change had higher psychometric indices, and these relationships were not obtained in the comparison group. These findings suggest developmental heterogeneity with respect to brain maturation as measured by the EEG and also suggest that test-retest EEG data can clarify a maturational lag diagnosis.

Naming-speed processes, timing, and reading: a conceptual review.

This article integrates the findings in the special issue with a comprehensive review of the evidence for seven central questions about the role of naming-speed deficits in developmental reading disabilities. Cross-sectional, longitudinal, and cross-linguistic research on naming-speed processes, timing processes, and reading is presented. An evolving model of visual naming illustrates areas of difference and areas of overlap between naming speed and phonology in their underlying requirements. Work in the cognitive neurosciences is used to explore two nonexclusive hypotheses about the putative links between naming speed and reading processes and about the sources of disruption that may cause subtypes of reading disabilities predicted by the double-deficit hypothesis. Finally, the implications of the work in this special issue for diagnosis and intervention are elaborated.

Diagnosis and antibiotic treatment of acute otitis media: report from International Primary Care Network.

STUDY OBJECTIVE: The relation between a history of disorders suggestive of acute otitis media, symptoms, and findings of an examination of the tympanic membrane and doctors' certainty of diagnosis. Also, to examine differences in prescribing habits for acute otitis media among doctors from different countries. DESIGN: Questionnaires were completed by participating doctors for a maximum of 15 consecutive patients presenting with presumed acute otitis media. SETTING: General practices in Australia, Belgium, Great Britain, Israel, The Netherlands, New Zealand, Canada, Switzerland, and the United States. PATIENTS: 3660 Children divided into the three age groups 0-12 months, 13-30 months, and greater than or equal to 31 months. MAIN OUTCOME MEASURES: General practitioners' responses to questions on their diagnostic certainty and resolution of patients' symptoms after two months. RESULTS: The diagnostic certainty in patients aged 0-12 months was 58.0%. This increased to 66.0% in those aged 13-30 months and 73.3% in those aged greater than or equal to 31 months. In all age groups diagnostic certainty was positively associated with the finding of a tympanic membrane that was discharging pus or bulging. Redness of the membrane and pain were also associated with certainty in patients aged 13-30 months, and a history of decreased hearing or recent upper respiratory infection was positively associated in patients aged greater than or equal to 31 months. The proportion of patients prescribed antibiotics varied greatly among the countries, from 31.2% in The Netherlands to 98.2% in both Australia and New Zealand, as did the duration of treatment. Patients who did not take antibiotics had a higher rate of recovery than those who did; the rate of recovery did not differ between different types of antibiotic. CONCLUSIONS: Doctors' certainty of diagnosis of acute otitis media was linked to patient's age. Improved criteria or techniques for diagnosing acute otitis media, especially in very young children, need to be developed. Antibiotic treatment did not improve the rate of recovery of patients in this study.