Evaluation of the aortic root by MRI: insights from patients with homozygous familial hypercholesterolemia

Hepatitis C chronically infects approximately 1.5% of Americans and is the most common clinical problem facing hepatologists. Since the virus was initially described in 1989, development of an effective therapy has been challenging. Although several different therapeutic agents have been used, no therapy has been shown to reliably eradicate the virus. Interferon-alpha, a cytokine with immunostimulatory and anti-viral properties, has become the therapy of choice for patients with chronic hepatitis C infection. Trials assessing the efficacy of interferon-alpha have characterized host and viral factors predictive of responses to treatment. A thorough understanding of these predictive factors is requisite to providing cost-effective therapeutic decisions for the patient with chronic hepatitis C infection.     

Global folds of highly deuterated, methyl-protonated proteins by multidimensional NMR

The development of 15N, 13C, 2H multidimensional NMR spectroscopy has facilitated the assignment of backbone and side chain resonances of proteins and protein complexes with molecular masses of over 30 kDa. The success of these methods has been achieved through the production of highly deuterated proteins; replacing carbon-bound protons with deuterons significantly improves the sensitivity of many of the experiments used in chemical shift assignment. Unfortunately, uniform deuteration also radically depletes the number of interproton distance restraints available for structure determination, degrading the quality of the resulting structures. Here we describe an approach for improving the precision and accuracy of global folds determined from highly deuterated proteins through the use of deuterated, selectively methyl-protonated samples. This labeling profile maintains the efficiency of triple-resonance NMR experiments while retaining a sufficient number of protons at locations where they can be used to establish NOE-based contacts between different elements of secondary structure. We evaluate how this deuteration scheme affects the sensitivity and resolution of experiments used to assign 15N, 13C, and 1H chemical shifts and interproton NOEs. This approach is tested experimentally on a 14 kDa SH2/phosphopeptide complex, and a global protein fold is obtained from a set of methyl-methyl, methyl-NH, and NH-NH distance restraints. We demonstrate that the inclusion of methyl-NH and methyl-methyl distance restraints greatly improves the precision and accuracy of structures relative to those generated with only NH-NH distance restraints. Finally, we examine the general applicability of this approach by determining the structures of several proteins with molecular masses of up to 40 kDa from simulated distance and dihedral angle restraint tables.     

Direct administration of insulin-like growth factor to fetal rhesus monkeys (Macaca mulatta)

A potential treatment for the amelioration of fetal growth failure is insulin-like growth factor-I (IGF-I). To address concerns of safety and efficacy, IGF-I (80 microg/kg; GroPep Pty.) was administered i.p. to healthy rhesus monkey fetuses via ultrasound guidance every other day between gestational days (GD) 110-120 and 130-140 (third trimester; term = approximately GD 165 +/- 10; n = 6). Pregnancies were monitored sonographically, and fetal/maternal blood samples were collected for complete blood counts, immunophenotyping, and biochemical analyses. Blood samples, external measures of the fetus and newborn, and tissue and organ weights were collected at fetal necropsy (GD 150; n = 2) or at term delivery of neonates (GD 160; n = 4). The results of these investigations have shown no evidence of hypoglycemia in the fetus or dam during the course of treatment. Circulating concentrations of fetal, but not maternal, IGF-I increased with treatment (approximately 80 to approximately 1015 ng/ml), and there was no evidence of a change in serum IGF-II or an increase in IGF binding protein-3 compared with historical control values. Fetal lymphocytes and select red cell parameters increased, and a significant elevation in circulating B cells and CD4/CD8 ratios in fetal lymph nodes was shown. Although no changes were detected in body weights, increases in thymic, splenic, and kidney weights and small intestine lengths occurred. Thus, administration of IGF-I to the fetal monkey is safe and results in 1) transient increases in circulating IGF-I, 2) a significant effect on fetal hematopoietic and lymphoid tissues, and 3) an increase in select fetal organ weights and measures. These data suggest that IGF-I may represent a potential candidate for therapeutic treatment of growth-compromised human fetuses in utero.     

Importance of the Arg-Gly-Asp triplet in human thrombin for maintenance of structure and function

Site-directed mutagenesis was employed to assess the importance of the Arg-Gly-Asp triplet that comprises residues 197 to 199 in the B-chain of thrombin. Properties of the R197E and the D199E variants were compared with those of zeta-thrombin and the inactive S205A variant wherein the active site Ser is replaced by Ala. Relative to zeta-thrombin, the R197E thrombin variant under the assay conditions used exhibits 26% activity toward a small chromogenic substrate, 13% activity in the activation of protein C in the presence of thrombomodulin, < 3% activity in processing fibrinogen, and 1% activity in inducing platelet activation. Thus, the substrate specificity of thrombin was altered by the R197-->E replacement. The D199E variant was essentially inactive. It exhibited only 0.02% of the activity of thrombin toward the chromogenic substrate and its reactivity toward the active site-directed alkylating agent D-Phe-Pro-Arg-CH2Cl was 10,000-fold lower than that of thrombin. Like the inactive S205A thrombin variant, the D199E variant antagonized the interactions of thrombin with hirudin and thrombomodulin, but was a less effective antagonist. The dependence of the antagonism of the thrombin-thrombomodulin interaction on the concentration of D199E thrombin variant provided evidence suggesting the presence of two or more domains in thrombin that independently interact with their counterparts in thrombomodulin. Although the S205A thrombin variant antagonized the action of thrombin on platelets no such activity could be demonstrated for the D199E variant in the concentration range studied (< 800 nm). Comparison of the circular dichroism spectra of zeta-thrombin, the D199E, R197E, and S205A variants indicated that subtle differences in conformation exist between the D199E variant and the other thrombins. These differences in conformation might well account for the altered behavior of the D199E variant with respect to its interactions toward thrombomodulin, hirudin, and platelets.     

Fluorescence in situ hybridization analysis of chromosome 1p36 deletions in human MYCN amplified neuroblastoma

Artificial illumination is an important factor in the management of layers. In this study, a new monochromatic light system was developed for egg layers. Prelaying pullets (Lohmann) were marked and housed in nine light and temperature control rooms (15 battery cages, 3 hens per cage; n = 45), divided into three light treatments: 0.1 and 0.01 W/m2 light intensity using light emitting diode (LED) lamps and 0.1 W/m2 using mini-fluorescent bulbs (PL) (control). In each of the LED rooms, three wavelengths were tested: 560 (n = 9), 660 (n = 9), 880 (n = 6), and 660 intermitted lighting (15 min light 45 min dark, 660IN) (n = 9). Birds were exposed to 12 h light and 12 h of darkness using PL lamps. At 21 wk of age, the light period was increased to 12.75 h by using 5.5 h of LED lamps and 7.25 of PL light source for Groups 1 and 2, the third group received 12.75 h of PL light. Until 28 wk of age, light hours increased by 0.5 h/w using LED light for Groups 1 and 2 and PL source for the third group, reaching 16 h of light at 28 wk of age. Egg production and feed consumption were recorded daily; egg components were recorded weekly for 10 mo. A significant reduction in egg production was observed in all 880nm groups; no differences in egg production and quality were found in the other groups. Feed consumption was significantly lower by 7% in all 0.01 W/m2 groups. We suggest that an important reduction in rearing costs of laying hens may be obtained by using this system.     

Necrotizing enterocolitis

Advances in neonatal care have resulted in an enlarging population of vulnerable premature newborns at risk for necrotizing enterocolitis (NEC). This article presents data supporting a unifying hypothesis for the initiation of NEC based on bacteria as the inciting agent(s), and the preterm baby as the vulnerable host. Facts and controversies concerning the pathology, microbiology, clinical presentation, management and outcome of infants afflicted with NEC are presented.     

Further studies of the mixed acetals of nucleosides

We reported in 1988 on a new nucleoside modification reaction: the exocyclic amino groups of (d)adenosine and (d)cytidine react rapidly at ambient temperature with acetaldehyde and alcohols to give stable mixed acetals (N-ethylethoxy-acetal). NH2 + O = CH(CH3) + ROH-->NH-CH(CH3)-O-R + H2O. Here we report in detail on the occurrence of this reaction in very dilute aqueous solution (ie under biological conditions), on its mechanism and kinetics, on the mixed acetal formation with other aldehydes and other nucleic acid components, and on the question of whether these adducts are mutagenic.     

Superior vena cava syndrome after heart transplantation: percutaneous treatment of a complication of bicaval anastomoses

The aims of this study were to establish a working rabbit heart model of regional myocardial ischaemia in which electrophysiologic parameters and arrhythmogenesis could be correlated and to explore the mechanisms underlying the antiarrhythmic activity of lignocaine. Monophasic action-potential duration (MAPD90), effective refractory period (ERP), and conduction delay were measured at three ventricular sites in isolated hearts paced at 3.3 Hz. The hearts were treated before and throughout 30 min of ischaemia and 15 min of reperfusion with a vehicle or 20 microM lignocaine. In both groups, ischaemia produced a similar shortening in MAPD90. Lignocaine decreased ERP shortening during ischaemia from -56+/-4 to -32+/-6 ms. An ischaemia-induced increase in conduction delay was greater in the lignocaine than the control group (49+/-7 vs. 11+/-2 ms). Ischaemia-induced dispersion of repolarisation was reduced by lignocaine from 66+/-4 to 32+/-7 ms, and dispersion of refractoriness was decreased from 57+/-6 to 16+/-3 ms. Lignocaine decreased inducibility of ventricular fibrillation (VF) during ischaemia from 86 to 25%. We conclude that, in this model, the antiarrhythmic activity of lignocaine during regional ischaemia is associated with an increase in ischaemia-induced conduction delay and reduced dispersion of repolarisation and refractoriness.