Optical spectroscopy of nicotinoprotein alcohol dehydrogenase from Amycolatopsis methanolica: a comparison with horse liver alcohol dehydrogenase and UDP-galactose epimerase

The NADH absorbance spectrum of nicotinoprotein (NADH-containing) alcohol dehydrogenase from Amycolatopsis methanolica has a maximum at 326 nm. Reduced enzyme-bound pyridine dinucleotide could be reversibly oxidized by acetaldehyde. The fluorescence excitation spectrum for NADH bound to the enzyme has a maximum at 325 nm. Upon excitation at 290 nm, energy transfer from tryptophan to enzyme-bound NADH was negligible. The fluorescence emission spectrum (excitation at 325 nm) for NADH bound to the enzyme has a maximum at 422 nm. The fluorescence intensity is enhanced by a factor of 3 upon binding of isobutyramide (Kd = 59 microM). Isobutyramide acts as competitive inhibitor (Ki = 46 microM) with respect to the electron acceptor NDMA (N,N-dimethyl-p-nitrosoaniline), which binds to the enzyme containing the reduced cofactor. The nonreactive substrate analogue trifluoroethanol acts as a competitive inhibitor with respect to the substrate ethanol (Ki = 1.6 microM), which binds to the enzyme containing the oxidized cofactor. Far-UV circular dichroism spectra of the enzyme containing NADH and the enzyme containing NAD+ were identical, indicating that no major conformational changes occur upon oxidation or reduction of the cofactor. Near-UV circular dichroism spectra of NADH bound to the enzyme have a minimum at 323 nm (Deltaepsilon = -8.6 M-1 cm-1). The fluorescence anisotropy decay of enzyme-bound NADH showed no rotational freedom of the NADH cofactor. This implies a rigid environment as well as lack of motion of the fluorophore. The average fluorescence lifetime of NADH bound to the enzyme is 0.29 ns at 20 degreesC and could be resolved into at least three components (in the range 0.13-0.96 ns). Upon binding of isobutyramide to the enzyme-containing NADH, the average excited-state lifetime increased to 1.02 ns and could be resolved into two components (0.37 and 1.11 ns). The optical spectra of NADH bound to nicotinoprotein alcohol dehydrogenase have blue-shifted maxima compared to other NADH-dehydrogenase complexes, but comparable to that observed for NADH bound to horse liver alcohol dehydrogenase. The fluorescence lifetime of NADH bound to the nicotinoprotein is very short compared to enzyme-bound NADH complexes, also compared to NADH bound to horse liver alcohol dehydrogenase. The cofactor-protein interaction in the nicotinoprotein alcohol dehydrogenase active site is more rigid and apolar than that in horse liver alcohol dehydrogenase. The optical properties of NADH bound to nicotinoprotein alcohol dehydrogenase differ considerably from NADH (tightly) bound to UDP-galactose epimerase from Escherichia coli. This indicates that although both enzymes have NAD(H) as nonexchangeable cofactor, the NADH binding sites are quite different.     

Making mistakes when predicting shifts in species range in response to global warming

Many attempts to predict the biotic responses to climate change rely on the 'climate envelope' approach, in which the current distribution of a species is mapped in climate-space and then, if the position of that climate-space changes, the distribution of the species is predicted to shift accordingly. The flaw in this approach is that distributions of species also reflect the influence of interactions with other species, so predictions based on climate envelopes may be very misleading if the interactions between species are altered by climate change. An additional problem is that current distributions may be the result of sources and sinks, in which species appear to thrive in places where they really persist only because individuals disperse into them from elsewhere. Here we use microcosm experiments on simple but realistic assemblages to show how misleading the climate envelope approach can be. We show that dispersal and interactions, which are important elements of population dynamics, must be included in predictions of biotic responses to climate change.     

Gallbladder diverticulum: a case report and review of the literature

BACKGROUND: The clinical significance of the 14 and 6 positive spikes (PS) electroencephalographic (EEG) phenomenon is not well established. This study was performed to provide further data regarding the clinical correlates of the PS, particularly attention-deficit/hyperactivity and somatic symptoms. METHODS: Diagnostic information gathered through structured interviews was compared among four groups of psychiatric inpatients aged 4-17 years who underwent an EEG examination over a 2-year period. Groups consisted of patients with: 1) 14 and 6 PS (n = 25); 2) epileptic discharges (n = 29); 3) slow-wave abnormalities (n = 23); and 4) a normal EEG group age and sex matched to the 14 and 6 PS group (n = 25). RESULTS: Attention-deficit hyperactivity disorder (ADHD) symptoms were significantly more frequent in the PS group (chi 2 = 2.96, p < .05) compared to the other three groups combined. Although somatic symptoms were not increased in the 14 and 6 PS group, anxiety symptoms tended to occur more in this group (chi 2 = 3.50, p < .06). CONCLUSIONS: The PS profile appears associated with ADHD symptoms. Possible treatment implications of this finding (e.g., use of anticonvulsants in ADHD patients with PS) need to be explored.     

"Death by proxy": ethics and classification in epidemiology

Epidemiology is reductionist in that it usually relies on creating categories of people or risk factors. Classification must be undertaken as part of any study, however by the act of choosing groups, individuals are (potentially) consigned to either a higher or lower risk group. We discuss this from an ethical perspective and consider: (a) whether the groupings commonly chosen genuinely represent the risk factor of interest, (b) the implications for individuals when consigned to groups and (c) the implications for epidemiology.     

Modulation of whole body protein metabolism, during and after exercise, by variation of dietary protein

The aim of this study was to investigate dietary protein-induced changes in whole body leucine turnover and oxidation and in skeletal muscle branched chain 2-oxo acid dehydrogenase (BCOADH) activity, at rest and during exercise. Postabsorptive subjects received a primed constant infusion of L-[1-13C,15N]leucine for 6 h, after previous consumption of a high- (HP; 1.8 g . kg-1 . day-1, n = 8) or a low-protein diet (LP; 0.7 g . kg-1 . day-1, n = 8) for 7 days. The subjects were studied at rest for 2 h, during 2-h exercise at 60% maximum oxygen consumption, then again for 2 h at rest. Exercise induced a doubling of both leucine oxidation from 20 micromol . kg-1 . h-1 and BCOADH percent activation from 7% in all subjects. Leucine oxidation was greater before (+46%) and during (+40%, P < 0.05) the first hour of exercise in subjects consuming the HP rather than the LP diet, but there was no additional change in muscle BCOADH activity. The results suggest that leucine oxidation was increased by previous ingestion of an HP diet, attributable to an increase in leucine availability rather than to a stimulation of the skeletal muscle BCOADH activity.     

Value of the pretransplant evaluation in predicting toxic day-100 mortality among blood stem-cell and bone marrow transplant recipients

PURPOSE: To determine the value of pretransplant studies in predicting day 100 nonrelapse toxic mortality following high-dose therapy. PATIENTS AND METHODS: A retrospective review of 383 consecutive hematopoietic stem-cell transplants was performed with attention to toxic mortality and pretransplant factors. Univariate log-rank analysis was used to yield the most significant cut-off values for individual factors. Multivariate analysis using Cox proportional hazards regression determined factors independently predictive of early toxic death. RESULTS: Nonrelapse toxic mortality before day 100 occurred in 23 of 383 (6.0%) transplant recipients. Factors associated with an increased risk of toxic death by univariate analysis included forced expiratory volume in 1 second (FEV1) less than 78% of predicted (P = .0002), allogeneic versus autologous transplant (P = .0003), diffusion capacity of carbon monoxide less than 52% of predicted (P = .002), serum creatinine concentration greater than 1.1 mg/dL (P = .003), Eastern Cooperative Oncology Group performance status greater than 0 (P = .006), preparative regimen containing total-body irradiation versus chemotherapy alone (P = .006), marrow versus blood stem cell (P = .01), serum ALT greater than 50 IU/L (P = .02), diagnosis of hematologic disorder versus solid tumor (P = .06), serum bilirubin level greater than 1.1 mg/dL (P = .08), left ventricular ejection fraction (P = .09), and growth factor use (P = .09). In the multivariate model, transplant type (relative risk, 4.2), FEV1 (relative risk, 4.5), performance status (relative risk, 3.7), serum creatinine (relative risk, 3.8), and serum bilirubin (relative risk, 3.7) were found to be independent predictors of early toxic mortality. CONCLUSION: The pretransplant evaluation is a useful tool to identify patients at risk for early toxic mortality following high-dose therapy.     

Rituximab chimeric anti-CD20 monoclonal antibody therapy for relapsed indolent lymphoma: half of patients respond to a four-dose treatment program

PURPOSE: The CD20 antigen is expressed on more than 90% of B-cell lymphomas. It is appealing for targeted therapy, because it does not shed or modulate. A chimeric monoclonal antibody more effectively mediates host effector functions and is itself less immunogenic than are murine antibodies. PATIENTS AND METHODS: This was a multiinstitutional trial of the chimeric anti-CD20 antibody, IDEC-C2B8. Patients with relapsed low grade or follicular lymphoma received an outpatient treatment course of IDEC-C2B8 375 mg/m2 intravenously weekly for four doses. RESULTS: From 31 centers, 166 patients were entered. Of this intent-to-treat group, 48% responded. With a median follow-up duration of 11.8 months, the projected median time to progression for responders is 13.0 months. Serum antibody levels were sustained longer after the fourth infusion than after the first, and were higher in responders and in patients with lower tumor burden. The majority of adverse events occurred during the first infusion and were grade 1 or 2; fever and chills were the most common events. Only 12% of patients had grade 3 and 3% grade 4 toxicities. A human antichimeric antibody was detected in only one patient. CONCLUSION: The response rate of 48% with IDEC-C2B8 is comparable to results with single-agent cytotoxic chemotherapy. Toxicity was mild. Attention needs to be paid to the rate of antibody infusion, with titration according to toxicity. Further investigation of this agent is warranted, including its use in conjunction with standard chemotherapy.     

Technical report: Initial experiences with an experimental solid-state universal digital X-ray image detector

This paper presents a brief technical evaluation and first review of clinical experiences with an experimental direct digital X-ray image detector designed to support both dynamic and snap-shot imaging. Derivatives of this type of image detector can potentially fulfil the majority of the fluoroscopic and radiographic imaging requirements of clinical radiology departments, and initial results suggest that imaging systems using the new technology will provide a high quality dose-efficient solution to the search for a universal digital X-ray image detector.     

Exogenous norepinephrine induces an enhanced microproteinuric response in microalbuminuric insulin-dependent diabetes mellitus

Exogenous norepinephrine (NE) increases intraglomerular pressure in animal experiments, but it is unknown whether NE induces a microproteinuric response in humans. Moreover, it has not been studied whether possible microproteinuric and renal hemodynamic changes induced by NE are altered in insulin-dependent diabetes mellitus (IDDM) complicated by microalbuminuria. Therefore, the microproteinuric and renal hemodynamic responses to exogenous NE infusions were measured in eight matched normoalbuminuric IDDM patients (group D1), microalbuminuric IDDM patients (group D2), and control subjects (group C). As anticipated, mean arterial pressure (MAP)-NE dose-response curves were significantly shifted leftward in groups D1 and D2 compared with group C (P < 0.05), indicating a higher systemic NE responsiveness in IDDM. On separate days, NE or placebo was infused at individually determined NE threshold doses (T; delta MAP = 0 mmHg), 20% pressor doses (20% P; delta MAP = 4 mmHg), and pressor doses (P; delta MAP = 20 mmHg), with measurement of urinary albumin (UalbV), IgG excretion (UIgGV), GFR (by 125I-iothalamate), and effective renal plasma flow (by 131I-hippurate). At NE pressor dose, UalbV and UIgGV rose in all groups (P < 0.05 to 0.01), whereas urinary beta 2-microglobulin was unchanged. The increases in UalbV and UIgGV were more pronounced in the microalbuminuric group than in the other groups (P < 0.05). An NE dose-dependent fall in effective renal plasma flow and rise in filtration fraction were found in all groups (P < 0.05 to 0.001 for all), whereas GFR did not change significantly. The renal hemodynamic dose-response relationship was similar in the groups. In conclusion, exogenous NE acutely promotes glomerular protein leakage, and it is plausible that intraglomerular NE effects contribute to this phenomenon. The microproteinuric response is enhanced in microalbuminuric IDDM despite unaltered renal hemodynamic responsiveness, which may reflect a specific NE response or a general effect of vasopressor stimuli to promote glomerular protein leakage in patients with a preexistent defect in glomerular permselectivity.