Acylcarnitines in plasma and blood spots of patients with long-chain 3-hydroxyacyl-coenzyme A dehydrogenase defiency.

The acylcarnitines in plasma and blood spots of 23 patients with proven deficiency of long-chain 3-hydroxyacylcoenzyme A dehydrogenase were reviewed. Long-chain 3-hydroxyacylcarnitines of C14:1, C14, C16 and C18:1 chain length, and long-chain acylcarnitines of C12, C14:1, C14, C16, C18:2 and C18:1 chain length were elevated. Acetylcarnitine was decreased. In plasma, elevation of hydroxy-C18:1 acylcarnitine over the 95th centile of controls, in combination with an elevation of two of the three acylcarnitines C14, C14:1 and hydroxy-C16, identified over 85% of patients with high specificity (less than 0.1% false positive rate). High endogenous levels of long-chain acylcarnitines in normal erythrocytes reduced the diagnostic specificity in blood spots compared with plasma samples. The results were also diagnostic in asymptomatic patients, and were not influenced by genotype. Treatment with diet low in fat and high in medium-chain triglyceride decreased all disease-specific acylcarnitines, often to normal, suggesting that this assay is useful in treatment monitoring.

Acylcarnitines in fibroblasts of patients with long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency and other fatty acid oxidation disorders.

Mitochondrial fatty acid oxidation disorders cause hypoglycaemia, hepatic dysfunction, myopathy, cardiomyopathy and encephalopathy. Despite their recognition for more than 15 years, diagnosis and treatment remain difficult. To help design rational diagnostic and therapeutic strategies, we studied the pathophysiology of accumulating metabolites in a whole-cell system. Acylcarnitines were quantified in cells and media of cultured fibroblasts after incubation with L-carnitine and fatty acids. Following incubation with palmitate, long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD)-deficient fibroblasts compared with controls showed elevation of hydroxypalmitoyl- and palmitoyl-carnitine and reduction of C10- and shorter acylcarnitines, and following incubation with linoleate an increase in C14:2-, C18:2- and hydroxy-C18:2- acylcarnitines and reduction in C10:1-acylcarnitines. Hydroxyacylcarnitines remained more intracellular compared to corresponding saturated acylcarnitines. Incubation with decanoate and octanoate showed absence of hydroxylated acylcarnitines and correction of secondary metabolic disturbances, suggesting that optimal treatment should include medium-chain triglycerides of these chain lengths. Fibroblasts of patients with other fatty acid oxidation disorders showed distinct elevations of disease-specific acylcarnitines. This acylcarnitine analysis allows the diagnosis of LCHAD deficiency and its differentiation from other fatty acid oxidation disorders, which can pose difficulties in vivo. The strategy has allowed in-depth analysis with different substrates, providing suggestions for the rational design of treatment trials.

Kinetic analysis of chloride conductance in frog skeletal muscle at pH 5.

At pH 5 the steady-state chloride chord conductance in frog skeletal muscle rises to an asymptotic maximum at very negative voltages and approaches an asymptotic minimum at positive voltages. When a two-pulse test paradigm is used, the conductance computed from steady-state currents during the first (conditioning) voltage step are not duplicated by the conductance at the onset of a second (test) step. If the test step is to a more negative voltage than the conditioning step the steady-state conductance is overestimated; if it is to a less negative voltage the conductance is underestimated. In some fibres the initial currents accompanying steps from the resting potential are inwardly rectified. From this it was inferred that chloride channel conductance is voltage dependent: in those fibres in which no such initial inward rectification was observed it was inferred that at rest the voltage-dependent chloride channels are all closed. Time-dependent ("gated") changes of conductance could be reasonably described by a first-order process, but the relaxations were not simple exponentials. Simulation of the experimental set-up predicted the type of deviation from exponentiality seen experimentally, although the observed deviations were often more pronounced than those predicted.

Effects of microgravity on tissue perfusion and the efficacy of astronaut denitrogenation for EVA.

The prevention of astronaut decompression sickness (DCS) during extravehicular activity (EVA) from the Shuttle Orbiter entails basic questions about how the efficacies of pre-EVA denitrogenations are affected by physiological responses and adaptations to microgravity. Many of these questions may be answered, without requiring inflight decompression experiments, when suitable inflight measurements of N2 elimination from spacecrew breathing 100% O2 can be analyzed using an N2 elimination/DCS risk correlation which has been calibrated in ground-based studies. In order to pursue this approach in our laboratory, a potentially flight-applicable, breath-by-breath method for measuring N2 elimination from human subjects breathing 100% O2 for 2-3-h periods has been developed. The present report describes this development with particular emphasis on required methodological accuracy and its achievement in view of certain properties of mass spectrometer performance. A method for the breath-by-breath analysis of errors in measured N2 elimination profiles is also described.

A likelihood analysis of experiments to test altitude decompression protocols for shuttle operations.

The principle of maximum likelihood and the method of linear regression both are used to fit mathematical models to experimental data, but likelihood can be applied to binary data such as the outcome of a decompression, whereas linear regression cannot. Maximum likelihood was applied to 548 individual altitude exposures from 30 experimental pressure profiles tested by NASA and the Air Force. Twelve decompression models were studied including modified Haldane models and models which assume that stationary bubbles cause Type I decompression sickness. The data was best represented by a model in which a bubble in tissue is surrounded by a diffusion barrier, but this representation was statistically indistinguishable from a single tissue Haldane model with a halftime of 508 min. By providing a quantitative measure of the agreement between theory and data, the principle of maximum likelihood offers an opportunity for improving the understanding of decompression mechanisms and for developing safer and faster decompression procedures.

Decompression induced nitrogen elimination.

A method for measuring nitrogen elimination after air diving has been developed in which a subject breathes air instead of oxygen or helium-oxygen. Accuracy is improved with this method because only nitrogen absorbed during the dive is eliminated. Nitrogen stored in the lungs and tissues at sea level is unaffected. Measurements were made with a closed-circuit breathing apparatus using a spirometer as a counterlung. The oxygen partial pressure in the apparatus was controlled at 0.209 +/- 0.003 atm. The spirometer volume was recorded periodically with the subject holding his breath at functional residual capacity. Increases in spirometer volume were used to define a nitrogen elimination curve. Elimination measurements were made after resting and exercising dives to 60, 100, and 130 fsw (2.8, 4.0, and 4.9 atm) at the U.S. Navy no-decompression exposure limits. Exercise during a dive increased the volume of nitrogen eliminated after the dive, but results for both resting and exercising divers were variable. Possible causes of this variability include bubble formation and changes in blood flow.