Paracorporeal artificial heart in postoperative heart failure.

A pneumatically driven artificial heart with a tubular silicone rubber membrane and disc valves was used for functional heart replacement in the paracorporeal mode. A fluidic drive system allows adjustment of the heart rate, positive and negative pressures and systole/diastole ratio. Since August, 1977, the artificial heart has been used in four patients with refractory postoperative heart failure not responding to volume loading, pH and electrolyte correction, catecholamines and intra-aortic balloon pumping. Large cannulae were placed in the atria and great vessels. The ventricles were fixed on the chest paracorporeally. The assist system was used as a left heart bypass in one patient and as a biventricular bypass in three other patients. After 48-72 hours, the ventricular function recovered in three patients, permitting removal of the artificial heart. One patient died of cerebral complications six weeks later; the other two recovered completely and were released in good condition. Profound postoperative heart failure can be completely reversed by the use of the paracorporeal artificial heart; the advantage of the system lies in the simplicity of its implantation and removal.


Round window niche melanocytes and webby tissue. Possible collateral route of vascular and perilymph circulation.

Two hundred thirteen human temporal bones were studied for characteristics of the round window niche. In 163 specimens melanocytes were found within the attachments of the round window membrane and in the niche, most prominently at the level of the cochlear aqueduct opening. Sixty-two specimens demonstrated discrete webby foci within the lining of the niche, 16 of which completely or partially covered the tympanic surface of the round window membrane, unassociated with fibrous tags or adhesions, and in some cases contained congested vascular channels and melanocytes. The webby foci were sometimes associate; with melanocytes, while melanocytes were present without demonstrable webby foci. These findings may represent a collateral route of vascular or perilymph-related drainage from the inner ear.


Half-list speech discrimination measures in hearing aid evaluations.

The usefulness of half-list speech discrimination tests for evaluating hearing aids was assessed. Test and retest speech discrimination scores (SDSs) were obtained from 23 adults with moderate hearing impairment using four preselected hearing aids. Reliability coefficients were generally lower for half-lists than for full-lists, and the rankings of the hearing aids on the basis of SDSs were less consistent for the half-lists than for the full-lists. Individual subject differences between half-list SDSs were 10% or greater in more than one third of the cases. It was concluded that half-lists derived from the NU-6 full-lists should not be used to differentiate among hearing aids.


Multiple epiglottic cysts.

A 66-year-old man had five cystic lesions of the epiglottis, which were discovered at autopsy. He had complained of hoarseness and dysphagia, but no laryngeal examination was performed because of his terminal condition due to widespread adenocarcinoma that did not involve the larynx. To our knowledge, this is the fourth reported case of multiple laryngeal ductal cysts. To develop a concept of the pathogenesis of these cysts, the normal salivary gland-duct epithelial histologic characteristics were studied in 17 epiglottises that were obtained from children and adults at autopsy. We conclude that the cysts arise from the collecting ducts rather than the smaller interlobular or intralobular ducts.


Myocardial infarction--the problems and prognosis: Paper 3--A summary.

In the pre-hospital phase of myocardial infarction, the overriding problem is our present inability to alter the fact that approximately 50 per cent of all deaths from infarction occur prior to the patient's arrival in hospital, despite the knowledge that these deaths are due largely to potentially correctible arrythmias.


Aspects of myocardial ischaemia: sudden death.

The majority of patients who die suddenly have significant coronary artery disease, and it seems likely that myocardial ischaemia, in consequence of these diseased arteries, plays an aetiological role in causation of sudden death.


Myocardial infarction: mortality reduction through coronary care?

It appears that the mortality from coronary heart disease, particularly in men between the ages of thirty and sixty-four, has fallen slightly in recent years. The reasons for this are not clear, but some of this mortality reduction has been attributed to better management of patients, arising through the introduction of coronary care and other forms of medical interference. It is equally possible that this reduction of overall mortality is related to changes in the community's habits.


Protection from morbidity and mortality from first infarcts: the case for beta-blockade.

The original concept of alpha and beta sympathetic receptors put forward by Alquist in 1948 suggesting that beta receptors were situated mainly in the heart and alpha receptors peripherally, gave rise to the theory that selective blockade of beta receptors might have the effect of relieving the heart from excessive sympathetic stimulation. This possibility was developed by Black and Stephenson with the birth of the first beta-blocking drugs--pronethalol and propranolol. The importance of such a drug was foreseen by Chamberlain who showed subsequently that propranolol and cervical sympathectomy reduced an exercise tachycardia to the same extent. Pronethalol was not developed for clinical use and propranolol was in any event found to be the more active drug with no demonstrable hazards. Propranolol was marketed for clinical use as 'Inderal' by the Pharmaceuticals Division of Imperial Chemical Industries, UK, where this original work had been done.


Surgery and the aged.

Surgery for the aged presents a special challenge for all members of the hospital team. The tone of a geriatric hospital is unique. It has usually been adapted specifically for the needs of the elderly. Quiet kindness replaces excessive urgency; a slower time scale allows unrushed opportunities for patient care and contact; and a friendly family spirit helps replace what is missing in the lives of many elderly patients.


The place of surgery in rheumatoid arthritis.

The medical and surgical management of patients with rheumatoid disease has improved over recent years. Despite the improvements, much disability and pain arise from the condition. The major surgical advances in recent years have been in the field of joint replacement, and these advances are continuing. Joint replacement units have evolved, allowing further specialization in this rapidly advancing field. Close liaison between the general practitioner, rheumatologist, orthopaedic surgeon and allied health professionals is most important in achieving the best possible results in the management of these patients.


Crystal induced arthritis: gout and pseudogout.

Intrasynovial deposits of monourate crystals in the presence of serum hyperuricaemia, and calcium pyrophosphate dihydrate (CPPD) crystals, are responsible for gout and pseudogout respectively. Identification of these by synovial fluid analysis is described. The clinical features, minimum investigations, history-taking, management and drug regimes of gout and pseudogout are discussed. Periodic review of the patient is stressed. Other intrasynovial crystals are briefly outlined.


Intestinal transport of monosaccharide after biliary diversion in the rat.

Sugar absorption is increased in rats with a bile fistula but approaches normal values with the addition of bile salt. It has therefore been suggested that bile salts have a physiological role in decreasing intestinal absorption of monosaccharides. In experiments using rats, jejunal and ileal uptake of arbutin, a glucose analogue was increased 5 days after creating a bile fistula but normal by the 10th day after operation. Bile fistula rats ate only about one third of the intake of normal rats in the first 5 days after operation. Control animals fed the same amount as the bile fistula group showed a similar increase in jejunal and ileal arbutin uptake. In both groups, on the 5th post-operative day, addition of taurocholate depressed arbutin uptake towards normal. In normal rats, taurocholate depressed arbutin uptake in the ileum but not the jejunum. These results suggest that increased monosaccharide uptake in bile fistula rats is related to semi-starvation and is not a specific effect of bile salts.


Turnover rates of muscle protein in cardiac and skeletal muscles of dog, fowl, rat and mouse: turnover rate related to muscle function.

Turnover rates of muscle protein in cardiac and skeletal muscle have been measured in dogs using a continuous infusion of 14C-tyrosine, and in fowls, rats and mice after a pulse label of either 3H-leucine or 3H-aspartate. In all species the total mixed protein from ventricle turned over the most rapidly. Tonic muscles predominantly comprised of red fibres were intermediate in rate and twitch muscles comprised of predominantly white fibres had the slowest rates of turnover. This same progression of turnover rates was also seen in myofibrillar and sarcoplasmic protein fractions and with myosin. The RNA concentrations of the muscle tissue were highly correlated with the turnover rates. It is suggested that the turnover rate of muscle protein is related to the function of the muscle.


Chromosome damage and DNA repair induced in human fibroblasts by UV and cholesterol oxide.

In human fibroblasts, cholesterol oxide induced a similar degree of chromosome damage (8.6% of metaphases) and DNA repair synthesis (8-10% of cells with lightly-labelled nuclei) as low doses of ultraviolet light (UV), but did not produce single-strand DNA breaks or DNA damage detectable by inhibition of thymidine incorporation. Chromosome aberrations were detected up to 8 weeks after treatment with cholesterol oxide and UV. Combined treatments had almost additive effects on the frequency of chromosome aberrations but not on repair synthesis. Multiple daily doses of UV did not cause more aberrations than a single dose. Attempts to transform two fibroblast strains from normal donors and three derived from melanoma patients using single and combined treatments of UV, cholesterol oxide and hyperthermia (40 degrees) were unsuccessful.


Effect of hydrocortisone on the response of rat lymphocytes to phytohaemagglutinin.

The proliferative response of rat peripheral blood lymphocytes and spleen cells to phytohaemagglutinin (PHA) was studied following treatment with single or multiple doses of hydrocortisone. A transient lymphopaenia with a rapid recovery at 24 h occurred with both dose schedules. Hydrocortisone in vitro caused a dose-dependent depression of blood lymphocyte response to PHA but enhanced the response of spleen cells when the steroid was added after the mitogen. An inverse relationship between blood and splenic lymphocyte responses to PHA occurred after a single dose of hydrocortisone. Blood lymphocytes from multiple dose hydrocortisone-treated rats had an enhanced response to PHA. Serum collected within 5 min of injection from single dose-treated rats depressed normal rat blood lymphocyte responses. After multiple doses of hydrocortisone serum enhanced blood lymphocyte responses to PHA. The primary antibody response to sheep erythrocytes was only affected by near toxic doses of hydrocortisone.


Suppression of skin reactivity in rats bearing IgE or IgG2a immunocytomas.

Passive cutaneous anaphylaxis (PCA) and reversed PCA have been examined in rats bearing IgE or IgG2a immunocytomas. PCA reactivity was completely abolished in rats bearing an IgE immunocytoma, whereas only a slight suppression of PCA reactivity was found in rats bearing an IgG2a immunocytoma. Reversed PCA reactivity was only slightly decreased in rats bearing IgE or IgG2a immunocytomas.


Sensitisation of gut-associated lymphoid tissue during oral immunisation.

The time course of appearance of sensitised lymphocytes was studied in rabbits fed 0.1% bovine serum albumin (BSA). Antigen reactive cells (ARC) were detected in Peyer's patches at 1 week, prior to both the appearance of primed cells in the spleen and the detection of circulating anti-BSA antibody. Systemic immunisation with 2 mg BSA induced sensitised cells in the spleen and Peyer's patches. No evidence of an early appearance of sensitised cells in the mucosal cell population was found. These data support the postulates that the initial lymphocyte sensitisation to ingested soluble antigen occurs in Peyer's patches, that this lymphoid tissue provides a significant contribution to systemic immunity following oral and parenteral immunisation, and that the mucosal lymphocyte population is not involved in the early phase of sensitisation to ingested antigen.


Fusion and hybridization of marsupial and eutherian cells. Activity of heterokaryons.

Marsupial and eutherian nuclei in heterokaryons were shown to synthesize DNA and RNA, apparently at control levels, and heterokaryons were found to contain marsupial hypoxanthine-guanine phosphoribosyltransferase. The two types of nucleus in heterokaryons were also able to undergo synchronous chromosome condensation. our results provide no evidence for suppression of nucleic acid synthesis, gene expression or mitosis in marsupial x eutherian heterokaryons.


The RNA genome of Tilligerry virus.

RNA extracted from Tilligerry virus, a member of the Eubenangee group of orbiviruses, separated into ten segments after electrophoresis in acrylamide gels. The molecular weights of most segments differed from the corresponding segments of Eubenangee virus.


In vitro growth of ovine squamous cell carcinoma.

Cultured by fragment explantation following a selective trypsinisation procedure, 22/31 (71%) specimens of ovine squamous cell carcinoma yielded pure strains of epithelial cells which to date have passaged vigorously to the 29th level of passage. In contrast to the diploid nature of epithelial cell strains derived from foetal sheep skin, the cell strains derived from skin tumors were highly aneuploid.


Comparative stability of alanine aminotransferase in rat plasma and hepatocyte suspensions.

Alanine aminotransferase (ALT) activity in rat plasma was stable for up to 56 days when stored at -25 degrees. High activity plasma samples were less stable. ALT activity was markedly unstable when derived from isolated hepatocyte preparations, and declined to 8-16% of initial values after 28-56 days storage at -25 degrees.


A cell culture vaccine against bovine ephemeral fever.

A vaccine was prepared from cell culture fluids harvested from the twelfth passage of the 919 strain of bovine ephemeral fever (BEF) virus in Vero cell cultures. Cattle were vaccinated subcutaneously with various combinations of strain 919 virus and adjuvants. Neutralising antibodies were assayed at various times after vaccination and some cattle were challenged by intravenous inoculation with the virulent 417WBC strain of BEF virus. Strain 919 virus of the third and twelfth passage levels in Vero cells produced neither fever, clinical illness nor detectable viraemia in 5 calves inoculated intravenously. Nor could viraemia be detected in 5 heifers receiving vaccine subcutaneously. When the vaccine was administered mixed with aluminium hydroxide adjuvant, the production of neutralising antibodies increased with an increase in the volume of vaccine from 2.5 ml to 10 ml and the response to 2 injections was significantly better than the response to a single injection. The neutralising antibody response was decreased when vaccine was diluted in phosphate buffered saline. The neutralising antibody response following 2 subcutaneous vaccinations with strain 919 virus mixed with aluminium hydroxide adjuvant was higher than that following intravenous inoculation with virulent virus. The vaccine-induced antibodies persisted for at least 12 months, and revaccination at this time led to an increase in the titre of neutralising antibody. Antibodies induced by a single subcutaneous administration of strain 919 virus mixed with Freund's complete adjuvant persisted for at least 40 weeks; those induced by vaccine containing Freund's incomplete adjuvant had virtually disappeared within 16 weeks. All these calves responded to vaccination with aluminium hydroxide-containing vaccine with increases in levels of neutralising antibodies. Of 26 vaccinated calves challenged with virulent BEF virus, 24 remained clinically normal. Two developed brief periods of pyrexia on the seventh day after challenge, but no other clinical signs. One of these calves had a viraemia that was demonstrated only by intravenous inoculation of a susceptible calf. The remaining calf had no detectable viraemia. All of 7 unvaccinated calves developed severe clinical BEF within 5 days of challenge. No disease attributable to the 919 virus occurred in 24 vaccinated pregnant heifers or their newborn calves.


Evaluation of new and currently used diagnostic procedures for bovine brucellosis.

The indirect haemolysis test (IHLT) and the rough antigen complement-fixation test (RCFT) were compared with several conventional tests using serum samples from 9 cows known to be infected with Brucella abortus. In 7 cows all the tests except the RCFT (which was developed to detect antibodies resulting from 45/20 vaccination) became positive and remained so until the cows were autopsied 6 months after infection. In the other 2 cows the Rose Bengal test was occasionally negative and the titre of the serum agglutination test only reached 100 iu briefly. The routine complement-fixation test, the IHLT and the anti-bovine globulin test reached diagnostic levels at most of the weekly test intervals but occasional low titres occurred in them all. Only very low or negative titres were detected by the RCFT.


Breed differences in the response of young beef bulls to electro-ejaculation.

A total of 702 electro-ejaculation attempts are recorded on young (16 to 31 months) beef bulls of 3 breeds and 3 breed crosses (Brahman cross 100, Africander cross 93, Shorthorn cross 109, Brahman 81, Africander 89, and Hereford 230. Four hundred and forty of these attempts were conducted on bulls of the first 5 of these breeds which had been reared under similar conditions on the same property. The machine employed was a Watson Transtimulator, powered by a 12-volt automobile battery, and connected to a 6.25 cm diameter bipolar ring electrode probe, 37.5 cm in length. The voltages at which the sperm free (sample A) and sperm rich (sample B) samples were initiated were 4.8 +/- .2 and 7.7 +/- .2 volts respectively. No breed differences occurred. Breed differences occurred in the proportion of bulls showing a severe reaction to the electricity (P less than 0.001) with the Africander (32.6%; P less than 0.001) and Ax (28.0%; P less than 0.01) having the highest proportions of severe reactions. These 2 breed groups also had the highest proportions of electro-ejaculation failures (respectively, 23.6%; P less than 0.05 and 19.4%; n.s.). Overall, bulls of Bos indicus derivation did not differ from those of Bos taurus derivation in electro-ejaculation failures. During stimulation, erection was achieved by 19.8% of bulls and failure to collect an assessable ejaculate occurred in 15%. No breed differences occurred in either category. Collapse during stimulation occurred in 5.3% of electro-ejaculation attempts (overall P less than 0.05) with the SH breed group having highest representation (12.8%; P less than 0.001).


The influence of the time of treatment in relation to feeding on the efficacy of amoxycillin therapy in dogs and cats.

A field trial comparing the clinically assessed response to amoxycillin therapy in 148 fasted and 129 non-fasted dogs and cats suffering clinically diagnosed bacterial infections is reported. A satisfactory response to treatment was obtained in 88.8% of these cases. The results indicate that the clinical response to amoxycillin therapy was not affected by the concurrent administration of the antibiotic with food.


Evaluation of oral iron galactan as a method of iron supplementation for intensively housed sucking piglets.

Iron supplementation of piglets with oral galactan given as a single dose within 24 hours of birth was evaluated in a series of on farm trials. The growth rate of piglets receiving this treatment was faster than that of piglets receiving single injections of iron dextran or iron galactan at 3 days of age, or ferrous sulphate crystals orally at weekly intervals. Mean values for red cell parameters of oral iron galactan supplemented piglets at 2 to 4 weeks of age were lower than those of injected piglets but there was no clinical evidence of anaemia in any of the piglets.


The effect of anthelmintic treatment and other management factors on the incidence of breech strike in Merino sheep.

Repeated scouring and soiling of the breech region with faeces rendered mulesed, Merino sheep susceptible to breech strike. Anthelmintic treatment of ewes before and after lambing and of lambs 6 to 8 weeks old reduced intestinal trichostrongylosis, scouring and consequent breech strike. Wether lambs tended to be more susceptible than ewe lambs and sheep grazing long pastures were struck more frequently than those on short pastures.


The importance of the radical mules operation and tail length for the control of breech strike in scouring Merino sheep.

While the radical Mules operation reduced the incidence of breech strike from 59% to 13% in scouring sheep, the problem was further alleviated by docking lambs' tails at the correct (medium) length. Docking lambs' tails very short strongly predisposed radically mulesed ewes and wethers to faecal soiling and breech strike (20% struck). Radically mulesed sheep with tails docked midway down the vulval orifice in ewes, and the corresponding length in wethers, were rarely struck (3%) and did not appear to require crutching to prevent breech strike associated with scouring or urine staining.


Preparation of teaser bulls by dorsal scrotal penile deflection.

A simple, quick and reliable technique of preparing teaser bulls has been developed. Four Bos indicus aged between 1 year 6 months and 2 years were subjected to this method by deflecting their penes backwards about 2 to 3 cm posterior and dorsal to the attachment of the scrotum. No serious postoperative complications were recorded. The sexual behaviour and libido of the bulls did not change after subjecting them to this technique.


Objective assessment of prior air traffic control-related experience through the use of an occupational knowledge test.

The Occupational Knowledge Test (OKT) 101-B was administered to 784 air traffic control trainees who entered the FAA Academy's 16-week training course in 1976. All trainees completed the nonradar laboratory portion of the training and, in addition, completed a preemployment questionnaire. Based on responses to the questionnaire, the trainees were assigned to one of three experience groups, corresponding to groups given credit for experience using Civil Service Commission (CSC) selection procedures. It was found that the OKT was highly correlated with experience (r = 0.64) and, in addition, had a higher correlation with successful completion of the nonradar lab than did experience (r = 0.25 vs. 0.12). It was determined that use of an OKT score of 75 or above to assign extra credit would result in a failure rate of 3.1% for those receiving credit, while use of the current experience scale would result in a failure rate of 7.6% for those receiving extra credit on the CSC selection battery. The results held up for a cross-validation sample of 432 trainees who entered the Academy during 1977. Based on the results, it is recommended that an OKT score of 75 or above be used to assign extra credit for experience in the selection of air traffic controllers.


Quantifying fatigue in working divers.

Thirty professional divers involved in training, bounce dives, working saturations, and deep experimental saturations were observed over a 1-month period. The subjects themselves performed simple twice-daily measurements of oral temperature, heart rate, breathhold time, handgrip strength, length and quality of sleep, and a subjective estimation of fatigue. The data were analysed with respect to the type of work done by the diver, separating diving from nondiving days. The results showed that subjective estimation of fatigue corresponded to the increment of evening oral temperature and heart rate over morning values. Both inspiratory and expiratory breathhold times decreased from morning to evening in those situations deemed most tiring by the divers. Sleep was generally of average duration and quality; however, diving during the daytime was associated with a decreased amount of sleep in the 24-h period including the following night. Sleep was also of poorer quality during periods of saturation diving. In addition to such group variations, individual divers showed significant performance changes on the various tests, demonstrating the value of this approach to the practical question of deciding when a man is too tired to dive safely. Extension of this method can, we hope, aid in the definition of safe working rotations for professional divers.


Luminance thresholds of the water-immersed eye.

Luminance thresholds were measured for the water-immersed eye as a function of adaptation level, viewing distance, and turbidity of the water, and compared to standard adaptation curves. In water, thresholds varied as a function of initial adaptation level, and the rate of adaptation paralleled that for the eye in air. However, thresholds were about 1-2 log units higher for the water-immersed eye than for the eye in air at all adaptation levels.


Ripcord release capability of female parachutists.

With the growth in popularity of skydiving as a sport has come an associated increase in parachuting accidents, some of them due to failure to deploy the main or reserve parachutes. The absence of appropriate strength data has prompted a study of the pull force capabilities of female parachutists on a ripcord release in varying positions. Pull forces exerted for periods of 0.25, 1, 1.5, 2, and 2.5 s during a 5-s pull are presented in percentile form. A substantial percentage of female parachutists could not exert the maximum ripcord release pull force permitted by the relevant parachute specification. The relatively low level of strength exhibited by the weakest groups of the population emphasizes the need for such basic information in equipment design.


Effect of vitamin and amino acid supplements on human performance during heavy mental and physical work.

This paper presents data describing the effects of vitamin and amino acid supplements on the health state and performance of men doing heavy mental and physical work. Experiments on seven test volunteers showed that a 20-d intake of a mixture of vitamins, amino acids, and mineral substances optimized the health state and increased the performance of test subjects doing heavy work.


Workload and fatigue--in-flight EEG changes.

Continuous recordings were made of the EEG and ECG of one pilot during a series of instrument flights in a feeder-type transport aircraft. The flights were arranged to contain epochs of distinctly differing levels of workload. In addition, some flights were made after a night of sleep deprivation and others were made as the second and third flights of the day. Subjectively, there appeared to be marked differences in performance between the two types of tired flight. The EEG analyses showed changes that correlated well with differences in workload. In the highest workload areas during fresh flights, EEG activity increased by approximately a factor of 4 over that of the preflight resting values. This large increase did not occur in the tired flights. Further experiments are planned using flights in aircraft and in simulators using several subjects.


Comparison of the vigilance performance of men and women using a simulated radar task.

The present study examined the question of possible sex differences in the ability to sustain attention to a complex monitoring task requiring only a detection response to critical stimulus changes. The visual display was designed to approximate a futuristic, highly automated air traffic control radar display containing computer-generated alphanumeric symbols. There were 26 men and 26 women tested, each over a 2-h session. Sixteen targets appeared on the screen at all times, with 10 signals (a designated change in the alphanumerics) randomly presented during each 0.5 h of the test session. Detection latency to the signals increased significantly during the session, but there was no evidence of any significant differences between the sexes in the magnitude or pattern of this increase. The results are discussed in terms of a general decline in alertness that was apparently equal for both sexes.


Effects of fracture trauma, estrone treatment, and 2-G environment on the epiphyseal cartilage zones of developing avian radii.

Female birds subjected to closed fracture of the right radius at 2 weeks post-hatching were allowed to heal for 14 d at either earth gravity (1 G) or a 2-G hypergravity state, with saline-injected controls and 0.2- and 0.4-mg estrone (estrogen) treated groups. Animals were sacrificed following the last injection and radii were routinely processed histologically for micrometer measurements of the mid-coronal height and width of the cartilage zones of the proximal and distal epiphyses. Proximal epiphyses of estrone-treated, normo-gravity chicks showed a decreased growth in height of the articular, proliferating, and hypertrophic cartilage zones and an increased interstitial growth of the hypertrophic and calcified layers of estrone-treated, 2-G animals. Growth in height of the cartilage layers of the distal epiphyses of fractured radii was primarily stimulated when estrone-treated chicks were also exposed to the 2-G state. Appositional growth of all cartilage layers of the proximal and distal epiphyses of normo-gravity birds was increased by 0.2 mg estrone. In addition, growth in the width of all cartilage layers of the distal epiphyses of 2-G animals was increased with the administration of 0.4 mg estrone.


U.S. fatal general aviation accidents due to cardiovascular incapacitation: 1974-75.

A study was undertaken to determine the relative impact of inflight cardiovascular incapacitation among general aviation pilots with respect to general aviation flight safety. During calendar years 1974-75, the National Transportation Safety Board reports reveal that 13 U.S. general aviation pilots died of cardiovascular incapacitation during flight. The analysis of these accidents will bear on any suggested changes in pilot medical screening procedures for cardiovascular disease, as well as on pilot safety education programs. Of the 13 cases noted above, nine pilots were flying alone. Of the remaining multiple occupant cases, the nonpilot wife of one deceased victim managed to land the aircraft. Eighteen deaths resulted from the inflight incapacitations. The ages of the pilots ranged from 33-68 years, with both a mean and a median of 52. Postmortem examinations revealed extensive coronary disease (atherosclerosis) in 12 cases (no pilot autopsy data is available in the case where the passenger landed the aircraft). Of these 12 cases, five demonstrated recent occlusions. In four more, evidence of old infarcts was revealed by the postmortem examination. It is concluded that these 13 inflight cardiovascular incapacitations, occurring among a total of 1,404 fatal general aviation accidents in the 1974-75 period, constitute such a small proportion (0.93%) of the documented fatal general aviation accidents that extensive additional cardiovascular screening procedures are not justified at present on cost/yield basis.


Movement by helicopter of patients with decompression sickness.

Rapid movement of a patient with decompression sickness sometimes poses problems when the site of the hyperbaric treatment facility is located a considerable distance away. Six cases of aviator decompression sickness were diagnosed in altitude chamber participants during an 18-month period. Five cases were uncomplicated decompression sickness and the sixth case was of central nervous system decompression sickness. All cases were transferred by low-level helicopter flight. No complications were noted when the helicopter stayed within 200 ft (61 m) AGL of the take-off point. Symptoms of decompression sickness did worsen however, when this altitude was exceeded. This study shows that movement of patients with decompression sickness by low-level helicopter flight is both safe and effective, especially when pressurized aircraft is neither available nor practical.


Bleeding duodenal ulcer and the flier.

Over the years, the aeromedical disposition of aircrew members with bleeding duodenal ulcer has been a vexing problem because two essential questions cannot be categorically answered: Is the risk of rebleeding less with medical or with surgical treatment? In either case, is the risk of rebleeding low enough to permit, with reasonable prudence, the resumption of aircrew duties by the aviator? To try to answer these questions, the author has reviewed a portion of the literature and a number of medical records of patients with bleeding duodenal ulcer. With this information, the author draws his conclusions and offers recommendations for the aeromedical disposition of such aircrewmen.


The nitrogenase system of Spirillum lipoferum.

The nitrogenase system of Spirillum lipoferum was separated into three components, the normal Mo-Fe and Fe proteins as well as an activating factor for the Fe protein. The rate of activation is increased by Mn2+ or by an excess of Mg2+, and the process requires ATP.


Comparative studies on intrinsic factor and cobalophilin in different parts of the gastrointestinal tract of the pig.

The vitamin B(12) binders in the pig pyloric mucosa gastric and intestinal juice from the upper gastrointestinal tract were fractionated into only two molecular forms, classified as intrinsic factor and cobalophilin. The unsaturated vitamin B(12)-binding power due to cobalophilin was lower in the intestinal than in the gastric juice. Electrofocusing revealed that intrinsic factor and cobalophilin in the intestinal juice contained more of the ;neutral'-type isoproteins, and the suggestion is made that this is due to enzyme activity. The gastric-juice intrinsic factor contained more acidic isoproteins, which supports the hypothesis that carbohydrate is added on to the polypeptide chain of this protein before it is secreted into gastric juice. The gastric- and intestinal-juice cobalophilins, studied also by electrofocusing, differed from that of pyloric mucosa and they appeared to be of salivary origin. With regard to molecular dimensions there was no significant difference between the intrinsic factors and cobalophilins from all sources studied. All cobalophilins had molecular weights by the formula of Svedberg of approx. 92500, Stokes radii of 4.62nm and sedimentation coefficients of 5.15S. The corresponding values for the intrinsic factors were 63600, 3.57nm and 4.38S. In addition, the intrinsic factors exhibited similar avidities for binding to the solubilized ileal intrinsic-factor receptor. Also the intrinsic factors and cobalophilins, irrespective of their source, bound to the analogous specific xenoantibodies with the same avidity. The present results demonstrate that intrinsic factor remains practically unaltered during its passage through the proximal intestine and render unlikely the speculations made about the presence of an endogenous binder for intrinsic factor as well as the existence of a ;pancreatic intrinsic factor'. In addition, they are compatible with the theory that the interference by undegraded cobalophilin may be the reason for the abnormal vitamin B(12) absorption observed in patients with pancreatic insufficiency.


The role of nitro groups in the binding of nitroaromatics to protein MOPC 315.

Two series of dinitrophenyl haptens, in which chlorine replaces one or both nitro groups, were used to investigate, by a combination of high-resolution 1H n.m.r. and fluorescence quenching, the presence of groups in the combining site of protein MOPC 315, which form hydrogen bonds to the aromatic-ring substituents of the hapten. The large differences in binding constants on successive replacement of nitro groups were shown to be due to specific hapten-substituent-protein interactions by (a) showing that there was little difference in the interaction between these haptens and 3-methylindole (a model for the residue tryptophan-93L with which the hapten stacks in protein MOPC 315), (b) proving by 1H n.m.r. that the mode of hapten binding is constant and (c) showing that the differences in Kd were consistent with the relative hydrogen-bonding capacities of chlorine and the nitro moiety. In this way it was established that each nitro group forms a hydrogen bond. Furthermore, from consideration of the 1H n.m.r. chemical shifts of several dinitrophenyl haptens and their trinitrophenyl analogues, it was shown that there is no distortion of the o-nitro group on binding to the variable fragment of protein MOPC 315.


Protein thiolation and reversible protein-protein conjugation. N-Succinimidyl 3-(2-pyridyldithio)propionate, a new heterobifunctional reagent.

A heterobifunctional reagent, N-succinimidyl 3-(2-pyridyldithio)propionate, was synthesized. Its N-hydroxysuccinimide ester group reacts with amino groups and the 2-pyridyl disulphide structure reacts with aliphatic thiols. A new thiolation procedure for proteins is based on this reagent. The procedure involves two steps. First, 2-pyridyl disulphide structures are introduced into the protein by the reaction of some of its amino groups with the N-hydroxysuccinimide ester sie of the reagent. The protein-bound 2-pyridyl disulphide structures are then reduced with dithiothreitol. This reaction can be carried out without concomitant reduction of native disulphide bonds. The technique has been used for the introduction of thiol groups de novo into ribonuclease, gamma-globulin, alpha-amylase and horseradish peroxidase. N-Succinimidyl 3-(2-pyridyldithio)propionate can also be used for the preparation of protein-protein conjugates. This application is based on the fact that protein-2-pyridyl disulphide derivatives (formed from the reaction of non-thiol proteins with the reagent) react with thiol-containing proteins (with native thiols or thiolated by, for example, the method described above) via thiol-disulphide exchange to form disulphide-linked protein-protein conjugates. This conjugation technique has been used for the preparation of an alpha-amylase-urease, a ribonuclease-albumin and a peroxidase-rabbit anti-(human transferrin) antibody conjugate. The disulphide bridges between the protein molecules can easily be split by reduction or by thiol-disulphide exchange. Thus conjugation is reversible. This has been demonstrated by scission of the ribonuclease-albumin and the alpha-amylase-urease conjugate into their components with dithiothreitol. N-Succinimidyl 3-(2-pyridyldithio)propionate has been prepared in crystalline form, in which state (if protected against humidity) it is stable on storage at room temperature (23 degrees C).


Formation and subsequent removal of O6-methylguanine from deoxyribonucleic acid in rat liver and kidney after small doses of dimethylnitrosamine.

1. The amounts of 7-methylguanine and O(6)-methylguanine present in the DNA of liver and kidney of rats 4h and 24h after administration of low doses of dimethylnitrosamine were measured. 2. O(6)-Methylguanine was rapidly removed from liver DNA so that less than 15% of the expected amount (on the basis of 7-methylguanine found) was present within 4h after doses of 0.25mg/kg body wt. or less. Within 24h of administration of dimethylnitrosamine at doses of 1mg/kg or below, more than 85% of the expected amount of O(6)-methylguanine was removed. Removal was most efficient (defined in terms of the percentage of the O(6)-methylguanine formed that was subsequently lost within 24h) after doses of 0.25-0.5mg/kg body wt. At doses greater or less than this the removal was less efficient, even though the absolute amount of O(6)-methylguanine lost during 24h increased with the dose of dimethylnitrosamine over the entire range of doses from 0.001 to 20mg/kg body wt. 3. Alkylation of kidney DNA after intraperitoneal injections of 1-50mug of dimethylnitrosamine/kg body wt. occurred at about one-tenth the extent of alkylation of liver DNA. Removal of O(6)-methylguanine from the DNA also took place in the kidney, but was slower than in the liver. 4. After oral administration of these doses of dimethylnitrosamine, the alkylation of kidney DNA was much less than after intraperitoneal administration and represented only 1-2% of that found in the liver. 5. Alkylation of liver and kidney DNA was readily detectable when measured 24h after the final injection in rats that received daily injections of 1mug of [(3)H]dimethylnitrosamine/kg for 2 or 3 weeks. After 3 weeks, O(6)-methylguanine contents in the liver DNA were about 1% of the 7-methylguanine contents. The amount of 7-methylguanine in the liver DNA was 10 times that in the kidney DNA, but liver O(6)-methylguanine contents were only twice those in the kidney. 6. Extracts able to catalyse the removal of O(6)-methylguanine from alkylated DNA in vitro were isolated from liver and kidney. These extracts did not lead to the loss of 7-methylguanine from DNA. 7. The possible relevance of the formation and removal of O(6)-methylguanine in DNA to the risk of tumour induction by exposure to low concentrations of dimethylnitrosamine is discussed.


Purification and properties of succinyl-coenzyme A-3-oxo acid coenzyme A-transferase from sheep kidney.

CoA-transferase (succinyl-CoA-3-oxo acid CoA-transferase, EC 2.8.3.5) isolated from sheep kidney was purified to homogeneity. The purified enzyme has a specific activity of approx. 200 units/mg. A mol.wt. of 110000 was obtained by gel filtration on Sephadex G-200, and a lower mol.wt. of 102000 was determined by analytical ultracentrifugation. A sedimentation coefficient of 5.6S was also determined. A subunit mol.wt. of 56000 was obtained by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Isoelectric focusing of sheep kidney extracts indicated the presence of a single band of CoA-transferase activity with pI9.0. However, isoelectric focusing of purified CoA-transferase showed the presence of two peaks of CoA-transferase activity with pI values of 8.7 and 8.4, suggesting the presence of proteolytic activity during purification. Evidence for sheep kidney CoA-transferase being a dimer of two identical subunits has been obtained from sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, the amino acid composition, peptide 'mapping' and N-terminal analysis.


Structure and enzymic activity of ribonuclease-A esterified at glutamic acid-49 and aspartic acid-53.

The dimethyl ester of bovine pancreatic ribonuclease-A (dimethyl RNAase-A), the initial product of esterification of RNAase-A in anhydrous methanolic HCl, was isolated in a homogeneous form. The two carboxy functions esterified in this derivative are those of glutamic acid-49 and aspartic acid-53. There were no changes in the u.v.-absorption spectral characteristics, the accessibility of the methionine residues, the resistance of the protein to proteolysis by trypsin and the antigenic behaviour of RNAase-A as a result of the esterification of these two carboxy groups. Dimethyl RNAase-A exhibited only 65% of the specific activity of RNAase-A, but still had the same K(m) value for both RNA and 2':3'-cyclic CMP. However, the V(max.) was decreased by about 35%. On careful hydrolysis of the methyl ester groups at pH9.5, dimethyl RNAase-A was converted back into RNAase-A. Limited proteolysis of dimethyl RNAase-A by subtilisin resulted in the formation of an active RNAase-S-type derivative, namely dimethyl RNAase-S, which was chromatographically distinct from dimethyl RNAase-A and had very nearly the same enzymic activity as dimethyl RNAase-A. Fractionation of dimethyl RNAase-S by trichloroacetic acid yielded dimethyl RNAase-S-protein and dimethyl RNAase-S-peptide, both of which were inactive by themselves but regenerated dimethyl RNAase-S when mixed together. Dimethyl RNAase-A-peptide was identical with RNAase-S-peptide. RNAase-S-protein could be generated from dimethyl RNAase-S-protein by careful hydrolysis of the methyl ester groups at pH9.5. The interaction of dimethyl RNAase-S-protein with RNAase-S-peptide appears to be about 4-fold weaker than that between the RNAase-S-protein and RNAase-S-peptide. Conceivably, the binding of the S-peptide ;tail' of dimethyl RNAase-A with the remainder of the molecule is similarly weaker than that in RNAase-A, and this brings about subtle changes in the geometrical orientation of the active-site amino acid residues of these modified methyl ester derivatives. It is suggested that these changes could be responsible for the generation of the catalytically less-efficient RNAase-A and RNAase-S molecules (dimethyl RNAase-A and dimethyl RNAase-S respectively).


The oxygenation of [3-methyl-3H]desacetoxycephalosporin C [7beta-(5-D-aminadipamido)-3-methylceph-3-em-4-carboxylic acid] to [3-hydroxymethyl-3H]desacetylcephalosporin C by 2-oxoglutarate-linked dioxygenases from Acremonium chrysogenum and Streptomyces clavuligerus.

Cell-free extracts of Acremonium chrysogenum and Streptomyces clavuligerus oxidize the 3-methyl group of desacetoxycephalosporin C to a 3-hydroxymethyl group. The enzyme responsible for this reaction in these organisms was purified 20- and 30-fold respectively by chromatography on DEAE-cellulose. The enzymes, which were assayed with [3-methyl-3H]desacetoxycephalosporin C as substrate, have the properties expected of 2-oxoglutarate-linked dioxygenases. They require 2-oxoglutarate, Fe2+ cations and a mixture of reducing agents (dithiothreitol and ascorbate) for full activity. The enzyme from A. chrysogenum, but not that S. clavuligerus, is activated about 10-fold when it is preincubated with a reaction mixture from which either desacetoxycephalosporin C or 2-oxoglutarate is omitted. Fe2+ cations seem to play a key role in this activation. Both enzymes seem highly specific for cephalosporins with the natural 7beta-(5-D-aminoadipamido) side chain and are likely to be responsible for the oxidation of the 3-methylcephem nucleus in vivo.


The interaction of purified acetylcholinesterase from pig brain with liposomes.

The binding of pig brain acetylcholinesterase to artificial phospholipid membranes was investigated at different temperatures. Calculation of the thermodynamic parameters revealed a small negative enthalpy change, but a large negative change in the free energy and a large positive change in the entropy on binding. The large entropy change might be interpreted as being responsible for forming the enzyme-membrane complex and was indicative of hydrophobic interactions between lipid and protein. This conclusion would also favour the hypothesis that the enzyme was an integral protein. Further support for this theory was provided by the study of acetylcholinesterase binding to liposomes containing the phospholipid 1,2-dimyristoyl-sn-glycero-3-phosphocholine. Lowering the temperature below the transition temperature or incorporating cholesterol into the liposomes decreased enzyme binding. Both factors could be interpreted as decreasing the fluidity of the hydrocarbon side chains of the phospholipids, causing an increase in bilayer thickness due to closer packing of side chains. This membrane condensation would certainly not favour the binding of integral protein molecules.


Amino acid sequence of the N-terminal 108 amino acid residues of the B chain of subcomponent C1q of the first component of human complement.

The amino acid sequence of the N-terminal 108 residues of the B chain of subcomponent C1q of the first component of human complement was determined. The B chain has a blocked N-terminal amino acid, which was judged to be 5-oxopyrrolidine-2-carboxylic acid. A collagen-like region of 84 residues was found, which started at position B-6, and all of the six hydroxylysine residues and 12 hydroxyproline residues present in the chain were found in this region. Four of the six hydroxylysine residues may be glycosylated. The repeating nature of the collagen-like region is broken at position B-9, where alanine is found in a position where glycine would be expected. The exact position of the interchain disulphide bond joining the A and B chains of human subcomponent C1q was shown to be between residues A4 and B4.


Effect of heparin on thrombin inactivation by antithrombin-III.

The inactivation of thrombin by heat and by its physiological inhibitor, antithrombin-III, shows quite different dependence on heparin concentration. Heparin at 250 microgram/ml protects thrombin against heat inactivation, and thrombin behaves heterogeneously in this reaction. In the absence of heparin, the thermodynamic activation parameters change with temperature (deltaH+ = 733 kJ/mol and 210 kJ/mol at 50 and 58 degrees C respectively). When heparin is present, heat inactivation of the protected thrombin species proceeds with deltaH+ = 88 kJ/mol and is independent of temperature in the same range. On the other hand, heparin at 0.125-2.5 microgram/ml accelerates the thrombin-antithrombin-III reaction. Thrombin does not show heterogeneity in this reaction and the time courses at any heparin concentration and any temperature between 0 and 37 degrees C appear to follow first-order kinetics. Activation enthalpy is independent of heparin concentration or temperature, deltaH+ = 82-101 kJ/mol, varying slightly with antithrombin-III concentration and thrombin specific activity. Heparin seems to exert its effect by increasing activation entropy. On the basis of these data we suggest a mechanism of action of heparin in the thrombin-antithrombin-III reaction which accounts for all the important features of the latter and seems to unify the different hypotheses that have been advanced.


Removal by transamination and scission of residues from the peptide representing the copper-transport site of serum albumin.

The peptide Asp-Ala-His-NH-Me was subjected to removal of its N-terminal residue by transamination and scission. Despite the high affinity of the peptide for Cu2+ ions, they catalysed its transamination smoothly. Two main transamination products were found, a complication previously observed with another peptide with an N-terminal aspartic residue, but their scission gave a single product, Ala-His-NH-Me. This was subjected to a further cycle of transamination and scission, and gave a single product after each step. For scission of transaminated peptides it proved unnecessary to remove them from transamination reagents provided that transamination was stopped with EDTA before adding the scission reagent.


Protein and lipid components of the pigeon erythrocyte membrane.

The plasma membrane of the nucleated pigeon erythrocyte was isolated by a method that is simple, reproducible and minimally disruptive, the final preparation consisting of whole cell 'ghosts', recovered at over 40% yield. Alternative methods, which yield membrane fragments, were also tested and some of their possible disadvantages demonstrated. Analysis of the protein components of the isolated membranes by gel elctrophoresis in the presence of sodium dodecyl sulphate revealed that their composition is very similar to that of the proteins of human erythrocyte membranes. However, two major proteins are unique to the nucleated cell membrane; these have apparent mol.wts. of 97000 and 57000. Also, the bands designated 4.2 (74500 mol.wt.) and 6 (35000 mol wt.) by Steck [(1974) J. Cell Biol. 62, 1-19] for the human cell membrane are absent from pigon cell membrane. Glycosylated membrane proteins could not be detected in gels stained with the periodate-Schiff-base procedure. Analysis of membrane phospholipids revealed the same components known to be present in mammalian erythrocytes, though in different proportions. These findings are discussed in the light of known physiological and biochemical differences between avian and mature mammalian erythrocytes.


Solubilization of isolated central-nervous-system myelin preparations by the amniotic detergent sodium dodecyl sulphate.

The mechanism for the solubilization of isolated central-nervous-system myelin by sodium dodecyl sulphate was studied in detail. The release of protein and phospholipid to the 100000 g x 1 h supernatant fraction is dependent on the total amount of detergent relative to the amount of membrane present and on the ionic strength of the solubilization system. Gel-filtration analysis of supernatant fractions indicate that at suboptimal concentrations of detergent these contain lipid-protein complexes. The complete dissociation of the individual protein components from lipid is dependent on the total amount of sodium dodecyl sulphate present in the system. The results indicate that for the analysis of membrane components in sodium dodecyl sulphate it is essential that sufficient detergent is present.


Molecular weight and subunit size of rabbit mammary-gland fatty acid synthetase.

1. The molecular weights of fatty acid synthetases isolated from lactating rabbit, rat, cow and goat mammary glands were estimated by sucrose gradient centrifugation and compared by chromatography on Sepharose 6B. 2. The values obtained for all four enzymes were in the same range (0.40 X 10(6)-0.55 X 10(6)) as that found for other mammalian and avian fatty acid synthetases. The molecular weight found for the rabbit mammary enzyme therefore differs from published values of approx. 0.9 X 10(6). 3. The molecular weights of the subunits of these four synthetases were 225000-242000. Again, the value for the rabbit mammary enzyme differs from published values.


Interactions between different corneal proteoglycans.

Proteoglycans were extracted from bovine cornea with 4M-guanidinium chloride and purified by CsCl-density-gradient centrifugation. Under associative conditions two fractions were found: one capable of forming assemblies of high molecular weight and another lacking this property. The heavier fraction (density 1.59 g/ml) was eluted as a single retarded peak from Sepharose 2B, but on DEAE-Sephadex chromatography, gave two peaks: the first (eluted with 0.75 M-NaCl) contained mainly proteochondroitin sulphate and the second (eluted with 1.25 M-NaCl) mainly proteokeratan sulphate. Each of these proteoglycans was more retarded on Sepharose 2B than was the original sample from density-gradient centrifugation. Re-aggregation was obtained by recombination of the two fractions. The lighter fraction (density 1.44 g/ml), containing predominantly keratan sulphate chains, was eluted from DEAE-Sephadex as a single peak with 1.25 M-NaCl and was retarded on Sepharose 2B: this fraction was not able to form aggregates with proteochondroitin sulphate. Chemical analyses of the carbohydrate and protein moieties of the proteoglycans from DEAE-Sephadex confirmed that, in the cornea, different subunits are present with characteristic aggregation properties and hydrodynamic volumes.


Structural studies on the glycoproteins from bovine cervical mucus.

The depolymerization of bovine cervical glycoprotein resulting from cleavage of disulphide bonds. Pronase digestion and both procedures sequentially was assessed by using gel filtration. Cleavage of disulphide bonds followed by Pronase digestion produced more extensive depolymerization than did either treatment alone, and gel filtration of the products resulted in two major peaks of glycosylated material on Sepharose CL-2B and Sepharose 4B. The glycopolypeptides in both peaks had similar sugar and sulphate compositions, but they migrated to different extents on gel electrophoresis. Electrophoretic studies indicated that both glycopolypeptides were derived from the same glycoprotein molecule and not from a mixture of two similar glycoproteins. Pronase digestion of glycoproteins in which the disulphide bonds had been labelled with iodo-[1-14C]acetamide revealed that most of the cysteine residues were situated in regions susceptible to Pronase. The results show the presence of two types of structural regions in bovine cervical glycoprotein, namely 'naked' peptide or non-glycosylated regions and glycopolypeptide subunit regions in which glycopolypeptides of two different sizes predominate. Comparison of the cervical glycoproteins isolated from mucus secreted during oestrus and pregnancy, by the methods outlined above, did not reveal any structural differences in the glycoproteins to explain the different physical properties of the mucus secreted under these conditions.


Properties of human tissue isoferritins.

1. Human liver ferritin was separated by preparative isoelectric focusing into six fractions. 2. Except for the least acidic fraction the reactivity with antibody against spleen ferritin increased with rising pI, but with antibody against heart ferritin the reactivity decreased. 3. The highest iron content was found in the most acidic isoferritins and progressively decreased with rising pI. 4. Iron uptake was studied in apoferritin prepared from heart and liver ferritin fractions separated by ion-exchange chromatography. There was good correlation between the rate of iron uptake and pI. The most acidic fractions took up iron more rapidly than did the more basic ones. 5. Ferritin was prepared from heart, liver, spleen and kidney. There was little difference on isoelectric focusing between ferritin obtained from normal tissues and the corresponding iron-loaded tissues from patients who had received multiple blood transfusions. The iron-loaked heart ferritin invariably contained relatively more of the basic isoferritins. Normal and iron-overloaded heart ferritins were separated into isoferritin fractions by ion-exchange chromatography, and in each case there was a fall in iron content as the pI increased. The iron content of ferritin from the iron-overloaded heart was higher throughout than that from normal heart. 6. There is a relationship between the rate of iron uptake by apoferritin and pI, and this probably accounts for the variation in iron content of the isoferritins found in human liver and heart.


Haem degradation in abnormal haemoglobins.

The coupled oxidation of certain abnormal haemoglobins leads to different bile-pigment isomer distributions from that of normal haemoglobin. The isomer pattern may be correlated with the structure of the abnormal haemoglobin in the neighbourhood of the haem pocket. This is support for haem degradation by an intramolecular reaction.


Proteins from sciatic-nerve myelin in quaking and jimpy mice.

Myelin from two neurological mutants in mice was isolated from sciatic nerves and its protein composition analysed. In Quaking mice, two intrinsic myelin proteins P1 and P2 were drastically decreased, whereas the major myelin protein P0 was unaffected. A normal protein composition was found in sciatic myelin from Jimpy mice.


A difference in the specificities of human liver N-acetyl-beta-hexosaminidases A and B detected by their activities towards glycosaminoglycan oligosaccharides.

N-Acetyl-beta-hexosaminidases A and B differ in their activities towards oligosaccharides prepared from glycosaminoglycans. Trisaccharides from hyaluronic acid and desulphated chondroitin 4-sulphate were hydrolysed by N-acetyl-beta-hexosaminidase A, but not by N-acetyl-beta-hexosaminidase B.


Oestrogen-induced cholesterol and fatty acid biosynthesis in Xenopus laevis liver during vitellogenic response.

1. Oestradiol-17beta induces livers of Xenopus laevis (South African clawed toad) to synthesize and secrete into the serum large quantities of the egg-yolk-protein precursor, vitellogenin. The peak of this response occurs 9-16 days after hormone treatment [Dolphin, Ansari, Lazier, Munday & Akhtar (1971) Biochem. J.124, 751-758]. It is now shown that 6 days after hormone treatment a 120-160-fold stimulation of the synthesis of cholesterol and fatty acid compared with control values occurred. 2. A cell-free system, derived from Xenopus liver, which synthesizes squalene and fatty acid is described. By using this system, several hundredfold stimulation of incorporation of [(14)C]acetate into squalene was recorded 6 days after the administration of oestradiol-17beta, compared with a 3-4-fold stimulation of incorporation of [(3)H]mevalonate compared with control values. It is argued that oestradiol-17beta must affect enzyme(s) catalysing step(s) between acetate and mevalonate in the biosynthetic pathway to cholesterol. 3. In incubation of liver slices in vitro, most of the lipid and cholesterol synthesized in response to the steroid hormone was associated with those subcellular fractions that contained membranes. Moreover, pulse-labelling experiments in vivo showed that 70% of this lipid and cholesterol was retained in the liver. The remainder appeared in the serum, where it was equally distributed between vitellogenin and vitellogenin-free serum. 4. G.l.c. analyses of the cholesterol content of liver microsomal fractions of Xenopus laevis indicated that the cholesterol content was at least 50% higher in microsomal fractions obtained from livers that had been exposed to oestradiol-17beta. Meanwhile, g.l.c. analysis of the lipid moiety of secreted vitellogenin showed that up to 35% of its lipid was cholesterol.


Comparative biochemistry of beta-oxidation. An investigation into the abilities of isolated heart mitochondria of various animal species to oxidize long-chain fatty acids, including the C22:1 monoenes.

Rates of acylcarnitine oxidation by isolated heart mitochondria from various animal species were measured polarographically, and by using a spectrophotometric assay [see Osmundsen & Bremer (1977) Biochem. J. 164, 621-633]. Polarographic measurements do not give a correct guide to abilities to beta-oxidize very-long-chain acylcarnitines, in particular C22:1 fatty acylcarnitines. 2. No significant species differences were detected in the abilities to beta-oxidize various C22:1 fatty acylcarnitines. Significant species differences were, however, detected when rates of beta-oxidation were correlated with rates of respiration brought about by very-long-chain acylcarnitines. We concluded that some aspects of oxidative metabolism (possibly the oxidation of tricarboxylic acid-cycle intermediates) are inhibited by very-long-chain fatty acids in some species (e.g. the rat and the cat but not in others (e.g. the pig and the rabbit). 3. It is proposed that the pattern of variation of rates of oxidation of various acylcarnitines (as measured spectrophotometrically) of various chain lengths can be used as a guide to the chain-length specificities of the acyl-CoA dehydrogenases of beta-oxidation (EC 1.3.99.3).


Glutamine metabolism in the kidney during induction of, and recovery from, metabolic acidosis in the rat.

Experiments were carried out on rats to evaluate the possible regulatory roles of renal glutaminase activity, mitochondrial permeability to glutamine, phosphoenolpyruvate carboxykinase activity and systemic acid-base changes in the control of renal ammonia (NH(3) plus NH(4) (+)) production. Acidosis was induced by drinking NH(4)Cl solution ad libitum. A pronounced metabolic acidosis without respiratory compensation [pH=7.25; HCO(3) (-)=16.9mequiv./litre; pCO(2)=40.7mmHg (5.41kPa)] was evident for the first 2 days, but thereafter acid-base status returned towards normal. This improvement in acid-base status was accompanied by the attainment of maximal rates of ammonia excretion (onset phase) after about 2 days. A steady rate of ammonia excretion was then maintained (plateau phase) until the rats were supplied with tap water in place of the NH(4)Cl solution, whereupon pCO(2) and HCO(3) (-) became elevated [55.4mmHg (7.37kPa) and 35.5mequiv./litre] and renal ammonia excretion returned to control values within 1 day (recovery phase). Renal arteriovenous differences for glutamine always paralleled rates of ammonia excretion. Phosphate-dependent glutaminase and phosphoenolpyruvate carboxykinase activities and the rate of glutamine metabolism (NH(3) production and O(2) consumption) by isolated kidney mitochondria all increased during the onset phase. The increases in glutaminase and in mitochondrial metabolism continued into the plateau phase, whereas the increase in the carboxykinase reached a plateau at the same time as did ammonia excretion. During the recovery phase a rapid decrease in carboxykinase activity accompanied the decrease in ammonia excretion, whereas glutaminase and mitochondrial glutamine metabolism in vitro remained elevated. The metabolism of glutamine by kidney-cortex slices (ammonia, glutamate and glucose production) paralleled the metabolism of glutamine in vivo during recovery, i.e. it returned to control values. The results indicate that the adaptations in mitochondrial glutamine metabolism must be regulated by extra-mitochondrial factors, since glutamine metabolism in vivo and in slices returns to control values during recovery, whereas the mitochondrial metabolism of glutamine remains elevated.


Mechanism of dealkylation of clionasterol in the insect Tenebrio molitor.

1. 25-3H- and 26-14C-labelled (24S)-24-ethylcholest-5-en-3beta-ol (clionasterol) were synthesized from (24S)-24-ethylcholesta-5,25-dien-3beta-ol. 2. These labelled substrates were mixed and administered, together with the hypocholesterolemic agent, triparanol citrate, to Tenebrio molitor larvae. 3. The 3H label from the clionasterol substrate was retained in both the desmosterol and the cholesterol isolated from the larvae. 4. Location of this 3H label in the desmosterol showed that dealkylation of the clionasterol involved 3H migration from C-25 to C-24. A possible mechanism for dealkylation is presented.


Metabolism of the surfactants sodium undecyltriethoxy sulphate and sodium dodecyltriethoxy sulphate in the rat.

The metabolic fates of the synthetic surfactants, sodium [1-(14)C]undecyltriethoxy sulphate and sodium [1-(14)C]dodecyltriethoxy sulphate were studied in the rat. Both compounds were extensively metabolized regardless of the route of administration, oral, intraperitoneal or intravenous. Short-chain radioactive products were eliminated in the urine: the major metabolite of the dodecyl homologue in the urine was identified as (-)O(2)C(14)CH(2)- (OC(2)H(4))(3)OSO(3) (-) by n.m.r. and g.l.c.-mass spectrometry, whereas the major metabolite of the undecyl homologue in the urine was tentatively identified as (-)O(2)CCH(2) (14)CH(2)- (OC(2)H(4))(3)OSO(3) (-). In contrast with experiments with the dodecyl derivative, when [1-(14)C]undecyltriethoxy sulphate was administered to rats, appreciable amounts of radioactivity were recovered as (14)CO(2) in expired air. Whole-body radioautography implicated the liver as the major site of metabolism of both surfactants. The nature of the metabolic products establishes that both compounds are degraded by omega,beta-oxidation. Cleavage of the ether linkage proximal to the sulphate moiety may account for the small amounts of (14)CO(2) recovered in expired air after the administration of [1-(14)C]dodecyltriethoxy sulphate. It is suggested the substantial amounts of (14)CO(2) recovered after [1-(14)C]-undecyltriethoxy sulphate administration originate from (-)O(2) (14)C(OC(2)H(4))(3) OSO(3) (-), an unstable product of omega,beta-oxidation. An n.m.r. spectrum of the metabolite identified as 2-(triethoxy sulphate)acetic acid and a mass spectrum of the trimethylsilyl derivative of the parent alcohol of that metabolite have been deposited as Supplementary Publication SUP50086 (5 pages) at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1978) 169, 5.


Evidence for the existence of different pools of microsomal phosphatidylinositol by the use of phosphatidylinositol-exchange protein.

1. The phosphatidylinositol-exchange protein from bovine brain was used to determine to what extent phosphatidylinositol in rat liver microsomal membranes is available for transfer. 2. The microsomal membranes used in the transfer reaction contained either phosphatidyl[2-(3)H]inositol or (32)P-labelled phospholipid. The (32)P-labelled microsomal membranes were isolated from rat liver after an intraperitoneal injection of [(32)P]P(i). The (3)H-labelled microsomal membranes and rough- and smooth-endoplasmic-reticulum membranes were prepared in vitro by the incorporation of myo-[2-(3)H]inositol into phosphatidylinositol by either exchange in the presence of Mn(2+) or biosynthesis de novo in the presence of CTP and Mg(2+). 3. Tryptic or chymotryptic treatment of the microsomes impaired the biosynthesis de novo of phosphatidylinositol. It was therefore concluded that the biosynthesis of phosphatidylinositol and/or its immediate precursor CDP-diacylglycerol takes place on the cytoplasmic surface of the microsomal membrane. 4. Under the conditions of incubation 42% of the microsomal phosphatidyl[2-(3)H]inositol was transferred with an estimated half-life of 5min; 38% was transferred with an estimated half-life of about 1h; the remaining 20% was not transferable. Identical results were obtained irrespective of the method of myo-[2-(3)H]inositol incorporation. 5. Both measurement of phosphatidylinositol phosphorus in the microsomes after transfer and the transfer of microsomal [(32)P]phosphatidylinositol indicate that phosphatidyl[2-(3)H]-inositol formed by exchange or biosynthesis de novo was homogeneously distributed throughout the microsomal phosphatidylinositol. 6. We present evidence that the slowly transferable pool of phosphatidylinositol does not represent the luminal side of the microsomal membrane; hence we suggest that this phosphatidylinositol is bound to membrane proteins.


Utilization by the isolated perfused rat liver of N-acetyl-D-[1-14C]galactosamine and N-[3H]acetyl-D-galactosamine for the biosynthesis of glycoproteins.

The isolated perfused rat liver system has been used to monitor the utilization of N-[3H]acetyl-D-galactosamine and N-acetyl-D-[1-14C]galactosamine for the biosynthesis of radiolabelled glycoproteins, which are subsequently secreted into the plasma. Both radiolabels appear in a number of different glycoproteins, predominantly as sialic acid and N-acetylglucosamine. The ratio of labelled sialic acid to labelled N-acetylglucosamine varies for different glycoproteins, but the bulk of N-acetyl-D-galactosamine is incorporated without deacetylation.


Kinetics of polyamine synthesis and turnover in mouse fibroblasts.

Kinetics of polyamine synthesis and degradation were studied in mouse fibroblasts growing in suspension culture. The approach was to prelabel cells with radioactive polyamines and to observe the rate of loss of radioactivity and the rate of decrease in specific activity of these compounds in cells. Radioactive putrescine declined with a half-life of 1.5-2h, whether derived directly from exogenous putrescine or indirectly from ornithine. Much of this turnover was due to excretion, the kinetics of which suggested that a steady-state was being established between putrescine inside and outside the cells. Within 5h of medium change, cells growing at a density of 5x10(5)cells/ml had supplied putrescine to the medium to a concentration of about 1mum. When cells were prelabelled with either putrescine or spermidine, radioactivity in cell spermidine declined with a half-life of 60h. This rate of turnover is sufficient to provide all the spermine required by the cell. Spermine synthesis was the only observed reaction of spermidine, although some excretion into the growth medium was detected. Spermine was not degraded at a detectable rate as long as cells were growing exponentially; in stationary phase, degradation to spermidine, which was excreted, became significant. The half-lives of the specific activities of spermine, spermidine and putrescine were 24, 15 and 1.5h respectively. From these values, the rate of synthesis of each was calculated. Spermidine was synthesized at 6.8 times the rate of spermine, and putrescine was synthesized at 0.46nmol/10(6)cells per h, twice the rate of spermidine. The significance of these kinetic parameters is discussed.


Analytical subcellular fractionation of needle-biopsy specimens from human liver.

1. Fragments (2-20 mg wet wt.) of closed needle-biopsy specimens from human liver were disrupted in iso-osmotic sucrose and subjected to low-speed centrifugation. The supernatant was layered on a linear sucrose-density gradient in the Beaufay small-volume automatic zonal rotor. The following organelles, with equilibrium densities (g/ml) and principal marker enzyme shown in parentheses, were resolved: plasma membrane (1.12-1.14; 5'-nucleotidase); lysosomes (1.15-1.20; N-acetyl-beta-glucosaminidase); mitochondria (1.20; malate dehydrogenase); endoplasmic reticulum (1.17-1.21; neutral alpha-glucosidase); peroxisomes (1.22-1.24; catalase). 2. The distribution of particulate alkaline phosphatase and, to a lesser degree, leucine 2-naphthylamidase followed that of 5'-nucleotidase. gamma-Glutamyltransferase was associated with membranes of significantly higher equilibrium density than was 5'-nucleotidase. 3. The distribution of 12 acid hydrolases was determined in the density-gradient fractions. beta-Glucosidase had a predominantly cytosolic localization, but the other enzymes showed a broad distribution of activity throughout the gradient. Evidence was presented for two populations of lysosomes with equilibrium densities of 1.15 and 1.20 g/ml, but containing differing amounts of each enzyme. Further evidence of lysosomal heterogeneity was demonstrated by studying the distribution of isoenzymes of hexosaminidase and of acid phosphatase. 4. The resolving power of the centrifugation procedure can be further enhanced with membrane perturbants. Digitonin (0.12 mM) selectively disrupted lysosomes, markedly increased the equilibrium density of plasma-membrane components and lowered the density of the endoplasmic reticulum, but did not affect the mitochondria or peroxisomes. Pyrophosphate (15 mM) selectively lowered the equilibrium density of the endoplasmic reticulum.


Developmental changes in the activity of lipoprotein lipase (clearing-factor lipase) in rat lung, cardiac muscle, skeletal muscle and brown adipose tissue.

The lipoprotein lipase activity of the lung, skeletal muscle, heart muscle and brown adipose tissue of the rat was studied during the period from late foetal to adult life. The enzyme activity in all four tissues emerged substantially during the first 24th after birth. Subsequently, heart and lung enzyme activity remained relatively constant per unit wet weight of tissue. The enzyme activity present in brown adipose tissue and skeletal muscle was elevated per unit weight of tissue during suckling compared with other periods of life. Delivery of near-term foetuses stimulated the emergence of enzyme activity in all four tissues with the same time course as that evoked by normal delivery. The significance of the presence of the enzyme in the tissues and the activity changes which occurred during development are discussed in relation to possible mechanisms of control.


Induction of rat kidney gluconeogenesis during acute liver intoxication by carbon tetrachloride.

1. Glucose production from L-lactate was completely inhibited 24h after carbon tetrachloride treatment in liver from 48h-starved rats. The activities of phosphoenolpyruvate carboxykinase, fructose diphosphatase and glucose 6-phosphatase were decreased by this treatment in fed and starved rats, whereas lactate dehydrogenase activity was only decreased in fed animals. 2. The production of glucose by renal cortical slices from fed rats previously treated with carbon tetrachloride was enhanced when L-lactate, pyruvate and glutamine but not fructose were used as glucose precursors. Renal phosphoenolpyruvate carboxykinase activity was increased in this condition. 3. This increase was counteracted by cycloheximide or actinomycin D, suggesting that the effect was due to the synthesis de novo of the enzyme. 4. The pattern of hepatic gluconeogenic metabolites in treated animals was characterized by an increase in lactate, pyruvate, malate and citrate as well as a decrease in glucose 6-phosphate, suggesting an impairment of liver gluconeogenesis in vivo. 5. In contrast, the profile of renal metabolites suggested that gluconeogenesis was operative in the treated rats, as indicated by the marked increase in the content of phosphoenolpyruvate, 2-phosphoglycerate, 3-phosphoglycerate and glucose 6-phosphate. 6. It is postulated that renal gluconeogenesis could contribute to the maintenance of glycaemia in carbon tetrachloride-treated rats.


Effects of amino acids, ammonia and leupeptin on protein synthesis and degradation in isolated rat hepatocytes.

Protein synthesis in isolated rat hepatocytes, as measured by the incorporation of [14C]-valine at constant specific radioactivity, proceeded at a rate of 0.3-0.5%/h in an unsupplemented medium, i.e. only about one-tenth the rate of protein degradation (4%/h). Leupeptin, which inhibits lysosomal protein degradation (previously found to be 75% of the total degradation in hepatocytes), had no effect on protein synthesis, showing that endogenous protein degradation supplied amino acids in excess of the substrate requirements for protein synthesis. The inhibition of protein synthesis by NH4Cl (another inhibitor of lysosomal protein degradation) as well as the stimulation by a physiological amino acid mixture must therefore represent indirect effects, either on general energy metabolism, or on unknown regulatory processes.


Chromatin structure through the cell cycle. Studies with regeneration rat liver.

Liver nuclei were prepared through the first cell cycle in partially hepatectomized young rats showing 30% parenchymal cell synchrony. To determine if nucleosome structure altered during this period, liver nuclei from sham-operated rats were compared with nuclei isolated at various times after partial hepatectomy. These nuclei were exposed to deoxyribonuclease I (EC 3.1.4.5), deoxyribonuclease II (EC 3.1.4.6) or micrococcal nuclease (EC 3.1.4.7) and the nucleosome-associated DNA length was ascertained. In no case was a difference in the DNA lengths associated with nucleosome structure observed. Differences were observed with regard to the histones and their relative association with nuclear material. When nuclei from normal rat livers were incubated in hypo-osmolar medium 9% of histone 1 and 4% of the other histones were released. These released histones, unlike those remaining bound to the nuclei, showed high [3H]adenosine and [3H]acetate uptakes in vivo. [32P]P1 uptake was also much greater into released than bound histones 1 and 3, but was not different for histone2A. At 3.5-4.5 h after partial hepatectomy, the release of histone 1 was trebled and that of histone 4 doubled. By 13.5 h, when phosphorylation of the bound forms of histones 2A and especially 1 was increased, no further changes in histone release in hypo-osmolar medium were found. The released histones from partially hepatectomized livers had indistinguishable [3H]adenosine uptakes from controls. The roles are discussed of phosphorylation and ADP-ribosylation in labilizing histone binding.


Modeccin, the toxin of Adenia digitata. Purification, toxicity and inhibition of protein synthesis in vitro.

1. Modeccin, the toxin of Adenia digitata (Modecca digitata), was purified from the roots of this plant by affinity chromatography on Sepharose 4B. 2. This toxin is a protein with mol.wt. 57000, which on treatment with 2-mercaptoethanol can be dissociated into two subunits of mol.wts. 25000 and 32000. 3. Modeccin inhibits protein synthesis in vitro in a lysate of rabbit reticulocytes and in Ehrlich ascites cells; the effect on cells is decreased in the presence of lactose. 4. Dissociation of modeccin into subunits decreases the toxicity to animals and the inhibition of protein synthesis in cells, but enhances the inhibition of protein synthesis in the lysate system.


The purification and some properties of rusticyanin, a blue copper protein involved in iron(II) oxidation from Thiobacillus ferro-oxidans.

The 'blue' copper-containing protein rusticyanin was purified to homogeneity from cells of the chemolithotrophic bacterium Thiobacillus ferro-oxidans by (NH4)SO4 fractionation and ion-exchange chromatography. The protein, which is stable at low pH, consists of a single polypeptide chain of mol. wt. 16500 and possesses 0.79 (+/- 0.28)g-atom of Cu/mol. The protein, which does not contain arginine residues, has optical absorbance maxima at 287, 450, 597 and 750 nm and is generally similar to azurin. The isolated protein is reduced directly by Fe2+ with a 1:1 stoicheiometry to Cu. On reduction by Fe2+ the absorption peaks at 450, 597 and 750 nm are abolished, with the appearance of a new absorption band at 320 nm. The results obtained are consistent with rusticyanin being the initial acceptor of electrons from Fe2+ during respiratory iron oxidation.


Incorporation of ribonucleic acid in vitro into dense ribonucleoprotein-like materials by isolated rat liver nuclei.

A cell-free system of isolated rat liver nuclei is described which permits an active incorporation of newly synthesized RNA into 'dense' ribonucleoprotein-like materials. The reaction is stimulated with increasing amounts of cytosol protein isolated from rat liver. This indicates that cytosol protein plays an important role in the formation of such material.


The brush border of rabbit kidney, a cellular compartment free of glycolytic enzymes.

Activities of four enzymes of the glycolytic pathway, hexokinase, glyceraldehyde 3-phosphate dehydrogenase, pyruvate kinase and lactate dehydrogenase, were determined in a vesicular brush-border preparation from rabbit kidneys. The specific activities of the enzymes were decreased several-hundredfold in the brush-border preparation compared with a kidney homogenate, but the enzymes were not totally absent. Density-gradient centrifugation of the brush-border preparation yielded brush border of even higher purity and also a characteristic pattern of distribution for each of the contaminating intracellular membranes. The presence of hexokinase in the brush-border preparation could be traced to contaminating mitochondria, and that of glyceraldehyde 3-phosphate dehydrogenase, pyruvate kinase and lactate dehydrogenase to contaminating vesicles derived from the endoplasmic reticulum. The brush-border vesicles contained some ATP. An intravesicular concentration of 0.1mm was estimated, indicating that the vesicles had retained at least a part of their original content. Experiments in which fluorescein isothiocyanate-dextran (mol.wt. 20000) was present during cell lysis revealed that much, but not all, of the brush-border contents had been exchanged with the medium. The complete absence of glycolytic enzymes from brush-border vesicles, which had retained part of their original content, indicates that the brush border does not contain glycolytic enzymes in vivo and can be thought of as a compartment of its own, somehow separated from the cytoplasm.


Adenine nucleotides in foetal rat liver cells. Compartmentation and variation with age.

The digitonin method for the separation of cytosolic and mitochondrial fractions was applied to liver cells isolated from foetal rats. The cytosolic [ATP]/[ADP] ratio approximately doubles during the last 4 days of gestation, whereas the mitochondrial ratio remains constant. In the presence of oligomycin and added glucose, the cytosolic [ATP]/[ADP] ratio does not increase with age, but is still considerably higher than the mitochondrial ratio. Without added glucose, and when the glycogen content of foetal liver is still very low (more than 3 days before birth), the cytosolic [ATP]/[ADP] ratio in the presence of oligomycin becomes very low and equal to the mitochondrial ratio. It is concluded that the increasein the cytosolic [ATP]/[ADP] ratio during the last 4 days of gestation is solely due to enhanced mitochondrial activity in this period. Atractyloside and bongkrekic acid do not influence the O2 consumption, nor the [ATP]/[ADP] ratios in either compartment of foetal liver cells. Respiration of isolated foetal mitochondria, however, is strongly inhibited by both compounds. The implications of these findings are discussed.


The effect of dietary carbohydrate and fat on the activities of some enzymes responsible for glycerolipid synthesis in rat liver.

1. Male rats were fed for 14 days on diets containing (by wt.) 53% of starch, or on diets in which 20% of the starch was replaced by sucrose, corn oil or lard. 2. The hepatic activities of the microsomal glycerol phosphate acyltransferase, dihydroxyacetone phosphate acyltransferase, phosphatidate cytidylyltransferase, diacylglycerol acyltransferase and choline phosphotransferase, and of the soluble phosphatidate phosphohydrolase, were measured. 3. The soluble phosphatidate phosphohydrolase activity was higher in those rats fed on lard than in those fed on the starch diet. Choline phosphotransferase activity was higher in the rats fed on corn oil than in those fed on the starch diet. 4. The rate of hepatic glycerolipid synthesis was measured in vivo 1 min after injection of [1,3-3H]glycerol and [1-14C]palmitate into the portal veins. 5. The relative rate of phosphatidylcholine synthesis in vivo was increased after feeding with corn oil and the higher specific activity of choline phosphotransferase may contribute to this result. The equivalent rate of triacylglycerol synthesis was increased by feeding with lard rather than corn oil, and the increased activity of phosphatidate phosphohydrolase may partly explain this. The latter changes probably contribute to the increased concentration of triacylglycerol which other authors have observed in the livers and sera of animals fed on saturated and monounsaturated fats.


Effects of testosterone on messenger ribonucleic acid and protein synthesis in rat seminal vesicle.

In a previous report [Higgins et al. (1976) Biochem. J.158, 271-282] we described the effects of alterations in androgen status on the synthesis of two basic secretory proteins of the rat seminal vesicle. In the present paper we examine the effects of testosterone on the activity of mRNA in the seminal vesicle. Total cellular poly(A)-rich RNA was isolated and translated in a cell-free system prepared from wheat germ. Translation products were separated on denaturing polyacrylamide gels and the protein bands corresponding to the two basic secretory proteins were identified immunologically. Incorporation of radioactive methionine into these bands was taken as a measure of the individual mRNA activities. Total mRNA activity was estimated by radioactivity in total acid-precipitable material. The results show that 1 to 2 weeks after castration the activities of mRNA molecules for the basic secretory proteins were decreased 10-20-fold on a tissue basis. Testosterone given in vivo rapidly and substantially restores mRNA activity to normal. Since these changes correlate closely with variations in the rates of synthesis of the secretory proteins in whole cells it suggests that androgenic steroids control protein synthesis chiefly via mRNA availability. In this respect their action resembles those of other steroid hormones acting in other systems. However, these effects of testosterone on the mRNA molecules for the major secretory proteins could not be distinguished from those on total mRNA. Thus the proportion of the total mRNA population accounted for by the two specific mRNA molecules showed less than a 2-fold variation with androgen status. Similarly the two secretory proteins always accounted for 25-33% of general protein synthesis. This is in sharp contrast with the markedly differential effects of other steroid hormones controlling synthesis of major proteins in other well-studied systems. We interpret our results as indicating that testosterone regulates the mRNA population of the seminal vesicle as a whole.


Transverse organization of phospholipids across the bilayer of plasma-membrane subfractions of rat hepatocytes.

Phospholipase C treatment of vesicular subfractions of plasma membranes derived from the three functional domains of rat liver indicated that there is an asymmetric distribution of phospholipids across the bilayer of these membranes. The bile-canalicular and sinusoidal membranes were similar and different from the contiguous membrane.


Derivation of an electron-transport rate equation, energy-conservation equations and a luminescence-flux equation of algal and plant photosynthesis.

On the assumption that the photosynthetic electron-transport rate is sometimes limited on the water-splitting side of Q (the oxidized primary electron acceptor), and that Q reduction, as well as primary charge recombination, is not kinetically a monomolecular process, a rate equation, a luminescence-flux equation and several versions of energy-conservation equations are derived. The energy-conservation equations explain most, if not all, observed relationships between rate and fluorescence. In particular, by assuming that the limiting site on the water-splitting side of Q is uncoupler-sensitive, these equations explain the uncoupler-induced simultaneous stimulations of rate and fluorescence as well as inhibition of luminescence without additional assumption ad hoc for each individual phenomenon. A newly introduced parameter central to the derivation of these equations is the specific affinity between two electron carriers.


The metabolic fate of (2-14C)folic acid and a mixture of (2-14C)- and (3',5',9-3h)-folic acid in the rat.

The metabolism of [2-14C]folic acid over 13 days and a mixture of [2-14C]- and [3',5',9-3h]-folic acid in rats over a 6-day period is described. Both 14C and 3H are excreted in urine over the 6-day period, but 3H and 14C are only detectable in faeces for 2 days. A breakdown product of folic acid labelled with 3H only was found in some urine samples, but no metabolite corresponding to the part of the molecule containing 14C was detected. These experiments show that in the whole animal a substantial portion of orally administered folic acid undergoes scission shortly after administration [Blair Biochem. J. (1957) 68, 385-387] and that the retained folates are a shortage form for folate monoglutamates.


The role of lipid components of the diet in the regulation of the fatty acid composition of the rat liver endoplasmic reticulum and lipid peroxidation.

The fatty acid compositions of the lipids and the lipid peroxide concentrations and rates of lipid peroxidation were determined in suspensions of liver endoplasmic reticulum isolated from rats fed on synthetic diets in which the fatty acid composition had been varied but the remaining constituents (protein, carbohydrate, vitamins and minerals) kept constant. Stock diet and synthetic diets containing no fat, 10% corn oil, herring oil, coconut oil or lard were used. The fatty acid composition of the liver endoplasmic reticulum lipid was markedly dependent on the fatty acid composition of the dietary lipid. Feeding a herring-oil diet caused incorporation of 8.7% eicosapentaenoic acid (C(20:5)) and 17% docosahexaenoic acid (C(22:6)), but only 5.1% linoleic acid (C(18:2)) and 6.4% arachidonic acid (C(20:4)), feeding a corn-oil diet caused incorporation of 25.1% C(18:2), 17.8% C(20:4) and 2.5% C(22:6) fatty acids, and feeding a lard diet caused incorporation of 10.3% C(18:2), 13.5% C(20:4) and 4.3% C(22:6) fatty acids into the liver endoplasmic-reticulum lipids. Phenobarbitone injection (100mg/kg) decreased the incorporation of C(20:4) and C(22:6) fatty acids into the liver endoplasmic reticulum of rats fed on a lard, corn-oil or herring-oil diet. Microsomal lipid peroxide concentrations and rates of peroxidation in the presence of ascorbate depended on the nature and quantity of the polyunsaturated fatty acids in the diet. The lipid peroxide content was 1.82+/-0.30nmol of malonaldehyde/mg of protein and the rate of peroxidation was 0.60+/-0.08nmol of malonaldehyde/min per mg of protein after feeding a fat-free diet, and the values were increased to 20.80nmol of malonaldehyde/mg of protein and 3.73nmol of malonaldehyde/min per mg of protein after feeding a 10% herring-oil diet in which polyunsaturated fatty acids formed 24% of the total fatty acids. Addition of alpha-tocopherol to the diets (120mg/kg of diet) caused a very large decrease in the lipid peroxide concentration and rate of lipid peroxidation in the endoplasmic reticulum, but addition of the synthetic anti-oxidant 2,6-di-t-butyl-4-methylphenol to the diet (100mg/kg of diet) was ineffective. Treatment of the animals with phenobarbitone (1mg/ml of drinking water) caused a sharp fall in the rate of lipid peroxidation. It is concluded that the polyunsaturated fatty acid composition of the diet regulates the fatty acid composition of the liver endoplasmic reticulum, and this in turn is an important factor controlling the rate and extent of lipid peroxidation in vitro and possibly in vivo.


The influence of passive stretch on the growth and protein turnover of the denervated extensor digitorum longus muscle.

At 7 days after cutting the sciatic nerve, the extensor digitorum longus muscle was smaller and contained less protein than its innervated control. Correlating with these changes was the finding of elevated rates of protein degradation (measured in vitro) in the denervated tissue. However, at this time, rates of protein synthesis (measured in vitro) and nucleic acid concentrations were also higher in the denervated tissue, changes more usually associated with an active muscle rather than a disused one. These anabolic trends have, at least in part, been explained by the possible greater exposure of the denervated extensor digitorum longus to passive stretch. When immobilized under a maintained influence of stretch the denervated muscle grew to a greater extent. Although this stretch-induced growth appeared to occur predominantly through a stimulation of protein synthesis, it was opposed by smaller increases in degradative rates. Nucleic acids increased at a similar rate to the increase in muscle mass when a continuous influence of stretch was imposed on the denervated tissue. In contrast, immobilization of the denervated extensor digitorum longus in a shortened unstretched state reversed most of the stretch-induced changes; that is, the muscle became even smaller, with protein synthesis decreasing to a greater extent than breakdown after the removal of passive stretch. The present investigation suggests that stretch will promote protein synthesis and hence growth of the extensor digitorum longus even in the absence of an intact nerve supply. However, some factor(s), in addition to passive stretch, must contribute to the anabolic trends in this denervated muscle.


The biochemical basis for the conjugation of bile acids with either glycine or taurine.

All animals, except for the placental mammals, conjugate their bile acids exclusively with taurine. However, in certain of the placental mammals, glycine conjugates are also found. The basis for the appearance of glycine conjugation among the placental mammals was investigated. The reaction of choloyl-CoA with glycine and taurine, as catalysed by the soluble fraction from guinea-pig liver, had a high affinity for taurine and a poor affinity for glycine. The predominant synthesis of glycine conjugates in the guinea pig can be related to the fact that guinea-pig liver contains an unusually low concentration of taurine and a high concentration of glycine. Rabbits make exclusively glycine conjugates and their livers also contain low concentrations of taurine. However, the biochemical basis for their glycine conjugation is more straightforward than in the guinea pig in that the soluble fraction from rabbit liver has a high affinity for glycine and a poor affinity for taurine. Alternative-substrate-inhibition studies with glycine and taurine in soluble fractions from guinea-pig and rabbit liver revealed that glycine and taurine were mutually inhibitory. This suggests that there is only one enzyme for glycine and taurine conjugation in these tissues. The soluble fractions from bovine liver and human liver also made both glycine and taurine conjugates and evidence is presented that suggests that there is only one enzyme in these tissues too. Even the rat, which excretes mostly taurine conjugates, could make both glycine and taurine conjugates in vitro. However, in contrast with all of the placental mammals studied, the supernatant fraction from liver of the chicken, and other non-mammals, could not make glycine conjugates even in the presence of very high concentrations of glycine.


Differential light-scattering of granal and agranal chloroplasts and their fragments.

Intact (class-A) granal and agranal maize chloroplasts and chloroplast fragments were examined for differential scattering of circularly polarized light (measured at 90 degrees) and c.d. (circular dichroism) (measured at 0 degrees) by using a modified spectropolarimeter with a large acceptance angle. Useful c.d. information was obtained by making corrections for scattered light. Chloroplast fragments exhibited a large and characteristic differential scattering of circularly polarized light recognized in the presence of granal chloroplasts. It is confirmed that agranal chloroplasts do not have the intense 682 nm c.d. peak that is assigned to the presence of grana.


Disialoganglioside GDla of rat brain subcellular particles during development.

The increase observed in the amount of the disialoganglioside GDlof the rat cerebrum during development between 21 and 81 days of age accounted for nearly 40% of the overall increase in total ganglioside in the tissue during the same period. Subcellular fractionation showed the microsomal fraction to contribute by far the most towards this increase in Cerebral ganglioside GDla. It is suggested that microsomal ganglioside GDla may serve as a marker for dendritic arborization in the rat cerebrum.