Calcium sensitivity of hybrid complexes of muscle myosin and Physarum proteins. 1. A myosin-actin hybrid complex was used to study actin-associated calcium sensitivity of a "cytoplasmic" actomyosin. The approach should be generally applicable. 2. Low salt extracts of Physarum polycephalum contain actin which remains in solution after centrifugation at 46 000 times g or at 100 000 times g for 1 h. The actin was precipitated by the addition of muscle myosin to the supernatants and detected in the hybrid complex by electron microscopy, sodium dodecyl sulfate gel analysis, super-precipitation and activation of the myosin ATPase activity. Actin was also precipitable from high speed supernatants of brain tissue or platelets. 3. The hybrid complexes from Physarum possessed 1.5-5-fold calcium dependency which could be removed by washing. Reincubation of the washed complex with concentrated wash solution resulted in high calcium sensitivity. On sodium dodecyl sulfate gels, unwashed complexes from Physarum contained high molecular weight material in addition to bands of molecular weights less than actin. The bands in the size range of 39 000 to 18 000 were primarily lost from the Physarum complex concomitantly with loss of calcium dependence. 4. When the Physarum supernatants were made 40 mM in MgCl2, precipitates were formed containing actin which possessed calcium sensitivity which was also lost on washing with low ionic strength solutions. This calcium dependency was partially reversed by the addition of desensitized rabbit actin to the precipitate before assay. 5. calcium regulation of actomyosin in Physarum is mediated primarily by factors that are bound to the actin component. The regulatory factors are soluble in low salt buffers. The molecular weights of the polypeptide chains of several of these factors are similar to those of the troponin polypeptides of striated muscle. In Physarum but not in platelet or brain a prominent polypeptide chain of approx. 55 000 molecular weight also occurs which coprecipitates with the hybrid complex and which is not easily removed.


Leucocyte myosin and its location in the cell. The intracellular location of the binding site of antibody against purified myosin prepared from equine leucocytes was investigated in neutrophils and lymphocytes by electron microscopy using peroxidase-labelled antibody method. The myosin extracted from equine leucocytes could bind skeletal muscle F-actin and the formed complex showed the biophysical and biochemical properties and electron microscopic appearance of actomyosin. On immunodiffusion, the leucocyte myosin formed a single precipitin line with its antibody prepared in rabbits. The antibody also formed single precipitin lines with myosins from lymphocytes and thrombocytes, fusing with each other. The antibody against the leucocyte myosin did not react with myosins from skeletal or arterial smooth muscle. The specificity of the antibody was further established by determination of K+-EDTA-activated ATPase activity remained in the supernate of antigen-antibody mixture. Under electron microscope, the intracellular immunoreactive products of peroxidase labelled antibody were found in cytoplasm of neutrophils and lymphocytes incubated with antibody against leucocyte myosin, but not in neutrophils or lymphocytes treated with IgG from normal rabbits.


Interaction of actin with phalloidin: polymerization and stabilization of F-actin. The cyclic peptide phalloidin, one of the toxic components of Amanita phalloides prevented the drop of viscosity of F-actin solutions after the addition of 0.6 M KI and inhibited the ATP splitting of F-actin during sonic vibration. The data concerning ATP splitting are consistent with the assumption (a) that only 1 out of every 3 actin units of the filaments needs to be combined with phalloidin in order to suppress the contribution of these 3 actins to the ATPase activity of the filament and (b) that all actin units of the filaments can combine with phalloidin with a very high affinity. -halloidin did not only stabilize the actin-actin bonds in the F-actin structure but it also increased the rate of polymerization of G-actin to F-actin. The ability of F-actin to activate myosin ATPase was not affected by phalloidin. The tropomyosin-troponin complex did not prevent the stabilizing effect of phalloidin on the F-actin structure.


Denaturation map of the circular mitochondrial genome of Neurospora crassa. A denaturation map of mitochondrial DNA from the wild type strain 5256 of Neurospora crassa was constructed by computer analysis of the contour length distribution of single- and double-stranded regions of nineteen circular and three full length linear molecules after partial denaturation. The data suggest that mitochondrial DNA in this strain is a homogeneous population of a circular molecule of molecular weight 41 - 10(6) with an asymmetric distribution of AT-rich regions, and that linear molecules derive from this genome by random breaks during isolation.


Synchronous reversible alterations in enzymatic activity (conformational fluctuations) in actomyosin and creatine kinase preparations. The phenomenon of synchronism of oscillations of actomyosin and creatine kinase activity in the whole volume of the enzyme preparations was analysed. The synchronous "conformational oscillations" were observed in concentrated gels of actomyosin and in diluted actomyosin and creatine kinase solutions (ATP-creatine N-phosphotransferase, EC 2.7.3.2). The macromolecules of proteins studied may be in two or four conformational states differing enzymatic activity. Large fluctuations become possible in a range of conditions wherein two or four different states, or conformers, are equiprobable. The synchronization of conformational changes of separate macromolecules is maintained with energy derived, for instance, from some oxidative process or dilution of the solution, the process being displayed as conformational oscillations.


Proteoglycans of mineralizing rib and epiphyseal cartilage. Rib cartilage from growing guinea pigs and epiphyseal cartilage from Beagle puppies were separated into three fractions, representing non-mineralized, low mineralized, and high mineralized, tissue, by centrifuging finely ground material in acetone/bromoform density gradients. Following extraction under dissociative conditions, the proteoglycans were fractionated by density gradient ultracentrifugation under associative and dissociative conditions. With the onset of mineralization, the cartilage lost approximately half its content of proteoglycans. The proteoglycans remaining in the calcified cartilage differed in composition and in size from those of nonmineralized tissue. With the increased mineral content of the tissues the ratios of protein to polysaccharide, of chondroitin sulfate to keratan sulfate, and of 4-sulfate to 6-sulfated chondroitin sulfate increased in the proteoglycan fraction. Furthermore, gel chromatograms indicated decreased proportions of very high molecular weight proteoglycans, in mineralized tissue.


Alterations in the composition and bacteriophage-binding properties of walls of Staphylococcus aureus H grown in continuous culture in simplified defined media. A nutritional mutant of Staphylococcus aureus H has been isolated and grown in media in which the only amino acids are arginine, cysteine, glutamic acid and proline. Walls of the bacteria grown in such media in continuous culture under potassium limitation differ in composition from walls of the bacteria grown in batch culture in rich nutrient broth in that they contain less glycine, the peptidoglycan component is less highly cross-linked and the teichoic acid component contains a reduced proportion of N-acetylglucosaminyl substituents. Walls of the potassium-limited bacteria retain the ability to bind bacteriophage 52a but are more susceptible to the action of lytic peptidases than are wall samples in which the peptidoglycan is more highly cross-linked. Teichoic acid was present in walls of the bacteria grown under phosphate limitation in the defined medium and these walls were also able to absorb bacteriophage 52a.


Comparison of plasma membranes and endoplasmic reticulum fractions obtained from whole white adipose tissue and isolated adipocytes. The influence of the mode of preparation upon some of the characteristics of white adipose tissue plasma membranes and microsomes has been reported. Plasma membrane fractions prepared from mitochondrial pellet were shown to have higher specific activities of (Mg2+ + Na+ + K+)-ATPase than plasma membranes originating in crude microsomes. Isolation of fat cells by collagenase treatment was found to result in a decrease in specific activity of the plasma membrane enzymes; in plasma membranes prepared from isolated fat cells, the specific activity values obtained for (Mg2+ + Na+ +k+)-ATPase and 5'-nucleotidase were only 42% and 6.3% respectively of those obtained in plasma membranes prepared from whole adipose tissue. Purification of whole adipose tissue crude microsomes by hypotonic treatment caused extensive solubilization of the endoplasmic reticulum marker enzymes, NADH oxidase and NADPH cytochrome c reductase. The lability of endoplasmic reticulum marker enzymes, however, was found to be greatly diminished in the preparations from isolated fat cells. The possibility that NADH oxidase and NADPH cytochrome c reductase activities found in the plasma membranes are microsomal enzymes adsorbed by the plasma membranes is discussed. The peptide patterns as well as the NADH oxidase and NADPH cytochrome c reductase activity patterns of plasma membranes and purified microsomes were compared by means of sodium dodecyl sulfate or Triton X-100 polyacrylamide gel electrophoresis.


Perinatal changes of interorgan differences in cell metabolism. Differences of metabolic qualities and capacities among organs and tissues of perinatal rabbits were investigated. A suitable alternative for the measurement of substrate utilization of single organs seems to be the total amount of certain key enzyme activities calculated for whole organs. The immediate perinatal period is the object of this study with adult animals serving as a reference. We have selected the phosphofructokinase activity to represent the upper segment of glycolytic reactions, 3-hydroxyacyl-CoA dehydrogenase as a key enzyme for beta-oxidation of fatty acids and citrate synthase activity to represent the Krebs cycle activity. In fetal, newborn, and adult rabbits we analyzed liver, kidneys, heart, lung, brain, brown and white adipose tissue, stomach and intestines, skin, a representative sample of skeletal muscles, and of bones. The organ weight distribution, total amount of protein, and DNA was determined for the same ten organs. The share each organ contributes to the total body key enzyme activity shows the importance of each metabolic capacity which is represented by the three enzymes. In perinatal adipose tissue the very high potential for energy production through utilization of Krebs cycle reactions is striking. The same tissue has a high capacity to oxidize fatty acids. The skeletal muscle represents the biggest capacity of glycolytic reactions in all age groups. After birth the metabolic profile of the whole organism shows a marked and steep increase of glycolytic capacity, whereas the capacity to oxidize fatty acids decreases slowly.


Age-dependent change in the gel-electrophoretic pattern of proteoglycans of human growth cartilage. The gel-electrophoretic pattern of dissociated proteoglycans was studied in 7 fetuses, 5 premature newborns, 4 term newborns, 5 infants and 5 children. The tibial growth cartilage was extracted with 4 M guanidinium chloride. After dialysis against 8 M urea at pH 7 the proteoglycans were obtained by ion chromatography in urea on DEAE cellulose and submitted to gel electrophoresis on polyacrylamide agarose gels. Gel electrophoresis of proteoglycans showed a different pattern in fetuses from that found in children. The change occurs in the first months of extrauterine life.


Ultrastructure of the contractile system of striated skeletal muscle and the processes of muscular contraction. II. Releasing system and mechanisms of muscular contraction. The tropomyosin-troponin system of the thin filaments acts as a regulator of the interaction of myosin with actin by a process of inhibition. This inhibition is released by Ca2+ ions. Muscle contraction occurs when Ca+2 ions are released from the sarcoplasmic reticulum into the myofibril, where they interact with the troponin of the thin filaments. Muscular contraction is due to the sliding of the two kinds of filaments past each other in the hexagonal network of the myofibril. Many hypotheses have been formulated concerning the intimate molecular mechanisms responsible for the sliding of the filaments past each other, but none of these can be considered to be entirely satisfactory.


A rapid and sensitive 125I-fibrin solid-phase fibrinolytic assay for plasmin. 125I-fibrinogen, adsorbed to polystyrene tubes at low ionic strength and treated with thrombin, serves as a substrate for a rapid, convenient, and sensitive test tube assay for plasmin and activators and inhibitors of this enzyme. 125I-labeled digestion products released from the 125I-fibrin-polystyrene matrix are readily separated and quantitated and behave, on gel permeation, in the same manner as plasmin-generated degradation products from an unlabeled conventional fibrin clot. The 125I-fibrin, in probable non-cross-linked form, is firmly bound to the polystyrene and is resistant to nonspecific release, with control (no enzyme) values equivalent to 15.2 ng +/- 1.2 (SD) fibrin (1% of the total bound 125I-fibrin). This fact permits consistent detection of lysis of 30-50 ng 125I-fibrin, which exceeds published sensitivities (1000-5000 ng) using 125I- or fluorochrome-labeled fibrin clots as substrate. The sensitivity for plasmin (0.2 mug/ml) is tenfold greater than that of the fibrin-plate method (2.0-2.5 mug/ml), while sensitivities for streptokinase and urokinase activation of plasmin are 0.02 U/ml and 0.04 CTA U/ml, respectively (sensitivity of fibrin-plate method, 0.5 U/ml for both). The method provides a reasonable analogue of the solid-phase nature of fibrin under physiologic conditions, and the ease of preparation of large batches of tubes makes the method suitable for large-scale screening of factors modulating the plasminogen-plasmin system.


The resistance of fibrinogen and soluble fibrin monomer in blood to degradation by a potent plasminogen activator derived from cadaver limbs. The effect of a cadaver-derived vascular plasminogen activator (VA) on the degradation of fibrinogen, soluble fibrin monomer, and fibrin was studied and compared with the effect of equivalent fibrinolytic potencies of streptokinase (SK), urokinase (UK), and plasmin. The proteolytic activity of the three activators and plasmin was determined by a standard fibrin plate assay and was expressed in CTA units from a UK reference curve. Fibrinogen degradation was measured by clottable protein determinations and by an electrophoretic technique sensitive to small changes in the molecular weight of fibrinogen. When VA was incubated in plasma, no degradation of fibrinogen occurred, whereas rapid fibrinolysis took place after the plasma was clotted. By contrast, equivalent potencies of SK, UK, and plasmin caused extensive fibrinogenolysis. Since the plasmin added and that formed by the three activators had equivalent fibrinolytic activity, the failure of VA to induce fibrinogen degradation was attributed to antiactivators rather than antiplasmins. VA activity in plasma was consumed by clotting, whereas the antiactivator activity remained in the serum, suggesting dissociation of the VA-antiactivator complex on the fibrin clot. Fibrinogen and its soluble derivatives resisted degradation by VA in plasma because a solid phase appeared necessary for the complex to dissociate. The findings indicated that the degradation of fibrinogen or soluble fibrin in blood as a result of plasminogen activation by VA was unlikely to occur due to a large excess of antiactivator activity. Alternative pathways for their catabolism are discussed.


Clinical trial of glibornuride in diabetes. Glibornuride is an addition to the second-generation sulphonylureas, that has been shown in clinical trials to be effective and non-toxic. Twenty-three diabetics who were poorly controlled on other oral hypoglycaemic agents and seven newly diagnosed diabetics were treated with glibornuride. The efficacy and lack of toxicity of this drug was confirmed, but there was no evidence to suggest that it is significantly more potent than other sulphonylureas. It does not seem to represent a significant therapeutic advance.


Occupational hazards in window cleaning. Accidental falls involving window cleaners treated at the Middlesex Hospital over five years are reviewed. Failure to use safety belts and the lack of suitable anchorage points were contributary factors in all 20 patients. The use of protective equipment and the provision of anchorage points should be enforced. While the doctor's duty is to treat injuries he also has the opportunity to draw attention to their prevention.


Lepromatous leprosy presenting with polyarthritis, myositis, and immune-complex glomerulonephritis. A Pakistani man aged 19 years was admitted to a rheumatological unit in the United Kingdom with acute widespread polyarthritis accompanied by night sweats and fever. Preliminary examination suggested Reiter's disease, but further investigation showed acute glomerulonephritis with uraemia. The possibility of periarteritis nodosa, and the prominence of muscle tenderness in the legs, led to biopsies of striated muscle and skin, in both of which were changes typical of lepromatous leprosy, with many Mycobacterium leprae on Ziehl-Neelsen staining. Serum showed IgG-IgM cryoglobulinaemia without antiglobulin activity, and in the recovery phase renal biopsy showed a resolving proliferative glomerulonephritis with linear IgG and IgM immunofluorescence and granular deposits of C3. Clinical signs subsided rapidly under steroid treatment and subsequent progress on anti-leprosy drugs was uneventful. The term erythema nodosum leprosum is inadequate and misleading as a title for a common and important immune-complex reaction of lepromatous leprosy, in which numerous body systems may be involved.


Comparison between laparoscopic sterilization and tubal ligation. During 1970-3, 1910 laparoscopic sterilizations and 531 tubal ligations were performed in Aberdeen. Common complications occurred with 61 (3.2%) and 99 (18.6%) of the operations respectively, but the pregnancy rate was much higher (2.2%) after laparoscopic sterilization.


Staphylococcal polysaccharide products inhibitory to Staphylococcus aureus bacteriophage. Four products derived from Staphylococcus aureus cultures were partially purified and tested for inhibitory activity against staphylococcal phage. Phage inhibition, a specific stable phenomenon, was concentration dependent. All inhibitory products contained carbohydrate and amino acids, the most active (phage 73 lysate product) having a high carbohydrate content. Galactose, glucosamine, five or six amino acids, and possibly 3-O-methylglucose and a uronic acid were found as components in all active preparations. However, the exact nature of the active material remains undetermined.


Value and limitations of vectorcardiography in cardiac diagnosis. The vectorcardiograpm improves the display of the cardiac potential by clearer demonstration of the orientation and magnitude of forces at each moment. The changes in different scalar leads are correlated. It is generally a more sensitive tool for detecting electrocardiographic abnormalities. However, it shares many of the pitfalls of the conventional 12 lead electrocardiogram. The specificity of the diagnosis, in most instances, is not improved.


The value of apexcardiography in cardiac  diagnosis. As can be seen from the aforementioned examples, apexcardiography may be helpful in cardiac diagnosis. It will, however, not yield much of value if done in isolation and with only an accompanying ECG. When it is part of a carefully chosen battery of noninvasive graphic records, the ACG may provide useful ancillary information in a variety of clinical situations. It should be recorded by a physician knowledgable concerning the patient's condition, so that the area most suitable for recording may be selected as well as an appropriate array of other graphic tracings. Care in the selection and periodic testing of equipment for adequacy of time constant and absence of air leaks in the system is necessary to guard against misleading distortions and artifacts.


Activation of the hypertrophic right ventricle in the dog. Right ventricular hypertrophy (RVH) of mild, moderate, or severe degree was produced in six dogs following systolic overload of the right ventricle by surgical banding of the pulmonary trunk. Activation of the myocardium and specialized conducting tissue of the right ventricle was studied using intramural multi-electrodes or an exploring electrode, first with the heart in situ and then with the heart in a modified Langendorff perfusion circuit. Normal epicardial and intramural activation patterns were found in RVH, and the prolonged excitation time was found to be due to the increased muscle mass. No delay in activation was found in any part of the specialized conducting tissue of the right ventricle. Late activated Purkinje fibres were found in the outflow tract of the right ventricle in the dogs with hypertrophy and in a control series of normal dogs. The present electrocardiographic criteria for complete and incomplete right bundle branch block (RBBB) are based on widening of the QRS complex and the rSR pattern. This type of change can occur also in RVH. Because it has been shown here that no delay occurs in the specialized conducting system of the right ventricle in RVH secondary to systolic loading; the application of criteria to electrocardiograms that also fulfil the criteria for RVH may be misleading. It is suggested that in such cases the diagnosis of a conduction disturbance must also depend upon other methods, and that the terms "incomplete' and "complete' RBBB should be handled with care in this context.


Glucagon-induced changes in the action potential, contraction, and Na+-K+-ATPase of cardiac muscle. The effects of glucagon in concentrations of 0.294 times 10(-6) mol/l, 1.47 times 10(-6) mol/l; 2.94 times 10(-6) mol/l, 5.8 times 10(-6) mol/l, and 1.47 times 10(-5) mol/l on the simultaneously recorded action potentials and contractions; and microsomal and sarcolemmal Na+-tk+-atpase in the myocardium of the guinea pig, rabbit, dog, and pig were investigated. Glucagon in all the concentrations produced an inhibition of the Na+-K+-ATPase associated with an increase in the contractility and shortening of the duration of action potential in dog myocardium. The increase in contraction was concentration-dependent up to a certain concentration. Inhibition of sarcolemmal ATPase was more than that of microsomal ATPase. In none of the concentrations did glucagon produce any significant changes in the Na+-K+-ATPase. In none of the concentrations did glucagon produce any significant changes in the Na+-K+-ATPase, contractility, and action potential duration in the myocardium of guinea pig, rabbit, or pig. These results suggest that glucagon-induced positive inotropic effect might be due to an increase in the Ca++ influx as a result of inhibition of membrane Na+-K+-ATPase. Shortening of the action potential duration might also be due to an increased efflux of potassium as a result of an inhibition of Na+-K+-ATPase.


A simplified parameter of LV function in children with congenital heart disease. Myocardial state in children with various congenital anomalies was assessed using pressured-derived parameters; Vmax and Vpm. A simple approach, based on the relationship between peak value of the first derivative of the left ventricular isovolumic pressure and its associated total pressure, for identifying patients with impaired myocardial function is described.


Isometric training of young rats--effects upon hind limb muscles. Histochemical, morphometric, and electron microscopic studies. The soleus, rectus femoris, and gastrocnemius muscles of young rats trained isometrically for 4 weeks were studied by light and electron microscopy.--The percentage of fast-twitch oxidative muscle fibers decreased at the cost of the fast-twitch glycolytic fibers in the rectus femoris muscle. The percentages of the slow-twitch oxidative fibers did not change significantly in any of the muscles studied. The changes in the areas of the muscle fibers were specific for the muscle and the fiber type and indicate geometrical rearrangements of the fibers in the trained muscles. The Z and M lines were broader in the soleus (containing about 85% slow-twitch oxidative fibers) than in the rectus femoris muscle (containing about 90% fast-twitch glycolytic fibers), while the sarcomere length and the pseudo-H zone were similar. The length of the myosin filaments appeared to be slightly shorter in the fast rectus femoris than in the slow soleus muscle.--The hypothesis on the temporal progress of muscle adaptation to training (Müller, 1974) was substantiated. Correlations between biochemical (Exner et al., 1973a) and histochemical parameters measuring the oxidative capacity were preserved during adaptation to training. The comparison of the histochemical results with the physiological data on similar animals (Exner et al., 1973a) suggests a complex relationship between the contraction time and the percentage of fast-twitch muscle fibers.


Retinal capillary junctions: ultrastructural tight junction artefacts induced by sodium ions and membrane reduction in streptozotocin diabetes. Retinal capillary junctions were analysed in normal and diabetic rats and in a human retina with the electron microscope. Diabetes mellitus was induced with streptozotocin. The retinae were fixed in Palade's osmium tetroxide containing sodium or calcium ions and block-stained in uranyl acetate. With Ca-fixation, no significant difference in interendothelial cleft width was detected between retinal layers or between normal and diabetic retinae. Diabetes caused a narrowing of the clefts in the Na-fixed tissue X +/- SE, n=375; Normal: 78.6 +/- 300 A; Diabetic: 57.7 +/- 2.42 A; p less than 0.001). A significant correlation was found between cleft width and the length of the tight junctions or zonulae occludentes (p less than 0.001). In the nerve fibre layer of the Na-diabetic retina, where cleft narrowing was greatest, there was an increase in length of the zonulae occludentes from 22.8 +/- 2.2% to 41.6 +/- 3.7% (p less than 0.001). Ca-fixation prevented these changes, indicating that at least some zonulae occludentes were interendothelial extraction artefacts. In the normal retina, endothelial cell membrane thickness was greater with Ca- than Na-fixation (p less than 0.001). Diabetes caused a decrease in membrane thickness of Ca-fixed tissue (p less than 0.001). The diabetic decrease in membrane thickness may explain the increased fragility and increased permeability of diabetic capillaries. Calcium binding by endothelial cell membranes is of primary importance in anticoagulation which is defective in diabetes.


Fine structure of single fibres of human skeletal muscle. Individual muscle fibres were separated from freeze-dried needle biopsies and classed as type I or type II fibres according to their myofibrillar ATP-ase. Portions of the same fibres were processed for electron microscopy and their fine structure examined. Type I fibres were found to have thicker Z-bands and more mitochondria and lipid droplets than the type II fibres.


[The effects in vivo of some sex hormones on the growth of tibias of normal and chondrodystrophic (Creeper) chick embryos]. Various sex hormones (testosterone, oestradiol, dehydroepiandrosterone), in the form of suspensions in oil, are injected into the allantois of normal and chondrodystrophic (Creeper) chick embryos of 6 days 1/2 incubation. Observations made on each of the phenotypes on the 12th day of incubation show that oil injected alone causes a significant decrease in the dry weight of the tibiae. This inhibitory effect is partially abolished when the oil is injected together with the sex hormones, testosterone being the most effective.


Echocardiographic evaluation of pericardial effusion in myxedema. Incidence and biochemical and clinical correlations. Pericardial effusion is a recognized consequences of myxedema. Its incidence is unknown, primarily because of past difficulties in establishing the diagnosis. We studied 33 hypothyroid patients by echocardiography. Ten of the 33 patients (30%) had positive echoes for pericardial effusion. Seven of these ten patients had enlarged hearts on chest X-ray. Five patients had cardiac enlargement but no echo evidence of pericardial effusion. Serum concentrations of thyroxine, 1.8+/-0.3 vs 1.5+/-0.1 mcg/dl and of thyroid stimulating hormone, 34+/-4 vs 38+/-5 muU/ml did not differ in the groups with and without pericardial effusion, respectively. However, the pericardial effusion group had significantly slower heart rates on ECG than those without pericardial effusion: 53+/-8 vs 68+/-2 beats/min, P less than 0.05. Low voltage was present in five of the ten patients with pericardial effusion and five of the 23 nonpericardial effusion patients. None of the patients with pericardial effusion developed tamponade. Seven patients with pericardial effusion were restudied after periods of thyroxine replacement therapy ranging from six months to two years. All were euthyroid and had negative echoes on follow-up, but two still showed cardiomegaly on chest X-ray (both had associated coronary artery disease). We conclude that pericardial effusion occurs frequently in patients with myxedema. Tamponade is uncommon and the effusions disappear with thyroid replacement therapy. Cardiomegaly on chest X-ray and low voltage on ECG are not reliable indicators of pericardial effusion.


Echocardiography of the "floppy" aortic valve. Report of a case. The echocardiographic features of a patient with severe aortic regurgitation due to a "floppy" aortic valve are presented. The salient abnormality observed was marked fluttering of the aortic valve cusps in diastole. Fluttering of the anterior leaflet of the mitral valve and left ventricular enlargement were noted. The diagnosis of a "floppy" aortic valve was substantiated at surgery.


Acidic glycosaminoglycans in human kidney tissue. The acidic glycosaminoglycans (AGAG) in normal human kidneys were fractionated on Dowex 1-X2 columns and analysed by electrophoretic separation in three buffers on cellulose acetate membranes and gel filtration on Sephadex G-100 columns, before and after digestion with chondroitinases and streptomyces hyaluronidase. Thin-layer chromatography was also performed to separate glucosamine from galactosamine moieties. Enzymatic digestion combined with electrophoretic characterization indicated that heparan sulfates exist as the main AGAG which accounted for two-fifths of the total AGAG. Hyaluronic acid and dermatan sulfates accounted for one-fourth and one-sixth of the total kidney AGAG, respectively. Chondroitin sulfate isomers (4-sulfate and 6-sulfate) consisted of the residual one-sixth of the total AGAG. An oversulfated chondroitin sulfate was detected in a small amount by demonstration of the unsaturated disulfated disaccharide after digestion with chondroitinase-ABC but not with chondroitinase-AC.


A rapid radioimmunoassay for determination of dehydroepiandrosterone sulphate in human plasma. A rapid radioimmunoassay for the determination of dehydroepiandrosterone sulphate (DHEA-S) in diluted human plasma is described. The radioimmunoassay is performed using an antiserum to DHEA-3-hemisuccinate-BSA. A charcoal-dextran mixture is used for separation of the bound from the free fraction. The reliability criteria of the method in terms of precision, accuracy, sensitivity and specificity have been evaluated. The mean level of DHEA-S in plasma samples from normal men (age range 24-37) is 241-5+/-72-5 mug/dl. In normal premenopausal women the highest values are in the second third of the menstrual cycle (corresponding to the ovulatory phase). In normal old men (age range 64-86) and in post-menopausal women (age range 55-88) the mean values are, respectively, 51-9+/-33-8 and 53-8+/-34-0. The DHEA-S levels found in several cases of adreno-cortical disorders are reported. In primary adrenal insufficiency and in Cushing's syndrome due to adrenal adenoma or bilateral multinodular hyperplasia low values are found; in Cushing's syndrome due to bilateral adrenal hyperplasia normal values are usually found.


Some further observations on the proteoglycans from ox nasal cartilage and pig aorta. Extracts of nasal cartilage prepared by dissociative (3 M MgCl2, 4M guanidinium chloride) procedures usually contain two species of proteoglycan macromolecules with widely differing sedimentation coefficients. This paper describes the isolation of the slower sedimenting component and compares its properties with those of the proteoglycan obtained by disaggregation of the faster of the two sedimenting materials. Additional evidence which illustrates the degradation of proteoglycans by homogenizing procedures (disruptive) is also presented; in particular, fractions obtained by gel permeation chromatography on Sepharose-4B of homogenized extracts, show sedimentation coefficients varying from about 8S to about 24S whereas dissociatively extracted materials treated similarly, give only two types of material with sedimentation coefficients of about 11-12S and 44-47S respectively. Because of the differences between the proteoglycans isolated from cartilage by disruptive and dissociative procedures, the protein-bound chondroitin sulfates obtained from pig aorta by both procedures have been examined; these two preparations are similar with respect to the methods used for characterization. In particular, both materials sediment as single boundaries in the ultracentrifuge with sedimentation coefficients (So20,w) of 5.4S (from homogenized extract) and 5.3S (from dissociative extract); further, their amino acid and carbohydrate compositions are almost identical.


Tissue sampling as a potential source of error in experimental studies of cartilage. Although it might seem trite to point out that tissue sampling is a potential source of experimental error, this survey disclosed that even experienced investigators in fact often work with cartilage that is contaminated by non-cartilaginous tissue of which they were unaware. Twenty-two specimens ranging from chick embryo sternum to bovine nasal septum were studied by serial sectioning. Eighteen of the 22 contained extraneous tissue comprising from 3 to 50% of the cross-sectional area. The impact of the contamination depends on the use being made of the material and probably is greatest in cell culture studies because chondrocytes and fibroblasts have large differences in population doubling time. Several approaches for minimizing the error are suggested by the findings. Histological examination of specimen material is thus a desirable quality control procedure in the design and interpretation of experiments on cartilage as well as other tissues.


The uronidic linkages in heparitin sulfate. Beta-glucuronidase preparations obtained from various sources were shown to liberate free D-glucuronic acid from heparitin sulfate oligosaccharides. Some free L-iduronic acid could also be detected in the hydrolyzates indicating the presence of an iduronidase in the preparations. The data presented indicate that heparitin sulfate contains a substantial amount of beta-D-glucuronidic and some alpha-L-iduronidic linkages.


Biosynthesis of glycosaminoglycans: uridine diphosphate glucose 4'-epimerase from cornea and epiphysial-plate cartilage. UDP-glucose 4'-epimerase (EC 5.1.3.2.) was extracted from newborn-pig epiphysial-plate cartilage and whole bovine cornea. The formation of radioactive UDP-galactose from UDP[U-14C]glucose was demonstrated by radioautography after separation of the sugar nucleotides by paper chromatography or t.l.c. The pH optimum and the Km values for UDP-glucose, UDP-galactose and NAD+ were determined in both tissues. UDP-galactose and UDP-glucuronic acid formation after incubation with different UDP-glucose concentrations was followed; the same experiment was carried out using different UDP-galactose concentrations and following the formation of UDP-glucose and UDP-glucuronic acid. At equilibrium, the ratio UDP-glucose/UDP-galactose reaches a value of about 3.5. The results obtained seem to indicate that UDP-glucose 4'-epimerase activity is strongly dependent on that of UDP-glucose dehydrogenase. The physiological meaning of UDP-glucose 4'-epimerase in glycosaminoglycan biosynthesis in the two tissues under study is discussed on the basis of the Km values of UDP-glucose 4'-epimerase and UDP-glucose dehydrogenase and on the basis of the rate of UDP-glucose and UDP-galactose utilization.


Depolymerization and de-N-acetylation of glycosaminoglycuronans by the action of alkali in the presence of sodium borohydride. Depolymerization of Ch-4-S, Ch-6-S, HA and HP was observed by gel filtration after treatment with 2-10 N NaOH or Ba(OH)2 in the presence of NaBH4 at 80-83 degrees for 1-5 hr. Depolymerized products were isolated after these treatments. 4-Deoxy-alpha-L-threo-4-enohexopyranosyluronic acid residue as the nonreducing end group was demonstrated by the H-1 and H-4 proton signals in the NMR spectra and by the absorption at 225-230 nm. L-Gulonic acid, which was produced from D-glucuronic acid by the action of NaBH4, was detected as the reducing end group in the depolymerized products of Ch-6-S, but no hexosaminitol was detected. De-N-acetylation to the extent of 34-50% occurred after treatment with 2.5 N NaOH in the presence of NaBH4, and almost complete de-N-acetylation accompanied by considerable destruction was observed with Ch-4-S and Ch-6-S after treatment with 5.5-10.0 N NaOH. A reaction mechanism is proposed for the depolymerization of glycosaminoglycuronans by the action of alkali.


Microheterogeneity of chondroitin sulfates from various cartilages. Chondroitin sulfate fractions were isolated from different animal cartilages, including whale, cattle, sheep, ray and shark, by Dowex 1 chromatography followed by ethanol fractionation. Although each preparation showed a single spot when electrophoresed on cellulose acetate, both 4- and 6-sulfated disaccharides were present in chondroitinase digests of each. In particular, the main fraction of bovine tracheal chondroitin sulfate (SO4/Ga1N = 1) gave both the disaccharides in nearly equal amounts, and its IR spectrum showed absorption bands at 820 and 850 cm-1. This fraction yielded three types of tetrasaccharides after digestion with testicular hyaluronidase. Structural studies on these tetrasaccharides, using P. vulgaris chondro-4-sulfatase followed by chondroitinase, showed that one of them is a hybrid consisting of the 4- and 6-sulfated residues. In the light of these facts, a nomenclature for chondroitin sulfates is discussed.


The distribution of sulfated uronic acid and hexosamine residues in heparin and heparan sulfate. Heparins from various sources and heparan sulfate from umbilical cords have been subjected to Smith-degradation and reaction with nitrites. These procedures were effective for providing data relating to the distribution of sulfated iduronic acid residues in the molecule. Results indicated that heparins may have, as a prominent structural feature of the molecule, non-sulfated uronic acid distributed in single sequences, much as had been shown previously for N-acetylglucosamine residues. Sulfated uronic acid, however, may occur in multiple sequences of up to 5 or 6 residues. Heparan sulfate was found to have a major proportion of its ester sulfate on iduronic acid rather than hexosamine units, thereby having sections similar to those in heparins, though in considerably lower proportion.


Bovine periodontal ligament. An invesitation of the collagen, glycosaminoglycan and insoluble glycoprotein components at different stages of tissue development. Periodontal ligaments from unerupted, partially erupted and mature teeth were extracted with 0.15 M NaCl. The major reducible collagen cross-link in each insoluble fraction was dehydrodihydroxylysinonorleucine; the dehydroydroxylysinonorleucine contents were smaller. There was no significant difference in the quantities of these cross-links relative to collagen contents in the three speciments, but one of the precursors, hydroxyallysine, markedly decreased in the older tissue. The amino acid compositions of the trypsin-resistant insoluble fractions were generally characteristic of collagen. Analyses of separated glycopeptides revealed the presence of insoluble non-collagenous glycoproteins and collagen hexoses. The latter were lower in the mature ligament. Hyaluronic acid progressively decreased relative to chondroitin sulphate on eruption and maturation. A hyaluronidase-resistant glycosaminoglycan, probably dermatan sulphate, occurred in the NaCl-insoluble fraction of the mature ligament and in appreciable amounts in all NaCl extracts.


Biological significance of helical conformation of acid polysaccharides. Optical rotary dispersion spectra have been studied of methylene blue complexes of glycosaminoglycans and different derivates of chondroitin 4-sulphate (free and protein bound complexes of different molecular weights and degrees of sulphation). Structural requirements of Ch-4S induction of the extrinsic Cotton effect in methylene blue were formulated. This Cotton effect is supposed to reflect the right-handed helical structure of the polysaccharide molecules. Ch-4S displayed induced Cotton effect both in free and in protein bound form; molecular weight should exceed 2-3000 (at least 4-6 disaccharide units), degree of sulphation should exceed 3% sulphate-sulphur content (more than 0.5 sulphate per one disaccharide unit). For comparison the effect of different derivatives of Ch-4S on the rate of in vitro collagen fibril formation was investigated. Close correlation was found between the macrostructural characteristics and the possible biological effect of Ch-4S. The biological significance of the macrostructural properties of Ch-4S is discussed.


The need for modification of the polyvinyl sponge model of connective tissue growth. Histoligic and biochemical studies in the rabbit. Quantitative histopathologic and biochemical comparisons were made between polyvinyl sponge capsular and sponge tissue in the rabbit on different days after subcutaneous implantation. Up to 9 days the predominant cell type in the capsular tissue is the fibroblast and in the sponge it is the neutrophil. During this time period the sponge tissue shows lower rates of (14C) proline and (14C) cytidine incorporation and lower rates of total (14C) collagen synthesis than the surrounding capsule. Different gel electrophoretic patterns of isolated radioactive proteins are found in sponge and capsule at 6 days. These biochemical differences appear to be related to the small number of fibroblasts, relative to granulocytes present in sponges during the first 9 days after implantation. It is suggested that future biochemical investigations of the early phase of connective tissue reactions (first 9 days) in this model utilize sponge capsular tissue within 1 cm of the sponge edge instead of the sponge and its contents.


Age related changes in the synthesis of connective tissues in the rabbit. The synthesis of collagen and non-collagen proteins was studied in rapidly growing male New Zealand rabbits. Animals were divided into sexually immature (i.e. 4 and 8 week old) and mature (i.e. 15 and 26 week old) groups. Comparisons of aorta, lung, and liver protein synthesis and accumulation were made to determine the effect of aging on both collagen and non-collagen proteins. In all tissues there appeared to be an age-related decrease in the relative percent of collagen synthesized. In the lung and liver the relative decrease in percent collagen synthesis resulted from an age-related increase in the synthesis rate of non-collagen proteins. In the aorta the decrease in percent collagen synthesis was primarily a result of a dete of accumulation of tissue proteins correlates more directly with the growth and degree of differentiation of a tissue than with direct animal age.


Collagen and mechanical strength in various organs of rats treated with D-penicillamine or amino-acetonitrile. After oral treatment with D-penicillamine (D-Pc) or with aminoacetonitrile (AAn) for 10 days, mechanical and chemical parameters were studied simultaneously in various organs of Sprague Dawley rats. Tensile strength of skin strips and of tail tendons, breaking strength of femur bones and tensile strength of granuloma tissue (induced by implanted glass rods) were measured and calculated. In the same tissue the soluble collagen fractions and the insoluble collagen were determined. Total collagen and the ratio insoluble vs. soluble collagen were calculated. Tensile strength of skin, tendon and granuloma tissue were greatly reduced by D-Pc treatment but only minimally influenced by AAN treatment. On the other hand only AAN significantly reduced the breaking strength of bone. All these changes were closely correlated with the content of insoluble collagen in the respective tissues. The correlation coefficients to total collagen were similar but lower. The correlation coefficients between strength and the ratio insoluble vs. soluble collagen were generally still lower. Earlier findings in aged and corticoid treated rats, proving that insoluble collagen content determines mechanical strength of connnective and supporting tissue thus could be confirmed.


Extent and topography of the acetylation of calcified chicken-bone collagen. Acetic anhydride in anhydrous acetone-triethylamine reacts with decalcified bone collagen in two ways. It completely acetylates the e-amino groups of lysine and hydroxylysine and causes a progressive change in the protein matrix, such that increasing amounts of degraded peptides become soluble in alkaline phosphate solution. The peptides produced have molecular weights ranging from 100,000 to less than 2,000 daltons. The number of peptides obtained indicates that there are about 10 bonds cleaved per chain. Ball-mill grinding, particle size of the decalcified bone fragments, and age of the chicken do not affect the fraction of collagen that can be solubilized. The appearance of peptides begins at the point where 90% of the e-amino groups are modified. Both the phosphate-soluble and phosphate-insoluble fractions were essentially completely N-acetylated. Calcified bone under the same conditions is rapidly acetylated to only 45% of the total e-amino groups. With further additions of reagent, the degree of modification of the phosphate-insoluble matrix stabilizes at 50 +/- 4%. At the same time, progressively larger amounts of peptides, soluble in alkaline phosphate solution, are produced. The peptides, which approach 100% N-acetylation, have a distribution of molecular weight from 2000 to 75,000 daltons, indicating 10-15 disrupted peptide bonds per chain. The peptides are initially derived from the more accessible regions of the matrix and subsequently are produced from increasingly inaccessible regions. Chromatography of samples of calcified bone collagen, which were acetylated nearly to completion, indicate that the lysyl and hydroxylysyl residues which are most difficult to modify in the calcified bone are located along continuous regions of the collagen chains. Chromatography of partially acetylated samples also show that all regions of the chains are not uniformly accessible to modification, and further, no single region of every alpha chain is exposed to an identical environment.


Chemosterilant (apholate)-induced ultrastructural changes during oogenesis in Aedes aegypti. The effect of chemosterilant, apholate, on-oogenesis has been studied in Aedes aegypti. Treatment of larvae to 20 ppm of the chemical induced ultrastural changes in the presumptive and primary follicles of the adult ovary. These changes comprised condensation of chromatin, disruption of nuclear envelope and extensive degeneration as evidenced by numerous myelin figures and residual bodies. In some primary follicles, where cellular degeneration was restricted to epithelial cells, no arrest in development was observed. However, in comparison with controls, these follicles also were retarded. Larval treatment with 30 ppm apholate completely supressed ovariolar development. High incidence of autophagy was observed in tissues at both dose levels.


[The reasons for the resistance to treatment in childhood. Presentation of the problem based on a 2 1/2-year patient study with 65 treatment-resisting children, aged 3 to 14 years]. On the basis of the experience gained with 65 children trying to resist dental treatment, the problems causing and influencing this behavior of children having an aversion against the dentist are presented. Apart from factors resulting from specific situations or specific ages, factors were found that are due to parental education, social environment, and serious psychical disturbances of the child.


Depressed duodenal calcium absorption in the diabetic rat: restoration by Solanum malacoxylon. Duodenal calcium absorption is depressed in alloxan and streptozotocin diabetic rats taking normal amounts to dietary vitamin D. Depression of absorption appears to be at least in part the result of altered metabolism of vitamin D with failure to form 1,25-dihydroxycholecalciferol (1,25-(OH)2D3), the vitamin D metabolite that acts directly on duodenum to stimulate calcium absorption. The South American plant Solanum malacoxylon causes extensive soft tissue calcification when ingested by cattle. An extract of this plant restores calcium absorption depressed by dietary strontium blockage of 1,25-(OH)2D3 formation in chicks. We gave an aqueous extract of S. malacoxylon to diabetic rats and restored duodenal calcium absorption to normal. These findings provide further evidence of the ability of a factor in the S. malacoxylon extract to mimic the actions of 1,25-(OH)2D3 on duodenal calcium transport and reinforce the hypothesis that abnormal vitamin D metabolism is an important determinant of depressed duodenal calcium absorption in diabetes.


Steroid metabolism of foetal tissues. I. Metabolism of pregnenolone-4-14C by human foetal ovaries. The metabolism of pregnenolone-4-14C of ovaries from human foetuses ranging in age from the 14th through the 42th week of gestation was studied by a double isotope method in vitro. Progesterone was found as the main metabolite. The rate of conversion is correlated with histochemical findings of the same material and comparison with the FSH- and LH-concentration in human foetal pituitaries is made. As further metabolites only 17alpha-hydroxypregnenolone and dehydroepiandrosterone were isolated. For comparison tissue from a polycystic ovary was investigated with the same method.


Steroids in umbilical cord plasma from normal term deliveries. Quantitation of nine steroids in 36 cord plasma samples and four plasma pools is reported and an attempt to isolate and quantitate oestetrol is described. A wide range of values was found, which did not show correlations to the weight of the newborn or of the placenta. Comparison with data from the literature are made and correlations among the steroid levels according to the sex of the foetus are looked for. The possible relationship of difference in HCG and steroid concentration according to the sex of the foetus is discussed.


Toxicology of octabromobiphenyl and decabromodiphenyl oxide. Decabromodiphenyl oxide (DBDPO) and octabromobiphenyl (OBBP) perform well as fire-retardant additives for thermoplastics. Both compounds have low acute oral toxicity and low skin absorption toxicity. They are neither primary skin irritants or skin sensitizers and are only mildly irritating to the eyes. A 30-day dietary feeding study in rats established 8 mg DBDPO/kg-day as an unequivocal no-effect level and 80 mg/kg-day as a marginal effect level. A no-effect level was not established for OBBP in a comparative study. A 2-yr rat study providing 0.1 mg DBDPO/kg-day in the diet revealed the bromine concentration reached a plateau in the liver within 30 days, while the concentration in adipose tissue slowly increased. A comparable OBBP study revealed bromine concentration in the liver and adipose tissue increased steadily and rapidly with no attainment of a plateau during 180 days of the study. Neither compound produced an accumulation of bromine in other tissues. After administration of 14C DBDPO, all 14C activity was eliminated via the feces within 2 days. After administration of 14C OBBP, 62% was eliminated with a half-life of less than 24 hr; the half-life for the remainder was greater than 16 days. In a teratology study, 10, 100, or 1000 mg DBDPO/kg-day had no effect in rats. Reproductive capacity of rats was not effected at 3, 30, or 100 mg DBDPO/kg-day. No effects were observed on cytogenetic examination of bone marrow cells of parents and weanlings from the reproduction study.


Polymers in contact with the body.

Polymers in contact with the body. The clinical use of polymeric materials in the body to repair and restore damaged or diseased tissues and organs is substantially increasing on an annual basis. Concomitant with this use is an increase in materials related research on medically used polymers. Information on the historical and current clinical use of polymeric materials is provided in order to establish a basis for the philosophy and problems encountered in assessing the acceptability of various polymers in the biological system. The requisite properties which must be demonstrated by a polymer in contact with the body are discussed from two viewpoints, i.e., the effects of the material on the stability of the host and the effects of the host on the stability of the material. In addition, the effects of synthesis, processing, storage, sterilization, implantation, and possible degradation of polymers are discussed, poly(ethylene terephthalate) being used as an example.


Regulation of muscle contraction: bindings of troponin and its components to actin and tropomyosin. The bindings of troponin components to actin and tropomyosin has been studied by cosedimentation with actin and affinity chromatography. It is shown that troponin binds to actin and tropomyosin in the presence and absence of calcium but the binding to actin is sensitive to ionic strength. Troponin-I + C binds to actin-tropomyosin in the absence of calcium but not to actin or tropomyosin alone. Troponin-I binds to actin and the binding is improved in the presence of tropomyosin even though troponin-I does not bind to tropomyosin alone. Troponin-C does not bind to actin or tropomyosin. The results suggest that the binding of troponin by actin is influenced by tropomyosin. A model of regulation by troponin is proposed.


Lipophilic proteins encoded by mitochondrial and nuclear genes in Neurospora crassa. Mitochondrial proteins soluble in neutral chloroform-methanol (2:1) were separated from lipids by ether precipitation and resolved by Sephadex G-200 filtration in the presence of dodecylsulfate into two major fractions eluting in the excluded region (peak I) and in a region of an apparent molecular weight 8000 (peak II). Residual phospholipids are found only in peak II. Peak I consists of several aggregated small polypeptides of molecular weights around 8000, which can be disaggregated by mild oxidation with performic acid. Cycloheximide stimulates almost two-fold incorporation of radioactive phenylalanine into peak I proteins but inhibits labelling of peak II proteins by 95%. Chloramphenicol and ethidium bromide inhibit the synthesis of peak I proteins by 70% and 95% respectively, but do not affect labelling of peak II proteins. At least 30% of the translation products of mitochondrial DNA in vitro behave like peak I proteins: they are soluble in organic solvents, they aggregate in dodecylsulfate buffer after removal of phospholipids and they contain species of molecular weights around 8000 that disaggregate upon oxidation. The data strongly suggest that the proteins of peak I are encoded by mitochondrial genes and synthesized on mitochondrial ribosomes, whereas the proteins of peak II are encoded by nuclear genes and synthesized on cytoplasmic ribosomes. Both groups of lipophilic proteins are very similar in their molecular weights, but the mitochondrially coded peak I proteins have the unique property of forming large heat-stable aggregates in the presence of dodecylsulfate.


Biogenesis of mitochondrial membranes in Neurospora crassa. Mitochondrial protein synthesis during conidial germination. The conidia of Neurospora crassa entered logarithmic growth after a 1-h lag period at 30 degrees C. Although [14C]leucine is incorporated quickly early in growth, cellular protein data indicated that no net protein synthesis occurred until after 2 h of growth. Neurospora is known to produce ethanol during germination even though respiratory enzymes are present. Also, Neurospora mitochondria isolated from cells less than 3-h old are uncoupled. Since oxygen uptake increased during germination, was largely cyanide-sensitive, and reached a maximum at 3 h, it is hypothesized that during early germination the uncoupled electron transport chain merely functions to dispose of reducing equivalents generated by substrate level ATP production. The rate of protein synthesis in vitro by mitochondria isolated from 0-8-h-old cells increased as did cell age. Mitochondrial protein synthesis in vivo, assayed in the presence of 100 mug cycloheximide/ml, increased from low levels in the cinidia to peak levels at 3-4 h of age and then slowly decreased. The rate of mitochondrial protein synthesis in vivo was linear for at least 90 min in 0-4-h-old cells, but declined after 15 min of incorporation in 6 and 8-h-old cells. The products of mitochondrial protein synthesis in vivo were analyzed with dodecylsulfate gel electrophoresis and autoradiography. Early in germination 80% of the synthesis was of two small proteins (molecular weights 7200 and 9000). At 8 h 85% of the radioactivity was in 10 larger proteins (12 200 to 80 000). Within the high-molecular-weight class, proteins of between 12 000 and 21 500 molecular weight were preferentially lavelled early in germination, whereas after 8 h of growth proteins of 27 500 to 80 000 molecular weight were preferentially labelled. It is hypothesized that the 7200 and 9000-molecular-weight products of mitochondrial protein synthesis combine with other proteins to form the larger proteins found later in growth. The availability of these other proteins in cells of different ages could affect the rate of mitochondrial protein synthesis in vivo.


Morphogenesis of the membrane-bound electron-transport system in sporulating Bacillus megaterium KM. The properties of electron transport systems present in soluble and particulate fractions of spores of Bacillus megaterium KM?HAVE BEEN COMPARED WIth those of similar fractions prepared from exponential-phase vegetative cells of this organism. The timing and localization of modifications of the electron transport system occurring during sporulation have been investigated by using a system for separating forespores from mother cells at all stages during development [8]. Spore membranes contained cytochromes a + a3, and o at lower concentrations than in vegetative membranes, and in addition cytochrome c, which was not found in exponential-phase vegetative membranes. An NADH oxidase activity of similar specific activity was found in both spore and vegetative membranes but DL-glycerol 3-phosphate and L-malate oxidase activities were found only in vegetative membranes. A soluble NADH oxidase of low specific activity was found in spores and vegetative cells which probably involves a flavoprotein reaction with oxygen because the activity was stimulated by FAD or FMN and difference spectra of concentrated soluble fractions showed spectra typical of a flavoprotein. Particulate NADH oxidase was sensitive to all classical inhibitors of electron transport tested whereas soluble NADH oxidase was insensitive to many of these inhibitors. Cytochrome c was formed between stage I and II of sporulation and this coincided with a five-fold increase in NADH-cytochrome c reductase activity. Forespore membranes had lower contents of cytochromes than sporangial cell membranes but similar levels of NADH and L-malate oxidases; DL-glycerol 3-phosphate oxidase activity could not be detected in either membranes by stage III of sporulation. This characterization of spore electron transport systems provides a basis for suggestions concerning initial metabolic events during spore germination and the effect of a number of germination inhibitors.


Preparation and properties of vertebrate smooth-muscle myofibrils and actomyosin. A new technique for obtaining a myofibril-like preparation from vertebrate smooth muscle has been developed. An actomyosin can be readily extracted from these myofibrils at low ionic strength and in yields 20 times as high as previously reported. The protein composition of all preparations has been monitored using dodecylsulfate-gel electrophoresis. By this method smooth muscle actomyosin showed primarily only the major proteins, myosin, actin and tropomyosin, while the myofibrils contained, additionally, three new proteins not previously described with polypeptide chain weights of 60000, 110000 and 130000. The ATPase activities of both the myofibrils and actomyosin preparations are considerably higher than previously described for vertebrate smooth muscle. They are sensitive to micromolar Ca2+ ion concentrations to the same degree as comparable skeletal and cardiac muscle preparations, even though troponin-like proteins could not be identified in these smooth muscle preparations. From the latter observation and the presence of Ca2+-sensitivity in tropomyosin-free actomyosin it is suggested that this calcium sensitivity is, as in some invertebrate muscles, a property of the myosin molecule.


Changes in the structure, composition and function of sarcoplasmic-reticulum membrane during development. The structure, chemical composition and function of the microsomal fraction, isolated by differential centrifugation and purified on sucrose gradients, from muscle of fetal, newborn and young rabbits were characterized and compared with those of sarcoplasmic reticulum vesicles from adult muscle. Negative staining shows that the microsomal vesicles isolated from muscles of embryos and newborn animals are smooth, in contrast to vesicles obtained from adult muscle which contain 4-nm particles on their surface. The particles appear first in the microsomal vesicles from muscles of 5--8-day-old rabbits. Their number increases with the age of the animals. Ca2+-pump protein, with molecular weight about 100000, accounts for 10% of the total protein content in sarcoplasmic reticulum membrane, isolated at the earliest stages of development analysed. Its amount increases continuously with the rabbit's age to the adult value of about 70% of total sarcoplasmic reticulum protein. The low amount of 100000-dalton protein and lack of 4-nm surface particles in sarcoplasmic reticulum vesicles obtained from fetal and newborn rabbits are strictly correlated with the low activity of Ca2+-dependent ATPase and the ability to take up Ca2+. These activities rise in parallel with the age of the rabbits. On the other hand, Mg2+-dependent ATPase activity is very high at the early stages of development and declines continuously to a low value in sarcoplasmic reticulum from adult muscle. The sarcoplasmic reticulum membrane from fetal and newborn rabbits contains a higher amount of lipids as compared with the membrane present in the muscle of adult animals. The ratio of both phospholipid to protein and neutral lipid to protein decreases with the age of the rabbits. The composition of sarcoplasmic reticulum phospholipids also changes during development.


Studies of energy transport in heart cells. Mitochondrial isoenzyme of creatine phosphokinase: kinetic properties and regulatory action of Mg2+ ions. 1. The kinetic properties of mitochondrial creatine phosphokinase (Km for all substrates and maximal rates of the forward and reverse reaction) have been studied. Since (a) Km value for MgADP- (0.05 mM) and creatine phosphate (0.5 mM) are significantly lower than Km for MgATP2- (0.7 mM) and creatine (5.0 mM) and (b) maximal rate of the reverse reaction (creatine phosphate + ADP leads to ATP + creatine) equal to 3.5 mumol times min-1 times mg-1 is essentially higher than maximal rate of the forward reaction (0.8 mumol times min-1 times mg-1), ATP synthesis from ADP and creatine phosphate is kinetically preferable over the forward reaction. 2. A possible regulatory role of Mg2+ ions in the creatine phosphokinase reaction has been tested. It has been shown that in the presence of all substrates and products of the reaction the ratio of the rates of forward and reverse reactions can be effectively regulated by the concentration of Mg2+ ions. At limited Mg2+ concentrations creatine phosphate is preferably synthesized while at high Mg2+ concentrations (more ATP in the reaction medium) ATP synthesis takes place. 3. The kinetic (mathematical) model of the mitochondrial creatine phosphokinase reaction has been developed. This model accounts for the existence of a variety of molecular forms of adenine nucleotides in solution and the formation of their complexes with magnesium. It is based on the assumption that the mitochondrial creatine phosphokinase reactions mechanism is analogous to that for soluble isoenzymes. 4. The dependence of the overall rate of the creatine phosphokinase reaction on the concentration of total Mg2+ ions calculated from the kinetic model quantitatively correlates with the experimentally determined dependence through a wide range of substrates (ATP, ADP, creatine and creatine phosphate) concentration. The analysis of the kinetic model demonstrates that the observed regulatory effect of Mg2+ on the overall reaction rate can be expained by (a) the sigmoidal variation in the concentration of the MgADP- complex resulting from the competition between ATP AND ADP for Mg2+ and (b) the high affinity of the enzyme to MgADP-. 5. The results predicted by the model for the behavior of mitochondrial creatine phosphokinase under conditions of oxidative phosphorylation point to an intimate functional interaction of mitochondrial creatine phosphokinase and ATP-ADP translocase.


On the primary structure of human plasminogen and plasmin. Purification and characterization of cyanogen-bromide fragments. Most of the cyanogen bromide fragments obtained from human plasminogen and plasmin have been purified using combinations of gel filtration and ion-exchange chromatography. The purified fragments have been characterized by molecular weight determination (dodecyl sulphate electrophoresis), amino acid analysis, carbohydrate analysis and direct NH2-terminal amino acid sequence determination. Since some of the purified fragments were compounds with uncompletely cleaved methionyl bonds it was possible to clarify the organization of most of the cyanogen bromide fragments in the plasminogen molecule. The fragment containing the arginyl-valyl bond cleaved during the second step of the activation process is further identified. It is also shown that the microheterogeneity that normally exists in human plasminogen probably has its origin in several sites. One such site is situated in the light (B) chain of plasmin, while another is situated in the carboxyterminal part of the heavy (A) chain. Neither of these sites seems to contain sialic acid.


Proteoglycans of guinea-pig coastal cartilage. Fractionation and characterization. Proteoglycans were extracted, in a yield of about 90%, from costal cartilage of young, growing guinea-pigs. Three solvents were used in sequence: 0.4 M guanidine - HCl, pH 5.8, 4 M guanidine - HCl, pH 5.8, and 4 M guanidine - HCl/0.1 M EDTA, pH 5.8. The proteoglycans were purified and fractionated by cesium chloride density gradient ultracentrifugation under associative and dissociative conditions. Gel chromatography on Sepharose 2 B of proteoglycan fractions from associative centrifugations showed the presence of both aggregated and monomer proteoglycans. The ratio of aggregates to monomers was higher in the second extract than in the other two extracts. Dissociative gradient centrifugation gave a similar distribution for proteoglycans from all three extracts. Thus, with decreasing buoyant density there were decreasing ratios of polysaccharide to protein, and of chondroitin sulfate to keratan sulfate. In addition, there was with decreasing density an increasing ratio of chondroitin 4-sulfate to chondroitin 6-sulfate. Amino acid analyses of dissociative fractions were inaccordance with previously published results. On comparing proteoglycan monomers of the three extracts, significant differences were found. Proteoglycans, extracted at low ionic strength, contained lower proportions of protein, keratan sulfate, chondroitin 6-sulfate and basic amino acids than those of the second extract. The proteoglycans of the third extract also differed from those of the other extracts. The results indicate that the proteoglycans of guinea-pig costal cartilage exist as a very polydisperse and heterogenous population of molecules, exhibiting variations in aggregation capacity, molecular size, composition of protein core, degree of substitution of the protein core, as well as variability in the type of polysaccharides substituted.


Transport ATPases of cardiac sarcolemma in 20,25-diazacholesterol induced myopathy. 20,25-Diazacholesterol, known to induce myotonia in skeletal muscle, also affects cardiac muscle as can be concluded from the development of cardiomegaly. At the same time (Na+, K+) ATPase of cardiac sarcolemmal membranes of the 20,25-diazacholesterol treated rats showed an increased activity as compared with control animals (91 percent and 46 percent stimulation respectively). The Ca++ stimulated ATPase showed the same tendency (96 percent and 64 percent stimulation). In the plasma of the treated rats creatine phosphokinase activity was found to be elevated whereas the amount of protein-bound iodine was decreased, a finding that is common in myotonic dystrophy.


Reversible and irreversible phases of serotonin depletion by 4-chloroamphetamine. Pretreatment of rats with an agent that inhibits uptake into serotoninergic neurons [Lilly 110140: 3-(p-trifluoromethylphenoxy)-N-methyl-3-phenylpropylamine hydrochloride] prevented the depletion of brain serotonin by 4-chloroamphetamine, presumably by preventing the entry of 4-chloroamphetamine into the serotonin neuron. When the uptake inhibitor was given after 4-chloroamphetamine, the lowering of both serotonin and tryptophan hydroxylase levels in brain was reversed. Serotonin levels returned to normal after 110140 administration to 4-chloroamphetamine-treated rats at a rate similar to the calculated rate of serotonin turnover in rats treated with 110140 alone. Progressively less reversibility of the 4-chloroamphetamine effect occurred when 110140 was injected at 8, 16 and 24 hr after 4-chloroamphetamine, and no reversibility was observed when 110140 was injected at 32 or 48 hr after 4-chloroamphetamine. These findings indicate that the depletion of brain serotonin by 4-chloroamphetamine is initially reversible but that there is a gradual transition into an irreversible effect between 24 and 48 hr after 4-chloroamphetamine injection. Apparently the prolonged presence of 4-chloroamphetamine inside the serotonin neuron produced by continual reuptake of 4-chloroamphetamine is required for the semi-permanent depletion of brain serotonin stores.


Noradrenaline stimulation of a membrane pump in frog skeletal muscle. Resting membrane potentials were recorded from the frog sartorius before and during exposure to noradrenaline (NA; 10(-2) M) in the presence or absence of ouabain (10(-3) M) and during manipulation of the extracellular potassium concentration. In potassium-free Ringer, NA produced no significant change in the resting membrane potential. With the extracellular potassium concentration at 2.5 or 5mM, NA caused a hyperpolarization. This reversed to a depolarization when extracellular K+ was greater than 7 mM. In all cases NA-induced changes were blocked by ouabain. These effects suggest that NA stimulates a membrane cation pump, the Na : K coupling ratio of which is affected by the available ion concentrations.


Reversible inhibition of (Na+ + K+)-ATPase with a cardiac glycoside. The effect of a semisynthetic cardiac glycoside, Actinogen (Ay22241), on Na+ + K+ - ATPase was studied. Ay22241 was found to be as an effective inhibitor of the enzyme as ouabain, Ay22241 inhibition was a time dependent process and was completely reversible. While ouabain inhibition was also time dependent, it was only partially reversible. This reversibility with Ay22241 should make it a useful tool in studying the mode of action of cardiac glycosides.


Influence of drugs on striatal and limbic homovanillic acid concentration in the rat brain. Homovanillic acid (HVA) was measured in the corpus striatum aan the limbic structures nucleus accumbens and olfactory tubercle of the rat, under normal conditions and after different drug treatments. Clozapine, thioridazine, morphine and physostigmine induced a similar percentage HVA increase in the three brain structures studied. Haloperidol and pimozide induced a higher percentage increase of HVA in the corpus striatum and nucleus accumbens when compared with the olfactory tubercle. Oxotremorine induced the highest HVA levels in the nucleus accumbens. Probenecid induced a significantly higher percentage accumulation of HVA in the limbic structures, especially in the olfactory tubercle. The HVA rise seen after haloperidol was suppressed by pretreatment with p-chlorophenylalanine or amino-oxyacetic acid in all structures studied. After atropine or trihexylphenidyl treatment the HVA rise induced by haloperidol was slightly suppressed in the limbic structures only. Our results suggest that not only under normal conditions but also after treatment with various types of drugs, dopamine metabolism as reflected by the HVA levels, is closely related in the different rat brain structures.


Interaction of leukocytes and endotoxin with the plasmin and kinin systems. Leukocytes can generate a substance that, when added to some partially purified human kininogen, is capable of forming kinins. The addition of endotoxin or polystyrene latex particles to the incubated leukocytes doubled the amount of kinin generated. Certain preparations of kininogen, however, failed to allow kinin formation by the leukocytes. No evidence could be found that an activator of prekallikrein or a kallikrein was present in the granulocyte preparations. However, the addition of highly purified plasminogen to inactive kininogen preparations restored their ability to generate kinins in the presence of leukocytes. All the kininogen preparations that allowed kinin formation when incubated with leukocytes contained plasminogen. These data suggest that a plasminogen activator is present on the leukocyte surface. This activator activates plasminogen to form plasmin which in turn acts on kininogen to release a kinin and thus provides a mechanism for the formation of kinins in inflammatory exudates and during endotoxemia.


Stimulation of vitamin D-dependent Ca-ATPase and of intestinal caldium absorption by diphenylhydantoin. The effects of diphenylhydantoin (DPH) and ouabain were studied in vitro on Mg-ATPase, Ca-ATPase and alkaline phosphatase (AlPase) in isolated brush borders from rat jejunum, and in vivo on intestinal calcium absorption. Vitamin D-deficient, -repleted and normal rats were used in this study. Repletion of deficient animals with vitamin D restored Ca-ATPase activity and AlPase activity partly. Ca-absorption was normalized by repletion with the vitamin. DPH greatly stimulated Ca-ATPase activity in vitro and Ca-absorption in vivo, but it inhibited AlPase activity. Mg-ATPase was not affected by vitamin D, nor by DPH. Ouabain had no consistent effect on any of the parameters studied. It was concluded that Ca-ATPase, and not AlPase, is involved in the transport of calcium through the jejunal microvillous membrane, and that DPH enhances Ca-absorption by activation of Ca-ATPase.


Studies of the hemopoietic microenvironment. IV. In vivo microscopic and histochemical study of allografts of bone marrow in the hamster cheek pouch chamber. Within a transparent chamber enclosing subcutaneous tissue of the hamster cheek pouch, allogeneic femoral marrow was grafted. This permitted in vivo microscopic study of the grafts for 14 days. Five to seven days after grafting, blood flow was established within sinusoids arranged in interconnecting polygonal networks, a pattern characteristic of marrow. All vessels appeared to have a complete and continuous endothelial lining. The graft contained foci of erythropoiesis, granulopoiesis, and megakaryocytopoiesis, but no osteogenic activity. Degeneration of the grafts began to occur between days 9 and 13 after implantation. Generally, this was preceded by infection within the chamber, fibroblastic proliferation, and an accumulation of acid mucopolysaccharide in the stroma. The concomitant loss of hemopoiesis and accumulation of acid mucopolysaccharides was consistent with our hypothesis reported previously that excessive concentrations of acid mucopolysaccharide in the hemopoietic microenvironment are not conducive to supporting hemopoiesis, especially erythropoiesis. Degeneration of the grafts probably was the result of infection since there was no evidence of an immunologic response to the graft, and in chambers not containing grafts, similar infections and changes in the connective tissue were observed. The technique of grafting marrow into the hamster cheek pouch chamber provides a model for long-term, in vivo microscopic study of bone marrow. However, methodologic improvements, especially in the control of infections, are needed.


Chalone regulation of the epidermal cell cycle. Purified epidermal G2 chalone does not inhibit DNA synthesis or influx of S-phase cells and is therefore cell cycle phase-specific, inhibiting only the flow of cells into M-phase.


Effect of albumin on uncoupling of oxidative phosphorylation by chinoform in rat liver mitochondria. Addition of serum albumin diminished the uncoupling effect of chinoform on oxidative phosphorylation in rat liver mitochondria. Upon increasing the concentration of magnesium ions in the medium, the action of serum albumin was diminished. These results indicate that serum albumin combines with chinoform in competition with magnesium ions.


The effects of glycosaminoglycans on surface-mediated interactions of lymphoid cells. The effect of chondroitin sulphate (CS) was studied in two cell surface-mediated interactions, the complement-independent contact reaction and antibody-induced capping of H-2 antigens. CS inhibited contact cytotoxicity when present in the reaction medium, but preincubation in CS of either killer cells or target cells had no effect. The capacity of CS-pretreated target cells to absorb H-2 antibodies was not reduced. However, a similar pretreatment resulted in an increased readiness to capping as induced by H-2 antibody followed by an anti-mouse immunoglobulin; in control cells time taken for the same percentage of caps to be formed was considerably longer than in CS-pretreated cells. A possible mechanism of this CS effect and the general biological role of glycosaminoglycans (particularly in immune interactions) is discussed.


Glucose turnover in the exercising dog with chemically induced diabetes and the effect of methylprednisolone. Dogs with indwelling polyethylene arterial and venous catheters ran on a treadmill (slope 15 per cent, speed 100 m./min.). Diabetes was produced by alloxan or by a combination of alloxan and streptozotocin. Glucose turnover was measured according to the primed constant-rate infusion technics with 2-3H-glucose as tracer. In resting diabetic dogs plasma glucose varied between 200 and 650 mg./100 ml. There was a direct linear correlation between the hepatic glucose output (Ra) and the plasma glucose level. Exercise increased both Ra and the clearance rate (CR) of glucose; however, Ra could not match the rate of disappearance (=renal loss plus glucose uptake of the muscle), causing the plasma glucose to decline more rapidly than in the running control dogs. Two to three days' treatment with methylprednisolone (MP, 3-3.2 mg./kg./day) caused a higher resting glucose level and a higher Ra. Exercise greatly increased the plasma glucose concentration, partly because MP enhanced the hepatic response but mainly because it essentially prevented the rise of CR. It is concluded that (a) in chemically induced diabetes, the variable glucose level is the result of a variable rate of hepatic glucose output; (b) the increase of Ra by MP treatment does not increase the plasma glucose in the normal dog but significantly aggravates the alloxan diabetes, (c) diabetes reduces the effect of exercise on the glucose uptake of the muscle, and this effect is potentiated by the inhibitory action of the glucocorticoid. This latter becomes unmasked only when the insulin secretion is impaired.


Barbituates and biliary function.

Barbituates and biliary function. This paper reviews the principal effects of phenobarbital on biliary function. Phenobarbital administration is followed by an increase in bile flow. This is mainly due to an increase in the bile salt-independent fraction of canalicular bile flow possibly through an increase in canilicular Na+-K+ ATPase activity. In addition, bile salt excretion may be increased. This effect of barbiturates on choleresis appears to be independent of microsomal enzyme induction. Barbiturates increase the uptake, storage and excretion of various dyes, for example sulfobromophthalein. Phenobarbital increases bilirubin clearance by the liver; it enhances bilirubin-UDP-glucuronyl transferase activity; whether the influence on bilirubin clearance is related to the effect on the enzyme is unknown. The influence of phenobarbital on biliary lipids is markedly different from one species to the other. In the rhesus monkey and in the rat, the relative concentration of cholesterol is decreased; in the hamster it is increased, and in man it appears largely unaffected. These effects of phenobarbital have been utilized in the treatment of chronic unconjugated hyperbilirubinemia and of certain cholestatic syndromes. Phenobarbital alone has been useful, so far, in the treatment of cholesterol gallstones.


Three chromosome abnormalities (trisomy 21, XXY, and a de nova reciprocal translocation) in a child with 48, XXY, + 21,T(6;10)(P22-24;P12). An infant with chromosomally normal parents was found to have double aneuploidy and a reciprocal translocation between chromosomes number 6 and 10. This cooccurrence may represent a very rare coincidence or may indicate that primary nondisjunction and chromosome rearrangements may be more than coincidentally related.


Studies on the polymorphism of C3, Tf and Bg in Down's syndrome and other diseases. The distribution of phenotypes C3, Ff and Bg was investigated in sera of patients with Down's syndrome, oligophrenia, Wilson's disease and heart infarct. Quantitative determination of the concentration of C3 and C4 components of human complement was also carried out in these patients. The results are compared to healthy controls and are discussed with already reported data from other authors. Despite differences in the percentage distribution of various phenotypes in the patients' sera as compared to that of the controls, no statistically significant association could be established.


Types of enzymatic overdosing in trisomy 21: erythrocytic superoxide dismutase-AJ and phosphoglucomutase. A comparative study was carried out on the hemolysates of 6 trisomic 21 and 6 normal subjects, by electrophoresis in starch gel, determining by a combined staining method both SOD-A (former IPO-dimer) and PGM activity. The enzymes were found statistically to be in a hyperactive status, the ratio of trisomic to normal values being approximately equalt to 1.4. SOD-A supraactivation is the effect of a genic dose, as demonstrated in earlier works (Sichitiu, 1973; Sichitiu et al., 1974; Sinet et al., 1974), whereas PGM hyperactivity appears to be modified secondarily, the same as the activity of other cellular enzymes in Down's disease.


Familial translocation t(3p-;21q+) associated with both Down's and Sturge-Weber's syndrome in unbalanced state. A boy with both Down's and Sturge-Weber's syndrome was found to have a partial trisomy 21 as a consequence of a familial translocation t(3p-;21q+) which is not reciprocal. Judging from the structure of the involved chromosomes studied by banding and photometrical techniques, the loss of relatively large material of 21q is to be suggested. The meiotic segregation appears to depend on the involved 3p segment and not on the involved centromere of No. 21 as actually expected. The pedigree of the family shows 6 balanced carriers through 3 generations in addition to the propositus. The risk of having offspring with Down's syndrome obviously concerns female carriers in the first place, whereas the male carriers rather produce balanced carriers. Of the additional Sturge-Weber's syndrome there was no cytogenetical cause as expected.


A radioisotopic method to measure delayed type hypersensitivity in the mouse. II. Cell transfer studies. Sensitized lymphoid cells could transfer to normal non-sensitized (naive) mice by 24-48 h after antigen challenge in the ear, the capacity to incorporate, at the site of antigen deposition, 5-iodo-2'deoxyuridine-125I in amounts significantly above those obtained in control mice. This was associated with a mononuclear cell infiltration in the pinna. In contrast to lymphoid cells, serum antibodies were unable to transfer a 24- or 48 hour ear reaction. The cells responsible were T lymphocytes as demonstrated by successful transfer following enrichment for T lymphocytes, and abrogation of transfer following treatment with anti-theta serum and complement. Transfer was achieved whether the naive recipients were normal, T-cell deprived, pretreated with cyclophosphamide, or lightly irradiated but not when they were heavily irradiated. Adoptive transfer of the 24-hour ear response was demonstrated with three different antigenic systems. The time-response curves were different with each system although peak reactions were obtained 5 days after sensitization of the donors in all cases. The specificity patterns of the 24-hour ear reaction on transfer were similar to those obtained in the sensitized donors. The results of these studies indicate that the radioisotopic ear method can, under defined conditions, demonstrate the existence of a state of delayed type hypersensitivity in the donors.


Experimental results to immunomesenchymal suppression. Wet weight of cotton-pellet granuloma and 35SO4-uptake in the acid mucopolysaccharides (AMPS) of granulation tissue of rats and the number of plaque forming cells (PFC) of mice were investigated after the treatment with antilymphocyte globulin, prednisolone, cyclophosphamide and azathioprine. In this way, the effect of these substances could be observed in a mesenchymal reaction and in an immune reaction. The substances are mesenchymosuppressive as well as immunosuppressive, but the present results show that the effective dosage is higher for mesenchymal suppression than for immunosuppression. Antilymphocyte globulin did not cause an alteration of the 35SO4-uptake in AMPS. These experimental findings confirm the clinical experience in the treatment of the so-called autoimmunological diseases which are primary localized in connective tissue. These diseases need a higher dosage of cytostatic substances, glucocorticoids and antilymphocyte globulin in times of acute inflammatory exacerbation than in the chronic phase.


Alteration of membrane integrity by delta1-tetrahydrocannabinol. Delta1-tetrahydrocannabinol was found to be a potent inhibitor of some membrane-bound enzymes, such as Mg-ATPase, Na-K-ATPase and acetylcholinesterase. At a given concentration, the degree of inhibition varied for each enzyme; the inhibition was more pronounced for the enzymes that are parts of the membranes. As the kinetic parameters of these enzymes are functions of the membrane composition and organization, these parameters were studied in vitro in the presence of THC. Although the Mg-ATPase was inhibited by THC, there was no change in the allosteric behaviour of the enzyme, indicating that the alterations caused by THC did not affect the enzymatic structure. The Na-K-ATPase and acetylcholinesterase had a different allosteric behaviour as compared to controls; these modifications were like the alterations caused by the decrease in membrane fluidity. These results suggest the fact that THC is incorporated in the membranes and causes alterations in the physical organization of the membranes.


[Systemic changes in occupational vitiligo]. In three patients with occupational vitiligo caused by paratertiary butylphenol--the first cases observed in the BRD--a functional disorder of the liver was found. In two of these patients splenomegaly was seen. The laboratory findings, the laparoscopic examination and the result of the liver-biopsy are described. It is assumed that these changes as well as the depigmentation and a functional disorder of the thyroid (described elsewhere) are caused by the systemic action of inhaled paratertiary butylphenol.


Malformation of the canine mitral valve complex. Twenty-nine dogs, including 13 Great Danes and 5 German Shepherd Dogs and averaging 7.3 months age, were diagnosed clinically and radiographically as having mitral regurgitation. Alterations of the mitral valve complex included enlarged anulus; short thick leaflets, with an occasional cleft; short and stout or long and thin chordae tendineae; upward malposition of atrophic or hypertrophic papillary muscles; insertion of one papillary muscle directly into one or both leaflets; and diffuse endocardial fibrosis, occasionally with jet lesions in te left atrium. Other cardiac anomalies included dysplasia of the tricuspid valve (5 dogs), patent ductus afteriosus (2 dogs), aortic stenosis (2 dogs), and ventricular septal defect (1 dog).


Disseminated oesophagostomiasis in the rhesus monkey. Disseminated oesophagostomiasis was diagnosed at necropsy of 5 rhesus monkeys (Macaca mulatta). Nodular lesions were in visceral organs as well as in the colon and mesentery. Lesions varied greatly with respect to degree of organ involvement and damage due to parasite invasion. In 2 monkeys, nodular lesions were in the colon, mesentery, liver, kidneys, abdominal wall, urinary bladder, and uterus; in 1 of these 2 monkeys, a nodular lesion inhabited by a parasite was in a lung. The ectopic nodules in the less severely affected monkeys were scattered in the liver, kidneys, and abdominal wall.


A multichannel implantable telemetry system for flow, pressure, and ECG measurements. The design of telemetry systems for chronic implantation within the body have progressed from simple single-channel devices to complex multichannel systems over the last 15 yr. Although chronic measurements of temperature, ECG, and pressure have been reported with good results, measurements such as dimension and blood flow have been difficult because of heavy power requirements. The design to be described is a multiplex system that will measure up to eight individual parameters simultaneously, including blood flow. Flow is measured using an electromagnetic transducer, and by special design, the normal high power requirements of the electromagnetic technique have been reduced to a few hundred milliwatts. The system is powered by two NiCad, rechargeable batteries which are periodically recharged through the intact skin by induction at 250 kHz to an implanted pickup coil. The presently constructed units have been configured to measure ECG, two pressures, temperature and ascending aortic flow.


Control of depth and frequency of breathing during baroreceptor stimulation in cats. In 10 tracheotomized anesthetized cats during steady-state inhalation of various concentrations of CO2 and O2, the acute respiratory response to baroreceptor stimulation produced by transient inflation of a balloon placed in the descending aorta was studied. The latter induced a sudden rise in mean arterial pressure, ranging from 62 to 95 mmHg. At all PACO2 levels above 30 mmHg, elevation in arterial pressure was accompanied by an immediate drop in tidal volume (VT) and prolongation of the durations of inspiration (Ti) and total breath (Ttot). Breaths obtained during baroreceptor stimulation fell along the same VT vs. Ti and VT vs. Ttot relationships obtained in the normotensive state, suggesting that the lung volume-related vagal control of Ti and Ttot is unaffected by changes in arterial pressure. Since, for a given change in arterial pressure, a constant reduction in VT was obtained at all PACO2 levels above 30 mmHg, it can be concluded that the interaction between PACO2 and arterial pressure is additive. In three cats, at PACO2 levels below 30 mmHg, aortic obstruction resulted in brief periods of apnea. Following apnea, the control of Ti and Ttot was transiently offset, describing hysteresis pathways on the VT vs. Ti and VT vs. Ttot relationships.


Diaphragmatic activity and lung liquid flow in the unanesthetized fetal sheep. For some time it has been suggested that breathing movements are made "in utero" and recently measurements of tracheal pressure and lung liquid flow in chronic fetal preparations have led to the hypothesis that rapid changes in these parameters are the result of respiratory muscle activity. To test this hypothesis diaphragmatic electrical activity was measured in seven chronic unanesthetized fetal sheep preparations and correlated with lung liquid flow and tracheal pressure. Diaphragmatic activity led to a fall of tracheal pressure and movement of a small volume of lung liquid into the lung. After the activity ceased, tracheal pressure returned to normal and flow diminished to zero or was directed out of the lung. The breathing pattern was unassociated with the net movement of lung liquid out of the lung. A histogram of the interval between breaths revealed a changing pattern of activity throughout gestation. The pattern was significantly altered after premature delivery of one animal with a respiratory problem. These observations provide evidence that respiratory muscles are active "in utero" and that the pattern of activity changes throughout gestation.


Mu-induced polarity in the Escherichia coli K-12 ent gene cluster: evidence for a gene (entG) involved in the biosynthesis of enterochelin. A strain of Escherichia coli K-12 has been isolated that carries a Mu bacteriophage-induced mutation in the ent gene cluster. Nutritional tests together with examination of the compounds accumulated by the mutant strain indicated that the mutant was blocked both in the synthesis of 2,3-dihydroxy-benzoate and its subsequent conversion into enterochelin. Enzymic complementation assays of the mutant with several mutants each affected in one of the ent genes showed that the Mu-induced mutant was entA-, entB-, entC+, entD+, entE+, and entF+. Since the mutant produced the entD, entE, and entF gene products but was unable to produce enterochelin from 2,3-dihydroxybenzoate, it must therefore be affected in an additional protein concerned with this conversion. It is therefore postulated that the Mu-induced mutation affects a previously unrecognized gene, entG. Genetic experiments indicate that the mutation in strain AN462 which affects the three ent genes is the result of a single insertion of Mu in the ent gene cluster. This polarity mutant therefore provides evidence that three of the ent genes are part of an operon.


Purine base transport in Neurospora crassa. Observations presented in this paper point to the presence of dual transport mechanisms for the base adenine in Neurospora crassa. Competition for transport, as well as growth inhibition studies using an ad-1 auxotroph, show that the purine bases adenine, guanine, and hypoxanthine share at least one transport mechanism which is insensitive to adenosine, cytosine, and a variety of other purine base analogues. On the other hand, uptake of adenine by an ad-8 mutant strain unable to transport [8-14C]hypoxanthine at any concentration was not inhibited by guanine or hypoxanthine. This observation demonstrates the existence of an adenine-specific transport system which was also found to be insensitive to inhibition by other purine base analogues, adenosine or cytosine. Recombination analysis of ad-8 by wild-type crosses showed that the inability to transport [8-14C]hypoxanthine was a consequence of the ad-8 lesion or a closely linked mutation. Saturation plots of each system gave intermediary plateaus and nonlinear reciprocal plots which, based on comparison with pure enzyme kinetic analysis, suggest that either each system consists of two or more uptake systems, at least one of which exhibits cooperativity, or that each system is a single uptake mechanism which possesses more than two binding sites where the relative affinity for the purine base first decreases and then increases as the sites are filled.