Issue published November 1, 1967

T he absorption of fat was studied in five male subjects with cannulation of the thoracic duct in the neck by the administration of doubly labeled monoglycerides, or triglyceride as well as labeled free glycerol or labeled free oleic acid, by gastric or duodenal intubation.Total recoveries of the administered glyceride radioactivity from the lymph lipids ranged from 35 to 53% for the glycerol label (tritium) and from 35 to 57% for the fatty acid label (¹⁴C). The recovery of administered radioactive free glycerol in lymph lipids was only 4.1%, even when given in mixture with bile salts, fatty acid, and monoglyceride.A comparison of the isotope ratios of the two components (glycerol and fatty acid) of the lymph glycerides with the ratios of these components of the original meal glyceride showed little change during the initial period of fat absorption, indicating that the doubly labeled monoglycerides passed into the lymph intact. During the later part of the period of major fat absorption, the ratios in lymph lipids changed due to loss of glycerol representation, indicating monoglyceride hydrolysis and portal venous diversion of free glycerol.Confirmation of the intact nature of 2-monoglyceride during absorption was made by analyzing the amount and position of the labeled fatty acid in the lymph triglycerides. The percentage of labeled fatty acid in the various positions of the lymph triglycerides was virtually identical with that of the meal during the initial period of fat absorption and then changed reflecting isomerization of fatty acids and subsequent complete hydrolysis of the glycerides.The 2-monoglyceride pathway appears to be the major route of fat absorption for man during normal digestion and absorption of dietary triglyceride.

W e have investigated the possibility that alterations in the duration of the systolic preejection period can be used to estimate adrenergic influences on the human left ventricle. The preejection period was determined from high speed, simultaneous recordings of the phonocardiogram, carotid pulse tracing, and electrocardiogram. The preejection period was shortened by isoproterenol, epinephrine, and moderate doses of norepinephrine—all of which activate beta adrenergic receptors—and by cedilanid-D. It was unaltered by changes in heart rate induced by atropine and right atrial electrical pacing. Beta adrenergic receptor blockade by propranolol abolished the shortening effects of the three catecholamines but did not inhibit that due to cedilanid-D. Vasoconstriction, both alpha adrenergic (epinephrine and norepinephrine after propranolol) and nonadrenergic (angiotensin), prolonged the preejection period. Most of the shortening of the preejection period by beta adrenergic receptor activating agents and cedilanid-D and all of the prolongation accompanying pharmacologic vasoconstriction occurred after the onset of the first heart sound, thereby excluding changes in electrical-mechanical delay as a major factor in the observed preejection period responses. Shortening of the preejection period by beta adrenergic activity induced with isoproterenol was dose-related. Increasing doses of propranolol produced parallel shifts to the right in the isoproterenol dose-response curve.In 37 normal resting subjects intravenous propranolol (10 mg) prolonged the preejection period an average of 10 (SE ± 1) msec. In six patients with psychogenic sinus tachycardia and a patient with a pheochromocytoma the presence of excessive beta adrenergic influences on the left ventricle was demonstrated by the finding of an initially short preejection period which responded with an abnormally great prolongation to beta adrenergic receptor blockade.

W hen paroxysmal nocturnal hemoglobinuria (PNH) erythrocytes were exposed to H₂O₂ they lysed excessively and formed greater than normal quantities of lipid peroxides when compared to red cells of normal subjects and patients with most types of hematologic disease. It was also shown that lytic sensitivity to acidified serum was related to the enhanced lytic sensitivity to H₂O₂. If the lipid of PNH cells was first extracted then exposed to ultraviolet radiation more lipid peroxides were formed than in extracts of normal red blood cells. The possible explanations for these findings and their relationship to the PNH hemolytic mechanism are discussed.

A possible role for adenylcyclase in insulin secretion was investigated. Isoproterenol, a predominantly β-adrenergic agent, when mixed with an α-adrenergic blocking agent (phenoxybenzamine), stimulated insulin secretion from pieces of the rat's pancreas in vitro. Theophylline, caffeine, 3′5′-cyclic AMP, glucagon, adrenocorticotropin (ACTH), and thyrotropin (TSH), all of which are thought to act through the adenylcyclase systems in the liver and adipose tissue, also stimulated insulin secretion in vitro; oxytocin and vasopressin, which do not stimulate lipolysis in adipose tissue, were inactive. In all cases, stimulation of insulin secretion could not be detected when glucose was absent or present in only low concentrations (less than 100 mg/100 ml) and was maximal at high levels of glucose (300 mg/100 ml). When pancreatic tissue was obtained from normoglycemic rats and contained no detectable glycogen in the Islets, the stimulant effects of glucose and of theophylline were reduced or abolished by mannoheptulose and 2-deoxyglucose. When tissue was derived from rats infused for 8-10 hr with glucose and contained glycogen, theophylline, even in the absence of glucose, stimulated secretion and this effect was reduced by 2-deoxyglucose but not by mannoheptulose. It is suggested that the β-cell contains an adenylcyclase system through which phosphorylase and possibly phosphofructokinase could be activated; and that insulin secretion could depend upon and be regulated by hormones and other substances which influence the rate at which glycolysis proceeds within the β-cell.

T he production and peripheral metabolism of cortisol have been studied in 10 cirrhotics and 11 controls after i.v. tracers of cortisol-¹⁴C. The findings were as follows: (a) Total urinary excretion of radioactivity was normal (81% of the dose) but a decreased fraction was present as glucosiduronates: 18-47% of the dose (average 34%) compared to a normal average of 54%. (b) There was a distinctively abnormal pattern of cortisol metabolites, not previously observed in other illnesses: tetrahydrocortisone was decreased to 14% of the enzyme hydrolysate (normal 26%); cortolones were increased to 34% (normal 19%), owing entirely to an increase in cortolone (20α) formation, since β-cortolone (20β) was not significantly increased; Reichstein's substances U and epi-U were increased, averaging 2.6% for the former and 4.7% for the latter; tetrahydrocortisol, allotetrahydrocortisol and cortols were normal. This pattern was independent of the degree of decreased glucosiduronate formation and also independent of the presence or absence of a portacaval shunt. (c) Cortisol production, determined by isotope dilution, was normal in each of six cirrhotic patients. From these data, taken in conjunction with our previously reported findings concerning the influence of norethandrolone on cortisol metabolism, the following conclusions were drawn: (a) Cirrhotic patients have decreased A-ring reduction of cortisone to tetrahydrocortisone and correspondingly increased 20-ketone reduction of cortisone to Reichstein's substances U and epi-U and then to the cortolones. (b) Intrahepatic cholestasis, a regular pathophysiological feature of cirrhosis, may be responsible for the observed abnormal cortisol metabolite pattern in this disease. (c) The slowed metabolic turnover rate of cortisol in cirrhosis may be due to decreased transport and/or binding of cortisol to its intracellular metabolic sites rather than to abnormalities of any specific metabolizing enzymes.

A n inspiratory fall in systolic arterial pressure of more than 10 mm Hg (pulsus paradoxus) was noted in 30 of 61 patients with shock. Inspiratory right atrial pressures and total blood volumes were significantly lower in patients with pulsus paradoxus. Rapid infusion of dextran in 22 patients usually was effective in reversing the exaggerated inspiratory fall in systolic pressure. Total peripheral vascular resistance tended to be higher in the patients with pulsus paradoxus and administration of vasoconsrictor drugs often accentuated the respirator pressure variation.Respiratory effects on blood flow in the aorta, pulmonary artery, and venae cavae were studied in anesthetized, closed-chest dogs. In the control state, pulmonary arterial flow increased during inspiration but aortic flow remained nearly constant. After hemorrhage a sharp inspiratory fall in aortic flow was associated with decreased central blood volume and attenuation of the usual inspiratory increase in venae caval and pulmonary arterial flows. The respiratory changes in aortic flow after hemorrhage could be attributed both to depletion of the pulmonary reservoir and to alterations in pulmonary inflow related to changes in systemic venous return.These data indicate that blood volume depletion may precipitate pulsus paradoxus both in the anesthetized dog and in the critically ill patient. The occurrence of pulsus paradoxus may aid in the clinical recognition of the common syndrome of occult hypovolemia in patients with shock in the absence of signs of blood loss.

D ietary carbohydrate accentuation of endogenous triglyceride production has been studied in 33 patients. A broad and relatively continuous spectrum of steady-state plasma triglyceride concentrations was produced in 31 of the 33 subjects during 3 wk of a high carbohydrate (fat-free) liquid formula diet. Two patients developed plasma triglyceride concentrations in excess of 2000 mg/100 ml, and these were the only patients we have studied in which carbohydrate induction of hypertriglyceridemia seemed to be associated with a defect in endogenous plasma triglyceride removal mechanisms. In the remaining 31 patients the degree of hypertriglyceridemia was highly correlated with the insulin response elicited by the ingestion of the high carbohydrate formula (P < 0.005). No significant correlation existed between fasting plasma triglyceride concentration and either plasma glucose or free fatty acid concentrations after the high carbohydrate diet, nor was the degree of hypertriglyceridemia related to degree of obesity. It is suggested that hypertriglyceridemia in most subjects results from an increase in hepatic triglyceride secretion rate secondary to exaggerated postprandial increases in plasma insulin concentration.

T he effects of estriol and estradiol on the plasma levels of cortisol- and thyroxine-binding globulin activity, and on the secretion rates of aldosterone and cortisol were studied in man. The metabolite estriol had no consistent or significant influence on plasma levels of the hormone-binding globulin activities; the hormone estradiol increased these binding capacities significantly, as expected. Cortisol secretion rate rose slightly after estriol but was unchanged after estradiol. Both compounds induced substantial increases in the aldosterone secretion rate of most treated subjects. The mechanism of this apparently paradoxical effect of estrogens is not clear; it is suggested that the “salt-retaining” action of estrogens is mediated in part by the rapid enhancement of aldosterone output which follows their administration in man. Balance experiments in four subjects suggest that both estradiol and estriol may induce a transient early natriuresis in man; but other mechanisms for estrogen stimulation of aldosterone secretion may be operative as well.

R eticulocytes from newborn infants with Rh isoimmune hemolytic disease actively incorporated radioactive amino acids in vitro into hemoglobins F and A. Approximately 50% of the reticulocytes appeared capable of synthesis of both of these hemoglobins within the same cell, as demonstrated by the selective elution technique of Betke and Kleihauer. An isoleucine analogue, L-O-methylthreonine, inhibited the incorporation of a variety of amino acids into hemoglobin F, without significantly affecting the synthesis of hemoglobin A. The inhibition was prevented upon concomitant addition of L-isoleucine to the medium. These observations suggest that an independent biosynthetic apparatus is present in the cell for the synthesis of each of these two hemoglobins. Because isoleucine is present only in the gamma chains of hemoglobin F, the inhibitory effect of the analogue on the synthesis of this hemoglobin must represent a selective effect on the production of gamma chains.

S era from 55 patients with systemic lupus erythematosus were studied to clarify the significance of the patterns of nuclear fluorescence observed. The sera in which the IgG fraction produced a peripheral pattern of nuclear fluorescence were found to contain complement-fixing antibodies to native DNA and to DNA-histone complexes. This correlation did not exist when complement-fixing activity was compared to the IgM nuclear patterns. Sera which contained only complement-fixing antibodies to denatured DNA and which did not react with native DNA or nucleoprotein did not produce the peripheral pattern of nuclear fluorescence. The data suggest that single strands of DNA were not the reactive groups in the nucleus responsible for the peripheral pattern. The results support the conclusion that DNA within a DNA-protein complex may be the nuclear antigen responsible for the peripheral pattern of nuclear fluorescence.Analysis of the clinical data revealed that a close correlation existed between the presence of IgG peripheral pattern, complement-fixing antibodies to DNA and histone-DNA complexes, and clinical manifestation of active disease.

H emoglobin C is less soluble than hemoglobin A in red cells, in hemolysates, and in dilute phosphate buffer. Its relative insolubility may be explained by electrostatic interactions between positively charged β6-lysyl groups and negatively charged groups on adjacent molecules. Red cells from patients with homozygous hemoglobin C (CC) disease exhibit aberrant physical properties which suggest that the cells are more rigid than normal erythrocytes. They pass through membrane filters less readily than normal red cells do, and their viscosity is higher than that of normal cells. Differences from normal cells are exaggerated if mean corpuscular hemoglobin concentration (MCHC) is increased, by suspension in hypertonic salt solution. Increased rigidity of CC cells, by accelerating their fragmentation, may be responsible for formation of microspherocytes. These small dense cells are exceptionally rigid, and probably are even more susceptible to fragmentation and sequestration. Rigidity of CC cells can be attributed to a “precrystalline” state of intracellular hemoglobin, in which crystallization does not occur, although the MCHC exceeds the solubility of hemoglobin in hemolysates.

I nhalation of aerosols of citric acid, histamine phosphate, or carbon dust, or air cooled to - 20°C or rapid respiratory maneuvers (inspiration or expiration) results in an increase in airway resistance in some patients with asthma or bronchitis. It has been shown previously in animals that stimulation of cough receptors results in bronchoconstriction through efferent cholinergic pathways. In the patients studied, the administration of atropine sulfate, which would block such pathways, abolished the bronchoconstrictor effects of all the stimuli except large doses of histamine, which may exert a direct effect on airway smooth muscle. These data suggest that sensitized cough receptors may be involved in triggering reflex airway constriction in such patients.

T he repair of cytochrome oxidase depletion during the treatment of copper deficiency was studied in the rat. The purpose of this study was to distinguish the role of new cell production from the possibly more specific role of mitochondrial turnover in determining the rate of this repair.In rats on a copper-deficient regimen until 2.5-3 months of age, activities of cytochrome oxidase expressed as per cent of control were as follows: skeletal muscle (quadratus lumborum), 18%; heart, 27%; liver, 34%; and intestinal mucosa, 34%. After 2-3 days of dietary supplementation with cupric acetate, repair of decreased cytochrome oxidase activity in intestinal mucosa is complete. Histochemical studies indicated that this repair starts in the newly differentiating cells at the base of the villus and then progresses toward the tip of the villus at a rate approximating the normal rate of migration of the mucosal cells. In liver and skeletal muscle, cytochrome oxidase activity returned to control values after 10-15 days of treatment with cupric acetate. In heart muscle, control values were approached more slowly as indicated both by activity of the enzyme and by mitochondrial difference spectra which reflect enzyme concentration.Although cytochrome oxidase repair in the intestine appeared to be limited by the rate of production of new mucosal cells, the rate of repair in liver and skeletal muscle was several times too rapid to be accounted for by known rates of new cell production. Incorporation of tritiated thymidine into DNA in these tissues in both the deficiency state and during repair indicated no major differences in new cell production compared to that of control animals. However, the time required for cytochrome oxidase repair in liver was similar to the turnover reported for other mitochondrial constituents in this tissue. The rate of cytochrome oxidase repair may therefore be more directly determined by the rate of synthesis of new mitochondrial material than by the rate of production of new cells.

I n an effort to elucidate the relation, if any, between thyroid abnormality and congenital deafness in Pendred's syndrome, an experiment was designed to study the effects of hypothyroidism on middle and inner ear hearing structures, including the auditory nerve and its central projection, in developing chick embryos. Propylthiouracil (PTU), 2 mg, was injected into the albumin of fertile chick eggs on the 10th incubation day. Single doses of L-thyroxine (range 1-100μg) were inoculated in a similar manner, either alone or with PTU. Control inocula included sterile saline or water. After hatching, each chick was examined for obvious malformations. The thyroid glands, middle and inner ear mechanisms, auditory nerve, and brainstem were studied grossly and with different histologic staining techniques. When compared to controls, chicks exposed to PTU on their 10th incubation day exhibited: increased mortality, delayed hatching, reduced size, incomplete yolk sac absorption, and death within 5 days unless exogenous thyroid hormone was provided in the first 24-48 hr after hatching. Specific, consistent, morphologic alterations were observed in their thyroid glands as well as in the sensory hair cells of the acoustic papilla and cells of the spiral ganglion of the cochlea. Our data also indicate that if 50-75 μg of L-thyroxine is given simultaneously with (or as long as 120 hr after) the PTU injection on the 10th incubation day, one cannot detect the gross defects, marked thyroid lesions, or abnormal histology in cells of the cochlea and its ganglion. A relationship between embryonic thyroid gland function and the hearing mechanism of the chick embryo is suggested.

T hree members of a family who have erythrocytosis and a new hemoglobin, designated hemoglobin Yakima, are described.The abnormal hemoglobin is characterized by the substitution of histidine for aspartic acid at residue 99 in the β-chain.Of three possible structure-function relations which would account for the increased oxygen affinity of hemoglobin Yakima, only two seem likely. These are: (a) an intrachain shift in the normal relations between the F and G helices and the heme group, or (b) an effect of the substituted side chain at a region of contact between nonpolar residues of the α- and β-chains which favors the oxyhemoglobin quarternary structure.

E rythrocytosis without clinical illness was noted in a man and his two daughters. Their blood contained approximately 62% hemoglobin A and 38% a new hemoglobin, designated hemoglobin Yakima. The oxygen affinity of whole blood from each subject was greatly increased and heme-heme interactions were impaired. At 37°C and a plasma pH of 7.40, the oxygen pressure required to produce 50% saturation of hemoglobin with oxygen was only 12 mm Hg as compared with a normal of 26 mm Hg. The high oxygen affinity of this blood is attributed to the presence of hemoglobin Yakima; and the increased oxygen affinity was shown to be characteristic of the isolated abnormal hemoglobin. A Bohr effect was present in hemoglobin Yakima.Arterial oxygen pressure, oxygen consumption, and cardiac output at rest were normal. With respect to oxygen delivery to tissues, the increased hemoglobin concentration appears to be the major compensation for the marked displacement of the oxygen-hemoglobin equilibrium curve, although other factors may contribute. The finding of high normal quantities of erythropoietin in the urine is consistent with this degree of erythrocytosis.

S era from patients with extrahepatic biliary obstruction were found to have an abnormal lipoprotein (obstructive lipoprotein) which failed to react with antibodies to normal lipoproteins of d < 1.063. Preparations of this abnormal lipoprotein made by a combination of immunoprecipitation and multiple polyanion precipitations revealed a high content of free cholesterol (26%) and phospholipids (61%) but only trace amounts of cholesterol esters and triglycerides. Protein content varied from 13% to a corrected low of 5% when ultracentrifugation was also performed. Amino acid analyses of the latter preparations resembled that of lipoproteins of d < 1.006.The reasons underlying the apparent unreactivity of the abnormal lipoprotein were explored. No evidence could be found for soluble antigen-antibody complexes of γ-globulin and the abnormal lipoprotein, nor for inhibition of antigen-antibody complex formation by serum factors. Purified preparations of obstructive lipoprotein did not react with antisera to high- or low-density lipoproteins prepared from normal sera. Moreover, rabbits immunized with the abnormal lipoproteins produced specific antibodies to this lipoprotein which reacted with a d < 1.006 lipoprotein in normal sera. All other lipoprotein fractions from normal sera were unreactive. It is not known whether this lipoprotein is abnormal by virtue of the presence of a unique peptide or because of secondary alterations in lipoprotein structure.

T he relative reactivities with native and denatured DNA of 35 lupus sera were investigated by quantitative complement fixation and precipitin studies and showed great variations. The use of purified native DNA demonstrated that, in at least 22 of these 35 sera, the anti-DNA antibodies reacted with the native form, independently of denatured contaminants. Systemic lupus sera were shown to contain three main types of DNA antibodies: those reacting only with denatured DNA, those reacting to the same extent with both forms of DNA, and those reacting preferentially with native DNA. In some instances, the latter antibodies fix complement and precipitate only with native DNA but are inhibited by the denatured form. This finding points to the importance of conformation in the antigenic structure of DNA. The simultaneous occurrence of different varieties of DNA antibodies was demonstrated in several sera. Evidence was obtained that some of these human antibodies to DNA can belong to the IgM class. Thus, DNA antibodies from systemic lupus patients differ in many respects from most of the experimentally produced antibodies capable of reacting with DNA.