Issue published December 1, 1971

V asopressin increased adenyl cyclase activity in homogenates of both inner and outer renal medulla of the rat. It also increased the concentration of cyclic 3′,5′-adenosine monophosphate (AMP) in slices of both inner and outer medulla but not in renal cortex. In the inner medulla, a concentration of prostaglandin E₁ (PGE₁), which was ineffective by itself significantly reduced the stimulation of adenyl cyclase activity and cyclic AMP concentration induced by vasopressin. These results are consistent with the hypothesis that PGE₁ can compete with vasopressin for adenyl cyclase-binding sites. However, the findings in the outer medulla suggest the situation is more complex. Although 10^-8 M PGE₁ had no effect by itself and inhibited the vasopressin-induced elevation of cyclic AMP, larger amounts of PGE₁ increased both adenyl cyclase activity and cyclic AMP levels. The maximum effect on the latter parameter was at least 6 times as great as that of maximum amounts of vasopressin.

P atients with glucose-6-phosphate dehydrogenase (G6PD) deficiency of red blood cells (RBC) may develop sudden hemolytic anemia during infection. Since phagocytizing polymorphonuclear leukocytes (PMN) are known to generate hydrogen peroxide, we explored the influence of this oxidant product of PMN on juxtaposed G6PD-deficient and normal RBC. The oxidant stress induced by phagocytosis depleted G6PD-deficient RBC of reduced glutathione (GSH) and this was associated with rapid removal of these cells from the circulation by the liver and spleen. No such effect was observed on normal RBC. Phagocytizing chronic granulomatous disease (CGD) PMN which lack hydrogen peroxide generation, failed to diminish GSH level in G6PD-deficient RBC. Thus, PMN can pose as a source of oxidant damage to G6PD-deficient RBC due to hydrogen peroxide generated during phagocytosis.

T he effects on thyroid function of an inhibitor of tyrosine dehalogenase, 3-nitro-L-tyrosine (MNT) have been investigated in rats. In preliminary studies, marked inhibition of iodotyrosine deiodination was demonstrated in rats drinking 8 mM MNT. A series of experiments was then performed in which rats received Remington low iodine diet and 8 mM MNT as drinking fluid. This regimen had the following effects, compared to the effects of a low iodine diet alone: (a) a decrease in serum protein-bound iodine, elevation of serum thyrotropin level, goiter, and growth inhibition all prevented or reversed by iodine supplements: (b) on initiation of MNT, a 2- to 3-fold increase in the rate of release of radioiodine from the thyroid and concomitant urinary excretion of large amounts of organic iodine: and (c) after 2 wk of MNT, a greatly increased rate of thyroidal uptake and release of ¹³¹I, an increase in the ratio of monoiodotyrosine-¹³¹I to diiodotyrosine-¹³¹I in thyroid proteolysates and the appearance of labeled iodotyrosines in serum.Acute administration of MNT intraperitoneally to rats on either an iodine-deficient or iodine-sufficient diet did not inhibit thyroidal uptake of ¹³¹I or alter the distribution of ¹³¹I among thyroidal iodoamino acids.It is concluded that MNT is an effective inhibitor of iodotyrosine deiodination in vivo, without other important actions on thyroid function. Thus, MNT treatment affords a model for the human dehalogenase defect. By provoking iodotyrosine secretion and consequent urinary loss of iodine, MNT can exaggerate the effects of a low iodine intake, producing goitrous hypothyroidism despite a rapid rate of iodine turnover in the thyroid.

T he kinetics of ribosomal RNA transcription and processing were assessed in chronic lymphocytic leukemia (CLL) lymphocytes during the initial phases of their delayed response to phytohemagglutinin. When compared to cultures of normal lymphocytes, CLL cultures developed normal augmentations in 45S rRNA precursor transcription and cleavage after a 1 hr incubation with PHA. However, failure to conserve 18S RNA subunits persisted in the CLL cultures. Subsequently the PHA-induced progressive rise in 45S RNA transcription became aborted and the over-all rate of RNA synthesis lagged far behind the levels attained by normal cultures incubated with PHA for 48 hr. CLL cultures responding to PHA in a delayed fashion exhibited efficient conservation of 18S RNA at 168 hr. In normal cultures, PHA-induced conservation of 18S RNA appeared to be independent of any effect on 45S ribosomal RNA precursor transcription. Therefore, the sluggish growth response of CLL lymphocytes was associated with a defect in one of the important mechanisms regulating assembly of new ribosomes.

S ulfonamides reduced substantially the ability of normal human neutrophils to kill strains of Candida albicans and Candida tropicalis, and impaired to a lesser extent their activity against Staphylococcus aureus 502A and Serratia marcescens. Sulfonamides also inhibited (a) iodination of Candida cells by normal neutrophils; (b) candidacidal activity in cell-free systems containing purified human myeloperoxidase, hydrogen peroxide, and potassium iodide; and (c) accumulation of molecular iodine in analogously constructed cell-free systems. In contrast to these effects on reactions catalyzed by myeloperoxidase, sulfonamides exerted relatively little effect on the levels of microbicidal activity manifested by human neutrophils that lacked myeloperoxidase. Sulfonamides appear to influence the function of human neutrophils predominantly by interfering with myeloperoxidase-mediated pathways. Certain basic and clinical implications of these data are discussed.

F ive healthy young men were studied during 24-30 wk of continuous bed rest. During the first 12 wk of bed rest, untreated subjects increased calcium excretion in the urine by 109 mg/day and in the feces by 147 mg/day. The rate of total body calcium loss was 0.5-0.7% per month. Losses of central calcaneus mineral, assessed by gamma ray transmission scanning, occurred at a tenfold higher rate, whereas the mineral content of the radius did not change. Changes in phosphorus balance resembled the calcium pattern, and increased excretion of nitrogen and hydroxyproline also occurred during bed rest. Upon reambulation, the subjects' calcium balance became positive in 1 month and recovery of their calcaneus mineral was complete within 10-20 wk.Treatment with potassium phosphate supplements (1327 mg P/day) entirely prevented the hypercalciuria of bed rest, but fecal calcium tended to increase. During the first 12 wk, calcium balance was slightly less negative (mean - 193 mg/day) than during bed rest without added phosphate (mean - 267 mg/day). This effect was not seen during the second 12 wk of bed rest. The patterns of magnesium excretion were similar to those of calcium. Fecal and urinary phosphorus excretions were doubled, and phosphorus balance became positive (+ 113 mg/day). Mineral loss from the central calcaneus was similar to that of untreated subjects. It is concluded that this form of phosphate supplementation reduces urinary calcium excretion but does not prevent bone loss during bed rest.

Q uantitative morphologic methods were used to measure the effects of feeding a low phosphorus diet to intact and thyroparathyroidectomized rats on several processes of bone mineralization and turnover. In severely hypophosphatemic animals, the matrix formation rate was decreased, the osteoid maturation rate was decreased, which indicated a delay in the onset of mineralization, the initial rate of mineralization was decreased, and the endosteal osteoclastic bone resorption rate was increased. In moderately hypophosphatemic animals, there was a substantial increase in bone resorption but no change in formation or in mineralization. The increase in endosteal bone resorption was due to an increase in the linear rate of bone resorption and particularly to an increase in the length of the endosteal resorbing surface. The magnitude of the increase in bone resorption was similar in thyroparathyroidectomized and intact rats indicating that neither parathyroid hormone nor calcitonin is involved in this change. This, together with the finding that there was a strong negative correlation (r = -0.99) between the per cent endosteal resorbing surface and the serum phosphorus, supports the view that the increased resorption was due to hypophosphatemia. This inverse relationship between endosteal resorbing surface and serum phosphorus appeared to hold for values of serum phosphorus above normal. The resorptive response to hypophosphatemia, as previously shown for the resorptive response to excess endogenous parathyroid hormone, was partially inhibited by vitamin D deficiency. Increased resorption occurred at levels of serum phosphorus where no changes were observed in bone formation, mineralization, or growth, suggesting that this resorptive response functions as a homeostatic mechanism to maintain serum and intracellular phosphorus concentrations.

T he fecal elimination and enterohepatic circulation of bile acid was studied in 11 patients. 10 patients with varying degrees of ileal disease or resection and 1 patient with pancreatic insufficiency and no ileal disease. A new technique was employed which involved the nearly simultaneous administration of cholic acid-¹⁴C and a nonabsorbable marker. ⁵¹CrCl₃. Each individual stool specimen was collected for 36-96 hr and analyzed separately. Assay of the radioactivity of each isotope allowed the accurate determination of an excretion rate for both cholic acid and ⁵¹Cr. The difference between these rates was used to calculate an absorption coefficient for cholic acid. In addition, bile acid concentration measured by the steroid dehydrogenase technique, and the water content of each stool was determined.The patients were divided into groups depending upon how much small intestine was resected or diseased: six patients with less than 100 cm of ileal resection or disease (group A), and five patients with more than 100 cm of ileal disease or resection (group B). The ⁵¹Cr excretion rate was similar in the two groups, but cholic acid-²⁴C excretion rates were significantly more rapid in group B than in group A. The cholic acid absorption coefficient was essentially normal in the patient with pancreatic insufficiency, moderately decreased in group A patients, and extremely low or zero in group B patients. It was inversely related to the length of intestine diseased or resected.Daily fecal bile acid excretion was normal to twice normal in group A patients and 2-8 times normal in group B patients. In all patients with ileal disease or resection, there was a direct correlation between fecal bile acid, fecal mass, and fecal water. Each millimole of additional bile acid in the stool was associated with an increase in stool water of 11 moles (P < 0.01).These studies show that the kinetics of bile acids in the enterohepatic circulation can be accurately studied in patients with extensive ileal resection. The regular relationship between fecal bile acid and fecal mass and water suggests, but does not prove, a critical role of bile acid in determining stool water.

U sing a new in vitro method of measuring the chemotaxis of polymorphonuclear leukocytes from peripheral blood, a chemotactic index has been calculated. The mean chemotactic index of 320 in 24 patients with definite rheumatoid arthritis, was significantly less (P < 0.0005) than the mean of 555 in 24 normal controls matched for age and sex.The mean chemotactic index of 435 in eight patients with juvenile rheumatoid arthritis was also significantly less (P < 0.01) than that of 553 in similarly matched controls.The chemotactic index could not be correlated with age, sex, disease activity, drugs used in treatment, latex titer, immunoglobulin levels, or protein coating on the cells. However, there was a correlation between the chemotactic index and the serum complement B_1e/B_1a value (P < 0.01) in 17 patients with adult onset rheumatoid arthritis. Although the serum complement B_1e/B_1a values were within the normal range, the lowest chemotactic indices were associated with the lowest complement values.The chemotactic indices in three patients with severe connective tissue disease (seropositive rheumatoid arthritis, systemic lupus erythematosus, and polymyositis) returned to normal after 5 days' treatment with 60 mg of prednisolone per day. Incubation of the cells from patients with rheumatoid arthritis with hydrocortisone in vitro failed to alter the chemotactic indices.Prior incubation of normal cells with purified rheumatoid factor complexes, rheumatoid serum, or macromolecules of iron dextran impaired their chemotaxis. It is suggested that phagocytosis of complexes in vivo is a possible mechanism by which the chemotaxis of the polymorphonuclear leukocytes of patients with rheumatoid arthritis is impaired.This impairment in chemotaxis may explain the increased incidence of bacterial infection, both during life and as a cause of death in these patients.

I n order to evaluate separately changes in vascular tone occurring in arteries and veins, we measured pulmonary capillary red blood cell (RBC) concentration under zone II (waterfall) conditions in isolated dog lungs rapidly frozen with Freon 12. The lungs were frozen while being perfused from artery to vein and from vein to artery breathing normal and hypoxic gas mixtures and during infusions of serotonin and histamine. Changes in capillary RBC concentration which occurred during the experimental conditions indicated an alteration in vascular resistance upstream from the capillaries. Alveolar hypoxia caused a significant decrease in capillary RBC concentration during forward perfusion, but no change from the control values during reverse perfusion. Serotonin infusion caused a decrease in RBC concentration during forward perfusion comparable with that of hypoxia and a small but significant decrease during reverse perfusion. Histamine infusion caused no change in RBC concentration from control values during forward perfusion, but a large decrease during reverse perfusion. We conclude that vasoconstriction occurs (a) exclusively in arteries during alveolar hypoxia, (b) predominantly in arteries but to a lesser extent in veins during serotonin infusion, and (c) exclusively in veins during histamine infusion.

I mmunologic responses to bacteriophage ϕX 174 were studied in 26 patients with immunodeficiency diseases. In eight cases of infantile X-linked agammaglobulinemia, there was prolonged circulation of phage and no detectable antibody response. The remaining 18 patients cleared phage normally and produced antibodies. 10 of these patients made only IgM antibody in spite of repeated immunization; all of these have recurrent respiratory tract infections and require treatment with gamma globulin and antibiotics. Eight patients made both IgM and IgG antibody; they experience either milder or no infections, and only one requires treatment with gamma globulin.Prolonged circulation of bacteriophage ϕX 174 and the absence of a detectable antibody response appear to be distinguishing characteristics of X-linked agammaglobulinemia if severe combined immunodeficiency can be excluded.

T he effects of extracellular fluid volume expansion on intestinal transport of salts and water were studied in dogs by perfusing loops of bowel in vivo. Saline loading caused depression of duodenal and jejunal absorption with net secretion of salt and water into the lumen. Studies of unidirectional transport of ²²Na⁺ revealed that the negative net sodium flux was due primarily, and perhaps exclusively, to increased serosal to mucosal transport, for mucosal to serosal sodium transport was not changed during volume expansion. Net transport of water and potassium paralleled net sodium flux. Administration of deoxycorticosterone did not affect the intestinal response to saline loading. Hemodilution, accomplished by equilibrating the dogs' blood with a reservoir of saline, did not affect intestinal absorption, but isotonic, iso-oncotic expansion of the extracellular fluid produced by reinfusing the saline-blood mixture from the reservoir resulted in negative net transport of water, sodium, and potassium by the duodenum. It is suggested that the small bowel is capable of secreting salts and water through intercellular spaces, and that this process is stimulated by extracellular fluid volume expansion.

T he mechanism by which agents that inhibit bacterial cell wall synthesis produce a synergistic effect against enterococci when combined with aminoglycoside antibiotics has not been elucidated. Using ¹⁴C-labeled streptomycin, it could be shown that uptake of this aminoglycoside antibiotic was markedly enhanced in enterococci growing in the presence of penicillin or other agents which inhibit the synthesis of bacterial cell walls. There was no enhancement of streptomycin uptake when the cells were incubated with antibiotics which primarily affect the bacterial cell membrane or inhibit protein synthesis. Increased streptomycin uptake was produced by penicillin only in actively growing bacteria. These observations are consistent with the hypothesis that enterococci exhibit a natural barrier to the entry of streptomycin which can be overcome by agents which inhibit cell wall synthesis, thus producing a synergistic effect.

T he effects of ouabain (G-strophanthin) 20 μg/kg, on left ventricular (LV) pressure (P), diameter (D), velocity of contraction (dD/dt), and dP/dt were studied in conscious dogs instrumented with ultrasonic diameter gauges and miniature pressure gauges. The effects of ouabain were compared on separate occasions in the same dogs after cardiac depression with propranolol, 3.0 mg/kg, and also after general anesthesia with Na pentobarbital, 30 mg/kg. Maximal pressor effects were observed in the first 10 min, but maximal effects on the contractile state occurred at 30 min after ouabain. At this time, in conscious dogs, ouabain had increased LV isolength systolic pressure by 5%, LV isolength velocity by only 9%, and LV (dP/dt)/P by 21%, while end systolic diameter (ESD) decreased slightly and end diastolic diameter (EDD) and heart rate (HR) were unchanged. After anesthesia, ouabain increased LV systolic pressure by 8%, velocity 32%, (dP/dt)/P by 47%, and ESD decreased by 1.2 mm while EDD rose slightly and HR fell by 26 beats/min. Returning HR to control with atrial pacing decreased EDD 0.9 mm below control. After cardiac depression with propranolol, ouabain caused responses similar to those observed in the anesthetized dogs. Thus, the cardiac glycoside was found to exert only minor inotropic effects on the nonfailing heart of conscious dogs but far more striking inotropic responses in the anesthetized state.

P lasma insulin dynamics were evaluated in 10 patients with primary hyperparathyroidism before and after parathyroidectomy and correction of hypercalcemia. Before surgery fasting plasma insulin concentrations and insulin responses to administered glucose, tolbutamide, and glucagon were significantly greater than postoperative values. Hyperinsulinemia was not associated with altered glucose curves during glucose or glucagon tolerance tests, but a relatively greater insulin response to tolbutamide resulted in an increased hypoglycemic effect following its administration. The glucose-lowering action of intravenous insulin was slightly impaired before treatment. Intramuscular injections of parathormone to six normal men for 8 days induced mild hypercalcemia and hypophosphatemia and reproduced augmented plasma insulin responses to oral glucose and intravenous tolbutamide. 4-hr intravenous infusions of calcium to another group of six normal men raised serum calcium concentrations above 11 mg/100 ml. This did not alter glucose or insulin curves during oral glucose tolerance but markedly accentuated insulin responses to tolbutamide and potentiated its hypoglycemic effect. When highly purified parathormone was incubated with isolated pancreatic islets of male rats, glucose-stimulated insulin secretion was unaffected.These findings suggest that chronic hypercalcemia of hyperparathyroidism sustains a form of endogenous insulin resistance that necessitates augmented insulin secretion to maintain plasma glucose homeostasis. This state is insufficient to oppose tolbutamide-induced hypoglycemia because of an additional direct, selective enhancement of hypercalcemia on pancreatic beta cell responsiveness to the sulfonylurea. The possible direct role of parathormone in these events has not been established.

T he effect of phenobarbital on bilirubin excretion was studied in rats with different capacities for bilirubin conjugation. Drug treatment induced substantial increases in bilirubin UDP-glucuronyl transferase activity in the liver of both normal and heterozygous Gunn rats, but not homozygous Gunn rats in which enzyme activity is completely absent. However, enhancement of bilirubin excretion in vivo was observed only in heterozygous Gunn rats. In these animals the maximum capacity to excrete bilirubin into bile (T_max), like the activity of the conjugating enzyme, was half normal; phenobarbital caused an increase in T_max to levels characteristic of normal animals, with a twofold rise in the excretion of conjugated pigment. This appeared to be largely unrelated to enhancement of bile flow, and there was no stimulation of alternate pathways of bilirubin excretion.Conjugated bilirubin was consistently recovered from the plasma and urine of both untreated normal and heterozygous Gunn rats infused with unconjugated pigment. The quantities thus recovered comprised a similar fraction of the total pigment conjugated in both types of animal. Moreover, there were linear correlations between T_max and both the rate of bile flow and the activity of the conjugating enzyme over the range of values represented by control rats of both types. These findings suggest that the process by which conjugated bilirubin is secreted into the bile is closely related to conjugation and limits the final excretory rate at different levels of pigment excretion. The phenobarbital effect uniquely observed in heterozygous Gunn rats appears to be mediated primarily by enhancement of the limited capacity for bilirubin conjugation with an associated rise in functional secretory capacity.

T his study was designed to develop a method for quantitative assessment of infarct size in the conscious animal based on serial changes of serum creatine phosphokinase (CPK) activity. From 11 experiments in which myocardial CPK was injected intravenously in conscious dogs, the average CPK distribution space and average CPK fractional disappearance rate from serum were found to be 11.4% of body weight and 0.48% min respectively. In other experiments, myocardial infarction was produced in 22 conscious dogs by constriction of a left coronary artery snare and serum CPK activity was determined at frequent intervals for 24 hr. Since myocardial CPK depletion reflects infarct size, infarct size was determined directly by analysis of myocardial CPK content in the same animals 24 hr after coronary artery occlusion. CPK released from the infarct was determined from observed changes in serum CPK activity analyzed according to a model taking into account the fraction of CPK released from an infarct and the rates of appearance and disappearance of CPK activity from serum. Infarct size was calculated on the basis of observed changes in serum CPK and compared to infarct size determined directly by analysis of myocardial CPK depletion. Agreement was close and results from all experiments fit the equation: [infarct size (g) determined from serum CPK] = 1.13 × [infarct size (g) determined from myocardial CPK] - 1.3, r = 0.96, n = 22. The method described is useful for accurate assessment of infarct size in the conscious animal and for detection of modification of infarct size produced by pharmacologic interventions.

P revious studies have demonstrated that considerable amounts of parenterally administered cardiac glycosides are excreted in the bile and reabsorbed across the intestinal mucosa in several species. It is currently believed that the more prolonged action of nonpolar digitalis glycosides is due to their retention and recycling in the enterohepatic circulation. This report describes studies carried out to evaluate the effects of pharmacologic interruption of this enterohepatic cycle with the intraluminal sequestering agent cholestyramine.Cholestyramine was found to bind substantial quantities of digitoxin-³H and digoxin-³H in vitro and this binding was only modestly inhibited by the presence of bile. Administration of cholestyramine to rats by intragastric catheter before the subcutaneous injection of the LD₁₀₀ dose of digitoxin (10 mg/kg) resulted in a 70% survival rate. Further, oral administration of cholestyramine to rats before the subcutaneous injection of digitoxin-³H resulted in accelerated fecal excretion of radioactivity and lower levels of digitoxin-³H and metabolites in brain tissue compared to controls. Similarly, pretreatment of guinea pigs with cholestyramine orally before the injection of digitoxin in dosages of 10.0 and 4.0 mg/kg resulted in a 25 and 70% survival rate respectively as compared to survival rates of 0 and 30% in control animals. Cholestyramine pretreatment of guinea pigs was also accompanied by lower levels of digitoxin-³H and metabolites in heart and liver 90 min after injection of digitoxin-³H. Cholestyramine therapy did not result in significant changes in serum potassium levels excluding the possibility that drug-induced hyperkalemia might have affected the cardiac uptake of digitoxin.The data obtained in this study indicate that cholestyramine treatment affords a significant degree of protection against lethal digitoxin intoxication in rats and guinea pigs. It is suggested that cholestyramine binds appreciable amounts of digitoxin in the intestinal lumen resulting in reduced reabsorption, increased fecal excretion, and lower tissue levels of glycoside in critical organs. The protective effects of cholestyramine appear to be mediated by interruption of the enterohepatic circulation of digitoxin.

P revious studies of digitalis glycoside metabolism and excretion have indicated that these compounds undergo a significant enterohepatic cycle in some species. It has been suggested that the existence of such a cycle in man contributes to the prolonged action of certain cardiac glycosides. Previous studies have demonstrated that cholestyramine binds digitoxin and digoxin in vitro and accelerates the metabolic disposition of digitoxin in rats and guinea pigs, presumably by interrupting the enterohepatic circulation.In order to assess the role of the enterohepatic circulation in the metabolism of digitalis glycosides in humans, maintenance doses of cholestyramine were administered to 7 of 15 normal human subjects beginning 8 hr after digitalization with 1.2 mg of digitoxin-³H. All subjects had frequent measurements of serum radioactivity, left ventricular ejection time (LVET), and electromechanical systole (QS₂), the latter recorded as the interval from onset of Q wave to first major component of second heart sound. Measurement of the LVET and QS₂ intervals affords a sensitive index of the cardiac response to digitalis. In addition, chloroform extraction of serum was performed to separate unchanged digitoxin and active metabolites from cardioinactive metabolites of digitoxin. Cholestyramine treatment resulted in reduction in half-life to total serum radioactivity from 11.5 to 6.6 days, and in chloroform-extractable radioactivity from 6.0 to 4.5 days, as compared to controls. In addition, cholestyramine treatment was accompanied by more rapid return to base line values of digitoxin-induced changes in the LVET and QS₂ intervals. A significant positive correlation was found between QS₂ values and chloroform-extractable radioactivity, the latter reflecting unchanged digitoxin-H³ (r=0.64; P=<0.01).The results indicate that administration of cholestyramine to digitalized human subjects accelerates the metabolic disposition of digitoxin and abbreviates the physiologic response to the glycoside. This effect is presumably mediated by interruption of the enterohepatic circulation of digitoxin by cholestyramine.

I n vivo and in vitro studies of granulocyte chemotaxis were performed in three patients with the Chediak-Higashi syndrome. Rebuck skin windows showed a decreased accumulation of leukocytes at an inflammatory site. Studies in Boyden chambers documented a cellular defect in granulocyte chemotaxis. The chemotactic response of Chediak-Higashi cells by this technique averaged approximately 40% of normal and was consistently reduced using several different chemotactic stimuli. This deficit was magnified by shortening the chamber incubation time or by decreasing the pore size of the micropore filter and was independent of granulocytopenia. No abnormalities of passive motility, adhesiveness, viability, or pH optimum for migration were found in these cells. Chediak-Higashi serum contained no inhibitors of chemotaxis and was capable of generating normal amounts of chemotactic factors with the exception of one patient with the accelerated phase of the disease. Heterozygotes for the Chediak-Higashi trait had normal chemotactic function. This cellular defect in chemotaxis may contribute to the marked susceptibility to pyogenic infections which is so characteristic of patients with the Chediak-Higashi syndrome.

T he effect of atrial contraction on left ventricular function in six patients with varying degrees of mitral stenosis was determined by utilizing the pressure gradient technique to measure instantaneous aortic blood flow and pressure. Aortic flow was measured as ventricular rate was controlled by right ventricular pacing to create A-V (atrioventricular) dissociation at varying rates (90-150 beats/min). At each heart rate, beats with preceding P waves, effective atrial systole, were grouped according to the duration of the P-R interval. Beats without P waves served as controls. There was always a significant increase in stroke volume, created by effective atrial systole, but the P-R interval at which it took place was different for each patient. There was no difference in the stroke volume for beats preceded by P waves having a P-R interval within the range of 0.05-0.20 sec. These beats were grouped for each patient, subjected to regression analysis, and compared to control beats. The absolute and percent change created by effective atrial systole was inversely proportional to the severity of the disease as determined by mitral valve orifice size.Effective atrial systole plays less of a role in augmenting left ventricular function in patients with mitral stenosis than in patients with normal valves.

A radioimmunoassay for monkey placental lactogen (MPL) was developed to study the factors controlling the secretion of MPL. The sensitivity of the assay was 1 ng MPL per ml. Human and monkey growth hormone, and human placental lactogen (HPL) showed minimal cross-reactions with MPL. Maternal MPL concentrations as measured in 40 rhesus monkeys increased progressively throughout pregnancy to a mean of 5000 ng/ml at term while umbilical vein MPL was less than 50 ng/ml. After term delivery maternal MPL concentrations decreased rapidly with a t½ of 20 min.After fetectomy but with retention of the placenta, MPL concentrations decreased by 25% reaching a plateau over a 6 hr period. Experimental abruption of the secondary placenta within 1 hr produced a 50% decrease in MPL concentration. After ligation of the fetal vessels supplying the secondary placental disc, MPL increased transiently and then decreased to levels significantly below those of the control period.These studies suggest MPL secretion is not directly controlled by the fetus but is sensitive to changes in placental blood flow. The pregnant rhesus monkey serves as a useful model for investigating factors which may regulate HPL secretion because of the close similarity between MPL and HPL secretion.

H ypocalcemia in the hypomagnesemic state in man is usually attributed to refractoriness of end-organs to the calcemic action of parathyroid hormone. We studied the responsiveness of end-organs to bovine parathyroid extract (PTE) in magnesium-depleted and control dogs by the following three methods after thyroparathyroidectomy: (a) assessment of the calcemic response to a set dose of PTE (0.3 U/kg per hr); (b) assessment of PTE dose required to attain normocalcemia; (c) evaluation of regression lines of plasma calcium concentration on PTE dose. The calcemic response of magnesium-depleted thyroparathyroidectomized puppies to a set dose of PTE was similar to that of control puppies. There was no significant difference in the dose of PTE required to attain normocalcemia nor in the dose-response relations between the plasma calcium concentration and the PTE dose. In a group of magnesium-depleted puppies with intact thyroid and parathyroid glands, the dose of PTE required to attain normocalcemia was similar to that required in thyroparathyroidectomized animals, indicating calcitonin was not a factor contributing to hypocalcemia. We conclude that hypocalcemia in magnesium-depleted puppies is not due to refractoriness of end-organs to the calcium-mobilizing action of parathyroid hormone. Defective synthesis or diminished secretion of parathyroid hormone is suggested as an explanation.

S erum triiodothyronine (T₃) has been measured by radioimmunoassay and corroborated by analysis of the identical samples with a previously described gas-liquid chromatographic technique. Special features of the radioimmunoassay procedure which permit determinations in unextracted serum include the use of a T₃-free serum preparation for the construction of the standard curve and of tetrachlorothyronine to inhibit binding of T₃ to thyroxine-binding globulin.T₃ values by radioimmunoassay were 138 ±23 ng/100 ml (mean ±SD) in 82 normal subjects, 62 ±9 ng/100 ml in 45 hypothyroid patients, and 494 ±265 ng/100 ml in 60 patients with toxic diffuse goiter. In the hypothyroid group, the range was similar in patients with both primary and secondary hypothyroidism. There was no overlap between the three thyroidal states. Elevated T₃ levels were seen in 40 cases that appeared clinically hyperthyroid but had normal serum thyroxine (T₃) determinations, a syndrome we have called T₃ toxicosis. Values obtained with radioimmunoassay agreed closely with those we had previously found by gas-liquid chromatography which were 68 ±2 ng/100 ml in hypothyroidism, 137 ±23 ng/100 ml in normal subjects, and 510 ±131 ng/100 ml in untreated toxic diffuse goiter.Since T₃ is very potent and its level varies in different clinical states, accurate T₃ measurements are required to assess a patient's thyroid status properly. The radioimmunoassay for T₃ appears to be sufficiently sensitive, precise, and simple to permit its routine clinical application for this purpose.

T o clarify the controversial renal action of calcitonin (CT) and a possible interrelationship between CT and parathyroid hormone, eight patients with untreated surgical hypoparathyroidism were studied. Various calcitonins, i.e. extracted porcine, synthetic porcine, synthetic human, and synthetic salmon CT in doses of 150 Medical Research Council U or 1.5 mg were infused over a 3 hr period. Subsequently, six of the same subjects received 500 USP U parathyroid extract (PTE) (Eli Lilly & Co., Indianapolis, Ind.) in 3 hr and later a combination of CT and PTE. In addition, two patients were given an infusion of ammonium phosphate with the aim of producting a phosphaturia of comparable degree as seen after CT and PTE, thus differentiating hormonal from nonhormonal influences on cation excretion. A protocol of serial clearance (C) studies using the patients as their own controls was followed. Serum and urinary inorganic phosphate (P), calcium (Ca), magnesium (Mg), sodium (Na), potassium (K), and creatinine (Cr) were determined and the clearance values calculated.All CT peptides caused a uniform, immediate and significant increase of C_P, C_Na, C_K, C_Ca, and C_Mg, PTE evoked a rise of C_P, C_Na, and C_K, but C_Ca and C_Mg were reduced, the Ca and Na figures being not statistically significant. The administration of both CT and PTE resulted in a summation of individual hormone effects on Ca and Mg excretion. Phosphate infusion on the other hand induced an isolated phosphaturia but no concomitant changes of the urinary cations.The hypoparathyroid data demonstrate that calcitonin enhances urinary elimination of P, Na, K, Ca, and Mg independently of parathyroid action, CT and PTE act qualitatively similarly on P, Na, and K excretion, while an antagonism seems to exist for the renal handling of Ca and Mg.

A rterial concentration and net exchange across the leg and splanchnic bed of 19 amino acids were determined in healthy, postabsorptive subjects in the resting state and after 10 and 40 min of exercise on a bicycle ergometer at work intensities of 400, 800, and 1200 kg-m/min. Arterio-portal venous differences were measured in five subjects undergoing elective cholecystectomy.In the resting state significant net release from the leg was noted for 13 amino acids, and significant splanchnic uptake was observed for 10 amino acids. Peripheral release and splanchnic uptake of alanine exceeded that of all other amino acids, accounting for 35-40% of total net amino acid exchange. Alanine and other amino acids were released in small amounts (relative to net splanchnic uptake) by the extrahepatic splanchnic tissues drained by the portal vein.During exercise arterial ananine rose 20-25% with mild exertion and 60-96% at the heavier work loads. Both at rest and during exercise a direct correlation was observed between arterial alanine and arterial pyruvate levels. Net amino acid release across the exercising leg was consistently observed at all levels of work intensity only for alanine. Estimated leg alanine output increased above resting levels in proportion to the work load. Splanchnic alanine uptake during exercise exceeded that of all other amino acids and increased by 15-20% during mild and moderate exercise, primarily as a consequence of augmented fractional extraction of alanine. For all other amino acids, there was no change in arterial concentration during mild exercise. At heavier work loads, increases of 8-35% were noted for isoleucine, leucine, methionine, tyrosine, and phenylalanine, which were attributable to altered splanchnic exchange rather than augmented peripheral release.The data suggest that (a) synthesis of alanine in muscle, presumably by transamination of glucose-derived pyruvate, is increased in exercise probably as a consequence of increased availability of pyruvate and amino groups; (b) circulating alanine serves an important carrier function in the transport of amino groups from peripheral muscle to the liver, particularly during exercise; (c) a glucose-alanine cycle exists whereby alanine, synthesized in muscle, is taken up by the liver and its glucose-derived carbon skeleton is reconverted to glucose.

A rterial concentrations and net substrate exchange across the leg and splanchnic vascular bed were determined for glucose, lactate, pyruvate, and glycerol in healthy postabsorptive subjects at rest and during 40 min of exercise on a bicycle ergometer at work intensities of 400, 800, and 1200 kg-m/min.Rising arterial glucose levels and small decreases in plasma insulin concentrations were found during heavy exercise. Significant arterial-femoral venous differences for glucose were demonstrated both at rest and during exercise, their magnitude increasing with work intensity as well as duration of the exercise performed. Estimated glucose uptake by the leg increased 7-fold after 40 min of light exercise and 10- to 20-fold at moderate to heavy exercise. Blood glucose uptake could at this time account for 28-37% of total substrate oxidation by leg muscle and 75-89% of the estimated carbohydrate oxidation.Splanchnic glucose production increased progressively during exercise reaching levels 3 to 5-fold above resting values at the heavy work loads. Close agreement was observed between estimates of total glucose turnover during exercise based on leg glucose uptake and splanchnic glucose production. Hepatic gluconeogenesis—estimated from splanchnic removal of lactate, pyruvate, glycerol, and glycogenic amino acids—could supply a maximum of 25% of the resting hepatic glucose production but could account for only 6-11% of splanchnic glucose production after 40 min of moderate to heavy exercise.It is concluded that: (a) blood glucose becomes an increasingly important substrate for muscle oxidation during prolonged exercise of this type: (b) peripheral glucose utilization increases in exercise despite a reduction in circulating insulin levels: (c) increased hepatic output of glucose, primarily by means of augmented glycogenolysis, contributes to blood glucose homeostasis in exercise and provides an important source of substrate for exercising muscle.

A new immunoglobulin A abnormality, absence of assembly of α-chain and light-chain, was found in an adult female suffering from recurrent upper respiratory infection and tonsillitis since childhood, but otherwise healthy. The IgA abnormality was manifest in her serum by the presence of free α-chains, in her saliva by the presence of α-chains bound to secretory piece, and in her urine by the presence of free α-chains and free light-chains. The serum IgG and IgM were found to be complete, containing both heavy-chains and light-chains.Evidence for this immunoglobulin A abnormality was also found in the proposita's mother and elder son, demonstrating it to be a hereditary disorder.Studies performed with patient's tonsillar cells in short-term culture, using amino acids-¹⁴C, revealed synthesis and secretion of both free α-chains and free light-chains, in addition to synthesis and secretion of normally assembled IgG and IgM.

N oradrenaline synthesis and metabolism of dopamine was evaluated in three patients with familial dysautonomia and compared with that of six normal subjects. Each patient and subject was infused with 104.8 μCi of dopamine-2-¹⁴C dissolved in 1000 ml of physiological saline. The urine was collected during the infusion period and at intervals thereafter. Using a specially designed flow monitor system, the various biosynthetic and metabolic products of dopamine were separated, identified, and their radioactivity measured. The results indicate that in familial dysautonomia the synthesis of noradrenaline is significantly decreased; this is reflected by a decrease in recovery of radioactive noradrenaline as well as various metabolic products of noradrenaline, i.e. 3-methoxy-4-hydroxymandelic acid (MOMA), normetadrenaline, and normetadrenaline conjugate. Concomitant with this decrease in noradrenaline synthesis, there was a shift towards dopamine metabolism as reflected by an increase in the recovery of primary and secondary dopamine metabolites; 3,4-dihydroxyphenylacetic acid (DOPAC), 3-methoxy-4-hydroxyphenylacetic acid (HVA), 3-methoxytyrosine, and respective conjugates, etc. Whereas all dysautonomic patients showed the same general metabolic pattern as was expected, they varied in degree.

A fter incubation with polystyrene latex beads for 5 min. the cyclic 3′,5′-adenosine monophosphate (cyclic AMP) content of human peripheral blood leukocyte suspensions was increased severalfold. Preparations enriched in mononuclear cells and containing only 0-20% polymorphonuclear leukocytes (PMN) and no visible platelets exhibited a quantitatively similar response. Purified fractions of cells containing 85-90% PMN responded to polystyrene beads with a much smaller increase in cyclic AMP content. Phagocytosis of paraffin oil emulsion in the unfractionated mixed human leukocyte preparation was associated with little or no change in cyclic AMP levels. There was no change in cyclic AMP content of rabbit alveolar macrophages or guinea pig PMN during phagocytosis of polystyrene beads. All of these observations are consistent with the view that particle uptake per se does not increase cyclic AMP levels in phagocytic cells. It seems probable that the increase in cyclic AMP concentration that results when unfractionated human blood leukocytes are incubated with polystyrene beads occurs in cells other than PMN.

T ransmembrane potential difference (pd) was studied in isolated perfused segments of rabbit proximal convoluted tubules. At perfusion flow rates above 10 nl/min the pd was -5.80 ±0.3 mv (lumen negative) when perfusing with isosmolal ultrafiltrate of same rabbit serum as the bath. That this pd is generated by transport activity of the tubule is supported by three separate observations: (a) pd reversibly decreased with cooling from 37°C to 25°C; (b) pd decreased when 10^-5 M ouabain was added to the bath and reversed to control levels when ouabain was removed; and (c) heating to 47°C irreversibly decreased pd to zero. The magnitude of the pd was related to perfusion flow rate at slower rates than 10 nl/min. A decrease in flow rate was associated with a decrease in pd. The tubular geometry and transmembrane hydrostatic pressure were ruled out as the mediating factors governing the magnitude of observed pd.

A lthough congenital adrenal hyperplasia due to 3β-hydroxysteroid dehydrogenase deficiency generally reveals a predominance of Δ⁵-3β-hydroxysteroids, on occasion substantial quantities of pregnanetriol have been found as well. It appears that the latter steroid more often occurs in the subjects who have survived beyond infancy. The use of the measurement of pregnanetriol alone may therefore not be relied upon as a sole determinant of the specific form of defective steroidal biogenesis. It is more characteristic of the 21-hydroxylase deficiency. However when both Δ⁵-pregnenetriol and pregnanetriol are measured the ratio of the former to the latter is always considerably below 1.0 in 21-hydroxylase deficiency and always above 1.0 in 3β-hydroxysteroid dehydrogenase. Furthermore, 11-ketopregnanetriol has been found only in the urine of subjects with the 21-hydroxylase deficiency. Thus, these two forms of defective steroidal biogenesis may be distinguished by the measurement of these three urinary steroidal metabolites.

F unctional messenger RNA for human hemoglobin synthesis was prepared from reticulocyte lysates of patients with homozygous beta thalassemia and sickle cell anemia. The messenger RNA stimulated the synthesis of human globin chains by a cell-free system derived from Krebs mouse ascites cells. In the presence of beta thalassemia messenger RNA, the system synthesized much less beta chain than alpha chain whereas in the presence of sickle cell anemia messenger RNA, nearly equal amounts of alpha and beta chains were synthesized. The beta/alpha synthetic ratios obtained in the cell-free system were similar to those obtained by incubating intact beta thalassemia and sickle cell anemia reticulocytes in the presence of radioactive leucine. The experiments provide direct evidence of a defect in messenger RNA for beta chains as a cause for the decreased synthesis of beta chains observed in beta thalassemia.