Endogenous plasma growth hormone concentrations were measured in 23 children who were receiving daily corticosteroid therapy and in 10 control asthmatic children who had not received steroids for at least 8 months. The growth hormone concentrations were similar in the two groups of patients both during the fasting state and after insulin-induced hypoglycemia. 12 children, who were studied while receiving a large dose of prednisone and again 2 wk after steroid withdrawal, also showed no change in growth hormone concentration in relation to corticosteroid therapy. These findings suggest that deficiency of growth hormone is not the major mechanism responsible for the dwarfism of corticosteroid-treated children.
Helen G. Morris, Jacqueline R. Jorgensen, Shirley A. Jenkins
The effects of administered human growth hormone (HGH) were evaluated in dwarfed, prepubertal children who were receiving long-term corticosteroid therapy for a chronic disease. During 11 complete metabolic balance studies, the eight corticosteroid-treated children demonstrated impaired response to large doses of HGH with minimal nitrogen and no phosphorus retention. In contrast, two hypopituitary subjects and two asthmatic children not receiving corticosteroid responded to the same preparations of HGH with nitrogen, potassium, and phosphorus retention. Six corticosteroid-treated children were given large doses of HGH (40-120 mg/wk for 4 to 8 months and showed no improvement in their retarded rate of growth, whereas the hypopituitary subjects showed accelerated growth during administration of 10-15 mg of HGH/wk. It is concluded that dwarfism in steroid-treated children results from corticosteroid-induced antagonism of the effects of HGH at the peripheral tissue level.
Helen G. Morris, Jacqueline R. Jorgensen, Harold Elrick, Richard E. Goldsmith
A method is described for the quantitation of megakaryocytes. In this technic bone marrow sections obtained at the time of maximal marrow uptake of previously injected plasma bound 59Fe are counted for radioactivity as well as the number of megakaryocytes and nucleated red cells. We calculated the total megakaryocytes by relating the ratio of megakaryocyte: radioactivity with the total marrow 59Fe activity at the time of sampling. A multiple counting correction factor which is related to cell diameter and section thickness is required for quantitation of cell numbers from section material. The normal megakaryocyte number for the rat is 11.0 × 106/kg, and 6.1 × 106/kg for man.
Lawrence A. Harker
Measurements of megakaryocyte number, volume, nuclear number, and cytoplasmic granulation were compared in animals with induced thrombocytosis, normal platelet counts, and induced thrombocytopenia for 4- and 10-day periods. Thrombopoiesis, as measured by megakaryocytic mass and 35S incorporation into platelets, appeared to be regulated by the demand for circulating platelets. In addition, two mechanisms were operative in the alteration of thrombopoiesis. The first mechanism involved the regulation of endomitosis in the immature megakaryocytes which in turn determined the subsequent volume of platelet-producing cytoplasm. The second mechanism regulated the number of megakaryocytes formed from the precursor, “stem cell,” compartment.
Laurence A. Harker
The adherence to collagen of rabbit platelets labeled in vivo with 35SO4= has been studied both in vitro and in vivo. The young platelets are labeled with 35SO4= 2-3 days after administration of the isotope to the animals. We exposed platelet-rich plasma (ethylenediamine-tetraacetate, EDTA, as anticoagulant), prepared from blood taken from rabbits 54 hr after giving the 35SO4=, to collagen in vitro. There was a fall in the specific radioactivity of the nonadherent platelets which indicated a selective adhesion of young platelets to the collagen. In experiments designed to have most of the 35S label in the oldest platelets it was found that exposure of plasma containing these platelets to collagen resulted in an increase in the specific radioactivity of the nonadherent platelets. Similar observations were obtained when glycine-14C was used as a platelet label. However, when DF32P (di-isopropyl phosphorofluoridate-32P), which is thought to label platelets of all ages equally, was used, the adherence of platelets to collagen did not result in any changes in the specific activity of the nonadherent platelets. In in vivo studies in which we infused a collagen suspension into rabbits 54 hr after giving 35SO4= we found that the specific radioactivity of the platelets remaining in the circulation fell. This did not occur when we infused the collagen 96 hr after giving the 35SO4=. The results from these studies indicate that young platelets adhere to collagen more readily than older platelets.
J. Hirsh, M. F. Glynn, J. F. Mustard
Experiments were designed to compare the distribution of free and antibody-bound unlabeled insulin to the distribution of free and antibody-bound insulin-125I. The insulin antibody was incorporated in a specific immune precipitate similar to the one used by Hales and Randle for the radioimmune assay of insulin. Insulin which was not bound by the specific immune precipitate was measured by the immune hemolysis inhibition assay. This report contains evidence that the addition of the unlabeled insulin in the radioimmune assay results in relatively more insulin-125I which remains free and less bound by antibodies than is the case with the unlabeled insulin. Methods are described for the separation of an electrophoretically homogeneous iodoinsulin from samples of crude iodoinsulin with average incorporations of less than 0.2 atoms iodine per molecule. These purified iodoinsulin fractions have a markedly attenuated biological activity. Evidence is presented which supports the postulate that only a portion of the antibodies in guinea pig insulin antiserum are capable of effectively binding with purified iodoinsulin.
Edward R. Arquilla, Henri Ooms, Karen Mercola
Distribution of intrarenal blood flow was studied in 12 dogs anesthetized with Nembutal. Medullary blood flow was estimated by local clearance of hydrogen gas from the outer medulla measured polarographically with needleshaped platinum electrodes, and by local clearance of 85Kr and mean transit time of 32P-labeled erythrocytes measured with a small semiconductor detector placed in the outer medulla. Cortical blood flow was estimated from cortical red cell transit time and from total renal blood flow measured by electromagnetic flowmeter.
Knut Aukland, Mats Wolgast
This study utilized rates with hereditary hypothalamic diabetes insipidus (D.I.) in order to explore possible mechanisms which prevent full urinary concentration after acute administration of vasopressin in hypothalamic D.I. and which correct this concentrating defect with prolonged therapy.
Avery R. Harrington, Heinz Valtin
The mechanisms of reflex vasodilation were studied in an innervated canine hindlimb preparation which was perfused at a constant rate. Reflex vasodilation was produced by suddenly increasing the pressure in the trunk by the intravenous injection of norepinephrine, with consequent stimulation of the baroreceptors. When the basal vasoconstrictor tone exerted by the sympathetic nervous system on the systemic arterial bed was minimized, either by pretreatment with the alpha adrenergic blocking agent phenoxybenzamine or with reserpine, which depletes endogenous catecholamine stores, reflex vasodilation was virtually abolished. Administration of cocaine, a drug which blocks reuptake of norepinephrine by the nerve terminals, significantly reduced reflex vasodilation, the response after cocaine averaging 47% of the vasodilator response in the control period. Cocaine also potentiated the vasoconstriction caused by intra-arterially administered norepinephrine but attenuated the vasoconstriction induced by tyramine. The antihistamine, tripelennamine, had effects similar to those of cocaine. It is suggested, therefore, that reflex vasodilation results from a sudden decrease in the level of norepinephrine at the neuroeffector junction, which is a consequence of the cessation of norepinephrine secretion, together with continued and possibly augmented uptake. When the uptake mechanism is impaired, either by the administration of cocaine or tripelennamine, the magnitude of reflex vasodilation is diminished. It does not appear necessary to postulate active secretion of a vasodilator substance to account for reflex vasodilation.
Gerald Glick, Andrew S. Wechsler, Stephen E. Epstein
Sodium excretion was studied in a group of patients with chronic renal disease, (a) on constant salt intakes of varying amounts with and without mineralocorticoid hormone administration and, (b) after acute extracellular fluid volume expansion. The lower the steady-state glomerular filtration rate (GFR), the greater was the fraction of filtered sodium excreted on both a 3.5 and 7.0 g salt diet; and the lower the GFR, the greater was the change in fractional excretion in the transition from the 3.5 to the 7.0 g salt diet. This regulatory capacity did not appear to be influenced by mineralocorticoid hormone administration. After acute expansion of extracellular fluid (ECF) volume, the increment in sodium excretion exceeded the concomitant increment in filtered sodium in six of nine studies and in the remaining three studies, the increment in excretion averaged 59% of the Δ filtered load (i.e., only 41% of the increase in filtered sodium was reabsorbed). During saline loading, the decrease in fractional reabsorption of sodium tended to vary inversely with the steady-state GFR, although all patients received approximately the same loading volume. When an edema-forming stimulus was applied during saline infusion, the natriuretic response was aborted and the lag time was relatively short. When GFR and the filtered load of sodium were increased without volume expansion, the Δ sodium excretion averaged only 19% of the Δ filtered load; moreover, changes in fractional sodium reabsorption were considerably smaller than those observed during saline loading. The data implicate the presence of a factor other than GFR and mineralocorticoid changes in the modulation of sodium excretion in uremic man.
Eduardo Slatopolsky, Ivan O. Elkan, Carol Weerts, Neal S. Bricker
The dog behaves like man in his ability to utilize dietary hemoglobin iron and, therefore, is an excellent model in which to study the mechanisms of absorption. Heme is taken up intact into the epithelial cell of the small intestine but the iron appears in the plasma in a nonheme form. A substance is present in mucosal homogenates which is capable of releasing iron from a hemoglobin substrate in vitro. This has a molecular weight greater than 64,000, and appears to behave as an enzyme. There is no difference in the in vitro, effective concentration of the hemesplitting substance in the mucosa of iron-loaded and iron-deficient dogs to explain in vivo changes in iron absorption. However, the rate at which the heme-splitting substance works in vivo appears to be increased by the removal of the nonheme-iron-end product from the epithelial cell to the plasma. Reduction of the heme-iron content within the epithelial cell may then enhance uptake from the lumen. These studies suggest that the labile nonheme-iron content of the intestinal epithelial cell determines its ability to accept heme as well as ionized iron from the lumen.
Lewis R. Weintraub, Morton B. Weinstein, Hans-Jurg Huser, Sheila Rafal
The effect of norepinephrine on exogenous vasopressin antidiuresis was investigated in water-loaded subjects. After an initial 2 to 3 hr period of water loading (phase 1), 10-100 mU of vasopressin per hr were infused at a constant rate for 1 hr (phase 2) followed by infusion of 10-100 mU of vasopressin per hr plus 600 μg of l-norepinephrine per hr for 1 hr (phase 3). Endogenous creatinine clearance, osmolal clearance, and free water clearance (in milliliters/minute) and sodium and chloride excretion (in milliequivalents/minute) were measured. In 10 subjects given 10-20 mU of vasopressin per hr during phases 2 and 3, free water clearance decreased significantly from phase 1 to phase 2 (9.3 to 0.15, P = 0.001) and increased during phase 3 norepinephrine infusion to 4.7 ml/min (P = 0.001). A comparable decrease in phase 2 free water clearance was observed in four subjects given 50 or 100 mU of vasopressin per hr during phases 2 and 3 (P < 0.01); however, the phase 3 norepinephrine infusion in these subjects was not associated with an increase in free water clearance. Creatinine clearance, osmolal clearance, and sodium and chloride excretion were unchanged throughout the studies in both groups of subjects.
D. A. Fisher
In eight sheep, uterine and umbilical blood flows and oxygen uptakes, the transplacental flow-limited clearance of an inert molecule, pH values, and oxygen pressures, saturations, and capacities in the main placental vessels have been measured during maternal air breathing and oxygen inhalation. The mean ±SEM percentage changes during oxygen inhalation were +4.6 ±8.4 for the umbilical flow, +2.8 ±8.7 for the uterine flow, and +4.6 ±6.2 for the clearance. None of these changes are statistically significant. Oxygen uptake rose slightly in two cases and remained unchanged in the others. In all cases the oxygen pressures, saturations, and contents rose significantly in the uterine and umbilical vessels with oxygen inhalation.
Frederick C. Battaglia, Giacomo Meschia, Edgar L. Makowski, Watson Bowes
To study the nature of the antigen-antibody complexes which initiate the specific wheal-and-flare (W & F) reaction in sensitized man, a homologous series of bivalent, oligovalent, and multivalent benzylpenicilloyl (BPO) haptens were quantitatively compared for their effectiveness in eliciting W & F in BPO-sensitized human subjects.
Bernard B. Levine, Anthony P. Redmond
The metabolic fate of the tyrosine hydroxylase inhibitor, α-methyl-para-tyrosine (α-MPT), was studied after oral administration of single and multiple doses to patients with pheochromocytoma and essential hypertension. No major urinary excretion product was found other than the drug itself, which accounted for 44-88% of the fate of single or repeated oral doses. Though less than 1% of the administered drug could be recovered in the urine as catechol metabolites, it was possible to identify α-methyldopa, α-methyldopamine, and α-methylnorepinephrine and to quantify the excretion of the first two of these compounds. This minor route of metabolism required revision of methodology (presented herein) for measuring urinary catecholamines during α-MPT treatment since these compounds produce spurious fluorescence in routine methods of assay for catecholamines. The catechol metabolites probably are not present in sufficient amounts to contribute to the biochemical effects of the drug. Determination of plasma concentrations of α-MPT during maintenance therapy and considerations of the kinetics of enzyme inhibition enabled a calculation to be made of the degree of inhibition of catecholamine synthesis to be expected in the patient. This was calculated to be about 75% for the highest doses employed and is similar in magnitude to experimentally determined values.
Karl Engelman, Eric Jéquier, Sidney Udenfriend, Albert Sjoerdsma
Alpha methyltyrosine (α-MPT) was administered to 52 patients from 4 days to 10 months; 22 patients were cases of pheochromocytoma and 20 had essential hypertension. Inhibition of catecholamine synthesis in the range of 50-80% was achieved with divided daily drug dosage of from 1.0 to 4.0 g. Striking clinical benefit was noted in patients with pheochromocytoma in whom the drug was used in preparation for surgery and during chronic medical management. The drug appeared to have limited usefulness when used in essential hypertension, unless added to existing therapy with conventional agents. No beneficial effects were noted in thyrotoxicosis, glaucoma, and Raynaud's phenomenon. Untoward effects in order of decreasing incidence were: sedation (with insomnia on withdrawal), anxiety, tremor, diarrhea, and galactorrhea. Drug crystalluria, which has been observed in animals and is currently restrictive of clinical trials, was not observed in these studies. Evidence is presented that the minor conversion of α-MPT to methyldopa probably does not contribute significantly to the central and peripheral effects of the drug.
Karl Engelman, David Horwitz, Eric Jéquier, Albert Sjoerdsma
Intimal sterol synthesis was examined in isolated human arterial segments obtained at surgery or at postmortem examination. The tissues were incubated with acetate-1-14C and mevalonate-2-14C and the incorporation of these precursors into sterols was determined. Intimal sterols were isolated by multiple chromatographic techniques and purified by bromination and oxidation procedures. The results indicate that the arterial intima can incorporate acetate and mevalonate into cholesterol, cholestanol, and squalene. Cholestanol was the major sterol synthesized by the arterial wall, but cholesterol production was also consistently observed. The findings suggest that local synthesis is a potential source of sterol accumulation within the arterial wall.
Aram V. Chobanian
When purified human C′1 esterase is injected intradermally in man, increased vascular permeability results. This effect is not blocked by soybean trypsin inhibitor and is not abolished by pretreatment with the antihistamine, pyribenzamine, or by compound 48/80. Thus, the effect is not due to the release of endogenous histamine. The decreased permeability response of individuals with a specific hereditary deficiency of C′2 is evidence for the complement-dependent nature of this reaction. The apparently normal response to intradermal C′1 esterase developed by individuals with an acquired specific deficiency of C′3 suggests that the vasoactive substance may be derived from one of the early reacting complement components. Characteristic attacks of angioedema have been provoked by the intradermal injection of human C′1 esterase in two individuals with hereditary angioneurotic edema. Patients with hereditary angioneurotic edema are unresponsive to intradermal injections of C′1 esterase immediately after attacks.
Martin R. Klemperer, Virginia H. Donaldson, Fred S. Rosen
During continuous intraduodenal infusion of emulsified fat in rats, a steady state of intestinal absorption is achieved.
Susanne Bennett Clark, Peter R. Holt
A technique for the in vitro culture of rheumatoid synovial tissue with 14C-amino acids and isolation and quantitation of the newly synthesized immunoglobulins has been developed. This technique has been used to compare immunoglobulin synthesis of 12 rheumatoid synovia with that of synovia from nonarthritic patients and with that of normal human lymph nodes and spleen. In addition, the spleen of a patient with Felty's syndrome has also been examined. Immunoglobulin synthesis in rheumatoid synovia has been shown to be quantitatively and qualitatively similar to that of normal human spleen and lymph nodes although somewhat less active than the Felty's syndrome spleen examined. 79% of the immunoglobulin produced in rheumatoid synovia was of the IgG type, whereas IgM comprised 10% and IgA, 11% of the total. Less than 10% of the IgM synthesized was found to be rheumatoid factor. A fraction containing approximately 90% of its radioactivity in the form of IgG has been obtained for further studies.
J. Donald Smiley, Charlotte Sachs, Morris Ziff
Sensitization of human recipients with transplantation antigens (leucocytes, skin, or kidney allografts) has resulted in the appearance of serum hemagglutinins directed against sheep, guinea pig, and rat erythrocytes. Such hemagglutinins have been identified as IgG and IgM antibodies. Their appearance was not related to AB0 erythrocyte group incompatibility between donors and recipients, and the antibodies were not of the Forssman or Paul-Bunnel type. The antibody responses appeared to be primarily directed against antigen(s) present on rat erythrocytes, but shared to varying extents by other species. The peak antibody titers occurred in association with allograft rejection. In this regard, they may be of interest as a possible early warning system for the diagnosis and prompt management of rejection crises in clinical organ transplantation.
F. T. Rapaport, K. Kano, F. Milgrom
The biological half-life of manganese and some factors influencing it have been studied in man. The disappearance of manganese from the body in normal subjects is described by a curve having two exponential components. An average of 70% of the injected material was eliminated by the “slow” pathway. The half-time characterizing this component showed a small variation in normal subjects and had an average value of 39 days. The half-time for the “fast” component also showed a small variation and had an average value of 4 days.
John P. Mahoney, Walter J. Small
To study the process of hormone action, we have developed an in vitro system utilizing minced oviduct from estrogen-treated chicks incubated in tissue culture medium. Progesterone added to the medium induced synthesis of a specific protein, avidin, that continued for up to 96 hr. During this period there was no increase in total oviduct protein, ovalbumin, or lysozyme, which suggests the specificity of the progesterone effect. The induction process was dependent on new protein synthesis, since cycloheximide inhibited the induction completely. Actinomycin D in doses that prevented nuclear RNA synthesis, but not general protein synthesis, inhibited avidin production 70-90%. Avidin synthesis was not affected by 5-fluorouracil. The rate of DNA synthesis examined by thymidine-3H pulse labeling was not stimulated during avidin induction. Hydroxyurea (an inhibitor of DNA synthesis) and colchicine (a mitotic inhibitor) did not prevent induction. Studies utilizing uridine-3H pulses showed an effect on rapdly labeled nuclear RNA coincident with induction. Nuclear RNA polymerase activity increased before avidin induction. Since avidin was the only new protein synthesized in response to progesterone, the early stimulation of nuclear RNA synthesis and RNA polymerase activity would suggest a mechanism of action for this steroid at the transcription level of protein synthesis.
Bert W. O'Malley, William L. McGuire
The recent isolation of highly purified human pituitary luteinizing hormone (LH) has permitted the development of a sensitive and specific radioimmunoassay for this hormone in plasma. Results of this immunoassay system employing anti-LH serum agree closely with previous reports for the measurement of plasma LH in which immunoassays employing cross-reactive antisera to human chorionic gonadotropin were used. The immunoassay and bioassay of LH in several crude and partially purified pituitary and urinary extracts show acceptable agreement. The sensitivity of the LH immunoassay (0.2 mμg/ml) is adequate to measure LH levels in almost half of all prepuberal children and in all but a few normal adults. A small, but significant, rise in plasma LH level occurs at pubescence in both boys and girls. In women, plasma LH level varies with both age and the phase of the menstrual cycle. The mean LH concentration in nine normal women during the follicular phase (1.2 mμg/ml was found to be significantly higher than during the luteal phase (1.0 mμg/ml). At midcycle, the mean peak LH level was 10.2 mμg/ml. In a large group of normal women, the mean plasma LH concentration rose significantly at menopause to a level of 5.8 mμg/ml during the fifth decade and 10.5 mμg/ml during the seventh decade. A small, but significant, rise in plasma LH concentration also occurred in men from the third and fourth decades (0.7 mμg/ml to the seventh and eighth decades (1.7 mμg/ml). Both estrogen and testosterone suppress plasma LH levels, but marked variation in response exists. The immunoassay serves as a useful diagnostic tool in evaluating men with gonadal failure, amenorrheic women of reproductive age, and postmenopausal women suspected of hypopituitarism. From the half-time disappearance of LH-131I in plasma (mean 69 min) and the calculated volume of distribution (2.5-2.8 liters) it has been determined that approximately 30 μg of LH is secreted per day in men, and in women except at midcycle, at which time the release of LH is estimated to be 10-15 times this basal rate.
Don S. Schalch, Albert F. Parlow, Robert C. Boon, Seymour Reichlin
Concurrent synthesis of two or more hemoglobins occurs in normal man, the human hemoglobinopathies, and certain animal species. Duck erythrocytes produced in response to acutely induced anemic hypoxia (hemolysis or blood loss) contained reciprocally altered proportions of Hb I (α2I β2I) and Hb II (α2II β2II); the relative proportion of Hb II was 50-100% increased. Relative rates of synthesis of the two hemoglobins remained proportional to their new concentrations throughout erythroid maturation. This information favors the proposal that relatively increased activity, not delayed decay, of biosynthetic processes responsible for net synthesis of Hb II had occurred. These studies support the concept that the individual biosyntheses of multiple hemoglobins, presumably under genetic control, are potentially manipulable, and they provide evidence for one mechanism leading in a reproducible fashion to alterations in net synthesis in vivo.
John F. Bertles, Thomas A. Borgese