These experiments were designed to determine whether fasting and feeding were associated with differing rates of protein synthesis in the rat pancreas. It has been established that feeding, acetylcholine, or cholecystokinin-pancreozymin administration was associated with enhanced rates of digestive enzyme secretion; however, the literature is unclear as to effects of such stimulation on enzyme synthesis. Rats fed ad lib. or fasted 24, 48, or 72 hr were used for this study. Pancreases were removed and incubated in tissue culture medium with l-phenylalanine-14C, and incorporation into TCA-insoluble material as well as purified amylase was measured. Compared with fed controls, fasting 24, 48, and 72 hr was associated with 29%, 39%, and 35% decreases in incorporation of l-phenylalanine-14C into protein. Decreases of similar magnitudes were apparent whether the data were expressed in terms of protein or DNA. Pancreatic amylase isolated from rats fasted 48 hr contained 57% fewer counts of l-phenylalanine-14C than amylase isolated from fed rats. Moreover, rats fasted for 24 hr and given bethanechol chloride incorporated greater amounts of l-phenylalanine-14C into protein than fasted controls. Studies were performed to exclude changes in pool size of precursor (l-phenylalanine-14C) or product (amylase) in accounting for decreases associated with fasting. These studies demonstrate that fasting was associated with decreased rates of pancreatic amylase and protein synthesis in rats.
J. A. Morisset, P. D. Webster
The experiment was carried out on 3-month old puppies. Control animals received a diet normal in calcium and vitamin D. The diet for one group of experimental animals was deficient in both calcium and vitamin D, while another experimental group was fed a diet deficient in calcium but with adequate vitamin D. The response of these animals to injected parathyroid extract was evaluated over a 4 month period. The serum calcium response fell after approximately 20 days in both the calcium-deficient (vitamin D-replete) and the calcium- and vitamin D-deficient animals. The effect on the parathyroid response of the addition of calcium or vitamin D in vitamin D- and calcium-deficient animals was also evaluated. The addition of vitamin D to rachitic animals did not restore the response to parathyroid extract; however, in calcium- and vitamin D-deficient animals with normal calcium levels there was a restored response to parathyroid extract. Morphologic studies were made of the bone at various times during the experimental period; the presence of osteoid tissue correlated with the absence of a response to injected parathyroid extract. The results suggest that parathyroid hormone acts independently and requires the presence of mineralized bone for its action in raising the serum calcium. Vitamin D appears to be important in the mineralization of new bone tissue.
Jenifer Jowsey
The infusion of hyperimmune agglutinating antibodies into man or animals causes spherocytosis and hemolysis. The mechanism of spherocytosis was studied in rats given rabbit anti-rat red cell antiserum intravenously. During the 18 hr after antibody infusion, a time before the onset of reticulocytosis, hematocrits fell from 40.6 to 27.6%. However, no change occurred in mean cell volume, mean cell hemoglobin content, or the red cell concentrations of potassium or adenosine triphosphate (ATP). There was a progressive loss of membrane constituents and membrane surface area which followed first order kinetics. At 18 hr membrane cholesterol had decreased 23.5%, phospholipid 26.3%, protein 4.7%, and surface area (calculated from a measure of osmotic fragility) 14.2%. There was no change in the per cent composition of the various phospholipids. Similar changes occurred in animals splenectomized before receiving antibody.
R. A. Cooper
Measurements of plasma renin activity (PRA) in renal vein blood from the ischemic kidney are reported to be generally higher than from the contralateral kidney. Importance of factors other than renin content of renal venous plasma has not been investigated. Initial rate measurements of angiotensin generation with added excess of homologous renin (Plasma Renin Substrate Activity [PRSA]-20 min) were made in bilateral renal venous plasma from 31 patients suspected of suffering from unilateral renal hypertension. The mean values from the involved vs. the contralateral kidney were 551 vs. 331 ng respectively of angiotensin II equivalents generated per milliliter of plasma per 20 min of incubation. The measurement of maximal angiotensin generation under the same conditions with incubation prolonged to 3 hr (PRSA-180 min), however, were bilaterally equal in renal venous plasma from selected patients with renal hypertension who showed distinct differences in PRSA-20 min and PRA measurements. Prior extraction of plasma lipids did not significantly change the bilateral renal venous PRSA-20 min determinations. Stimulation of endogenous renin release in normal dogs did not change the PRSA determinations. The data suggest strongly the presence of a “renin activating” mechanism in the renal venous plasma from the involved kidney of patients with renal hypertension.
Mohinder P. Sambhi, Charles E. Wiedeman
Humoral immunity to Hemophilus influenzae, type b was studied in normal human adults by means of assays for serum bactericidal and opsonizing activities against the organism and for passive hemagglutinating activity using erythrocytes sensitized with polyribophosphate, the type-specific capsular antigen. Hemagglutinating activity was detectable in about 60% of the 114 sera tested. Serum bactericidal and opsonizing activities were found in all sera tested; the levels in some sera, however, were quite low. The antibacterial activities were due not only to antibodies directed against the polyribophosphate capsule but also to antibodies that appear to be directed against somatic antigens. Type b strains differed in their susceptibility to the antisomatic antibodies of particular sera but were uniformly sensitive to anticapsular antibody.
Porter Anderson, Richard B. Johnston Jr., David H. Smith
In human volunteers, single injections of purified polyribophosphate elicited antibodies detectable by passive hemagglutination and by serum bactericidal and opsonizing activities against viable Hemophilus influenzae, type b. All three activities rose by 2 wk to maximal levels, at which they remained for at least 6 months. Doses of 1 μg elicited antibody responses in nearly all recipients; higher doses of the antigen, however, produced larger increases in titer. Booster doses of 1 μg given at 6 months did not further increase the antibody titers. A tuberculin-like response was often observed at the site of injections given intradermally.
Porter Anderson, Georges Peter, Richard B. Johnston Jr., Leslie H. Wetterlow, David H. Smith
A reproducible microbiologic assay of microgram quantities of idoxuridine (IDU) in serum, urine, or cerebrospinal fluid is presented. The antiviral assay is not interfered with by type-specific antibody or interferon. During slow intravenous infusions of idox-uridine (4 mg/min) in patients with suspected diagnoses of Herpesvirus hominis encephalitis, the rate of inactivation and/or removal of drug exceeded its administration. During several rapid infusions of idoxuridine (50 mg/min) significant quantities of the drug were found in serum, urine, and cerebrospinal fluid. Idoxuridine is not significantly bound to serum proteins and is not deiodinated in fresh serum or urine in vitro to inactive products (iodouracil, uracil, iodide). It is rapidly excreted into the urine. Inactivation of IDU occurs in tissues. This antiviral assay of IDU in body fluids should be applicable to other viruses and potential antiviral agents.
A. Martin Lerner, Elizabeth Jane Bailey
Mediators of acute immunologic injury have been studied in vivo by producing arthritis in rabbit knee joints. A reversed passive Arthus lesion was produced by injecting antibody into the joint space and antigen intravenously. Injury was assessed by measuring leakage of serum proteins and circulating radiolabeled proteins into the joint space and by the accumulation of neutrophils in the joint fluid. Inflammatory exudate was recovered for study by a standardized irrigation technique.
Claude V. Deshazo, Peter M. Henson, Charles G. Cochrane
During intravenous administration of varying doses of angiotensin II antibody to anesthetized rats, apparently specific vascular receptors were characterized. These receptors compete with administered antibody to bind circulating angiotensin. This competitive phenomenon was used to evaluate the affinity of these receptors for angiotensin. Apparent vascular receptor affinity was defined by the amount of antibody required to block the blood pressure response to exogenous angiotensin. It was found that this receptor affinity varies directly with sodium intake so that the amount of antibody required to block was eightfold greater in normal animals on a high sodium intake, as compared with those on a low sodium intake. Sodium dependence of receptors was also demonstrated in nephrectomized animals, in desoxycorticosterone (DOC)-treated rats, and in chronic renal hypertension. Thus the observed changes in receptor affinity were usually inversely related to measured endogenous angiotensin II levels. Ganglionic blockade increased antibody requirement eightfold. All of these changes were consistent, with no overlap observed in response of individual animals from different groups. These results may explain the variation in pressor activity of angiotensin associated with changes in salt balance and ganglionic blockade.
Hans R. Brunner, Paul Chang, Ronald Wallach, Jean E. Sealey, John H. Laragh
A reliable, relatively simple method for isolation and quantification of disaturated lecithins is described. In rabbit lung, 34% of the lecithins were disaturated, in alveolar macrophages, 19%. More than 95% of the fatty acids of the disaturated lecithins from lung and alveolar macrophages was palmitic. Hence, the disaturated lecithins from these sources were essentially all dipalmitoyl lecithin.
Robert J. Mason, Gary Huber, Martha Vaughan
Serum follicular-stimulating hormone (FSH) and luteinizing hormone (LH) as determined by radioimmunoassay, were correlated with sexual development in 29 patients with hypopituitarism (ages 14.2-29.9 yr).
Robert Penny, Thomas P. Foley Jr., Robert M. Blizzard
Washed human platelets were incubated with 0.1-1.0 U/ml human thrombin and the effects on adenyl cyclase activity and on a platelet membrane protein (designated thrombin-sensitive protein) were studied. Adenyl cyclase activity was decreased 70-90% when intact platelets were incubated with thrombin. The T½ for loss of adenyl cyclase activity was less than 15 sec at 1 U/ml thrombin. There was no decrease of adenyl cyclase activity when sonicated platelets or isolated membranes were incubated with these concentrations of thrombin. Loss of adenyl cyclase activity was relatively specific since the activities of other platelet membrane enzymes were unaffected by thrombin. Prior incubation of platelets with dibutyryl cyclic adenosine monophosphate (AMP), prostaglandin E1, or theophylline protected adenyl cyclase from inhibition by thrombin.
G. N. Brodie, Nancy Lewis Baenziger, Lewis R. Chase, Philip W. Majerus
Radioactive antigen binding tests have been developed to measure quantitatively the antibody response of 167 adults, 84 children, and 51 infants to several different preparations of group A and group C meningococcal polysaccharides. Almost all the adults injected responded and the geometric mean responses were approximately 15 μg/ml of antibody protein in individuals vaccinated subcutaneously with two preparations of group A vaccine. The geometric mean antibody concentration after immunization with two preparations of group C vaccine was approximately 35 μg/ml. Most children aged 7 yr responded to immunization with two group A vaccines, and their mean response was only slightly less than that seen in adults. There was no difference between the subcutaneous and the intradermal route if both were given with jet gun. The majority of infants aged 6-13 months responded to a preparation of group A vaccine and the geometric mean titer was approximately 1.2 μg/ml. Adults, children, and infants responded significantly less to a preparation of group A polysaccharide which was of low molceular weight.
Emil C. Gotschlich, Michel Rey, Rene Triau, Kenneth J. Sparks
The effect of beta adrenergic stimulation on renal-diluting capacity was examined in the dog. Beta adrenergic stimulation with intravenous isoproterenol significantly increased urinary osmolality (UOsm) and decreased free water clearance (CH2O), and these effects were rapidly reversible with cessation of the infusion. This antidiuretic effect of systemic beta adrenergic stimulation was comparable in innervated and denervated kidneys and was not associated with alterations in glomerular filtration rate or renal vascular resistance. Renal perfusion pressure was maintained constant in all of the experiments. The same dose of isoproterenol, which produced the antidiuretic effect and markedly stimulated cardiac beta adrenergic receptors when infused intravenously, was not found either to increase UOsm or to decrease CH2O when infused directly into the renal artery. Removal of the source of production and release of antidiuretic hormone (ADH) was, however, found to abolish the effect of intravenous isoproterenol on UOsm. A small effect on CH2O persisted and appeared to be related to an increase in arterial hematocrit. Thus, the results of the study exclude a major role of alterations in renal hemodynamics and renal innervation in the antidiuretic response to beta adrenergic stimulation with isoproterenol. They also provide no support for the hypothesis that beta adrenergic stimulation may directly alter the water permeability of the renal tubular epithelium. Rather the results suggest that the primary mechanism of the antidiuretic effect of beta adrenergic stimulation involves the integrity of the hypothalamoneurohypophyial system and the release of ADH.
Robert W. Schrier, Roger Lieberman, Robert C. Ufferman, Judith A. Harbottle
The pathways of bile acid synthesis in man were evaluated by studying the metabolism of 7α-hydroxycholesterol-4-14C and 26-hydroxycholesterol-16, 22-3H administered parenterally to individuals requiring external biliary drainage. Techniques for the identification of metabolites were thin-layer chromatography, column chromatography, gas-liquid chromatography with stream splitting, and crystallization to constant specific activity. It was found that both compounds were rapidly metabolized to bile acids and excreted in bile. Of the total radioactivity recovered in bile as bile acids, 87% of the 26-hydroxycholesterol-3H and 90% of the 7α-hydroxycholesterol-14C was found to be metabolized to both chenodeoxycholate and cholate. Compared to 7α-hydroxycholesterol, a greater proportion of 26-hydroxycholesterol was found to be metabolized to chenodeoxycholate.
Karl E. Anderson, Engeline Kok, Norman B. Javitt
The failure of blood flow to return to the kidney following a transient period of ischemia has long been recognized. The cause of this “no-reflow” has been investigated in the rat after a transient period of total obstruction of the renal arteries. The vascular pattern of the kidneys as visualized with silicone rubber injection shows a diffuse patchy ischemia throughout the kidney, which persists after release of the obstructed renal artery. Electron microscopic studies of ischemic kidneys showed that all cellular elements were swollen and limiting the available vascular space. Functional studies revealed an increase in plasma urea nitrogen and creatinine after 1 hr or longer ischemic periods. The ischemia, cell swelling, “no-reflow,” and subsequent renal dysfunction occurring after obstruction to the renal arteries were corrected by the administration of hypertonic mannitol, but were unaffected by an equivalent expansion of the extracellular fluid volume either with isotonic saline or isotonic mannitol, showing that the osmotic effect was primary. The hypothesis is presented that ischemic swelling of cells may occlude small blood vessels so that recirculation does not resume even after the initial cause of the ischemia is no longer present; solutes which do not penetrate cell membranes are able to shrink swollen cells, increase the available vascular space and thus permit reflow of blood to the ischemic organ.
Jorge Flores, Donald R. DiBona, Clyde H. Beck, Alexander Leaf
Glucose reabsorption was measured in dogs in which sodium reabsorption was stimulated by obstruction of the thoracic inferior vena cava or inhibited by volume expansion with Ringer's lactate. Glucose reabsorption was much higher during periods of enhanced sodium reabsorption than during sodium diuresis. The relationship of glucose reabsorption to glomerular filtration rate was examined using data from animals that had fractional sodium excretion rates of less than 1%. Under this condition the relationship of glucose reabsorption to glomerular filtration rate is highly linear. When points obtained during sodium diuresis (CNa/GFR>0.1) are plotted on the same graph, glucose reabsorption at any given glomerular filtration rate is much less than during antidiuresis. Glucose reabsorption divided by glomerular filtration rate varies inversely with fractional sodium excretion. This study demonstrates that glomerular tubular balance for glucose exists in the dog and that this balance is changed when sodium reabsorption changes.
Neil A. Kurtzman, Martin G. White, Philip W. Rogers, James J. Flynn III
Cerebrotendinous Xanthomatosis is a rare, inherited disease characterized by an extraordinary accumulation of cholestanol in all tissues, xanthomatous deposits in the brain, lungs, and Achilles tendons, premature atherosclerosis, and low plasma cholesterol concentrations. In two patients with the disease, the biosynthesis of cholestanol was examined by different techniques. After cholesterol-4-14C was injected intravenously into one patient, cholestanol and cholesterol isolated from the bile on 3 different days over the ensuing week contained significant radioactivity. The specific radioactivity-time curves for cholesterol-14C and cholestanol-14C suggested a precursor product relationship and provided additional evidence for the transformation of cholesterol into cholestanol. The second patient received intravenously a mixture of mevalonate-2-14C and stereospecifically labeled mevalonate-3R,4R-3H. Again cholesterol and cholestanol were isolated from the bile, and the 3H/14C ratio in both sterols was almost the same. This experiment again demonstrated that the biosynthetic path of cholestanol proceeded through cholesterol and not directly from earlier 5α-H-saturated precursors. These two independent lines of evidence indicate that the extraordinary deposition of cholestanol in Cerebrotendinous Xanthomatosis arises from cholesterol presumably through the accentuation of the normal biosynthetic pathway.
Gerald Salen, Alan Polito
Tumors from patients with the syndrome of inappropriate secretion of antidiuretic hormone (SIADH) have been found to contain large amounts of the antidiuretic hormone vasopressin. A lung tumor from a patient with hyponatremia most likely due to SIADH was removed at surgery and found to contain 23.5 mU vasopressin/g wet weight by radioimmunoassay Slices of this tumor were incubated with phenylalanine-3H. Arginine vasopressin-3H was purified from the incubate by Sephadex G-25 column chromatography in two different systems, performic acid oxidation, and gradient elution column chromatography with diethylaminoethyl Sephadex. As oxidation of vasopressin results in drastic conformational change with breaking of the ring of the cyclic polypeptide and addition of two cysteic acid residues per molecule, the radioactive material which eluted coincident with vasopressin both before and after this procedure was considered to be arginine vasopressin-3H. To our knowledge this is the first demonstration of in vitro biosynthesis of vasopressin by a tumor from a patient with SIADH.
Jack M. George, Charles C. Capen, Audra S. Phillips
Fetal renal function in the sheep was investigated in a chronic preparation which permitted repeated evaluations of urine flow and osmolality as well as renal clearances in animals which were unanesthetized and remote from acute surgical stress. Measurements of fetal blood pressure, pH, osmolality, fetal growth in utero, and final outcome did not indicate an adverse effect of the experimental procedure on the fetus.
Edwin L. Gresham, John H. G. Rankin, Edgar L. Makowski, Giacomo Meschia, Frederick C. Battaglia
Highly specific antisera to triiodothyronine (T3) were prepared by immunization of rabbits with T3-bovine serum albumin conjugates. Antisera with T3 binding capacity of up to 600 ng/ml were obtained. The ability of various thyronine derivatives to inhibit the binding of T3-125I to anti-T3 serum was found to vary considerably. l-T3, d-T3 and several triiodoanalogues were potent inhibitors of the reaction. Little inhibition of T3-125I binding was produced by l-thyroxine (T4) or other tetraiodo- analogues, thyronine or iodotyrosines. Chromatography of several T4 preparations indicated that most of their very slight activity could be ascribed to contamination with T3.
Jeffrey Lieblich, Robert D. Utiger
All cells in the intestinal villus of the rat are capable of synthesizing protein from amino acid precursors (l-leucine). Moreover, polyribosomes from both crypts and villi are equally able to incorporate l-leucine into protein. Unlike other tissues, e.g. liver, there is no diurnal variation of protein synthesis in the intestine of the unfed rat, whether leucine is administered intraluminally or intravenously.
David H. Alpers
The process of removal of triglyceride from the plasma may involve a sequential conversion of larger to smaller glyceride-rich lipoproteins. This has been studied within the species of lipoproteins comprising the very low density lipoproteins (VLDL) which transport the bulk of endogenously formed triglyceride. Palmitic acid-14C which was used to label the plasma glycerides was administered either as a prolonged constant infusion or as a pulse label. The specific activity-time curves of triglyceride fatty acids (TGFA) were analyzed both in total VLDL and in two subfractions of VLDL. The nature of the curves for total VLDL that were observed during the constant infusions were consistent with slow isotopic equilibration of precursors of VLDL-TGFA or with the presence of a precursor-product relationship between different components of VLDL-TGFA. The curves did not indicate any detectable differences in (fractional) turnover rates of independently metabolized pools of VLDL-TGFA. Differences in the specific activity-time curves of TGFA in two subfractions of VLDL (Sf > 100 and Sf 20-100) were consistent with a precursor-product relationship between TGFA in the two subfractions; again there was no indication of significant differences in (fractional) turnover rates. The specific activity-time curves of TGFA in the two subfractions of VLDL that were obtained with single injections of radio-palmitate showed a consistent difference in the rates at which TGFA became labeled in the two subfractions, being slower in the Sf 20-100 fraction. The findings from all experiments when considered together, were compatible with a precursor-product relationship that suggested that larger VLDL were converted to progressively smaller species as triglyceride was being removed.
Philip J. Barter, Paul J. Nestel
The antihypertensive function of the renal medulla was tested on accelerated (malignant) hypertension of the rabbit. A procedure for the development of accelerated hypertension of the rabbit of lethal proportions within 3 wk was established. This procedure consisted of the application of a rigid clip with a fixed and unyielding gap to the left renal artery and removal of the right kidney. Three additional manipulations, other than simple nephrectomy, were performed on the right kidney after application of the rigid clip to the left renal artery. These were: (a) a sham operation, (b) removal of the kidney and separation of the renal cortex and its autotransplantation in a fragmented state, and (c) removal of the kidney and separation of the renal medulla and its autotransplantation in a fragmented state. After the sham-operated kidney and autotransplanted renal medulla, the standardized accelerated hypertension did not develop, whereas after autotransplanted renal cortex it did. After a period of protection against accelerated hypertension, removal of either the sham-operated kidney or the renomedullary transplants was followed by a prompt rise in arterial pressure and death of the animal. Thus, the antihypertensive action of renomedullary tissue was similar to that of the whole kidney. The main cell type noted in the protective renomedullary transplants had the microscopic characteristics of the lipid-containing interstitial cells. These cells occurred in clusters, often were near capillaries, and appeared hyperplastic. It is suggested that the renomedullary interstitial cell is the most eligible cell for exertion of the renomedullary antihypertensive action. Since vasoactive lipids are extractable from the renal medulla and its interstitial cells, the hypothesis that interstitial cells secrete antihypertensive substance(s) is attractive.
E. E. Muirhead, B. Brooks, J. A. Pitcock, P. Stephenson
Blood flow through aorta-to-coronary artery bypass grafts has been measured selectively in 16 patients at or within 6 wk after operation. Inert gas desaturation curves were obtained from coronary venous blood samples after a 7-15 min infusion of dissolved H2 directly into the graft. Samples were analyzed chromatographically and curves resolved to 1-3% of initial H2 concentrations. Average flow per unit volume (F/V) was 67±21 (sd) ml/min per 100 g. Semilogarithmic plots showed F/V to be distributed heterogeneously in every case. In nine studies at operation, H2 measurements of average F/V were combined with electromagnetic measurements of total flow to estimate revascularized tissue mass. Electromagnetic flows ranged from 25 to 170 ml/min and averaged 69 ml/min. Tissue mass ranged from 46 to 155 g and averaged 88 g. We conclude that bypass grafts provide nutritive flow to significant amounts of myocardium at and shortly after operation. However, nutritive flow is not distributed evenly throughout the revascularized segment. The majority of the segment has a F/V within the accepted range of normal but there remain areas in which F/V is reduced significantly. The combination of inert gas and electromagnetic techniques allows a revascularized area to be characterized in terms of total flow, F/V, and tissue mass.
David G. Greene, Francis J. Klocke, George L. Schimert, Ivan L. Bunnell, Stephen M. Wittenberg, Thomas Lajos