Effect of nitroglycerin and dipyridamole on regional left ventricular blood flow during coronary artery occlusion.

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Abstract

Coronary vasodilators have been variously reported to increase, decrease, or have no effect upon blood flow to ischemic myocardium. Consequently, the effects of two different types of dilators, nitroglycerin (TNG) and dipyridamole, were studied with radioactive microspheres in open-chested dogs after coronary artery ligation. Given as a bolus i.v. injection 0.4 mg TNG resulted in an increase in blood flow to nonischemic areas of myocardium and a preservation of flow to ischemic regions, despite a fall in blood pressure. 5 min later blood pressure and nonischemic flow were back to base line, and a small selective increase in flow to ischemic myocardium was found (0.15-0.18 ml/min per g, P less than 0.05). During an 0.2 mg/min infusion of TNG, and also after 1 mg/kg i.v. dipyridamole, ischemic flow was maintained in the face of a 20-30% reduction in blood pressure. In this setting, nonischemic flow was unchanged during TNG and doubled after dipyridamole. With the addition of methoxamine in both dilator groups, blood pressure returned to base line while flow to ischemic areas increased above base-line values (TNG, 0.16-0.20 ml/min per g, P less than 0.01; dipyridamole, 0.18-0.31 ml/min per g, P less than 0.05). Epicardial ST segment elevations increased during TNG infusion and were unchanged after dipyridamole, but with addition of methoxamine, ST segments became less elevated in both drug groups, concomitant with the observed increase in collateral blood flow. These data indicate that both types of coronary vasodilators, when used in conjunction with methoxamine to support blood pressure, reduce collateral resistance, increase collateral flow, and reduce epicardial ST-segment elevations.

Authors

L C Becker

The Sézary syndrome: a malignant proliferation of helper T cells.

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Abstract

The Sézary syndrome is a frequently lethal disease characterized by circulating malignant cells of thymus-derived (T)-cell origin. The capacity of circulating malignant lymphocytes from patients with this syndrome to synthesize immunoglobulins and to function as helper or suppressor cells regulating immunoglobulin synthesis by bone marrow-derived (B) lymphocytes was determined. Peripheral blood lymphocytes from normal individuals had geometric mean immunoglobulin synthetic rates of 4,910 ng for IgM, 1,270 ng for IgA, and 1,625 ng for IgG per 2 X 10(6) cells in culture with pokeweed mitogen for 7 days. Purified normal B cells had geometric mean synthetic rates of 198 ng for IgM, 145 ng for IgA, and 102 ng for IgG. Leukemic cells from patients with the Sézary syndrome produced essentially no immunoglobulins. Adding normal T cells to normal B cells restored their immunoglobin producing capacity. Leukemic cells from four of five patients tested had a similar capacity to help immunoglobulin synthesis by purified normal B cells. Additionally, Sézary cells from one patient studied induced a nearly 10-fold increase in IgA synthesis by lymphocytes from a child with ataxia telangiectasia and selective IgA deficiency. Furthermore, these Sézary cells induced more than a 500-fold increase in IgG and IgA synthesis by lymphocytes from a child with Nezelof's syndrome. When Sézary cells were added to normal unfractionated lymphocytes, they did not suppress immunoglobulin biosynthesis. In addition, unlike the situation observed when large numbers of normal T cells were added to purified B cells, there was no depression of immunoglobulin synthesis at very high malignant T-cell to B-cell ratios. These data support the view that Sézary T cells do not express suppressor cell activity. The results presented in this paper suggest that neoplastic lymphocytes from the majority of patients with the Sézary syndrome originate from a subset of T cells programmed exclusively for helper-like interactions with B cells in their production of immunoglobulin molecules.

Authors

S Broder, R L Edelson, M A Lutzner, D L Nelson, R P MacDermott, M E Durm, C K Goldman, B D Meade, T A Waldmann

The lymphocyte beta-adrenoceptor in normal subjects and patients with bronchial asthma: the effect of different forms of treatment on receptor function.

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Abstract

beta-Adrenoceptor function has been compared in lymphocytes of normal subjects, asthmatic patients taking large doses of beta-adrenergic bronchodilators, and comparable asthmatics treated exclusively with nonadrenergic medication. The effect of prolonged administration of beta-adrenoceptor agonists on receptor function in normal subjects has also been examined. beta-receptor response in each situation was quantitated by changes in levels of cyclic AMP, measured by a protein-binding assay. Dose response curves to isoproterenol (10 nM-0.1 mM) have been constructed for each group. Maximal increase in cyclic AMP in lymphocytes from normal subjects (393.2+/-44.0%) and in asthmatics on nonadrenergic preparations (408.3+/-46.7%) was significantly greater (P less than 0.001) than in asthmatics taking large doses of beta-sympathomimetics (67.5+/-24.2%). Depression of the cyclic AMP response appeared to correlate with the degree of exposure to beta-adrenergic agonists but not with the prevailing severity of the patient's asthma. Withdrawal of beta-adrenergic drugs was followed by a reversion of the cyclic AMP response to normal values, which suggests that the depression was drug-induced rather than an inherent feature of the disease. This interpretation was confirmed by the finding that prolonged exposure of normal subjects to high doses of a beta-adrenergic agonist caused a marked and significant (p less than 0.001) reduction in the cyclic AMP response, very similar to that seen in asthmatics on large doses of adrenergic bronchodilators. A possible link between drug-induced changes in the cyclic AMP response and the rise in the United Kingdom asthma death rate in the 1960's is discussed.

Authors

M E Conolly, J K Greenacre

Relationship between epicardial ST-segment elevation and myocardial ischemic damage after experimental coronary artery occlusion in dogs.

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Abstract

The relationship between early and late epicardial electrocardiographic changes as well as those in regional myocardial blood flow (MBF) and the severity of myocardial damage was determined in 12 anesthetized dogs with left anterior descending coronary artery ligation. Radioactive microspheres (15 mum) were used to measure regional MBF at 15 min (early) and 24 h (late) after coronary occlusion. Severity of myocardial damage was assessed by the extent of myocardial creatine phosphokinase depletion 24 h after coronary ligation. There was a close linear correlation between myocardial creatine phosphokinase activity and regional MBF both early (r=0.93, 2P less than 0.001) and late (r=0.88, 2P less than 0.001). An inverse but less precise relationship existed between acute epicardial ST-segment elevation and early (r=-0.41, 2P less than 0.001), or late (r=0.35, 2P less than 0.05) regional MBF. Similarly, a weak correlation was found between myocardial creatine phosphokinase (IU/mg protein) at 24 h and early epicardial ST (millivolt) elevation (r=-0.36, 2P less than 0.02). In the center zones of the infarct with MBF 1/10 of normal, about 35% of the areas with normal QRS width had no epicardial ST-segment elevation 15 min after coronary occlusion. About 44% of the areas which developed pathological Q-waves in the electrocardiogram at 24 h had no ST elevation 15 min after coronary ligation. Late evolution of abnormal Q-waves occurred almost invariably in areas in which the early MBF was reduced to less than 50% of normal and in areas which subsequently had myocardial creatine phosphokinase levels reduced to less than 60% of normal. After coronary occlusion, the severity of the ultimate myocardial damage, which was directly proportional to the degree of reduction in MBF, was therefore not reliably predicted by the early epicardial ST-segment elevation. The data obtained in these studies suggest the need for caution in the use of acute ST-segment elevation as a predictive index of the extent or severity of myocardial ischemic damage.

Authors

M K Heng, B N Singh, R M Norris, M B John, R Elliot

Bone resorption in organ culture: inhibition by the divalent cation ionophores A23187 and X-537A.

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Abstract

The ionophores A23187 and X-537A were used as probes to investigate the possible role of calcium uptake by bone as a mediator for the stimulation of bone resorption induced by parathyroid hormone (PTH) and other agents in cultured mouse calvaria. The ionophores alone at concentrations from 1 nM to 20 muM did not stimulate bone resorption, nor did they potentiate bone resorption stimulated by submaximal concentrations of PTH after either brief (15-60 min) or extended (1-3 day) exposure to the ionophores. Unexpectedly, we found that the ionophores inhibit in a dose-dependent manner bone resorption stimulated by PTH and a wide variety of other compounds (prostaglandin E2, 1alpha-hydroxycholecalciferol, 3-isobutyl-1-methyl-xanthine, and phorbol myristate acetate). This inhibition was not due to irreversible damage to the bones by the ionophores, because the inhibition was reversible even after 24 h of treatment. Inhibition of bone resorption by the ionophores was observed in media of both high and low calcium concentration, indicating that the inhibition was not due to a critical extracellular calcium concentration. Inhibition by the ionophores differs qualitatively in several ways from that produced by calcitonin, a natural inhibitor of bone resorption. Furthermore, A23187 at 1.0 mug/ml had no effect on the accumulation of cyclic AMP in the medium of either control, PTH- or calcitonin treated calvaria. We conclude that the ionophores A23187 or X537A do not stimulate bone resorption nor potentiate the effects of stimulators of bone resorption; instead they are inhibitors of bone resorption stimulated by a wide variety of compounds.

Authors

J L Ivey, D R Wright, A H Tashjian Jr

Videometric analysis of regional left ventricular function before and after aortocoronary artery bypass surgery: correlation of peak rate of myocardial wall thickening with late postoperative graft flows.

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Abstract

The peak rate of systolic wall thickening (pdTw/dt) in regions of the left ventricle was determined by biplane roentgen videometry in 60 patients before and a median of 14 mo after aorto-coronary bypass graft surgery. The left ventricular ejection fraction, stroke volume, and end-diastolic volume and pressure did not change significantly after surgery in the presence of patent or occluded grafts (P greater than 0.05). Statistically significant increases occurred in the peak rate of systolic wall thickening regions supplied by patent bypass grafts, and significant decreases occurred in regions with occluded grafts (P less than 0.01). Of 42 preoperatively hypokinetic regions (pdTw/dt greater than 0 less than 5.0 cm/s) supplied by a patent graft, 30 improved by an average of 2.6 cm/s after operation; 18 returned to normal. Failure of 24 hypokinetic regions to improve to normal was associated with myocardial infarction in 11 or with late postoperative graft blood flows of less than 60 ml/min measured by videodensitometry, in 10. All seven preoperatively akinetic (pdTw/dt=0) or dyskinetic (pdTw/dt less than 0) regions did not improve after the operation despite the fact that, in five of the seven, coronary bypass flows were over 60 ml/min. All eight preoperatively hypokinetic regions supplied by coronary artery graft flows of less than or equal 40 ml/min failed to improve to normal after operation. All nine preoperatively hypokinetic regions supplied by coronary artery graft flows of over 60 ml/min improved to normal after surgery. Late postoperative coronary artery bypass graft flows, the functional status of the myocardium, the status and distribution of the native coronary circulation, and decreased regional function elsewhere in the ventricle must all be considered when regional left ventricular function is interpreted.

Authors

J H Chesebro, E L Ritman, R L Frye, H C Smith, D C Connolly, B D Rutherford, G D Davis, G K Danielson, J R Pluth, D A Barnhorst, R B Wallace

A study of the intrarenal recycling of urea in the rat with chronic experimental pyelonephritis.

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Abstract

The concentrating ability of the kidney was studied by clearance and micropuncture techniques and tissue slice analyses in normal rats with two intact kidneys (intact controls), normal rats with a solitary kidney (uninephrectomized controls), and uremic rats with a single pyelonephritic kidney. Urinary osmolality after water deprivation for 24 h and administration of antidiuretic hormone was 2,501+/-217 and 2,874+/-392 mosmol/kg H2O in intact and uninephrectomized control rats, respectively, and 929+/-130 mosmol/kg H2O in pyelonephritic rats (P less than 0.001 compared to each control group). Fractional water reabsorption and concentrating ability were significantly decreased in the pyelonephritic group, and, to achieve an equivalent fractional excretion of urea, a greater fractional excretion of water was required in the pyelonephritic rats than in the control rats. Whole animal glomerular filtration rate was 1.57+/-0.19 ml/min and 1.39+/-0.18 ml/min in intact and in uninephrectomized controls, respectively, and 0.30+/-0.07 ml/min in pyelonephritic rats (P less than 0.001 compared to each control group). Single nephron glomerular filtration rate was 35.6+/-3.8 nl/min in intact control rats and was significantly increased (P less than 0.05) in both uninephrectomized (88.0+/-10.8 nl/min) and pyelonephritic rats (71.5+/-14.4 nl/min). In all groups fractional water delivery and fractional sodium delivery were closely comparable at the end of the proximal convoluted tubule and at the beginning of the distal convoluted tubule. In contrast, fractional urea delivery out of the proximal tubule was greater in the intact control group (73+/-8%) than in either the uninephrectomized (52+/-2%) or the pyelonephritic group (53+/-3%) (P less than 0.005). Fractional urea delivery at the early part of the distal tubule increased significantly to 137+/-11% and 93+/-6% of the filtered load in intact control and uninephrectomized control rats, respectively (P less than 0.001 compared to the late proximal values of each group), but failed to increase significantly in pyelonephritic rats (65+/-13%), indicating interruption of the normal recycling of urea in the latter group. Analysis of tissue slices demonstrated a rising corticopapillary gradient for total tissue water solute concentration as well as for tissue water urea concentration in both groups of control rats. In contrast, the pyelonephritic animals exhibited no similar gradients from cortex to papilla. These data indicate that the pyelonephritic kidney fails to recycle urea and accumulate interstitial solute. The latter must inevitably lead to a concentrating defect.

Authors

R M Gilbert, H Weber, L Turchin, L G Fine, J J Bourgoignie, N S Bricker

Potasssium transport in human blood lymphocytes treated with phytohemagglutinin.

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Abstract

We have confirmed that phytohemagglutinin (PHA) rapidly enhances the uptake of potassium (K+) by human blood lymphocytes. PHA, however, did not produce an increase in lymphocyte K+ concentration. The apparent steady-state of cell K+ concentration despite the marked increase in uptake of 42K+ could be explained by either an increase in K+-K+ exchange or an increase in concentrative (active) K+ accumulation in association with an increase in the leak of K+ from the cell. We compared, therefore, the uptake of 42K+ with the decrement in cellular K+ content when active transport was inhibited by ouabain. These studies established that K+-K+ exchange was negligible in human blood lymphocytes and that the increase in 42K+ uptake after PHA treatment represented concentrative transport. Our studies did indicate that 42K+ exodus from PHA treated lymphocytes increased markedly from 19 to 38 mmol-1 cell water-1-h-1. Within the same time period K+ influx into PHA-treated lymphocytes increased from 20 to 38 mmol-1 cell water-1-h-1. Thus, PHA produces a marked increase in the permeability of the lymphocyte membrane to K+, and the increase in active K+ influx in PHA-treated lymphocytes may represent a homeostatic response by the membrane K+ transport system to the increase in K+ efflux. Increased K+ turnover was observed at the lowest concentrations of PHA which produced an observable increase in [3H]thymidine incorporation into DNA. Thus, PHA produces an increase in K+ permeability that closely parallels its mitogenic effect. The rapid increase in K+ influx preceding blastogenesis and mitogenesis is required, therefore, to maintain normal intracellular K+ concentration. An adequate intracellular K+ concentration is essential for the synthetic processes required for cell transformation or division.

Authors

G B Segel, M A Lichtman

Modulation of the cyclic AMP content of rat renal inner medulla by oxygen: possible role of local prostaglandins.

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Abstract

The lower O2 tension and more active anerobic metabolism that pertain in the inner medulla (IM) of kidney relative to cortex (C) are well recognized, but there is no evidence that O2 availability constitutes a limiting or regulatory factor in IM metabolism or function. In the present in vitro study, we examined the effects of O2 on adenosine 3',5'-monophosphate (cAMP) metabolism in slices of rat renal C and IM. After a 20-min incubation of slices in Krebs Ringer bicarbonate buffer with 95% O2 + 5% CO2 serving as the gas phase, the cAMP content of IM was 6-10 fold higher than that of C in either the presence or absence of 2 mM 1-methyl-3-isobutylxanthine in the incubation media. In slices of IM incubated for 20 min with 1-methyl-3-isobutylxanthine, cAMP was 22.5+/-SE 2.48 pmol/mg wet weight at 95% O2 and 4.37 without O2. Oxygenation of O2-deprived IM increased cAMP twofold in 2 min, an effect fully expressed in 5 min (fivefold increase). Further, cAMP of IM rose progressively and significantly over a range of atmospheric O2 content from 0 to 50% conditions which should reproduce and encompass O2 tensions that pertain in tissues in vivo. By contrast, basal cAMP content of C varied less than twofold in the presence of 95% versus no O2, implying that O2 modulation of cAMP was specific for IM. Indomethacin and meclofenamate, structurally distinct inhibitors of prostaglandin synthesis, both significantly decreased basal cAMP accumulation in oxygenated slices of IM but not of C. Meclofenamate also reduced basal adenylate cyclase activity determined in homogenates prepared from slices of IM which had been incubated at 95% O2. In slices of IM previously exposed to indomethacin or meclofenamate at 95% O2, a maximally effective concentration of exogenous prostaglandin E1 restored cAMP and adenylate cyclase activity to levels which approximated those observed at 95% O2 in the absence of an inhibitor of prostaglandin synthesis. These results suggest that O2 enhancement of cAMP content in IM may be mediated at least in part by local prostaglandins.

Authors

F R DeRubertis, T V Zenser, P A Craven, B B Davis

Assessment of group B streptococcal opsonins in human and rabbit serum by neutrophil chemiluminescence.

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Abstract

The factors important in host defense against group B streptococci are not well understood. The role of antibody and complement in the prevention of serious infection by these organisms is not known because, to date, a reliable measure of functional opsonic activity has not been developed. Recently, it has been shown that neutrophils produce a chemiluminescence after ingestion of particulate matter, and that this event can be detected and quantitated in a liquid scintillation system. We have adapted the chemiluminescence procedure to examine rabbit hyperimmune and human serum for the presence of group B streptococcal opsonins. Group B streptococci of types Ia, II, and III that were opsonized in homologous but not heterologous type serum produced a peak in chemiluminescence when added to normal human neutrophils. Such activity was correlated, in each instance, with ingestion of bacteria by neutrophils and deposition of immunoglobulin and C3 on the bacterial surface as detected by indirect immunofluorescence. With this assay, we have examined sera from colonized and diseased patients for the presence of opsonins to types Ia, II, and III group B streptococci. Maternal sera often contained type-specific opsonins which resided in the IgG fraction and which crossed the placenta to appear in paired cord specimens. 63% of patients colonized with group B streptococci had serum opsonins to their colonizing type of organism. In contrast, none of the 15 patients with sepsis or meningitis had opsonins directed against their infecting strain. These data suggest that the lack of type-specific opsonins to group B streptococci may be one of the important factors in determining host susceptibility to systemic infection with strains of this group.

Authors

V G Hemming, R T Hall, P G Rhodes, A O Shigeoka, H R Hill

Cutaneous antigen-stimulating lymphokine production by lymphocytes of patients with progressive systemic sclerosis (scleroderma).

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Abstract

Cell-mediated immunity to skin extracts was studied by the macrophage migration inhibition test, lymphocyte transformation, and direct cytotoxicity to skin fibroblasts, in normal individuals and patients with progressive systemic sclerosis. The latter included 18 individuals with diffuse scleroderma and 12 with the CREST syndrome, a variant form of systemic sclerosis in which there is more limited involvement of the skin. Controls consisted of 13 patients with other connective tissue diseases and 16 normal individuals. Phosphate-buffered saline and 3 M KCl extracts of both normal and sclerodermatous skin were used as antigens. No evidence of lymphocyte reactivity was found by the lymphocyte transformation and direct cytotoxicity test procedures. However, the lymphocytes of patients with diffuse scleroderma did respond to extracts of both normal and sclerodermatous skin in the migration inhibition assay. 10 of 16 patients (62.5%) had migration indices below 2 SD of the normal range, 1 of 10 CREST patients and 1 of 13 patients with other connective tissue diseases showed similar reactivity. Antisera specific for immunoglobulin-bearing lymphocytes (B lymphocytes) and T lymphocytes were used to characterize the lymphocytes found in skin biopsies of patients with diffuse scleroderma. T lymphocytes made up the majority of lymphocytes in the skin infiltrates. These findings suggest that lymphocytes sensitized to skin extracts are present in patients with diffuse scleroderma. The cell-mediated immune reaction to skin antigens may be a factor in the pathogenesis of diffuse scleroderma.

Authors

H Kondo, B S Rabin, G P Rodnan

Flaujeac factor deficiency. Reconstitution with highly purified bovine high molecular weight-kininogen and delineation of a new permeability-enhancing peptide released by plasma kallikrein from bovine high molecular weight-kininogen.

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Abstract

Flaujeac trait is the functional deficiency of a plasma protein of the intrinsic coagulation, kinin-forming, and plasma fibrinolytic pathways. The Flaujeac factor in man has been isolated and tentatively identified as a kininogen of high molecular weight (HMW). Highly purified bovine HMW-kininogen, but not bovine low molecular weight kininogen, repaired Flaujeac factor deficiency. The two subspecies of this molecule, HMW-kininogen a and HMW-kininogen b, also corrected Flaujeac factor deficiency. When bovine HMW-kininogen was incubated with bovine plasma kallikrein, kinin-free HMW-kininogen, bradykinin, and a glycopeptide fragment (peptide 1-2; 12,584 daltons) were rapidly released. None of these fragmentation products corrected Flaujeac factor deficiency alone or in mixtures. The function of HMW-kininogen appeared to depend upon the structural integrity of the native molecule. When injected in concentrations of 2 pmol-8 nmol/0.1 ml, peptide 1-2 caused increased vascular permeability in rabbits, rats, or guinea pigs. The enhanced permeability was maximal within 1-2 min and terminated in 5-10 min, differing from that of bradykinin or histamine. Injected together in equimolar amounts, peptide 1-2 and bradykinin produced a synergistic permeability response which was immediate in onset as well as prolonged in duration. Peptide 1-2 is a rapidly acting, highly basic glyco-peptide which mediates increased vascular permeability in a complementary and synergistic manner with bradykinin.

Authors

R T Matheson, D R Miller, M J Lacombe, Y N Han, S Iwanaga, H Kato, K D wuepper

The role of insulin and glucagon in the regulation of basal glucose production in the postabsorptive dog.

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Abstract

The aim of the present experiments was to determine the role of insulin and glucagon in the regulation of basal glucose production in dogs fasted overnight. A deficiency of either or both pancreatic hormones was achieved by infusin somatostatin (1 mug/kg per min), a potent inhibitor of both insulin and glucagon secretion, alone or in combination with intraportal replacement infusions of either pancreatic hormone. Infusion of somatostatin alone caused the arterial levels of insulin and glucagon to drop rapidly by 72+/-6 and 81+/-8%, respectively. Intraportal infusion of insulin and glucagon at rates of 400 muU/kg per min and 1 ng/kg per min, respectively, resulted in the maintenance of the basal levels of each hormone. Glucose production was measured using tracer (primed constant infusion of [3-3H]glucose) and arteriovenous difference techniques. Isolated glucagon deficiency resulted in a 35+/-5% (P less than 0.05) rapid and sustained decrease in glucose production which was abolished upon restoration of the plasma glucagon level. Isolated insulin deficiency resulted in a 52+/-16% (P less than 0.01) increase in the rate of glucose production which was abolished when the insulin level was restored. Somatostatin had no effect on glucose production when the changes in the pancreatic hormone levels which it normally induces were prevented by simultaneous intraportal infusion of both insulin and glucagon. In conclusion, in the anesthetized dog fasted overnight; (a) basal glucagon is responsible for at least one-third of basal glucose production, (b) basal insulin prevents the increased glucose production which would result from the unrestrained action of glucagon, and (c) somatostatin has no acute effects on glucose turnover other than those it induces through perturbation of pancreatic hormone secretion. This study indicates that the opposing actions of the two pancreatic hormones are important in the regulation of basal glucose production in the postabsorptive state.

Authors

A D Cherrington, J L Chiasson, J E Liljenquist, A S Jennings, U Keller, W W Lacy

Translation and stability of human globin mRNA in Xenopus oocytes.

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Abstract

Human globin messenger RNA (mRNA) prepared from erythroid cells of patients with sickle cell anemia has been translated in Xenopus laevis oocytes. Addition of hemin to the injected mRNA causes total globin synthesis to increase and the ratio of alpha- to betas-globin synthesis (alpha/betas ratio) to approach unity. To determine the effect of the length of the poly-(A) segment on human globin mRNA stability, 10 S globin mRNA was fractionated into poly-(A)-poor fractions by oligo (dT)-cellulose column chromatography. When oocytes are injected with each of these fractions, translation of the poly-(A)-rich globin mRNA is sustained for a longer period than that of the poly-(A)-poor mRNA. Regardless of the mRNA fraction injected, the alpha/betas ratio of the synthesized globin decreases as the injected oocytes are incubated for longer periods. The results indicate that in frog oocytes poly-(A)-rich mRNA has greater translational stability than poly-(A)-poor mRNA, AND beta-mRNA has greater stability than alpha-mRNA with comparable poly-(A) content.

Authors

G M Maniatis, F Ramirez, A Cann, P A Marks, A Bank

Umbilical uptake of amino acids in the unstressed fetal lamb.

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Abstract

The whole blood concentrations of 22 amino acids were measured in a chronic, unstressed fetal lamb preparations. Samples were taken daily from the umbilical artery, umbilical vein, and maternal artery over the latter quarter of gestation. 73 sets of samples (from the umbilical artery and vein and the maternal artery) from 13 animals were analyzed for amino acid levels. Oxygen contents were determined simultaneously in 48 sets (umbilical artery and vein) to relate fetal oxygen consumption to amino acid uptake via the umbilical circulation. The results indicate that there is no umbilical uptake of the acidic amino acids, glutamate and aspartate; there is, in fact, a net flux of glutamate out of the fetus into the placenta. As both of these amino acids are major constituents of body proteins, the data indicate that they are formed within the fetus. The umbilical uptake of some neutral and basic amino acids (e.g., valine, leucine, isoleucine, arginine, phenylalanine, and tyrosine) is in considerable excess of estimated growth requirements, suggesting that some amino acids undergo extensive transamination and oxidative degradation in the fetus. Finally, the net uptake of nitrogen, carbon, and calories by the growing ovine fetus in the form of amino acids, glucose, and lactate is compared to estimated requirements as determined in previous studies.

Authors

J A Lemons, E W Adcock 3rd, M D Jones Jr, M A Naughton, G Meschia, F C Battaglia

Correlation between insulin receptor binding in isolated fat cells and insulin sensitivity in obese human subjects.

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Abstract

This study examined the relationship between receptor binding of insulin in a metabolically significant target tissue in vitro and sensitivity to insulin in vivo in obese human subjects. Specific insulin binding was measured at 24 degrees C in isolated enlarged fat cells obtained from 16 patients, by observing the effect of increasing concentrations of unlabeled insulin on the binding of [125I]insulin. Scratchard plots of the binding data were curvilinear with an upward concavity, similarity shaped, and essentially parallel. Kinetic studies on the dissociation of [125I]insulin from fat cells indicated that these curvilinear Scratchard plots could be explained by the presence of site:site interactions of the negative cooperative type. Differences in binding between individual patients were predominantly due to differences in the numbers of receptor sites whether expressed in relation to cell number, cell volume, or cell surface area. These findings were not accounted for by differences in [125I]insulin degradation. Acute exposure of adipose tissue to insulin in vitro had no significant effect on [125I]insulin binding to isolated cells. The number of receptor sites was directly correlated with insulin sensitivity in vivo, measured as the rate constant (Kitt) for the fall in blood glucose after intravenous insulin, and was inversely correlated with the level of fasting plasma insulin. These findings corroborate those from other studies using human mononuclear leukocytes and various tissues from the obese mouse, which indicate that decreased insulin binding is a characteristic feature of insulin resistance in obesity.

Authors

L C Harrison, F I Martin, R A Melick

Characterization of antibodies to the insulin receptor: a cause of insulin-resistant diabetes in man.

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Abstract

We have characterized the circulating inhibitor of insulin receptor binding found in several patients with a new syndrome of extreme insulin resistance. The inhibitor is an immunoglobulin by multiple criteria, including precipitation by 33% ammonium sulfate, migration on G-200 Sephadex gel filtration and DEAE chromatography, and immuno-precipitation with specific anti-human immuno-globulins. Although predominantly IgG, some activity is found in the IgM fraction of the immunoglobulins in one patient. The inhibitory immunoglobulins reacted with antisera to both kappa and lambda light chain determinants and are therefore polyclonal. In addition, activity is retained in the F(ab')2 fraction of pepsin-digested IgG. Evidence suggests that these antibodies are directed at determinants on or near the insulin receptor, and that they are responsible for the observed clinical insulin resistance.

Authors

J S Flier, C R Kahn, D B Jarrett, J Roth

Effects of fasting on insulin binding, glucose transport, and glucose oxidation in isolated rat adipocytes: relationships between insulin receptors and insulin action.

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Abstract

Insulin binding, glucose transport, and glucose oxidation were studied in isolated adipocytes obtained from fasting rats. Fasting led to an increase in the overall binding affinity for insulin, while the number of receptor sites per cell remained constant. Glucose oxidation was markedly attenuated during fasting. Basal rates of oxidation decreased by about 50%, while insulin-stimulated rates decreased 6 to 10-fold. Glucose transport was assessed by measuring initial uptake rate of 2-deoxy-glucose. Fasting led to a 40-50% decrease in the apparent maximal transport capacity (Vmax) of 2-deoxy-glucose uptake with no change in apparent Km. A progressive decrease in basal and insulin-stimulated rates of 2-deoxy-glucose uptake was seen from 24-72 h of starvation and a significant correlation (r=0.85, P less than 0.001) existed between basal and maximal insulin-stimulated uptake rates in individual animals. When 2-deoxy-glucose uptake was plotted as a function of insulin bound, due to the decrease in maximal uptake capacity, cells from fasting animals took up less hexose for any amount of insulin bound. When the insulin bound was plotted as a function of the percent insulin effect on uptake, control cells and cells from 24-h-fasted rats gave comparable results, while cells from 48- and 72-h-fasted animals still took up less hexose for any amount of bound insulin. The effects of fasting on 3-O-methyl glucose uptake were comparable to the 2-deoxy-glucose data. In conclusion: (a) insulin binding is increased during fasting due to an increased overall binding affinity with no change in receptor number; (b) glucose oxidation is severely impaired during fasting; (c) 2-deoxy-glucose uptake decreases with fasting due to a decrease in maximal transport capacity (Vmax) with no change in Km; (d) the decrease in glucose oxidation is much greater than the decrease in glucose transport, indicating impaired intracellular oxidative metabolism; and (e) coupling between insulin receptors and the glucose transport system is normal after 24 h of fasting but is impaired at 48 and 72 h.

Authors

J M Olefsky

Renal resistance to parathyroid hormone during phosphorus deprivation.

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Abstract

Because previous studies have demonstrated that renal inorganic phosphate reabsorption is enhanced in rats after dietary phosphorus deprivation, we studied the effects of parathyroid hormone (PTH) upon inorganic phosphate reabsorption in acutely thyroparathyroidectomized rats stabilized on a low phosphorus diet to determine if the phosphaturic response to PTH is impaired during phosphorous depletion. Acutely thyroparathyroidectomized phosphorus-deprived rats responded only minimally to PTH, whereas similarly prepared animals stabilized on a high phosphorus diet exhibited a large phosphaturic response. Base-line urinary cyclic AMP values and PTH-induced increases in cyclic AMP excretion were similar in both groups. In other experiments, dibutyryl cyclic AMP elicited a greatly diminished phosphaturic response in phosphorus-deprived rats, as compared to their high phosphorus counterparts. These results indicate that the renal phosphaturic responses to PTH and cyclic AMP are impaired during dietary phosphorus deprivation. The impaired phosphaturia would contribute to phosphorus conservation and to the replenishment of inorganic phosphate stores after phosphorus depletion.

Authors

T H Steele, B A Stromberg, J L Underwood, C A Larmore

Regulation of low density lipoprotein receptor activity in freshly isolated human lymphocytes.

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Abstract

Circulating human lymphocytes freshly isolated from venous blood of 15 normal subjects exhibited a low capacity to bind, take up, and degrade 125I-labeled low density lipoprotein (LDL). However, when these cells were incubated for 72 h in the absence of lipoproteins, they gradually acquired in increased number of high affinity cell surface receptors for LDL. The increase in the number of LDL receptors was associated with a 16-fold increase in the rate at which the cells were able to take up and degrade the lipoprotein. The LDL binding and degradation processes that developed in normal lymphocytes exhibited the following characteristics; (a) high affinity (saturation was achieved at LDL concentrations below 50 mug protein/ml); (b) specificity (unlabeled LDL was much more effective than human high density lipoprotein or other plasma proteins in competing with 125I-LDL for binding to the LDL receptor); and(c) feedback regulation (the increase in the number of LDL receptors that appeared after incubation of freshly isolated lymphocytes in lipoprotein-deficient medium was prevented by exposure of the cells to either LDL or a mixture of 25-hydroxycholesterol plus cholesterol but not to HDL). Freshly isolated lymphocytes obtaine from three subjects with the homozygous form of familial hypercholesterolemia failed to develop normal amounts of LDL receptor activity when incubated in medium devoid of lipoproteins. The current data indicate: (a) that the LDL receptors that appear on the surface of cholesterol-deprived, normal human lymphocytes are genetically identical to the previously characterized LDL receptors of cultured human fibroblasts and long-term lymphoid cells and (b) that at least one cell type in the human body, the circulating human lymphocyte, has the capacity to produce a high affinity LDL receptor that mediates the cellular uptake and degradation of plasma LDL.

Authors

Y K Ho, S Brown, D W Bilheimer, J L Goldstein

Quantitation of human gamma globin genes and gamma globin mRNA with purified gamma globin complementary DNA.

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Abstract

Complementary DNA (cDNA) specific for gamma-globin nucleotide sequences has been prepared by hybridizing total cDNA made from cord blood messenger RNA (mRNA) as template to an excess of normal adult human globin mRNA and recovering the single-stranded cDNA from hydroxylapatite. The specificity of the gamma cDNA for gamma mRNA sequences is strongly supported by the hybridization of this cDNA at low Cot values (Co, concentration of RNA and t, time in seconds) to RNA samples containing large amounts of functional gamma globin mRNA and the lack of hybridization to RNA samples containing little, if any, gamma-globin mRNA. The absence of cross-hybridization of gamma cDNA with alpha, beta, and delta mRNAs is demonstrated by the complete hybridization of the gamma cDNA to mRNA samples completely lacking either alpha or beta and delta mRNA. An estimate of the number of gamma-globin genes in human cellular DNA was obtained by hybridization of purified gamma cDNA to DNA from spleen and white blood cells of normal and beta-thalassemia subjects and measurement of the percent of gamma cDNA hybridized at saturation. The results indicate that there are between one and two gamma-globin genes per total haploid gene DNA equivalent obtained from both normal and beta-thalassemia subjects. These values are consistent with genetic evidence for the presence of multiple gamma gene loci in human cells. The finding that the number of gamma-globin genes in beta-thalassemia DNA is similar to that in nonthalassemia DNA indicates that a deletion of gamma-globin genes cannot account for either the inadequate gamma-globin synthesis or indirectly for the decreased or absent beta-globin synthesis in beta-thalassemia cells.

Authors

F Ramirez, J V O'Donnell, C Natta, A Bank

Action of cholecystokinin and cholinergic agents on membrane-bound calcium in dispersed pancreatic acinar cells.

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Abstract

In dispersed acinar cells prepared from guinea pig pancreas, cellular uptake of 45Ca was moderately rapid and reached a steady state by 60 min. At the steady state, 69% of total cellular 45Ca was membrane-bound. In acinar cells preloaded with 45Ca and then incubated with COOH-terminal octapeptide of cholecystokinin (CCK-OP) or carbamylcholine, total cellular 45Ca decreased by approximately 40% within 5-10 min and then steadily increased to control values by 60 min. Under identical conditions, membrane-bound 45Ca decreased by 40% within 5-10 min and remained constant for the duration of the incubation. Free cellular 45Ca did not change during the initial 30 min but then increased steadily to values three times those in control cells by 60 min. In cells preloaded with 45Ca and then incubated with EDTA, the loss of total cellular radioactivity stimulated by CCK-OP could be accounted for by loss of membrane-bound 45Ca. CCK-OP failed to alter total cellular uptake of 45Ca when both tracer and peptide were added at the beginning of the incubation. Under identical conditions, membrane-bound 45Ca was not altered by CCK-OP during the first 30 min of incubation but was significantly below control values after this time. The effect of CCK-OP on free cellular 45Ca was the same as in cells preloaded with the tracer. These results suggest that CCK-OP causes release of 45Ca from a membrane-bound compartment that equilibrates slowly with extracellular fluid and that the change in free cellular 45Ca is a secondary effect.

Authors

H T Shelby, L P Gross, P Lichty, J D Gardner