Calcium and sodium permeability of erythrocytes from patients with untransfused α- or β- thalassemia major has been studied and compared to mature erythrocytes or control cells with comparable reticulocytosis. Isotopic Na+ influx was increased a mean fourfold greater than normals and threefold greater than reticulocyte rich control. Passive net leak of Na+ into thalassemic cells incubated with ouabain was also increased corresponding to their greater 22Na+ influx. Erythrocyte Na+ and K+ concentrations and cell water content per unit volume of cells were normal. Quantitation of active cation pumps in the cell membrane by the technique of [3H]ouabain binding showed a 2.6- to 9.9-fold increase above normal. Inward Ca2+ movement was studied in cells with absent Ca2+ pumping produced by depletion of either ATP or Mg2+-ions. Calcium uptake by ATP depleted thalassemic cells was increased 12-fold above normals and 3.6-fold above reticulocyte-rich controls. The Ca2+ uptake by Mg2+-depleted thalassemic cells was also increased above normal confirming that erythrocyte Ca2+ permeability is increased in this disease. Osmotic fragility measurements show that the surface area to volume ratio of thalassemic erythrocytes was increased by 15 to 25% above mature erythrocytes. The increased passive cation permeability of thalassemic erythrocytes cannot be explained by either reticulocytosis or an increased surface area to volume ratio of these cells. Moreover, erythrocyte Na+ and Ca2+ influxes in congenital dyserythropoietic anemia (CDA type 2) were increased 2- and 14-fold, respectively, above normal. The increased cation fluxes and cation pump numbers in thalassemic and congenital dyserythropoietic anemia erythrocytes are consistent with the hypothesis of membrane immaturity arising from rapid marrow transit times, a concept previously advanced to explain the persistence of i-antigen on these cells.
J. S. Wiley
The syntheses of triglyceride and its precursors were increased when liver homogenates of ketotic diabetic rats were incubated with [U-14C]-glycero 3-phosphate and cofactors. Triolein sonicates produced a concentration-dependent inhibition of the synthesis of both diglyceride and triglyceride, whereas monoolein sonicates had no effect. Rat serum very low density lipoproteins, like triolein sonicates, inhibited the synthesis of diglyceride and triglyceride. Furthermore, the intracellular form of very low density lipoproteins, namely nascent very low density lipoproteins, also inhibited the synthesis of diglyceride and triglyceride. A higher apparent I50 (concentration of inhibitor that produces 50% inhibition of activity) was observed in liver homogenates of ketotic diabetic rats for inhibition of triglyceride or diglyceride synthesis by triolein sonicates, serum very low density lipoproteins, high density lipoproteins, and nascent very low density lipoproteins. Insulin treatment of the diabetic rats reversed the I50 values to control. In studies on the site of inhibition of triglyceride synthesis in the overall biosynthetic pathway, serum very low density lipoproteins produced a concentration-dependent inhibition of liver cytosolic phosphatidate phosphohydrolase activity. A higher I50 value was obtained with the hepatic enzyme of the diabetic rats. This higher I50 value was reversed to control by insulin treatment of the diabetic rats. These results indicated that the activity of this enzyme was less sensitive to inhibition by very low density lipoproteins in the ketotic diabetic state. The reduced sensitivity of phosphatidate phosphohydrolase activity to triglyceride inhibition observed in the present studies could explain our previous observation of an increased rate of triglyceride synthesis in ketotic diabetic liver homogenates.
V K Murthy, J C Shipp
Murine schistosomiasis is a granulomatous disease associated with high serum and granuloma angiotensin I-converting enzyme (ACE) activity. SQ 14225, a specific competitive inhibitor of ACE, was administered to normal mice and mice infected with Schistosoma mansoni to determine whether this compound could inhibit granuloma ACE activity and modify the size of the granulomatous response to schistosome eggs. Peroral administration of SQ 14225 for 5 wk to infected mice with peak granulomatous responses decreased ACE activity in isolated liver granulomas. Treated mice demonstrated a decrease in granuloma size in the liver, colon, and ileum, and hydroxyproline concentration of isolated liver granulomas was increased. Mean diameters of synchronous pulmonary granulomas, induced by the pulmonary embolization of schistosome eggs into normal and sensitized mice, were decreased by a similar dose of SQ 14225. Withdrawal of SQ 14225 from unsensitized mice with 2-wk-old synchronous pulmonary granulomas induced an increase in inflammation. Infected, but not normal mice receiving SQ 14225 demonstrated reduced portal pressure, liver weight, and body weight. Both normal and infected mice experienced dipsogenesis, expanded intravascular volume, and increased serum ACE. These observations suggest that SQ 14225 can partially inhibit the granulomatous response to schistosome eggs and the pathological manifestations of schistosomiasis. It is possible that ACE has an inflammatory role in granulomatous inflammation.
J V Weinstock, M N Ehrinpreis, D L Boros, J B Gee
Human lymphocytes from elderly and young donors were cultured with phytohemagglutinin (PHA) or concanavalin A. Cultures from old donors produced less T cell growth factor (TCGF) and incorporated less tritiated thymidine (3H-Tdr) than did similar cultures from young donors in the presence of either mitogen. Furthermore, the response of lymphocytes from elderly donors to TCGF was impaired. Thus, PHA-activated T cells from aged donors showed no increase tritiated thymidine incorporation when incubated with exogenous human TCGF. In contrast, addition of exogenous human TCGF to PHA-activated peripheral blood leukocytes from younger individuals increased tritiated thymidine incorporation by 30-50%. The impaired response to TCGF was associated with decreased binding of TCGF by PHA-activated cells from old donors. TCGF production or responsiveness was not associated with the presence of "suppressor" activity in elderly T cell preparations. These studies suggest a possible molecular mechanism for the impaired proliferative response of elderly human T cells. These data lend support to the hypothesis that defects in the capacity to either produce or respond to TCGF may be a fundamental cause of immune deficiency.
S Gillis, R Kozak, M Durante, M E Weksler
It has been suggested that the phorbol ester tumor promoters act via the receptor-effector system for epidermal growth factor (EGF), since they interact with the EGF receptor system and mimic many of the effects of EGF in cultured cells. We have studied the interaction of phorbol esters with the EGF-responsive MCF-7 human breast cancer cell line. Similar to other systems, phorbol esters inhibit EGF binding in MCF-7 cells in a manner paralleling their potency as tumor promoters in mice. The effect is specific for EGF since the membrane binding of insulin is unaffected. Like EGF, the potent phorbol ester 12-O-tetradecanoyl-13-phorbol acetate (TPA) stimulates protein synthesis as indicated by a twofold increase in [3H]leucine incorporation into protein after 24 h in TPA. Cell morphology, however, is significantly different with TPA treatment. After 24-48 h in TPA, cells become markedly enlarged with increased cytoplasmic vacuolization and increased membrane microvilli. This is reflected in a fourfold increase in the protein/DNA ratio (control 13.1; TPA 55.9). Furthermore, TPA inhibits cell division in media with or without serum, and prevents growth stimulation by EGF. Low TPA concentrations (1.0 ng/ml) are active, and 10 ng/ml results in maximal inhibition of cell replication. Other phorbol esters inhibit MCF-7 cells relative to their tumor promoting activity in vivo and their ability to inhibit EGF binding in these cells. After 24 h in TPA, incorporation of [3H]thymidine into DNA is markedly reduced and the thymidine labeling index falls (33% to 2%) indicating very few S-phase cells. Growth inhibition is reversible by removing TPA from the medium. Similar inhibitory effects are seen with the two other human breast cancer cell lines studied, ZR75-1 and MDA-MB-231. In conclusion, phorbol esters may interact with the EGF receptor domain in MCF-7 human breast cancer cells, but they have distinct effects on cell morphology and growth suggesting alternative pathways of action. The antineoplastic activity of these compounds needs further investigation.
C. Kent Osborne, Barbara Hamilton, Marianne Nover, Jeanette Ziegler
The purpose of these experiments was to determine whether alterations in preproinsulin messenger (m)RNA activity could account for changes in insulin biosynthesis during fasting and refeeding. Rats were fasted 4 d and then fed for 6, 8, 24, or 48 h. With fasting, body weight decreased 25%, plasma glucose decreased from 6.1 to 2.2 mM, and pancreatic insulin content fell to 40% that of fed animals. Islet RNA decreased to 50% and protein to 55% that of control animals, while islet DNA content remained unchanged. After 6 h of refeeding, islet RNA content increased and was not significantly different from controls.
Stephen J. Giddings, John Chirgwin, M. Alan Permutt
The vascular effects of arginine vasopressin (AVP) were examined in conscious Sprague-Dawley rats. In six control rats, synthetic AVP at a dose of 40 ng/kg, injected as an intravenous bolus, resulted in a rise in mean arterial blood pressure (BP) from 127 to 149 mm Hg (P < 0.005). No tachyphylaxis was observed after a second AVP bolus administered 30 min later, as BP increased from 125 to 150 mm Hg, P < 0.005. In a second group of six rats, 1-deamino penicillamine, 2-(O-methyl) tyrosine AVP ([dPTyr (Me)]AVP), was administered intravenously at a dose of 10 μg/kg, just before the second AVP bolus. In this group of studies BP rose from 124 to 150 mm Hg (P < 0.01) after the first AVP bolus, but not after the second AVP bolus, which was administered after [dPTyr (Me)]AVP (129 vs. 129 mm Hg, NS). To assess the effect of this AVP pressor antagonist on BP in rats with suppressed endogenous vasopressin, six water-diuresing rats (mean urinary osmolality, 99 mosmol/kg H2O) were administered the analogue at the same dose as the first group of rats. The analogue exerted no demonstrable effect on mean BP (128 before vs. 129 mm Hg after [dPTyr (Me)]AVP, NS). In these rats, mean radioimmunoassayable levels of AVP were at or below the detectable limits of our assay (0.5 pg/ml). In contrast, six rats in which endogenous AVP was stimulated by fluid deprivation for 24 h (mean urinary osmolality, 2,489 mosmol/kg H2O and mean AVP level of 21.6 pg/ml) had a marked fall in BP when administered the AVP analogue. In these animals [dPTyr (Me)]AVP caused a fall in BP from 124 to 110 mm Hg (P < 0.005). This fall in blood pressure was due to a fall in peripheral vascular resistance (0.35 vs. 0.30 mm Hg/ml per min per kg, P < 0.02) after [dPTyr (Me)]AVP, as cardiac index remained unchanged.
Gary A. Aisenbrey, William A. Handelman, Patricia Arnold, Maurice Manning, Robert W. Schrier
Beginning on day 4 ex ovo, and every 3 d thereafter, genetically dystrophic Line 413 chickens were given intraperitoneal injections (4 mg/kg body wt) of a protease inhibitor, leupeptin, pepstatin, or antipain. Experimental chickens received protease inhibitors dissolved in a water:ethanol:dimethyl sulfoxide solution (50:40:10, vol:vol:vol). Control untreated animals received diluent injections.
Michael S. Hudecki, Catherine M. Pollina, Reid R. Heffner
The importance of the spleen in host defense against pneumococcal bacteremia has been suggested by a number of experimental models as well as the occurrence of the syndrome of overwhelming pneumococcal sepsis in asplenic individuals. We studied the mechanism of splenic protection against pneumococcal bacteremia using a guinea pig model. Rates of removal of pneumococci from the blood stream in normal and splenectomized guinea pigs were compared with the extent of hepatic and splenic sequestration of radiolabeled organisms for three different types of pneumococci. A relationship was found between the virulence of a pneumococcus for normal guinea pigs, the extent to which it is cleared by the spleen, and the magnitude of the defect in blood stream sterilization induced by splenectomy. The spleen plays an increasingly important role in the clearance of progressively more virulent organisms, for which hepatic clearance cannot compensate. Thus, the division between hepatic and splenic clearance of bacteremia is a key determinant of the outcome of experimental pneumococcal infection.
E J Brown, S W Hosea, M M Frank
The intrapulmonary instillation into rat lung of enzymes that generate oxygen metabolites results in acute lung injury. The injection of xanthine oxidase and xanthine produces acute lung injury that, in the presence of superoxide dismutase, but not in the presence of catalase, can be significantly diminished, suggesting that O2- has the capacity to injure the lung. Instillation of a generator of H2O2, namely glucose oxidase, will, in sufficient quantities, produce acute injury that is not neutrophil-dependent. When either a low dose of glucose oxidase alone or lactoperoxidase alone is employed, little lung injury occurs. However, instilling the combination of the two enzymes produces severe, acute injury that can be blocked in a dose-dependent manner by catalase, but not by superoxide dismutase. Purified human leukocytic myeloperoxidase, but not horseradish peroxidase, will substitute for lactoperoxidase in the model of lung injury. The lung damaging effects of these enzymes cannot be attributed to the presence of contaminating proteases. Acute lung injury produced by the instillation of glucose oxidase and lactoperioxidase progresses to interstitial fibrosis. These studies represent a direct application of generators of oxygen metabolites to the in vivo induction of lung injury. The data suggest that rat lung is susceptible to injury by a variety of oxygen metabolites, including O2-, H2O2 and its lactoperoxidase or myeloperoxidase-produced derivatives. The studies also indicate that lung injury produced by oxygen metabolites can result in interstitial pulmonary fibrosis.
K J Johnson, J C Fantone 3rd, J Kaplan, P A Ward
An animal model was used to determine the basis for the increase in purine biosynthesis that results from hepatic depletion of purine nucleotides, such as seen in patients with type I glycogen storage disease or following fructose administration. Mice were injected intravenously with glucose or fructose, 2.5 mg/g of body weight, and the animals were killed at 0, 3, and 30 min following carbohydrate infusion. Fructose, but not glucose, administration led to a threefold increase in [14C]glycine incorporation into hepatic purine nucleotides documenting an increase in the rate of purine biosynthesis in the liver of fructose-treated animals. In the fructose, but not the glucose-treated animals, there was a reduction in the hepatic content of purine nucleotides that are inhibitory for amidophosphoribosyltransferase, the enzyme that catalyzes the first reaction unique to the pathway of purine biosynthesis. PP-ribose-P, an important metabolite in the control of purine biosynthesis, was increased 2,3-fold in liver following fructose, but not glucose administration. In conjunction with the decrease in inhibitory nucleotides and increase in PP-ribose-P 29% of amidophosphoribosyltransferase was shifted from the large inactive to the small active form of the enzyme. Results of these studies demonstrate that the end-products of the pathway, purine nucleotides, control the activity of the enzyme that catalyzes the first reaction leading to purine nucleotide synthesis either through a direct effect of purine nucleotides on the enzyme, through an indirect effect of the change in nucleotides on PP-ribose-P synthesis, or a combination of these effects. The resultant changes in amidophosphoribosyltransferase conformation and activity provide a basis for understanding the increase in purine biosynthesis that results from hepatic depletion of purine nucleotides.
M Itakura, R L Sabina, P W Heald, E W Holmes
Most previous compartmental models describing bilirubin transport and metabolism in the liver have been validated solely by analysis of the plasma disappearance of radiolabeled bilirubin in human subjects. We now have determined the transport kinetics of a bilirubin tracer pulse by analysis of plasma, liver, and bile radioactivity data from 30 intact rats. Plasma [3H]bilirubin disappearance was best described by the sum of three exponentials, and a six-compartment model, derived by simulation analysis, was necessary and adequate to describe all experimental data. Examination of the injected radiolabeled bilirubin by extraction with hexadecyltrimethylammonium bromide and thin-layer chromatography revealed that 6.6% (mean) of the original pigment had been degraded to labeled nonbilirubin derivatives during preparation of the tracer dose. This material exhibited a significantly longer half-life (mean 50.6 min) of the plasma terminal exponential than that of authentic radiobilirubin (20.6 min). In isolated perfused rat liver, the kinetics of [3H]bilirubin in perfusate and bile readily fitted the proposed model. Compatibility of the model with the data obtained, both in the isolated liver and in vivo, required that a fraction of bilirubin conjugated in the liver be deconjugated and returned to the plasma. Deconjugation of bilirubin glucuronides was evaluated directly by infusion of bilirubin monoglucuronides, containing 14C in the glucuronosyl group, into rats with an external bile fistula. Since metabolic degradation of hydrolyzed 14C-labeled glucuronic acid yields 14CO2, this was measured in expired air. Whereas 86% of the administered labeled pigment was recovered in bile, 7% of the label appeared in 14CO2. These findings directly validate a portion of the proposed kinetic model and suggest that hepatic deconjugation of a small fraction of bilirubin glucuronides is a physiological event. Deconjugation may also account, at least in part, for the presence of increased concentrations of unconjugated bilirubin in the plasma of patients with cholestasis.
J Gollan, L Hammaker, V Licko, R Schmid
Investigative data have suggested that the extrapancreatic actions of the sulfonylureas may be paramount in their chronic antidiabetic action. The present study examines the effects of chronic sulfonylurea treatment on in vivo insulin action. Peripheral insulin levels, hepatic glucose production (Ra), and overall glucose disposal (Rd) were studied in six awake, normal dogs given both 0.5 and 1.0 mU/kg per min pork insulin for 2.5 h. This produces stable hyperinsulinemia from 15 to 150 min. Fasting euglycemia was held constant by the glucose clamp technique and averaged 99% basal glucose in all studies. Ra and Rd were determined from infusion of [3-(3)H]glucose, begun 90 min prior to insulin infusion. 10 mg of the sulfonylurea glipizide, was given daily to the test animals for the 10 to 20 d following appropriate control studies, then was withheld for 24 h, and the dogs were restudied. Glipizide treatment did not significantly alter basal glucose turnover, Ra, mean glucose values, or mean insulin levels as determined by radioimmunoassay. Increase in Rd above basal glucose turnover in response to insulin (delta Rd) was significantly (P less than 0.05) increased by glipizide treatment at both insulin dosage levels (paired analysis). At 1.0 mU/kg per min insulin, delta Rd rose from 2.6 mg/kg per min before glipizide to 6.5 mg/kg per min after glipizide treatment. At 0.5 mU/kg per min insulin, delta Rd went from 1.1 mg/kg per min before glipizide to 2.2 mg/kg per min after glipizide treatment. Glipizide treatment doubled the effects of insulin on Rd, while showing no significant effect upon insulin suppression of Ra. We conclude that a significant extrapancreatic chronic action of glipizide lies in its ability to selectively potentiate Rd.
W S Putnam, D K Andersen, R S Jones, H E Lebovitz
To study the influence of hypometabolism on evolving myocardial infarction in a model with intact autoregulation, we investigated 53 awake dogs after coronary artery occlusion. Severe hypothyroidism was induced by the intravenous administration of 131I. Animals were instrumented to obtain hemodynamic measurements, and regional myocardial blood flow was measured with radioactive microspheres. Infarct size was determined by the creatine kinase depletion method, and dysrhythmia analysis was performed from 24-h Holter monitor tapes in animals matched for infarct size. The microarchitecture of hypothyroid myocardium was determined by the electron microscope. Before coronary occlusion, mean systemic pressure in hypothyroid dogs was reduced by 14% and cardiac output reduced by 32%, with no change in left ventricular end-diastolic pressure, first derivative of left ventricular pressure rise, (dP/dt), or heart rate. After coronary occlusion, there was deterioration in hemodynamic measurements in both groups, with lower absolute levels of mean systemic blood pressure and cardiac output obtained in hypothyroid dogs. Hypothyroidism was detrimental to evolving infarction with a 36% increase in infarct size present in hypothyroid dogs (30 +/- 2%) compared to euthyroid controls (22 +/- 3%), P less than 0.05. Dysrhythmias were more severe in hypothyroid dogs. There were no changes in the relationship between regional myocardial blood flow and the extent of infarction after coronary occlusion. Abnormalities in microarchitecture were present in hypothyroid dog myocardium. Severe hypometabolism in this model was associated with alterations in hemodynamics, more severe dysrhythmias and changes in microarchitecture. The combined effect of these alterations resulted in an overall detrimental influence of hypothyroidism on evolving myocardial necrosis in this model.
R P Karlsberg, D A Friscia, W S Aronow, S S Sekhon
Amyloid fibril protein has been isolated from the tissues of a patient of Swedish ancestry with autosomal dominant heredofamilial amyloidosis. After solubilization in guanidine HCl, a significant amount of the protein was contained in a homogeneous low molecular weight fraction. Molecular weight of approximately 14,000, amino acid analysis, double immunodiffusion analysis and immunoelectrophoresis all supported this material being a prealbumin-related protein. Automated sequence analysis gave a mixture of amino acids at each step, suggesting an heterogeneous NH2-terminus. After cleavage of the protein with cyanogen bromide, a homogeneous peptide was obtained with the sequence Val-Val-Val-Leu-Asp-Ala-Val-Arg-Gly-Thr-Pro- corresponding in 9 of the 11 positions analyzed with the known sequence of human prealbumin, starting with position 14. Antiserum raised to the amyloid protein reacted with normal human prealbumin. After absorption with normal human serum, this antiserum continued to detect a determinant in the amyloid patient's serum, suggesting an abnormal serum prealbumin, which may be the precursor of the fibril protein in this type of heredo-familial amyloidosis. Indirect immunohistochemical studies on kidney tissue from the patient with amyloidosis showed marked staining with anti-prealbumin and anti-heredofamilial amyloid protein, but not with anti-AA or anti-kappa antisera. No genetic association between this family and amyloidosis and Portuguese families with familial amyloid polyneuropathy is known.
M D Benson
Pyrroline-5-carboxylate reductase, which converts pyrroline-5-carboxylate to proline, has been identified in human erythrocytes. The level of pyrroline-5-carboxylate reductase activity in these cells is comparable to the activity levels of major erythrocyte enzymes. The physiologic function of the enzyme in erythrocytes cannot be related to its function in other tissues, i.e., producing proline for protein synthesis. We examined the kinetic properties of erythrocyte pyrroline-5-carboxylate reductase and compared them to the properties of the enzyme from proliferating cultured human fibroblasts. We found that the kinetic properties and regulation of the erythrocyte enzyme are distinctly different from those for human fibroblast pyrroline-5-carboxylate reductase. These characteristics are consistent with the interpretation that the function of the enzyme in human erythrocytes may be to generate oxidizing potential in the form of NADP+.
G C Yeh, S C Harris, J M Phang
The precise mechanisms for paroxysmal reentrant supraventricular tachycardia (PSVT) initiation during right ventricular premature stimulation (V2 method) were analyzed in 14 consecutive patients with Wolff-Parkinson-White Syndrome in whom the PSVT was inducible during retrograde refractory period studies. 9 patients had left-sided and the remaining 5 of 14 had right-sided ventriculo-atrial (VA) accessory pathway (AP). At the basic cycle lengths (V1V1) ranging from 550 to 900 ms (mean, 657.1±139.5), closely coupled V2 (mean V1V2, 357.3±59.2 ms, range 320-500) produced retrograde His bundle (H2) activation via the bundle branches and retrograde atrial (A2) activation via the AP. As the V1V2 were further shortened, the V2 showed a retrograde block in the His Purkinje system (HPS) and conducted to the atria via AP in 9 of 14 cases. Subsequently, the A2 impulse conducted anterograde over the atrioventricular node-HPS to initiate a PSVT or an atrial echo response in all nine cases. In none of the patients was a PSVT induced by V2 when the latter produced retrograde H2 activation via the bundle branches. In 10 of 14 cases, however, the retrograde H2 was followed by a V3, due to macroreentry in the HPS. The V3 in turn blocked retrogradely in the HPS while producing A3 via the AP to initiate a PSVT or an atrial echo response in 9 of 10 cases. Retrograde block of V2 and/or V3 in the HPS resulted in PSVT initiation in 13 of 14 cases, whereas in the remaining 1 case the exact mechanism was not clear. In none of the patients in this series was the PSVT initiated with a retrograde block of V2 in the atrioventricular node with or without concomitant retrograde A2 activation via the AP. We conclude that within the ranges of cycle lengths tested, a retrograde block of V2 and/or V3 in the HPS is the most common mechanism for initiation of PSVT during ventricular premature stimulation in patients with the Wolff-Parkinson-White Syndrome.
Masood Akhtar, Mohammad Shenasa, Donald H. Schmidt
Arterial concentrations and splanchnic exchange of glucose, amino acids, lactate, pyruvate, and glycerol were determined in 14 hyperthyroid patients and 12 healthy controls. Seven of the patients were restudied after 5-12 mo of medical management at which time there was chemical and clinical evidence of a euthyroid state. The arterial level of glucose was slightly higher (+10%) in the patient group and the glycerol concentration was three times greater among the patients. The plasma levels of the glycogenic amino acids, alanine, glycine, and serine were decreased by 20-30%, while the concentrations of leucine, isoleucine, and tyrosine were increased by 20-80%. The levels of lactate and pyruvate were similar in patients and controls as were insulin and glucagon concentrations. Splanchnic glucose output in the patient group was 35% lower than in controls. However, total splanchnic uptake of glucogenic precursors was 100% higher than in controls and showed a direct linear correlation with serum triiodothyronine. Total precursor uptake could account for 75% of splanchnic glucose output in the patients, compared to 26% in controls. The increase in uptake of lactate, alanine, and other amino acids was due to a 35-80% rise in splanchnic fractional extraction plus a 20% rise in estimated hepatic blood flow. When the patients were restudied after medical treatment splanchnic exchange of glucose and glucose precursors had reverted to normal values. The present findings demonstrate that in hyperthyroidism (a) total splanchnic glucose output is reduced in relation to controls, (b) splanchnic uptake of gluconeogenic precursors is accelerated, largely due to a rise in fractional extraction of precursor substrates and to a smaller extent, as a result of an increase in hepatic blood flow, and (c) these changes revert to normal when a euthyroid state has been achieved.
J Wahren, A Wennlund, L H Nilsson, P Felig
The platelet Fc receptor, a membrane receptor for immune complexes or aggregated immunoglobulin (Ig)G, was compared in normal and myeloproliferative platelets. Washed platelets from 11 normal donors and 27 patients were incubated with fluorescein-conjugated ovalbumin-anti-ovalbumin complexes and examined by phase and fluorescence microscopy. Only 3.2±1% of the normal platelets stained, whereas 76±16% of the myeloproliferative platelets stained with the immune complex. The fluorescent staining was mediated by a platelet Fc receptor, as shown by the absence of platelet staining with immune complex containing antibody preincubated with Staphylococcal protein A to block the Fc region. In addition, no staining occurred with antigen or antibody alone or after preincubation of platelets with aggregated IgG. Platelets from normal or myeloproliferative donors did not stain with the immune complexes when the incubation was performed in plasma. The increased expression of Fc receptors on myeloproliferative platelets was corroborated by studies of [14C]serotonin release by immune complexes or aggregated IgG in 8 patients and 17 normal donors. Serotonin uptake was similar in both groups. Myeloproliferative platelets released significantly more serotonin than normal platelets at each concentration of immune complex or aggregated IgG; in addition, myeloproliferative platelets released serotonin in response to much smaller concentrations of immune complex or aggregated IgG. [14C]Serotonin release by myeloproliferative platelets was not increased above that of normal platelets when thrombin was used as the stimulus. The results were independent of patient age, sex, therapy, hematocrit, or platelet size. Interaction of circulating immune complexes with platelets bearing increased Fc receptors may contribute to the abnormal hemostasis associated with the myeloproliferative syndromes.
Anne Moore, Ralph L. Nachman
Patients with porphyrias have varying degrees of photosensitivity, associated with elevated levels of porphyrins in plasma, erythrocyte, urine and/or feces. To investigate the role of complement in the pathogenesis of cutaneous lesions, varying amounts of uroporphyrin were added to normal human serum (0.1-10 microgram/ml), and the mixtures were then exposed to 405 nm irradiation. Such treatments result in the diminution of total hemolytic complement activity and hemolytic titers of C1, C4, C2, C3, and C5; furthermore, cleavage products of C3 and C5 were detected. Chemotactic activity for human polymorphonuclear leukocytes was generated that was inhibitable by incubation with anti-C5, but not with anti-C3 antisera. No chemotactic activity was generated in Mg++-EGTA treated serum nor in C4-deficient guinea pig serum. These data indicate that irradiation with 405 nm light of normal human serum containing uroporphyrin results in activation of the complement system via the classical pathway, and the generation of complement (C5)-derived chemotactic activity for human polymorphonuclear leukocytes.
H W Lim, H D Perez, I M Goldstein, I Gigli
In patients with an autosomal recessive diamino acid transport disorder, lysinuric protein intolerance (LPI), we measured plasma and urinary amino acids basally, and during intravenous infusion of citrulline at two rates. Compared with controls, the patients' plasma citrulline concentrations rose similarly, but urinary citrulline excretion increased excessively. Their plasma arginine and ornithine levels rose subnormally, but massive argininuria and moderate ornithinuria appeared. The excretion rates of the third diamino acid lysine and other amino acids remained practically unaltered, thus excluding mutual competition as the cause for the increases. The results suggest that (a) in the normal kidney reabsorption involves partial conversion of citrulline to arginine and ornithine (metabolic run-out), (b) in LPI, the diamino acid transport defect is located at the basolateral cell membrane of the renal tubules; this inhibits the efflux of arginine and ornithine, increasing their cellular concentration, which in turn inhibits the metabolic disposal of citrulline, and causes leakage of arginine, ornithine, and citrulline into the tubular lumen.
J Rajantie, O Simell, J Perheentupa
Insulin-induced hypoglycemia by unknown mechanism(s) increases plasma arginine vasopressin (AVP) levels in humans. Mechanisms for increased AVP levels during central nervous system glucoprivation were investigated by administering 20-min i.v. infusions of 2-deoxy-d-glucose (50 mg/kg), a competitive inhibitor of glucose utilization, or normal saline (sham), to 24 normal volunteers. Some of the infusions were administered in combination with neuropharmacological blocking agents (placebo). The behavioral, physiological, metabolic, and hormonal correlates of 2-deoxy-d-glucose (2DG)-induced gluco-privation and AVP secretion were studied in a group (n = 5) pretreated for 1 wk with either mazindol (1 mg per os three times per day), a potent norepinephrine and dopamine-reuptake blocker, or placebo. A second group (n = 5) received either propranolol (3 mg/3 min followed by 80 μg/min) or normal saline infusion before and during 2DG administration. With 2DG alone, plasma AVP levels increased from 1.3±0.3 pg/ml at base line to a peak of 4.5±1.4 pg/ml at 60 min and remained elevated for 150 min. From 30 to 180 min after 2DG administration, the 2DG-infused volunteers increased their water intake in comparison with sham-infused volunteers. Marked increases in epinephrine and slight increases in norepinephrine were associated with increases in plasma glucose and renin activity and decreases in plasma potassium. Plasma sodium and osmolality increased transiently and mean arterial pressure (MAP) fell. These changes, however, were small and inconstant and could not account for the observed increases in thirst and AVP levels. Pretreatment with mazindol prevented the decrease in MAP and the increase in plasma renin activity (PRA) following 2DG infusions without modifying increased thirst, water intake, or AVP responses to glucoprivation. Pretreatment with propranolol effectively blocked β-adrenoreceptors as evidenced by increased MAP and plasma epinephrine, and abolition of the RPA increases during 2DG-induced glycoprivation, but did not suppress AVP and thirst responses. A cervical cord-sectioned patient lacking descending sympathetic out-flow had a potentiated thirst response to 2DG-induced glucoprivation in the absence of increases in sodium, catecholamines, and PRA. Thus 2DG administration activates mechanisms for increased thirst and AVP which are unrelated to changes in peripheral catecholamines, MAP, PRA, and osmolality.
D. A. Thompson, R. G. Campbell, U. Lilavivat, S. L. Welle, G. L. Robertson
Induction of antigen-specific and non-specific (polyclonal) humoral immune responses in vitro was investigated in peripheral blood mononuclear cells of aged (65-85 yr) and young (20-30 yr) volunteers. In vitro immunization of lymphocytes with antigen (sheep erythrocytes) was performed in a recently described microculture system, and anti-sheep erythrocyte plaque forming cells were quantitated in a direct hemolytic plaque assay. Immunoglobulin secreting cells, induced polyclonally with pokeweed mitogen, were quantitated in a reverse hemolytic plaque assay. Significant depressions of antigen-specific as well as polyclonal responses were noted in relation to advancing age. Antigen-specific responses were more frequently depressed than polyclonal responses. T cell mitogen concanavalin A (Con A) was used to amplify functions of autologous immunoregulatory T cells. Addition of 10 microgram/ml Con A to lymphocytes of young donors at culture initiation resulted in activation of suppressor cells and abrogated antigen-specific responses. Delayed addition of Con A, on the other hand, enhanced responses, presumably because of activation of helper T cells. Similar manipulations of lymphocyte cultures from aged donors showed failure of Con A to suppress antigen-specific responses in approximately half of the responders. In many nonresponders, responses within normal range were elicited by the delayed addition of Con A to their lymphocyte cultures. Deviations beyond the range of expected responses were noted in 32.5% of the co-cultures between pokeweed mitogen stimulated young and aged cells. Our findings suggest that age-related deficiencies of B cell function are frequently associated with dysfunction of immunoregulatory T cells and are only occasionally due to intrinsic defects of B cells.
S G Pahwa, R N Pahwa, R A Good
To see whether methylprednisolone would affect the pulmonary vascular response to endotoxemia, we studied responses to endotoxemia in the presence and absence of methylprednisolone in the same chronically instrumented, unanesthetized sheep. Infusion of Escherichia coli endotoxin (0.70-1.33 μg/kg) caused an initial period of marked pulmonary hypertension followed several hours later by a long period of increased vascular permeability when pulmonary vascular pressures were near base line (base-line pulmonary artery pressure (PPa) = 21±1 cm H2O SE, left atrial pressure (Pla) = 1±3; experimental PPa = 20±3, Pla = 3±4; P = NS), lung lymph flow (˙Qlym) was high (base-line ˙Qlym = 7.2±0.2 ml/h; experimental ˙Qlym = 23.2±1.0; P < 0.05) and lymph/plasma protein concentration (L/P) was high (base-line L/P = 0.65±0.04; experimental L/P = 0.79±0.05; P < 0.05). When methylprednisolone (1.0 g + 0.5 g/h i.v.) was begun 30 min before the same dose of endotoxin was infused, the initial pulmonary hypertension was less and the late phase increase in lung vascular permeability was prevented (experimental PPa = 24±1, Pla = 1±1, ˙Qlym = 10.0±0.4; L/P = 0.56±0.03). ˙Qlym and L/P were significantly (P < 0.05) lower than with endotoxin alone. Methylprednisolone began during the initial pulmonary hypertensive response to endotoxin also prevented the late phase increase in lung vascular permeability, but the drug had no effect once vascular permeability was increased. We conclude that large doses of methylprednisolone given before or soon after endotoxemia prevent the increase in lung vascular permeability that endotoxin causes, but do not reverse the abnormality once it occurs.
Kenneth L. Brigham, Ronald E. Bowers, Charles R. McKeen
Acute caffeine in subjects who do not normally ingest methylxanthines leads to increases in blood pressure, heart rate, plasma epinephrine, plasma norepinephrine, plasma renin activity, and urinary catecholamines. Using a double-blind design, the effects of chronic caffeine administration on these same variables were assessed. Near complete tolerance, in terms of both humoral and hemodynamic variables, developed over the first 1-4 d of caffeine. No long-term effects of caffeine on blood pressure, heart rate, plasma renin activity, plasma catecholamines, or urinary catecholamines could be demonstrated. Discontinuation of caffeine ingestion after 7 d of administration did not result in a detectable withdrawal phenomenon relating to any of the variables assessed.
D Robertson, D Wade, R Workman, R L Woosley, J A Oates
Severe endotoxemia, a condition where microembolization and intravascular coagulation are thought to play important roles, was treated experimentally with prostacyclin (PGI2). In a study of 24 dogs, 8 control animals injected with 1.75 mg·kg−1 of endotoxin died within 24 h. Six animals given intravenous aspirin 100 mg/kg, 30 min after endotoxin died. 9 of 10 dogs infused with 100 ng PGI2·kg−1·min−1 for 3 h, given 30 min after the injection of endotoxin survived 24 h (P < 0.025). Injection of endotoxin resulted in a: (a) maximal 62% fall in mean arterial pressure (P < 0.001); (b) transient doubling of mean pulmonary arterial pressure (P < 0.001); (c) initial 70% drop in cardiac index (P < 0.001); (d) decline in blood platelets from 213,700 to 13,700/mm3 (P < 0.001), and leukocytes from 7,719 to < 750/mm3 (P < 0.001); (e) depressed urine output (P < 0.001); (f) 34% decrease in blood fibrinogen (P < 0.01) and an increase in fibrin degradation products > 50 μg/ml (P < 0.001); (g) fivefold increase in circulating cathepsin D titer (P < 0.005) and (h) increase in blood norepinephrine (P < 0.005), dopamine (P < 0.005), and epinephrine (P < 0.001). Aspirin treatment led to an increase in mean arterial pressure (P < 0.001) and mean pulmonary arterial pressure (P < 0.005), but cardiac index, urine flow, platelets, leukocytes, fibrin degradation products, and cathepsin D levels remained similar to untreated controls. After infusion of PGI2 there was a: (a) prompt increase of cardiac index to base-line levels; (b) late increase in mean arterial pressure (P < 0.005) after the discontinuation of PGI2 treatment (c) restoration of urine output; (d) increase in circulating platelets to levels still below base line but above untreated control animals (P < 0.05); (e) no effect on circulating leukocyte levels; (f) fall in fibrin degradation products to 11.2 μg/ml (P < 0.05); (g) decline in cathepsin D levels to values 60% lower than the untreated controls (P < 0.025); and (h) reduction in plasma norepinephrine levels to base line at 4 h (P < 0.005). Although the mode of PGI2 action is not clear, it is effective in the treatment of experimental endotoxemia.
Michael M. Krausz, Takayoshi Utsunomiya, Giora Feuerstein, John H. N. Wolfe, David Shepro, Herbert B. Hechtman
3,3′,5′-triiodothyronine, (rT3), is easily measured in human amniotic fluid (AF) during the second and third trimesters. To determine if AF rT3 levels are maintained by either maternal or fetal thyroid function, or both, models of fetal hypothyroidism (FH), maternal hypothyroidism (MH), and combined maternal and fetal hypothyroidism (MFH) were developed in pregnant rats. Hormone analyses of maternal and fetal serum and AF were performed at term. Thyroxine (T4) and 3,3′,5-triiodothyronine (T3) were not detectable in the sera and AF of term fetuses in all groups. MFH rats were prepared by administration of methimazole to the dams, and in some experiments, by maternal thyroidectomy and a low iodine diet as well. In the MFH groups from the three experiments serum thyrotropin (TSH) was markedly elevated in the dams and in the fetuses. FH rats were prepared by administering T4 by various routes to dams treated according to the MFH protocols and serum TSH was elevated in fetal serum. Analysis of FH maternal serum T4, T3, and TSH concentrations suggested mild maternal hyperthyroidism or hypothyroidism depending upon the schedule of T4 administration. The MH groups were prepared by maternal thyroidectomy and in all experiments the fetuses had normal serum TSH concentrations. The degree of maternal hypothyroidism in the MH and MFH groups was equivalent. The mean concentration of AF rT3 in normal rats in three experiments was 28.4±2.5 ng/dl (±SEM). In the three experiments, AF rT3 was undetectable or markedly reduced in the MH and MFH rats and was normal in the FH rats. These results in the amniotic fluid could not be explained by transfer of rT3 from fetal serum to the AF because fetal serum rT3 concentrations in these various models did not correlate with AF rT3 concentration. Furthermore, infusion of large doses of rT3 in MFH dams resulted in a 35-fold elevation in maternal serum rT3 concentration, a twofold elevation in fetal serum rT3 concentration, and only a minimal increase in AF rT3. These studies demonstrated that, in the rat, the maternal thyroid has the dominant role in maintaining AF rT3, whereas little effect of fetal thyroid status on AF rT3 could be demonstrated. Transfer of maternal rT3 or of fetal rT3 derived from maternal T4 to the AF do not appear to be the mechanisms whereby the maternal thyroid maintains AF rT3.
Mohamed M. El-Zaheri, Apostolos G. Vagenakis, Lee Hinerfeld, Charles H. Emerson, Lewis E. Braverman
The frequencies and levels of antibodies to Epstein-Barr virus (EBV)-specific antigens were determined in paired sera and synovial fluids from patients with rheumatoid arthritis (RA) and in sera from patients with other connective tissue diseases; i.e., systemic lupus erythematosus, progressive systemic sclerosis, and osteoarthritis (OA). The specimens were also tested for the presence of antibodies to RA-associated nuclear antigen. Compared to healthy controls, the patients' sera showed increased frequencies of elevated antibody titers (≥320) to Epstein-Barr viral capsid antigen, a correspondingly enhanced (twofold to threefold) geometric mean titer, and an increased frequency of antibodies at elevated titers (≥10), usually to the restricted component and rarely the diffuse component of the early antigen complex. Levels of antibody to the EBV-associated nuclear antigen were within the normal range. Enhancement of antibody titers was more pronounced in seropositive RA patients (i.e., positive for rheumatoid factor) than in those who were not. Enhancement was also found in systemic lupus erythematosus and progressive systemic sclerosis. Antibody to RA-associated nuclear antigen was detected at an increased frequency only in the group of seropositive RA patients (90%), as compared to 8-15% in the other connective tissue diseases and 6-8% in healthy controls. The antibody titers in the synovial fluids equaled or were at most twofold higher or lower than those in the sera. In addition, levels of EBV-specific antibodies were studied serially over a period of 6-10 mo in patients with RA and OA. Parameters of disease activity were determined and compared to antibody levels. EBV-specific antibodies in sera of OA patients remained constant and within normal limits throughout the study. Although EBV-specific antibodies were often elevated in RA patients, they also remained constant, with the exception of three patients, who showed gradual increases in one of the four antibodies, which did not correlate with disease activity.
Margaret A. Alspaugh, Gertrude Henle, Evelyne T. Lennette, Werner Henle
Using the standing droplet technique in the renal proximal convolution and simultaneous microperfusion of the peritubular capillaries, the zero net flux transtubular concentration difference of taurocholate (ΔCTC−) at 45 s was determined as a measure of active bile acid reabsorption in vivo. Starting with 0.1 mmol/liter taurocholate in both perfusates the control ΔCTC− of 0.042 mmol/liter fell to 0.006 mmol/liter (P < 0.001) when the Na+ concentration in the perfusates was reduced to zero. Removal of bicarbonate from the perfusates to alter pH had no influence on ΔCTC−. When glycocholate was added to the perfusates ΔCTC− was decreased, while probenecid increased ΔCTC−.
Frederick A. Wilson, Gerhard Burckhardt, Heini Murer, Gerhard Rumrich, Karl J. Ullrich
To enable its immunohistologic localization, angiotensin converting enzyme (EC 126.96.36.199) from human lung was solubilized by trypsinization and purified ∼2,660-fold to apparent homogeneity from a washed lung particulate fraction. The specific activity of pure enzyme was estimated to be 117 μmol/min per mg protein with the substrate hippuryl-l-histidyl-l-leucine. Consistent with previously described lung enzyme studies, catalytic activity was strongly inhibited by EDTA, O-phenanthroline, SQ 20,881, and SQ 14,225 and increased by CoCl2. SQ 20,881 was a somewhat more potent inhibitor than SQ 14,225, unlike rabbit lung enzyme. The Michaelis constant (Km) with hippuryl-l-histidyl-l-leucine was 1.6 mM. The molecular weight was estimated at 150,000 from sucrose density gradient centrifugation. Sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed a single polypeptide chain estimated at 130,000 daltons.
Joan Friedland, Emanuel Silverstein, Martin Drooker, Charlotte Setton
18 patients with malignant effusions were treated with continuous intraperitoneal, intrapleural, or intrapericardial infusion of methotrexate (MTX) 30 mg/m2 per d combined with simultaneous intravenous administration of leucovorin at a dose rate calculated to yield an equimolar concentration in the serum. In the serum the geometric mean steady-state MTX concentration was 0.95 microM, whereas it was 24 microM in the peritoneal, 213 microM in the pleural, and 434 microM in the pericardial cavities. Mean clearance was 6.6 ml/min from the peritoneal cavity, 2.6 ml/min from the pleural cavity, and 0.14 ml/min from the pericardial cavity. Leucovorin provided sufficient protection to allow the duration of infusion to be escalated from 24 to 120 h before myelosuppression was encountered. Marrow thymidylate synthetase activity was inhibited by an average of 46% compared to 86% inhibition in malignant cells in the effusions. Flow cytometric analysis showed no perturbation of the cell cycle phase distribution of marrow cells. All eight of the evaluable patients have responded: three received no other form of therapy, five also received systemic hormonal or chemotherapy. This study demonstrated that tumors confined to third space body fluids can be given very high concentration x time exposures to MTX with minimal systemic toxicity.
S B Howell, B B Chu, W E Wung, B M Metha, J Mendelsohn
At 48 h after bilateral nephrectomy in rats there is a two- to threefold increase in the number of adrenal angiotensin II receptors and a decrease in Kd of smooth muscle angiotensin II receptors. These changes have been attributed to the absence of circulating angiotensin II. Serum K+, which increases after nephrectomy may be an important and overlooked modulator. Therefore, the present experiments were designed to assess the role of K+ as a regulator of angiotensin II receptors after nephrectomy. Serum K+ was controlled with Na polystyrene sulfonate (Kayexalate), a resin designed to exchange Na+ for K+ in the gastrointestinal tract. Acutely nephrectomized rats were divided into two groups: experimental animals received Kayexalate resin every 12 h for four doses, and controls received Kayexalate exchanged with KCl in vitro before gavage. There was a significant positive correlation serum K+ and aldosterone (r = 0.78, P less than 0.001). Kayexalate maintained a normal serum K+ of 5.9 +/- 0.2 meq/liter (n = 27), aldosterone 25 +/- 3 ng/dl (n = 27) and adrenal receptor concentration of 934 +/- 156 fmol/mg protein (n = 4). Control animals had significantly higher serum K+ of 10.5 +/- 0.4 meq/liter (n = 23), aldosterone 435 +/- 32 (n = 23), and adrenal receptors of 2726 +/- 235 fmol/mg protein (n = 4). There was a linear relationship between serum K+ and number of adrenal receptors (r = 0.87). No such relationship was present in uterine smooth muscle. Therefore, these studies demonstrate that K+ modulates the number of adrenal but not smooth muscle angiotensin II receptors after nephrectomy. This is the first evidence that potassium modulates angiotensin II receptors independently of changes in angiotensin II blood levels.
J G Douglas
The plasma concentrations of dehydroepiandrosterone, androstenedione, and dehydroepiandrosterone sulfate decrease during the first year of life, remain low during childhood, and then increase during adrenarche. To determine whether alterations in adrenal enzyme activity might explain the changing secretory pattern of the adrenal androgens, we measured human adrenal microsomal 3 beta-hydroxysteroid dehydrogenase-isomerase, 17,20-desmolase, 17-hydroxylase, and 21-hydroxylase activities. 12 adrenals from individuals aged 3 mo to 60 yr were studied. The patients were divided into three groups based upon the age of the patient when the adrenal glands were obtained: group 1, infants aged 3--8 mo (n = 3); group 2, preadrenarchal or early adrenarchal children aged 2--9 yr (n = 4); and group 3, adults aged 20--60 yr (n = 5). The mean activity of the 17,20-desmolase, 17-hydroxylase, and 21-hydroxylase fell by 50% and that of 3 beta-hydroxysteroid dehydrogenase-isomerase activity rose 80% from group 1 to 2. A fourfold increase in 17,20-desmolase (P less than 0.002) and 17-hydroxylase (P less than 0.001) activity and a doubling in 21-hydroxylase activity (P less than 0.005) occurred between groups 2 and 3. We conclude that the decline in plasma adrenal androgens after birth appears to be associated with a rise in 3 beta-hydroxysteroid dehydrogenase-isomerase and a fall in 17,20-desmolase and 17-hydroxylase activity. The subsequent increase in plasma adrenal androgen concentration during adrenarche is coincident with a rise in 17,20-desmolase and 17-hydroxylase activity.
R J Schiebinger, B D Albertson, F G Cassorla, D W Bowyer, G W Geelhoed, G B Cutler Jr, D L Loriaux
Although the liver is the major site of erythropoietin (Ep) production in the fetus, this function is assumed by kidneys in the adult. The mechanisms underlying the liver to kidney switch of Ep formation are not understood. We studied the natural progression of this transition in sheep by measuring Ep production in response to anemia in normal and bilaterally nephrectomized fetal and newborn sheep beginning at about 80 d gestation (normal gestation: 140 d). Removal of both kidneys before induction of anemia did not affect Ep formation up to about 120 d of gestation. A significant reduction (29%, P < 0.02) in Ep synthesis was first noted at about 130 d of gestation (initiation of switch). This level of nephrectomy-induced reduction of Ep formation persisted until about 15 d after birth. Thereafter, bilateral nephrectomy caused further significant decreases (P < 0.05) in Ep production, gradually resulting in near total absence of Ep production at about day 40 postpartum (completion of switch). Chronic administration of testosterone (12 mg/wk) or estradiole benzoate (1.5 mg/d, 5 d/wk) to the fetus/newborn beginning at 85-90 d of gestation enhanced or suppressed erythropoiesis, respectively, but failed to affect the time at which the liver to kidney switch was initiated and/or completed. By contrast, a significant delay (P < 0.001) in the onset, but not completion of the switch occurred in animals that were either thyroidectomized or rendered chronically anemic beginning in the second third of the gestation period. Administration of thyroxin (1.2 mg/d, 5 d/wk) to thyroidectomized fetus/newborns not only prevented the delay in the initiation of the switch, but also accelerated the rate at which the switch was completed. These results demonstrate that in sheep (a) the liver to kidney switch of Ep production is initiated in utero during the last third of the gestation period, but is completed after birth, (b) this transition occurs gradually; the assumption of Ep producing capacity by the kidney is not preceded by an abrupt loss of hepatic Ep formation; and (c) the switch is not affected by changes in sex hormone levels during the prenatal-postnatal growth periods, but is profoundly influenced by alterations in thyroid hormone and oxygen supply-demand levels.
Esmail D. Zanjani, Joao L. Ascensao, Philip B. McGlave, Mussa Banisadre, Robert C. Ash
The mechanism responsible for K transport in the mammalian colon is controversial. Experiments were performed to determine whether K secretion involves active as well as passive driving forces in controls and in animals with a marked increase in K secretion. In these experiments a steady-state solution was established in proximal and distal colon of both control rats and animals fed a K-enriched diet during in vivo luminal perfusion, to compare the observed luminal [K] with predicted equilibrium [K] when net water and electrolyte movement approached zero. Transmural potential difference was measured simultaneously. A difference between the predicted equilibrium and observed luminal [K] under this condition indicates active transport. In controls the observed [K] of 20 mmol/liter in proximal colon markedly exceeded the predicted value of 6.2 +/- 0.3, mean +/- SE, indicating active secretion. In contrast, the observed [K] in distal colon of 5 mmol/liter was less than predicted (10.0 +/- 1.0), suggesting active absorption. In K-loaded animals active K secretion was demonstrable and increase above control in both segments of colon. In proximal colon the observed [K] rose to 40 mmol/liter, compared to a predicted value of 7.2 +/- 0.3, whereas in distal colon the observed [K] was 50 mmol/liter vs. a predicted value of 7.0 +/- 0.8. These studies suggest active K secretion in proximal, but not in distal colon of control animals. Further, these data suggest that the increase in the capacity for K secretion that occurs in response to chronic K loading involves stimulation of an active mechanism in both proximal and distal colon.
A S Kliger, H J Binder, C Bastl, J P Hayslett
The purpose of this study was to determine the locus of interaction of angiotensin peptides with the sympathetic nervous system leading to alterations in jejunal sodium and water transport. At low physiological doses, angiotensin II (AII) stimulates jejunal sodium and water absorption, while at high doses peptide inhibits absorption and/or stimulates secretion. Both the stimulation of jejunal transport and the inhibition of absorption were expressed in adrenalectomized rats. However, the stimulation of jejunal water absorption was abolished and a potentiated inhibition of transport was expressed in peripherally sympathectomized rats (intact adrenal medulla) and in normal rats after administration of guanethadine, phentolamine, and prazosin. The angiotensin analog (Sar1 Leu8)-AII has low efficacy and is a potent competitive antagonist of the parent peptide in pressor and myotropic systems, but is a full agonist with even greater potency than AII in stimulating jejunal transport. The increased water transport in response to (Sar1 Leu8)-AII is not secondary to enhanced renal renin release, as the analog also stimulated jejunal transport in the presence of captopril and after bilateral nephrectomy. The stimulation of absorption in response to (Sar1 Leu8)-AII alone or together with AII was abolished by phentolamine. These data demonstrate that AII-increased intestinal absorption is secondary to the release of norepinephrine from nerve endings in the jejunum and that AII inhibition of absorption is not mediated by the sympathetic nervous system. The analog (Sar1 Leu8)-AII is a full agonist in the stimulation of jejunal transport (increased norepinephrine release), but antagonizes the inhibitory response to high doses of AII. Angiotensin peptides are potent modulators of intestinal sodium and water absorption.
N R Levens, M J Peach, R M Carey
To explore the control of thyroid hormone metabolism in brain during maturation, we have measured iodothyronine deiodination in homogenates of rat cerebrum, cerebellum, and hypothalamus from 1 d postnatally through adulthood. Homogenates were incubated with 125I-l-thyroxine (T4) + [131I]3,5,3′-l-triiodothyronine (T3) + 100 mM dithiothreitol. Nonradioactive T4, T3, and 3,3′,5′-triiodothyronine (rT3) were included, as appropriate. The net production rate of [125I]T3 from T4 in 1-d cerebral homogenates was similar to the rate in adult cerebral homogenates (9.9±2.5[SEM]% vs. 8.9±1.2% T4 to T3 conversion in 2 h). Production of T3 was not detectable in 1-d cerebellar and hypothalamic homogenates. The net T3 production rate in adult cerebellar homogenates was twice as great as, and that in adult hypothalamic homogenates similar to, the rate in cerebral homogenates.
Michael M. Kaplan, Kimberlee A. Yaskoski
Myocardiopathy is common in uremia, but its cause in unknown. Excessive entry of calcium in heart cells by catecholamines has been shown to cause necrosis of myocardium. The high blood levels of parathyroid hormone (PTH) in uremia may also enhance entry of calcium into heart cells and exert deleterious effects on the heart. We examined the effect of PTH on rat heart cells grown in culture. Both amino-terminal (1-34) PTH and intact (1-84) PTH, but not the carboxy-terminal (53-84) PTH produced immediate and sustained significant rise in beats per minute and the cells died earlier than control. The effect was reversed if PTH was removed from medium, and was abolished by inactivation of the hormone. There was a dose-response relationship between both moieties of PTH and the rise in heart beats, but the effect of 1-84 PTH was significantly greater than that of 1-34 moiety. PTH stimulated cyclic AMP production within 1 min, and cyclic AMP remained significantly elevated thereafter. The effect of PTH required calcium, was mimicked by calcium ionophore, was prevented by verapamil and was not abolished by alpha- or beta-adrenergic blockers. PTH action was additive to phenylephrine and synergistic with isoproterenol. Sera from uremic parathyroidectomized rats did not effect heart beats, but sera from uremic rats with intact parathyroid glands or from uremic-parathyroidectomized rats treated with PTH had effects similar to PTH. Data indicate that (a) heart cell is a target organ for PTH and may have receptors for the hormone; (b) PTH increases beating rate of heart cells and causes early death of cells; (c) PTH effect appears to be due to calcium entry into heart cells; (d) the locus of action through which PTH induces calcium entry is different from that for catecholamines; and (e) uremic serum has no effect unless it contains PTH. Data suggest that myocardial damage may occur in uremia due to prolonged exposure to very high blood levels of PTH, and assign new dimensions to PTH toxicity in uremia.
E Bogin, S G Massry, I Harary
Cyclosporin A (CS-A), a selective inhibitor of T lymphocytes, is reported here to prevent S antigen (S-Ag) induced uveitis in Lewis rats. The S-Ag, found in all mammalian retinas, is uveitogenic under experimental conditions and patients with certain uveitic entities demonstrate cell mediated responses to this antigen. Daily treatment with CS-A (10 mg/kg) begun on the same day as S-Ag immunization totally inhibited the development of the uveitis in this experimental autoimmune model. Moreover a greater CS-A dose (40 mg/kg) efficiently prevented the disease process when therapy was started 7 d after S-Ag immunization. Anti-S-Ag antibody titers were observed to be similar in rats either protected or not protected with CS-A. Our data support strongly the need for T cell participation in this disease model. Since ocular inflammatory disease is an important cause of visual impairment, the data further suggest that CS-A may be useful in the treatment of patients with intractable uveitis.
R B Nussenblatt, M M Rodrigues, W B Wacker, S J Cevario, M C Salinas-Carmona, I Gery
We have recently demonstrated enhanced alpha-adrenergic responsiveness assessed electrophysiologically in ischemic and reperfused myocardium. This study was performed to determine whether ischemia alters alpha 1-adrenergic receptor number (Bmax) of affinity (KD) based on [3H]prazosin binding. Within 30 min after occlusion, Bmax increased in ischemic regions to 207% of control to 27 +/- 2 fmol/mg protein, with the increase persisting (+ 141% of control) during early reperfusion (2 min), before returning to control base-line values (13 +/- 1.6) after 15 min of reperfusion. KD was not altered at any interval studied. Beta receptor number of ([3H]dihydroalprenolol) and Na+-K+ ATPase activity were comparable in control compared to ischemic myocardium although beta-receptor Bmax and KD in both regions decreased during early reperfusion. Thus, the enhanced alpha-adrenergic responsivity previously recognized with ischemia and reperfusion is correlated with an increase in alpha 1-adrenergic receptors.
P B Corr, J A Shayman, J B Kramer, R J Kipnis