To gain insight into a possible role for antibody-dependent cell-mediated cytotoxicity in vivo, we examined the ability of leukocytes from 28 patients with primary immunodeficiency and from 20 normal controls to lyse three different types of antibody-coated targets in vitro. Mean cytotoxic indices ±1 SD elicited by unfractionated mononuclear cells from normal controls were 28.74±13.26 for human HLA antibody-coated lymphocyte targets, 42.79±8.27 for rabbit IgG antibody-coated chicken erythrocyte targets, and 47.58±10.34 for human anti-CD (Ripley)-coated O+ erythrocyte targets. Significantly (P=<0.05) lower than normal mean cytotoxic indices against lymphocyte targets were seen with effector cells from 10 patients with X-linked agammaglobulinemia (3.7±4.33), in 10 with common variable agammaglobulinemia (16.05±7.74), in 3 with immunodeficiency with hyper IgM (18.41±4.88), and in 2 with severe combined immunodeficiency (3.94±0.3). Antibody-dependent cytotoxicity against chicken erythrocytes was significantly (P=<0.05) lower than normal only in the common variable agammaglobulinemic group (33.33±12.3) and against human erythrocytes only in the common variable (34.36±9.59) and hyper IgM (27.54±0.66) groups. Rosette and anti-F(ab′)2 depletion studies with normal leukocytes indicated that a nonadherent, nonphagocytic, non-Ig-bearing, non-C receptor-bearing, Fc receptor-bearing lymphocyte was the only effector capable of lysing HLA antiboyd-coated lymphocyte targets. Patients with infantile X-linked agammaglobulinemia and severe combined immunodeficiency appear to have a marked deficiency in this type of effector cell function.
S. Ozden Sanal, Rebecca H. Buckley
The d- and l-isomers of glyceraldehyde are equally effective in the inhibition of SS erythrocyte sickling in vitro. The following compounds at a concentration of 20 mM were ineffective in inhibiting sickling: glyceraldehyde-3-phosphate, d-erythrose, d-ribose, d-fructose, d-glucose, d-sucrose, dihydroxyacetone, and methylglyoxal. Glyceraldehyde does not reverse the sickling of cells in the deoxy state. The properties of purified hemoglobin after treatment with glyceraldehyde and of the hemoglobin isolated from treated cells are very similar; these results suggest that glyceraldehyde itself is the reactive species within the erythrocyte. Erythrocyte glutathione is decreased by treatment in vitro with the aldehyde.
Alan M. Nigen, James M. Manning
This study was designed to compare the effect of [des-Aspartyl1]-angiotensin II ([des-Asp]-A II) and angiotensin II (A II) on blood pressure and aldosterone production in man under conditions of normal and low sodium (Na) intake. Seven normal male subjects in balance on constant normal Na intake (UNa V 160.3±5.0 meq/24 h) for 5 days received A II and [des-Asp]-A II infusions on two consecutive days; 1 mo later they were restudied after 5 days of low Na intake (UNa V 10.5±1.6 meq/24 h). Each dose was infused for 30 min, sequentially. During normal Na intake, [des-Asp]-A II from 2 to 18 pmol/kg per min increased mean blood pressure from 85.2±3 to 95.3±5 mm Hg and plasma aldosterone concentration from 5.2±1.1 to 14.3±1.9 ng/100 ml. During low Na intake, the same dose of [des-Asp]-A II increased mean blood pressure from 83.7±3 to 86.7±3 mm Hg and plasma aldosterone concentration from 34.4±6.0 to 51.0±8.2 ng/100 ml. In contrast, A II from 2 to 6 pmol/kg per min during normal Na intake increased mean blood pressure from 83.3±4 to 102.3±4 mm Hg and plasma aldosterone concentration from 7.0±2.2 to 26.8±2.0 ng/100 ml; during low Na intake, A II increased mean blood pressure from 83.0±3 to 96.0±4 mm Hg and plasma aldosterone concentration from 42.0±9.7 to 102.2±15.4 ng/100 ml. A II and [des-Asp]-A II were equally effective in suppressing renin release. Plasma cortisol and Na and K concentration did not change.
Robert M. Carey, E. Darracott Vaughan Jr., Michael J. Peach, Carlos R. Ayers
Experiments were performed on 36 plasma-expanded Munich-Wistar rats to examine the effects of acute hypercalcemia on the determinants of glomerular ultrafiltration. Elevation of total plasma calcium concentration to an average value of 13.2 +/- 0.5 mg/dl, by acute infusion of calcium chloride into nonthyroparathyroidectomized (non-TPTX) rats, resulted in significant declines in single nephron and whole kidney glomerular filtration rate. These declines were due primarily to a fall in the glomerular capillary ultrafiltration coefficient (Kf), to a mean value approximately 60% below that determined in the pre-infusion period. These changes were not seen in a separate group of sham-treated non-TPTX rats. It is of interest that these effects of acute hypercalcemia were largely abolished in rats that underwent acute TPTX before hypercalcemia. Infusion of a submaximally phosphaturic dose of parathyroid hormone, together with calcium chloride, into a second group of acute TPTX rats, however, had the effect of reproducing the striking declines in filtration rate and Kf noted in non-TPTX rats given calcium chloride alone. These findings suggest that the decline in filtration rate associated with hypercalcemia is due largely to the reduction in Kf, the latter dependent upon the presence of parathyroid hormone.
H D Humes, I Ichikawa, J L Troy, B M Brenner
Bombesin caused depolarization of rat or mouse pancreatic acinar cell membrane, reduction of membrane resistance, and a steep rise in amylase output from superfused pancreatic fragments. These effects were similar to those previously described for acetylcholine, cholecystokinin, and gastrin. The dose-response curves for these three effects of bombesin were very similar, with effects being detectable at concentrations of about 30 pM and maximal effects at about 10 nM. The equilibrium potential for the membrane action of bombesin, i.e., the membrane potential at which bombesin did not cause any change in membrane potential, was -16 mV. Similar values for equilibrium potential were obtained with acetylcholine, caerulein and pentagastrin. Bombesin in the higher dose range (10 nM) caused electrical uncoupling of acinar cells within an acinus, i.e., a marked increase in junctional membrane resistance. Similar uncoupling effects were observed after acetylcholine, caerulein, and pentagastrin stimulation. In conclusion, bombesin acts on the pancreatic acinar plasma membrane in exactly the same way as acetylcholine and cholecystokinin-pancreozymin. The electrical uncoupling caused by stimulation is evidence for an increase in cytosol free calcium ion concentration.
N Iwatsuki, O H Petersen
Cobalamin (Cbl; vitamin B12) malabsorption in pancreatic insufficiency can be partially corrected by bicarbonate and completely corrected by pancreatic proteases but the mechanisms involved are unknown. Because saliva contains enough R-type Cbl-binding protein (R protein) to bind all of the dietary and biliary Cbl, it is possible that R protein acts as an inhibitor of Cbl absorption and that pancreatic proteases are required to alter R protein and prevent such inhibition. To test this hypothesis we studied the ability of R protein and intrinsic factor (IF) to compete for Cbl binding and ability of pancreatic proteases to alter this competition.
Robert H. Allen, Bellur Seetharam, Elaine Podell, David H. Alpers
Hodgkin's disease (HD) is associated with a deficit in T-cell immunity characterized by skin test anergy and decreased lymphocyte responses to phytohemagglutinin (PHA). To investigate this mitogen hyporesponsiveness in HD, we separated peripheral blood mononuclear cells on Ficoll-Hypaque gradients and determined their response to various suboptimal concentrations of PHA. As was expected, patients with HD demonstrated marked mitogen hyporesponsiveness relative to normal controls; however, if the cell suspensions were first passed through glass wool columns to remove adherent cells, the PHA responsiveness of the hyporesponsive HD cells was markedly increased. In contrast, the responsiveness of normal controls was decreased so that the responses of nonadherent normal and HD cells were statistically indistinguishable. Evidently, a glass wool-adherent suppressor cell had been removed from patients with HD, while a glass wool-adherent cell which enhanced mitogenic responses had been removed from normal controls during column passage. Previous to column depletion, patients with HD had decreased proportions of E-rosettes and increased proportions of cells with surface α-fetoprotein; however, the proportion of these cells was not changed after column passage. Significant changes with column depletion of glass wool-adherent cells in HD were recorded in the proportions of monocytes (13.2 vs 5.8%) and lymphocytes with C-3 receptors (12.6 vs. 7.8%). The only significant change in normal controls was a decrease in the proportion of monocytes (10 vs. 1.7%). To determine if glass-adherent cells would have a suppressor effect, HD-adherent cells were added in progressively increasing numbers to mononuclear cell suspensions depleted of glass wool-adherent cells. PHA responsiveness returned toward predepletion levels. In summary, patients with HD possess a glass wool-adherent suppressor cell which is responsible at least in part for in vitro mitogen hyporesponsiveness.
W. L. Sibbitt Jr., A. D. Bankhurst, R. C. Williams Jr.
We have compared the ability of human serum and peripheral lymph to suppress the activity of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMG-CoA reductase), to activate cholesteryl ester synthesis, and to compete with 125I-labeled low density lipoprotein (LDL) for binding to LDL receptors in cultured human fibroblasts. Whole lymph was active in all three tests and the activity per unit volume in lymph was approximately equal to 1/10th that in serum. All three biologic activities in lymph were confined to the d less than 1.063 g/ml fraction. Whole lymph had no significant effect on HMG-CoA reductase activity in fibroblasts from a patient with homozygous familial hypercholesterolemia, whose cells lack LDL receptors. The LDL-like biologic activity per unit mass of immunologically active apoprotein B was approximately the same in lymph as in serum. The current data indicate that functionally active LDL is present in lymph and that the concentration of this lipoprotein is approximately equal to 1/10th that in serum.
D Reichl, N B Myant, M S Brown, J L Goldstein
To examine the electrostatic effects of fixed negative charges on the glomerular capillary wall, polydisperse [3H]DEAE dextran, a polycationic form of dextran, was infused into 10 Munich-Wistar rats. Fractional clearances of DEAE ranging in radius from 18 to 44Å were determined in these rats, together with direct measurements of the forces and flows governing the glomerular filtration rate of water. These results were compared with data previously obtained in Munich-Wistar rats receiving tritiated neutral dextran (D) and polyanionic dextran sulfate (DS). Measured values for the determinants of the glomerular filtration rate of water in rats given DEAE were found to be essentially identical to those in rats given either D or DS. In addition, DEAE was shown to be neither secreted nor reabsorbed. Fractional clearances of polycationic DEAE were increased relative to both D and DS, the increase relative to D being significant for effective molecular radii ranging from 24 to 44Å.
Michael P. Bohrer, Christine Baylis, H. David Humes, Richard J. Glassock, Channing R. Robertson, Barry M. Brenner
Pulmonary fibrosis was induced in eight baboons with bleomycin; five untreated animals were controls. After 45-65 U/kg of bleomycin, lung volumes and diffusing capacity were reduced, and static lung pressure-volume curves were shifted to the right. Right middle lobes were resected at this time in five bleomycin-treated and two control animals. Compared to controls, right middle lobes from bleomycintreated animals had increased weight and contained increased amounts of total protein, collagen, elastin, and DNA; synthesis of collagen and noncollagen protein were also elevated. Occasional alveolar septae were edematous and infiltrated by mononuclear inflammatory cells; a slight increase in collagen was demonstrable histologically. Four of six treated animals died with extensive diffuse interstitial fibrosis after 95 U/kg of bleomycin. Biochemical analyses revealed significantly elevated lobar contents of dry weight, protein, elastin, and collagen. Two animals survived 95 U/kg of bleomycin and were terminated 6 mo after treatment. In these animals, physiologic studies were indicative of restrictive lung disease, but lung histology was nearly normal. Lung weight, total protein, and DNA had returned to control values, but collagen and elastin were increased in amount and concentration. Bleomycin induces an intense inflammatory response in the lung. During this inflammation, connective tissue proliferation occurs in concert with proliferation of other tissue components. Cessation of bleomycin treatment is followed by resolution of inflammation manifested by decreases in tissue mass, cellular content, and nonconnective tissue protein. Collagen and elastin deposited during inflammation are less successfully removed during resolution, leading to a stage characterized by increased concentrations of these proteins. A similar sequence of tissue alterations may occur in idiopathic diffuse interstitial fibrosis of man in response to various lung injuries.
We have recently described a new animal model of arthritis induced by intradermal injection of a distinct type of collagen found in cartilage (type II collagen). Since immunologic sensitivity to collagen could play a role in the pathogenesis of this type II collagen-induced arthritis in rats, the ability of purified types of native collagens to induce cellular and humoral responses was quantified by antigeninduced tritiated thymidine incorporation into lymphocytes by collagen and passive hemagglutination, respectively. Rats injected intradermally with native heterologous or homologous type II collagens in adjuvant developed type-specific cellular as well as humoral reactivity. Types I and III collagens were less immunogenic than was type II. The latter collagen induced brisk cellular and humoral responses that were equivalent whether complete Freund's adjuvant or incomplete Freund's adjuvant were employed. Both responses could be induced by native type II collagens modified by limited pepsin digestion, indicating that they are not attributable to determinants in the telopeptide regions of the molecule. Thus, these studies demonstrate the unique immunogenic as well as arthritogenic properties of the type II collagen molecule and indicate that both result from a helical conformation of its structurally distinct α-chains. Further, they suggest that type II collagen may, by humoral or cellular mechanisms, provoke or perpetuate inflammation in other arthritic diseases.
David E. Trentham, Alexander S. Townes, Andrew H. Kang, John R. David
The effect of corticosteroids and cytotoxic chemotherapeutic agents on the excretion of Bence Jones protein was determined for periods of 1 - 62 mo in 29 patients with multiple myeloma and Bence Jones proteinuria. The amount of protein present in 24-h urine specimens collected before treatment and at frequent intervals during monthly treatment cycles was determined. Striking variations occurred in the amount of Bence Jones protein excretion; these changes were especially evident when 75 mg of prednisone were given daily for 7 days as part of a monthly chemotherapeutic regimen. Within the 7-day period seven patients showed essentially no decrease (<25%), whereas 13 and 9 patients had a moderate decrease (25-75%) or a marked decrease (>75%), respectively, in Bence Jones proteinuria as compared to pre-treatment values. The decrease in excretion of Bence Jones protein during this period was attributed mainly to corticosteroid therapy because of the transient nature of the response in most patients and the lack of such response in three patients when the hormone was omitted. Biosynthetic studies were performed to determine in vitro the effect of corticosteroids on Bence Jones protein synthesis. Plasma cells obtained from the bone marrow of 13 patients were incubated in a growth medium containing 14C-labeled lysine and isoleucine and prednisone in concentrations up to 240 μg/ml, and the amount of Bence Jones protein synthesized was determined immunochemically. No differences in viability were apparent between untreated and prednisone-treated cells. The type of response exhibited by an individual patient in the percent decrease of Bence Jones protein excreted after 7 days of prednisone treatment was comparable to the percent decrease in newly-synthesized Bence Jones protein secreted by tumor cells when cultured in the presence of prednisone at a concentration of 120 μg/ml. The marked differences in the capacity of corticosteroids to affect Bence Jones protein synthesis appear to reflect a biochemical heterogeneity among plasma cell neoplasms.
Continuously recorded bipolar electrograms were obtained simultaneously from epi-, endo-, and mid-myocardial regions of the ischemic and normal zones of cat left ventricle in vivo after coronary occlusion, analyzed by computer, and compared to regional cyclic AMP levels. Regional cyclic AMP content was used as an index of the combined local effects of: (a) efferent sympathetic nerve discharge; (b) release of myocardial catecholamines due to ischemia; and (c) circulating catecholamines. Ischemia resulted in a progressive increase in pulse width and rise time and a decrease in rate of rise of voltage (dV/dt) of the local electrograms from ischemic zones reaching a maximum within 2.4±0.3 min (mean±SE) at the time of onset of severe ventricular dysrhythmias, all of which returned toward control before the cessation of the dysrhythmia (33.5±1.5 min after coronary occlusion). Increases in cyclic AMP in ischemic zones preceded corresponding increases in the frequency of premature ventricular complexes (PVCs). Propranolol inhibited the increases in cyclic AMP and reduced the frequency of PVCs in animals without ventricular fibrillation. In animals with ventricular fibrillation, cyclic AMP was significantly elevated in normal and ischemic zones compared to animals with PVCs only. Electrical induction of PVCs or ventricular fibrillation in ischemic and nonischemic hearts failed to increase cyclic AMP. The results suggest that the changes in regional adrenergic stimulation of the heart may contribute to perpetuation of ventricular dysrhythmia and the genesis of ventricular fibrillation early after the onset of myocardial ischemia.
Peter B. Corr, Francis X. Witkowski, Burton E. Sobel
The absence of normal high density lipoproteins (HDL) in Tangier disease is well established, but the properties of very low density lipoproteins (VLDL) and low density lipoproteins (LDL) in this disorder have not been well defined. The profiles obtained by analytic ultracentrifugation and the chemical composition, morphology, and electrophoretic mobility of Tangier and normal VLDL and LDL were compared. Apolipoproteins were fractionated by gel chromatography and characterized by amino acid analysis, polyacrylamide-gel electrophoresis, and immunochemical reactivity.
Robert J. Heinen, Peter N. Herbert, Donald S. Fredrickson
We have used purified, 125I-labeled human transcobalamin II (TC II), saturated with cobalamin (Cbl), to study the uptake process for the TC II-Cbl complex by intact normal cultured human skin fibroblasts. We have also investigated the possibility that a defect in one step of this process underlies that inborn error of Cbl metabolism—designated cbl C—in which mutant cells are unable to retain Cbl intracellularly or convert it to its coenzyme forms. TC II-Cbl binding at 4°C reached a plateau after 3-4 hr; 95% of the bound 125I was releasable with trypsin. Binding of TC II-Cbl at 4°C could be inhibited by human and rabbit TC II-Cbl and human TC II devoid of Cbl but not by other Cbl-binding proteins, albumin, or free Cbl. Specific binding reached saturation at ≅5 ng TC II/ml (0.13 nM) and could be inhibited by ethylene glycol-bis (β-aminoethyl ether) N,N,N′,N′- tetraacetic acid. At 37°C, the TC II-Cbl complex was internalized as shown by a progressive decrease in the trypsin-releasable fraction of bound 125I. After 2 h at 37°C, increasing amounts of acid-soluble 125I were found in the incubation medium indicating that the labeled TC II was being degraded. Chloroquine, an inhibitor of lysosomal proteolysis, prevented this degradation. The binding, internalization, and degradation of TC II-Cbl by cbl C cells was indistingusihable from that by control cells. Our studies provide additional support for the concepts: (a) that the TC II-Cbl complex binds to a specific cell surface receptor through a site on the TC II; (b) that the interaction between the receptor and TC II is calcium dependent; (c) that the TC II-Cbl is internalized via endocytosis; (d) that the degradation of TC II and release of Cbl from the complex occurs in lysosomes. We also conclude that the defect in cbl C must reside at some step beyond this receptor-mediated uptake process.
Pamela Youngdahl-Turner, Leon E. Rosenberg
After the intravenous injection of unconjugated [3H]bilirubin into normal Sprague-Dawley and Wistar R rats, radiolabeled bile pigments rapidly accumulated in the liver. By 1.5 min after injection, an average of 36% of the injected isotope was present in liver homogenates. Between 3 and 15 min, 37-64% of the total intrahepatic radiolabeled bilirubin was conjugated, as demonstrated by extraction of label into the polar phase of a solvent partition system. This indicates both rapid conjugation, and accumulation of conjugated bilirubin within the liver cell. Fluorometric determination of the dissociation constants of purified bilirubin and its mono- and diglucuronides for homogeneous preparations of two human and four rat glutathione S-transferases, including ligandin, revealed avid binding of all three bile pigments to this class of proteins. Hence, the observation that the intrahepatic bile pigment pool contains substantial amounts of conjugated bilirubin can be attributed to the high binding affinities observed. Thin-layer chromatographic analysis of the 3H-pigments produced by p-iodoaniline diazotization of homogenates and cytosol demonstrated that the intrahepatic pool of conjugated bilirubin was almost exclusively monoglucuronide. Examination of radiolabeled bilirubin conjugates excreted in bile during the first 20 min after injection of [3H]bilirubin showed no preferential excretion of diglucuronide. These studies indicate that (a) both bilirubin and its monoglucuronide accumulate within the liver cell as ligands with the glutathione S-transferase; and (b) bilirubin diglucuronide does not significantly accumulate within the general intrahepatocellular pool of protein-bound bile pigments. The latter observation is compatible with the formation and excretion of bilirubin diglucuronide directly from the canalicular pool of the liver cell.
Allan W. Wolkoff, Jeanne N. Ketley, Jeanne G. Waggoner, Paul D. Berk, William B. Jakoby
The potent synthetic androgen methytrienolone (R 1881), which does not bind to serum proteins, was utilized to characterize binding to receptors in human androgen responsive tissues. Cytosol extracts prepared from hypertrophic prostates (BPH) were utilized as the source of receptor for the initial studies. High affinity binding was detected in the cytosol of 29 of 30 samples of BPH (average number of binding sites, 45.8±4.7 fmol/mg of protein; dissociation constant, 0.9±0.2 nM). This binding had the characteristics of a receptor: heat lability, precipitability by 0-33% ammonium sulfate and by protamine sulfate, and 8S sedimentation coefficient. High affinity binding was also detected in cytosol prepared from seminal vesicle, epididymis, and genital skin but not in non-genital skin or muscle. However, similar binding was demonstrated in the cytosol of human uterus. The steroid specificities of binding to the cytosol of male tissues of accessory reproduction and of uterus were similar in that progestational agents were more effective competitors than natural androgens. Binding specificities in cytosol prepared from genital skin were distinctly different and were similar to those of ventral prostate from the castrated rat in that dihydrotestosterone was much more potent than progestins in competition. Thus binding of R 1881 to the cytosol of prostate, epididymis, and seminal vesicle has some characteristics of binding to a progesterone receptor.
Mani Menon, Catherine E. Tananis, L. Louise Hicks, Edward F. Hawkins, Martin G. McLoughlin, Patrick C. Walsh
A 70-yr-old mildly diabetic white male was discovered to have an elevated level of serum free glycerol in the range of 75 mg/dl and to excrete about 13 g of free glycerol in the urine per 24 h. During a 24-h fast the urine glycerol loss increased to 21.5 g per 24 h. Studies carried out in vitro using leukocytes prepared from the patient's blood which were incubated with [14C]glycerol demonstrated an almost complete absence of glycerol oxidation to 14CO2 and of glycerol phosphorylation, in contrast to control studies with leukocytes collected from normal subjects. Homogenates of the patient's leukocytes contained negligible activity of ATP:glycerol phosphotransferase (glycerokinase EC 18.104.22.168) as measured by a direct spectrophotometric method. Marked hyperglycerolemia has thus far been detected in one brother and in one son of the daughter of this patient. This evidence suggests an x-linked recessive inheritance pattern of the trait. There is a high prevalence of diabetes mellitus in this family.
C I Rose, D S Haines
The pattern of retrograde His-Purkinje conduction was evaluated in 28 patients using ventricular extrastimuli. In each patient progressive prolongations of His-Purkinje conduction (S2H2) which appeared as ventricular extrastimuli were induced at closer coupling intervals (S1S2). There was an inverse linear relationship of S2H2 to S1S2 which was cycle length-dependent: i.e., at any S1S2 interval the resultant S2H2 was less at shorter drive cycle lengths. The degree of S2H2 delay varied widely (from 30 to 340 ms) and was unrelated to the presence of bundle branch block, H-V intervals, or capability of ventriculoatrial conduction. Prolongation of S2H2 was independent of intraventricular (muscle) conduction delay; such delay was usually absent at most, and occasionally all, S1S2 coupling intervals during which S2H2 was lengthening. Furthermore, in two patients both left and right ventricles were activated before the timed depolarization of the His bundle occurred, demonstrating that under the stress of extrastimuli, the impulse conducts through ventricular muscle with less delay than through the His-Purkinje system. We conclude that the His-Purkinje system typically displays slow conduction response to ventricular stress. The site of this conduction delay is probably at the distal "gate".
M E Josephson, J A Kastor
Perfusion studies of the normal human jejunum were performed to test whether dihydroxy bile acids and hydroxy fatty acids inhibit the absorption of oleic acid, since previous reports documented their inhibitory effects on the absorption of several other organic solutes. 3 mM deoxycholate and 7 mM glycodeoxycholate inhibited the absorption of 3 mM oleic acid in isotonic micellar solutions while inducing net fluid secretion. Similarly, fractional absorption of oleic acid decreased in the presence of hydroxy fatty acids. However, only the changes induced by 2 mM ricinoleic acid could be distinguished from changes induced by an increase in total fatty acid concentration. Under all experimental conditions, close linear relationships existed between net water movement and fractional absorption of glucose, xylose, and fatty acids, as well as between the absorption rates of these solutes. In contrast, net fluid secretion induced by hypertonic D-mannitol (450 mosmol/liter) had no effect on solute absorption. Our data and observations in the literature do not allow formulation of a hypothesis which would adequately define all effects of dihydroxy bile acids and fatty acids on intestinal transport processes. The observations help explain the malabsorption of fat and other nutrients in patients with the blind loop syndrome.
R Wanitschke, H V Ammon
Mediator release from rat peritoneal and human lung mast cells as well as human leukemic basophils was examined to determine whether super-oxide (O−2) was concomitantly generated. Immunologic or nonimmunologic stimulation of each preparation induced parallel release of histamine and O−2 within 2 min. O−2 production was quantitated by superoxide dismutase (SOD)-inhibitable chemiluminescence and cytochrome c reduction. SOD was detected in basophil and mast cell lysates and was also released by rat mast cells stimulated by anti-IgE. Secretory granules isolated from purified rat mast cells released histamine, O−2, and SOD upon exposure to cations. Thus, both superoxide radicals and SOD may play a role in host defenses involved in immediate hypersensitivity reactions.
William R. Henderson, Michael Kaliner
The studies so far reported on the metabolic clearance rate of insulin in human diabetes mellitus have given conflicting results, probably because they have been conducted on few patients and have used a variety of experimental techniques and data treatments. We investigated the kinetics of insulin distribution and degradation in 35 normal subjects and in 42 nonketotic, nonobese, overtly diabetic patients, of whom 26 were above 40 yr old and 16 were 40 yr old or less at diagnosis. The design of the study combined (a) the use of a tracer to perturb minimally the steady state and to avoid glucose infusion; (b) the preparation of purified [125I]-monoiodoinsulin, which has a metabolic behavior similar to that of native insulin; and (c) noncompartmental analysis of the plasma immunoprecipitable 125I-insulin disappearance curves, which were recorded for 2 h after pulse i.v. injection of the tracer.
Renzo Navalesi, Alessandro Pilo, Eleuterio Ferrannini, Paolo Cecchetti, Antonio Masoni
To test the hypothesis that in both the liver and renal cortex of the fructose-loaded rat, severity of depletion of inorganic phosphate (Pi), and not the magnitude of accumulation of fructose-1-phosphate (F-1-P), determines the severity of the dose-dependent reduction of ATP, we intraperitoneally injected fed rats with fructose, 20 and 40 μmol/g, alone, and at the higher load, in combination with (a) sodium phosphate, 20 μmol/g, administered shortly beforehand or subsequently or, (b) adenosine, 2 μmol/g, administered beforehand. The following observations were made: (a) With fructose loading alone, at the higher load, both Pi and total adenine nucleotides (TAN) were reduced by one third in the renal cortex and (as previously observed) by two thirds in the liver; and at either load, the reduction of ATP (and TAN) and the accumulation of F-1-P were less severe in the renal cortex than in the liver. (b) Prior phosphate loading largely prevented the reductions of ATP and TAN in the renal cortex and significantly attenuated them in the liver, yet doubled the renal cortical accumulation of F-1-P. (c) Adenosine loading substantially attenuated the reductions of ATP, TAN, and Pi only in the renal cortex. (d) ATP varied directly with Pi (P < 0.001, r = 0.98) in the domain of control and reduced values of Pi taken from both liver and renal cortex. (e) As judged from tissue and plasma concentrations of fructose and glucose, and tissue concentrations of fructose-6-phosphate and glucose-6-phosphate, the rate at which the renal cortex and liver converted fructose to glucose was much lower at the higher fructose load. (f) Prior phosphate loading prevented this decrease in rate in the renal cortex and attenuated it in the liver; adenosine loading attenuated it only in the renal cortex. We conclude that in both the renal cortex of the fructose-loaded rat: (a) Depletion of Pi is critical to the causation of the reductions in both ATP and TAN and, at the higher fructose load, of a decrease in the rate at which ATP is regenerated. (b) The severity of depletion of Pi determines the severity of these disturbances. (c) By differentially mitigating the depletion of Pi, prior phosphate loading largely prevents these disturbances in the renal cortex, and attenuates them in the liver; and adenosine loading attenuates them only in the renal cortex.
R. Curtis Morris Jr., Kathleen Nigon, Elizabeth B. Reed
Sulfasalazine (salicylazosulfapyridine), an agent widely used for the treatment of ileitis and colitis, is also a competitive inhibitor of intestinal folate transport (1, 2). The mechanism of action of sulfasalazine remains uncertain. To further explore the mechanism of sulfasalazine action, the interaction of the drug with the folate recognition site was tested with three enzymes: dihydrofolate reductase, methylenetetrahydrofolate reductase, and serine transhydroxymethylase, each catalyzing a reaction involving a different folate derivative. Each of these enzymes was inhibited by sulfasalazine in the same concentration range as that previously observed to inhibit intestinal folate transport; the kinetic data are consistent with a competitive mode of inhibition. Specificity of inhibition was demonstrated by the finding that the reduction of the pteridine ring of pteroylheptaglutamic acid by dihydrofolate reductase was subject to inhibition, whereas the hydrolysis of the γ-glutamyl peptide side chain by chicken pancreas conjugase was not affected. These results are interpreted to indicate that sulfasalazine interferes with a folate recognition site which is common to these enzymes and to the intestinal transport system. Sulfasalazine, therefore, has certain properties of an antifolate drug.
Jacob Selhub, G. Jeelani Dhar, Irwin H. Rosenberg