Showing papers in "Journal of Clinical Investigation in 1972"

The metabolism of low density lipoprotein in familial type II hyperlipoproteinemia

Abstract: The metabolism of low density lipoprotein (LDL, beta lipoprotein) was studied in 10 normal individuals and 10 patients with familial type II hyperlipoproteinemia using purified radioiodinated LDL. Over 97% of the label was bound to the protein moiety of LDL and therefore the turnover data reflect the fate and distribution of LDL-apoprotein. Comparison of the metabolic behavior of biologically screened and unscreened labeled LDL preparations in dogs as well as the analysis of the urinary excretion of radioiodide derived from labeled LDL degradation in humans indicated that no significant denaturation resulted from the isolation, purification, and labeling techniques. The plasma concentration of LDL-cholesterol in normals was 105+/-21 mg/100 ml (mean +/-1 SD) in contrast to 254+/-47 mg/100 mg in patients with type II hyperlipoproteinemia; these values corresponded to LDL-apoprotein concentrations of 63+/-13 mg/100 ml and 153+/-30 mg/100 ml, respectively. Despite these differences in concentration, the synthetic rate of LDL-apoprotein in both groups was not significantly different (14.43+/-1.75 mg/kg per day in normals vs. 15.01+/-1.71 mg/kg per day in type II) nor was there any difference in the fraction of the total exchangeable LDL which was in the intravascular space (68.4+/-4.3% vs. 73.3+/-5.2%). However, the fractional catabolic rate of LDL in normal individuals differed significantly from that of patients with type II hyperlipoproteinemia (0.462+/-0.077/day in normals vs. 0.237+/-0.044/day in type II) and correspondingly the biological half-life of LDL was significantly prolonged (3.08+/-0.35 days normals vs. 4.68+/-0.44 days in type II). These data indicate that the pathologic elevation of plasma LDL concentration in the individuals with type II hyperlipoproteinemia studied here is due to a decreased fractional rate of LDL degradation rather than to an abnormality of LDL synthesis. This defect of catabolism may be the primary defect in type II hyperlipoproteinemia or, alternatively, may be secondary to an underlying abnormality in lipid metabolism.

The Human Rosette-Forming Cell as a Marker of a Population of Thymus-Derived Cells

Abstract: A BS T R A C T Sheep red blood cells can surround, in vitro, some human peripheral blood lymphocytes in a formation called a rosette. The number of rosetteforming cells (RFC) in 50 normal persons had a wide range (4-40%). The organs of 13 human fetuses (11-19 wk conceptional age) were examined for the presence of RFC. The thymus possessed the highest percentage of RFC, the maximum being 65% of total thymocytes in two 1516 wk fetal specimens. Blood RFC were always present and their number slightly increased in the oldest fetuses. The bone-marrow showed 0-8% in the six fetuses studied. RFC were found in the spleen around the 13th wk and in the liver around the 17th wk of gestation. These observations lead to the hypothesis that human blood RFC may be chiefly thymic derived. Studies of patients with immunological disorders support this hypothesis: one patient with Nezelof syndrome had no blood RFC and four patients with Wiskott-Aldrich syndrome had a low number of blood RFC (1 and 1.5%). Patients with acquired hypogammaglobulinemia showed a normal percentage of RFC. With the fetal thymocytes, the percentage of inhibition with anti-i, serum increased with the fetal age to become complete in the oldest fetuses studied. Incubation of the oldest fetal thymocytes or the blood lymphocytes with anti-e serum or anti-ja serulmi completely inhibited the rosette formation. These results suggest that u-chain determinants are presenit on human fetal thymocytes and blood RFC. The significance of the presence of y-chain determinants on these cells is uniclear.

The Role of Cell Swelling in Ischemic Renal Damage and the Protective Effect of Hypertonic Solute

Abstract: The failure of blood flow to return to the kidney following a transient period of ischemia has long been recognized. The cause of this "no-reflow" has been investigated in the rat after a transient period of total obstruction of the renal arteries. The vascular pattern of the kidneys as visualized with silicone rubber injection shows a diffuse patchy ischemia throughout the kidney, which persists after release of the obstructed renal artery. Electron microscopic studies of ischemic kidneys showed that all cellular elements were swollen and limiting the available vascular space. Functional studies revealed an increase in plasma urea nitrogen and creatinine after 1 hr or longer ischemic periods. The ischemia, cell swelling, "no-reflow," and subsequent renal dysfunction occurring after obstruction to the renal arteries were corrected by the administration of hypertonic mannitol, but were unaffected by an equivalent expansion of the extracellular fluid volume either with isotonic saline or isotonic mannitol, showing that the osmotic effect was primary. The hypothesis is presented that ischemic swelling of cells may occlude small blood vessels so that recirculation does not resume even after the initial cause of the ischemia is no longer present; solutes which do not penetrate cell membranes are able to shrink swollen cells, increase the available vascular space and thus permit reflow of blood to the ischemic organ.

A threshold distribution hypothesis for packet storage of insulin and its mathematical modeling

Abstract: Phases of insulin release were studied in the perfused pancreas during a variety of glucose stimulation patterns. Patterns included staircase stimulations, constant prolonged single steps, restimulations, and ramp functions. Except at low concentrations, prolonged single steps of glucose elicited early spikes of insulin and a slowly rising second phase. Total insulin in the initial spikes increased with higher glucose concentrations. However, the time-related pattern of these spikes was similar in all cases; ratios of initial secretion rate to total insulin released were constant. Total insulin released in this early phase approximated a sigmoidal function of glucose concentration; mathematical differentiation of this function gave a skewed bell-shaped distribution curve. Staircase stimulations caused insulin to be released as a series of transient spikes which did not correlate with the increment of glucose but rather to the available insulin for a given glucose concentration minus that released in previous steps. The sum of total insulin released as spikes in a staircase series leading to a given glucose concentration was the same as when that concentration was used as a single step. Interrupted prolonged glucose infusions indicated the second phase of insulin release could prime the pancreas and that the first and second phases were interrelated. When glucose was perfused as ramp functions of slow, increasing, concentration, phasic response disappeared.A previous two-compartmental model was expanded to include a threshold or sensitivity distribution hypothesis. This hypothesis proposes that labile insulin is not stored in a homogeneous form but as packets with a bell-shaped distribution of thresholds to glucose. These packets respond quickly when their threshold levels to glucose are reached or exceeded. Data from single step stimulations were utilized for constructing a mathematical model which simulated satisfactorily the various stimulation patterns.

Ventilation with end-expiratory pressure in acute lung disease

Abstract: In 10 patients with severe, acute respiratory failure we studied the effects of positive end-expiratory pressure when intermittent positive pressure ventilation (IPPV) with inspired oxygen (F(IO2)) up to 0.5 failed to maintain arterial oxygen tension (P(aO2)) above 70 torr.Positive end-expiratory pressures (PEEP) of 0, 5, 10, and 15 cm H(2)O were applied for 30-min periods each and in random order. Blood gas exchange, lung volumes, compliance, and hemodynamics were studied at each level of PEEP. P(aO2) (F(IO2) = 1.0) rose linearly with elevation of PEEP, the mean increase being from 152 to 347 torr, or 13 torr/cm H(2)O PEEP. Mean functional residual capacity (FRC) was 1.48+/-0.78 liters at zero PEEP (i.e., IPPV) and the increase was essentially linear, reaching 2.37 liters at 15 cm H(2)O PEEP. P(aO2) and FRC showed a close correlation. Total and lung static compliance were greater during ventilation with high than with low levels of PEEP. The increase in P(aO2) correlated with the specific lung compliance. Dynamic lung compliance decreased progressively with rising levels of PEEP except for an increase with 5 and 10 cm H(2)O PEEP in patients with initial values of 0.06 liter/cm H(2)O or higher. Cardiac index fell in some patients and rose in others and there was no correlation of mean cardiac index, systemic blood pressure, or peripheral vascular resistance with level of PEEP. The most probable explanation for the effect of PEEP on P(aO2) and compliance is recruitment of gas exchange airspaces and prevention of terminal airway closure.

The neutrophilic leukocyte in wound repair: A study with antineutrophil serum

Abstract: The role of the neutrophilic leukocyte in wound healing was investigated by observing the progress of repair in the absence of these cells. Circulating neutrophils were eliminated in guinea pigs by the administration of antineutrophil serum (ANS) 24 hr before wounding and by daily injections throughout a 10 day period of healing. Control animals received normal rabbit serum at the same dose levels and times. The wounds consisted of six linear incisions in the dorsal skin of the animals.The contents of 24-hr neutropenic and control wounds were compared by quantitating the major cellular and extracellular wound components using a histometric technique. At 24 hr, there were no differences between control and neutropenic wounds in the per cent of total wound volume occupied by mononuclear leukocytes and fibrin. The neutropenic wounds had no neutrophils, had a significantly decreased volume of fluid space, and an increased volume of red cells, as compared with controls. The differences in numbers of erythrocytes and amount of fluid space in these two sets of wounds may be related to substances within neutrophils that promote lysis of erythrocytes or affect vascular permeability. In spite of the lack of neutrophils in the ANS-treated animals during the 10 days of healing, no differences were observed between the control and neutropenic wounds relative to the rate of wound debridement or the extent of repair. The wounds from the two groups of animals were identical in cellularity and degree of connective tissue formation. These observations support the notion that neither wound debridement nor the formation of granulation tissue are dependent upon the presence of neutrophils. A neutrophil response in early wounds is not an essential antecedent to the infiltration of monocytes, as suggested by previous investigations.

Coronary Artery Reperfusion

Abstract: The effects of coronary artery reperfusion 3 hr after coronary occlusion on contractile function and the development of myocardial damage at 24 hr was studied experimentally. In 14 control and 6 reperfused dogs, relationships between epicardial ST segment elevation 15 min after coronary occlusion and myocardial creatine phosphokinase activity (CPK) and histologic appearance 24 hr later were examined. The electrocardiograms were recorded from 10 to 15 sites on the left ventricular epicardium and transmural samples for CPK and histology were obtained from the same sites where epicardial electrocardiograms had been recorded. An inverse relation existed between ST segment elevation (mv) 15 min after occlusion and log CPK activity (IU/ mg of protein) 24 hr later, log CPK = - 0.06ST + 1.26. In dogs subjected to coronary artery reperfusion, there was significantly less CPK depression (log CPK = - 0.01ST + 1.31, [P < 0.01]) than that expected from the control group. In the control group 97% of specimens showing ST segment elevations over 2 mv at 15 min showed abnormal histology 24 hr later. In contrast, in the reperfused group 43% of sites exhibiting elevated ST segment at 15 min showed abnormal histology 24 hr later. In six additional dogs it was shown that the paradoxical movement of the left ventricular wall could be reversed within 1 hr of perfusion. Therefore, by enzymatic and histologic criteria, as well as by functional assessment, coronary artery reperfusion 3 hr after occlusion resulted in salvage of myocardial tissue.

Predominance of histocompatibility antigen HL-A8 in patients with gluten-sensitive enteropathy.

Abstract: HL-A phenotypes were determined in 24 unrelated patients with gluten-sensitive enteropathy (GSE) using a lymphocyte microcytotoxicity test 21 of the 24 patients had HL-A8 in the second segregant series, a frequency of 0875 In contrast, the HL-A8 frequency in 200 normal individuals was 0215 (difference significant at P < 0002), and in 6 patients with villous atrophy due to tropical sprue or hypogammaglobulinemia the HL-A8 frequency was 017 (difference from normal not significant) The HL-A types in the families of three HL-A8 positive patients with GSE indicated that the HL-A8 antigen was inherited as an autosomal dominant Frequencies of the other HL-A antigens in the GSE group did not differ significantly from that of the normal group These findings are compatible with the hypothesis that GSE is due to the presence of an abnormal "immune response (Ir) gene," leading to the production of pathogenic antigluten antibody or, alternatively, to the presence of a particular membrane configuration leading to the binding of gluten to epithelial cells with subsequent tissue damage

Abnormal Bactericidal, Metabolic, and Lysosomal Functions of Chediak-Higashi Syndrome Leukocytes

Abstract: Phagocytic, antimicrobial, and metabolic functions were studied in leukocytes obtained from three patients with the Chediak-Higashi syndrome (CHS) and compared to normals, individuals, heterozygous for Chediak-Higashi syndrome, and two subjects with chronic granulomatous disease of childhood (CGD). Chediak-Higashi syndrome leukocytes showed normal ingestion of a variety of bacteria, Candida albicans, and polystyrene latex particles. Intracellular destruction was significantly impaired for Staphylococcus aureus, Group D streptococci, and a rough strain of Type II pneumococci over a 2 hr incubation. Killing of Serattia marcescens was consistently delayed at 1 hr whereas that of Escherichia coli and C. albicans appeared normal, unless the incubations were shortened to 20 min. Examination of the rates of killing indicated that the greatest defect occurred in the first 20 min of contact between Chediak-Higashi syndrome cells and bacteria. Separation of Chediak-Higashi syndrome granulocytes from monocytes revealed that the former were most defective in bactericidal activity. After phagocytosis, Chediak-Higashi syndrome granulocytes displayed a normal burst in oxygen consumption and oxidation of glucose-1-(14)C and glucose-6-(14)C and formate-(14)C. Oxidation of glucose-1-(14)C by non-phagocytizing Chediak-Higashi syndrome granulocytes and monocytes averaged 2-3 times normal, whereas glucose-6-(14)C and formate-(14)C oxidation were not significantly increased by resting cells. Iodination of intracellular protein by Chediak-Higashi syndrome leukocytes was significantly increased above normal in both the resting and phagocytizing state. Electron microscopic histochemistry revealed that almost all peroxidase activity was localized to the giant granules in Chediak-Higashi granulocytes, and after bacterial ingestion there was a failure of delivery of peroxidase to many phagosomes. Upon longer incubation more phagosomes acquired peroxidase activity, presumably through a fusion process, although many giant granules remained intact. The contrasting patterns and kinetics of the killing defects and the differing metabolic properties of Chediak-Higashi syndrome and chronic granulomatous disease leukocytes emphasize the pleiomorphic nature of inherited disorders of leukocyte function.

The Amino Acid Sequence of a Major Nonimmunoglobulin Component of Some Amyloid Fibrils

Abstract: A B S T R A C T The coxnplete amino acid sequence of a protein, acid soluble fraction, (ASF) which constitutes up to 50% of amyloid fibrils from a patient with familial Mediterranean fever has been obtained. Partial amino acid sequences of three other proteins from patients with secondary amyloidosis were identical in the regions studied except for an alanine-valine interchange in one. The ASF contains no cysteine, does not resemble any known immunoglobulin, and has not been detected as yet in myeloma-associated amyloid.

The role of low pressure baroreceptors in reflex vasoconstrictor responses in man

Abstract: Studies were performed on 11 healthy men to evaluate the role of low pressure baroreceptors in the reflex forearm vasoconstrictor responses (plethysmography) to venous pooling produced by lower body negative pressure. Lower body negative pressure (LBNP) at - 5, - 10, - 20, and - 40 mm Hg lowered central venous pressure by 42, 59, 74, and 93%, respectively, and decreased forearm vascular conductance by 24, 29, 34, and 40%, respectively. The decreases in forearm blood flow and conductance during the low levels of venous pooling (LBNP - 5 and - 10 mm Hg) occurred without significant changes in arterial pressure, arterial dP/dt. and heart rate. These results with the low levels indicate that maneuvers which decrease venous return and central venous pressure in man can influence forearm vascular tone without significant changes in the determinants of carotid and aortic baroreceptor activity. During high levels of venous pooling (LBNP - 20 and - 40 mm Hg), significant decreases in arterial pressure and dP/dt and significant increases in heart rate accompanied the further reductions in central venous pressure, forearm blood flow, and forearm vascular conductance. About 73% of the decrease in conductance during venous pooling at LBNP - 40 mm Hg, which was sufficient to decrease arterial pressure and activate high pressure baroreceptor reflexes, occurred during low levels of venous pooling at LBNP - 10 mm Hg without changes in arterial pressure. This suggests that much of the forearm vasoconstriction with the high levels of venous pooling, which were sufficient to decrease arterial pressure, may be accounted for by reflexes originating in areas other than high pressure baroreceptors. The results of these studies suggest that low pressure baroreceptors exert an important influence on forearm vascular tone during decreases in venous return and central venous pressure in man.

Splanchnic and peripheral glucose and amino acid metabolism in diabetes mellitus

Abstract: Splanchnic and leg exchange of glucose, lactate, pyruvate, and individual plasma amino acids was studied in diabetics 24 hr after withdrawal of insulin and in healthy controls. Measurements were made in the basal postabsorptive state and during the administration of glucose at a rate of 2 mg/kg per min for 45 min. In the basal state, net splanchnic glucose production did not differ significantly between diabetics and controls. However, splanchnic uptake of alanine and other glycogenic amino acids was 1½-2 times greater in the diabetics, while lactate and pyruvate uptake was increased by 65-115%. Splanchnic uptake of these glucose precursors could account for 32% of hepatic glucose output in the diabetics, as compared to 20% in the controls. This increase in precursor uptake was a consequence of a two- to threefold increment in fractional extraction of these substrates inasmuch as arterial levels of alanine, glycine, and threonine were reduced in the diabetics, while the levels of the remaining substrates were similar in the two groups. Peripheral output of alanine and other glycogenic amino acids as reflected in arterio-femoral venous differences was similar in both groups. An elevation in arterial valine, leucine, and isoleucine was observed in the diabetics, but could not be accounted for on the basis of alterations in splanchnic or peripheral exchange of these amino acids. Administration of glucose (2 mg/kg per min) for 45 min resulted in an 80% reduction in splanchnic glucose output in controls, but failed to inhibit hepatic glucose release in the diabetics despite a twofold greater increment in arterial glucose levels. In both groups no consistent changes in arterial glucagon were observed during the infusion. It is concluded that in nonketotic diabetics (a) total splanchnic output of glucose is comparable to controls, but the relative contribution of gluconeogenesis may be increased by more than 50%; (b) accelerated splanchnic uptake of glucose precursors is a consequence of increased hepatic extraction of available substrates rather than a result of augmented substrate supply; and (c) the failure of glucose infusion to inhibit hepatic glucose output suggests that the exquisite sensitivity of the liver to the infusion of glucose in normal man is a consequence of glucose-induced insulin secretion.

Hormonal control of the renal conversion of 25-hydroxycholecalciferol to 1,25-dihydroxycholecalciferol.

Abstract: Isolated renal tubules from vitamin D-deficient chicks catalyse the in vitro conversion of 25-hydroxycholecalciferol to 1,25-dihydroxycholecalciferol. This conversion is stimulated by 5 x 10(-10) M bovine parathyroid hormone, or by 10(-6) M cyclic AMP. It is inhibited by 10(-9) M porcine calcitonin. It is concluded that these hormonal controls of the synthesis of the renal hormone 1,25-dihydroxycholecalciferol are of particular physiological significance in coordinating the activities of the various organs involved in extracellular calcium homeostasis.

Effect of Cholera Enterotoxin on Ion Transport across Isolated Ileal Mucosa

Abstract: The effects of cholera enterotoxin on intestinal ion transport were examined in vitro. Addition of dialyzed filtrate of Vibrio cholerae (crude toxin) to the luminal side of isolated rabbit ileal mucosa caused a delayed and gradually progressive increase in transmural electric potential difference (PD) and shortcircuit current (SCC). A similar pattern was observed upon addition of a highly purified preparation of cholera toxin, although the changes in PD and SCC were smaller. Na and Cl fluxes across the short-circuited mucosa were determined with radioisotopes 3-4 hr after addition of crude toxin or at a comparable time in control tissues. The toxin caused a net secretory flux of Cl and reduced to zero the net absorptive flux of Na. Similar flux changes were observed when either crude or purified toxin was added in vivo and tissues were mounted in vitro 3-4 hr later. Additon of D-glucose to the luminal side of toxin-treated mucosa produced a large net absorptive flux of Na without altering the net Cl and residual ion fluxes. Adenosine 3',5'-cyclic phosphate (cyclic AMP) and theophylline had previously been shown to cause a rapid increase in SCC and ion flux changes similar to those induced by cholera toxin. Pretreatment of ileal mucosa with either crude or purified cholera toxin greatly reduced the SCC response to theophylline and dibutyryl cyclic AMP, which, together with the flux data, suggest that both cyclic AMP and cholera toxin stimulate active secretion by a common pathway. Inhibition of the SCC response to theophylline was observed after luminal but not after serosal addition of toxin. In vitro effects of cholera toxin correlated closely with in vivo effects: heating toxin destroyed both; two V. cholerae filtrates which were inactive in vivo proved also to be inactive in vitro; PD and volume flow measurements in isolated, in vivo ileal loops of rabbit revealed that the PD pattern after addition of toxin is similar to that seen in vitro and also correlates closely with changes in fluid movement. The results suggest that stimulation by cholera toxin of a cyclic AMP-dependent active secretory process of the intestinal epithelial cells is a major cause of fluid loss in cholera.

The pathogenesis of Shigella Diarrhea: I. Enterotoxin production by Shigella dysenteriae 1

Abstract: A strain of Shigella dysenteriae 1, freshly isolated from a patient with dysentery in Guatemala in August 1969, was found to elaborate an enterotoxin into the liquid of broth cultures. Partial purification of the enterotoxin by ultrafiltration on graded polymeric membranes and Sephadex gel filtration (Pharmacia Fine Chemicals, Inc., Piscataway, N. J.) suggested an approximate molecular weight of 55,000-60,000. The partially purified toxin was heat-labile, pronase sensitive, and activated by alkaline pH, and it elicited fluid production in ligated rabbit ileal segments; it failed, however, to cause increased vascular permeability in skin. When the activities of equal weights of identically prepared Vibrio cholerae and S. dysenteriae enterotoxins were compared in the rabbit ileum the latter caused a significantly smaller volume response with increased concentrations of potassium, chloride, and protein. The previously described neurotoxic (mouse lethal) factor was also present and eluted from Sephadex G-150 with the enterotoxin. If these biological activities prove to be possessed by a single molecular species, it is suggested that it be renamed Shigella enterotoxin in recognition of the physiologically more relevant biological action.

Effect of age, sex, and sites on the cellularity of the adipose tissue in mice and rats rendered obese by a high-fat diet.

Abstract: Cell size and number of parametrial fat pads were determined in Swiss mice made obese by means of a high-fat diet (40% lard w/w) given ad lib. This diet and a control were introduced to two groups of mothers during gestation and lactation, and sucklings were given the same diets as their mothers at weaning and throughout life.2-wk old mice suckled by mothers fed a high-fat diet have fatter parametrial pads. This difference is due solely to an increase in fat cell size. After weaning, until the 18th wk, the two groups differed with a striking fat cell enlargement seen in the obese group. Later on, whereas cell numbers did not change in the control group, a constant and uninterrupted increase in number is shown in those of obese mice until the 52nd wk. Hyperplasia was observed only in adults. When the high-fat diet was introduced to adult rats it also triggered an increase in fat cell number. Three sites of fat pads were compared in both sexes at 32 wk of age. All sites increased in weight in the high-fat fed group. This was due to: hyperplasia in perirenal site, hypertrophy in epididymal and subcutaneous sites, and hyperplasia plus hypertrophy in the parametrial one. So, in each sex, adipose sites in the obese mice reacted to the diet in a site-specific way. It was concluded that the level of fat in a diet is involved in both formation and maturation of new fat cells and in the regulation of fat cell lipid content. The two processes may be separated or may act together according to the adipose tissue site.

Coronary Artery Reperfusion: II. REDUCTION OF MYOCARDIAL INFARCT SIZE AT 1 WEEK AFTER THE CORONARY OCCLUSION

Abstract: The question of whether or not the size of an area of myocardial infarction, measured at 1 wk after coronary occlusion, can be influenced by coronary artery reperfusion was examined in dogs. In seven control experiments the anterior descending coronary artery was ligated, while in seven other studies the occlusion was released after 3 hr. In all animals calibrated photographs were used to assess the zone of hypoperfusion and the acutely injured area of epicardial ST segment elevation, as well as the extent of damage at postmortem 1 wk later. In control dogs, the gross infarct size at postmortem averaged 63.8+/-7.3% of that predicted from the acutely injured zone. However, in reperfused hearts the average gross infarct size at 1 wk was only 10.2+/-4.4% of that predicted. Transmural specimens were obtained at autopsy for histology and measurement of myocardial creatine phosphokinase (CPK) activity from sites initially used for epicardial electrocardiography. In control animals, there was a direct relationship between the degree of ST segment elevation and the degree of cell necrosis in transmural histologic sections. ST segment elevation also predicted myocardial CPK (international units per milligram protein): log CPK = - 0.0613 ST + 1.17 (r = 0.66, n = 56 sites). In the reperfused animals, log CPK = - 0.166 ST + 1.36 (r = 0.69, n = 46 sites) showing almost complete preservation of CPK activity at 1 wk, sparing being most prominent in the epicardial zone. Similarly, there was a good correlation between myocardial CPK activity and the histological assessment of cell destruction, the degree of cell damage = - 0.152 CPK + 3.86 (r = 0.86; n = 102 sites). Thus, control dogs showed severe myocardial CPK depletion and histologic evidence of extensive cell destruction, whereas animals subjected to coronary artery reperfusion had little CPK depletion and much less evidence of myocardial cell necrosis 1 wk later.

Identification of the colony-stimulating cell in human peripheral blood

Abstract: Bone marrow colony formation in soft gel culture may be stimulated by substances elaborated by human peripheral blood leukocytes. In order to determine the cell type responsible for colony stimulation, peripheral leukocytes were separated by Ficoll-Hypaque gradients and differential glass adhesion. Morphologic, histochemical, and functional criteria were applied to determine the purity of the monocyte, lymphocyte, and neutrophil fractions. Using these cells as feeder layers and as a source of conditioned medium, evidence was obtained that the monocyte is the colony-stimulating cell of human peripheral blood. Activity greater than that of mixed white cells was obtained with monocyte underlayers, and only monocyte- and macrophage-conditioned media were shown to have significant colony-stimulating activity.

Food iron absorption measured by an extrinsic tag.

Abstract: The paper describes the use of an extrinsic tag of inorganic radioiron to determine the total absorption of nonheme iron from a complete meal. The method was developed by measuring the iron absorbed from vegetable foods containing biosynthetically incorporated (55)Fe (intrinsic tag) and from (59)Fe added as a small dose of inorganic iron to the same meal (extrinsic tag). In studies with maize, black bean, and wheat, a consistent extrinsic: intrinsic radioiron absorption ratio averaging 1.10 was observed. Similar results were obtained with either ferrous or ferric iron as the extrinsic tag, and with doses of the latter ranging from 0.001 to 0.5 mg iron added to a test meal containing 2-4 mg of food iron. Adding the radioiron at different stages in preparation of the test meal also had little effect. Separate administration of the extrinsic tag was less satisfactory when small portions of a single food were employed, but with a complete meal, the separate dose was preferable. The extrinsic tag provided a valid measure of absorption despite marked differences in the iron status of the subject, and with wide changes in absorption imposed by adding desferrioxamine or ascorbic acid to the test meal. These findings indicate that there is a common pool of nonheme iron, the absorption of which is influenced by various blocking or enhancing substances present in the meal.

Human Serum Activities against Hemophilus influenzae, Type b

Abstract: Humoral immunity to Hemophilus influenzae, type b was studied in normal human adults by means of assays for serum bactericidal and opsonizing activities against the organism and for passive hemagglutinating activity using erythrocytes sensitized with polyribophosphate, the type-specific capsular antigen. Hemagglutinating activity was detectable in about 60% of the 114 sera tested. Serum bactericidal and opsonizing activities were found in all sera tested; the levels in some sera, however, were quite low. The antibacterial activities were due not only to antibodies directed against the polyribophosphate capsule but also to antibodies that appear to be directed against somatic antigens. Type b strains differed in their susceptibility to the antisomatic antibodies of particular sera but were uniformly sensitive to anticapsular antibody.

Phenobarbital-induced alterations in vitamin D metabolism.

Abstract: The metabolic fate of intravenously injected vitamin D3-1,2-3H (D3-3H) was studied in two normal individuals on chronic phenobarbital therapy. Silicic acid column chromatography of lipid-soluble plasma extracts obtained serially for 96 hr after D3-3H injection demonstrated a decreased plasma D3-3H half-life and increased conversion to more polar metabolites. The polar metabolites formed included several with chromatographic mobility similar to known biologically inactive vitamin D metabolites and one with chromatographic mobility identical to 25-hydroxycholecalciferol. Disappearance of this latter material was also accelerated. A child with rickets and a normal volunteer studied before and after a 2 wk course of phenobarbital therapy demonstrated similar alterations in D3-3H metabolism. When liver microsomes from 3-wk-old Sprague-Dawley rats treated with phenobarbital were incubated with D3-3H, polar metabolites were produced with chromatographic mobility similar to the plasma D3-3H metabolites from phenobarbital-treated humans. Similar incubations employing 25-hydroxy-cholecalciferol-26-27-3H as the substrate also demonstrated an increased conversion to polar metabolites. The data suggest that the reported increased incidence of osteomalacia observed in patients on chronic anticonvulsant therapy may be the result of an accelerated conversion of vitamin D and its active metabolite, 25-hydroxycholecalciferol, to polar metabolites by druginduced liver microsomal enzymes.

Characterization of the kinetics of the passive and active transport mechanisms for bile acid absorption in the small intestine and colon of the rat

Abstract: Bile acid uptake occurs via passive diffusion in all regions of the intestine and via active absorption in the ileum. Determination of the passive permeability coefficient for ionized monomers (*P-) demonstrated that permeability decreased by a factor of 3.4, 6.8, and 8.1 for the addition of a hydroxyl, glycine, or taurine group, respectively, to the steroid nucleus. Removal of the negative charge increased permeation by a factor of 4.4; however, permeability coefficients for the protonated monomers showed the same relative decrease with addition of a hydroxyl group. The calculated incremental free energies of solution (δΔFW→1) associated with these additions equaled + 757 (hydroxyl), + 1178 (glycine), and + 1291 (taurine) cal/mole. Passive permeability coefficients for the transverse colon showed the same relative relationships among the various bile acids. After making appropriate corrections for passive permeability across the ileum, apparent values for the maximal transport velocity (*Vmax) and Michaelis constant (*Km) of the active transport system were measured. *Vmax depended upon the number of hydroxyl groups on the steroid nucleus; values for the trihydroxy bile acids were high (1543-1906 pmoles/min per cm) while those for the dihydroxy (114-512 pmoles/min per cm) and monohydroxy (45-57 pmoles/min per cm) acids were lower. In contrast, *Km values were related to whether the bile acid was conjugated; unconjugated bile acids had values ranging from 0.37 to 0.49 mM, while values for the conjugated bile acids were approximately half as high (0.12-0.23 mM).

Use of pulmonary hydrogen (H 2 ) measurements to quantitate carbohydrate absorption: Study of partially gastrectomized patients

Abstract: A technique was developed to quantitate the absorption of ingested carbohydrate by means of continuous measurements of pulmonary H(2) excretion. This technique is based on the observation that H(2) is produced in the colon when carbohydrate is fermented by colonic bacteria, and this H(2) is then excreted by the lungs. The quantitative relationship of pulmonary H(2) excretion to unabsorbed carbohydrate was studied in nine subjects. After ingestion of 6.5, 13, and 26 g of lactulose (a nonabsorbable disaccharide), H(2) excretion increased linearly, averaging (+/-1 SEM) 13+/-3.5, 23+/-7.2, and 49+/-7 ml per 2 hr. Because of consistent individual differences in H(2) excretion per gram of lactulose, the variability of this linear response was less in a given subject, with the H(2) excretion after 6.5 g and 26 g lactulose dosages averaging 55+/-4.2% and 214+/-16% of that observed after the 13 g dose. It was further demonstrated with fecal homogenates, as well as in rats after direct intracecal instillation of carbohydrate, that there was no significant difference in the rate of H(2) formation from lactulose as compared with the normally ingested sugars. Thus, a subject's H(2) excretion after a 13 g dose of lactulose can be used as a standard to convert H(2) excretion after ingestion of other carbohydrates into grams of carbohydrate not absorbed. Application of this technique to seven partially gastrectomized patients indicated all subjects malabsorbed a portion of a 100 g dose of glucose whereas six of seven completely absorbed a 25 g dose. Malabsorption of physiologic quantities of various carbohydrates was clearly demonstrated in one subject. This technique appears to provide quantitative information on carbohydrate malabsorption not readily obtained by presently available techniques.

The mechanism of intestinal uptake and transcellular transport of IgG in the neonatal rat.

Abstract: The transport of immunoglobulins across the intestinal mucosa of neonatal rats provides an excellent model for the study of transcellular protein transport. The mechanism of intestinal uptake and transcellular transport of plasma proteins has been studied in 12-14-day old rats using intraduodenally administered radioiodinated proteins. Appreciable quantities of rat IgG, mouse IgG, rabbit IgG, and all four subclasses of human IgG were taken up by the intestinal wall (19-54% of administered dose at 4 hr) and transported to the animal (10-35% of administered dose at 4 hr). In contrast there was little or no uptake of human IgM, IgA, and IgE and little or no transport of human IgM, IgA, IgD, IgE, albumin, transferrin, and ceruloplasmin. Both the uptake and transport of labeled IgG were significantly inhibited by unlabeled IgG. Further insight into the transport process was obtained from the observation that an appreciable proportion of the label of IgG in intestinal wall homogenates, but not in plasma or intestinal washings, migrated in a sucrose ultracentrifugation gradient much more rapidly than did the administered 7S molecules. This pattern was not observed with other proteins studied. This apparent binding of labeled IgG was also markedly inhibited by unlabeled IgG. In subcellular fractionation studies of intestinal homogenates the complexed labeled IgG was shown to be associated predominantly with cell membrane rather than cell sap fractions. In addition IgG could be shown to bind to purified enterocyte microvillous membranes in vitro. IT IS CONCLUDED THAT IN THE NEONATAL RAT: (a) the major processes involved in both intestinal uptake and transport of IgG are specific and saturable; (b) intestinal transport is associated with complexing of IgG molecules with membranes, most probably with enterocyte microvillous membranes; and (c) the part of the IgG structure involved in this process is probably similar to that involved in the concentration-catabolism effect but is not identical to that mediating other non-antigen combining functions of IgG. Our data are consistent with the existence of specific receptors for IgG on enterocyte microvillous membranes of the neonatal rat. Such receptors would be necessary for the specific uptake and transport of these molecules.

Metabolic relationships among the plasma lipoproteins: Reciprocal changes in the concentrations of very low and low density lipoproteins in man

Abstract: The changes in other plasma lipoproteins which accompany alterations in very low density lipoproteins (VLDL) were studied in 31 normal and hyperlipidemic men and women who underwent weight reduction, carbohydrate induction, or clofibrate treatment. Plasma lipids and individual lipoprotein cholesterol concentrations were measured serially during control and treatment periods. Low density lipoprotein (LDL) protein was determined by radial immunodiffusion. Oppositely directed changes in VLDL and LDL were found with each of the three metabolic perturbations. Changes in high density lipoprotein (HDL) cholesterol generally paralleled those in LDL but were less consistent. Two patients with type III hyperlipoproteinemia failed to demonstrate reciprocal increases in LDL despite more than 40% reduction in plasma glycerides or VLDL with weight reduction or clofibrate therapy. After clofibrate therapy, LDL increased in proportion to the absolute decrease in VLDL cholesterol during treatment. LDL protein changed relatively less than did LDL cholesterol. The mechanism for the interdependency of plasma VLDL and LDL concentrations over the long term is not known and may be the result of altered rates of interconversion of these lipoproteins, or to feedback inhibition by VLDL of LDL production and release.

The renal handling of low molecular weight proteins: II. Disorders of serum protein catabolism in patients with tubular proteinuria, the nephrotic syndrome, or uremia

Abstract: The present study was directed toward determining the role of the kidney in the metabolism of various classes of serum proteins and to define the urinary protein excretion patterns and the pathogenesis of disorders of protein metabolism in patients with proteinuria. To this end, the metabolic fates of a small protein, lambda-L chain (mol wt 44,000), and a protein of intermediate size, IgG (mol wt 160,000), were studied in controls and patients with renal disease. Controls metabolized 0.28%/hr of circulating IgG and 22.3%/hr of circulating lambda-L chain. All the IgG and 99% of the lambda-L chain was catabolized with the remaining lambda-L chain lost intact into the urine. The kidney was shown to be the major site of catabolism for small serum proteins. Three distinct disorders of protein metabolism were noted in patients with renal tubular disease and tubular proteinuria, glomerular disease (the nephrotic syndrome), and disease involving the entire nephrons (uremia), respectively. Patients with renal tubular disease had a 50-fold increase in the daily urinary excretion of 15-40,000 molecular weight proteins such as lysozyme and lambda-L chains. Serum IgG and lambda-L chain survivals were normal; however, the fraction of the over-all lambda-L chain metabolism accounted for by proteinuria was increased 40-fold whereas endogenous catabolism was correspondingly decreased. Thus, tubular proteinuria results from a failure of proximal tubular uptake and catabolism of small proteins that are normally filtered through the glomerulus. Patients with the nephrotic syndrome had a slight increase in lambda-L chain survival whereas IgG survival was decreased and the fraction of IgG lost in the urine was markedly increased. Here, abnormal glomerular permeability to proteins of intermediate size is the basic abnormality. Patients with uremia had a normal IgG survival but a four to 10-fold prolongation of lambda-L chain survival due to loss of entire nephrons, the major site of metabolism of these proteins. This results in an increase (up to 10-fold) in the serum concentration of lambda-L chain, lysozyme, and other small biologically active proteins, a phenomenon that may be of importance in causing some of the manifestations of the uremic syndrome.

Site of Airway Obstruction in Asthma as Determined by Measuring Maximal Expiratory Flow Breathing Air and a Helium-Oxygen Mixture

Abstract: Because maximum expiratory flow-volume rates in normal subjects are dependent on gas density, the resistance between alveoli and the point at which dynamic compression begins (Rus) is mostly due to convective acceleration and turbulence. We measured maximum expiratory flow-volume (MEFV) curves in asthmatics and chronic bronchitics breathing air and He-O2. In the latter and in some asthmatics, MEFV curves did not change, indicating that Rus is mostly due to laminar flow. Therefore, the point at which dynamic compression begins must be further upstream than in normal subjects and the site of obstruction must be in small airways. In other asthmatics, flow increased normally indicating obstruction in larger airways. The response to He-O2 did not correlate with initial values of pulmonary resistance, the initial MEFV curves or the response to bronchodilators. We conclude that the site of airway obstruction varies among asthmatics and that the site of obstruction is not detectable by measurement of the usual parameters of lung mechanics.

Estrone sulfate: production rate and metabolism in man.

Abstract: Since estrone sulfate (E(1)S) is present at high concentration in plasma, we have examined the parameters of the plasma estrone, estradiol, E(1)S system. The metabolic clearance rate of E(1)S was 157 liter/day (range 70-292) in men and women. Estimated plasma production rates of E(1)S were (mugrams per day): men, 77; women, early follicular phase, 95; women, early luteal phase, 182. The conversion of plasma estrone and estradiol to E(1)S was measured and from these data and the metabolic clearance rates of the estrogens, the transfer factors were rho(E) (1) (E) (1) (S) = 0.54 and rho(E) (2) (E) (1) (S) = 0.65. Using average production rates, all plasma E(1)S could be shown to be derived from plasma estrone and estradiol. The conversion of plasma E(1)S to plasma estrone and estradiol was studied. The calculated transfer factors were: rho(E) (1) (SE) (1) = 0.21, rho(E) (1) (SE) (2) = 0.014. Essentially, similar data were obtained when E(1)S was given by mouth to two subjects. WE CONCLUDE: (a) E(1)S is a major circulating plasma estrogen and has a long plasma half-life; (b) the large contributions of estrone and estradiol to plasma E(1)S are more than sufficient to account for all the circulating plasma E(1)S.

Glucagon-stimulating activity of 20 amino acids in dogs

Abstract: The effect of 20 L-amino acids upon pancreatic glucagon secretion has been studied in conscious dogs. Each amino acid was administered intravenously over a 15 min period in a dose of 1 mmole/kg of body weight to a group of four or five dogs. Pancreatic glucagon and insulin were measured by radioimmunoassay. 17 of the 20 amino acids caused a substantial increase in plasma glucagon. Asparagine had the most glucagon-stimulating activity (GSA), followed by glycine, phenylalanine, serine, aspartate, cysteine, tryptophan, alanine, glutamate, threonine, glutamine, arginine, ornithine, proline, methionine, lysine, and histidine. Only valine, leucine, and isoleucine failed to stimulate glucagon secretion, and isoleucine may have reduced it. No relationship between glucagon-stimulating activity and insulin-stimulating activity was observed. The amino acids which enter the gluconeogenic pathway as pyruvate and, which are believed to provide most of the amino acid-derived glucose, had a significantly greater GSA than the amino acids which enter as succinyl CoA or as alpha-ketoglutarate. However, pyruvate itself did not stimulate glucagon secretion. The R-chain structure of the amino acid did not appear to be related to its GSA, except that the aliphatic branched chain amino acids, valine, leucine, and isoleucine, were devoid of GSA.

Circadian rhythm in serum parathyroid hormone concentration in human subjects: correlation with serum calcium, phosphate, albumin, and growth hormone levels

Abstract: A circadian variation in serum calcium, albumin and PTH concentration in normal subjects has been demonstrated. The levels of the three blood constituents were remarkably constant during the day, but striking night and early morning changes occurred. Serum calcium levels were highest at 8:00 p.m. and reached a nadir between 2:00 and 4:00 a.m. Serum albumin levels were parallel to those of serum calcium. PTH levels began to rise after 8:00 p.m., reached the highest levels between 2:00 and 4:00 a.m., and fell to baseline values by 8:00 a.m. The nocturnal fall in serum calcium levels appears to be secondary to dilution of serum proteins by increasing blood volume. The nocturnal rise in PTH levels appears to be independent of serum calcium levels within the normal range but it can be abolished by induced hypercalcemia. Serum phosphate levels were lowest between 8:00 a.m. and 10:00 a.m. and highest between 2:00 a.m. and 4:00 a.m. The data presented suggest that circadian changes in serum phosphate levels are not mediated in toto by parathyroid hormone but they are exaggerated when the secretion of this hormone is inhibited. They are independent of growth hormone levels and activity but they are greatly modified during a prolonged fast.