Issue published August 1, 1970

G roups of male nonobese juvenile diabetics with recent onset, short term (1-8 yr), and long-term (12-30 yr) diabetes as well as comparable nondiabetic controls were studied during exercise experiments. The chosen exercise load, 450 kg/min for 20 min never induced changes in serum growth hormone in our nondiabetic control subjects.The principal results of the study were as follows: (a) an immediate high rise in serum growth hormone followed the commencement of exercise in all diabetics. The increase and pattern of serum growth hormone was not related to the duration of the diabetes. (b) The abnormal growth hormone response to exercise in diabetics was observed when the patients were in poor control as well as when they were in clinically excellent control (fasting blood glucose level between 100 and 140 mg/100 ml). (c) However, the abnormal serum growth hormone response was significantly diminished when exceedingly strict control was achieved (fasting blood glucose level between 60 and 100 mg/100 ml). In two of these experiments an entirely normal growth hormone pattern was obtained. (d) The change in serum growth hormone pattern during regulation was totally unrelated to the changes in serum free fatty acid patterns. A normal free fatty acid level and exercise pattern was obtained much earlier during the improved control. (e) Fasting serum growth hormone levels were also significantly raised in the juvenile diabetics irrespective of the diabetes duration. (f) Fasting serum growth hormone levels were also significantly decreased during regulation. Furthermore, a significant correlation between blood glucose and fasting serum growth hormone concentration was established. (g) In the juvenile diabetics a significant increase in serum insulin was observed at the point of time when exercise was concluded.

P reparations of lipid droplets and droplet-free tissue residue (cytoplasm + membranes + nuclei) were obtained by homogenization and centrifugal separation from intimal fatty streak lesions of aortic atherosclerosis of 21 adults who had died suddenly. Neutral lipids and phospholipids were analyzed by quantitative thin-layer chromatography and cholesteryl ester fatty acids by gas-liquid chromatography. Optical properties of droplets were evaluated by differential counting and sizing procedures with the polarizing microscope. The droplets occurred in mixtures of two forms distinguished by their optical properties, anisotropic (i.e. liquid crystals) and isotropic (true liquids). Both forms had average diameters of about 1.8 μ, with a range of 0.5-5μ. The proportions of the two forms varied with temperature as individual droplets changed their form; anisotropic forms averaged 83.7% at °C and 37.8% at 37°C, with isotropic forms being 16.3 and 62.2% respectively. The proportions of anisotropic forms at 22°C decreased with age. These forms were not separated for chemical analysis. The droplets contained about half the lipid in the lesions. The composition of the lipids of the droplet mixture was remarkably uniform and strikingly different from that of the droplet-free residue, respectively: cholesteryl esters 94.9% vs. 38.7%, free cholesterol 1.7% vs. 18.6%, total phospholipids 1.0% vs. 38.6%, and triglycerides 2.4% vs. 4.0%. The proportions of individual phospholipids, with the exception of lysolecithin, were also different between the preparations. In the droplets only the proportions of lecithin correlated positively with the proportion of anisotropic forms (at 22°C). Droplet cholesteryl esters were particularly rich in oleic acid and when compared to residue esters had more palmitoleic (+0.7%), oleic (+12.3%), and eicosatrienoic (+2.4%) and less palmitic (-2.2%), linoleic (-12.4%), and arachidonic (-1.6%) acids. The proportions of most individual fatty acids of droplets and residue correlated positively. The lipids of the residue closely resemble those reported for the normal intima.The observations that these droplets are prominent in the morphology of the fatty streak lesions, and that their high content of oleate-rich cholesteryl esters is similar to that reported for analysis of the whole lesions, suggest that the droplets may be involved in the pathogenesis of the fatty streak lesions of artherosclerosis in man.

T here was a rapid net uptake of free fatty acid (FFA) by human platelets when long-chain FFA, bound to human serum albumin, were incubated with platelet suspensions. Results from experiments in which both palmitate and albumin were labeled indicated that the fatty acid dissociated from the protein during uptake. Much of the FFA taken up by the platelet in short-term incubations remained in unesterified form, i.e., it was recovered as platelet FFA. As the incubation continued, increasing amounts of FFA were oxidized to CO₂ and incorporated into platelet lipid esters, particularly lecithin. Essentially all of the fatty acid that was incorporated into the platelet FFA fraction was released rapidly from the cells when they were exposed to a medium containing FFA-free albumin. The magnitude of uptake into the platelet FFA fraction was similiar at 0° and 37°C. Likewise, the rate and magnitude of FFA release from the platelet were similar at 0° and 37°C. Therefore, it is likely that both FFA uptake and FFA release occur by energy-independent mechanisms. The major effect of increasing the FFA concentration of the incubation medium was increased fatty acid uptake into the platelet FFA fraction. Similar results occurred when platelets were incubated in human plasma containing increasing amounts of added palmitate. At a given extracellular FFA concentration, considerably more of the saturated fatty acids, palmitate and stearate, were taken up as platelet FFA than either oleate or linoleate.

T wo O-hydroxylysyl glycosides, Hyl-Gal-Glc and Hyl-Gal, have been isolated from normal human urine and shown to be identical to two glycosides isolated from alkaline hydrolysates of collagen. A relatively sample and reproducible analytical procedure has been devised to measure the levels of these glycosides in human urine. By the use of this procedure it was shown that a normal diet has only a small effect on 24-hr urinary excretion levels of these glycosides indicating an endogenous origin. Urinary glycoside levels appear to be highest in children, roughly paralleling collagen turnover as indicated by urinary hydroxyproline levels. Collagen turnover equivalents calculated from urinary hydroxylysyl glycoside levels were found to be significantly larger than collagen turnover equivalents calculated from urinary hydroxyproline levels. This suggests that urinary glycosides are more quantitative indicators of collagen metabolism than urinary hydroxyproline.The ratio of Hyl-Gal-Glc to Hyl-Gal was measured in urines of diseased as well as normal individuals and a bimodal distribution was found. Alkaline hydrolysates of different human connective tissue collagens showed that only bone collagen, of the collagens examined, had a low ratio of Hyl-Gal-Glc to Hyl-Gal compared to human urine. Other collagens examined had higher ratios than found in human urine. On the basis of these results it is postulated that the bimodal distribution of glycoside ratios represents two populations of collagen turnover, the lower ratio population having a high bone collagen turnover, the lower ratio population having a high bone collagen turnover relative to the second population. Examination of the types of subjects making up the two populations supports this hypothesis. These data suggest that urinary O-hydroxylysyl glycoside excretion, in addition to providing a more quantitative estimate of collagen turnover than urinary hydroxyproline, may prove to be of value as a specific means of studying the metabolism of bone collagen.

S erial determinations of plasma renin activity, sodium balance, urinary potassium excretion rate, and plasma sodium and potassium concentration were done in five dogs during dietary-induced potassium depletion and repletion. Duration of depletion for the different animals ranged from 5 to 7 wk.Plasma renin activity increased in all animals during depletion, with rises being demonstrated as early as the 1st depletion day in two of the dogs. Maximum values in the five dogs were recorded from the 2nd to the 28th day of depletion. Early in depletion, changes in renin activity did not correlate with changes in sodium balance. Late in the course of depletion, plasma renin activity decreased concurrently with a progressive retention of sodium. However, in every case increased renin activity persisted throughout depletion despite development of sodium retention sufficient to inhibit renin release in normal dogs.Potassium repletion resulted in a prompt decrease in renin activity to predepletion values.This study indicates that potassium deficiency has a stimulatory effect on renin release that is independent of any effect on sodium balance.

T he concentrations of plasma glucose, free fatty acids, insulin, growth hormone, and placental prolactin in subhuman primate fetal and maternal plasma were examined following intravascular administration of insulin and glucagon to the fetus and mother. The neonatal plasma responses to these same stimuli were also examined.Fetal plasma glucose concentrations were minimally altered by direct fetal insulin injections, whereas neonatal glucose levels declined with similar injections. In both instances, however, plasma free fatty acid levels declined following insulin. When the amount of insulin given the fetus was increased, fetal plasma glucose concentrations did decline. Combined intravascular insulin injections and infusions in the mother were associated with a disappearance of the initial maternal to fetal plasma glucose concentration gradient and a nearly parallel fall in both maternal and fetal plasma glucose levels. It was concluded that insulin was biologically active in the fetus. Obtunded fetal plasma glucose responses to direct fetal insulin administration may be a function of placental transfer of glucose from the maternal pool.Maternal plasma placental prolactin and fetal plasma growth hormone levels were unchanged in the presence of sustained maternal and fetal hypoglycemia. However, neonatal plasma growht hormone levels did increase in response to hypoglycemia. The observed bidirectional placental barrier to transfer of radioisotopically labeled growth hormone indicated that fetal plasma growth hormone was solely of fetal origin. These data suggested further that a change in the growth hormone-releasing mechanism may occur from fetal to neonatal life.Direct maternal intravascular glucagon administration led to augmentation in both maternal and fetal plasma insulin and glucose levels. Direct fetal glucagon injections enhanced both maternal and fetal plasma insulin levels. These simultaneous changes in both plasma pools were consistent with the demonstration of a bidirectional placental transfer of radioisotopically labeled glucagon. The role of endogenously produced glucagon in these studies remains to be clarified.

T he phagocytic, bactericidal, and metabolic capabilities of circulating blood leukocytes from three adults (two males, one female) with hypogammaglobulinemia and recurrent pneumonia, chronic sinusitis, and intestinal giardiasis were studied. These functions were found to be normal when leukocytes from the patients were incubated in media containing normal human serum. Phagocytosis of Staphylococcus albus and polystyrene balls by both patient and normal leukocytes was diminished when the cells were incubated in hypogammaglobulinemic plasma. A similar defect in opsonization by patient plasma was also noted for pneumococci, Escherichia coli and variably with Staphylococcus aureus. Both patient and normal sera had equivalent levels of heat-labile S. albus opsonins; normal serum, however, contained heat-stable S. albus-specific absorbable opsonins in significantly greater quantities to account for its superior opsonic capacity. The addition of commercial gamma globulin or purified IgG to hypogammaglobulinemic sera restored full S. albus opsonic activity. The relevancy of these observations to the impaired host defenses in these patients will be discussed.

L eft ventricular performance in 19 patients with acute myocardial infarction has been evaluated by measuring left ventricular response in terms of cardiac output, stroke volume, work, and power to progressive elevation of filling pressure accomplished by progressive expansion of blood volume with rapid infusion of low molecular weight dextran. Such infusion can elevate the cardiac output, stroke volume, work, and power and thus delineate the function of the left ventricle by Frank-Starling function curves. Left ventricular filling pressure in the range of 20-24 mm Hg was associated with the peak of the curves and when the filling pressure exceeded this range, the curves became flattened or decreased. An increase in cardiac output could be maintained for 4 or more hr. Patients with a flattened function curve had a high mortality in the ensuing 8 wk. The function curve showed improvement in myocardial function during the early convalescence. When left ventricular filling pressure is monitored directly or as pulmonary artery end-diastolic pressure, low molecular weight dextran provides a method for assessment of left ventricular function.

C hromium is a trace metal of importance in human physiology and, in addition, as 51-chromate, has been extensively used as a label in the study of blood cell pool sizes and intravascular kinetics. The transport characteristics of 51-chromate were investigated in normal human leukocytes. Chromate uptake is unidirectional over a 1 hr incubation with extracellular chromate concentrations up to 200 μmoles/liter. Under these conditions, intracellular 51-chromium is in a form in which it is nonexchangeable. Influx is temperature sensitive with a Q₁₀ of approximately 2 and may be energy dependent since a variety of metabolic poisons strongly inhibit uptake. The unidirectional influx of chromate follows Michaelis-Menten kinetics; the maximum velocity is 52 mμmoles/g dry weight of cells per min and the chromate concentration at which influx velocity is half maximal is 87 μmoles/liter. This transport mechanism is highly specific for chromate; other divalent tetrahedral anions only slightly inhibit influx at concentrations up to 10 times that of chromate. Metavanadate, however, competitively inhibits chromate influx at equimolar concentrations. Exposure of cells to unlabeled chromate leads to inhibition of subsequent influx of 51-chromate. It is suggested that this is due to a primary inhibitory effect of chromate on cellular energy metabolism.

T he ability of products of digestion to stimulate pancreozymin secretion in man was investigated using a bioassay procedure, based on duodenal perfusion, which quantified the total outputs of pancreatic enzymes evoked by intraduodenal stimuli under steady-state conditions. Patterns of response resulting from physiologic intraduodenal concentrations of test material were basal output (with isotonic saline), washout of enzymes (with dextrose, micellar fatty acid, and amino acids), and sustained output of enzymes (with amino acids and micellar fatty acid). The sustained secretion of pancreatic enzymes found during the 2nd hr of perfusion and subsequently was characteristic of pancreozymin-induced secretion. The enzyme output in response to a mixture of essential and nonessential amino acids was significantly higher than that evoked by micellar fatty acid and was comparable with that resulting from the maximally tolerated dose of pancreozymin given by vein.Perfusion with essential amino acids caused enzyme outputs comparable to those induced by perfusion with the original amino acid mixture, whereas perfusion with nonessential amino acids had no effect. When the essential amino acids were tested individually, only phenylalanine, methionine, and valine caused significant increases in pancreatic enzyme output; the effect of tryptophan was indeterminate. However, the pancreatic enzyme output was less in response to these three essential amino acids than to mixtures containing all of them.

I ntravenous injection of polyinosinic acid/polycytidylic acid [(poly rI)·(poly rC)] offered significant protection against intranasal challenge of young mice with vesicular stomatitis virus (VSV). Optimal protection was obtained when a single dose was administered 2 hr before virus challenge, but repeated doses were effective when started as late as 3 days after virus challenge. The therapeutic ratio or ratio of maximum tolerated dose to minimum effective dose for a single intravenous injection of (poly rI)·(poly rC) 2 hr before virus inoculation was ≥8 mg/kg:0.004 mg/kg or ≥200.Dose-response curves for interferon production and antiviral protection by (poly rI)·(poly rC) were closely parallel. Equivalent doses of poly rI or poly rC alone did not exert any interferon-inducing capacity or protective effect on intranasal VSV challenge. Several factors, which are known to potentiate or antagonize interferon production, increased or decreased the interferon-inducing capacity and antiviral protection of either (poly rI)·(poly rC) or maleic acid/divinyl ether copolymer (MA/DVE) in parallel. Interferon production and antiviral protection by MA/DVE were enhanced by arginine but abolished by prior treatment with MA/DVE; DEAE-dextran (intraperitoneally), kinetin riboside and isopentenyladenosine, and prior injection of endotoxin reduced both interferon production and antiviral protection by (poly rI)·(poly rC).Treatment with exogenous interferon in amounts which closely mimicked the levels of circulating interferon produced endogenously by an effective dose of (poly rI)·(poly rC) gave protection against intranasal VSV which was identical with that dose of (poly rI)·(poly rC). This strongly suggests that interferon production accounts for the whole protective effect of (poly rI)·(poly rC) in the intranasal VSV assay.

A patient who had received multiple transfusions for complications of acute hemorrhagic pancreatitis developed a potent factor V anticoagulant with bleeding due to defective hemostasis. Despite its potency, the anticoagulant disappeared within 15 days of its first manifestation. A second patient with adenocarcinoma of the colon developed an anticoagulant to factor V postoperatively after a single blood transfusion. The anticoagulants appeared to react stoichiometrically with factor V in normal plasma in vitro. They had the physicochemical properties of immunoglobulins, and their activity was neutralized by antihuman immunoglobulin antiserum. One anticoagulant appeared to be slightly more active against homologous than against autologous factor V, but it also inhibited heterologous factor V. Both anticoagulants progressively inactivated intrinsic prothrombin activator formed from normal reagents in the incubation mixture of the thromboplastin generation test, thus confirming that factor V is required for the effective action of the intrinsic prothrombin activator. Since the anticoagulants were immunoglobulins whose activity was consumed in their reaction with factor V, consumption of anticoagulant activity was used to detect factor V antigenic material in test materials. Human serum without factor V clotting activity was found to consume anticoagulant activity, i.e., to contain inactive factor V antigenic material. Plasma from two patients with hereditary factor V deficiency (parahemophilia) failed to consume significant anticoagulant activity. Thus, the lack of factor V activity in these patients represents a deficiency of factor V molecules rather than the synthesis of a defective molecule with impaired clotting activity.

T he immunologic response of graft recipients to antilymphocyte globulin has been studied. The clearance from the serum of ¹²⁵I-labeled antilymphocyte globulin was studied in 15 graft recipients previously treated with antilymphocyte globulin and in 4 control patients not previously treated with antilymphocyte globulin. The mean serum half-life of antilymphocyte globulin was 7.2 days in control patients, 3.8 days in 13 renal graft recipients, and 22 hr in 2 heart graft recipients. All but one of the antilymphocyte globulin-treated patients had rapid clearance. Patients treated with equine antilymphocyte globulin had rapid clearance of rabbit and goat antilymphocyte globulin as well as horse antilymphocyte globulin. All patients with rapid clearance of antilymphocyte globulin had circulating antibodies to xenogeneic gamma globulin. Two patients with rapid clearance of antilymphocyte globulin had circulating complexes of antilymphocyte globulin.Five renal graft recipients were treated with aggregate-free equine gamma globulin before antilymphocyte globulin therapy in an attempt to induce tolerance to xenogeneic gamma globulin. In these five patients neither rapid clearance of antilymphocyte globulin nor significant titers of circulating antibody to xenogeneic gamma globulin developed. The induction of tolerance to xenogenic gamma globulin may benefit patients treated with antilymphocyte globulin.

A ntagonism of the antihypertensive action of guanethidine by the tricyclic antidepressants, desipramine and protriptyline, has been demonstrated in controlled studies. These antidepressants also prevent the effect of the related ring-substituted guanidinium adrenergic neuron blockers, bethanidine and debrisoquin. That the rise in blood pressure when desipramine is added to guanethidine therapy is not due simply to a pressor action of the two drugs in combination was demonstrated by the lack of an increase in blood pressure when guanethidine was added to desipramine therapy.Investigations were conducted to determine whether antagonism of guanethidine's clinical effect could result from blockade by the tricyclic antidepressants of the norepinephrine pump in the adrenergic neuron membrane, thereby preventing the uptake of guanethidine into the neuron by this pump. Like guanethidine, the indirectly acting pressor amine, tyramine, enters the neuron via the norepinephrine pump. Desipramine, protriptyline, and amitriptyline in clinical doses all were found to block the pressor action of tyramine while potentiating the pressor effect of norepinephrine. The amino acid, methyldopa, does not enter the neuron via the norepinephrine pump, and its antihypertensive action is not altered by concomitant administration of tricyclic antidepressants. It is concluded from the evidence in this investigation together with the results of previous studies in experimental animals that clinical doses of desipramine-like drugs inhibit the norepinephrine pump in the peripheral adrenergic neuron in man and thereby prevent uptake of guanethidine to its site of action.