Canine marrow cells were incubated with transferrin-bound 59Fe, and the partition of cellular iron was studied by chromatographic and gel filtration methods. Splitting-off of iron from the stromal fraction was avoided by lysing the cells in Tris HCl buffer at pH 8.6. Cellular iron was divided into four major compartments: stroma, microsomes, main hemoglobin, and fraction I. The iron in fraction I was found in ferritin, heme proteins, and low molecular weight iron.
Joseph V. Primosigh, E. Donnall Thomas
Suspensions of erythrocytes from patients with hemoglobin (Hb) CC disease showed an increased viscosity and decreased filterability suggesting a less deformable cell. Hemolysates prepared from Hb CC erythrocytes had an increased viscosity compared with hemolysates of normal cells, suggesting that the increased viscosity of Hb CC cells in serum was the result of an increased internal viscosity of the cell. These abnormal rheological properties of Hb CC erythrocytes were associated with a decreased content of cations and an abnormality of cell water. The fraction of the cell volume, which is water in Hb CC cells, was 95.5% of normal. The amount of cell water in Hb CC cells available for osmotic equilibrium, termed solvent water, was only 67% of that in normal cells. The smaller amount of solvent water in Hb CC cells indicates a greater amount of water bound to protein.
John R. Murphy
Blood samples containing antibodies to DNA were obtained from patients with systemic lupus erythematosus (SLE) and rabbits immunized with denatured DNA complexed to methylated bovine serum albumin. The immunoglobulin fractions from these sources did not decrease the over-all template activity of singlestranded DNA with DNA polymerase or DNA-dependent RNA polymerase. In competition studies, both DNA polymerase and DNA-dependent RNA polymerase inhibited the binding of DNA antibodies to single-stranded DNA, as evidenced by inhibition of micro-complement fixation. These findings suggest that antibodies to DNA fail to decrease denatured DNA template activity because the enzymes which use a single-stranded DNA template can displace or block the antibodies from the denatured DNA as a result of greater binding affinity to the denatured DNA. The anti-DNA antibodies associated with SLE, therefore, may not be involved in the pathogenesis of the intracellular abnormalities associated with the disease.
John N. Whitaker, Jason L. Starr
Purine metabolism was studied in fibroblasts cultured from three patients with gout in an attempt to determine the biochemical bases of their disease. The rate of purine biosynthesis de novo was normal in one line of cells, but the rate of catabolism of adenine nucleotides to hypoxanthine and inosine was greatly increased. The rate of purine biosynthesis de novo was increased in two lines of cells, and this was associated with increased concentrations of 5-phosphoribosyl 1-pyrophosphate. Purine synthesis was also less sensitive than normal to feedback inhibition. The catabolism of inosinate synthesized de novo was increased.
J. Frank Henderson, Frederick M. Rosenbloom, William N. Kelley, J. Edwin Seegmiller
Fecal bile acid and neutral sterol excretion rates were determined in five healthy young men when serum cholesterol changes were induced by isocaloric substitution of an unsaturated (safflower oil) for a saturated fat (butter). The isotope balance method was used after the intravenous injection of cholesterol-4-14C. A feces extraction method is presented which permits essentially complete separation of fecal neutral sterols and bile acids.
Richard B. Moore, Joseph T. Anderson, Henry L. Taylor, Ancel Keys, Ivan D. Frantz Jr.
The dynamics of CO2 exchange in the lungs of man was studied by observing the rate of disappearance of a stable isotope of CO2 (13CO2) from the alveolar gas during breath holding. Over 50% of the inspired isotope disappeared within the first 3 sec followed by a moderately rapid logarithmic decline in which one-half of the remaining 13CO2 disappeared every 10 sec.
Richard W. Hyde, Ricardo J. M. Puy, William F. Raub, Robert E. Forster
Long-acting thyroid stimulator (LATS) increased glucose oxidation and 32P incorporation into phospholipids in in vitro experiments with dog thyroid slices. The time course of the response was different from that obtained with thyroid-stimulating hormone (TSH), but was very similar to the delayed effect observed in vivo. During a 45 min incubation, TSH, but not LATS increased glucose oxidation, whereas in longer experiments up to 6 hr, both substances augmented 14CO2 production. Amounts of pooled human gamma globulin equivalent to LATS were inactive. Although TSH stimulated 32P incorporation into phospholipids during a 2 hr incubation, LATS was ineffective. In longer incubations, from 4½ to 8 hr, LATS did increase 32P incorporation. The stimulatory effect of LATS was not abolished by anti-TSH antibody capable of neutralizing human TSH. Effects of LATS were also obtained with beef and pig thyroid slices. In addition to stimulation of glucose oxidation in dog thyroid slices, LATS occasionally also stimulated glucose oxidation in dog spleen and liver slices. Despite a 54-fold increase in LATS concentration, a satisfactory dose-response curve could not be demonstrated when 14CO2 production was measured.
James B. Field, Adrienne Remer, Gail Bloom, Joseph P. Kriss
The effect of infusions of hyperoncotic solutions on fractional sodium reabsorption by the proximal tubule of the dog was studied by the recollection micropuncture method. Tubule fluid to plasma inulin concentration ratios were measured for identified proximal tubule segments before and after infusion of 25% albumin or dextran solutions. Results were compared with changes in fractional reabsorption during saline diuresis. Plasma volume increased 66% ± SE 5.8 after infusion of albumin solution and 94% ± SE 8.2 after infusion of dextran solution. Fractional sodium reabosorption by the proximal tubule was depressed after infusion of both of these hyperoncotic solutions. Nevertheless, changes in sodium excretion after infusion of albumin and dextran were small. In contrast, after infusions of isotonic sodium chloride solution, which increased plasma volume 61% ± SE 5.8, a decrease in fractional reabsorption of 50.7% ± SE 7.2 was associated with large changes in sodium excretion.
Stuart S. Howards, Bernard B. Davis, Franklyn G. Knox, Fred S. Wright, Robert W. Berliner
The mechanisms of the conversion of cholesterol into bile acids in man were studied by examining the metabolism of cholesterol-1,2-3H, cholest-5-ene-3β,7α-diol-7β-3H, tritiumlabeled 7α-hydroxycholest-4-en-3-one, 7α,12α-dihydroxycholest-4-en-3-one, and cholest-5-ene-3β,7α,12α-triol in fractions of liver homogenates. The 20,000 g supernatant fluid catalyzed the conversion of cholesterol into cholest-5-ene-3β,7α-diol, 7α-hydroxycholest-4-en-3-one, 7α-12α-dihydroxycholest-4-en-3-one, and 5β-cholestane-3α,7α,12α-triol. In the presence of microsomal fraction fortified with NAD+, cholest-5-ene-3β,7α-diol was converted into 7α-hydroxycholest-4-en-3-one, and when this fraction was fortified with NADPH small amounts of cholest-5-ene-3β-7α,12α-triol were formed. 7α-Hydroxycholest-4-en-3-one was metabolized into 7α-12α-dihydroxycholest-4-en-3-one in the presence of microsomal fraction fortified with NADPH and into 5β-cholestane-3α,7α-diol in the presence of 100,000 g supernatant fluid. Cholest-5-ene-3β,7α,12α-triol was converted into 7α,12α-dihydroxycholest-4-en-3-one in the presence of microsomal fraction fortified with NAD+. The 100,000 g supernatant fluid catalyzed the conversion of 7α,12α-dihydroxycholest-4-en-3-one into 5β-cholestane-3α,7α,12α-triol. The sequence of reactions in the conversion of cholesterol into 5β-cholestane-3α,7α-diol and 5β-cholestane-3α,7α,12α-triol, the subcellular localization of the enzymes, and the cofactor requirements were found to be the same as those described for rat liver.
Ingemar Björkhem, Henry Danielsson, Kurt Einarsson, Gunnar Johansson
A procedure for bioassaying parathyroid hormone-like activity in human urine has been developed. 24-hr urine samples were concentrated with dry Sephadex G-25 and bioassayed in the young thyroparathyrocauterized mouse by the measurement of whole blood calcium. Recovery of biological activity and radioiodinated beef parathyroid hormone was over 80%. Normal subjects usually excreted less than 30 U (USP) of activity per day while 18 patients with proven primary hyperparathyroidism excreted a mean of 182 U/day (USP). The activity was not found in 7 patients with hypoparathyroidism or in 5 patients with carcinoma of the breast, but was present in 9 patients with uremia and in 5 with carcinoma of the lung and hypercalcemia.
John E. Bethune, Randolph A. Turpin
The optimal conditions for the incorporation of acetate-1-14C and palmitic acid-1-14C into platelet lipids have been described. In buffer incubations with acetate there was a sharp pH optimum at 6.8; in plasma incubations, there was a broad pH optimum between 6.8-7.4. Maximal incorporation of acetate occurred at a final concentration of 1.5 mmoles/liter. In buffer, no labeled lipids were released from platelets into the medium. In plasma, 40% of newly formed lipids was recovered in the plasma. 75% of the incorporated acetate could be recovered in ceramide, lecithin, and free fatty acids. Platelet fatty acids were formed both by de novo synthesis and chain elongation. The fatty acids formed by de novo synthesis exchanged with plasma free fatty acids. In buffer incubations no turnover of newly labeled lipids occurred, but in the plasma incubations exchange of newly labeled lecithin with plasma lipids was demonstrable. Palmitic acid-1-14C added to plasma was incorporated into platelet lipids. The distribution among the lipid classes of palmitate taken up from plasma was the same as that of palmitate formed intracellularly by de novo synthesis.
Daniel Deykin, Richard K. Desser
Purified staphylococcal alpha toxin was found to inhibit the active transport of sodium across the isolated toad bladder when applied to the serosal but not to mucosal surface. Heating or the addition of specific antitoxin abolished this effect. Low temperatures reduced this activity significantly. Application of vasopressin to the bladder serosa shortly after toxin resulted in only weak and transient stimulation of sodium transport; once maximal toxin activity had been established, exposure to the hormone was without effect. Transport in bladders previously stimulated by vasopressin was rapidly inhibited by alpha toxin. Concentrations that suppressed active sodium transport completely within 30-45 min produced a significant increase in oxygen consumption by minced bladder tissue within the same period; antitoxin neutralized this activity. 2,4-dinitrophenol also inhibited sodium transport and stimulated oxygen consumption by the toad bladder. The addition of 2,4 dinitrophenol to bladder tissue in which respiration was maximally stimulated by alpha toxin resulted in a further increase in respiratory rate. The addition of toxin to bladder tissue after its exposure to a concentration of 2,4 dinitrophenol known to uncouple oxidative phosphorylation produced a significant stabilization but no increment in respiratory rate. The data are consistent with the previously suggested action of staphylococcal alpha toxin on cell membranes and suggest that energy-dependent transport processes are inhibited. The stimulation of oxygen consumption may be due to an additional effect on oxidative phosphorylation.
James J. Rahal Jr., Martin E. Plaut, Louis Weinstein
There is a profound need, on both clinical and physiologic grounds, for a measure of the contractile state of the intact ventricle. Such a measure can be obtained by evaluating the force-velocity relationship with a correction for myocardial fiber length. The force-velocity relation can be expressed as the ratio of maximum rate of pressure rise to maximum isovolumetric pressure, a quantity which was described by Hill as the maximum rate of proportional rise of pressure and which is similar to the velocity constant of a chemical reaction. Division of this ratio by an estimate of ventricular circumference corrects for variations due to differences in initial fiber length.
Martin J. Frank, Gilbert E. Levinson
The normal relationship between red cell mass measured, with 51chromium-labeled red cells, and arterial oxygen saturation (SaO2) over the range from 97.3 to 83.4% was examined by studying 73 normal men residing at sea level and altitudes of 1600 and 3100 m. A simple, linear relationship between SaO2 and red cell mass was found over the entire range (r = - 0.7524, P < 0.001). In contrast, a correlation between red cell mass and arterial O2 tension was found only over the lower half of the range of O2 tensions where SaO2 was also decreased (r = - 0.7731, P < 0.005). This suggested that O2 saturation rather than tension is the more important determinant of the erythropoietic response to chronic hypoxia. If this response is regulated by tissue O2 tension, then it will be influenced by O2 transport, which, in turn, is a function of blood flow and arterial O2 content, and hence SaO2. In nine patients with chronic obstructive airway disease the relationship between red cell mass and SaO2 was also determined and was found to be steeper than in the normal subjects (P < 0.05).
John V. Weil, Gail Jamieson, Donald W. Brown, Robert F. Grover
The present studies demonstrate that iodine depletion increases the sensitivity of the thyroid to the goitrogenic effects of thyrotropin. Iodine depletion was induced by feeding rats a low iodine diet containing propylthiouracil for 10-14 days before hypophysectomy. Accumulation of iodine in the thyroid after hypophysectomy was prevented by continuing the antithyroid drugs in the diet. Doses of thyrotropin as low as 3 mU/100 g of body weight per day produced significant thyroid enlargement in 3-7 days in iodine-depleted rats. Adding propylthiouracil or perchlorate to the diet during treatment with thyrotropin did not reduce or augment the goitrogenic response to thyrotropin in iodine-depleted rats. Increasing the level of circulating iodide also did not reduce the goitrogenic response to thyrotropin. The increased sensitivity of the iodine-depleted thyroid gland may provide an explanation for the development of thyroid enlargement without requiring an increased level of circulating thyrotropin.
George A. Bray
In adult patients with hereditary fructose intolerance (HFI) fructose induces a renal acidification defect characterized by (a) a 20-30% reduction in tubular reabsorption of bicarbonate (T HCO3-) at plasma bicarbonate concentrations ranging from 21-31 mEq/liter, (b) a maximal tubular reabsorption of bicarbonate (Tm HCO3-) of approximately 1.9 mEq/100 ml of glomerular filtrate, (c) disappearance of bicarbonaturia at plasma bicarbonate concentrations less than 15 mEq/liter, and (d) during moderately severe degrees of acidosis, a sustained capacity to maintain urinary pH at normal minima and to excrete acid at normal rates. In physiologic distinction from this defect, the renal acidification defect of patients with classic renal tubular acidosis is characterized by (a) just less than complete tubular reabsorption of bicarbonate at plasma bicarbonate concentrations of 26 mEq/liter or less, (b) a normal Tm HCO3- of approximately 2.8 mEq/100 ml of glomerular filtrate, and (c) during acidosis of an even severe degree, a quantitatively trivial bicarbonaturia, as well as (d) a urinary pH of greater than 6.
R. Curtis Morris Jr.
The effect of cholestyramine on the fecal excretion of bile acids and neutral sterols was measured in a hypercholesterolemic patient on a low fat, high polyunsaturated fatty acid-containing diet after the intravenous injection of cholesterol-4-14C. A significant (16%) lowering of serum cholesterol concentration was accompanied by a 3.2-fold increase in fecal bile acid excretion but no change in neutral sterol output. The increased bile acid loss was adequate to account for the observed fall in serum cholesterol level. The implications of these findings were discussed.
Richard B. Moore, Catherine A. Crane, Ivan D. Frantz Jr.
Internal carotid artery blood flow and arterial pressure were measured with a sine-wave electromagnetic flowmeter and a pressure transducer in 22 patients during control period and after the intravenous and intracarotid administration of norepinephrine, epinephrine, and angiotensin. Intravenous infusion of both norepinephrine and angiotensin was accompanied by an increase in cerebral vascular resistance. Administration of norepinephrine, epinephrine, and angiotensin into the internal carotid artery failed to alter blood flow immediately. However, when the systemic blood pressure increased, a concomitant passive rise in blood flow did not occur. Thus, at this time cerebral vascular resistance was significantly increased. It is concluded that these drugs do not have a direct action on the cerebral vessels, but that the increased cerebral vascular resistance after their administration is due to autoregulation or to a combination of autoregulation and reduced arterial carbon dioxide pressure (PCO2) secondary to hyperventilation.
Joseph C. Greenfield Jr., George T. Tindall
Factors involved in the hyperlipidemia of nephrosis have been studied in seven patients. The turnover of triglyceride was measured in plasma very low density lipoproteins after the injection of glycerol-14C. The turnover of esterified cholesterol was measured in whole plasma and in very low density lipoproteins after the injection of mevalonic acid-2-3H.
I. F. C. McKenzie, P. J. Nestel
The effect of acute hypertension on sodium reabsorption by the proximal tubule was studied in rats by means of micropuncture methods. Hypertension was induced by bilateral carotid artery ligation and cervical vagotomy. Within a few minutes after blood pressure rose (30-60 mm Hg above control levels), a moderate natriuresis and diuresis began. Proximal sodium reabsorption, measured by two independent methods, was found to be markedly suppressed, both in absolute amount per unit length and per unit of tubular volume (C/πr2). The ratio between tubular volume and glomerular filtration rate (GFR) (πr2d/V0) was found to be increased. These observations indicate that the inhibition of proximal sodium reabsorption induced by hypertension cannot be explained by the tubular geometry hypothesis of sodium regulation.
Karl M. Koch, Hagop S. Aynedjian, Norman Bank
An immunoadsorption technique employing a rabbit antiserum specific for human serum prealbumin has been devised to remove thyroxine (T4)-binding prealbumin (TBPA) from serum completely without affecting the T4-binding activity of thyroxine-binding globulin (TBG) or the concentration of the other major proteins in serum. As judged from the proportion of T4 associated with the antigen-antibody precipitate, only about 15% of the endogenous T4 is bound by TBPA, a value considerably less than that indicated by electrophoretic methods. As judged from the increase in the proportion of free T4 that followed immunoadsorption of TBPA, TBPA does act as one determinant of the proportion of free T4 but is far less important than TBG in this respect. A decrease in the T4-binding capacity of TBPA cannot solely account for the increase in the proportion of free T4 in the sera of ill patients, since a comparable increase does not occur in normal sera after complete removal of TBPA. From data obtained in normal and abnormal sera before and after immunoadsorption of TBPA, estimates of the equilibrium constants for the interactions between T4 and its binding proteins, as they exist in serum, have been derived. The values obtained were: KALB, 6.2 × 105; KTBPA, 2.3 × 108; and KTBG, 1.7 × 1010.
Kenneth A. Woeber, Sidney H. Ingbar
Antibodies directed against human follicle-stimulating hormone (FSH) were demonstrated in rabbit serum by neutralization of biological activity. Antibodies that bound FSH-131I were produced in rabbits and guinea pigs by repeated injections of FSH. By 131I immunochemical methods, we found that at least 90% of the FSH-131I-binding antibody failed to distinguish the four human glycoprotein hormones: FSH, luteinizing hormone, chorionic gonadotropin, and thyrotropin, purified as well as endogenous hormone in plasma. Neither growth hormone, adrenocorticotropin, nor a variety of glycoproteins or animal plasmas were able to react with these antibodies.
Sheldon Schlaff, Saul W. Rosen, Jesse Roth