Leon L. Miller

Determination of gastric acidity without intubation by use of cation exchange indicator compounds.

Summary The preparation of a cation exchange indicator compound called quininium resin indicator compound has been described. The rationale and method of its use have been stated. In vitro tests have demonstrated that the quininium cation present in this compound will be displaced by the hydrogen ions of dilute hydrochloric acid solutions and of the gastric juice. In vivo experiments have shown that, by administering this cation exchange resin indicator compound orally and noting the time of appearance of the quininium cation in the urine, the presence or absence of free hydrochloric acid in the gastric juice can be determined without subjecting the individual to intubation.

Alloxan Diabetes and Demonstrated Direct Action of Insulin on Metabolism of Isolated Perfused Rat Liver

The direct effect of insulin was studied in intact surviving livers removed from normal and alloxan-diabetic rats and perfused for 4 hours with rat blood containing acetate-1-C 14 . Changes due to diabetes per se were a ) decreased lipogenesis from acetate, b ) increased ureogenesis, and c ) increased incorporation of acetate into carbohydrate. The positive effects of insulin consisted of an at least partial correction of the depressed lipogenesis characteristic of diabetes and of fasting, and a net removal of glucose from the perfusate after the 1st hour. The action of insulin was inhibited in most of the experiments with ketotic liver donors, and also in many experiments in which the operative procedure was accompanied by excessive trauma. Insulin administration depressed gluconeogenesis from acetate and lowered ketogenesis in experiments with alloxan diabetic donors.

Effect of aflatox1n B1 on net synthesis of albumin, fibrinogen, and α1-acid glycoprotein by the isolated perfused rat liver☆

Abstract The isolated perfused rat liver has been used to study the influence of aflatoxin B 1 on net synthesis of the plasma proteins albumin, fibrinogen, α 1 -acid glycoprotein and α 2 -(acute phase) globulin. Aflatoxin B 1 was used because it may inhibit messenger RNA synthesis from DNA in a manner similar to that generally accepted for actinomycin D, thereby allowing estimates of half-lives of individual species of messenger RNA to be made by observation of changes in the rate of synthesis of individual proteins after administration of the drug. Livers of adult male Sprague-Dawley rats were perfused for 12 hr with defibrinated rabbit blood with l -lysine-1- 14 C and 500 mg glucose continually infused. Net changes in the specific plasma proteins were measured serologically. Aflatoxin B 1 was added to the perfusate (half the total dose at the outset and the other half infused); although a dose of 62.5 μg aflatoxin B 1 did not affect the net synthesis of albumin, fibrinogen, α 1 -acid glycoprotein or α 2 -(acute phase) globulin, doses of 125, 250, 500 or 1000 μg were associated with increasingly severe inhibition of synthesis manifest after 2–4 hr of perfusion. These data are consistent with short half-lives for messenger RNA for these plasma proteins; however, histological evidence of progressively more widespread parenchymal cell degeneration, necrosis and interstitial hemorrhage associated with doses of aflatoxin B 1 above 62.5 μg strongly suggests that the impaired protein synthesis may in part have been secondary to cytotoxic effects. l -Lysme-1- 14 C incorporation into hepatic protein was inhibited progressively by increasing doses of aflatoxin B 1 . Significant elevations in rate of 14 CO 2 production from l -lysine-1- 14 C, in urea synthesis and in α-amino acid nitrogen accumulation were observed in perfusions with more than 125 μg aflatoxin B 1 .

Chromatographic studies of amino acids in the eggs and embryos of various species

Abstract Many known amino acids have been found to occur free in very low concentration in ovarian frog eggs and larvae. In addition, several as yet unidentified ninhydrin reactive molecular species have been observed; of these, both “underglutamic acid” and “fast arginine” occur in relatively high concentration. The data obtained on hydrolysate studies indicated that there is a surprising similarity in the total amino acid composition of all the egg types, developmental stages and germ layers investigated (Table II). In only a few species specific differences in the distribution of ninhydrin reactive substances were encountered (Table III). As far as the amphibian egg is concerned, the chromatograms give no indication that new amino acids appear and others disappear in the course of embryonic and larval development. Those of the amino acids which were approached with semi-quantitative methods were found to be present in all of the tissue primordia of the amphibian embryo without showing appreciable local differences of concentration (Table I). Therefore, if there is to be an embryonic chemodifferentiation within the cellular protein components, it apparently does not occur at the level of the amino acids, but on a higher plane of molecular organization which still eludes a biochemical approach. Evidently, the egg receives from the maternal body a vast array of amino acids ready made, and the subsequent embryogenetic elaboration of structurally and metabolically specific proteins results from specific integrations and regroupings of these units rather than from their neo-formation.

Net biosynthesis of antithrombin III by the isolated rat liver perfused for 12–24 hours compared with rat fibrinogen and α-2 (acute-phase) globulin, antithrombin III is not an acute phase protein

Antithrombin III-heparin cofactor has been isolated from normal rat plasma, purified to homogeneity on acrylamide gel electrophoresis and used to prepare a monospecific antiserum in rabbits. Measurements of rat antithrombin III were made by a single radial immunodiffusion assay. Net synthesis of antithrombin III was investigated during 12- or 24-h perfusions of the isolated rat liver. In perfusions performed under basal conditions cumulative synthesis of antithrombin-III was observed to occur at a rate sufficient to replace the total circulating plasma antithrombin III in about 6 h. In perfusions performed under full supplementation conditions which greatly enhanced synthesis of fibrinogen and alpha-2 (acute-phase) globulin (known acute-phase reactant proteins) net synthesis of antithrombin III was not significantly greater than that observed in control perfusions. Although these prolonged perfusion studies conclusively demonstrate net synthesis of antithrombin III by the isolated rat liver, they afford no evidence that this protein is an acute-phase reactant.

Serum antitrypsin synthesis by the isolated perfused rat liver.

Summary The isolated perfused rat liver synthesizes antitrypsin activity in a linear fashion as long as 24 hr. The rate of synthesis may be ten times the rate necessary for physiologic levels in vivo. These data are compatible with the liver as the principal site of synthesis of serum antitryptic activity in the rat. The antitrypsin activity of the rat perfusate resembles that of normal human plasma in lability to heat (56°) and to acid (below pH 6) and in apparent molecular size (elution from Sephadex G-200). We thank Maria Luz Sevilla for expert technical assistance and Dr. Robert H. Schwartz for antitrypsin typing of the human serum. This paper is based on work partially performed under contract with the United States Atomic Energy Commission at the University of Rochester Atomic Energy Project, and partially by Grant No. 1-R01-AM110 MET and Contract No. NHLI 71-2221 from the National Institutes of Health, United States Public Health Service. It has been assigned Report No. UR-3490-519.

Studies on fibrinogen turnover before and after whole body x-irradiation in the rat.

Summary 1. The biological half-life for in vivo screened I131 labeled rat fibrinogen injected intravenously in normal rats was found to be 1.5 days. After 6–7 days the apparent biological half-life changed to 2.7 days. 2. The apparent whole-body turnover of I131 fibrinogen after whole-body X-irradiation was the same for both irradiated and control rats. However, the data on plasma I131 fibrinogen indicate that fibrinogen leaves the vascular space at an enhanced rate after whole-body irradiation.

The role of the carbon skeleton of lysine in the biosynthesis of hemoglobin

Abstract 1. 1. It has been shown that lysine-ϵ-C 14 , led to a dog, contributes significantly to the biosynthesis of the protoporphyrin IX in erythrocyte hemoglobin. 2. 2. The C 14 -activity of such hemoglobin protoporphyrin was found to be localized largely in carbon atoms C -10 and D -10 as ascertained by isolation of the two carbon atoms in question by means of the Schmidt reaction. 3. 3. A comparison of the specific millimolar C 14 -activities of protoporphyrin IX, and of glutamic and aspartic acids isolated from erythrocyte globin, suggests that the conversion of the carbon skeleton of lysine to the afore-mentioned metabolites occurs without preceding fragmentation through a five carbon intermediary metabolite which may be identical with α-ketoglutarate.

Increased Plasma IgA, sIgA, and C3- and IgA-Containing Immune Complexes With Renal Glomerular Deposits in Diabetic Rats

Male Sprague-Dawley rats were fasted 18 h and given streptozocin (STZ; 60 mg/kg body wt i.p.). The resultant diabetes mellitus, not treated with insulin, was associated with persistent manifoldly increased plasma IgA levels, as measured by single-radial immunodiffusion after reduction with dithiothreitol and alkylation with iodoacetamide. Also observed were concurrent increases in plasma levels of secretory IgA (sIgA) and of C3- and IgA-containing immune complexes (C3-IgA-CIC). After 104 days without insulin treatment, six of the diabetic rats were given daily injections of 2 U of insulin for 11 days. Insulin treatment was associated with a precipitous decrease in plasma levels of IgA, sIgA, and C3-IgA-CIC. Cessation of insulin treatment resulted in restoration of greatly increased levels of all three IgA-containing species. Histoimmunofluorescence studies of kidneys from untreated rats with diabetes of 192–324 days revealed glomerular capillary wall and mesangial deposits reacting strongly with anti-IgA (α-chain-specific) antiserum. Kidneys from two of the diabetic rats (324 days) were tested with anti-rat C3 and anti-rat secretory component (SC) antisera, and they reacted positively. Control kidneys from normal rats examined simultaneously were negative. The concurrent changes in plasma levels of three IgA-containing species in the untreated STZ-induced diabetic rat and the demonstration of abnormal immunoreactive IgA-containing renal glomerular deposits make this experiment an attractive model for studying the possible role of disturbed IgA metabolism in the pathogenesis of diabetic nephropathy.

The metabolism of hydrocortisone in the isolated perfused dog liver

Abstract Hydrocortisone was perfused through a male dog liver. Six compounds have been identified on the basis of presumptive evidence: Pregnane-3α, 17α,20α,21-tetrol-11-one, pregnane-3α, 11β,17α,21-tetrol-20-one, pregnane-3α,17α,21-triol-11,20-dione; Δ4-pregnene-11α, 17α, 21-triol-3, 20-dione; Δ4-pregnene-17α,21-diol-3,11,20-trione; and Δ4-androstene-11β-ol-3,17-dione.