Uptake Of 35S Research Articles

Gamma-Glutamyl amino acids. Transport and conversion to 5-oxoproline in the kidney.

Transport of gamma-glutamyl amino acids, a step in the proposed glutathione-gamma-glutamyl transpeptidase-mediated amino acid transport pathway, was examined in mouse kidney. The transport of gamma-glutamyl amino acids was demonstrated in vitro in studies on kidney slices. Transport was followed by measuring uptake of 35S after incubation of the slices in media containing gamma-glutamyl methionine [35S]sulfone. The experimental complication associated with extracellular conversion of the gamma-glutamyl amino acid to amino acid and uptake of the latter by slices was overcome by using 5-oxoproline formation (catalyzed by intracellular gamma-glutamyl-cyclotransferase) as an indicator of gamma-glutamyl amino acid transport. This method was also successfully applied to studies on transport of gamma-glutamyl amino acids in vivo. Transport of gamma-glutamyl amino acids in vitro and in vivo is inhibited by several inhibitors of gamma-glutamyl transpeptidase and also by high extracellular levels of glutathione. This seems to explain urinary excretion of gamma-glutamylcystine by humans with gamma-glutamyl transpeptidase deficiency and by mice treated with inhibitors of this enzyme. Mice depleted of glutathione by treatment with buthionine sulfoximine (which inhibits glutathione synthesis) or by treatment with 2,6-dimethyl-2,5-heptadiene-4-one (which effectively interacts with tissue glutathione) exhibited significantly less transport of gamma-glutamyl amino acids than did untreated controls. The findings suggest that intracellular glutathione functions in transport of gamma-glutamyl amino acids. Evidence was also obtained for transport of gamma-glutamyl gamma-glutamylphenylalanine into kidney slices.

Effect of inhibition of gamma-glutamyltranspeptidase by AT-125 (acivicin) on glutathione and cysteine levels in rat brain and plasma.

AT-125 (Acivicin) is an inhibitor of gamma-glutamyltranspeptidase (gamma-GTP) which initiates glutathione catabolism to cysteine. We measured plasma and brain glutathione and cysteine in rats treated with AT-125. Six h after AT-125 treatment, plasma glutathione had increased 6-fold and plasma cysteine had fallen significantly. Brain cysteine fell after 24 h of AT-125 treatment, and brain glutathione had also decreased 18%. AT-125 pretreatment inhibited brain uptake of 35S when it was given as 35S-GSH but had no effect when it was given as 35S-cysteine. These results suggest that plasma glutathione is catabolized by gamma-GTP, and cysteine derived from it is taken up by the brain. N-acetylcysteine was administered to AT-125 treated rats in an attempt to supply cysteine to the brain in the face of gamma-GTP inhibition. N-acetylcysteine supported brain glutathione levels, suggesting that it can serve as a source of cysteine under these conditions.

Augmentation of in Vitro Antibody Response by Disulfide Compounds I. Comparison between Intermolecular and Intramolecular Disulfides

The following disulfide compounds: 2-hydroxyethyldisulfide (2-MEox), 2-aminoethyidisulfide (cystamine) and oxidized dithiothreitol (DTTox) were found to augment the in vitro antibody response to sheep erythrocytes in murine lymphocytes as effectively as their reduced forms when they were added to the culture medium. We, however, found out that the mode of action of DTTox was greatly different from that of 2-MEox or cystamine. 2-MEox and cystamine showed augmenting effects on the proliferative response to lipopolysaccharide and on the uptake of (35S)-cystine by the lymphocytes. In contrast to these intermolecular disulfides, DTT0X, an intramolecular disulfide compound, was found to be inactive in these systems. 2-MEox and cystamine, but not DTTox, were reduced to thiol forms by the intact lymphocytes or by the cell homogenate. Thus, it is likely that DTTox did not behave as the reduced form in the lymphocyte culture in contrast to 2-MEox and cystamine, suggesting that the disulfide form itself plays an important role in augmenting effects of DTTox.

Sulphate uptake from surface water by peat

Time-dependent uptake of 35S]SO 4 2 − from surface water overlying cores of peat occurred by passive diffusion along a concentration gradient set up by SO 4 2 − metabolism in the peat. The limiting rate constant of SO 4 2 − uptake was related to concentration according to Michaelis-Menten kinetics. In peat cores taken from an area of mire submerged by surface water biologically-mediated uptake began immediately. But in cores taken from an adjacent area where the water table was about 5 cm below the peat surface, SO 4 2 − metabolism was slower and developed after a lag of about 2.3–4.0 days. Only about 2.2% of [ 35S]SO 4 2 − taken up by peat cores remained in the water-soluble pool, while about 11% was associated with acid-volatile H 2S. Most of the remainder appeared to be incorporated into organic matter. Less than 0.3% was released as H 2S into the gas phase. The experimental results are consistent with a flux into the peat of 3.28–7.71 g S m −2yr −1, comparable with 4.76–6.06 g S m −2yr −1 indicated by measurements of S content and age of the peat. The results suggest that uptake and metabolism of dissolved SO 4 2 − may be the major route of S incorporation into peat.

Human Excretion Patterns Following Occupational Uptake of 3H-, 14C- and 35S-labeled Compounds

Graphs of excretion patterns following occupational uptake of 3H, 14C and 35S labeled compounds in humans are presented. A brief description of New England Nuclear's bioassay program and analytical methods is given. One or two exponential regression curves are found to fit most sets of data. Exceptions where exponentials do not closely fit the data are discussed. Characteristic rate constants are estimated and used to establish retention equations. When groups of workers were exposed to the same source excretion parameters were not found to strongly correlate with weight or sex. Variations in individual excretion patterns following low level group exposure did not significantly effect individual dose commitment estimates. Excretion rates following uptake of 14C and 3H labeled compounds were found to span three orders of magnitude. This indicates that health physicists may tend to dissipate resources in unnecessary investigations if conservative ICRP recommendations are assumed.

The effect of temperature on the uptake and loss of anions by seedling roots ofZea mays L

Intact maize seedlings were examined for the uptake and leakage of labelled sulphate and phosphate anions affected by temperature. Control plants, grown at 25 °C were compared from the aspects of uptake capacity and leakage with plants incubated in nutrient solutions cooled to 15 °C and 5 °C, respectively. Short time intervals as well as 1–7 d exposure to cooling were used. Already after 1 h exposure at 5 °C and 5 h cooling at 15 °C and at 5 °C, considerable changes were manifested in anion uptake and leakage.The uptake of32P declined more than that of35S. So, after a 30 min uptake interval the uptake of35S decreased at 15 °C to 49.84% and at 5 °C to 6.05% comparing with the uptake at 25 °C, while the uptake of32P declined to 28.64% at 15 °C and to 4.45% at 5 °C. The leakage of both anions was the highest at 25 °C in absolute rates, but relatively most of the uptaken35S and32P was released at 5 °C. Longer exposure to a chilling temperature of 5 °C (1–7 days) resulted in two patterns of sulphate and phosphate uptake.

The use of sulphite solutions for studying the effects of SO 2 on higher plants

The effects of sulphite concentration and pH on 14Co 2 fixation or the uptake of 35S by needle segments of Pinus sylvestris were studied in factorial experiments. In addition, changes in the chemical composition of the sulphite solutions during the experiment were measured. Uptake of 14CO 2 was increased and standard errors decreased by incorporating 1·0–10 ppm of Tween 80 into the solutions used. Inhibition of 14CO 2 fixation by sulphite was significantly greater at low pH and high sulphite concentration, with a significant interaction. Uptake of Na 2 35SO 3 was greater at low pH and was linear with respect to the concentration of Na 2SO 3 carrier added between 10 −3 and 10 −4 m. In the absence of carrier, recovery of 35S at the end of the experiment was reduced. Oxidation of the sulphite solutions was rapid at high pH and low concentration, with a significant interaction. It was also accelerated by the presence of plant material. The use of sulphite solutions to predict responses of higher plants to SO 2 is discussed.

In vitro incorporation of ( 35S)-methionine in mitochondrial proteins of drosophila melanogaster

In order to study the interrelation between nuclear genome and mitochondria during biogenesis of mitochondrial proteins, mitochondria of D. melanogaster were isolated from whole insects. The fraction obtained showed respiratory control by ADP in the presence of pyruvate-malate substrates of up to 6–7. Conditions of in vitro uptake of ( 35S) methionine in polypeptides were studied. The energy necessary for protein synthesis is supplied by endogenous ATP. Bacterial and fungal contamination was severely repressed. Incorporation rates of 50–100 picomoles of( 35S) methionine mg protein −1 hour −1 could be measured under these conditions. Electrophoresis in the presence of SDS of proteins of the internal membrane showed, after 50 min of incubation, a significantly labelled band of proteins at 22 Kd. Other bands at 80, 52 and 40 Kd were labelled to a lesser degree.

Glycosaminoglycan and collagen synthesis in N-nitroso-N-methylurethane induced pulmonary fibrosis.

SummaryGlycosaminoglycan and collagen synthesis were studied in a hamster model of interstitial pulmonary fibrosis, induced by weekly subcutaneous injections of N-nitroso-N-methylurethane (NNNMU) for up to 16 weeks. Experimental and control animals were injected intraperitoneally with 35S or [3H]proline to label glycos-aminoglycans and collagen, respectively. The labeling studies were performed at 1 and at 3 months following completion of NNNMU treatment. Uptake of 35S into lung glycosaminoglycans was higher in diseased animals than in controls at both 1 and 3 months post-NNNMU. The experimental lungs had a significantly increased percentage of labeled dermatan sulfate and/or chondroitin 4-sulfate compared to controls at both time intervals. Additional studies performed only on the 1-month samples showed that this increase in percentage labeling was primarily in dermatan sulfate. NNNMU-treated lungs also had a significantly lower percentage of labeled heparin and/or heparan sulfate than did controls at 1 ...

Effect of pH on the Uptake of IIS-Sulfur from Sulfate, Sulfite, and Sulfide by Isolated Spinach Chloroplasts

The rates of uptake of 35S from S(VI), S(IV), and S(-II) solutions by isolated spinach chloroplasts were measured at a variety of solution pH values between pH 5 and pH 8. S(VI) showed a constant uptake rate of about 3.6 nmol yl-1 stroma space min-1 within the whole range of pH. S(IV) gave an uptake rate of a similar value at higher pH values, which increased to almost double as pH decreased to pH 5. S(-II) showed uptake rates ranging from 10 nmol ,ul-1 stroma space min-' at pH 7.5 to 35 nmol ,ul-1 stroma space min-' at pH 4.9. The increasing uptake of S(IV) and S(-II) with decreasing pH has been related to. the simple diffusion of S02 - H2O and H2S(aq), respectively, across the inner chloroplast membrane. It is not under metabolic control and with S02 - H2O it takes place in addition to the transport of the ionic forms via the phosphate translocator. With all sulfur compounds investigated an increase in uptake with increase in light intensity was found. Whereas the light triggered uptake of S(VI) and of S(IV) earlier was found to be related to ATP and to carrier-SH group supply, respectively, the light enhanced uptake of S(-II) is independent of both. The results point to the importance of pH in the medium for toxicity levels of S02 and of H2S, especially in aquatic environments.

Chlorpromazine Reverses Diarrhea in Piglets Caused by Enterotoxigenic Escherichia coli

The effect of chlorpromazine (CPZ) on diarrhea caused by enterotoxigenic Escherichia coli was tested in piglets since CPZ has been shown to be a potent antagonist to enterotoxins in vitro in a cell system and in vivo in a mouse model. Experimental diarrhea was induced in three litters of newborn piglets which were infected by mouth with 2 x 10(9)E. coli bacteria, which produce heat-labile (LT) and heat-stable (ST) enterotoxins. Treatment with CPZ given intramuscularly 1 h after the onset of diarrhea reversed fluid secretion in small intestine as well as dehydration, as judged by clinical criteria. A dose of 5 mg of CPZ per kg of body weight completely normalized the intestinal-fluid content measured 4 h after diarrhea developed, whereas 1 to 2 mg of CPZ per kg of body weight was somewhat less effective but still caused significant reduction of fluid (P < 0.001). Studies with radioactive [(35)S]CPZ showed preferential and dose-dependent uptake of (35)S in the intestinal mucosa, the radioactivity being evenly distributed in the membranes of both crypt and villus cells. The enzyme adenylate cyclase, which probably mediates the cellular effects of LT, was shown to have two- to threefold higher activity in the infected than in the uninfected animals. This activation was reduced about 50% by the CPZ treatment (2 mg/kg of body weight). In a preliminary field trial the effect of CPZ was tested in a spontaneous outbreak of diarrhea in piglets due to enterotoxinogenic E. coli. The animals were treated either with oral electrolyte solution and standard antimicrobial agents only (controls) or with 1 mg of CPZ per kg of body weight intramuscularly in addition to this treatment. The mean duration of diarrhea in CPZ-treated animals was significantly shorter, 4.1 h (n = 23), than that in controls, 7.2 h (P < 0.05).

Distribution of 35S labeled sulfated mucopolysaccharids in the recipient eye after corneal allografting

The release of 35S labeled corneal glycoprotein from an allogenic interlamellar corneal graft was studied in randomly selected rabbits. Simultaneously, the uptake of 35S by the recipient animal was investigated. These changes followed for a period of 80 h indicate that there is a constant 35S release fromthe donor cornea and that it is the recipient cornea which takes up the major part of the 35S from the donor. The iris, choroid, and sclera of the recipient also take up a certain percentage of the donor's 35S while the aqueous humor, the optic nerve, and the local lymph nodes take up only minimal amounts.

TRANSLOCATION AND TRANSFER OF NUTRIENTS IN VESICULAR‐ARBUSCULAR MYCORRHIZAS

SUMMARY Trifolium repens (white clover) or Allium cepa (onion) infected with the mycorrhizal fungus Glomus mosseae were grown in split‐plate cultures, so that nutrients could be supplied to the external hyphae only. The uptake and translocation of 32P, 65Zn and 35S by hyphae of the fungus was measured.A lag phase of 2‐3 days in the appearance of 32P in the shoot was caused by a delay in the uptake and translocation of P by the hyphae, rather than in transfer from fungus to host.The calculated maximum translocation rate of P for onion was 6.4 × 10−16 mole s−1, and the P flux in the hyphae about 2 × 10−10 mole cm−2 s−1. The molar amounts of P, S and Zn translocated were in the ratio of 35:5:1 and the mean fluxes in the ratio of 50:8:1 with clover.

Effect of pulmonary edema on drug transport and binding in rat lung.

The pulmonary absorption and uptake of (35S)phenol red ((35S)PR was measured in anesthetized rats with alpha-naphthylthiourea- (ANTU) induced lung edema. When (35S)PR solution was injected through a tracheal cannula in control animals and the percentage of the tracheal cannula in control animals and the percentage of the dose unabsorbed plotted semilogarithmically against time, an apparent first-order absorption rate was obtained. In contrast, in rats with ANTU-induced edema, the absorption time curve showed at least two different first-order components. Increasing the concentration of (35S)PR from 0.01 to 3 mM resulted in a decrease in the percentage absorption of the compound in controls compared with a relatively constant percentage absorption in edematous lungs. (35S)PR uptake by lung slices from ANTU-treated rats was decreased in the presence of IAA and a N2 atmosphere, and the dye accumulated at a faster rate and to a greater extent than in controls. The results suggest that although energy-dependent drug transport mechanisms remain intact, the porosity of the absorbing membranes and the extent of drug binding in the lung are increased markedly in the presence of edema.

Long-term effects of chondrocyte death on rabbit articular cartilage in vivo

The effects of freezing localized areas of rabbit articular cartilage in vivo were studied at two to twelve months morphologically (gross and microscopic examination, including scanning electron microscopy), histochemically (toluidine blue and safranin O), and metabolically (35S uptake). Up to six months the frozen cartilage was intact but appeared to be dead, as shown by the absence of stainable chondrocytes, severely decreased acid mucopolysaccharide content, and absence of significant uptake of 35S. At twelve months fibrillation and softening were evident, clusters of new chondrocytes with surrounding acid mucopolysaccharides were visible microscopically, and scanning electron microscopy revealed an irregular pattern of collagen fibers which were larger than normal, of varying thickness, and broken in some areas. These changes resembled those seen in degenerative joint disease.

The uptake of35S by hypothalamic and neurohypophysial proteins following intraventricular injection ofl-cysteine-35S-hydrochloride in rats dehydrated and reserpinized

In rats dehydrated for 48 h and injected intraventricularly with L-cysteine-35S-hydrochloride, the specific activity of TCA-precipitable material, both in the hypothalamus and neurohypophysis, was found to diminish under reserpine treatment.

LASMA AMINO ACID CONCENTRATIONS AND 35S-DISTRIBUTION IN THE SOW AND FETUS FOLLOWING INJECTION OF 35S-METHIONINE

Following maternal intravenous injection of 1.6 mg/kg body weight 35S-methionine, only a small proportion of the administered 35S reached fetal circulation. Fetal liver uptake of 35S was much less than that of maternal liver. A transient minor increase in the maternal plasma methionine concentration was not reflected in fetal plasma. Isoleucine, leucine, valine and phenylalanine were at lower concentrations in fetal plasma than in maternal plasma, with threonine, glutamic acid, proline, glycine, alanine and tyrosine generally higher in fetal blood. The proportion of 35S reaching the fetal circulation appeared to be cleared at a slower rate than the 35S from maternal plasma. The placenta appeared to function as an effective buffer against transient minor changes in the maternal plasma methionine concentration. The extent of the buffering capacity of the placenta against maternal plasma amino acid fluctuations requires further investigation.

AGE-DEPENDENT VARIATION IN CARTILAGE RESPONSE TO SOMATOMEDIN

The role of variation in end-organ cartilage response to somatomedin (SM) as a significant determinant of growth velocity has been investigated.The response of costal cartilage of foetal and post-natal rabbits to SM has been assessed by simultaneously measuring 35S and 3H-thymidine uptake in vitro. Cartilage from rabbits of known growth rate at 23 and 30 days after conception (gestation 33 days), and at 1, 7, 17, 28, 39, 64, 90 and more than 200 days after birth, was studied. An excellent correlation between thymidine uptake and the growth velocity of the costal cartilage was shown. The uptake of 35S was also closely related to growth velocity and the 3H/35S ratio declined as growth rate slowed.Serum SM of each age group was measured by the porcine cartilage technique of Van den Brande. The post-natal serum SM increased progressively with age to an adult plateau.This study suggests that the growth velocity of costal cartilage is determined by the combined effects of end-organ response and serum SM stimulus.

STABILITY OF SULFIDES OF MANGANESE, IRON, ZINC, COPPER, AND MERCURY IN FLOODED AND NONFLOODED SOIL

The stability of heavy metal sulfides (MnS, FeS, ZnS, CuS, and HgS tagged with 35S) in aerobic and anaerobic soil was studied. The sulfides are listed in decreasing order of solubility. Water extracts of each treated soil were made regularly during a 75-day incubation period. The radioactivity of the extracts was indicative of the stability of the water-soluble salts. For the aerobic soils, H2O-extractable 35S decreased with a decrease in solubility of the sulfide. The metal-sulfides were relatively stable in the anaerobic soil. The least soluble salt was the most stable. To study the oxidative capacity of rice (Oryza sativa L.) roots the 35S tagged metalsulfides were mixed with anaerobic soil to which rice was transplanted. The tagged soil was placed in the lower rooting zone and covered to a depth of 8 cm with unlabeled soil. The transplanted rice plants were grown for specific times and the aerial portions harvested. The uptake of 35S from the respective sulfide salt was determined. The apparent degree of stability and subsequent uptake of 35S was directly related to the solubility of the sulfide salt. No 35S was taken up from the Hg35S.

Autoradiographic study of chick limb cartilage in vivo and in vitro and its response to mitomycin C (MMC).

Effect of mitomycin C on mucopolysaccharide synthesis by the cartilage cells of chick limb bud (5-9 days old) was investigated both in vivo and in vitro by the uptake study of 35S. Control specimens (both in vivo and in vitro) showed higher concentration of silver grains, mostly intercellular, in the initial stages. Hypertrophy of cartilage cells at later stages reduced the matrix, leading to a corresponding decrease in the accumulation of silver grains. In the treated groups, both in vivo and in vitro, the silver grains were found to be less in number during the initial stage of growth than in the corresponding controls, due to the suppressive effect of the mitomycin C. However, during the later period of growth, uptake of 35S was more in the matrix of treated specimens, which was apparently not reduced due to lack of hypertrophy of cartilage cells. The mechanism of the suppressive effect of mitomycin C seems similar in vivo and in vitro.