Superoxide dismutase,reduced glutathione and TBA-reactive products in erythrocytes of patients with multiple sclerosis

• 1.1. Superoxide dismutase, reduced glutathione and lipid peroxides levels were determined in the erythrocytes of multiple sclerosis patients.
• 2.2. Superoxide dismutase activity and the malonyldialdehyde production rate were found to be significantly enhanced.
• 3.3. The isoelectric focusing pattern of Superoxide dismutase from multiple sclerosis and normal subjects erythrocytes was substantially overlapping.
• 4.4. Our results indicate the occurrence of a higher susceptibility of multiple sclerosis erythrocytes to lipid peroxidation.

     

Human GMP synthetase

• 1.1. The specific activity of GMP synthetase was measured in several human tissues and found to be highest in cultured skin fibroblasts, followed by bone marrow, leukocytes, erythrocytes. placenta, and liver.
• 2.2. The enzyme from fibroblasts was purified approximately 50-fold by ammonium sulfate fractionation and gel filtration.
• 3.3. The K_m values were determined to be 4.9μM for XMP, 270μM for ATP. and 340 μM for glutamine.
• 4.4. Ammonium sulfate could replace glutamine as the amino donor but was much less efficient.
• 5.5. The enzyme was specific for ATP as the energy source.
• 6.6. Unlike the calf thymus enzyme, the human enzyme has no requirement for a reduced sulfhydryl compound.
• 7.7. Human GMP synthetase is inhibited by ATP, dATP, azaserine, and hydroxylamine.

     

Fatty acid synthetase from pigeon red blood cells

• 1.1. Fatty acid synthetase has been purified 200-fold from pigeon erythrocytes.
• 2.2. The enzyme gave 2 major staining bands on disc gel electrophoresis corresponding to the complex and dissociated forms of the enzyme.
• 3.3. Sucrose density gradient centrifugation of the enzyme showed only one sedimenting peak and high performance liquid chromatography also showed only 1 major light absorbing peak.
• 4.4. The molecular weight of the enzyme was estimated to be 300,000–330,000 and the enzyme is comprised of 2 subunits of similar molecular weights.
• 5.5. The red blood cell fatty acid synthetase was found to be immunochemically nonidentical with the liver fatty acid synthetase.

     

Immobilization stress-induced antioxidant defense changes in rat plasma: Effect of treatment with reduced glutathione

• 1.1. We examined immobilization stress-induced antioxidant defense changes in rat plasma and observed the antioxidant effect of reduced glutathione (GSH) administration on these changes.
• 2.2. Immobilization stress induced severe bleeding in the stomach and a significant increase in plasma levels of thiobarbituric acid receives substances (TBARS).
• 3.3. Immobilization stress induced a significant decrease in plasma iron-binding, ironoxidizing protections and radical scavenging activity.
• 4.4. Plasma levels of ascorbic acid, ascorbyl radical and superoxide dismutase activity remained unchanged following immobilization stress.
• 5.5. Treatment with GSH showed a significant protective effect on stomach bleeding, on the increase in plasma TEARS, and on the decrease of iron-binding, iron-oxidizing protection and radical scavenging activity in plasma.
• 6.6. These results suggest that immobilization stress induces generation of reactive oxygen species and decreases the endogenous antioxidant defenses, which can be attenuated by extracellular administration of antioxidant GSH.

     

A comparative study of seafish erythrocytes and agglutinins from seaweeds

• 1.1. To date, only a few authors have assayed the agglutinic activity of marine algae against fish erythrocytes, and in these cases, mainly against freshwater fish.
• 2.2. For the first time, the hemagglutinic activity of 70 seaweeds (29 brown, 37 red and four green algae) against erythrocytes of 16 seaflsh species is reported.
• 3.3. The presence of agglutinins was demonstrated in 100% of algae assayed, against at least one of the different types of erythrocytes tested.
• 4.4. The results obtained confirm the presence of receptors for algae agglutinins on the surface of the erythrocytes of the fish studied.
• 5.5. This could be useful in establishing the origins of fish populations, as these serological differences could distinguish between populations of cultivated and wild fish.

     

Acute hepatic porphyria syndrome with porphobilinogen synthase defect

On the basis of metabolite and enzyme studies a new type of acute hepatic porphyria with porphobilinogen synthase defect and repeated intermittent acute manifestations, abdominal colics, tachycardia and hypertension, and a persistent neurological syndrome was found in two young male patients. The main characteristic features are the following:

• 1.1. High urinary δ-aminolevulinic acid excretion( ⪢ 1 mmol/24hr), slight increase of porphobilinogen (up to 25 μmol/24 hr) and high increase of porphyrins (up to 22 μmol/24 hr) with coproporphyrin dominance.
• 2.2. Normal fecal and liver porphyrins.
• 3.3. Slight increase of erythrocyte protoporphyrin.
• 4.4. Decrease of porphobilinogen synthase activity in erythrocytes in both cases below 1% of healthy and not lead-exposed persons; normal activities of uroporphyrinogen synthase and decarboxylase in erythrocytes.
• 5.5. Low-normal lead concentrations in blood and low-normal lead excretion in urine in both cases; normal lead content in bone.
• 6.6. Normal plasma and urinary amino acids.
• 7.7. Irrelevant hepatological (liver biopsy), general clinical chemical and hematological findings.
• 8.8. Diminished activity of porphobilinogen synthase in nearly all family members of both patients. From these investigations it can be concluded that there is no exogeneous, “toxic” cause of this porphyria. Porphobilinogen synthase in lead poisoning is not diminished to such an extent as demonstrated here; in contrast to lead intoxication, porphobilinogen synthase activity cannot be activated or reactivated by thiols. All clinical and pathobiochemical data point at a new enzymatic type of endogeneous acute hepatic porphyria with intermittent acute manifestations, clinically analogous to so-called acute intermittent porphyria. Porphyrin precursors and porphyrin excretion both reflects the enzymatic defect and the regulatory consequences starting with the induction of δ-aminolevulinic acid synthase.

     

Effects of disulphides and α-oxoglutarate on nuclear thiol formation and thiol content of chromatin in lysed rat spleen nuclei

• 1.1. During incubation of lysed rat spleen nuclei, there was a noted increase in total SH, accompanied by a slight decrease of chromatin SH and a more pronounced increase in SH of the non-sedimentable fraction.
• 2.2. During incubation, chromatin nucleolytic cleavage was completely inhibited by 1 mM cystamine and 1 mM oxidized glutathione (GSSG).
• 3.3. In nucleus lysates, glutathione reductase activity dependent on NADPH, and disulphide reductase NADH-dependent reducing disulphides of the non-sedimentable fraction, but not affecting GSSG, were demonstrated.
• 4.4. In the presence of 1 mM GSSG there was a transfer of SH from the non-sedimentable fraction to the rat spleen chromatin, while 1 mM 6,8-thioctic acid exerted an opposite effect. Cystamine (1 mM) was found to decrease SH content of either chromatin and the non-sedimentable fraction.
• 5.5. Reduced glutathione (GSH) and cysteamine (MEA) and CoA did not change chromatin SH.
• 6.6. In the presence of α-oxoglutarate, as in the presence of GSSG, an increase in chromatin SH was found, indicating the role of dihydrolipoate in the control of chromatin SH level.

     

The role of Zn2+ in pyridoxal phosphate mediated regulation of glutamic acid decarboxylase in brain

• 1.1. γ-Aminobutyric acid, a major inhibitory neurotransmitter in the CNS, is synthesized by glutamic acid decarboxylase which demonstrates an absolute requirement for pyridoxal phosphate.
• 2.2. At physiological concentrations, zinc stimulates the activity of pyridoxal kinase, enhancing the formation of pyridoxal phosphate, which in turn stimulates the activity of glutamic acid decarboxylase.
• 3.3. At pharmacological concentrations, zinc inhibits the activity of glutamic acid decarboxylase without inhibiting pyridoxal kinase.
• 4.4. These results suggest that zinc may play a role in pyridoxal phosphate-mediated regulation of glutamic acid decarboxylase.

     

Effect of inorganic phosphate on synthesis of 5-phosphoribosyl-1-pyrophosphate in cultured plant cells

• 1.1. Inorganic phosphate (Pi) was absorbed rapidly by suspension-cultured cells of Catharanthus roseus which had previously been cultured in Pi-free Murashige Skoog medium.
• 2.2. The intracellular levels of ATP, ADP and 5-phosphoribosyl-l-pyrophosphate (PRPP) increased markedly during the 24 hr which followed the addition of Pi (1.25mM).
• 3.3. Availability of PRPP in vivo, estimated by the measurement of nucleotide synthesis from [8-¹⁴C]adenine, was also increased by addition of Pi.
• 4.4. Only a 20% increase in the maximum catalytic activity of PRPP synthetase was observed in extracts of cells, prepared 24 hr after addition of Pi.
• 5.5. In contrast to results for mammalian PRPP synthetase, the activity of PRPP synthetase, partially purified from Catharanthus roseus, was inhibited by concentration of Pi greater than 5mM.
• 6.6. The mechanisms involved in the increased availability of PRPP and the synthesis of adenine nucleotides in the plant cells cultured in Pi-containing medium are discussed.

     

The effect of reduced salinity on tissue and plasma composition of the dogfish,Squalus acanthias

• 1.1. Dogfish (Squalus acanthias) were acclimated to reduced salinities and their plasma, muscle tissue and erythrocytes subsequently analysed.
• 2.2. Decrease in the osmolarity of the plasma was principally due to a fall in urea concentration and a significant fall in the concentrations of sodium and chloride.
• 3.3. Changes in the muscle and erythrocytes in dilute media were a decrease in urea, potassium, sodium and chloride concentrations.
• 4.4. The concentrations of the free amino acids in the muscle and the red blood cells decreased more than would be expected by the movements of water only.
• 5.5. The results were discussed in relation to the regulation of cellular volume and the involvement of the free amino acid pool of the tissues in this process.

     

Permeability of erythrocytes to glycerol and its acylated derivatives in the camel and dog

• 1.1. Glycerol and glyeerol derivatives transport was compared in human, dog and camel erythrocytes.
• 2.2. The camel erythrocytes had the slowest glyeerol transport, that of the dog erythrocytes was also slow, while the quickest transport occurred in human erythrocytes.
• 3.3. In glyeerol mono-, di- and triacetates the transport in camel and dog erythrocytes increased until in the triacetate the quickest transport was found.
• 4.4. Addition of copper ions had no inhibiting effect on glyeerol transport in camel erythrocytes but a striking effect was seen in human erythrocytes.
• 5.5. Changes in pH greatly affected transport in camel erythrocytes, a decline in pH enhanced and an increase in pH slowed the transport. Storage of camel erythrocytes did not affect the glyeerol transport in camel erythrocytes.
• 6.6. It is concluded that the mechanism of glyeerol transport in camel erythrocytes is one of a non-facilitated diffusion.

     

Effects of protein synthesis inhibitors on A- and L-site amino acid incorporation by bullfrog tadpole liver and tail fin cells

• 1.1. Cycloheximide and puromycin inhibited leucine transport and incorporation into isolated bullfrog tadpole tail and hepatic cells.
• 2.2. However, high concentrations of these 2 inhibitors did not affect alanine incorporation appreciably in either tissue.
• 3.3. NEM and DNP inhibited leucine and alanine incorporation in both cell types, but at different concentrations.
• 4.4. NEM stimulated leucine transport only in hepatocytes; alanine transport was inhibited by NEM in tail fin cells.
• 5.5. The results suggest different mechanisms of transport and protein synthesis for the 2 types of amino acids by tadpole liver and tail fin cells.

     

Regulation of horse-liver glutathione reductase

• 1.1. The enzyme was rapidly inactivated by NAD(P)H, GSH, dithionite or borohydride, while activity increased in the presence of NAD(P)+ or GSSG. NADH was more efficient for inactivation than NADPH. Redox inactivation required neutral or alkaline pH, was maximal at pH 8.5, and depended on the presence of metal cations.
• 2.2. GSSG and dithiothreitol fully protected the enzyme from inactivation at concentrations stoichiometric with NAD(P)H. Ten-fold higher ferricyanide or GSH concentrations were required to obtain partial protection. NAD+ or NADP+ were quite ineffective.
• 3.3. GSSG fully reactivated the inactive enzyme at 38°C and neutral to acidic pH (5.5–7.5). Reactivation by dithiothreitol was accomplished in short periods of time at pH 8.5 although the activity was progressively lost afterwards. Ferricyanide and GSH also reactivated the enzyme to different extents.

     

Effect of depletion of glutathione by buthionine sulfoximine on lipid peroxidation in rats acutely treated with ethanol

• 1.1. The effect of depletion of glutathione (GSH) by dl-buthionine-S,R-sulfoximine (BSO) on lipid peroxidation in rats acutely treated with ethanol was investigated.
• 2.2. BSO pretreatment has not been found to potentiate an increase in liver, brain and erythrocyte lipid peroxide levels.

     

γ-Glutamyltranspeptidase in echinoderm eggs and larvae: fertilization-dependent and developmentally-induced changes in specific activity

• 1.1. γ-Glutamyltranspeptidase is present in echinoderm eggs and larvae: in homogenates the level of activity is comparable to that of rat cerebral cortex.
• 2.2. In eggs of Lytechinus pictus, fertilization induces an early rapid and sustained (5 min–6 hr) 37% increase in the activity of γ-glutamyltranspeptidase in homogenate fractions.
• 3.3. Relative to these homogenate levels, the specific activity of γ-glutamyltranspeptidase are ≈60% lower in 40,000 g supernatant fractions and 2.7-fold higher in 40,000 g particulate fractions in both unfertilized and 15 min post-fertilized Lytechinus pictus eggs.
• 4.4. The subcellular distribution of γ-glutamyltranspeptidase is the same in both unfertilized and 15-min post-fertilized Lytechinus pictus eggs: 78% in 40,000 g particulate fractions, 22% in 40,000 g soluble fractions.
• 5.5. In both unfertilized and 15 min post-fertilized eggs of Lytechinus pictus the enzyme responds to heat (50 vs 37°C) by activation in a similar manner: 1.72- and 1.68-fold homogenates; 2.6- and 3.0-fold in supernatants; 1.97- and 1.90-fold in particulate fractions.
• 6.6. In homogenates of Pisaster ochraceous larvae, γ-glutamyltranspeptidase activity increases steadily during the course of larval development: relative to the low activity at day 5, activities exhibit an increase of 1.2-, 2.0-, 3.1- and 5.4-fold at days 10, 16, 22 and 28, respectively.

     

Ascorbate (vitamin c) and dehydroascorbate stimulation of copper induced hemolysis protective action of ceruloplasmin,albumin and apotransferrin

• 1.1. Copper ion induced lysis of rat erythrocytes was markedly stimulated by low concentrations of ascorbate and dehydroascorbate.
• 2.2. Ascorbate oxidase, superoxide dismutase, catalase or scavengers of hydroxyl radicals protected erythrocytes against copper-ascorbate stimulated lysis.
• 3.3. It is proposed that superoxide radicals and hydrogen peroxide cooperate in producing hydroxyl radicals, which are directly involved in hemolysis.
• 4.4. The serum proteins, ceruloplasmin. albumin and apotransferrin, also reduced the hemolytic action of copper-ascorbate, the order of effectiveness being; ceruloplasmin > albumin > apotransferrin.

     

A kinetic study of homocitrate synthetase activity in the yeast Saccharomycopsis lipolytica

• 1.1.|A rapid method for estimating the activity of the first enzyme of lysine biosynthesis in yeasts (acetyl-coenzyme A: 2-ketoglutarate C-acetyl transferase, EC 4.1.3.21) is described.
• 2.2.|In the wild type strain, the fixation of one substrate, S-acetyl coenzyme A, shows sigmoidal saturation kinetics. The initial rate experiments indicate that the reaction obeys an ordered mechanism, 2-ketoglutaric acid binding before S-acetyl coenzyme A.
• 3.3.|The activity is completely inhibited in vitro by lysine and by some lysine analogs, which all show cooperative binding and have an heterotrophic effect on 2-ketoglutaric binding sites. A second class of effectors is found, including 2-aminoadipic acid, pipecolid acid and dipicolinic acid, which all affect the cooperativity of S-acetyl coenzyme A binding sites.
• 4.4.|Two types of mutations which modify these inhibition patterns without affecting the catalytic activity are described. One results in a desensitization towards lysine and lysine analogs only. The other entirely abolishes the susceptibility towards the second type of inhibitors, without affecting the susceptibility to lysine.
• 5.5.|No variations of the specific activity could be detected in the wild type strain at all; mutants showing an increased or a reduced activity were isolated.
• 6.6.|Our results do not support the existence of isoenzymes at the level of homocitrate synthetase in this yeast.

     

Glycogen synthetase in the sea mussel Mytilus edulis L.—II. Seasonal changes in glycogen content and glycogen synthetase activity in the mantle tissue

• 1.1. The glycogen content of the mantle tissue reached a maximum in the summer (May–July) with levels of 41.0–53.5% of the dry tissue weight.
• 2.2. Seasonal changes in glycogen synthetase activity showed that the I-activity (independent of G6P) increased up to 10-fold in June as compared with December. The measured I-activity of glycogen synthetase was sufficient to account for the accumulation of mantle glycogen in the summer.
• 3.3. The I-activity of glycogen synthetase declined rapidly in July of each year. A possible role for the inhibition of glycogen synthetase by high levels of tissue glycogen is suggested.
• 4.4. The I-activity in the mantle tissue of mussels on the shore was higher than that for animals starved in the laboratory for 2–3 days. The differences were minimal in early May but increased markedly in late May–July. Starved mussels returned to the shore showed an increase in I-activity of glycogen synthetase.
• 5.5. Injection of 30 μmol glucose into the adductor muscle increased the concentration of glucose in the mantle fluid to 2.0–2.5 mM. A similar injection of 60 μ mol glucose resulted in a time-dependent increase in the I-activity of glycogen synthetase.
• 6.6. Injection of mussels with mammalian insulin or anti-insulin serum had no effect on the activity of glycogen synthetase. Our results are at variance with those of other workers who have used the mammalian hormone in molluscan studies (see Discussion).

     

Studies on glycogenolysis in carp liver: Evidence for an amylase pathway for glycogen breakdown

• 1.1. Glycogen-phosphorylase seems to be lacking in the carp liver. This enzymatic defect bears a resemblance to glycogen storage disease type VI, described in humans.
• 2.2. Carp liver homogenates exhibit an important γ-amylase (α-glucosidase, EC 3213) activity. By its pH curve and distribution in subcellular fractions of liver, this enzyme could be, to a large extent, of lysosomal origin.
• 3.3. During the strong hepatic glycogenolysis, which is induced in carp by insulin injections, the γ-amylase pathway could offer an explanation for glycogen breakdown in a tissue where glycogen phosphorylase is supposed to be absent.

     

Metabolic energy and cytoskeletal requirements for synthesis and secretion by acini from rat mammary gland—ii. Intracellular transport and secretion of protein and lactose

• 1.1. Iodoacetate, 2,4-dinitrophenol, cyanide and cycloheximide inhibited protein secretion as well as synthesis by acini (alveoli) from rat mammary gland. Cytochalasin B and vinblastine inhibited protein secretion and marginally reduced protein synthesis. Colchicine was without effect on protein synthesis but inhibited secretion.
• 2.2. Intracellular protein transport was altered during incubation with metabolic and cytoskeletal inhibitors. Cycloheximide, iodoacetate. 2,4-dinitrophenol and Cytochalasin B appeared to block protein synthesis on polysomes of rough endoplasmic reticulum. Vinblastine inhibited protein transport from rough endoplasmic reticulum to Golgi apparatus and colchicine appeared to cause accumulation of protein in several endomembrane fractions.
• 3.3. Iodoacetate reduced acinar lactose content but was without effect on lactose synthetase activity. Cyanide, cycloheximide and vinblastine reduced lactose synthetase activity but not tissue lactose concentration. Cytochalasin B reduced glucose incorporation but was without effect on lactose content and lactose synthetase activity. Colchicine and 2,4-dinitrophenol did not alter glucose incorporation, lactose content or lactose synthetase activity. Lactose secretion was inhibited by all metabolic and cytoskeletal inhibitors examined.
• 4.4. Results indicated that sustained protein secretion depended on continued protein synthesis and that lactose secretion was coupled to protein secretion.