Contribution to the genetics of serum β-lipoproteins in man

Summary Blood and serum from 17 Macaca mulatta were analysed for haptoglobins, transferrins, 6-phosphogluconate dehydrogenase, adenylate kinase, adenosine deaminase, phosphoglucomutase, acid phosphatase, glucose-6-phosphate dehydrogenase and phosphohexose isomerase. Compared to the human pattern, the AcPh and the ADA components of macaques are fast moving; AK, PGM, 6-PGD and G-6-PD have almost uniform and similar electrophoretic mobilities; and the Hp, Tf and PHI show differential mobilities. All these macaques possess similar karyotypes.Aided by UGC Junior Research Fellowship.     

Glucose-6-?hosphate dehydrogenase deficiency,haptoglobin and hemoglobin variants in dogs

Blood samples from 31 male and 34 female, adult, healthy dogs of different breeds were studied for erythrocytic G-6-PD activity. The hemolysates were also studied electrophoretically for G-6-PD, 6-PGD, and hemoglobin variants. Most of the plasma samples revealed a human Hp 1–1 type of band while three samples had an additional fast-moving band that disappeared on addition of an excess of hemoglobin and two samples were ahaptoglobinemic. G-6-PD deficiency was detected in eight samples, and it was more frequent in males than in females. The implications of G-6-PD deficiency with no difference in the electrophoretic pattern and of ahaptoglobinemia are discussed with respect to different genetic and clinical possibilities.     

Effects of prolactin and androgens on enzymes of carbohydrate metabolism in prostate of castrated bonnet monkeys Macaca radiata (Geoffroy).

Effects of prolactin (PRL), bromocriptine (Br), testosterone propionate (TP), dihydrotestosterone (DHT) and the combinations of these androgens with PRL/Br on the specific activities of caudal and cranial prostatic cellular enzymes involved in carbohydrate metabolism in castrated mature bonnet monkeys have been studied. Castration decreased all the enzymes studied such as hexokinase (HK), 6-phosphofructokinase (6-PFK), glyceraldehyde-3-phosphate dehydrogenase (G-3-PD), pyruvate kinase (PK), glucose-6-phosphate dehydrogenase (G-6-PD) and 6-phosphogluconate dehydrogenase (6-PGD) in the cranial and caudal prostates. PRL elevated the activities of all the enzymes above normal except G-3-PD of cranial lobe. In the caudal lobe, PRL brought back the activities of HK, PFK, PK, G-6-PD to normal and 6-PGD above normal except G-3-PD. TP/DHT treatment increased all the enzymes in both the lobes. PRL given along with TP/DHT further enhanced the androgen action with regard to HK, PK, G-6-PD and 6-PGD of cranial and PFK, G-3-PD, PK, G-6-PD and 6-PGD of caudal lobe. Br treatment did not produce any alteration of these enzymes in both the lobes. In the cranial lobe, during Br+TP/DHT treatment, the stimulating effects of androgen were unaffected on all the enzymes except PK. On the other hand in the caudal, the stimulatory effects of androgens were affected and the activities of HK, PFK, PK and 6-PGD were significantly decreased. The present results suggest that PRL has a direct as well as a synergistic action with androgens on enzymes of EMP and HMP shunt in the prostates of monkeys.     

Interstrain differences in red cell enzyme activities in mice and rats.

1. Interstrain differences in red blood cell enzyme activities were studied in mice (BALB/c, C57BL/6, C3H/He, DBA/2 and ddY) and rats (Donryu, F344/N, SD, Wistar and Wistar/ST), and were also compared with hamster, guinea-pig and rabbit. 2. The enzyme activities measured were: glutathione S-transferase (GST), glucose-6-phosphate dehydrogenase (G-6-PD), 6-phosphogluconate dehydrogenase (6-PGD), NADPH-diaphorase (ND), hexokinase (Hx), glutamate oxaloacetate transaminase (GOT), lactate dehydrogenase (LDH) and acetylcholinesterase (AChE). 3. There were marked variations in the activities of some red cell enzymes (e.g. GST, Hx, ND), while others (e.g. G-6-PD, 6-PGD) were much less variable both within different strains and species.     

The relationship between red cell aging and enzyme activities in experimental animals.

1. The relationship between red cell aging and enzyme activities was studied in rabbit, guinea-pig, hamster, rats (F344/N and SD), and mice (BALB/c and DBA/2). 2. The activities of six enzymes: glucose-6-phosphate dehydrogenase (G-6-PD), 6-phosphogluconate dehydrogenase (6-PGD), hexokinase (Hx), glutamate oxaloacetate transminase (GOT), lactate dehydrogenase (LDH) and acetylcholinesterase (AChE), were measured in the red cells of different ages which were obtained either by centrifugation or experimental anaemia. 3. Hx, AChE and GOT activities were much higher in younger red cells than in older cells, hence the activities of these enzymes may be used as an indicator of age of the cells.     

Activity of some glycolytic and pentose phosphate pathway enzymes during the development of adventitious roots

Activities of phosphofructokinase (PFK, EC 2.7.1.11), glyceraldehyde 3-phosphate (NAD) dehydrogenase [G-3-PD(NAD), EC 1.2.1.12], glucose 6-phosphate dehydrogenase (G-6-PD, EC 1.1.1.49), and 6-phosphogluconate dehydrogenase (6-PGD, EC 1.1.1.44) were determined in bean cuttings (Phaseolus vulgaris L. cv. Top Crop) over 4 days, encompassing adventitious root primordium initiation and development. Effects of applied auxin and “endogenous root-forming stimulus”(ERS) on enzyme activities, concentrations of reducing sugars, and primordium development were also determined during the first 4 days of propagation. Effects of auxin were determined through use of applied indole-3-acetic acid (IAA) or 2,3,5-triiodobenzoic acid. Effects of ERS were evaluated by means of decapitation of cuttings. Increased basipetal transport and increased metabolism of reducing sugars occurred in leafy cuttings in response to applied IAA and to ERS. Primordium development and activities of the four enzymes increased in leafy cuttings under conditions that simultaneously increased basipetal transport and metabolism of reducing sugars. Three types of enzyme activity response were found: (i) activity increased over time by ERS and by applied IAA [G-3-PD(NAD)], (ii) activity increased over time by ERS but not by applied IAA (PFK, G-6-PD), (iii) activity increased over time but not by ERS or applied IAA (6-PGD). Increases in G-3-PD(NAD), G-6-PD, and PFK activity in leafy cuttings were positively related to primordium development. 6-PGD activity increased in leafy cuttings during primordium development and may have supported it. However, equal increases occurred in decapitated cuttings, in which the long-term development of primordia was supressed. Results for G-3-PD(NAD) that were obtained in an experiment with jack pine (Pinus banksiana Lamb.) seedling cuttings were similar to results for the same enzyme in bean cuttings. G-3-PD(NAD) activity in naphthaleneacetic acid-treated jack pine cuttings increased with time, in comparison with untreated cuttings, before root emergence.     

Enzyme activities during imbibition and germination of seeds of tamarack (Larix laricina)

Activities of six enzymes from extracts of separated embryos and gametophytes of tamarack [ Larix laricina (Du Roi) K. Koch] seeds were assayed at various stages of imbibition and germination. On a per seed part basis, activities of 6-phosphogluconate dehydrogenase (6-PGD, EC 1.1.1.44), glucose-6-phosphate dehydrogenase (G-6-PD, EC 1.1.1.49), malate dehydrogenase (NAD⁺–MDH, EC 1.1.1.37), isocitrate dehydrogenase (NADP⁺–IDH, EC 1.1.1.42), soluble peroxidase (PER, EC 1.11.1.7), and acid phosphatase (ACP, EC 3.1.3.2) from both the embryo and gametophyte tissues generally increased slowly, following cold stratification for 30 days and imbibition under germinating conditions for 5 days, but then increased at a faster rate with emergence of the radicle and subsequent growth of the seedling. The rate of increase of enzyme activity was highest for PER. Soluble protein levels also increased with imbibition and germination, with about 3 times greater levels present in the gametophyte than in the embryo. Heat inactivation experiments showed that, except for G-6-PD, activities were stable up to 40°C. Inactivation occurred at lower temperatures for G-6-PD, while higher temperatures were required for PER. Incubation of extracts for 7 days at 4°C indicated that loss of enzyme activity was greatest for G-6-PD (3.9% remaining) and least for PER and ACP (94 and 95% remaining, respectively).     

Studies of enzymes connected with erythrocyte glutathione metabolism in a rural tropical population

Summary The activities of the erythrocyte enzymes hexokinase (HK), glucose-6-phosphate dehydrogenase (G-6-PD), 6-phosphogluconate dehydrogenase (6-PGD), glutathione reductase (GR) and glutathione peroxidase (GSH-PO) were determined in a group of 12 Europeans and in a group of 103 male Thai subjects in northern Thailand. In the Thai group there were 16 subjects with G-6-PD deficiency and 28 subjects with abnormally low levels of GR activity. A comparison of the enzyme activities in the different subgroups indicated that HK and 6-PGD are not influenced by G-6-PD deficiency whereas GR and GSH-PO activities are significantly higher in G-6-PD deficient subjects. In the group with low GR activity G-6-PD and GSH-PO showed a tendency to an elevation of activity when compared with the normal control group. Significant positive correlations exist between G-6-PD and 6-PGD in the normal group and between GR and GSH-PO in the G-6-PD deficient group. A negative correlation between GR and GSH-PO was present in the group with low GR activities. A study of the families of subjects with low activity of GR did not yield evidence for the existence of a deficiency polymorphism.

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Zusammenfassung Bei 12 Europäern und einer Gruppe von 103 männlichen thailändischen Versuchspersonen wurden die Aktivitäten der Erythrocytenenzyme Hexokinase (HK), Glucose-6-Phosphat-Dehydrogenase (G-6-PD), 6-Phosphogluconat-Dehydrogenase (6-PGD), Glutathion-Reduktase (GR) und Glutathion-Peroxidase (GSH-PO) bestimmt. In der Thai-Gruppe waren 16 Personen mit G-6-PD-Mangel und 28 Personen mit abnormal niedrigen Aktivitäten der GR. Ein Vergleich der Enzymaktivitäten in verschiedenen Untergruppen zeigte, daß HK und 6-PGD durch G-6-PD-Mangel nicht beeinflußt werden. Im Gegensatz hierzu sind die Aktivitäten der GR und der GSH-PO bei G-6-PD-Mangel signifikant erhöht. In der Gruppe mit erniedrigter GR-Aktivität bestand eine Tendenz zu erhöhten Werten für G-6-PD und GSH-PO. Die Korrelationen zwischen G-6-PD und 6-PGD in der Gruppe mit normaler G-6-PD und die zwischen GR und GSH-PO in der Gruppe mit G-6-PD-Mangel waren signifikant. In der Gruppe mit erniedrigter GR-Aktivität fand sich eine negative Korrelation zwischen GR und GSH-PO. Die Untersuchungen in Familien von Personen mit niedriger GR-Aktivität ergaben keinen sicheren Hinweis auf das Vorliegen eines GR-Mangel-Polymorphismus in der untersuchten Bevölkerung.

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Established and supported by Stiftung Volkswagenwerk, Hannover.     

The Metabolic and Phagocytic Activities of Leukocytes from Patients Receiving Corticosteroid and Radiation Therapy,and Patients with Bacterial Infections

Peripheral blood leukocytes from patients given corticosteroid or radiation therapy, as well as patients with bacterial or viral infections, were studied with regard to the selected enzyme activities of the hexose monophosphate shunt (HMS). Glucose-6-phosphate dehydrogenase (G-6-PD), 6-phosphogluconate dehydrogenase (6-PGD) and reduced nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase were assayed spectrophotometrically on mixed leukocyte suspensions in isotonic glycerol. Enzyme activities of G-6-PD and NADPH oxidase in patients receiving corticosteroid or radiation therapy were significantly lower than the enzyme activity of 6-PGD. In patients with bacterial infections, activities of the three enzymes increased but in patients with viral infections, only the activities of NADPH oxidase and G-6-PD were slightly decreased. Nitroblue tetrazolium (NBT) dye-reducing activities of neutrophils from patients receiving corticosteroid or radiation therapy were attenuated which coincides with the reduced activities of HMS enzymes. From these results, it is likely that the reduced activities of intraleukocytic HMS enzymes of patients receiving corticosteroid or radiation therapy are correlated with intracellular bactericidal activities which might result from the attenuated level of hydrogen peroxide production.     

Human platelet glucose-6-phosphate dehydrogenase. Total purification, kinetic studies and relationship with enzyme from other blood cells.

Human platelet G-6-PD has been highly purified, to homogeneity, and its kinetic, electrophoretic and immunological characteristics have been studied. Platelet G-6-PD differs from erythrocyte or leukocyte enzymes by an increased Michaelis constant for G-6-P and a slow activity at the acid pHs. By electrofocusing only a main active band (band a) of platelet G-6-PD was found. The incubation at 37 degrees C in the presence of NADP+ and dithiothreitol normalize Km-G-6-P of platelet G-6-PD; the incubation with boiled and ultrafiltered leukemic granulocyte extracts led to an anodisation of G-6-PD active forms, a decrease of the molecular specific activity and a further increase of Km-G-6-P; these last modifications are the same as those undergone by G-6-PD incubated in crude extracts of normal or leukemic granulocytes.     

Characteristics of a new abnormal variant of G-6-PD in human red cells.

Kinetic and electrophoretic properties were studied in 230--300 fold purified preparations of glucose-6-phosphate dehydrogenase (G-6-PD) from red cells of donors and patients with hemolytic anemia induced by G-6-PD deficiency. In abnormal variant of G-6-PD isolated from red cells of a patient with hemolytic anemia which had not before been described in the literature was found. The abnormal variant differs from the normal enzyme by a decreased Michaelis constant for G-6-P and NADP, by increased utilization of substrate-analogues (2-deoxy-G-6-P and deamino NADP in particular), by low heat stability, the character of pH dependence, and by the appearance of one band of G-6-PD activity during electrophoresis in polyacrylamide gel. The isolated abnormal variant of G-6-PD has been called "Kremenchug" according to the origin of the patient.     

The metabolic and phagocytic activities of leukocytes from patients receiving corticosteroid and radiation therapy, and patients with bacterial infections.

Peripheral blood leukocytes from patients given corticosteroid or radiation therapy, as well as patients with bacterial or viral infections, were studied with regard to the selected enzyme activities of the hexose monophosphate shunt (HMS). Glucose-6-phosphate dehydrogenase (G-6-PD), 6-phosphogluconate dehydrogenase (6-PGD) and reduced nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase were assayed spectrophotometrically on mixed leukocyte suspensions in isotonic glycerol. Enzyme activities of G-6PD and NADPH oxidase in patients receiving corticosteroid or radiation therapy were significantly lower than the enzyme activity of 6-PGD. In patients with bacterial infections, activities of the three enzymes increased but in patients with viral infections, only the activities of NADPH oxidase and G-6PD were slightly decreased. Nitroblue tetrazolium (NBT) dyereducing activities of neutrophils from patients receiving corticosteroid or radiation therapy were attenuated which coincides with the reduced activities of HMS enzymes. From these results, it is likely that the reduced activities of intraleukocytic HMS enzymes of patients receiving corticosteroid or radiation therapy are correlated with intracellular bactericidal activities which might result from the attenuated level of hydrogen peroxide production.     

Regulation and rate of the hexose monophosphate shunt in Rana ridibunda erythrocytes

1. Resting rates of Rana ridibunda erythrocyte glucose consumption and 14CO2 production from 1-14C-glucose were found to be significantly lower than the respective values in human erythrocytes. 2. In the presence of 1-14C-glucose Methylene Blue stimulated 14CO2 production 7-fold, while in the presence of 6-14C-glucose Methylene Blue stimulated 14CO2 production 1.2-fold. 3. The Km of G-6-PD for G-6-P and NADP were 29 and 12 microM, respectively while the Km of 6-PGD for 6-PG and NADP were 83 and 32 microM, respectively. The Ki of G-6-PD and 6-PGD for NADPH were 80 and 12 microM, respectively. 4. Excess amounts of NADP resulted in a significant decrease of 14CO2 production from 1-14C-glucose in total haemolysates. 5. ATP, ADP and fructose diphosphate inhibited both G-6-PD and 6-PGD, the latter being more sensitive than G-6-PD to their inhibitory effect, 2,3-DPG and reduced and oxidized glutathione showed a marked inhibitory effect on 6-PGD, while the phosphorylated trioses inhibited only G-6-PD. 6. Physiological concentrations of oxidized glutathione decreased the inhibition exercised by NADPH on G-6-PD. 7. The possible role of the two dehydrogenases in the regulation of the HMS is discussed.     

Effect of tuftsin on the activity of energy metabolism enzymes in peritoneal macrophages

Cytochemistry was used to measure the activity of succinate dehydrogenase (SOD), lactate dehydrogenase (LDH) and glucose-6-phosphate dehydrogenase (G-6-PDH) in rat peritoneal macrophages under the action of the endogenous immunostimulant tuftcin (tre-lys-pro-arg) during phagocytosis of latex particles and at rest. Tuftcin did not affect the activity of the study enzymes in non-phagocytic cells. Elevation of the peptide concentration to 0.25 micrograms/ml and higher in phagocytic macrophages activated G-6-PDH and lowered the activity of LDH. Tuftcin did not alter the activity of SOD in phagocytic macrophages.     

Gene dosage effects in human diploid and tetraploid fibroblasts

The effects of cell ploidy on the biochemical characteristics of cultured cells were compared using human diploid vs tetraploid fibroblasts isolated with a non-selective method. Their DNA replication was compared by thymidine incorporation, and DNA content by Feulgen staining and quantitative analysis. Their RNA and protein content, cell sizes and the specific activities of glucose-6-phosphate dehydrogenase (G-6-PD) and 6-phosphogluconate dehydrogenase (6-PGD) were assayed quantitatively. With the exception of RNA content, all other parameters demonstrated a 2-fold increase reflecting the increase in cell ploidy. These direct gene dosage effects on the genetic material and functional expression of the human genome were in contrast to previous observations in other species and validate the use of human intraspecific euploid hybrids for biochemical and genetic studies.     

Red cell antigen, serum protein and red cell enzyme polymorphisms in Karkar Islanders and inhabitants of the adjacent North Coast of New Guinea

Blood samples from the Waskia and Takia populations of Karkar Island, Papua New Guinea, and other nearby mainland populations, were tested for genetic variation in blood group, serum protein and red cell enzyme systems. Polymorphic variation was present in the ABO, P, MNS, Rh, Lewis, Duffy, Kidd and Gerbich blood group systems, in the Hp and Tf serum protein systems, and in the acid phosphatase, 6-PGD, ADA, PGM, MDH, and G-6-PD enzyme systems. A small number of variants was found in other systems: there were 4 Lu(a+), 1 Kp(a+), 2 C variants in the acid phosphatase system, 6 LDH variants, 1 ADA3-1 and 1 AK2-1 sample. All samples were negative for the red cell antigens Cw, Vw, He, K, Jsa, Dia, Wra, Rd and Marriott, and no variation was observed in the PHI enzyme system. The results are discussed in relation to those obtained on other Papua New Guinea populations.     

Effect of hypoxia on phosphoesterases,oxidative and glycolytic enzymes in the rabbit common carotid artery

Summary An enzyme-histochemical study was performed on the rabbit common carotid artery at periods ranging from one to seventeen days following double-ligation and injections of human , human pre- serum lipoproteins and a physiologic saline into the lumen. The alterations in enzyme activities compared to the contralateral carotid (control) were studied for DPN diaphorase, succinic acid dehydrogenase (SDH), lactic acid dehydrogenase (LDH), glucose-6-phosphate dehydrogenase (G-6-pH DH), ATPase, AMPase, acid and alkaline phosphatase. At earlier time intervals there was a general reduction in oxidative enzyme and ATPase activities concomitant with a general increase in LDH and acid phosphatase activities. At later times oxidative enzyme and ATPase activities returned to their control levels and actually increased in the thickened intima. LDH and acid phosphatase activities remained above the control levels, especially in the thickened intima. Focal areas, presumably necrotic, demonstrated complete loss of activity for all enzymes studied. AMPase activity did not differ from the controls throughout this study, while G-6-pH DH and alkaline phosphatase activities were found only sparsely in the adventitia. The same general pattern of alteration in enzyme activity was found regardless of the substance injected into the ligated artery. Arguments for the use of this experimental model for studies on the pathogenesis of atherosclerosis are given.     

Influence of L-thyroxine upon enzymatic activity in the renal tubular epithelium of the rat under normal conditions and mercury-induced lesions. II. Histochemical studies of lactate dehydrogenase, succinate dehydrogenase, malate dehydrogenase, unspecific esterase, and glucose-6-phosphate dehydrogenase.

Mercury-induced renal tubular lesions in the rat present histochemically with a decrease of succinate dehydrogenase (SDH), malate dehydrogenase (MDH), glucose-6-phosphate dehydrogenase (G-6-PD), and unspecific esterase (UE), but with an increase of lactate dehydrogenase (LDH), indicating a drop of energy supply as well as a switch from oxidative to glycolytic energy production. L-thyroxine has the same effect on SDH, G-6-PD, and LDH, but an inverse effect on MDH and UE, pointing to stimulation of gluconeogenesis. However, administration of L-thyroxine to animals which have been submitted to sublimate intoxication even further decreases the MDH and UE activity while raising or partly restoring the activity of LDH, SDH, and G-6-PD. This observation is interpreted as an attempt of the damaged epithelial cell, as the gluconeogenesis ceases, to gain relatively more energy supply for the benefit of the vitally indispensable tubular Na+ reabsorption.     

Activities of Glucose-6-phosphate, 6-phosphogluconate,and Isocitrate Dehydrogenases from Leishmania donovani Cultivated at 25 and 37 C*

SYNOPSIS. The activities of glucose-6-phosphate dehydrogenase (G-6-PD) (EC No. 1.1.1.49), 6-phosphogluconate dehydrogenase (PGD) (EC No. 1.1.1.44), and isocitrate dehydrogenase (ICD) (EC No. 1.1.1.42) from promastigotes of Leishmania donovani strain 3S grown at 25 C in modified Tobie's (mT) medium and from promastigotes of the 37 C-adapted substrain of this strain cultivated in the mT at 37 C were assayed at 25 and 37 C. At 25 C ICD from both the strain and the substrain had the highest, and PGD, the lowest activity; the activity of G-6-PD was intermediate, but much closer to that of ICD. Irrespective of the temperature of the assay, the activities of G-6-PD and ICD from the 37 C substrain were significantly higher than those of these enzymes from the parental strain; however, the activity of PGD from the 25 C strain was slightly higher than that of this dehydrogenase from the 37 C-adapted stock. No significant activity losses of G-6-PD and ICD from either the strain or the substrain were noted after incubation of the extracts in the presence of 0.25 M sucrose at 37 C for 2 hr. PGD was unstable in such extracts, but it could be rendered stable by the addition of 4 mM 6-phosphogluconate. G-6-PD was the least and ICD the most dependent on Mg²⁺ ions. In the 15–25 C range, the Q₁₀ values of the enzymes from the 25 C strain were 2.83, 2.5, and 2.63 for G-6-PD, PGD, and ICD, respectively. These values for the respective enzymes in the 25–35 C range were 2.06, 1.67, and 1.62. The Q₁₀ values of the enzymes from the 37 C substrain in the 15–25 C range were 2.06 for G-6-PD, 3.25 for PGD, and 2.77 for ICD; in the 25–35 C range, the corresponding values were 1.67, 1.46, and 1.83. Cultivation of the 37 C substrain at 25 C was accompanied by a drop in G-6-PD and ICD activities.     

Comparative changes in the lipogenic enzyme activities and in the in vivo fatty acid synthesis in liver and adipose tissues during the post-weaning growth of male rats

Changes in the specific activities of acetyl-CoA-carboxylase (ACX), malic enzyme (ME) and glucose-6-phosphate dehydrogenase (G-6-PD) were compared to changes in de novo lipogenesis measured by in vivo incorporation of [3H] of tritiated water into fatty acids of liver and of perirenal and dorsal subcutaneous adipose tissues. In the adipose tissues, the specific activities of the three enzymes rather closely followed fluctuations in the rate of fatty acid synthesis. In the liver, ACX and especially ME activities were satisfactory indicators of de novo lipogenesis; G-6-PD activity did not depend on de novo lipogenesis.