摘要:

AbstractPyruvate transport and carboxylation have been determined in mitochondria from liver and kidney cortex isolated from Wistar rats with acidosis produced by three different treatments: fasting, exercise and ingestion of ammonium chloride. Fasting for 48 h or swimming for 2 h resulted in an increased rate of CO2fixation by mitochondria from both organs incubated with pyruvate. This increase was accompanied by a rise in the rate of pyruvate transport in all cases except in mitochondria derived from the kidney of the fasted animals. Acute acidosis produced by the ingestion of ammonium chloride resulted in increases in pyruvate transport and carboxylation in kidney mitochondria, but a drop in pyruvate carboxylation was observed in mitochondria from the liver. The results are discussed in terms of the differential regulation of the mitochondria steps for gluconeogenesis from three carbon precursors in liver and kidney, taking into consideration the hormonal status of the animals and the prevailing available substrates in each condition.

ISSN:0263-6484    

DOI:10.1002/cbf.290120402

出版商:John Wiley&Sons, Ltd.    

年代:1994

数据来源: WILEY

摘要:

AbstractThe synthesis of uric acid from purine bases, nucleosides and nucleotides has been measured in reaction mixtures containing rat liver supernatant and each one of the following compounds at 1 mMconcentration (except xanthine, 0·5 mMand guanosine and guanine, 0·1 mM). The rates of the reaction, expressed as nanomoles of uric acid synthesized g−1of wet liver min−1were: ATP, 10; ADP, 37; AMP, 62; adenosine, 108; adenine 6; adenylo‐succinate, 9; IMP 32; inosine, 112; hypoxanthine, 50; GTP, 19; GDP, 19; GMP, 27; guanosine, 34; guanine, 72; XMP, 10; xanthosine, 24; xanthine, 144. These figures divided by 55 correspond to nanomoles of uric acid synthesized min−1per mg−1of protein. The rate of synthesis of uric acid obtained with each one of those compounds at 0·1 and 0·05 mMconcentrations was also determined. ATP (1 nM) strongly inhibited uric acid synthesis from 0·05 mMAMP (91 per cent) and from 0·05 mMADP (88 per cent), but not from adenosine. CTP or UTP (1 mM) also inhibited (by more than 90 per cent) the synthesis of uric acid from 0·05 mMAMP. Xanthine oxidase was inhibited by concentrations of hypoxanthine higher than 0·012 mM. The results favour the view that the level of uric acid in plasma may be an index of the energetic state of the organism. Allopurinol, besides inhibiting uric acid synthesis, reduced the rate of degradation of AMP. The ability of crude extracts to catabolize purine nucleotides to uric acid is an important factor to be considered when some enzymes related to purine nucleotide metabolism, particularly CTP synthase, are measured in cru

ISSN:0263-6484    

DOI:10.1002/cbf.290120403

出版商:John Wiley&Sons, Ltd.    

年代:1994

数据来源: WILEY

摘要:

AbstractThe activities of DNA topoisomerase I and II and the relative proportions of the histone H1 subtypes were investigated in rat liver which was undergoing hypertrophy and exhibiting increased transcriptional activity following the administration of dexamethasone. There was a rise in the level of activity of DNA topoisomerase I and a slight fall in that of DNA topoisomerase II. The relative proportions of the H1 subtypes were altered due to a preferential increase in H1.1. The results are discussed in relation to the effect of glucocorticoids on the transcription and replication of hepatic DNA.

ISSN:0263-6484    

DOI:10.1002/cbf.290120404

出版商:John Wiley&Sons, Ltd.    

年代:1994

数据来源: WILEY

摘要:

AbstractThe impact of season and temperature on frog liver γ‐glutamyltranspeptidase was assessed by measuring the activity of this enzyme in plasma membranes isolated from the livers ofRana pipiensobtained as summer and winter frogs; subjected to short‐term (3 weeks) temperature acclimation; and subjected to multiple‐temperature shifts. Plasma levels of T3were determined. γ‐Glutamyltranspeptidase was found to be 2·2‐fold higher in the summer frog relative to the winter frog; decreased by 44 percent in the summer frog by cold acclimation and increased by 1·7‐fold in the winter frog by warm acclimation; and increased by 1·9‐fold in the summer frog and 2·8‐fold in the winter frog subjected to multiple‐temperature shifts. Plasma T3levels were found to be 42‐fold higher in the summer frog relative to the winter frog; decreased by 42 percent by cold acclimation and increased by 2·9‐fold by warm acclimation; and decreased by 39 percent and 38 percent in the summer and winter frogs subjected to multiple temperature shifts. T3replacement during the last phase of the multiple‐temperature shift protocol, restored the plasma T3levels to 75 percent of the control levels and prevented the increase evoked by the multiple‐temperature shifts in γ‐glutamyl‐transpeptidase activity. Indeed, enzyme activity in the T3replaced state was 19 percent lower than in the control state. The involvement of thyroid hormone as a negative regu

ISSN:0263-6484    

DOI:10.1002/cbf.290120405

出版商:John Wiley&Sons, Ltd.    

年代:1994

数据来源: WILEY

摘要:

AbstractThe L/B/K type of mammalian alkaline phosphatase (ALP) is inhibited uncompetitively by nucleotides. A combination of adenosine and nicotinamide is more effective than either adenosine or nicotinamide alone, probably because a dinucleotide structure is necessary to trigger a conformational change accompanying binding of structures such as NADH. It has been suggested that a loop region containing residue 429 in the ALP polypeptide is important in the interaction of uncompetitive inhibitors with the enzyme. In the L/B/K isoenzyme, residue 429 is a histidine and is a potential target for modification. In an attempt to learn more about the molecular events accompanying inhibition of ALP by uncompetitive inhibitors, bovine kidney ALP was reacted with oxidized adenosine in the presence of nicotinamide to see if site‐directed modification occurs. Kidney ALP was irreversibly inactivated by oxidized adenosine but the reaction was slow. The site modified is likely to be close to the region of binding. Sequence data for the kidney enzyme shows that in the region of residue 429 there are no residues except His429itself that is likely to react with oxidized adenosin

ISSN:0263-6484    

DOI:10.1002/cbf.290120406

出版商:John Wiley&Sons, Ltd.    

年代:1994

数据来源: WILEY

摘要:

AbstractThe state of polymerization of actin and the organization of actin filaments is widely believed to be related to cellular transformation. Since the intracellular monomer (G) and filamentous (F) actin content reflects the state of microfilament polymerization, we measured the G/total actin ratio in primary cultures of normal and malignant human keratinocytes. In normal keratinocytes the mean value of this ratio was 0·30 ± 0·03 (mean ± SE,n= 15), while in basal cell carcinoma (BCC) keratinocytes it was 0·49 ± 0·03 (n= 8) and in squamous cell carcinoma keratinocytes (SCC) 0·5 ± 0·07 (n= 4), indicating a 1·7‐fold increase of the G/total actin ratio in malignant cells. These results imply that the proportion of polymerized actin is decreased markedly in malignant keratinocytes, suggesting alterations of microfilament structures which probably occur during the transformation process. This was supported by the morphological changes of microfilament structures as assessed by fluorescence microscopy. A different distribution of actin filaments in normal and malignant cells became evident; stress‐fibres were converging in patches at several points in SCC cells, when compared to normal keratinocytes. Furthermore, incubation of normal and malignant keratinocytes with cytochalasin B indicated differences in the resistance of their microfilament networks. After 1 h exposure to 10−6and 10−5Mcytochalasin B, microfilaments in normal cells appeared to be less affected than their counterparts in neoplastic cells. Even in a high excess of cytochalasin B (10−4M), normal keratinocytes preserved their shape, while both basal cell and SCC were totally disrupted. We concluded that the G/total actin ratio was significantly increased in malignant keratinocytes. This seems to be correlated with altered microfilament morphology and resistance to cytochalasin B treatment. Our results suggest that the process of malignant transformation may be characterized by changes in the state of the polymerization of actin and in the stability of the microfilament network indicating that both features could potentially serve as markers determine the transformed st

ISSN:0263-6484    

DOI:10.1002/cbf.290120407

出版商:John Wiley&Sons, Ltd.    

年代:1994

数据来源: WILEY

摘要:

AbstractA comparison has been made between the effects of 4‐hydroxy‐2,3‐trans‐nonenal (HNE) and 4‐hydroxy‐2,3‐trans‐octenal (HOE), two lipid peroxidation products, on the basal and GTPgammaS‐stimulated activities of phosphoinositide‐specific phospholipase C (PL‐C) of rat polymorphonuclear leukocytes. PL‐C activity was determinedin vitroby measuring the hydrolysis of [3H] phosphatidylinositol‐4,5‐bis‐phosphate (PtdIns‐P2) added as exogenous substrate to neutrophil plasma membranes. PL‐C was activated by concentrations of HNE ranging from 10−8to 10−6Mboth in the presence and in the absence of 2 × 10−5MGTPgammaS; HOE stimulated the enzymatic activity between 10−11and 10−8M; maximal stimulation was given by 10−11MHOE plus GTPgammaS. The aldehyde concentrations able to accelerate PtdIns‐P2breakdown displayed a good correspondence with those which have been reported to stimulate the oriented migration of rat neutrophils. Pretreatment of neutrophils with pertussis toxin prevented the stimulation of PL‐C by 10−11MHOE and by HOE plus GTPgammaS. Our results suggest that the chemotactic action of HNE and HOE might depend on the activation of PL‐C; furthermore a regulatory G protein appears to be in

ISSN:0263-6484    

DOI:10.1002/cbf.290120408

出版商:John Wiley&Sons, Ltd.    

年代:1994

数据来源: WILEY

摘要:

AbstractHyaluronan (HA) protected tendon fibroblasts against cell damage mediated by hydroxyl radicals (OH·) as demonstrated by release of51Cr from labelled cells. Protection afforded by high molecular mass (Mr) HA (1218 kDa) was much more effective than that provided by lower (176 kDa and 668 kDa)MrHA.OH· was generated by coupling H2O2produced by glucose oxidase: glucose to [Fe2+‐EDTA] chelate in a Fenton‐type system. The flux of OH· was measured by a spectrofluorimetric assay of salicylate produced by the reaction of benzoate with OH·. Cell damage caused by the OH· generating system was prevented in the presence of catalase, which destroyed H2O2. Damage caused in a standard incubation time increased with increased amounts of glucose oxidase.Protection against OH·‐mediated cell damage increased with increasing concentration of HA. The presence of HA did not interfere with the enzyme‐Fenton system, as monitored by production of gluconate. On the other hand, HA scavenged OH· produced by the enzyme‐Fenton system, as shown by competition with benzoate, which produced less salicylate in the spectrofluorimetric assay in the presence of HA.The reaction of OH· with HA was measured directly by a pulse radiolysis technique in which a hydrated electron (e aq−) produced OH· by the reaction with nitrous oxide. Second order rate constants obtained in distilled H2O or in phosphate buffer showed no dependence on HAMr. Similarly, fluorimetric assay of the flux of in the enzyme‐Fenton system confirmed that HA competed with benzoate, thus lowering salicylate production, and the flux was also independent of the molecular mass of HA.These results demonstrate that part of the HA‐mediated protection against enzyme‐Fenton produced OH· and other reactive oxygen‐derived toxic species was not a consequence of either the primary or secondary structure of HA, but rather depends on higher order HA organization. Some aspects of the formation of the HA meshwork (tertiary structure) areMrdependent. We therefore propose that cell‐anchored HA meshworks excluded relatively large enzyme molecules from the immediate environment of the cell, thus reducing the flux of OH· etc. at the cell s

ISSN:0263-6484    

DOI:10.1002/cbf.290120409

出版商:John Wiley&Sons, Ltd.    

年代:1994

数据来源: WILEY

摘要:

AbstractThe present report describes experiments to evaluate phospholipid methyltransferase activity in golden hamster spermatozoa incubated under different conditions. Washed cauda epididymal sperm were incubated with taurine, in the presence or absence of epinephrine. At various times, the sperm were separated, and phospholipid methyltransferase activity measured. Also, at each time, aliquots of the sperm suspension were assayed for motility, and acrosome reactions. Some sperm incubated in the presence of taurine and epinephrine were capacitated by 3·5 h, because about 40 per cent of them can undergo the acrosome reaction 10 min after addition of the fusogen lysophosphatidylcholine. In epinephrine‐free incubations the fusogen failed to stimulated acrosome reactions. On the other hand, epinephrine stimulated by twofold phospholipid methyltransferase activity from ‘0 time’ incubated sperm, in comparison to that observed in taurine‐treated cells. Enzyme activities from both taurine or epinephrine plus taurine‐treated cells decreased as the incubation time of the sperm suspension increased. Kinetic properties of the sperm phospholipid methyltransferase acvity were modified by the presence of taurine and epinephrine when S‐adenosylmethionine was used as the substrate. These results suggest that refined molecular events occur in the sperm cell during the acquisition of fertili

ISSN:0263-6484    

DOI:10.1002/cbf.290120410

出版商:John Wiley&Sons, Ltd.    

年代:1994

数据来源: WILEY

ISSN:0263-6484    

DOI:10.1002/cbf.290120412

出版商:John Wiley&Sons, Ltd.    

年代:1994

数据来源: WILEY