摘要:

AbstractSuccinate dehydrogenase activity was measured in rat pancreatic islet homogenates incubated in the presence of [1,4‐14C]succinate, the reaction velocity being judged through the generation of14CO2in the auxiliary reactions catalysed by pig heart fumarase and chicken liver NADP‐malate dehydrogenase. In the presence of 1·0 mM succinate, the reaction velocity averaged 5·53 ± 0·44 pmol min−1μg−1islet protein. TheKmfor succinate was close to 0·4 mM and the enzymic activity was restricted to mitochondria. These kinetic results indicate that, under the present experimental conditions, the activity of succinate dehydrogenase does not vastly exceed that of either NAD‐isocitrate dehydrogenase or the 2‐ketoglutarate dehydrogenase complex, at least when the latter enzymes are activated by ADP and/or Ca2+. Nevertheless, the activity of succinate dehydrogenase is sufficient to account for the increase in O2uptake evoked in intact islets by the monomethyl ester of succinic acid. It could become a rate‐limiting step of the Krebs cycle in models of

ISSN:0263-6484    

DOI:10.1002/cbf.290110302

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractThe effect of propranolol on the process of thyroid hormone formation was studied in a physiological culture system. Porcine thyroid follicles were preincubated with propranolol for 24 h. Iodide transport, iodine organification, andde novothyroid hormone formation were measured by incubating these follicles with the mixture of carrier‐free 0·1 μCi Na125I and 50 nMNaI for 2 to 6 h at 37°C. A concentration of propranolol greater than 100 μMinhibited iodide transport in a dose‐dependent manner; this inhibition was non‐competitive with iodide and independent of thyrotropin (TSH). Reduced iodine organification and thyroid hormone formation was seen with 150 μMpropranolol or greater. The inhibitory action of propranolol was not caused by beta‐blocking activity, sinceD‐propranolol (devoid of beta‐blocking activity) inhibited iodide transport, and other beta‐blockers (metoprolol, atenolol, and labetalol) did not inhibit iodide transport. The inhibition of iodide transport was most likely caused by membrane stabilizing activity since quinidine, which possess the same membrane stabilizing activity as propranolol, also inhibited iodide transport. TSH‐mediated cAMP generation and Na+K+ATPase activity, membrane functions for iodide transport, were unaffected by propranolol.Our study has shown, for the first time, that propranolol has a direct antithyroid action, namely inhibition of iodide transport in the int

ISSN:0263-6484    

DOI:10.1002/cbf.290110303

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractBeta‐glucosidase activity was evaluatedin situby means of fluorogenic probes in normal human fibroblasts and fibroblasts from homozygous carriers of the Gaucher trait. Probe internalization, targeting to lysosomes and post‐cleavage probe retention were the primary concerns. Internalization and targeting were attempted byin situphotosensitized labilization of lysosomal membranes, lysosomotropic detergents and the use of low density lipid (LDL) or the receptor ligand apolipoprotein E (ApoE). Post‐cleavage increase of fluorescence with fluoresceinyl (bis) betaglucopyranoside was appreciably above the rather large pre‐cleavage emission. In cells incubated overnight with nonylumbelliferylbetaglucoside (UG9) in the presence of bovine serum albumin and in the absence of ApoE, the probe was dealt with as a cytotoxic agent, accumulating in a paranuclear cap, most likely comprising elements of the endoplasmic reticulum (ER) and Golgi apparatus. Targeting of UG9 to lysosomes occurred within 1 to 3 h of preincubation in the presence of ApoE. There was some evidence of specificity, as Gaucher fibroblasts exhibited weaker cleavage of UG9 (by 50 per cent or more) compared to normal fibroblasts, but in the Gaucher cells there was some residual beta‐glucosidase activity. Cleavage of UG9 was nearly totally suppressed in Gaucher cells treated with the beta‐glucosidase inhibitor, conduritol B epoxide, for 24 h to 7 days. Suppression in the control fibroblasts was evident but to a lesser degree. Thein situmethod of fluorogenic assay established for beta‐glucosidase deficiency, is in principle applicable to enzyme deficiencies in other lysosomal storage diseases, or to evaluate enhanced enzyme activity following

ISSN:0263-6484    

DOI:10.1002/cbf.290110304

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractAn enriched preparation of rat hepatocyte couplets was obtaied by collagense perfusion and subsequent elutriation (>85 per cent couplets and triplets; viability of over 95 per cent).Canalicular secretory activity (the ability to accumulate cholyl‐lysyl‐fluorescein, CLF) was first apparent after 2 h of culture at 37°C and was present in over 80 per cent of the total population after 5–6 h. This remained almost constant for at least 4 h in both elutriated and directly plated cells. Initial storage of freshly prepared couplets at 4°C for up to 6 h prior to incubation had no adverse effect upon secretory function.Reduction of canalicular secretory activity occurred at a concentration of the hepatotoxic agent menadione (IC5017 μM) that was lower than that required to induce mild plasma‐membrane blebbing (IC5043 μM).This study has optimized and characterized the canalicular secretory effectiveness and stability of an enriched preparation of hepatocyte couplets, and established the feasibility of studies of toxic agents on hepatobiliary function in a heterogeneous population of hepatocytes. In this preparation other biochemical parameters can be assessed, thus complementing previous techniques using individu

ISSN:0263-6484    

DOI:10.1002/cbf.290110305

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractThe genomic 3′ structure of the gene coding for the human slow skeletal troponin T (TNNT1) gene, is reported. An intron of 912 nucleotides containing an Alu‐element has been identified and characterized. The complexity of the sequenced region suggests an alternative exon use. The present results may be valuable for further studies on the gene structure of TNNT1 and the related troponin gene fam

ISSN:0263-6484    

DOI:10.1002/cbf.290110306

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractArtemiauses the stored diguanosine tetraphosphate as a source of adenine and guanine nucleotides during development from the encysted gastrula to the free swimming larva. Further development of the larvae depends on a dietary source of purine rings. We have investigated the growth ofArtemiain axenic cultures supplemented with 0·6 mg ml−1of adenosine, guanosine, inosine or xanthosine. The total protein and soluble nucleotide content ofArtemiagrown in the presence of adenosine, guanosine or inosine was very similar, around (2 A260units and 500 mg protein) and (4 A260units and 1000 mg protein) after 4 and 6 days of postlarval development, respectively. The nucleotide pattern of those extracts subjected to HPLC were almost identical, the major peaks corresponding to ATP, ADP and AMP. Other nucleotides, not well characterized, were also present in those extracts. Mycophenolic acid (10 μg ml−1) inhibited the growth ofArtemia(as measured by their protein and soluble nucleotide content) in the presence of adenosine and inosine as the purine source, and had no appreciable effect in the presence of guanosine. A quantitative analysis of the chromatographic peaks obtained fromArtemiagrown in the presence of any of the three nucleosides ± mycophenolic acid showed that the effect of the antibiotic on each one of the chromatographic peaks was very similar, suggesting thatArtemia, and probably other organisms as well, tend to maintain a balance between all nucleotides and to adjust the overall level to the limiting step(s) in their rates of synthesis/interconversion. Xanthosine was not able to support the development ofA

ISSN:0263-6484    

DOI:10.1002/cbf.290110307

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractThe biosynthesis of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) in monocyte‐like leukemia U937 cells was monitored by adding [3H]choline, [14C]ethanolamine or [14C]glycerol to the culture media; incorporation into phospholipid (PL) increased with time. The effect of unsaturated fatty acids (UFA) on PC and PE synthesis was investigated by pretreating U937 cells for 72h with 10 μM18:1 (n–9), 18:2 (n–6), 18:3 (n–3), 20:4 (n–6) and 20:5 (n–3). The UFA caused no alteration in cell growth, as evidenced by light microscopy and the incorporation of [3H]thymidine and [3H]leucine. Total cellular uptake of radioactive precursors remained unaffected by all the treatments. Pretreatment with 20:5 resulted in approximately 25 per cent reduction in the incorporation of [3H]choline into PL, while no significant effect was detected with the other UFAs. 18:3, 20:4 and 20:5 depressed the incorporation of [14C]ethanolamine into PL by 34 per cent, 28 per cent and 49 per cent respectively. However, there was no redistribution of label with any of the treatments. 18:3, 20:4 and 20:5 also antagonized the stimulatory effect of endotoxin (LPS) on PC and PE synthesis. In addition, the incorporation from [14C]glycerol into PC and PE was reduced by 18:3, 20:4 and 20:5.Although the PL composition of the cells remained essentially unaffected, our study shows that chronic treatment of U937 cells withn–3 PUFA (20:5) depressed PC and PE synthesis, and 18:3 and 20:4 also caused inhibition o

ISSN:0263-6484    

DOI:10.1002/cbf.290110308

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractWe have tested the effect of alkaloids (cocaine, morphine) and enkephalins on neutral endopeptidase of peripheral blood mononuclear cells activated by lectins. When treated with concanavalin A and cocaine, peripheral blood mononuclear cells showed an enhanced activity (+110 per cent) of the membrane neutral endopeptidase, which was not related to the expression of the common acute lymphoblastic leukemia antigen at the cell surface, although both molecules have the identical amino acid sequence. Phytohemagglutinin‐P, morphine and synthetic enkephalins did not induce the activity of neutral endopeptidase nor the expression of common acute lymphoblastic leukemia antigen. Our findings suggested that the drugs of abuse, cocaine and morphine, affected specific membrane constituents without altering proliferation, subcellular localization of membrane enzymes or the surface immune phenotype of peripheral blood mononuclear cell

ISSN:0263-6484    

DOI:10.1002/cbf.290110309

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractA new continuous column culture system for adherent cells was developed using beads. The beads were packed in a column and an appropriate medium was continuously passed through. The whole system was kept under closed conditions.L cells and C6 cells were cultured by this new system. The number of cells increased linearly up to 16 days and reached a maximum at around 18 days. As the heat production remained constant for 16 days, it can be concluded that cells grown in this system had identical characteristics. The final concentration of cells reached was 1.0 × 108ml−1. The cells could grow both in the upward and the downward direction.Advantages of this system are: (1) Cells can be recovered in their adherent form on the beads; (2) cells can easily be collected from the column by trypsinization, and (3) cells remaining in the column after trypsinization can grow aga

ISSN:0263-6484    

DOI:10.1002/cbf.290110310

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractEndothelial and smooth muscle cells were isolated from porcine aorta and kept in short‐term culture. To determine the terminal carbohydate composition of the plasma membranes from both cell populations, the cells were incubated with a panel of fluorescein‐labelled lectins. Both cell populations shared a number of terminal carbohydrates, but theN‐galactosamine specific lectinWistaria floribundaagglutinin labelled only the endothelial cells. A lectin which selectively labelled smooth muscle cells was not found. Western blot analysis of isolated endothelial cell membrane glycoproteins indicated that most membrane glycoproteins are labelled byWistaria floribundaagglu

ISSN:0263-6484    

DOI:10.1002/cbf.290110311

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY