Pernicious anaemia and hypoparathyroidism in a patient with Kearns-Sayre syndrome with mitochondrial DNA duplication.
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Algeria↗
Biomedical subjects
L H Cohen
Publications and source records attributed to L H Cohen.
Assessment and prediction of stress-related growth.
This article reports the development of the Stress-Related Growth Scale (SRGS) and its use in a study examining determinants of stress-related positive outcomes for college students. Study 1 analyses showed that the SRGS has acceptable internal and test-retest reliability and that scores are not influenced by social desirability. Study 2 analyses showed that college students' SRGS responses were significantly related to those provided by friends and relatives on their behalf. Study 3 analyses tested the determinants of stress-related growth longitudinally. Significant predictors of the SRGS were (a) intrinsic religiousness; (b) social support satisfaction; (c) stressfulness of the negative event; (d) positive reinterpretation and acceptance coping; and (e) number of recent positive life events. The SRGS was also positively related to residual change in optimism, positive affectivity, number of socially supportive others, and social support satisfaction, lending further support to the validity of this new scale. Results have implications for current theory on stress-related positive outcomes.
Adaptation, Psychological↗
Different analogues of farnesyl pyrophosphate inhibit squalene synthase and protein:farnesyltransferase to different extents.
The inhibitory potency of farnesyl pyrophosphate analogues was investigated on two farnesyl pyrophosphate-consuming enzymes: squalene synthase, a secondary regulation site in the cholesterol synthesis pathway, and protein:farnesyl transferase, which plays a role in the function of Ras-proteins. For the transferase determination a rapid in vitro assay, using Sepharose-bound Ras-peptides, was developed. The distinct farnesyl pyrophosphate analogues showed a different order of potency in the inhibition of these two enzymes. Using the farnesyl transferase assay with pre-p21Ha-ras as substrate the same result was obtained. The difference observed in the in vitro assays was also reflected in the inhibition of cholesterol synthesis, protein prenylation in general and Ha-ras farnesylation in Rat-1.H-ras13 cells, a rat fibroblast cell line that overproduces human p21Ha-ras. This work shows that farnesyl pyrophosphate analogues can be developed for specific inhibition of different processes such as cholesterol synthesis and protein prenylation.
Alkyl and Aryl Transferases↗
Different effects of 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors on sterol synthesis in various human cell types.
The three vastatins examined, lovastatin, simvastatin and pravastatin, are equally strong inhibitors of the sterol synthesis in human hepatocytes in culture with IC50-values of 4.1, 8.0 and 2.0 nM, respectively. However, in the human extrahepatic cells: umbilical vascular endothelial cells, retinal pigment epithelial cells, cornea fibroblasts and granulosa cells, pravastatin was much less inhibiting the sterol synthesis than lovastatin or simvastatin. It was observed as well that longer incubation with the vastatins resulted in higher IC50-values. In order to show that the feedback regulation mechanism for 3-hydroxy-3-methylglutaryl-coenzyme A reductase was involved in this phenomena mRNA levels were measured in human vascular endothelial cells after incubation with the vastatins for 3.5 h and for 20 h. Indeed, lovastatin and simvastatin gave rise to higher levels of HMG-CoA reductase mRNA after 20 h than after 3.5 h of incubation. The differences observed in different human cell types can be explained by supposing that pravastatin is transported into the human hepatocyte via a liver-specific transporter. This was supported by the results of uptake experiments with 14C-labelled pravastatin and 14C-labelled simvastatin into human hepatocytes compared to that into human umbilical endothelial cells (as an example of an extrahepatic cell type). [14C]-Simvastatin was associated with both cell types, whereas [14C]-pravastatin was hardly associated with human endothelial cells, but to a similar extent as [14C]-simvastatin with human hepatocytes.
Blotting, Northern↗
Components of a system that ligates cyclin to ubiquitin and their regulation by the protein kinase cdc2.
Cyclin B, a positive regulatory subunit of the cdc2 protein kinase complex, is synthesized across the cell cycle and then rapidly degraded at the end of mitosis. Degradation of cyclin B is triggered by increased levels of active cdc2 and is required for exit from mitosis. It was shown previously that cyclin degradation is carried out by the ubiquitin system, but the components responsible for the specificity and regulation of cyclin-ubiquitin ligation have not been identified. The formation of ubiquitin-protein conjugates usually requires the sequential action of three enzymes: a ubiquitin-activating enzyme (E1), a ubiquitin-carrier protein (E2), and a ubiquitin-protein ligase (E3). In this work we employed a fractionation approach to identify the components of a clam oocyte system responsible for specific ubiquitination of cyclin and to determine which components are regulated by cdc2. Experimental conditions were established under which a fusion protein containing an amino-terminal fragment of cyclin B is ligated to ubiquitin only in extracts from M-phase but not from interphase cells. Fractionation of M-phase extracts by DEAE-cellulose and high speed centrifugation yielded three fractions that were all required for cell cycle stage-specific cyclin-ubiquitin ligation. Only one of these fractions could be replaced by a previously known enzyme of the ubiquitin system, E1. A second fraction contained a novel species of E2, termed E2-C, which acts in the ligation of ubiquitin to cyclin but not to other endogenous proteins. A third component is associated with particulate material. Whereas E2-C from either M-phase or interphase extracts is active, the particulate component is active only in M-phase. Incubation of the particulate fraction from interphase cells with the protein kinase cdc2 activates it for cyclin-ubiquitin ligation, after a lag of about 30 min. These findings suggest that the particulate fraction may contain an E3 enzyme that acts on cyclin, as well as additional factors activated by cdc2.
Animals↗
Pravastatin inhibited the cholesterol synthesis in human hepatoma cell line Hep G2 less than simvastatin and lovastatin, which is reflected in the upregulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase and squalene synthase.
The possible difference between lovastatin (mevinolin, MK-803), simvastatin (MK-733) and pravastatin (CS-514), all chemically-related competitive inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, were tested in the human hepatoma cell line Hep G2, which is often used as a model for the human hepatocyte. After an 18-hr incubation of the cells with the drugs, pravastatin (IC50 = 1900 nM) was less potent than simvastatin and lovastatin (IC50 = 34 and 24 nM, respectively) in inhibiting the sterol synthesis. As a consequence of this inhibition, the HMG-CoA reductase mRNA levels and squalene synthase activity, both negatively-regulated by sterols, were increased equally by simvastatin and lovastatin, whereas the induction by pravastatin was much less. In contrast, there were fewer differences between the compounds in inhibiting HMG-CoA reductase activity, when assayed directly in Hep G2 cell homogenates (IC50 values = 18, 61 and 95 nM for simvastatin, lovastatin and pravastatin, respectively). Moreover, in experiments with human hepatocytes in primary culture the IC50 values for inhibition of the cholesterol synthesis by simvastatin and pravastatin were of the same order of magnitude (23 and 105 nM, respectively). The results are therefore explained as follows: the three drugs act in the same way within the Hep G2 cell in terms of inhibiting HMG-CoA reductase and their subsequent effect on the feedback regulation of the cholesterol synthesis, i.e. increasing squalene synthase and HMG-CoA reductase mRNA. However, pravastatin seems to be less able to enter the cells compared with simvastatin and lovastatin, possibly because of the higher hydrophobicity of the latter compounds. The observation with human hepatocytes suggests that in Hep G2 cells a specific hepatic transporter is missing. On one hand the human hepatoma cell line Hep G2 has proved to be a good model for the study of the feedback regulation of enzymes of the cholesterol biosynthetic pathway such as HMG-CoA reductase and squalene synthase, but, on the other hand seems to be less suitable as a model for the study of specific uptake of drugs, e.g. the vastatins, in human hepatocytes.
Cholesterol↗
Different effects of the hypolipidemic drugs pravastatin and lovastatin on the cholesterol biosynthesis of the human ocular lens in organ culture and on the cholesterol content of the rat lens in vivo.
This study aimed to investigate the influence of the hypocholesterolemic drugs pravastatin and lovastatin on the cholesterol biosynthesis in the ocular lens. Two model systems were used: a human lens organ culture system and an in vivo rat lens system. For measurements of cholesterol and fatty acid synthesis rates, human lenses were incubated for 20 h in the presence of [14C]acetate. Pravastatin and lovastatin were added 1 h prior to the addition of the radioactive label. In order to avoid the influence of differences relating to individual donors, one lens from each donor was incubated without drug (control) and the other lens was incubated in the presence of the drug. Statistical analysis showed that the fatty acid synthesis rate was not influenced by the drug. For each lens pair the percentage inhibition of the cholesterol synthesis caused by the drug was calculated. Using various concentrations of the drugs, a dose-response curve was composed for the inhibition of the cholesterol synthesis. The experiments showed that in the human lens organ culture system, lovastatin was 100-fold more potent than pravastatin in inhibiting the cholesterol biosynthesis. To study the in vivo influence of vastatins on the cholesterol content of the developing lens, Wistar rats were weaned at day 21 of age and subsequently the pups were fed a control diet or drug-containing diet (10, 50 or 100 mg lovastatin/kg chow) for a 3-week period. At the end of diet intervention, doses of 50 or 100 mg lovastatin/kg chow had caused a reduction of about 20% of the lenticular cholesterol content compared with controls. No effect on the lens cholesterol content by pravastatin was observed. Both human ex vivo and rat in vivo experiments show that lovastatin much more strongly inhibits the lenticular cholesterol synthesis than does pravastatin.
Adult↗
Cholesterol content of the rat lens is lowered by administration of simvastatin, but not by pravastatin.
The influence of the 3-hydroxy-3-methyl-glutaryl coenzyme A (HMG-CoA) reductase inhibitors pravastatin and simvastatin on lens cholesterol metabolism was investigated in the rat. Short-term organ culture experiments with explanted lenses from 21-day-old Wistar rats showed that simvastatin was at least 35 times more effective than pravastatin in inhibiting cholesterol synthesis. In vivo the cholesterol content of the rat lens increased linearly with age. Experiments were designed to answer the question whether simvastatin and pravastatin inhibit lens cholesterol synthesis in vivo, which would result in a reduced cholesterol accumulation in the lens with age. Young Wistar rats were weaned at an age of 21 days and had ad libitum access to a chow supplemented with 10-100 mg vastatin kg-1 (drug consumption: 1.5-15 mg vastatin kg-1 body weight day-1, respectively) or no additions for 3 weeks. Both drugs induced the HMG-CoA reductase activity in rat liver microsomes (isolated after 1, 2 and 3 weeks of treatment) to a similar extent. This indicates that the two drugs inhibited hepatic cholesterol synthesis to a comparable extent. During the whole treatment period no significant differences between control and drug-treated animals could be observed when the wet weight and protein content of the lenses were considered. However, a striking difference between the control group and pravastatin group (50 mg drug kg-1 diet) on the one hand and the simvastatin group (50 mg drug kg-1 diet) on the other was observed when the cholesterol content of the lenses were compared as a function of age.(ABSTRACT TRUNCATED AT 250 WORDS)
Aging↗
Extrahepatic lipogenesis contributes to hyperlipidemia in the analbuminemic rat.
Hepatic lipid and apolipoprotein synthesis is increased in the nephrotic syndrome. Catabolism of triglyceride-rich lipoproteins is impaired in nephrotic syndrome but not in rats with hereditary analbuminemia (NA), suggesting that lipid synthesis should be increased by analbuminemia in the absence of proteinuria. In this study the rate of cholesterol and fatty acid synthesis in liver and extrahepatic tissue was measured in female NA and control Sprague-Dawley (SD) rats to determine whether lipid synthesis was indeed increased in isolated analbuminemia and to identify the site(s) of increased lipogenesis. We also measured the concentrations of apolipoproteins (apo) AI, B, and E in plasma, as well as the levels of the respective mRNAs in liver. Plasma cholesterol, triglycerides, and apo AI, B, and E were all increased severalfold in the NA rat (P < 0.001). Although liver apolipoprotein mRNA content was significantly increased (P < 0.001) for apo AI (643%), B (273%), and E (299%), 3-hydroxy-3-methylglutaryl-coenzyme A reductase activity in liver microsomes and hepatic cholesterol synthesis were not significantly increased in the NA rats. Hepatic fatty acid synthesis and intestinal cholesterol synthesis were not increased in the NA rats. Surprisingly, intestinal fatty acid synthesis was elevated by 60% (P < 0.01). The NA rats demonstrated approximately fourfold increases in the incorporation of 3H2O into circulating cholesterol and fatty acids (P < 0.001). A 56% increase in the synthesis of total nonsaponifiable lipid was found in the extravisceral carcass (P < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Pravastatin and simvastatin differently inhibit cholesterol biosynthesis in human lens.
PURPOSE: In the current study, the hypocholesterolemic drugs pravastatin and simvastatin were compared for their influence on cholesterol biosynthesis in the human lens. METHODS: For measurements of cholesterol and fatty acid synthesis rates, human lenses were incubated for 20 hr in the presence of [14C]-acetate, and pravastatin or simvastatin. Radiolabeled [14C]-cholesterol and [14C]-fatty acids were determined. To avoid the influence of individual differences, one lens from each donor was incubated without drug (control) and the other lens was incubated in the presence of drug. For each lens pair, the percentage inhibition of the cholesterol synthesis caused by the drug was calculated. Fatty acid synthesis was not influenced by the drugs. By comparing the fatty acid synthesis rate of the drug-incubated with the control lens of a pair, a predefined exclusion criterion was used to eliminate lens pairs in which the lenses had no comparable biosynthetic capacities. RESULTS: Using various concentrations of the drugs, a dose-response curve was constructed for the inhibition of the cholesterol synthesis. The IC50 values (drug concentration give 50% inhibition) were 0.5 mumol/l and 0.004 mumol/l for pravastatin and simvastatin, respectively. 3-Hydroxy-3-methylglutaryl coenzyme A reductase activity in microsomal membranes from human lens cortex was inhibited by simvastatin and pravastatin to the same extent. CONCLUSIONS: Under the conditions used in this study, cholesterol synthesis in human lenses is inhibited by simvastatin 100-fold more effectively than by pravastatin. This difference was likely due to differences in the intracellular exposure of the reductase to the drugs in intact human lenses.
Adolescent↗
Subcellular localization of squalene synthase in human hepatoma cell line Hep G2.
Using the Hep G2 cell line as a model for the human hepatocyte the question was studied whether Hep G2-peroxisomes could be able to synthesize cholesterol. Hep G2 cell homogenates were applied to density gradient centrifugation on Nycodenz, resulting in good separation between the organelles. The different organelle fractions were characterized by assaying the following marker enzymes: catalase for peroxisomes, glutamate dehydrogenase for mitochondria and esterase for endoplasmic reticulum. Squalene synthase activity was not detectable in the peroxisomal fraction. Incubation of Hep G2 cells with U18666A, an inhibitor of the cholesterol synthesis at the site of oxidosqualene cyclase, together with heavy high density lipoprotein, which stimulates the efflux of cholesterol, led to a marked increase in the activity of squalene synthase as well as HMG-CoA reductase, whereas no significant effect on the marker enzymes was observed. Neither enzyme activity was detectable in the peroxisomal density gradient fraction, suggesting that in Hep G2-peroxisomes cholesterol synthesis from the water-soluble early intermediates of the pathway cannot take place. Both stimulated and non-stimulated cells gave rise to preparations where squalene synthase activity was comigrating with the reductase activity at the lower density side of the microsomal fraction; however, it was also present at the high density side of the microsomal peak, where reductase activity was not detected.
Androstenes↗
A short list at the Royal Free.
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Discrimination, Psychological↗
Effects of DNA looping on pyrimidine dimer formation.
We have assessed the effects of DNA curvature on pyrimidine dimer (PD) formation by examining the pattern of PD formation in DNA held in a loop by lambda repressor. The loop region was composed of diverse DNA sequences such that potential PD sites occurred throughout the loop. PD formation in the loop occurred with peaks at approximately 10 base intervals, just 3' of where the bending of the DNA was inferred to be toward the major groove. This relationship between the peaks and the DNA curvature is essentially identical to that observed in the nucleosome. This indicates that DNA curvature is the major source of the periodicity of PD formation in the nucleosome, and supports an earlier model of the conformation of nucleosomal DNA based on PD formation. DNA loops containing diverse sequences should be of general value for assessing the effects of DNA curvature on DNA modification by other agents used to probe DNA-protein interactions and DNA conformation.
Base Sequence↗
Methylated ubiquitin inhibits cyclin degradation in clam embryo extracts.
A derivative of ubiquitin in which amino groups were blocked by reductive methylation was used to study the possible role of the ubiquitin pathway in the cell cycle-programmed degradation of cyclin. It was shown previously that methylated ubiquitin can be efficiently ligated to protein substrates, but cannot form polyubiquitin chains. In the well-characterized ubiquitin-dependent proteolytic system from reticulocytes, it was found that rates of protein breakdown obtained with methylated ubiquitin are generally slower than those with ubiquitin; and thus, this derivative can be used, in some cases, as an inhibitor of ubiquitin-dependent protein degradation. The addition of methylated ubiquitin to a cell-free system from fertilized clam oocytes inhibited the degradation of both cyclins A and B. That this was due to specific interference with ubiquitin function was indicated by the observation that the supplementation of excess ubiquitin completely overcame the inhibitory action of methylated ubiquitin on cyclin degradation. These findings suggest that polyubiquitin chain formation is required for cyclin degradation.
Animals↗
Biosynthetic capacity of the human lens upon aging.
Lipid and protein biosynthesis were investigated in organ culture of human lenses from subjects ranging from 8 to 95 years old. A marked increase of protein and cholesterol content of the human lens with age was found. No significant linear correlation of age with either cholesterol or fatty acid synthesis was observed. Moreover, there are no indications that either synthesis declines with increasing age of the subject. Notwithstanding the large variety in age of the subjects, cholesterol and fatty acid synthesis of the individual lenses were correlated with each other in a positive linear way. Non-cataractous lenses as old as 80 years revealed a normal protein synthesis pattern as determined by incorporation of [35S]methionine into protein and analysis by 2D-gel electrophoresis. In addition, they maintained a high protein biosynthetic capacity.
Acetates↗
Identification of a naturally occurring methyl-ester of phosphate, methyl-phosphorylcholine (methyl-2-(N,N,N trimethylamino) ethyl phosphate), in the eggs of the sea urchin S. purpuratus.
A novel metabolite of choline, phosphorylcholine methyl ester, has been identified in the eggs of S. purpuratus wherein it is present at approximately 1 mM concentration. To the best of our knowledge, this is the first instance of a phosphoryl-methyl-ester to be observed in nature. The compound appears to be species specific, since it has not been observed in other species such as L. pictus and P. depressus. In S. purpuratus its distribution is confined to the ovary, eggs and embryos, and is absent from young animals following metamorphosis.
Animals↗
Postnatal developmental profile of 3-hydroxy-3-methylglutaryl-CoA reductase, squalene synthetase and cholesterol 7 alpha-hydroxylase activities in the liver of domestic swine.
Activities of 3-hydroxy-3-methylglutaryl-CoA reductase, squalene synthetase and cholesterol 7 alpha-hydroxylase, measured in liver microsomal preparations from domestic swine between birth and adolescence, correlated strongly in individual animals. A synchronous increase was observed between 4 and 6 weeks after birth, i.e., immediately after weaning. Rise in activity was highest for HMG-CoA reductase (30-fold), and smallest for squalene synthetase (5-fold). In pubertal pigs (16 to 30 weeks old), activities of these enzymes had the same low values as in suckling piglets. The increase of both HMG-CoA reductase and squalene synthetase activities may be caused by the shift from high-cholesterol milk intake to a chow diet with low-cholesterol content. The rise in cholesterol 7 alpha-hydroxylase activity might be due to other dietary or hormonal factors.
Animals↗
Intrinsic religiousness and religious coping as life stress moderators for Catholics versus Protestants.
Two prospective studies were conducted to test the stress-moderating effects of intrinsic religiousness and overall religious coping on the depression and trait anxiety of Catholic and Protestant college students. Both studies found a significant cross-sectional interaction between controllable life stress and religious coping in the prediction of Catholics' depression, with religious coping serving a protective function at a high level of controllable negative events. Both studies also found a significant prospective interaction between uncontrollable life stress and intrinsic religiousness in the prediction of Protestants' depression; the relationship between uncontrollable stress and depression was positive for low intrinsic Protestants, flat for medium intrinsic Protestants, but negative for high intrinsic Protestants. The findings are discussed in terms of their implications for the role of religion in life stress adjustment.
Adaptation, Psychological↗