Biochemical medicine and metabolic biology最新文献

{"title":"High-Density Lipoprotein 3 Stimulates Phosphatidylcholine Breakdown and Sterol Translocation in Rat Aortic Smooth Muscle Cells by a Phospholipase C/Protein Kinase C-Dependent Process","authors":"Dusserre E.,&nbsp;Pulcini T.,&nbsp;Bourdillon M.C.,&nbsp;Berthezene F.","doi":"10.1006/bmmb.1994.1032","DOIUrl":"10.1006/bmmb.1994.1032","url":null,"abstract":"<div><p>The aim of this study was to elucidate signal transduction pathways following high-density lipoprotein 3 (HDL3) fixation to HDL high-affinity binding sites and leading to translocation of newly synthesized cholesterol to the plasma membrane pool for efflux. First, membrane phosphatidylcholine (PC) breakdown and 1,2-diacylglycerol (DAG) production were investigated following HDL3 or tetranitromethane (TNM)-HDL incubation with smooth muscle cells in culture. Second, newly synthesized cholesterol was labeled using [³H] mevalonolactone. Phospholipase C (PLC) and protein kinase C (PKC) were stimulated using carbachol and phorbol 12-myristate 13-acetate. Translocation and efflux of newly synthesized cholesterol were monitored using the cholesterol oxidase method and TNM-HDL as cholesterol acceptor. Results showed that: (1) native HDL3 but not modified HDL was able to stimulate PC breakdown and DAG formation; and (2) PLC and PKC stimulation using specific agents induce cholesterol translocation from intracellular to plasma membrane pool. Taken together, these two sets of results suggest that native HDL3 could induce cholesterol translocation by a PLC/PKC process in smooth muscle cells.</p></div>","PeriodicalId":8752,"journal":{"name":"Biochemical medicine and metabolic biology","volume":"52 1","pages":"Pages 45-52"},"PeriodicalIF":0.0,"publicationDate":"1994-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1006/bmmb.1994.1032","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18913857","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Application of Molecular Genetics in Public Health: Improved Follow-up in a Neonatal Hemoglobinopathy Screening Program","authors":"Zhang Y.H.,&nbsp;Mccabe L.L.,&nbsp;Wilborn M.,&nbsp;Therrell B.L.,&nbsp;Mccabe E.R.B.","doi":"10.1006/bmmb.1994.1030","DOIUrl":"10.1006/bmmb.1994.1030","url":null,"abstract":"<div><p>Newborn screening for the hemoglobinopathies has been shown to reduce morbidity and mortality, particularly for sickle cell anemia, by facilitating initiation of penicillin prophylaxis by 4 months of age. The purpose of the current investigation was to determine whether molecular genetic follow-up testing could be introduced into a neonatal hemoglobinopathy screening program and, if successfully introduced, whether it would reduce time to diagnostic confirmation. Between July 1, 1991, and October 7, 1992, 518 original dried blood specimens were referred from the Texas Department of Health Neonatal Hemoglobinopathy Screening Program for molecular genetic follow-up testing. Allele-specific cleavage (ASC) after amplification with matched and mismatched polymerase chain reaction primers was compared to allele-specific oligonucleotide (ASO) hybridization. By November 2, 1992, molecular genetic analyses were definitive in 506, and agreement was observed between ASC and ASO hybridization in all specimens analyzed. Approximately 13% of those initially screened FS were considered probable S/β-thal by DNA and RNA testing. Rapid molecular genetic analysis contributed to a substantial reduction of the mean age at confirmation by approximately 50%, to about 2 months of age. ASC is a reliable method for molecular genetic analysis of dried blood specimens, providing methodology which can be readily automated. An automated method is demonstrated that is based on microtiter plate technology and will significantly reduce labor intensity and costs, while increasing sample throughput. Even with current manual testing methods, DNA and RNA analysis of initial newborn screening specimens will reduce the age at confirmation well under 4 months, the age cut-off for effective initiation of penicillin prophylaxis.</p></div>","PeriodicalId":8752,"journal":{"name":"Biochemical medicine and metabolic biology","volume":"52 1","pages":"Pages 27-35"},"PeriodicalIF":0.0,"publicationDate":"1994-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1006/bmmb.1994.1030","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18530095","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Contribution to Control of Mitochondrial Oxidative Phosphorylation by Supplement of Reducing Equivalents","authors":"Kunz W.,&nbsp;Gellerich F.N.,&nbsp;Schild L.","doi":"10.1006/bmmb.1994.1035","DOIUrl":"10.1006/bmmb.1994.1035","url":null,"abstract":"<div><p>The influence of enhancing the supply of hydrogen donors on respiratory rates, NAD(P)H fluorescence, and membrane potential was investigated. Addition of 5 mM malate to mitochondria during oxidation of 10 mM isocitrate, oxoglutarate, succinate, proline, or glycerol-3-phosphate under steady-state conditions resulted in an inhibition of respiration, coincident with a decrease in both transmembrane electrical potential and percentage reduction of NAD(P). Half-maximum inhibition of NAD(P) reduction in the resting state of 10 mM isocitrate respiration was reached at 10 mM malate. This inhibition was concluded to be due to oxaloacetate formed immediately from malate by succinate dehydrogenase. Addition of 5 mM isocitrate caused higher respiratory rates, accompanied by an increase in both Δψ and percentage of NAD(P) reduction, in mitochondria oxidizing 10 mM oxoglutatate, glutamate, proline, hydroxybutyrate, glycerol-3-phosphate, or 0.025 mM palmitoyl carnitine. The half maximum increase in percentage NAD(P) reduction with 10 mM 2-oxoglutarate as primary substrate was found at 0.24 mM isocitrate. Within the citric acid cycle, succinate dehydrogenase and NAD-isocitrate dehydrogenase play an important role in changes in the rate of NADH formation. Therefore, they participate in flux control. Furthermore, mitochondrial aspartate aminotransferase and oxidoreductases of the β-oxidation pathway of fatty acids are additionally involved in adjusting the rate of NADH formation.</p></div>","PeriodicalId":8752,"journal":{"name":"Biochemical medicine and metabolic biology","volume":"52 1","pages":"Pages 65-75"},"PeriodicalIF":0.0,"publicationDate":"1994-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1006/bmmb.1994.1035","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18913159","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Enzyme Replacement Therapy for Gaucher Disease: Critical Investigations beyond Demonstration of Clinical Efficacy","authors":"Brady R.O.,&nbsp;Barton N.W.","doi":"10.1006/bmmb.1994.1026","DOIUrl":"10.1006/bmmb.1994.1026","url":null,"abstract":"<div><p>Enzyme replacement therapy is highly effective for patients with Type 1 Gaucher disease. In order to estimate the quantity of enzyme that would be necessary for clinical benefit, we conducted a single-infusion, dose-response study in nonsplenectomized patients with Gaucher disease. Biochemical and histologic changes were compared in liver biopsy specimens obtained before and 44 h following the infusion of varying quantities of enzyme. Based on the information obtained from this investigation, patients in our initial clinical efficacy trial were given 60 IU of macrophage-targeted glucocerebrosidase/kg body wt every other week. All patients had significant improvement of their anemia and reduction of splenomegaly after 6 months of treatment. In a subsequent investigation, 10 moderately symptomatic patients with intact spleens were given 10 IU of glucocerebrosidase/kg body wt every other week. After 6 months of treatment, only a portion of these patients had beneficial responses. We concluded that the rate and extent of response to enzyme replacement therapy in patients with Gaucher disease are dependent upon the quantity of enzyme administered. When treatment is initiated in patients with mild to moderately severe disease, a lower dose of enzyme can be selected. Moreover, the maintenance dose of glucocerebrosidase has been shown to be much less than the amount initially required to reduce the accumulated lipid. Some patients require enzyme infusions on only a monthly basis, and it is possible that even this frequency may eventually be reduced. These refinements in treatment strategy merit serious consideration for the long-term management of patients with Gaucher disease.</p></div>","PeriodicalId":8752,"journal":{"name":"Biochemical medicine and metabolic biology","volume":"52 1","pages":"Pages 1-9"},"PeriodicalIF":0.0,"publicationDate":"1994-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1006/bmmb.1994.1026","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18913848","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Vitamin B6 Metabolism and Diabetes","authors":"Rogers K.S.,&nbsp;Mohan C.","doi":"10.1006/bmmb.1994.1027","DOIUrl":"10.1006/bmmb.1994.1027","url":null,"abstract":"","PeriodicalId":8752,"journal":{"name":"Biochemical medicine and metabolic biology","volume":"52 1","pages":"Pages 10-17"},"PeriodicalIF":0.0,"publicationDate":"1994-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1006/bmmb.1994.1027","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18913851","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Sodium Chloride Cotransport at the Basolateral Membrane of Type II Pneumocytes","authors":"Shabarek F.,&nbsp;Xu F.,&nbsp;Oelberg D.G.","doi":"10.1006/bmmb.1994.1036","DOIUrl":"10.1006/bmmb.1994.1036","url":null,"abstract":"<div><p>Current evidence indirectly supports the hypothesis that Na⁺/Cl⁻ cotransport occurs at basolateral membranes of type II pneumocytes. To test this hypothesis, enriched apical and basolateral plasma membrane vesicles were prepared from adult bovine type II pneumocytes. Uptake of ²²Na⁺ or ³⁶Cl⁻ by these vesicle populations was monitored over time. Using enriched basolateral vesicles, substitution of formate^- for Cl⁻ nearly eliminated ²²Na⁺ uptake, and substitution of Tris⁺ for Na⁺ significantly reduced ³⁶Cl⁻ uptake. These observations of Cl⁻-dependent ²²Na⁺ uptake and Na⁺-dependent ³⁶Cl⁻ uptake demonstrated coupling between Na⁺ and Cl⁻ absorption at the basolateral membrane. Na⁺/Cl⁻ cotransport inhibitors, furosemide (1 mM) and bumetanide (0.1 mM), also reduced ²²Na⁺ and ³⁶Cl⁻ uptakes by enriched basolateral vesicles. By contrast, furosemide and bumetanide did not affect ²²Na⁺ and ³⁶Cl⁻ uptakes by enriched apical vesicles. Collectively, these observations support localization of Na⁺/Cl⁻ cotransport to the basolateral membranes of mature type II pneumocytes.</p></div>","PeriodicalId":8752,"journal":{"name":"Biochemical medicine and metabolic biology","volume":"52 1","pages":"Pages 76-83"},"PeriodicalIF":0.0,"publicationDate":"1994-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1006/bmmb.1994.1036","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18913160","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Impaired Ketogenesis in Carnitine Depletion Caused by Short-Term Administration of Pivalic Acid Prodrug","authors":"Abrahamsson K.,&nbsp;Eriksson B.O.,&nbsp;Holme E.,&nbsp;Jodal U.,&nbsp;Lindstedt S.,&nbsp;Nordin I.","doi":"10.1006/bmmb.1994.1028","DOIUrl":"10.1006/bmmb.1994.1028","url":null,"abstract":"<div><p>Long-term treatment with pivalic acid prodrug results in impaired ketone-body production. Therefore, it was of interest to investigate whether short-term treatment had any influence on the fatty acid oxidation. In this study six healthy males were given 1200 mg per day of pivmecillinam for 12 days to induce carnitine deficiency. The concentration of free carnitine in serum was reduced from a mean of 42.8 μmol/liter (range, 31-48) to 11.6 μmol/liter (range, 7.0-24), but the muscle carnitine concentration was not reduced. A 36-h fasting test was performed before and after drug administration to study the effect on ketone-body production. After treatment, the two subjects with the lowest level of serum free carnitine at the end of the fasting period had impaired ketogenesis. This indicates a carnitine deficiency in the liver which was reflected in the free carnitine concentration in serum and which could not be compensated for by mobilization of muscle carnitine. We conclude that there is a substantial risk to develop carnitine deficiency and impaired fatty acid oxidation in the liver during short-term treatment with drugs conjugated with pivalic acid.</p></div>","PeriodicalId":8752,"journal":{"name":"Biochemical medicine and metabolic biology","volume":"52 1","pages":"Pages 18-21"},"PeriodicalIF":0.0,"publicationDate":"1994-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1006/bmmb.1994.1028","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18913853","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Characterization of Wild-Type Human Medium-Chain Acyl-CoA Dehydrogenase (MCAD) and Mutant Enzymes Present in MCAD-Deficient Patients by Two-Dimensional Gel Electrophoresis: Evidence for Posttranslational Modification of the Enzyme","authors":"Bross P.,&nbsp;Jensen T.G.,&nbsp;Andresen B.S.,&nbsp;Kjeldsen M.,&nbsp;Nandy A.,&nbsp;Kolvraa S.,&nbsp;Ghisla S.,&nbsp;Rasched I.,&nbsp;Bolund L.,&nbsp;Gregersen N.","doi":"10.1006/bmmb.1994.1031","DOIUrl":"10.1006/bmmb.1994.1031","url":null,"abstract":"<div><p>Two-dimensional gel electrophoresis was used to study and compare wild type medium-chain acyl-CoA dehydrogenase (MCAD; EC 1.3.99.3) and missense mutant enzyme found in patients with MCAD deficiency. By comparing the patterns for wild-type and mutant MCAD expressed in <em>Escherichia coli</em> or in eukaryotic COS-7 cells we demonstrate that variants with point mutations changing the net charge of the protein can be readily resolved from the wildtype protein. After expression of the cDNA in eukaryotic cells two spots representing mature MCAD can be distinguished, one with an isoelectric point (p<em>I</em>) corresponding to that obtained for the mature protein expressed in <em>E. coli</em> and another one shifted to lower p<em>I</em>. This demonstrates that MCAD protein is partially modified after transport into the mitochondria and removal of the transit peptide. The observed p<em>I</em> shift would be compatible with phosphorylation of one aspartic acid residue per monomer. Comparison of pulse labeling and steady-state amounts of MCAD protein in overexpressing COS-7 cells confirms that K304E MCAD is synthesized and transported into mitochondria in amounts similar to the wild type protein, but is degraded much more readily. For wild-type MCAD, the spot representing the nonmodified form predominates after pulse labeling while that representing the modified form is relatively stronger in steady state, demonstrating that the modification occurs in mitochondria after the transit peptide has been removed. For K304E mutant MCAD, the nonmodified spot is relatively stronger both in pulse labeling and in steady state, indicating that either the efficiency of modification or the stability of the modified form is affected by the K304E mutation. Detection of both wild-type and K304E mutant MCAD was achieved in lymphoblastoid cells from patients and carriers of the mutation. Both spots for the wild-type but only the nonmodified spot for the K304E mutant could be detected. In lymphoblastoid cells from carriers, the intensity of the spot representing the mutant protein is much weaker than the two spots representing wild-type MCAD, emphasizing that the K304E mutant protein is more susceptible to degradation than wild-type MCAD. The absence of detectable amounts of modified K304E mutant MCAD protein in these cells suggest that the conclusion drawn from COS-7 cell expression is also valid in patient cells.</p></div>","PeriodicalId":8752,"journal":{"name":"Biochemical medicine and metabolic biology","volume":"52 1","pages":"Pages 36-44"},"PeriodicalIF":0.0,"publicationDate":"1994-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1006/bmmb.1994.1031","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"18913856","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Choline Deficiency in Cultured Adrenal Medullary Cells: Effect on Phosphatidylcholine Biosynthesis","authors":"Percy A.K.,&nbsp;Moore J.F.","doi":"10.1006/bmmb.1994.1022","DOIUrl":"10.1006/bmmb.1994.1022","url":null,"abstract":"<div><p>The effect of choline deficiency on the composition and biosynthesis of the major membrane phospholipids was examined in adrenal medullary cells maintained in suspension cultures. The amount and proportions of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) in these cells were not affected by the removal of choline from the culture media. However, the rate of biosynthesis of choline at the phosphatide level by the stepwise methylation of PE increased twofold within 24 h after choline was removed from the culture media, while ethanolamine incorporation into PE was increased by 50%. In contrast, the rate of incorporation of labeled choline into PC, presumably via CDP-choline, was virtually identical in cells that had been preincubated in the presence or absence of 1 mM choline. These results demonstrate that cultured cells of neural origin are capable of compensating for lack of exogenous choline by forming choline at the phosphatide level through the sequential methylation of PE. The hypolipidemic drug, DH-990, when added to the culture media, inhibited conversion of phosphatidylmonomethylethanolamine (PME) to PC, but had no effect on the N-methylation of PE. This differential effect indicates that the initial N-methylation of PE is catalyzed by an enzyme that is distinguishable from the enzyme(s) catalyzing the conversion of PME to PC.</p></div>","PeriodicalId":8752,"journal":{"name":"Biochemical medicine and metabolic biology","volume":"51 2","pages":"Pages 169-174"},"PeriodicalIF":0.0,"publicationDate":"1994-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1006/bmmb.1994.1022","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"19036978","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Structure and Expression of Mammalian Peroxisome Assembly Factor-1 (PMP35) Genes","authors":"Wilson G.N.,&nbsp;Bryant D.D.","doi":"10.1006/bmmb.1994.1018","DOIUrl":"10.1006/bmmb.1994.1018","url":null,"abstract":"<div><p>Mammalian genes encoding a 35-kDa peroxisomal membrane protein (PMP35, peroxisome assembly factor-1) are compared using the polymerase chain reaction and DNA sequencing. DNA sequencing of the 915 bp of the PMP35 coding regions was in complete agreement with previously published rat data and showed 36 and 133 nucleotide substitutions, respectively, in mouse and man. The 12 and 35 respective amino acid changes encoded by these nucleotide substitutions are clustered and compatible with putative membrane-spanning regions. Rat/human and rat/mouse comparisons yield silent mutation rates of 0.33 and 0.21% per site per million years and replacement mutation rates of 0.082 and 0.076%; transitions account for 67% (human/mouse) and 83% (rat/mouse) of nucleotide replacements among PMP35 genes. PMP35 gene expression in mouse tissues as measured by reverse transcriptase-PCR was responsive to clofibrate and disproportionately high in neural tissue.</p></div>","PeriodicalId":8752,"journal":{"name":"Biochemical medicine and metabolic biology","volume":"51 2","pages":"Pages 140-148"},"PeriodicalIF":0.0,"publicationDate":"1994-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1006/bmmb.1994.1018","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"19037025","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}