Mitochondrial membrane potential, transmembrane difference in the NAD redox potential and the equilibrium of the glutamate-aspartate translocase in the isolated perfused rat heart

The distribution of glutamate and aspartate and the mitochondrial membrane potential (Δψ) were studied in isolated rat heart mitochondria and in the intact perfused rat heart. The diffusion potential imposed by the glutamate-aspartate exchange through mediation of the electrogenic glutamate-aspartate translocator attained a value close to the mitochondrial Δψ measured from the distribution of triphenylmethylphosphonium ion (TPMP⁺) both in isolated mitochondria and in intact myocardium. Distributions of the Δψ probe and metabolites were determined by subcellular fractionation of the heart muscle in a non-aqueous medium. The results indicate that the glutamate-aspartate translocator is in near equilibrium in the myocardium. The diffusion potential of the glutamate-aspartate exchange, and the mitochondrial/cytosolic difference in the redox potentials of the free NAD⁺/NADH pools are equal allowing for experimental error. These data obtained from intact tissue can therefore be interpreted as supporting the notion of the transmembrane uphill transport of reducing equivalent from the cytosolic free NAD⁺/NADH pool being driven by the malate-aspartate cycle energized by the mitochondrial Δψ.     

A colcemid-sensitive mechanism involved in regulation of chromosome movements during meiotic pairing

Active movements of the chromosomes may be needed in the process, where homologous chromosomes find each other during the meiotic pairing. Because the components of the cytoskeleton are generally believed to be responsible for all movements in living nonmuscle cells, we have analyzed the regulation of the movements of zygotene chromosomes in the male rat by using specific inhibitors of the assembly of the various components of the cytoskeleton. — Colcemid, an inhibitor of microtubule formation, completely inhibited the chromosome movements in vitro at a concentration of 1 g/ml. This was associated with a damage of the nuclear envelope revealed by the electron microscopic analysis. Another inhibitor of microtubule formation, vinblastine, was ineffective below the level of general toxicity (100 g/ml). A specific microfilament inhibitor, cytochalasin B was similarly ineffective. — The findings suggest the presence of a specific colcemid-sensitive mechanism in the nuclear envelope of the zygotene spermatocytes, which regulates the movements of the chromosomes during meiotic pairing.     

Thermal inactivation and reactivation of an enzyme in vivo. Pantothenate hydrolase of Pseudomonas fluorescens

Thermal inactivation and reactivation of pantothenate hydrolase were studied in whole cells of Pseudomonas fluorescens. The enzyme is susceptible to thermal inactivation in whole cells at 37-40 degrees C, and is reactivated when the temperature is lowered again. Chloramphenicol does not prevent reactivation. The activation energy of enzyme inactivation in vivo is about 540kJ/mol. This activation energy is 220kJ/mol in vitro, but it is increased to 550-630kJ/mol by several metabolites, such as succinate, glyoxylate and oxalate. Generally, good carbon sources, causing rapid growth, protect the enzyme from thermal inactivation in vivo, and enable reactivation to occur at a fast rate. The enzyme is also inactivated below 35 degrees C, showing an activation energy of about 35kJ/mol. Good carbon sources prevent this inactivation as well, and cause slight reactivation. Glycine, although not utilized for growth, protects the enzyme well from this inactivation but not from inactivation at 37-40 degrees C, and prevents reactivation totally. From the activation energies of inactivation and the effects of the various carbon sources, it appears possible that changes in the concentrations of intracellular metabolites may be responsible for the changes in inactivation and reactivation.     

Vitamin C deficiency and risk of myocardial infarction: prospective population study of men from eastern Finland.

OBJECTIVE: To examine the association between plasma vitamin C concentrations and the risk of acute myocardial infarction. DESIGN: Prospective population study. SETTING: Eastern Finland. SUBJECTS: 1605 randomly selected men aged 42, 48, 54, or 60 who did not have either symptomatic coronary heart disease or ischaemia on exercise testing at entry to the Kuopio ischaemic heart disease risk factor study in between 1984 and 1989. MAIN OUTCOME MEASURES: Number of acute myocardial infarctions; fasting plasma vitamin C concentrations at baseline. RESULTS: 70 of the men had a fatal or non-fatal myocardial infarction between March 1984 and December 1992.91 men had vitamin C deficiency (plasma ascorbate < 11.4 mumol/l, or 2.0 mg/l), of whom 12 (13.2%) had a myocardial infarction; 1514 men were not deficient in vitamin C, of whom 58 (3.8%) had a myocardial infarction. In a Cox proportional hazards model adjusted for age, year of examination, and season of the year examined (August to October v rest of the year) men who had vitamin C deficiency had a relative risk of acute myocardial infarction of 3.5 (95% confidence interval 1.8 to 6.7, P = 0.0002) compared with those who were not deficient. In another model adjusted additionally for the strongest risk factors for myocardial infarction and for dietary intakes of tea fibre, carotene, and saturated fats men with a plasma ascorbate concentration < 11.4 mumol/l had a relative risk of 2.5 (1.3 to 5.2, P = 0.0095) compared with men with higher plasma vitamin C concentrations. CONCLUSIONS: Vitamin C deficiency, as assessed by low plasma ascorbate concentration, is a risk factor for coronary heart disease.     

Mycoflora of the Finnish “sauna” (bath-house)

The investigation concerned the mycoflora of the old, in some cases already discarded, bath houses (sauna) situated in various parts of Finland for the purpose of finding the fungus,Phialophora pedrosoi, which causes chromoblastomycosis in man. The material, 220 specimens, consisting of thin shavings from the wooden floors and platforms, was collected from 78 bath houses. P. pedrosoi was not found in this investigation; the followingPhialophora-species:P. fastigiata, P. lagerbergii, P. lignicola andP. verrucosa were isolated, however.In total 98 various fungus species were isolated, the number of isolates was 1052. Phycomycetes represented 9.2 % of the species and 15.2 % of the isolates, Ascomycetes 4.1 % and 3.4 %, Basidiomycetes 4.1 % and 0.7 % and Fungi imperfecti 82.6 % and 80.7 % respectively of the species and the isolates. The species most frequently isolated from specimens were:Trichoderma viride, Mucor mucedo, Stemphylium consortiale, Rhinocladiella atrovirens, Arthrobotrys superba andPhialophora verrucosa.In all, 13 nematode-destroying fungus species were isolated belonging to the genera:Nematoctonus, Cephalosporium, Harposporium, Dactylaria, Arthrobotrys andDactylella. 60 % of the bath houses were infected by nematodes. Gliodendron gen. nov. with the speciesGliodendron balnicolum sp. nov. is described and illustrated.

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Zusammenfassung Die Untersuchung betraf die Mykoflora von alten, in manchen Fällen bereits aufgegebenen Badehäusern, die sich in verschiedenen Teilen Finnlands befanden, um den Pilz,Phialophora pedrosoi, den Erreger der Chromoblastomykose, zu finden. Das Material bestand aus 220 Einzelstücken, Spänen von Holzfußböden, die in 78 Badehäusern gesammelt wurden,Phialophora pedrosoi wurde in diesen Untersuchungen nicht gefunden. Die folgendenPhialophora-Arten sind isoliert worden:P. fastigiata, P. lagerbergii, P. lignicola, undP. verrucosa. Insgesamt sind 98 verschiedene Pilzarten isoliert worden. Die Anzahl der Isolierungen war 1052. Phykomyceten stellten 9,2 % von den Arten und 15,2 % der Isolierungen dar: Askomyceten 4,1 % und 3,4 %; Basidiomyceten 4,1 % und 0,7 %; Fungi imperfecti 82,6 % und 80,7 %. Die häufigst isolierten Arten waren:Trichoderma viride, Mucor mucedo, Stemphylium consortiale, Rhinocladiella atrovirens, Arthrobotrys superba undPhialophora verrucosa. Alles in allem wurden 13 Nematoden zerstörende Arten isoliert, die zu den folgenden Gattungen gehörten:Nematoctonus, Cephalosporium, Harposporium, Dactylaria, Arthrobotrys undDactylella. Sechzig Prozent der Badehäuser waren von Nematoden infektiert.Gliodendron gen. nov. ist mit der ArtGliodendron balnicolum, sp. nov. beschrieben und abgebildet worden.

     

Advantages from diel vertical migration can explain the dominance of Gonyostomum semen (Raphidophyceae) in a small, steeply-stratified humic lake

In late summer, a large flagellated alga, Gonyostomum semen(Raphidophyceae), constituted most of the phytoplankton biomassin a small steeply-stratified humic lake. Its diel verticalmigration (DVM) was very distinct and extended at night intothe anoxic hypolimnion. After midsummer, the depletion of hypolimneticoxygen led to a gradual release of soluble reactive phosphorus(SRP) from the sediment, but one month later, irrespective ofcontinuing stratification, the concentrations again returnedto undetectable levels down to the bottom. As this coincidedwith the rapid increase in G.semen population, the latter wasprobably responsible for the depletion of SRP. The flux of SRPfrom the sediment to the epilimnion was virtually interrupted,making non-migrating phytoplankton dependent only on regeneratedand inflowing inorganic phosphorus. Besides nutrient availability,DVM also benefited G.semen in the reduction of metabolic andgrazing losses. In this lake, the remarkable multiple advantagesof DVM probably explain the dominance of the large G.semen inthe late summer phytoplankton biomass over much smaller algae.     

Magnesium dependence of the measured equilibrium constants of aminoacyl-tRNA synthetases

The apparent equilibrium constants (K') for six reactions catalyzed by aminoacyl-tRNA synthetases from Escherichia coli were measured, the equations for the magnesium dependence of the equilibrium constants were derived, and best-fit analyses between the measured and calculated values were used. The K' values at 1 mM Mg(2+) ranged from 0.49 to 1.13. The apparent equilibrium constants increased with increasing Mg(2+) concentrations. The values were 2-3 times higher at 20 mM Mg(2+) than at 1 mM Mg(2+), and the dependence was similar in the class I and class II synthetases. The main reason for the Mg(2+) dependence is the existence of PP(i) as two magnesium complexes, but only one of them is the real product. AMP exists either as free AMP or as MgAMP, and therefore also has some effect on the measured equilibrium constant. However, these dependences alone cannot explain the measured results. The measured dependence of the K' on the Mg(2+) concentration is weaker than that caused by PP(i) and AMP. Different bindings of the Mg(2+) ions to the substrate tRNA and product aminoacyl-tRNA can explain this observation. The best-fit analysis suggests that tRNA reacts as a magnesium complex in the forward aminoacylation direction but this given Mg(2+) ion is not bound to aminoacyl-tRNA at the start of the reverse reaction. Thus Mg(2+) ions seem to have an active catalytic role, not only in the activation of the amino acid, but in the posttransfer steps of the aminoacyl-tRNA synthetase reaction, too.     

Distribution of aquaporin 4 on sarcolemma of fast-twitch skeletal myofibres

The aquaporin 4 (AQP4) water channel is present on the sarcolemma of fast-twitch-type skeletal myofibres. We have examined the distribution of AQP4 in relation to sarcolemmal domain structure and found that AQP4 protein is not evenly distributed on the sarcolemma. Immunofluorescence staining of isolated single myofibres indicated a punctate staining pattern overlapping with the dystrophin glycoprotein complex, but with the transverse tubule openings being left clear. Myotendinous and neuromuscular junctions also lacked AQP4, despite their high content of the dystrophin glycoprotein complex. The destruction of caveoli with methyl-β-cyclodextrin did not change the distribution of AQP4 at the sarcolemma. Moreover, AQP4 did not float with the caveolar marker caveolin-3 in sucrose gradients after Triton X-100 extraction at 4°C. These data indicated that AQP4 was not associated with caveoli. Surprisingly, m. flexor digitorum brevis fibres, although of the fast-twitch type, often lacked AQP4. Furthermore, those fibres harbouring AQP4 at the sarcolemma showed a regionalized distribution, suggesting that large areas were devoid of the protein. Blockage of the synthesized proteins in the endoplasmic reticulum with brefeldin A showed that, in spite of its regionalized sarcolemmal distribution, AQP4 was synthesized along the entire length of the fibres. These results suggest functional differences in the water permeability of the sarcolemma not only between the fast-twitch muscles, but also within single muscle fibres.     

Type 2 diabetes whole-genome association study in four populations: the DiaGen consortium

Type 2 diabetes (T2D) is a common, polygenic chronic disease with high heritability. The purpose of this whole-genome association study was to discover novel T2D-associated genes. We genotyped 500 familial cases and 497 controls with >300,000 HapMap-derived tagging single-nucleotide-polymorphism (SNP) markers. When a stringent statistical correction for multiple testing was used, the only significant SNP was at TCF7L2, which has already been discovered and confirmed as a T2D-susceptibility gene. For a replication study, we selected 10 SNPs in six chromosomal regions with the strongest association (singly or as part of a haplotype) for retesting in an independent case-control set including 2,573 T2D cases and 2,776 controls. The most significant replicated result was found at the AHI1-LOC441171 gene region.