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A comparative assessment of the expression of the mitochondrial fission marker Drp1 and the autophagy marker LC3 in neurons and endothelial cells in the hippocampus and entorhinal cortex during progression of cognitive deficit was performed in animals with intrahippocampal administration of ß-amyloid. In both brain regions, the expression of Drp1 and LC3 in neuronal and endothelial cells was enhanced. The peak of cognitive impairment corresponded to the maximum expression of Drp1 and LC3 in hippocampal neurons and was preceded by an increase in the number of Drp1+ and LC3+ endothelial cells in this brain region. These data attests to a possible role of aberrant mitochondrial dynamics and autophagy of endothelial cells in the impairment of brain plasticity in the Alzheimer's type neurodegeneration.

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A conception on amyloidosis as a key factor of neuronal death in neurodegenerative diseases is stated. Experimental evidence is presented that amyloidosis is caused by alterations in the activity of a number of enzymes as well as conformational changes in pathogenic proteins. Arguments for amyloidosis as the universal biological mechanism of specific elimination of neurons showing changed metabolic and physiological status of cell differentiation are adduced. The final pattern of cell death seems to differ cardinally from that in both apoptosis and necrosis.

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Using gel filtration and ultracentrifugation, the quaternary structure of muscle creatine kinase (isoenzyme MM) has been shown to depend on pH, protein concentration and ionic strength of solution. In solution the enzyme can exist not only as a dimer, but also as a monomer and tetramer with molecular weights of 40 000 and 160 000, respectively. High dilution of the protein solution and pH 6.0--5.0 provide for the monomer formation; the dimer can arise under various conditions, while the tetramer requires pH 9.0 and high protein concentration. The monomer differs from the dimer by a higher enzymatic activity, lability and needs thiol to maintain its catalytic activity. Dissociation of creatine kinase into more active subunits accounts for the increase in specific activity induced by protein dilution. It is assumed that dissociation can have physiological significance as one of the mechanisms of creatine kinase activation in muscles.

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It is shown that lactate is a noncompetitive inhibitor of muscle creative kinase Ki 16 mmol with respect to MgATP. When inhibitors are applied in combination (lactate with glucose-6-phosphate or inorganic phosphate, glucose-6-phosphate with Pi or all three inhibitors together) a competition arises. When applied together the combined action is weaker than the sum of inhibition of three effectors in the case of separate application. The competition is revealed in model experiments with metabolite concentrations peculiar to resting and acting muscle. The supposition is made that formation of lactate, glucose-6-phosphate and inorganic phosphate in the muscle tissue can induce partial, but not complete inhibition of the creatine kinase activity.

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Inhibition of rabbit muscle creatine kinase by lactate, glucose-6-phosphate and PEP occurs immediately and depends on the concentration of the effectors and enzyme properties. Desensitization of creatine kinase to these effectors is described. Partial desensitization to G-6-P and lactate occurs after the enzyme preincubation with dithiothreitol. A complete loss of sensitivity to lactate and a significant loss of sensitivity to G-6-P and PEP occurs after the enzyme storage at -15 degrees during several weeks. The data obtained support our previously made assumption on the allosteric regulation of the MM isoenzyme of creatine kinase.

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Phosphoenolpyruvate (PEP) is a potent inhibitor of muscle creatine kinase. The inhibition of ATP formation is more pronounced at pH 7.0. The substrates, creatine and creatine phosphate, partially prevent the inhibition of the enzyme activity by PEP, creatine being as an effector.

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The dependence of the reaction rate on substrate concentrations at pH 8.0--7.5 does not submit the Michaelis-Menten kinetics. The dependence of v on Mg-ATP is described with a curve having an intermediate plateau. The dependence of v on the creatine concentration is expressed by a curve, which is not hyperbolic. In this case the index of the substrate concentration, (q), is variable, and it increases with the increase of creatine concentration from 1 to 2 (at the presence of effectors, PEP and ADP,--from 1 to 3.5). The specific creatine kinase activity increases 3--4-fold with protein dilution, but this effect is not observed in the presence of inhibitors to FDP and PEP. Creatine kinase is desensibilized with respect to FDP and PEP after a prolonged storage. The data obtained and the presence of an effector set indicate, that muscle creatine kinase is a regulated allosteric enzyme.

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The effect of adenylic acid, glucose-6-phosphate, fructose-1,6-diphosphate and phosphoenolpyruvate on creatine kinase isoenzymes (brain extract, muscle and heart extracts and purified muscle enzyme) was studied. These effectors, especially phosphoenolpyruvate, are shown to inhibit in different degree the reaction of ATP formation catalysed by creatine kinase from all tissues. The effectors do not inhibit the creatine phosphate synthesis in extracts, but depress purified creatine kinase. The interrelationship of the creatine kinase system and the key glycolytic enzymes (phosphofructokinase, hexokinase, pyruvate kinase) is discussed.

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