Α-glycerophosphate Dehydrogenase Research Articles

НЕКОТОРЫЕ МОРФОФУНКЦИОНАЛЬНЫЕ ОСОБЕННОСТИ МИТОХОНДРИЙ, ИХ СВЯЗЬ С РЕПРОДУКТИВНОЙ ФУНКЦИЕЙ ЖЕНЩИН

Introduction. Mitochondrial morphology and metabolism can have a decisive influence on energy metabolism and reproductive function in women. The objective of this study is to characterize the metabolic and energy metabolism processes in women of reproductive age and those in perimenopause. Methods. The study involved 100 women divided into two equal groups: women of reproductive age and women in perimenopause. Results. All participants had regular menstrual cycles. The duration of menopause in the second group was 1-3 years, with an onset age of 45 years. In total, 64% of the women in group 2 had neurohumoral abnormalities and obesity. Moderate to severe menopause symptoms were detected in 56% of women. Group 2 had reduced activity of mitochondrial enzymes in lymphocytes, such as succinate dehydrogenase and α-glycerophosphate dehydrogenase, while protein, glucose, and uric acid were above normal. Conclusion. This finding suggests that mitochondrial enzymes in group 2 become less active with age. At the end of the reproductive age, the intensity of the energy processes decreases. Cell energy metabolism remains an important area as further research, as this process can shift the timing of reproductive aging in women.

Mitochondrial Disorders in Stroke and Chronic Brain Ischemia

Stroke is the leading cause of disability and death in the adult population. Modern methods of treating patients with acute ischemic stroke include thrombolytic therapy with a narrow therapeutic window and endovascular thrombectomy. The development of other methods of treatment of brain hypoxia in the penumbra zone is relevant. Mitochondria, which are involved in immediate and delayed molecular mechanisms of adaptation to hypoxic stress in the cerebral cortex, primarily respond to hypoxia. Hypoxia induces reprogramming of the mitochondrial respiratory chain function and switching from oxidation of substrates of the respiratory chain complex I to succinate oxidation (complex II). The brain's need for succinate increases. Clinical studies have shown a positive effect of drugs containing succinates on the course of stroke. The study of mitochondrial function is carried out mainly in an experiment. In the present study of mitochondrial disorders in stroke and chronic brain ischemia in adult patients, the quantitative method proposed by A. G. Pearse was used to assess the activity of mitochondrial enzymes of peripheral blood lymphocytes, which are referred to as the enematic mirror of tissues. In acute cerebral ischemia, a compensatory increase in the activity of succinate dehydrogenase was observed on the first day, indicating the tension (increased activity) of the second complex of the mitochondrial respiratory chain.. These data confirm the need to prescribe succinic acid in the acute phase of stroke. At the same time, the dose of 250 mg per day is not sufficient for patients with increased body weight. The standard dose of the drug should be higher, taking into account the different body weight of patients. In patients with stroke, there was also a decrease in the activity of α-glycerophosphate dehydrogenase, which is involved in the fat metabolism of mitochondria, which is an indication for the appointment of carnitine. In chronic brain ischemia, the activity of succinate dehydrogenase and α-glycerophosphate dehydrogenase decreased, indicating indications for the appointment of idebenone and carnitine along with vasodilator therapy and endovascular thrombectomy. Thus, the results of a study of mitochondrial function in patients with acute and chronic brain ischemia are presented. Violations of complex II in the respiratory chain cycle and violation of fat metabolism were revealed, indicating indications for the appointment of energotropic therapy.

Дистальный артрогрипоз 5-го типа – артрогрипоз с офтальмоплегией, полиневропатией. Клинический случай

A clinical case of a 27-year-old patient with distal arthrogryposis of the 5th type – arthrogryposis with ophthalmoplegia, which was combined in a patient with polyneuropathy is presented. To assess tissue respiration (mitochondrial respiratory chain) and other types of metabolism in mitochondria, cytochemical analysis of lymphocytes in peripheral blood was carried out according to A. Pearse's method modified by R.P. Narcissov. The activity of four mitochondrial enzymes involved in carbohydrate metabolism (lactate dehydrogenase), amino acid metabolism (glutamate dehydrogenase), fatty acid metabolism (α-glycerophosphate dehydrogenase) and the second complex of the mitochondrial respiratory chain (succinate dehydrogenase) was assessed. A slight decrease in the activity of the enzyme succinate dehydrogenase, which is part of the second complex of the mitochondrial respiratory chain, was determined. The activity of the enzyme α-glycerophosphate dehydrogenase was more significantly reduced, an increase in the activity of lactate dehydrogenase was noted. In the presented observation, along with the typical manifestations of the disease (contractures of the hands and feet, ophthalmoplegia, ptosis of the eyelids, visual impairment), polyneuropathy with impaired sensitivity of the polyneuritic type was revealed. Thus, a patient with polyneuropathy had a hereditary type 5 arthrogryposis disease. Along with the typical manifestations of the disease, polyneuropathy with hyporeflexia and impaired sensitivity of the polyneuritic type was revealed. Secondary mitochondrial disorders were identified, which was the basis for the appointment of energotropic therapy.

Features of the recovery period of hypoxic lesion to the central nervous system in children of the first year of life with congenital heart disease

Aim. To study the features of recovery period of hypoxic lesion to the central nervous system (CNS) in children of the first year of life in the presence of congenital heart disease (CHD).Material and Methods. The study involved 80 children born full-term and premature with gestational status of 35–37 weeks with hypoxic damage to the CNS. The main observation group comprised 50 children with CHD (interventricular and atrial septal defects, open ductus arteriosus). All children underwent a comprehensive health assessment, standard echocardiography, and neurosonography at ages of five to seven days and one, three, and six months. Biochemical analysis included assessment of serum neurospecific enolase (NSE), succinate dehydrogenase (SDG), and α-glycerophosphate dehydrogenase (α-GPDH). The control group included 20 full-term newborns without CHD and CNS lesions.Results. The main manifestations in newborns with CHD and hypoxic damage to the CNS were the suppression syndrome, agitation, and hypertension-hydrocephalic syndrome. At the age of six months, a delay in motor development indicators persisted in 35% of children in the main group. The high NSE level in newborns with concomitant septal heart defects was associated with a decrease in the quantitative indicators of neuropsychic development (g = –0.6, p < 0.05). The children with CHD and hypoxic damage to the CNS in the first year of life were significantly more often (p < 0.05) deficient in weight and height. A decrease in the resistance level in the first year of life was observed in 40% of children from the main group, which significantly differed compared with group of children without CHD (p < 0.001). The newborns with hypoxic CNS and CHD lesions had a decrease in the activity of α-GPDH and SDG at the age of five to seven days; the low activity of SDG persisted at the ages of one and six months; the enzyme activity in children of the comparison group was normal (p < 0.05).Conclusion. Children with CHD had the features of clinical course of perinatal damage to the CNS in the acute and recovery periods, a slowdown in the rate of physical and neuropsychic development, a decrease in the resistance level, and impaired functional state of the body. The decreases in the activities of SDG and α-GPDH in children with hypoxic lesions to the CNS in the presence of CHD implied the disturbances in cellular bioenergetics and resulted in inadequate response to external factors.

Experimental substantiation of the possibility of using the cytochemical parameters of peripheral blood to assess the functional state of the organism of the pilot

The activity of succinate (mitochondrial), lactate, α-glycerophosphate, glucose-6-phosphate peripheral blood lymphocytes in rats under the conditions of modeling acute and chronic gravitational stress was studied by quantitative spectrophotometry of cytochemical reaction products to reveal the localization of oxidative enzymes. The dynamics of morphofunctional transformations is determined, the severity of which depends on the duration of the effect (the rotation time of the animals on the centrifuge). For acute gravitational stress, changes in the activity of mitochondrial succinate dehydrogenase are characteristic, and in the various stages of chronic gravitational stress, glycolysis (increase in lactate dehydrogenase and α-glycerophosphate dehydrogenase activity) and plastic metabolism (increase in glucose-6-fosfata dehydrogenase activity) are characteristic. These changes are explained by the inclusion of neurohumoral regulatory mechanisms that ensure synchronization of oxidative metabolic processes in the cells of organs of various body systems under unfavorable conditions of their functioning. Significant correlation links between the activity of lymphocyte enzymes with the same indices of neurons of the celiac sympathetic node, adenocytes of the cortical and medulla of the adrenal glands have been established. This suggests that the activity of lymphocyte enzymes can be used as indirect indicators of the processes occurring in the organs of the sipato-adrenomedular and pituitary-adrenocortical systems under conditions of acute and chronic gravitational stress.

НЕКОТОРЫЕ ПОКАЗАТЕЛИ МОРФОФУНКЦИОНАЛЬНОЙ АКТИВНОСТИ ЛИМФОЦИТОВ ПЕРИФЕРИЧЕСКОЙ КРОВИ У ЖЕНЩИН С ОСЛОЖНЕННЫМ ТЕЧЕНИЕМ БЕРЕМЕННОСТИ

The aim of the research was to study some indices of morphofunctional state of lymphocytes in the peripheral blood under a complicated course of pregnancy in women of Khabarovsk. 96 women were examined in different trimesters of gestation. Two main groups depending on the type of complications were formed: the first group consisted of pregnant women with vaginitis, the second group had women whose pregnancy was complicated with clinical manifestations in the form of edemas, proteinuria and hypertensive disorders. The group of comparison included the women without complications. In the peripheral blood of the women under study with the method of high tech automated digital system of blood smear test VISION HEMA there was done a morphometric assessment of lymphocytes and with histochemical methods there was found a level of fermentation activity of succinate dehydrogenase (SDH), α-glycerophosphate dehydrogenase (α-GPDH) and lactate dehydrogenase (LDH). By morphometric studies of lymphocytes of the peripheral blood in pregnant women of the first group in the III trimester of gestation the indices of the square of the cell, of cytoplasm, cytoplasm brightness, the square of the nucleus and the brightness of the nucleus were 102.33±9.83 mсm², 36.84±4.88 mсm², 181.39±6.94 units of optical density, 65.69±6.87 mсm², 97.44±6.88 units of optical density, respectively (in the group of the comparison these data were 121.31±4.44 mсm²; 46.14±2.52 mсm²; 167.74±3.44 units of optical density; 75.33±2.66 mkm², 88.00±3.36 units of optical density, respectively). In the second group of women the same data in the same period of pregnancy corresponded to the following values: 128.44±4.89 mсm², 57.15±3.87 mсm², 173.53±4.65 units of optical density, 70.95±3.39 mсm² and 91.13±4.93 units of optical density (in the group of comparison there were the following data: 109.92±3.57 mсm², 39.63±1.53 mсm², 158.96±2.63 units of optical density, 75.37±3.23 mсm², 82.16±2.58 units of optical density, respectively). By the results of histochemical studies the level of activity of enzymes SDH, α-GPDH, LDH in the first group of pregnancy was 15.38±2.10, 10.72±0.58 and 20.53±2.36 pellet/cell, respectively. And just the other way round in pregnant women of the second group there was revealed a statistically reliable suppression of fermentation activity of these enzymes: 9.11±0.75 (p<0.05); 9.61±1.06 (p>0.05); 8.31±1.20 (p<0.05) pellet/cell in regard to the data in the group of comparison (12.84±1.21; 12.18±1.13 and 13.09±0.78 pellet/cell, respectively). Thus the changes of morphometric indices characterizing parameters of the cell, the nucleus and lymphocytes cytoplasm in a complex study with their fermentation activity can be extra diagnostic criteria of the formation of different complications during pregnancy. The obtained data allow to justify in-time application of medications for the correction of metabolic disorders of immune competent cells with the aim to decrease unfavorable perinatal outcomes.

Resveratrol supplementation does not augment performance adaptations or fibre-type-specific responses to high-intensity interval training in humans.

The present study examined the effect of concurrent exercise training and daily resveratrol (RSV) supplementation (150 mg) on training-induced adaptations following low-dose high-intensity interval training (HIIT). Sixteen recreationally active (∼22 years, ∼51 mL·kg(-1)·min(-1)) men were randomly assigned in a double-blind fashion to either the RSV or placebo group with both groups performing 4 weeks of HIIT 3 days per week. Before and after training, participants had a resting muscle biopsy taken, completed a peak oxygen uptake test, a Wingate test, and a submaximal exercise test. A main effect of training (p < 0.05) and interaction effect (p < 0.05) on peak aerobic power was observed; post hoc pairwise comparisons revealed that a significant (p < 0.05) increase occurred in the placebo group only. Main effects of training (p < 0.05) were observed for both peak oxygen uptake (placebo - pretraining: 51.3 ± 1.8, post-training: 54.5 ± 1.5 mL·kg(-1)·min(-1), effect size (ES) = 0.93; RSV - pretraining: 49.6 ± 2.2, post-training: 52.3 ± 2.5 mL·kg(-1)·min(-1), ES = 0.50) and Wingate peak power (placebo: pretraining: 747 ± 39, post-training: 809 ± 31 W, ES = 0.84; RSV - pretraining: 679 ± 39, post-training: 691 ± 43 W, ES = 0.12). Fibre-type distribution was unchanged, while a main effect of training (p < 0.05) was observed for succinate dehydrogenase activity and glycogen content, but not α-glycerophosphate dehydrogenase activity or intramuscular lipids in type I and IIA fibres. The fold change in PGC-1α, SIRT1, and SOD2 gene expression following training was significantly (p < 0.05) lower in the RSV group than placebo. These results suggest that concurrent exercise training and RSV supplementation may alter the normal training response induced by low-volume HIIT.

Substrate cycling based fluorometric assay for dihydroxyacetone phosphate

A sensitive fluorometric assay for the determination of dihydroxyacetone phosphate (DHAP) is reported here. DHAP is reduced to l-glycerol-3-phosphate with NADH-dependent α-glycerophosphate dehydrogenase. DHAP recycling is provided by oxidation reaction catalysed by α-glycerophosphate oxidase to release hydrogen peroxide. The reaction of hydrogen peroxide with Amplex® Red reagent under horseradish peroxidase catalysis leads to the fluorescent product resorufin. The limit of detection of DHAP is estimated at 1pmol which is roughly 2250 fold more sensitive than the usual DHAP assay based on the detection of NADH by spectrophotometry. This assay is ready-to-use for automated medium-throughput screening.

The death and life of the resurrection drug.

In his own words, it was the most dramatic moment of his scientific career. At the back of an auditorium in Nairobi, Kenya, Cyrus Bacchi met Simon Van Nieuwenhove. Bacchi, at the time, was essentially a biochemist whose interest was the African trypanosome. Van Nieuwenhove was a clinician who had worked for many years in the field in Africa. The topic of their conversation was eflornithine—a small and simple compound that would eventually make a big impact. Bacchi had shown that the compound had an effect on the parasite that causes Human African Trypanosomiasis. Van Nieuwenhove was dosing patients with it. The story of eflornithine did not start with eflornithine.It started because no one knew how to purify an enzyme. Bacchi was working on his thesis, which centred on the α-glycerophosphate shuttle in hemoflagellates. The shuttle acts as a way to transport reducing equivalents from the cytosol to the mitochondrion. α-glycerophosphate dehydrogenase was the enzyme he spent much of his time trying to purify. The reason he could not purify it was it was hidden within another compartment of the trypanosome. It would not be until several years later, and published in 1977, that Fred Opperdoes discovered the glycosome [1]: the organelle that encapsulates glycolysis in trypanosomes, and the organelle that was hiding the enzyme. At every step of the purification process Bacchi lost activity in his enzyme extracts. Magnesium chloride, surprisingly, managed to boost activity. Bacchi looked into other nonmetallic compounds that would act as cations and eventually chose the polyamines: naturally occurring, nitrogen-containing cations whose concentration is closely controlled by the cell. Spermidine and spermine were found to be the best replacements for magnesium, yielding higher activities. With evidence that the enzyme he was trying to purify was most likely locked within the glycosome, Bacchi moved his attention to how the polyamines were made within the trypanosome. This was an area that had amassed a lot of knowledge everywhere apart from in trypanosomes. It was in 1677 that Anton van Leeuwenhoek first observed spermatozoa in humans, dogs, and a host of other organisms, and he later discovered crystals of spermine phosphate in human semen. Modern research had identified the biologically active polyamines—spermidine and spermine—in plants and many types of mammalian cells. The biochemical pathways that make and degrade the polyamines, along with some of their enzymes, had also been identified. At the time, nothing was known about polyamines in protozoa, and in trypanosomes in particular. Did these parasites contain polyamines? Could polyamine metabolism be a useful chemotherapeutic target?

HISTOCHEMISTRY, CAPILLARIZATION, AND MITOCHONDRIAL DENSITIES OF MUSCLE FIBERS ISOLATED FROM THE ILIOFIBULARIS MUSCLE OF AGAMA PALLIDA

ABSTRACT The histochemical and ultrastructural properties of the fiber types present in the iliofibularis muscle isolated from the lizard Agama pallida have been investigated. On the basis of the histochemical activities of alkaline myofibrillar ATPase (m-ATPase), succinic dehydrogenase (SDH), and α-glycerophosphate dehydrogenase (α-GPDH), the pale (outer) region of the iliofibularis was found to contain fast-twitch-oxidative glycolytic fibers, which stain dark for all three enzymes, and tonic fibers, which stain light for m-ATPase, dark for SDH, and moderate for α-GPDH. The cross-sectional area of the white region is 78%, three times the cross-sectional area of the red region (22%). The ultrastructure of the myofibrils and the sarcoplasmic reticulum in fast fibers was compared with that in tonic fibers. Fast glycolytic and fast oxidative glycolytic fibers had straight Z-lines, clear M-bands, and well developed sarcoplasmic reticulum. Furthermore, glycogen granules and lipid droplets were demonstrated in ...

High Ca2+ load promotes Hydrogen peroxide generation via activation of α-glycerophosphate dehydrogenase in brain mitochondria

H2O2 generation associated with α-glycerophosphate (α-GP) oxidation was addressed in guinea pig brain mitochondria challenged with high Ca2+ load (10μM). Exposure to 10μM Ca2+ induced an abrupt 2.5-fold increase in H2O2 release compared to that measured in the presence of a physiological cytosolic Ca2+ concentration (100nM) from mitochondria respiring on 5mM α-GP in the presence of ADP (2mM). The Ca2+-induced stimulation of H2O2 generation was reversible and unaltered by the uniporter blocker Ru 360, indicating that it did not require Ca2+ uptake into mitochondria. Enhanced H2O2 generation by Ca2+ was also observed in the absence of ADP when mitochondria exhibited permeability transition pore opening with a decrease in the NAD(P)H level, dissipation of membrane potential, and mitochondrial swelling. Furthermore, mitochondria treated with the pore-forming peptide alamethicin also responded with an elevated H2O2 generation to a challenge with 10μM Ca2+. Ca2+-induced promotion of H2O2 formation was further enhanced by the complex III inhibitor myxothiazol. With 20mM α-GP concentration, stimulation of H2O2 formation by Ca2+ was detected only in the presence, not in the absence, of ADP. It is concluded that α-glycerophosphate dehydrogenase, which is accessible to and could be activated by a rise in the level of cytosolic Ca2+, makes a major contribution to Ca2+-stimulated H2O2 generation. This work highlights a unique high-Ca2+-stimulated reactive oxygen species-forming mechanism in association with oxidation of α-GP, which is largely independent of the bioenergetic state and can proceed even in damaged, functionally incompetent mitochondria.

Rapid Determination of Myosin Heavy Chain Expression in Rat, Mouse, and Human Skeletal Muscle Using Multicolor Immunofluorescence Analysis

Skeletal muscle is a heterogeneous tissue comprised of fibers with different morphological, functional, and metabolic properties. Different muscles contain varying proportions of fiber types; therefore, accurate identification is important. A number of histochemical methods are used to determine muscle fiber type; however, these techniques have several disadvantages. Immunofluorescence analysis is a sensitive method that allows for simultaneous evaluation of multiple MHC isoforms on a large number of fibers on a single cross-section, and offers a more precise means of identifying fiber types. In this investigation we characterized pure and hybrid fiber type distribution in 10 rat and 10 mouse skeletal muscles, as well as human vastus lateralis (VL) using multicolor immunofluorescence analysis. In addition, we determined fiber type-specific cross-sectional area (CSA), succinate dehydrogenase (SDH) activity, and α-glycerophosphate dehydrogenase (GPD) activity. Using this procedure we were able to easily identify pure and hybrid fiber populations in rat, mouse, and human muscle. Hybrid fibers were identified in all species and made up a significant portion of the total population in some rat and mouse muscles. For example, rat mixed gastrocnemius (MG) contained 12.2% hybrid fibers whereas mouse white tibialis anterior (WTA) contained 12.1% hybrid fibers. Collectively, we outline a simple and time-efficient method for determining MHC expression in skeletal muscle of multiple species. In addition, we provide a useful resource of the pure and hybrid fiber type distribution, fiber CSA, and relative fiber type-specific SDH and GPD activity in a number of rat and mouse muscles.

Ecotype Differentiation in the Face of Gene Flow within the Diving Beetle Agabus bipustulatus (Linnaeus, 1767) in Northern Scandinavia

The repeated occurrence of habitat-specific polyphyletic evolved ecotypes throughout the ranges of widely distributed species implies that multiple, independent and parallel selection events have taken place. Ecological transitions across altitudinal gradients over short geographical distances are often associated with variation in habitat-related fitness, these patterns suggest the action of strong selective forces. Genetic markers will therefore contribute differently to differences between ecotypes in local hybrid zones. Here we have studied the adaptive divergence between ecotypes of the water beetle Agabus bipustulatus along several parallel altitudinal gradients in northern Scandinavia. This water beetle is well known for its remarkable morphological variation associated with mountain regions throughout the western Palaearctic. Two morphological ecotypes are recognised: a montane type with reduced flight muscles and a lowland type with fully developed muscles. Using a multilocus survey of allozyme variation and a morphological analysis with landmark-based morphometrics, across thirty-three populations and seven altitudinal gradients, we studied the local adaptive process of gene flow and selection in detail. Populations were sampled at three different elevations: below, at and above the tree line. The results indicate that the levels of divergence observed between ecotypes in morphology and allele frequencies at α-Glycerophosphate dehydrogenase relative to those shown by neutral molecular markers reflects local diversifying selection in situ. Four main lines of evidence are shown here: (1) A repeated morphological pattern of differentiation is observed across all altitudinal transects, with high reclassification probabilities. (2) Allele and genotype frequencies at the α-Gpdh locus are strongly correlated with altitude, in sharp contrast to the presumable neutral markers. (3) Genetic differentiation is two to three times higher among populations across the tree line than among populations at or below. (4) Genetic differentiation between ecotypes within independent mountain areas is reflected by different sets of allozymes.

Glycolytic and Non-glycolytic Functions of Mycobacterium tuberculosis Fructose-1,6-bisphosphate Aldolase, an Essential Enzyme Produced by Replicating and Non-replicating Bacilli

The search for antituberculosis drugs active against persistent bacilli has led to our interest in metallodependent class II fructose-1,6-bisphosphate aldolase (FBA-tb), a key enzyme of gluconeogenesis absent from mammalian cells. Knock-out experiments at the fba-tb locus indicated that this gene is required for the growth of Mycobacterium tuberculosis on gluconeogenetic substrates and in glucose-containing medium. Surface labeling and enzymatic activity measurements revealed that this enzyme was exported to the cell surface of M. tuberculosis and produced under various axenic growth conditions including oxygen depletion and hence by non-replicating bacilli. Importantly, FBA-tb was also produced in vivo in the lungs of infected guinea pigs and mice. FBA-tb bound human plasmin(ogen) and protected FBA-tb-bound plasmin from regulation by α(2)-antiplasmin, suggestive of an involvement of this enzyme in host/pathogen interactions. The crystal structures of FBA-tb in the native form and in complex with a hydroxamate substrate analog were determined to 2.35- and 1.9-Å resolution, respectively. Whereas inhibitor attachment had no effect on the plasminogen binding activity of FBA-tb, it competed with the natural substrate of the enzyme, fructose 1,6-bisphosphate, and substantiated a previously unknown reaction mechanism associated with metallodependent aldolases involving recruitment of the catalytic zinc ion by the substrate upon active site binding. Altogether, our results highlight the potential of FBA-tb as a novel therapeutic target against both replicating and non-replicating bacilli.

Thyroid hormone contributes to the hypolipidemic effect of polyunsaturated fatty acids from fish oil: in vivo evidence for cross talking mechanisms

n-3 polyunsaturated fatty acids (n-3 PUFA) from fish oil (FO) exert important lipid-lowering effects, an effect also ascribed to thyroid hormones (TH) and TH receptor β1 (TRβ1)-specific agonists. n-3 PUFA effects are mediated by nuclear receptors, such as peroxisome proliferator-activated receptors (PPAR) and others. In this study, we investigated a role for TH signaling in n-3 PUFA effects. Euthyroid and hypothyroid adult rats (methimazole-treated for 5 weeks) received FO or soybean oil (control) by oral administration for 3 weeks. In euthyroid rats, FO treatment reduced serum triglycerides and cholesterol, diminished body fat, and increased protein content of the animals. In addition, FO-treated rats exhibited higher liver expression of TRβ1 and mitochondrial α-glycerophosphate dehydrogenase (mGPD), at protein and mRNA levels, but no alteration of glutathione S-transferase or type 1 deiodinase. In hypothyroid condition, FO induced reduction in serum cholesterol and increase in body protein content, but lost the ability to reduce triglycerides and body fat, and to induce TRβ1 and mGDP expression. FO did not change PPARα liver abundance regardless of thyroid state; however, hypothyroidism led to a marked increase in PPARα liver content but did not alter TRβ1 or TRα expression. The data suggest that part of the effect of n-3 PUFA from FO on lipid metabolism is dependent on TH signaling in specific steps and together with the marked upregulation of PPARα in liver of hypothyroid rats suggest important in vivo consequences of the cross-talking between those fatty acids and TH pathways in liver metabolism.

Glucuronoxylomannan from Cryptococcus neoformans Down-regulates the Enzyme 6-Phosphofructo-1-kinase of Macrophages

The encapsulated yeast Cryptococcus neoformans is the causative agent of cryptococosis, an opportunistic life-threatening infection. C. neoformans is coated by a polysaccharide capsule mainly composed of glucuronoxylomannan (GXM). GXM is considered a key virulence factor of this pathogen. The present work aimed at evaluating the effects of GXM on the key glycolytic enzyme, 6-phosphofructo-1-kinase (PFK). GXM inhibited PFK activity in cultured murine macrophages in both dose- and time-dependent manners, which occurred in parallel to cell viability decrease. The polysaccharide also inhibited purified PFK, promoting a decrease on the enzyme affinity for its substrates. In macrophages GXM and PFK partially co-localized, suggesting that internalized polysaccharide directly may interact with this enzyme. The mechanism of PFK inhibition involved dissociation of tetramers into weakly active dimers, as revealed by fluorescence spectroscopy. Allosteric modulators of the enzyme able to stabilize its tetrameric conformation attenuated the inhibition promoted by GXM. Altogether, our results suggest that the mechanism of GXM-induced cell death involves the inhibition of the glycolytic flux.

Late onset glycogen storage disease Type II with “reducing body”-like inclusions

Skeletal muscle tissue from 3 patients with clinical diagnosis of limb girdle muscular dystrophy revealed a vacuolar myopathy with glycogen storage and lysosomal activity. A diagnosis of late onset GSD Type II was considered. An interesting finding was the presence of round to oval eosinophilic inclusions which reduced on menadione linked a-glycerophosphate dehydrogenase (MAG). There are only two reports in the literature describing similar inclusions in late onset GSD II. We report morphological findings of this rare disorder and compare the findings with earlier two reports.

Eastern grey kangaroo (Macropus giganteus) myofibres. 1. A simplified classification method using two commercially available antibodies

Skeletal muscles from eastern grey kangaroos (Macropus giganteus) were assessed for myofibre contractile and metabolic characteristics using immunocytochemical and histological staining of serial sections. Myofibre classification using monoclonal antibodies that typically bind to mammalian slow (clone WB-MHC), fast (clone MY-32) and Types 1, 2X and 2B (clone S5 8H2) myosin heavy chains was validated using acid- and alkali-preincubated myofibrillar ATPase, NADH and α-glycerophosphate dehydrogenase stains. Myofibres were classified as Type 1 (slow oxidative), Type 2A (fast oxidative-glycolytic), Type 2X/2B (fast glycolytic) or intermediate or transitional myofibre Types 2C (Type 1–Type 2A intermediate) and 2AX/B (Type 2A–Type 2X/2B intermediate). The Type 2 (fast) antibody (clone MY-32) used in the present study did not bind to a subset of fast myofibres in any of the eight kangaroo muscles investigated. These myofibres were identified as Type 2A using clone S5 8H2 and on the basis of the histochemical staining profile. Hence, a simplified immunostaining system using only clones WB-MHC (anti-Type 1) and MY-32 (anti-Type 2X/2B) allowed five myofibre types to be identified based on the staining pattern and intensity of staining for the two antibodies. It is concluded that the myofibres of muscles from kangaroos can be quickly classified into five types using two commercially available antibodies. This method is directly applicable for routine investigations into the myofibre properties of commercially important muscles from the kangaroo musculature and, when combined with enzymatic assays for oxidative and glycolytic activity, will allow for a better understanding of factors influencing the quality of meat from kangaroos.

An electrophoretic characterisation of three paratypes of Arion fagophilus De Winter, 1986, with notes on the subgeneric division of the genus Arion Férussac, 1819 (Mollusca, Pulmonata)

Three paratypes of Arion fagophilus were electrophoretically compared with nine other West European species of the genus Arion. Esterases, glutamate-oxaloacetate-transaminase, a-amylase, superoxide dismutase, phosphoglucomutase, fumarase, α-glycerophosphate dehydrogenase and phosphogluconate dehydrogenase were screened in the hepatopancreas and general proteins in the albumen gland. A. fagophilus appears to be a very well defined species, which may be assigned to the subgenus Kobeltia. The subgeneric division of the genus Arion is preliminary discussed with respect to electrophoretic and chromosomal data. It is concluded to retain four subgenera, which may be grouped two by two: Mesarion and Arion s. s. on one hand, and Kobeltia and Carinarion on the other. It is remarked that the electrophoretic differences between Norwegian and Belgian A. ater suggest a separation into two taxa.

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