Changes in the exercise-induced hormone response to branched chain amino acid administration

Summary It was the aim of the present experiment to detect possible effects of branched-chain amino acids (BCAA) on the endocrine response to 1 h of continuous running. Blood samples were collected from 14 long-distance runners (age 24–42 years) in two different trials performed at 1-week intervals. In both trials (E and P) blood samples were collected at the following times: 9 a.m. (basal values sample), 10.30 a.m. (sample 90), 11.30 a.m. (sample 150), 12.30 p.m. (sample 210); the athletes performed 1 h of running at a constant predetermined speed between samples 90 and 150. Following the basal sample a mixture containing BCAA (E trial), or not containing BCAA (P trial) was ingested. In both trials no hormone basal concentrations, except insulin, were changed before exercise. In P trial, following exercise (sample 150), human growth hormone (HGH), prolactin (PRL), adrenocorticotropic hormone (ACTH) and cortisol (C) increased, while testosterone (T) decreased. In sample 210, after 1 h of rest, while ACTH, PRL and HGH had recovered to basal concentrations, C remained elevated and T displayed a further decrease. In the E trial a similar pattern of change was observed in sample 150 for HGH, PRL, ACTH and C; in sample 210 HGH and PRL displayed significantly lower values than in the corresponding P trial samples. The T was not modified by the running exercise and increased during the recovery period. It is, therefore, suggested that BCAA administration before exercise affects the response of some anabolic hormones, mainly HGH and T.     

Modulating effect of Withania somnifera on TCA cycle enzymes and electron transport chain in azoxymethane-induced colon cancer in mice

The aim of the present investigation was to evaluate the efficacy of Withania somnifera on tricarboxylic acid (TCA) cycle enzymes and electron transport chain in azoxymethane-induced experimental colon cancer in mice. Azoxymethane at the dose of 15?mg/kg body weight was induced intraperitoneally once in a week for 28?days. The progression in colon tumor development was correlated with the appearance of the histological biomarker and aberrant crypt foci (ACF). Azoxymethane-induced colon cancer animals were treated with 400?mg/kg body weight of W. somnifera once in a week orally for 28?days. After the experimental period, the animals were killed and analyzed for TCA cycle key enzymes, such as isocitrate dehydrogenase (ICDH), succinate dehydrogenase (SDH), malate dehydrogenase (MDH), and α-keto glutarate dehydrogenase (α-KGDH). The modulating effect of W. somnifera on TCA cycle key enzymes and electron transport chain complexes were investigated against colon cancer induced by azoxymethane in Swiss albino mice. Decreased activities of TCA cycle key enzymes such as ICDH, SDH, MDH, and α-KGDH in colon cancer bearing animals were observed. W. somnifera administration normalized these enzyme levels in azoxymethane-induced experimental mice. These results suggested that W. somnifera is the promising chemotherapeutic agent for the treatment of colon cancer.     

Interaction between 2,6-dimethyl-3,5-dicarbethoxy-1,4-dihydropyridine and enzymes of the NADPH-specific electron transport chain of rat liver microsomes

2,6-Dimethyl-3,5-dicarbethoxy-1,4-dihydropyridine interacts with the NADPH-dependent electron transport system of rat liver microsomes: It forms a type 1 complex with the terminal oxidase (cytochrome P-450) and also definitely inhibits the activity of NADPH-cytochrome c reductase and methindione demethylase. In experiments in vivo repeated administration of the compound had no inducing action on microsoma enzymes.Laboratory of Biochemistry and Pharmacokinetics, Institute of Organic Synthesis, Academy of Sciences of the Latvian SSR, Riga. (Presented by Academician of the Academy of Medical Sciences of the USSR V. V. Zakusov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 83, No. 6, pp. 671–673, June, 1977.     

Evidence for the occurrence of respiratory electron transport in adult Brugia pahangi and Dipetalonema viteae

Mixed-sex adult stages of Brugia pahangi and Dipetalonema viteae, in the absence of exogenous substrate, consumed oxygen at rates of 4.18 +/- 0.38 and 2.12 +/- 0.20 ngatoms O2 min-1 mg-1 dry wt. respectively. When calculated on a unit dry weight basis the endogenous O2 consumption rates (E-QO2) of mature adult male macrofilariae of B. pahangi and D. viteae were significantly greater than those of mature females, although the E-QO2 calculated per individual worm was essentially similar irrespective of sex. When assayed as separate unisexual groups, the oxygen uptake of male and female macrofilariae of both species was inhibited by classical inhibitors of respiratory electron transport (RET), and showed classical substrate bypass phenomena in response to succinate and ascorbate, N,N,N',N'-tetramethyl-p-phenylenediamine with respect to the RET inhibitors rotenone (inhibitor of complex I) and antimycin A (inhibitor of complex III). Since male worms elicited similar responses to the classical RET inhibitors as did mixed-sex and/or adult female populations, the possibility that developmental stages contained within the female filariids were contributing in any significant manner to the overall responses observed with the RET inhibitors can be discounted. Such responses as observed with live-intact macrofilariae are normally elicited only by mitochondrial preparations and suggest that the cuticles of both species are permeable to rotenone, succinate, antimycin A, N,N,N',N'-tetramethyl-p-phenylenediamine, azide and cyanide. The uncoupler 2,4-dinitrophenol stimulated the endogenous rate of oxygen consumption (E-QO2) of intact B. pahangi at 33-160 microM, indicating the probable occurrence of RET-coupled oxidative phosphorylation. Higher concentrations of 2,4-dinitrophenol proved inhibitory. Respiratory studies on subcellular fractions substantiated the responses elicited by the intact parasites, suggesting the presence of antimycin A-sensitive and -insensitive RET pathways capable of utilising alpha-glycerophosphate, succinate, and malate as substrates. Both B. pahangi and D. viteae macrofilariae therefore probably possess branched RET-pathways bifurcating on the substrate side of RET-complex III. The rates of substrate oxidation in terms of QO2 mg-1 mitochondrial protein compare well with those observed with other nematode parasites.     

A hypothesis for a role for unsaturated fatty acids in electron transport and its potential application to understanding the mitochondrial respiratory chain

Unsaturated fatty acids are required for mitochondrial oxidative phosphorylation and electron transport, though their role has not been determined. We have considered the possibility that unsaturated fatty acids might facilitate electron transfer from non-heme iron to heme iron. Unsaturated fatty acids markedly enhanced the reduction of ferric cytochrome c by ferrous iron. Neither stearic acid nor methyl arachidonic acid were effective suggesting that both a double bond and the carboxylic acid group were essential. Thus unsaturated fatty acids can directly facilitate electron transfer from non-heme iron to heme iron. We hypothesize that unsaturated fatty acids may play a similar critical role in mitochondrial oxidative phosphorylation and other biological processes where rapid electron transfer occurs.     

The two mechanisms for antigenic variation in Trypanosoma brucei are independent processes

Antigenic switching in Trypanosoma brucei can occur either by the production of a telomeric copy of a variant surface glycoprotein (VSG) gene through a gene conversion mechanism or by the nonduplicative activation of a telomeric VSG gene. The 5 VSG gene telomeric copy that is expressed in IsTaR 1 variant antigenic type (VAT) 5 is retained in an inactive state following an antigenic switch to VAT A5. This inactive telomeric 5 VSG gene copy is absent following independent single antigenic switches to VATs 1A5 and 11A5. The inactive 5 VSG gene does not appear to have been replaced with the newly expressed VSG gene. Thus, inactive telomeric VSG genes that are capable of being expressed can be lost, presumably through gene conversion to new VSG genes. These results suggest that gene conversion of an inactive VSG gene does not obligately activate the new VSG gene. We conclude that the gene conversion and telomeric activation mechanisms for antigenic switching are separate and independent processes.     

Evidence for axonal transport of RNA in mammalian neurons

Summary Adult albino rabbits were injected into one eye with (³H) uridine. At various time intervals following the injections the amount of RNA-bound and acid soluble radioactivity was measured in the retrobulbar optic pathway. Evidence was obtained for a relatively slow transport of a small amount of macromolecular RNA along the axons of the retinal ganglion cells down to the nerve terminals.     

Mitochondrial electron transport chain dysfunction during development does not extend lifespan in Drosophila melanogaster

Since the initial identification of reactive oxygen species (ROS) as the major factor in aging, many studies have provided evidence for the central role of mitochondria in longevity. A few years ago, an unexpected finding showed that the inactivation of the mitochondrial respiratory chain (MRC) in Caenorhabditis elegans, during the developmental stages only, extended lifespan. Activation of this mitochondrial pathway affecting aging (MIT) is associated with several phenotypic features: increased longevity, increased time of development, decreased fertility/fecundity and reduced adult size. Here, we investigated this pathway in another model organism, Drosophila melanogaster. To assess the role of mitochondrial activity in the Drosophila aging process, we partially inactivated the MRC using RNA interference (RNAi) during larval stages. Developmental perturbation of the respiratory process prolonged development, increased lethality during developmental stage, reduced both fecundity and fertility and slightly reduced individual weight. However, in contrast to the nematode, this genetic intervention either shortened or had no effect on lifespan, depending on the level of gene inactivation. Thus, the effects of MRC disruption during development on aging differ between species. We discuss the possible origins of such differences.     

HLA-DM can partially replace the invariant chain for HLA-DR transport and surface expression in transfected endocrine epithelial cells

The function of HLA class II molecules as peptide presenters to CD4+ T cells depends on the expression of associated molecules such as the invariant chain (Ii) and DM responsible for the correct transport of and high-stability peptide binding to the class II dimers. In organs affected by autoimmune diseases, endocrine epithelial cells express class II molecules, which presumably are involved in the presentation of self-peptides to autoreactive T cells. We have transfected the rat insulinoma cell line RINm5F with different combinations of HLA-DR, Ii and HLA-DM cDNAs and have studied how Ii and DM affect the transport and stability of class II molecules expressed by the different transfectants. Immunofluorescence and biochemical analysis showed that cells transfected with DR and DM in the absence of Ii expressed mostly stable molecules in their surface, and showed a lower accumulation of DR molecules in the endoplasmic reticulum (ER) than cells expressing only DR. This suggests that, in the absence of invariant chain, DM molecules can not only exchange peptides other than class II-associated invariant chain peptide (CLIP) but may also be involved in the transport of class II molecules out of the ER towards the endosomal route. In addition, these data confirm that expression of DR alone or DR+Ii do not allow the formation of sodium dodecyl sulphate (SDS)-stable complexes, that cells expressing DR+Ii have most DR molecules occupied by CLIP and that Ii and DM molecules allow regular routing and peptide loading of class II molecules.     

ATP in cholinergic nerves: Evidence for the axonal transport of a stable pool

Summary The ATP content of the cholinergic electromotor nerves of Torpedo marmorata has been measured. After freezing and thawing of pieces of nerve, their ATP content dropped rapidly to about 5 % of that of pieces extracted immediately. The amount of ATP in nerves incubated in Torpedo Ringer remains at a high level over a long period. In nerves ligated for seven days there is no accumulation of total ATP above the ligature. If, however, the ligated nerves are frozen and thawed for 20 min before extraction, the residual ATP shows significant proximal accumulation at the ligature. This transport is not due to the accumulation of mitochondria at the ligature. Acetylcholine, measured in the same nerve extracts of nerve segments, is also axonally transported.The ATP which is resistant to hydrolysis is probably sequestered within a subcellular organelle. The possibility that its accumulation at ligatures may indicate movement of cholinergic synaptic vesicles which are known to contain ATP and acetylcholine is considered.     

牛磺酸拮抗同型半胱氨酸诱导的线粒体呼吸链自由基生成和同型半胱氨酸抑制线粒体牛磺酸转运体

目的:观察同型半胱氨酸(Hcy)对心肌线粒体电子漏及自由基生成的影响,以及观察牛磺酸的拮抗效应。方法: 分离大鼠心肌线粒体,超声波破碎线粒体制备亚线粒体,纯化制备猪心肌线粒体琥珀酸细胞色素C还原酶(SCR)重组体。分别用Hcy和/或牛磺酸共同孵育心肌线粒体、亚线粒体、SCR;用化学发光法测定H₂O₂^- 及O₂^- 生成;并用滤膜抽滤法观察线粒体膜牛磺酸转运体的性质及Hcy对牛磺酸转运功能的影响。结果: Hcy呈浓度依赖性地刺激大鼠心肌线粒体、亚线粒体氧自由基生成及SCR电子漏增加,牛磺酸自身不影响心肌线粒体、亚线粒体及SCR氧自由基生成,但呈浓度依赖性地抑制Hcy诱导的线粒体、亚线粒体及呼吸链重组体氧自由基生成。线粒体膜上存在Na+依赖性的牛磺酸转运体,Hcy呈浓度依赖性地抑制牛磺酸转运体对牛磺酸的转运功能。结论: 牛磺酸抑制Hcy刺激的线粒体呼吸链电子漏增加及氧自由基生成。线粒体膜上存在Na+依赖性的牛磺酸转运体,Hcy抑制线粒体膜上牛磺酸转运体对牛磺酸的转运功能。     

Mitochondrial energetic impairment in a patient with late‐onset glutaric acidemia Type 2

Glutaric acidemia type 2 (GA2), also called multiple acyl‐CoA dehydrogenase deficiency, is an autosomal recessive disorder of fatty acid, amino acid, and choline metabolism resulting in excretion of multiple organic acids and glycine conjugates as well as elevation of various plasma acylcarnitine species (C4–C18). It is caused by mutations in the ETFA, ETFB, or ETFDH genes which are involved in the transfer of electrons from 11 flavin‐containing dehydrogenases to Coenzyme Q₁₀ (CoQ₁₀) of the mitochondrial electron transport chain (ETC). We report a patient who was originally reported as the first case with primary myopathic CoQ₁₀ deficiency when he presented at 11.5 years with exercise intolerance and myopathy that improved after treatment with ubiquinone and carnitine. At age 23, his symptoms relapsed despite increasing doses of ubiquinone and he was shown to have biallelic mutations in the ETFDH gene. Treatment with riboflavin was started and ubiquinone was changed to ubiquinol. After 4 months, the patient recovered his muscle strength with normalization of laboratory exams and exercise tolerance. Functional studies on fibroblasts revealed decreased levels of ETFDH as well as of very long‐chain acyl‐CoA dehydrogenase and trifunctional protein α. In addition, the mitochondrial mass was decreased, with increased formation of reactive oxygen species and oxygen consumption rate, but with a decreased spared respiratory capacity, and decreased adenosine triphosphate level. These findings of widespread dysfunction of fatty acid oxidation and ETC enzymes support the impairment of a larger mitochondrial ETC supercomplex in our patient.     

Evidence for a branched pathway in the polarized cell division of Saccharomyces cerevisiae

Cells of Saccharomyces cerevisiae can choose a bud site in one of two different spatial patterns (axial or bipolar) determined by their mating type. Genes important for bud-site selection have been identified and a linear model describing the hierarchy of these genes was proposed. We have uncovered a new class of genes which is required only for the bipolar pattern. The phenotype of the corresponding mutants coupled with epistasis experiments with some budding mutants already described suggest the existence of specific genes for the bipolar pathway.     

Evidence for a perforin-mediated mechanism controlling cardiac inflammation in Trypanosoma cruzi infection

CD8+ T lymphocytes are considered an important cell population involved in the control of parasitaemia and mortality after Trypanosoma cruzi infection. However, despite recent developments in this field, the mechanism whereby this control is exerted is still not completely understood. Here we have used perforin knockout (-/-) mice infected with Y strain T. cruzi in order to evaluate specifically the participation of the perforin-based cytotoxic pathway in the destruction of cardiomyocytes, cellular inflammatory infiltration, and control of parasitaemia and mortality. We observed that although parasitaemia was equivalent in perforin (+/+) and (-/-) groups, survival rate and spontaneous physical performance were significantly lower in the perforin deficient mice. The cardiac inflammatory cell infiltration, mostly composed of CD8+ cells, was more evident in perforin (-/-) mice. Ultrastructural and immunofluorescence analysis, as well as plasma creatine kinase activity, revealed cardiomyocyte damage and necrosis, more evident in perforin (-/-) mice. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) assays performed in heart samples revealed similar and modest levels of apoptosis in both perforin (+/+) and (-/-) mice. These results indicate that perforin does not play a pivotal role in the control of parasitaemia and direct lysis of cardiomyocytes, but seems to be an important molecule involved in the control of cardiac inflammation and pathology induced by a highly virulent strain of T. cruzi.     

Pairing of VH gene families with the λ light chain: Evidence for a non-stochastic association

The frequencies at which four V_H gene families pair with the λ1 light (L) chain were determined by sequential hybridization of V_H- and λ1-specific DNA probes to mitogen-induced colonies of B cells. Analysis of pair frequencies indicates that the repertoire of λ L chain antibodies is generated by the stochastic pairing of smaller 3′-to-mid-locusV_H gene families (X-24, S107, Q52). However, the large 5′ V_H J558 family appeared to associate with the λ1L chain non-stochastically; the frequency of VhJ558/λ1⁺ colonies among all λ1⁺ colonies was significantly lower than the frequency of J558 expression among all (Cμ⁺) B cell colonies. This difference suggests that selection, either intrinsic at the level of rearrangement or heavy and L chain pairing, or extrinsic following surface immunoglobulin expression, may operate to shape the λ antibody repertoire prior to the introduction of exogenous antigen.     

A rapid method for the isolation of intact glycosomes from Trypanosoma brucei by Percoll -gradient centrifugation in a vertical rotor

A rapid method for the isolation of intact glycosomes from trypanosoma brucei is described. A post large-granule extract is subjected to density-gradient centrifugation on isotonic Percoll in a vertical rotor. This results in a good separation of the glycosomes, which equilibrated at 1.09 g/cm3, from other cellular components. Glycosomes isolated from such a gradient are 9-fold purified relative to the starting homogenate with a yield of 25%. Their glycolytic enzymes exhibit a high latency indicative of an intact glycosomal membrane.     

Evidence for selective axon-terminal uptake and retrograde transport of label in cortico- and rubrospinal systems after injection of 3H-proline

Summary In tracing the macaque's ascending spinal connections by anterograde axonal flow, perikaryal labelling was also observed within the n. ruber pars magnocellularis as well as in the sensorimotor cortex. While neuropil labelling was demonstrated after single injections of leucine or proline, labelling of perikarya was only evident following proline injections. Perikaryal labelling is most likely to be due to uptake of label by axons and/or axons terminals within the injection field and subsequent retrograde axonal transport of radioactive compound to the parent cell bodies. Advantages and disadvantages of the uptake of label by axon terminals are discussed in relation to the use of ³H-proline for tracing neuronal connections. The selectivity of this event and its relationship to the suggested inhibitory functional role of proline and its metabolites is not clear and needs further investigation.     

The use of lectin transport in the mouse central nervous system as an anterograde axonal marker for electron microscopy

Lesion-induced axonal degeneration and autoradiography-electron microscopy have been the only reliable anterograde axonal markers available for electron microscopic examination of neuronal circuitry. However, these methods have their limitations. Recently, Phaseolus vulgaris-leucoagglutinin (PHA-L) has been used as an anterograde axonal marker for light microscopy. This report describes the use of this lectin as an anterograde marker for electron microscopy. PHA-L was injected into mouse SmI cortex or ventrobasal thalamus. Using standard immunohistochemical techniques, the transported lectin was tagged with antibody, which was then visualized with avidin-biotin-horseradish peroxidase binding. Light microscopy demonstrated anterograde transport to predicted cortical regions. With the electron microscope, labeled axon terminals were seen forming asymmetric synapses with spines, dendrites and cell bodies.     

Evidence for an anterograde transport of the fluorescent tracer Fast Blue in the strio-nigro-striatal loop of the rat

The report is concerned with serendipitous observations which were made in the course of investigations on the connectivity of the claustrum. Stereotaxical injections of Fast Blue were made in the neostriatum. All major sources of striopetal fiber systems were found to contain fluorescent cell bodies. Quite unexpectedly, also fluorescent axons and terminals could be observed in brain regions known to receive a striofugal input including the globus pallidus, entopeduncular and subthalamic nucleus, and the substantia nigra, zona reticulata. It is concluded that Fast Blue may be used for both retrograde and anterograde neuronal tracing.