ABSTRACT
This study evaluated the plasma membrane integrity, acrosomal membrane integrity, and mitochondrial membrane potential of Nelore bull sperm from early puberty to early sexual maturity and their associations with sperm motility and vigor, the mass motility of the spermatozoa (wave motion), scrotal circumference, and testosterone. Sixty Nelore bulls aged 18 to 19 months were divided into four lots (n=15 bulls/lot) and evaluated over 280 days. Semen samples, collected every 56 days by electroejaculation, were evaluated soon after collection for motility, vigor and wave motion under an optical microscope. Sperm membrane integrity, acrosomal integrity, and mitochondrial activity were evaluated under a fluorescent microscope using probe association (FITC-PSA, PI, JC-1, H342). The sperm were classified into eight integrity categories depending on whether they exhibited intact or damaged membranes, an intact or damaged acrosomal membrane, and high or low mitochondrial potential. The results show that bulls have a low amount of sperm with intact membranes at puberty, and the sperm show low motility, vigor, and wave motion; however, in bulls at early sexual maturity, the integrity of the sperm membrane increased significantly. The rate of sperm membrane damage was negatively correlated with motility, vigor, wave motion, and testosterone in the bulls, and a positive correlation existed between sperm plasma membrane integrity and scrotal circumference. The integrity of the acrosomal membrane was not influenced by puberty. During puberty and into early sexual maturity, bulls show low sperm mitochondrial potential, but when bulls reached sexual maturity, high membrane integrity with high mitochondrial potential was evident.(AU)
Este estudo avaliou a integridade da membrana plasmática, da membrana acrossomal e o potencial da membrana mitocondrial de espermatozoides de touros da raça Nelore da puberdade até à maturidade sexual e suas correlações com motilidade, vigor, turbilhão dos espermatozoides, circunferência escrotal e testosterona. Sessenta touros da raça Nelore, com idade entre 18 e 19 meses, foram divididos em quatro lotes (n=15 touros / lote) e foram avaliados por 280 dias. Coletaram-se as amostras de sêmen a cada 56 dias por eletroejaculação e, logo após, foram determinados a motilidade, o vigor e o turbilhão dos espermatozoides por microscopia óptica. A integridade da membrana de esperma, da membrana acrosomal e da atividade mitocondrial foi avaliada por microscopia de fluorescência, utilizando-se associação das sondas (FITC-PSA, PI, JC-1, H342). Os espermatozoides foram classificados em oito categorias de integridade, dependendo se eles exibiram membranas plasmática e mitocondrial intactas ou danificadas e potencial mitocondrial elevado ou baixo. Os resultados mostram que os touros têm uma baixa quantidade de espermatozoides com membranas íntegras na puberdade, com baixa motilidade, vigor e turbilhão. No entanto, nos touros na maturidade sexual precoce, a integridade da membrana dos espermatozoides aumentou significativamente. A taxa de dano à membrana espermática foi negativamente correlacionada com a motilidade, o vigor, o movimento das ondas e a concentração de testosterona nos touros, e uma correlação positiva existiu entre a integridade da membrana plasmática e a circunferência escrotal. A integridade da membrana acrossomal não foi influenciada pela puberdade.(AU)
Subject(s)
Animals , Cattle , Cell Membrane , Mitochondrial Turnover , Sperm Motility , Testosterone , Puberty/metabolism , Sperm Count/veterinaryABSTRACT
<p><b>BACKGROUND</b>Mitochondrial dysfunction has been reported to be one of the contributing factors of sepsis-associated encephalopathy (SAE). Mitochondrial biogenesis controls mitochondrial homeostasis and responds to changes in cellular energy demand. In addition, it is enhanced or decreased due to mitochondrial dysfunction during SAE. The aim of this study was to explore the changes of mitochondrial biogenesis of astrocytes under septic conditions.</p><p><b>METHODS</b>Lipopolysaccharide (LPS; 50 ng/ml) and interferon-γ (IFN-γ; 200 U/ml) were incubated with astrocytes to model the effects of a septic insult on astrocytes in vitro. The mitochondrial ultrastructure and volume density were evaluated by transmission electron microscopy. Intracellular adenosine triphosphate (ATP) levels were detected by the firefly luciferase system. The expression of protein markers of mitochondrial biogenesis and the binding ability of mitochondrial transcription factor A (TFAM) were determined by western blot and electrophoretic mobility shift assays, respectively. The mitochondrial DNA (mtDNA) content was detected by real-time polymerase chain reaction.</p><p><b>RESULTS</b>The number of mildly damaged mitochondria was found to be significantly greater after treatment for 6 hours, as compared with at 0 hour (P < 0.05). The mitochondrial volume density was significantly elevated at 24 hours, as compared with at 0 hour (P < 0.05). The ATP levels at 6 hours, 12 hours, and 24 hours were significantly greater than those at 0 hour (P < 0.05). The protein markers of mitochondrial biogenesis were significantly increased at 6 hours and 12 hours, as compared with at 0 hour (P < 0.05). The TFAM binding activity was not significantly changed among the four time points analyzed. The mtDNA contents were significantly increased at 12 hours and 24 hours, as compared with at 0 hour (P < 0.05).</p><p><b>CONCLUSIONS</b>Under septic conditions, mitochondrial biogenesis of astrocytes increased to meet the high-energy demand and to promote mitochondrial recovery. Furthermore, the TFAM-DNA binding ability was not sensitive to sepsis-induced injury.</p>
Subject(s)
Animals , Rats , Astrocytes , Metabolism , Blotting, Western , Cells, Cultured , DNA, Mitochondrial , Genetics , Electrophoretic Mobility Shift Assay , Interferon-gamma , Pharmacology , Lipopolysaccharides , Pharmacology , Microscopy, Electron, Transmission , Mitochondrial Turnover , Physiology , Nitric Oxide , Metabolism , Reactive Nitrogen Species , Metabolism , Sepsis , Metabolism , Tumor Necrosis Factor-alpha , MetabolismABSTRACT
FUNDAMENTO: O programa de biogênese mitocondrial no coração parece apresentar remodelação adaptativa após estresse biomecânico e oxidativo. Os mecanismos adaptativos que protegem o metabolismo do miocárdio durante a hipóxia são coordenados, em parte, pelo óxido nítrico (NO). OBJETIVO: Observar a biogênese mitocondrial e expressão do óxido nítrico sintase (NOS) em corações de cardiopatia congênita com cianose; discutir a resposta mitocondrial à hipóxia crônica do miocárdio. MÉTODOS: Foram investigados 20 pacientes com defeitos cardíacos cianóticos (n = 10) ou acianóticos (n = 10). Foram estudadas amostras do miocárdio na via de saída ventricular direita, tomadas durante a operação. A análise morfométrica de mitocôndrias foi realizada por microscopia eletrônica de transmissão. A relação mtDNA/nDNA foi determinada com PCR em tempo real. Os níveis de transcrição da subunidade I da citocromo c oxidase (COXI), coativador-1α do receptor γ ativado por proliferador de peroxissoma (PGC-1α), o fator respiratório nuclear 1 (NRF1), e fator de transcrição mitocondrial A (Tfam) foram detectados por reação em cadeia da polimerase via transcriptase reversa (RT-PCR) ativado por fluorescência em tempo real. Os níveis proteicos de COXI e nNOS, iNOS e eNOS foram medidos por técnica de Western Blot. RESULTADOS: A densidade volumétrica mitocondrial (Vv) e a densidade numérica (Nv) foram significativamente elevadas em pacientes com cianose, em comparação com a cardiopatia congênita acianótica. MtDNA elevada e suprarregulação dos níveis de COXI, PGC-1 α, NRF1 e Tfam mRNA foram observadas em pacientes cianóticos. Os níveis de proteína de COXI e eNOS foram significativamente maiores no miocárdio de pacientes cianóticos que nos de acianóticos. Os níveis de transcrição do PGC-1α se correlacionam com os níveis de eNOS. CONCLUSÃO: A biogênese mitocondrial é ativada no miocárdio da via de saída ventricular na cardiopatia congênita com cianose, que ...
BACKGROUND: Mitochondrial biogenesis program in heart appears to exhibit adaptive remodeling following biomechanical and oxidative stress. The adaptive mechanisms that protect myocardium metabolism during hypoxia are coordinated in part by nitric oxide (NO). OBJECTIVE: To observe mitochondrial biogenesis and nitric oxide synthase (NOS) expression in hearts of congenital heart disease with cyanosis, discuss mitochondrial response to chronic hypoxia in myocardium. METHODS: 20 patients with cyanotic (n=10) or acyanotic cardiac defects (n=10) were investigated. Samples from the right ventricular outflow tract myocardium taken during operation were studied. Morphometric analysis of mitochondria was performed with transmission electron microscope. Relative mtDNA/nDNA ratio was determined with real-time PCR. Cytochrome c oxidase subunit I (COXI), peroxisome-proliferator-activated receptor γ coactivator-1α (PGC-1α), nuclear respiratory factor 1 (NRF1), and mitochondrial transcription factor A (Tfam) transcript levels were detected by real-time fluorescent RT-PCR. COXI and nNOS, iNOS and eNOS protein levels were measured with western blot. RESULTS: Mitochondrial volume density (Vv) and numerical density (Nv) were significantly elevated in patients with cyanotic compared to acyanotic congenital heart disease. Elevated mtDNA and up-regulated COXI, PGC-1α, NRF1 and Tfam mRNA levels were observed in cyanotic patients. Protein levels of COXI and eNOS were significantly higher in the myocardium of cyanotic than of acyanotic patients. PGC-1α transcript levels correlated with the levels of eNOS. Conclusion: Mitochondrial biogenesis is activated in right ventricular outflow tract myocardium in congenital heart disease with cyanosis, which could be the adaptive response to chronic hypoxia and possibly involves eNOS up-regulation. (Arq Bras Cardiol. 2012; [online].ahead print, PP.0-0).
Subject(s)
Adolescent , Child , Child, Preschool , Female , Humans , Male , Young Adult , Cyanosis/enzymology , Cyanosis/physiopathology , Heart Defects, Congenital/enzymology , Mitochondrial Turnover/physiology , Myocardium/metabolism , Nitric Oxide Synthase Type III/metabolism , DNA Copy Number Variations , DNA, Mitochondrial/chemistry , Gene Expression Regulation/physiology , Heart Defects, Congenital/physiopathology , Mitochondrial Size , Nitric Oxide Synthase/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Transcription Factors/metabolismABSTRACT
Mitochondrial dysfunction and endoplasmic reticulum (ER) stress are considered the key determinants of insulin resistance. Impaired mitochondrial function in obese animals was shown to induce the ER stress response, resulting in reduced adiponectin synthesis in adipocytes. The expression of inducible nitric oxide synthase (iNOS) is increased in adipose tissues in genetic and dietary models of obesity. In this study, we examined whether activation of iNOS is responsible for palmitate-induced mitochondrial dysfunction, ER stress, and decreased adiponectin synthesis in 3T3L1 adipocytes. As expected, palmitate increased the expression levels of iNOS and ER stress response markers, and decreased mitochondrial contents. Treatment with iNOS inhibitor increased adiponectin synthesis and reversed the palmitate-induced ER stress response. However, the iNOS inhibitor did not affect the palmitate-induced decrease in mitochondrial contents. Chemicals that inhibit mitochondrial function increased iNOS expression and the ER stress response, whereas measures that increase mitochondrial biogenesis (rosiglitazone and adenoviral overexpression of nuclear respiratory factor-1) reversed them. Inhibition of mitochondrial biogenesis prevented the rosiglitazone-induced decrease in iNOS expression and increase in adiponectin synthesis. These results suggest that palmitate-induced mitochondrial dysfunction is the primary event that leads to iNOS induction, ER stress, and decreased adiponectin synthesis in cultured adipocytes.