Embryo development is impaired by sperm mitochondrial-derived ROS

BACKGROUND: Basal energetic metabolism in sperm, particularly oxidative phosphorylation, is known to condition not only their oocyte fertilising ability, but also the subsequent embryo development. While the molecular pathways underlying these events still need to be elucidated, reactive oxygen species (ROS) could have a relevant role. We, therefore, aimed to describe the mechanisms through which mitochondrial activity can influence the first stages of embryo development. RESULTS: We first show that embryo development is tightly influenced by both intracellular ROS and mitochondrial activity. In addition, we depict that the inhibition of mitochondrial activity dramatically decreases intracellular ROS levels. Finally, we also demonstrate that the inhibition of mitochondrial respiration positively influences sperm DNA integrity, most likely because of the depletion of intracellular ROS formation. CONCLUSION: Collectively, the data presented in this work reveals that impairment of early embryo development may result from the accumulation of sperm DNA damage caused by mitochondrial-derived ROS.

Allergens of the urushiol family promote mitochondrial dysfunction by inhibiting the electron transport at the level of cytochromes b and chemically modify cytochrome c1

BACKGROUND: Urushiols are pro-electrophilic haptens that cause severe contact dermatitis mediated by CD8+ effector T-cells and downregulated by CD4+ T-cells. However, the molecular mechanism by which urushiols stimulate innate immunity in the initial stages of this allergic reaction is poorly understood. Here we explore the sub-cellular mechanisms by which urushiols initiate the allergic response. RESULTS: Electron microscopy observations of mouse ears exposed to litreol (3-n-pentadecyl-10-enyl-catechol]) showed keratinocytes containing swollen mitochondria with round electron-dense inclusion bodies in the matrix. Biochemical analyses of sub-mitochondrial fractions revealed an inhibitory effect of urushiols on electron flow through the mitochondrial respiratory chain, which requires both the aliphatic and catecholic moieties of these allergens. Moreover, urushiols extracted from poison ivy/oak (mixtures of 3-n-pentadecyl-8,11,13 enyl/3-n-heptadecyl-8,11 enyl catechol) exerted a higher inhibitory effect on mitochondrial respiration than did pentadecyl catechol or litreol, indicating that the higher number of unsaturations in the aliphatic chain, stronger the allergenicity of urushiols. Furthermore, the analysis of radioactive proteins isolated from mitochondria incubated with 3H-litreol, indicated that this urushiol was bound to cytochrome c1. According to the proximity of cytochromes c1 and b, functional evidence indicated the site of electron flow inhibition was within complex III, in between cytochromes bL (cyt b566) and bH (cyt b562). CONCLUSION: Our data provide functional and molecular evidence indicating that the interruption of the mitochondrial electron transport chain constitutes an important mechanism by which urushiols initiates the allergic response. Thus, mitochondria may constitute a source of cellular targets for generating neoantigens involved in the T-cell mediated allergy induced by urushiols.

Ultra-short-course and intermittent TB47-containing oral regimens produce stable cure against Buruli ulcer in a murine model and prevent the emergence of resistance for

Buruli ulcer (BU), caused by

Research journey of respirasome

Respirasome, as a vital part of the oxidative phosphorylation system, undertakes the task of transferring electrons from the electron donors to oxygen and produces a proton concentration gradient across the inner mitochondrial membrane through the coupled translocation of protons. Copious research has been carried out on this lynchpin of respiration. From the discovery of individual respiratory complexes to the report of the high-resolution structure of mammalian respiratory supercomplex IIIIIV, scientists have gradually uncovered the mysterious veil of the electron transport chain (ETC). With the discovery of the mammalian respiratory mega complex IIIIIV, a new perspective emerges in the research field of the ETC. Behind these advances glitters the light of the revolution in both theory and technology. Here, we give a short review about how scientists 'see' the structure and the mechanism of respirasome from the macroscopic scale to the atomic scale during the past decades.

Research journey of respirasome

Respirasome, as a vital part of the oxidative phosphorylation system, undertakes the task of transferring electrons from the electron donors to oxygen and produces a proton concentration gradient across the inner mitochondrial membrane through the coupled translocation of protons. Copious research has been carried out on this lynchpin of respiration. From the discovery of individual respiratory complexes to the report of the high-resolution structure of mammalian respiratory supercomplex IIIIIV, scientists have gradually uncovered the mysterious veil of the electron transport chain (ETC). With the discovery of the mammalian respiratory mega complex IIIIIV, a new perspective emerges in the research field of the ETC. Behind these advances glitters the light of the revolution in both theory and technology. Here, we give a short review about how scientists 'see' the structure and the mechanism of respirasome from the macroscopic scale to the atomic scale during the past decades.

Insulin resistance: is it a feedback mechanism or a therapeutic target in type 2 diabetes?

Insulin resistance is an essential pathological condition leading to hyperglycemia of blood, and hence insulin resistance is considered as a therapeutic target. The experimental evidence of studying the effect of naphthalene on Anadara granosa, a bivalve, indicated mitochondrial dysfunction and osmolarity change in them. This communication tries to suggest whether insulin resistance is a feedback mechanism to protect the influx of nutrients into the cells demanding osmoregulation of cells.

A case of imported furuncular cutaneous myiasis and cytochrome oxidase Ⅰ gene sequence analysis of the pathogenic Cordylobia anthropophaga / 中华皮肤科杂志

To report a case of imported furuncular cutaneous myiasis,and to analyze the sequence of the mitochondrial cytochrome C oxidase subunit Ⅰ (CO Ⅰ) gene of the pathogenic Cordylobia anthropophaga.A 33-year-old female patient had a travel history to Ghana and Cameroon in Africa 1 month prior to the presentation.No anti-mosquito measures were taken during her stay,and she hung up the laundries outside to dry for several times.Skin examination showed furuncular protuberances with diameters of 1-2 cm on the inner side of the left upper arm as well as on the outer side of the left chest,which were bright red and hard on palpation with irregular borders and a small hole on their central surface.Morphological identification revealed that the larva squeezed from the lesion was suspected as myiasis.After PCR amplification of the CO Ⅰ gene of the larva,an about 650-bp PCR product was acquired.Sequencing and BLAST analysis showed that this product was most closely related to the CO Ⅰ gene (GenBank accession number:FR719158.1) of Cordylobia anthropophaga isolated in Cameroon in 2010 with the sequence similarity being 99.84％,and they were grouped together on the phylogenetic tree.According to the clinical features and travel history of the patient and the sequencing results of the pathogenic Cordylobia anthropophaga,this case was confirmed as imported furuncular cutaneous myiasis caused by Cordylobia anthropophaga.

Cancer Energy Metabolism: Shutting Power off Cancer Factory

In 1923, Dr. Warburg had observed that tumors acidified the Ringer solution when 13 mM glucose was added, which was identified as being due to lactate. When glucose is the only source of nutrient, it can serve for both biosynthesis and energy production. However, a series of studies revealed that the cancer cell consumes glucose for biosynthesis through fermentation, not for energy supply, under physiological conditions. Recently, a new observation was made that there is a metabolic symbiosis in which glycolytic and oxidative tumor cells mutually regulate their energy metabolism. Hypoxic cancer cells use glucose for glycolytic metabolism and release lactate which is used by oxygenated cancer cells. This study challenged the Warburg effect, because Warburg claimed that fermentation by irreversible damaging of mitochondria is a fundamental cause of cancer. However, recent studies revealed that mitochondria in cancer cell show active function of oxidative phosphorylation although TCA cycle is stalled. It was also shown that blocking cytosolic NADH production by aldehyde dehydrogenase inhibition, combined with oxidative phosphorylation inhibition, resulted in up to 80% decrease of ATP production, which resulted in a significant regression of tumor growth in the NSCLC model. This suggests a new theory that NADH production in the cytosol plays a key role of ATP production through the mitochondrial electron transport chain in cancer cells, while NADH production is mostly occupied inside mitochondria in normal cells.

Desempenho fotossintético de folhas jovens e maduras de Vochysia cinnamomea (Vochysiaceae) em áreas de cerrado rupestre intactas e pós-queimada / Photosynthetic performance of young and mature leaves of Vochysia cinnamomea (Vochysiaceae) at intact and after fire areas of cerrado rupestre

O fogo exerce um papel importante como modelador dos ecossistemas de diversas formações vegetais, especialmente o Cerrado. Sua ação induz a brotação em diversas espécies, muitas vezes formando folhas mais vigorosas e fisiologicamente mais ativas. Nesse contexto, o presente trabalho investigou o desempenho fotossintético de folhas jovens e maduras de Vochysia cinnamomea Pohl (Vochysiaceae) em resposta à ação do fogo natural sobre uma área de cerrado rupestre da Serra da Canastra, Minas Gerais, comparando-se; (1) folhas jovens de área queimada e não queimada e (2) folhas jovens e maduras de um mesmo indivíduo. Foram analisadas folhas maduras de indivíduos de área não queimada cuja brotação ocorreu anterior à queimada, e folhas jovens de indivíduos de áreas queimadas e não queimadas cuja brotação ocorreu após a passagem do fogo. Foram coletados e analisados dados relativos às taxas fotossintéticas, rendimento quântico potencial e efetivo, taxa relativa de transporte de elétrons, assimilação líquida de CO2, condutância estomática, transpiração e teores de clorofila. Não houve diferenças significativas entre as folhas jovens da área queimada e da área não queimada em relação aos processos fisiológicos testados. Desta forma não há nenhum tipo de resposta rápida em relação à passagem do fogo. Entretanto, foram encontradas diferenças significativas quando comparadas folhas jovens e maduras. É fato que folhas jovens de V. cinnamomea são estruturalmente distintas de folhas maduras, são mais suculentas e pilosas enquanto as folhas maduras são mais coriáceas, glabras e com cutícula evidente. Estas diferenças estruturais e os estágios fisiológicos de maturação distintos se refletem nas características fisiológicas estudadas nestas folhas que, quando maduras apresentam-se com menor grau de fotoinibição, maior teor de clorofilas e maior assimilação líquida de CO2.

Post-fire effect plays an important role as a modulator of plant ecosystems, especially of the Cerrado. It induces leaf sprouting in several species, often forming vigorous and more physiologically active leaves. In the present study it was investigated the photosynthetic performance of young and mature leaves of Vochysia cinnamomea Pohl (Vochysiaceae) in response to the fire action in a "cerrado rupestre" area of Serra da Canasta, Minas Gerais, comparing; (1) young leaves from fired and intact area, and (2) young and mature leaves from the same plant. Mature leaves from intact area plants which leaf sprouting occurred before fire action in the adjacent area, and young leaves from intact and fired area which leaf sprouting occurred after fire action were analyzed. Data were collected and analyzed on photosynthetic rates, potential and effective quantum yield, electron transport rate, CO2 assimilation, stomatal conductance, transpiration and chlorophyll contents. There were no significant differences between the young leaves of the burned and unburned area in relation to physiological parameters tested. Thus there is no kind of quick response in relation to fire effects. However, significant differences between young and mature leaves were found. Young leaves are structurally distinct of the mature leaves, been more succulent and pilous while mature leaves are more coriaceous, glabrous and with thicked cuticle. The structural differences between young and mature leaves are reflected in distinct physiological performance. Mature leaves present lower degree of photoinhibition, higher pigment content and CO2 assimilation.

Detection of Mitochondrial Reactive Oxygen Species in Living Rat Trigeminal Caudal Neurons

Growing evidence suggests that mitochondrial reactive oxygen species (ROS) are involved in various pain states. This study was performed to investigate whether ROS-induced changes in neuronal excitability in trigeminal subnucleus caudalis are related to ROS generation in mitochondria. Confocal scanning laser microscopy was used to measure ROS-induced fluorescence intensity in live rat trigeminal caudalis slices. The ROS level increased during the perfusion of malate, a mitochondrial substrate, after loading of 2',7'-dichlorofluorescin diacetate (H2DCF-DA), an indicator of the intracellular ROS; the ROS level recovered to the control condition after washout. When pre-treated with phenyl N-tert-butylnitrone (PBN) and 4-hydroxy-2,2,6,6-tetramethylpiperidene-1-oxyl (TEMPOL), malate-induced increase of ROS level was suppressed. To identify the direct relation between elevated ROS levels and mitochondria, we applied the malate after double-loading of H2DCF-DA and chloromethyl-X-rosamine (CMXRos; MitoTracker Red), which is a mitochondria-specific fluorescent probe. As a result, increase of both intracellular ROS and mitochondrial ROS were observed simultaneously. This study demonstrated that elevated ROS in trigeminal subnucleus caudalis neuron can be induced through mitochondrial-ROS pathway, primarily by the leakage of ROS from the mitochondrial electron transport chain.

Novo papel da proteína XPC na regulação dos complexos da cadeia de transporte de elétrons e desequilíbrio redox / New role of XPC protein in regulating the electron transport chain complexes and redox unbalance

Espécies reativas de oxigênio (EROs) são normalmente e continuamente geradas em mitocôndrias, majoritariamente na cadeia de transporte de elétrons (CTE). Harman (1956, 1972 e 1992) teorizou que os radicais livres gerados nas mitocôndrias seriam a principal causa do envelhecimento. De fato, durante o envelhecimento é observado um desequilíbrio entre formação e remoção de EROs, que resulta em estresse redox. Essa condição favorece a formação de lesões oxidadas no DNA, acarretando em mutagênese ou morte celular. Diversos mecanismos moleculares cooperam para o reparo de DNA. Duas vias de reparo de DNA lidam com a maioria das lesões: o reparo por excisão de base (BER) e o reparo por excisão de nucleotídeos (NER). A via BER corrige pequenas modificações de bases que surgem de reações de desaminação, alquilação e oxidação. A via NER é mais versátil, reconhecendo lesões que distorcem a dupla hélice de DNA, como danos induzidos por luz UV e adutos volumos. Pacientes xeroderma pigmentoso (XP-A a XP-G) herdam mutações em um de sete genes que codificam proteínas envolvidas na via NER, ou em um gene que codifica uma polimerase translesão (XP-V). A doença é caracterizada por fotosensibilidade e incidência elevada de neoplasias cutâneas. A proteína XPC atua na etapa de reconhecimento da lesão de DNA na subvia de reparo global do genoma (GG-NER), e sua mutação dá origem aos sintomas clássicos de XP. Novas funções de XPC foram recentemente descritas: i) atuando como cofator na via BER auxiliando as DNA glicosilases OGG1, TDG e SMUG; ii) atuando como cofator transcricional de elementos responsivos a Oct4/Sox2, RXR e PPARα; e iii) na adaptação metabólica na transformação de queratinócitos. Então, propusemo-nos a investigar as relações entre XPC e a manutenção da integridade do DNA mitocondrial, a sensibilidade celular a estresse redox mitocondrial e possíveis alterações bioenergéticas e redox. Para tal, padronizamos um ensaio in vitro de cinética de incisão em DNA plasmidial a fim de investigarmos o possível papel de XPC no reparo de lesões oxidadas em mtDNA. Porém, nossos dados revelaram que XPC não se encontra em mitocôndrias. Apesar disso, células XP-C são mais sensíveis ao tratamento com azul de metileno (AM), antimicina A (AA) e rotenona (ROT), que geram estresse redox mitocondrial. A sensibilidade à AA foi completamente revertida em células corrigidas. Células XP-C apresentaram alterações quanto ao uso dos complexos mitocondriais, com diminuição da taxa de consumo de oxigênio (OCR) via complexo I e um aumento da OCR via complexo II, dependente da presença de XPC. Ademais, a linhagem XP-C apresentou um desequilíbrio redox mitocondrial com maior produção de EROs e menor atividade de GPx. O DNA mitocondrial de células XP-C apresentou níveis elevados de lesão e deleção, que no entanto não retornaram aos níveis encontrados em células selvagens na linhagem XP-C corrigida. Observamos uma acentuada diminuição da expressão de PPARGC1A, um importante regulador de biogênese mitocondrial. Contudo, não foi possível determinar o mecanismo de supressão da expressão de PPARGC1A. Por fim, identificamos que o tipo de mutação em XPC pode estar associado a expressão de PPARGC1A. Esse estudo abre novas possibilidade na investigação do papel de proteína XPC, à parte da instabilidade genômica, na adaptação metabólica e desequilíbrio redox em direção da progressão tumoral

Mitochondria continuously produce reactive oxygen species (ROS), mainly at the electron transport chain. Harman (1956, 1972 e 1992) proposed that normal aging is driven by increased mitochondrially generated free radicals. Indeed, during the course of aging there is an increased imbalance between formation and removal of ROS, leading to redox stress. This condition favours the formation of oxidized DNA lesions, given rise to mutations and cell death. Several molecular mechanisms cooperates to repair the DNA. Two DNA repair pathways deal with the majority of lesions: base excision repair (BER) and nucleotide excision repair (NER). The BER pathway corrects small base modifications that arise from deamination, alkylation and oxidation reactions. The NER pathway is more versitile, recognizing helix-distorting lesions, such as UV-induced damage and bulky adducts. Xeroderma pigmentosum (XP-A to XP-G) patients inherit mutations in one of seven protein-coding genes involved in NER pathway, or in a gene coding a translesion DNA polymerase (XP-V). Photosensitivity and a thousand-fold increased in the risk of developing cutaneous neoplasms are the main clinical features of XP. XPC protein functions in the recognition step of global genome NER (GG-NER) sub-pathway, and mutations in this gene lead to classical XP symptoms. Recently, it has been described that XPC acts: i) as a cofactor in BER pathway through functional interaction with DNA glycosylases OGG1, TDG and SMUG1; ii) as coactivator in transcription at Oct4/Sox2, RXR and PPARα responsive elements; iii) in metabolic shift during keratinocytes transformation. Thus, we sought to investigate a possible role for XPC in the maintenance of mtDNA integrity, cellular sensitivity to mitochondrial redox stress and eventual bioenergetic and redox changes. For this purpose, we established an in vitro plasmid incision assay to investigate the possible role of XPC in the repair of oxidized lesions in mitochondrial DNA. However, our data revealed that XPC did not localized in mitochondria. Nonetheless, XP-C cells are more sensitive to methylene blue, antimycin A (AA) and rotenone treatment, which induce mitochondrial redox stress. The XP-C sensitivity to AA was completely reverted in XPC-corrected cells. XP-C cells presented altered usage of mitochondrial complexes, with decreased oxygen consumption rate (OCR) via complex I and increased OCR through complex II, an XPC-dependent phenomenon. Furthermore, the XP-C cell line showed mitochondrial redox imbalance with increased ROS production and decrease GPx activity. MtDNA from XP-C cells accumulate lesions and deletions, which, however, were found at similar levels in the corrected cell line. We identified a sharp decrease in the expression of PPARGC1A, a master regulator of mitochondrial biogenesis. Nevertheless, it was not possible to determine the mechanism of suppression of PPARGC1A expression. Finally, our results suggest a possible link between the type of XPC mutation and PPARGC1A expression. This study unfolds new possible roles for XPC, aside from its established roles in genomic instability, in metabolic adaptation and redox imbalance towards tumour progression

Influence of ubiquinol-cytochrome C reductase core protein 1 on hypoxia/reoxygenation injury in cultured H9c2 cardiac myocytes / 重庆医学

Objective To construct the recombinant adenovirus vector containing ubiquinol-cytochrome C reductase core protein 1(UQCRC1) and to investigate the protective role of UQCRC1 against hypoxia/reoxygenation injury in H9c2 cardiac myocytes . Methods UQCRC1 gene was obtained from the cDNA library by PCR ,then was double-digested with restriction endonucleases SalⅠand XbaⅠand inserted into pAd Track-CMV .The identified plasmid of pAd Track-UQCRC1 was transfected into BJ 5183 contai-ning pAdEasy-1 .After screening the positive clone ,the plasmid was transfect into 293T cells with liposome to integrate and package the recombinant adenovirus .Finally ,these adenoviruses were transfected into H9c2 cardiac myocytes .The expressions of green fluo-rescence protein(GFP) ,UQCRC1 gene and protein were observed by RT-PCR and Western blot analysis .The cell viability and the LDH release were detect .Results The recombinant adenovirus-UQCRC1 was constructed successfully .The overexpression of UQCRC1 increased the cell viability(P<0 .05) and decreased the LDH release(P<0 .05) from H9C2 cardiac myocytes after suf-fering hypoxia/reoxygenation injury .Conclusion UQCRC1 has the protective effect on hypoxia/reoxygenation injury in H9c2 car-diac myocytes ,and the construction of recombinant adenovirus vector will lay the foundation for further studying the role of UQCRC1 in cardioprotection .

Influence of Electroacupuncture on COX Activity of Hippocampal Mitochondria in Senescence- accelerated Mouse Prone 8 Mice / 针灸推拿医学（英文版）

Objective: To observe the effect of electroacupuncture (EA) on cytochrome c oxidase (COX)activity of hippocampal mitochondria in senescence-accelerated mouse prone 8 (SAMP8) mice, and to explore the EA mechanism on Alzheimer disease (AD) in improving energy metabolic disorder. Methods: Twelve SAMP8 mice were randomly divided into a model group and an EA group, with six in each group. Six senescence-accelerated mouse resistance 1 (SAMR1) mice were prepared as blank group. Mice in the EA group received EA on Baihui (GV 20) and Yongquan (KI 1), once a day for 7 d as a course, altogether 3 courses with one day intervalbetween two courses. Mice in the model group and the blank group were manipulated and fixed as those in the EA group. After interventions, Morris water maze was employed to test spatial learning and memory ability to evaluate EA effect; spectrophotometry was used to detect the activity of hippocampal mitochondria COX. Results: Compared with the blank group, mean escape latenciesof the EA group and model group were prolonged significantly in Morris water maze tests (P Conclusion: It’s plausible that EA improves AD learning and memory ability by increasing mitochondria COX activity, protecting the structure and function, and improving energy metabolism.

The inhibitory effects of rotenone, 2, 4-dinitrophenol, oligomycin on the electron proton flow through “The membrane redox potential three state line” / Монголын Анагаах Ухаан

Rotenone is a specific inhibitor of the NADH dehydrogenase complex. In mitochondria, rotenone inhibitsthe oxidation of NADH to NAD, thereby blocking the oxidation of NAD and the substrates such asglutamate, alpha-ketoglutarate, and pyruvate. Rotenone also inhibits the mitochondrial respiratory chainbetween diphosphopyridine nucleotide and flavine.2, 4-Dinitrophenol – (DNP) is lipophilic weak acids that pick up a proton, transport across the mitochondrialinner membrane into the matrix, deprotonate, then exit as anions before repeating the catalytic cycle,and dissipating the proton gradient. In this situation, electrons continue to pass through the electrontransport system, reduce oxygen to water and metabolic rate, heat are increased, but ATP is lesssynthesized in this process.The macrolide antibiotic - oligomycin binds to the surface of the c8-10 ring of the Fo domain of ATPsynthase, making contact with two neighboring molecules and blocking proton flow, which explains theinhibitory effect on ATP synthesis. Intraperitoneal injection of oligomycin into the rat (0.5 mg per kg)reduces the oxygen consumption by about 50%; decreases ATP production by the aerobic pathway andincreases formation of lactate in blood serum. These changes may cause a decelerated metabolism andan increased formation of free radicals or ROS in membranes.

Impact of warm ischemia injury on mitochondrial morphology and function of rat donor liver after cardiac death / 中国组织工程研究

BACKGROUND:How to control functional activity of donor liver after cardiac death and maintain the optimal function of grafts are the key issues in organ transplantation study. OBJECTIVE:To preliminarily explore the effect of warm ischemia injury on the morphology and function of rat donor liver after cardiac death. METHODS:Cardiac death model was established in Sprague-Dawley rats and the successful models were divided into six groups:control group (warm ischemia for 0 minute), warm ischemia 10 group (warm ischemia for 10 minutes), warm ischemia 20 group (warm ischemia for 20 minutes), warm ischemia 30 group (warm ischemia for 30 minutes), warm ischemia 40 group (warm ischemia for 40 minutes) and warm ischemia 50 group (warm ischemia for 50 minutes). The rat liver specimens in each group were cut into ultrathin sections. The structure of liver cells was observed and photographed by electron microscopy. Flameng score was applied to analyze the degree of mitochondrial damage. Liver mitochondria were extracted and then spectrophotometry was used to assess the viability of cytochrome C oxidase. RESULTS AND CONCLUSION:Under electron microscopy, there were no significant changes in liver cells within 30 minutes of warm ischemia, nuclear membrane was intact, mitochondria mildly swel ed, no mitochondrial crista ruptured, and Flameng score was<2 points. With the extension of warm ischemia time, the cells became swel ing, nuclear chromatin condensated, apoptotic body was clearly visible, mitochondrial matrix coagulated, mitochondria exhibited vacuolation, and Flameng score was 3-4 points. The viability of cytochrome C oxidase showed no significant difference within 30 minutes of warm ischemia, but began to significantly decrease at 40 and 50 minutes. The mitochondrial structure and function after liver injury is not obviously affected by 30 minutes of warm ischemia, and significant changes appear after 40 minutes.

Fisiopatologia do Transtorno de Humor Bipolar e efeito do tratamento com lítio: enfoque em neuroproteção e função mitocondrial / Bipolar disorder pathophysiology and the effect of lithium treatment: focus on neuroprotection and mitochondrial function

Introdução: Diversas evidências apontam para um papel da disfunção mitocondrial no Transtorno de Humor Bipolar (THB), mas pouco se sabe sobre isso no THB de início recente. Na mitocôndria a atividade da cadeia transportadora de elétrons (CTE) atua juntamente com o ciclo do ácido cítrico na produção de energia, mas não está claro se estão alteradas no THB. O DNA mitocondrial (DNAmt) codifica diversas proteínas da CTE e está associado ao estresse oxidativo, mas nunca foi avaliado em pacientes no THB in vivo. O estresse oxidativo está associado ao THB e à disfunção mitocondrial, mas não se sabe muito das atividades das enzimas antioxidantes no THB de início recente. O óxido nítrico (NO) é uma molécula com efeitos neuromoduladores, mas com um papel no THB ainda não elucidado. O lítio é um tratamento padrão-ouro no THB, tendo mostrado efeitos neuroprotetores. Apesar disso, pouco se conhece do efeito do lítio na CTE, nas enzimas do ciclo do ácido cítrico, no conteúdo de DNAmt e na regulação de NO em humanos. Também não está claro o papel antioxidante do lítio no THB. Metódos: Pacientes com THB em depressão (n=31), não medicados em sua maioria (84%), foram tratados por 6 semanas com lítio. Antes e depois do tratamento, verificaram-se em leucócitos as atividades dos complexos I-IV da CTE, atividades das enzimas citrato sintase, succinato desidrogenase e malato desidrogenase e também o conteúdo de DNAmt; em plasma foram analisados os níveis de NO, substâncias reativas ao ácido tiobarbitúrico (TBARS) e as atividades de catalase (CAT), glutationa peroxidase (GPx), superóxido dismutase (SOD) e razão de SOD/CAT. Os pacientes com depressão bipolar foram comparados com 28 controles saudáveis. Resultados: Em comparação com controles, os pacientes com THB tiveram um aumento de GPx (p < 0,001) e CAT (p=0,005) e uma diminuição de SOD/CAT (p=0,001), sem outras diferenças nos demais biomarcadores. Pacientes com THB I mostraram uma diminuição de citrato sintase (p=0,02) e uma...

Background: Several evidences point to a role for mitochondrial dysfunction in Bipolar Disorder (BD), but few is known about it on short-term BD. In mitochondria the electron transport chain (ETC) acts jointly with citric acid cycle to produce energy, but it is not clear if they are altered in BD. Mitochondrial DNA (mtDNA) encodes several ETC proteins and is associated with oxidative stress, but it was never evaluated in BD in vivo. Oxidative stress is associated with BD and with mitochondrial dysfunction, but few is known about the activities of antioxidant enzymes in short-term BD. Nitric oxide (NO) is a molecule with neuromodulatory effects, but with an unclear role in BD. Lithium is a gold-standard treatment for BD, which has shown neuroprotective effects. However, few is known about lithium effect on ETC, citric acid cycle, mtDNA content, and NO regulation in humans. Also, lithium's antioxidant role in BD is unclear. Methods: Patients with BD depression (n=31) unmedicated in majority (84%) received lithium treatment for 6 weeks. Before and after treatment, in leukocytes the activities of ETC complex I-IV, citrate synthase, succinate dehydrogenase, and malate dehydrogenase, and mtDNA content were evaluated; in plasma, NO levels, thiobarbituric acid reactive substances (TBARS), the activities of catalase (CAT), glutathione peroxidase (GPx), and superoxide dismutase (SOD), and SOD/CAT ratio were evaluated. Bipolar depression patients were compared with 28 healthy controls. Results: When compared with controls, BD patients showed an increase in GPx (p < 0.001) and CAT (p=0.005) and a decrease in SOD/CAT (p=0.001), but showed no difference for other biomarkers. Patients with BD I showed a decrease in citrate synthase (p=0.02) and a slight decrease in mtDNA content (p=0.05) when compared to BD II; mtDNA content was slightly decreased in BD I compared to controls (p=0.05). From baseline to endpoint, there was an increase in ETC complex I activity (p=0.02),...

The mechanism of miR-181c induced neuroprotection by hypoxia preconditioning in rats / 中华神经科杂志

Objective To investigate the neuroprotective effect of miR-181c on hypoxia-preconditioned ischemia in rats and its mechanism.Methods Thirty-nine male SD rats were randomly divided into 5 groups of control group,sham-operated group,middle cerebral artery occlusion (MCAO)group,hypoxia-preconditioned group,hypoxia-preconditioned and MCAO group.Infarct volume and behavioral deficits were quantified.Real-time PCR was applied to detect the expression levels of miR-181c and Western blotting was used to verify the target protein of mt-cox1.Results Under the treatment of hypoxia-preconditioned,the neurological impairment was alleviated and the infarct volume was reduced significantly from 22.50％ ±2.96％ to 16.40％ ±3.13 ％ (t =5.26,P ＜0.01).The expression of miR-181c was decreased significantly in hypoxia-preconditioned and MCAO group than that in MCAO group (1.89 ± 0.14 vs 3.05 ± 0.26,t =6.10,P ＜ 0.01),and the expression of mt-cox1 protein was also significantly decreased (0.54 ± 0.07 vs 0.93 ± 0.04,t =8.01,P ＜ 0.01).Conclusion Hypoxia-preconditioned may attenuate the ischemic injury in SD rats,which may be related to the down-regulation of the expression of miR-181c,therefore increasing the expression of its targeted protein mt-cox1.

Anesthetic Management of a Patient with Mitochondrial Myopathy Undergoing Orthopedic Surgery

Mitochondria myopathies consist of large heterogeneous groups of disorders resulting from primary dysfunction of the mitochondrial respiratory chain and causing muscle disease. These disorders involve the multiple organ system and appear to be extensive variability in clinical presentation. Therefore anesthetic management of patients with mitochondrial myopathies should begin with careful preoperative investigation and preparation. We report successful general anesthesia in a 22-year-old man with mitochondrial myopathy undergoing orthopedic surgery.

Effects of Mitochondrial Reactive Oxygen Species on Neuronal Excitability in Rat Spinal Substantia Gelatinosa Neurons

Recent studies indicate that reactive oxygen species (ROS) are critically involved in persistent pain primarily through spinal mechanisms, and that mitochondria are the main source of ROS in the spinal dorsal horn. To investigate whether mitochondrial ROS can induce changes in membrane excitability on spinal substantia gelatonosa (SG) neurons, we examined the effects of mitochondrial electron transport complex (ETC) substrates and inhibitors on the membrane potential of SG neurons in spinal slices. Application of ETC inhibitors, rotenone or antimycin A, resulted in a slowly developing and slight membrane depolarization in SG neurons. Also, application of both malate, a complex I substrate, and succinate, a complex II substrate, caused reversible membrane depolarization and enhanced firing activity. Changes in membrane potential after malate exposure were more prominent than succinate exposure. When slices were pretreated with ROS scavengers such as phenyl-N-tert-buthylnitrone (PBN), catalase and 4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPOL), malate-induced depolarization was significantly decreased. Intracellular calcium above 100 microM increased malateinduced depolarization, witch was suppressed by cyclosporin A, a mitochondrial permeability transition (MPT) inhibitor. These results suggest that enhanced production of spinal mitochondrial ROS can induce nociception through central sensitization.

Construction of Cox7a2 fluorescent vector and its effect on cytochrome C oxidase activity in mouse Sertoli cell line TM4 / 中华泌尿外科杂志

Objective To construct Cox7a2 fluorescent vector and study its effect on cytochrome C oxidase (COX) activity in mouse Sertoli cell line TM4. Methods The coding region of Cox7a2 was amplified from mouse Sertoli cell line TM4 by RT-PCR. The PCR product was inserted into pEYFP-C1 vector with BamH I and EcoR I restriction site, and confirmed by DNA sequencing. The recombinant fusion protein vector was amplified by transforming into DH5a and transfected into TM4 cells. The protein expression was identified by Western blot. COX activity was measured by spectrophotometer 6, 12, 24 and 48 h after the transfection of recombinant vector into the TM4 cell line. Results The entire coding sequence of Cox7a2 was cloned with 252 bp length. Plasmid pEYFP-C1-Cox7a2 vector was constructed and the positive clones were verified by restriction enzymes digestion and DNA sequencing. The transfection efficiency of the TM4 cell line was about 70% and 37000 D fusion protein was obtained. The COX activities were (0.642±0.051), (0.542±0.049), (0.311±0.021) and (0.216±0.010) U/mg 6, 12, 24 and 48 h after the transfection of recombinant vector in the TM4 cell line. Meanwhile, the COX activities were (0.714±0.064) and (0.653±0.031) U/mg in non-tranfected and naked vector group respectively. Compared with the non-tranfected group, COX activity decreased significantly 12, 24 and 48 h after the transfection. Conclusions The recombint plasmid vector was successfully constructed. Cox7a2 gene has an inhibiting effect on COX activity and may play an important role in the regulation of COX activity in mouse Sertoli cell line TM4.