Taurine promotes axonal sprouting via Shh-mediated mitochondrial improvement in stroke

Purpose: Motor function is restored by axonal sprouting in ischemic stroke. Mitochondria play a crucial role in axonal sprouting. Taurine (TAU) is known to protect the brain against experimental stroke, but its role in axonal sprouting and the underlying mechanism are unclear. Methods: We evaluated the motor function of stroke mice using the rotarod test on days 7, 14, and 28. Immunocytochemistry with biotinylated dextran amine was used to detect axonal sprouting. We observed neurite outgrowth and cell apoptosis in cortical neurons under oxygen and glucose deprivation (OGD), respectively. Furthermore, we evaluated the mitochondrial function, adenosine triphosphate (ATP), mitochondrial DNA (mtDNA), peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PCG-1α), transcription factor A of mitochondria (TFAM), protein patched homolog 1 (PTCH1), and cellular myelocytomatosis oncogene (c-Myc). Results: TAU recovered the motor function and promoted axonal sprouting in ischemic mice. TAU restored the neuritogenesis ability of cortical neurons and reduced OGD-induced cell apoptosis. TAU also reduced reactive oxygen species, stabilized mitochondrial membrane potential, enhanced ATP and mtDNA content, increased the levels of PGC-1α, and TFAM, and restored the impaired levels of PTCH1, and c-Myc. Furthermore, these TAU-related effects could be blocked using an Shh inhibitor (cyclopamine). Conclusion: Taurine promoted axonal sprouting via Shh-mediated mitochondrial improvement in ischemic stroke.

Remote ischemic preconditioning-induced late cardioprotection: possible role of melatonin-mitoKATP-H2S signaling pathway

Purpose: Remote ischemic preconditioning (RIPC) confers cardioprotection against ischemia reperfusion (IR) injury. However, the precise mechanisms involved in RIPC-induced cardioprotection are not fully explored. The present study was aimed to identify the role of melatonin in RIPC-induced late cardioprotective effects in rats and to explore the role of H2 S, TNF-α and mitoKATP in melatoninmediated effects in RIPC. Methods: Wistar rats were subjected to RIPC in which hind limb was subjected to four alternate cycles of ischemia and reperfusion of 5 min duration by using a neonatal blood pressure cuff. After 24 h of RIPC or ramelteon-induced pharmacological preconditioning, hearts were isolated and subjected to IR injury on the Langendorff apparatus. Results: RIPC and ramelteon preconditioning protected the hearts from IR injury and it was assessed by a decrease in LDH-1, cTnT and increase in left ventricular developed pressure (LVDP). RIPC increased the melatonin levels (in plasma), H2 S (in heart) and decreased TNF-α levels. The effects of RIPC were abolished in the presence of melatonin receptor blocker (luzindole), ganglionic blocker (hexamethonium) and mitochondrial KATP blocker (5-hydroxydecanoic acid). Conclusion: RIPC produce delayed cardioprotection against IR injury through the activation of neuronal pathway, which may increase the plasma melatonin levels to activate the cardioprotective signaling pathway involving the opening of mitochondrial KATP channels, decrease in TNF-α production and increase in H2 S levels. Ramelteon-induced pharmacological preconditioning may also activate the cardioprotective signaling pathway involving the opening of mitochondrial KATP channels, decrease in TNF-α production and increase in H2 S levels.

Arsenic exposure and its implications in male fertility

Arsenic exposure is a global health concern. This toxic metalloid is ubiquitous in the environment and contaminates food and drinking water. Once ingested, it undergoes a complex metabolic process within the body, which contributes to its accumulation and reactivity. Arsenic toxicity stems from the induction of oxidative stress, inhibition of thiol-containing proteins, and mimicry of inorganic phosphates. Arsenic poisoning is associated with the development of reproductive disorders. In males, arsenic causes a reduction in testicular weight and alterations in steroidogenesis and spermatogenesis. Moreover, it reduces the number and quality of spermatozoa harvested from the cauda epididymis. The mitochondria are targets of arsenic toxicity because of the production of free radicals and their high content of cysteine-rich proteins and fatty acids. Mitochondrial dysfunction may contribute to reproductive disorders because this organelle is crucial for controlling testicular and epididymal events related to sperm production and maturation. All of these alterations mediated by arsenic exposure contribute to the failure of male reproductive competence by reducing gamete viability. This review describes the potential mechanisms of arsenic toxicity, its detrimental effects on male reproductive organs, and consequences on sperm fertility.(AU)

Through DNA sensors and hidden mitochondrial effects of SARS-CoV-2

The COVID-19 pandemic brought attention to studies about viral infections and their impact on the cell machinery. SARS-CoV-2, for example, invades the host cells by ACE2 interaction and possibly hijacks the mitochondria. To better understand the disease and to propose novel treatments, crucial aspects of SARS-CoV-2 enrolment with host mitochondria must be studied. The replicative process of the virus leads to consequences in mitochondrial function, and cell metabolism. The hijacking of mitochondria, on the other hand, can drive the extrusion of mitochondrial DNA (mtDNA) to the cytosol. Extracellular mtDNA evoke robust proinflammatory responses once detected, that may act in different pathways, eliciting important immune responses. However, few receptors are validated and are able to detect and respond to mtDNA. In this review, we propose that the mtDNA and its detection might be important in the immune process generated by SARS-CoV-2 and that this mechanism might be important in the lung pathogenesis seen in clinical symptoms. Therefore, investigating the mtDNA receptors and their signaling pathways might provide important clues for therapeutic interventions.(AU)

Through DNA sensors and hidden mitochondrial effects of SARS-CoV-2

The COVID-19 pandemic brought attention to studies about viral infections and their impact on the cell machinery. SARS-CoV-2, for example, invades the host cells by ACE2 interaction and possibly hijacks the mitochondria. To better understand the disease and to propose novel treatments, crucial aspects of SARS-CoV-2 enrolment with host mitochondria must be studied. The replicative process of the virus leads to consequences in mitochondrial function, and cell metabolism. The hijacking of mitochondria, on the other hand, can drive the extrusion of mitochondrial DNA (mtDNA) to the cytosol. Extracellular mtDNA evoke robust proinflammatory responses once detected, that may act in different pathways, eliciting important immune responses. However, few receptors are validated and are able to detect and respond to mtDNA. In this review, we propose that the mtDNA and its detection might be important in the immune process generated by SARS-CoV-2 and that this mechanism might be important in the lung pathogenesis seen in clinical symptoms. Therefore, investigating the mtDNA receptors and their signaling pathways might provide important clues for therapeutic interventions.(AU)

Citometria de Fluxo em andrologia: da pesquisa básica para a rotina clínica / Flow citometry in andrology: from basic research to clinical routine

Os primeiros estudos de espermatozoides com citometria de fluxo com espermatozoides iniciaram no final da década de 1970. Com os avanços tecnológicos, hoje contamos com equipamentos com alta sensibilidade e eficiência que, em conjunto com amplo catálogo de sondas fluorescentes, podemos mensurar com alta precisão características celulares. Como exemplo, destacam-se dano em membrana plasmática, atividade mitocondrial, produção de espécies reativas de oxigênio, dano ao DNA espermático e muito mais. Na presente revisão, as potencialidades e limitação para a implementação da citometria de fluxo na análise seminal de espécies domésticas são exploradas e comentadas.

The first flow cytometry studies with spermatozoa were published in the late 1970s. With the technological advances in the following years, today we have equipments with high sensitivity and efficiency that, together with a large number of commercially available fluorescent probes, allow us to measure different cell characteristics with high accuracy. Some of the most evaluated characteristics are plasma membrane damage, mitochondrial activity, production of reactive oxygen species, sperm DNA damage, and much more. In this review, the potentials and limitations for the implementation of flow cytometry in the seminal analysis of domestic species are explored and commented.

Evaluation of Indigo carmine on hepatic ischemia and reperfusion injury

Purpose To evaluate the effects of treatment with Indigo Carmine (IC) on rat livers subjected to ischemia-reperfusion injury. Methods The animals were subdivided into 4 groups: 1.SHAM group(SH) - saline; 2.SHAM group with IC-2mg/Kg(SHIC); 3.IR group - rats submitted to ischemia and reperfusion with saline(IR); 4.IR group with IC-2mg/Kg(IRIC). The IR protocol consists of liver exposure and administration of drug or saline intravenously, followed by 60 minutes of ischemia and 15 of reperfusion. Liver samples were collected for biochemical analysis. Results State 3 of mitochondrial respiration showed a significant worsening of the IRIC group in relation to all others. State 4 showed a difference between IRIC and SHIC. The Respiratory Control Ratio showed statistical decrease in IR and IRIC versus Sham. The osmotic swelling showed significant difference between SHxIR; SHICxIRIC and SHxIRIC. There was a significant increase in ALT in the IRIC group in relation to all the others. Concerning the nitrate dosage, there was a decrease in the group treated with IC(IRxIRIC). There was no difference regarding the dosage of Malondialdehyde. Conclusion IC was not able to protect mitochondria from IR injury and proved to be a potentiating agent, acting in synergy with the IR injury promoting damage to the hepatocyte membranes.(AU)

Avaliação distribuição mitocondrial na maturação in vitro de oócitos bovinos de ovários com baixa e alta contagem de folículos antrais / Mitochondrial distribution evaluation in vitro maturation of ovarian bovine oocytes with low and high antral follicle count

A Produção in vitro de embriões (PIVE) depende das etapas de maturação, fertilização e cultivo in vitro, sendo a maturação a etapa decisiva para o oócito atingir o estádio de mórula e blastocisto. Neste trabalho objetivou-se avaliar a qualidade de maturação in vitro, de acordo com a distribuição mitocondrial de oócitos provenientes de ovários de fêmeas de abatedouro, com baixa ou alta contagem de folículos antrais (CFA). Foram aspirados 243 ovários dos quais se selecionou 334 oócitos perante de cinco repetições para cada grupo. Os ovários foram separados em dois grupos: 1-baixa CFA (n=19), 2-alta CFA (n= 53) e os oócitos recuperados corados com a sonda MitoTracker Red. Os dados obtidos foram analisados pelo teste não paramétrico exato de Fisher (α=5%) mediante o software Statistical Analysis System for Windows (SAS Inst., Inc., Cary, NC).Em relação a localização das mitocôndrias o presente trabalho apresentou diferenças (p<0,001) entre oócitos imaturos e maturados, o que é desejável. Oócitos maturados apresentaram maior porcentagem de mitocôndrias dispersas no citoplasma e oócitos imaturos na periferia, independente do grupo experimental (baixa e alta CFA). Em oócitos imaturos, não houve diferença (p=0,1542) na localização das mitocôndrias entre os grupos baixa e alta CFA. No entanto, o grupo baixa CFA, após a maturação, apresentou menor quantidade de oócitos com mitocôndrias na periferia em relação aos de alta CFA (p = 0,0355), indicando melhor aquisição de competência desses oócitos para PIVE.

In vitro embryo production (PIVE) depends on the stages of maturation, fertilization and in vitro cultivation, being the decisive stage for the oocyte to reach the morula and blastocyst stage. This study aimed to evaluate the quality of in vitro maturation, according to the mitochondrial distribution of oocytes from ovaries of slaughter females with low or high antral follicle count (CFA). A total of 243 ovaries were aspirated from which 334 oocytes were selected for five replicates for each group. The ovaries were separated into two groups: 1-low CFA (n = 19) 2-high CFA (n = 53) and stained with the Mito Traker Red probe. Data obtained were analyzed by Fisher's exact nonparametric test ( α= 5%) using the Statistical Analysis System for Windows software (SAS Inst., Inc., Cary, NC). Regarding the location of mitochondria, the present study showed significant differences (p <0.001) between immature and mature oocytes, which is desirable. Mature oocytes had a higher percentage of dispersed mitochondria in the cytoplasm and immature oocytes in the periphery, regardless of the experimental group (low and high CFA). In immature oocytes, there was no difference (p = 0.1542) in the location of mitochondria between the low and high CFA groups. However, the oocytes of the low CFA group, after maturation, presented smaller number of oocytes with mitochondria in the periphery compared to the high CFA (p = 0.0355), indicating for this parameter a better competence of these oocytes for PIVE.

Avaliação distribuição mitocondrial na maturação in vitro de oócitos bovinos de ovários com baixa e alta contagem de folículos antrais / Mitochondrial distribution evaluation in vitro maturation of ovarian bovine oocytes with low and high antral follicle count

A Produção in vitro de embriões (PIVE) depende das etapas de maturação, fertilização e cultivo in vitro, sendo a maturação a etapa decisiva para o oócito atingir o estádio de mórula e blastocisto. Neste trabalho objetivou-se avaliar a qualidade de maturação in vitro, de acordo com a distribuição mitocondrial de oócitos provenientes de ovários de fêmeas de abatedouro, com baixa ou alta contagem de folículos antrais (CFA). Foram aspirados 243 ovários dos quais se selecionou 334 oócitos perante de cinco repetições para cada grupo. Os ovários foram separados em dois grupos: 1-baixa CFA (n=19), 2-alta CFA (n= 53) e os oócitos recuperados corados com a sonda MitoTracker Red. Os dados obtidos foram analisados pelo teste não paramétrico exato de Fisher (α=5%) mediante o software Statistical Analysis System for Windows (SAS Inst., Inc., Cary, NC).Em relação a localização das mitocôndrias o presente trabalho apresentou diferenças (p<0,001) entre oócitos imaturos e maturados, o que é desejável. Oócitos maturados apresentaram maior porcentagem de mitocôndrias dispersas no citoplasma e oócitos imaturos na periferia, independente do grupo experimental (baixa e alta CFA). Em oócitos imaturos, não houve diferença (p=0,1542) na localização das mitocôndrias entre os grupos baixa e alta CFA. No entanto, o grupo baixa CFA, após a maturação, apresentou menor quantidade de oócitos com mitocôndrias na periferia em relação aos de alta CFA (p = 0,0355), indicando melhor aquisição de competência desses oócitos para PIVE.(AU)

In vitro embryo production (PIVE) depends on the stages of maturation, fertilization and in vitro cultivation, being the decisive stage for the oocyte to reach the morula and blastocyst stage. This study aimed to evaluate the quality of in vitro maturation, according to the mitochondrial distribution of oocytes from ovaries of slaughter females with low or high antral follicle count (CFA). A total of 243 ovaries were aspirated from which 334 oocytes were selected for five replicates for each group. The ovaries were separated into two groups: 1-low CFA (n = 19) 2-high CFA (n = 53) and stained with the Mito Traker Red probe. Data obtained were analyzed by Fisher's exact nonparametric test ( α= 5%) using the Statistical Analysis System for Windows software (SAS Inst., Inc., Cary, NC). Regarding the location of mitochondria, the present study showed significant differences (p <0.001) between immature and mature oocytes, which is desirable. Mature oocytes had a higher percentage of dispersed mitochondria in the cytoplasm and immature oocytes in the periphery, regardless of the experimental group (low and high CFA). In immature oocytes, there was no difference (p = 0.1542) in the location of mitochondria between the low and high CFA groups. However, the oocytes of the low CFA group, after maturation, presented smaller number of oocytes with mitochondria in the periphery compared to the high CFA (p = 0.0355), indicating for this parameter a better competence of these oocytes for PIVE.(AU)

Oocyte mitochondria: role on fertility and disease transmission

Oocyte mitochondria are increased in number, smaller, and rounder in appearance than mitochondria insomatic cells. Moreover, mitochondrial numbers and activity are narrowly tied with oocyte quality because of the key role of mitochondria to oocyte maturation. During oocyte maturation, mitochondria display great mobility and cluster at specific sites to meet the high energy demand. Conversely, oocyte mitochondria are not required during early oogenesis as coupling with granulosa cells is sufficient to support gamete’s needs. In part, this might be explained by the importance of protecting mitochondria from oxidative damage that result in mutations in mitochondrial DNA (mtDNA). Considering mitochondria are transmitted exclusively by the mother, oocytes with mtDNA mutations may lead to diseases in offspring. Thus, to counterbalance mutation expansion, the oocyte has developed specific mechanisms to filter out deleterious mtDNA molecules. Herein, we discuss the role of mitochondria on oocyte developmental potential and recent evidence supporting a purifying filter against deleterious mtDNA mutations in oocytes.

Rivastigmine prevents injury induced by ischemia and reperfusion in rat liver

Purpose: To evaluate whether pre-treatment with rivastigmine is able to attenuate the I/R induced lesions in rat liver. Methods: SHAM animals or those submitted to I/R, non-treated or pre-treated with rivastigminine (2mg/kg) either 50 or 15 minutes before ischemia, were used. After I/R protocol, these animals were killed and their livers were harvested to measurement of the mitochondrial swelling as well as the malondialdehyde (MDA), nitrite and nitrate tissue concentration. Blood was also harvested for serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) determinations. Results: I/R promoted a significant increase of mitochondrial swelling in the studied animals. This increase of mitochondrial swelling was partially prevented by rivastigmine, but only if administered 50 minutes before ischemia. No significant modification of MDA, nitrite or nitrate tissue concentrations was observed in consequence of I/R, followed or not by rivastigmine treatments. In addition, I/R elevated both AST and ALT. These elevations of serum enzymes were not reversed by the different rivastigmine treatments. Conclusions: Rivastigmine administered 50 minutes before ischemia attenuates I/R-induced mitochondrial swelling, that indicates liver injury. This protective effect may be related to a greater stimulation of alfa7nAChR present in the Kupffer cells by the non-methabolized ACh, leading to an attenuation of I/R-induced inflammation.(AU)

Oocyte mitochondria: role on fertility and disease transmission

Oocyte mitochondria are increased in number, smaller, and rounder in appearance than mitochondria insomatic cells. Moreover, mitochondrial numbers and activity are narrowly tied with oocyte quality because of the key role of mitochondria to oocyte maturation. During oocyte maturation, mitochondria display great mobility and cluster at specific sites to meet the high energy demand. Conversely, oocyte mitochondria are not required during early oogenesis as coupling with granulosa cells is sufficient to support gametes needs. In part, this might be explained by the importance of protecting mitochondria from oxidative damage that result in mutations in mitochondrial DNA (mtDNA). Considering mitochondria are transmitted exclusively by the mother, oocytes with mtDNA mutations may lead to diseases in offspring. Thus, to counterbalance mutation expansion, the oocyte has developed specific mechanisms to filter out deleterious mtDNA molecules. Herein, we discuss the role of mitochondria on oocyte developmental potential and recent evidence supporting a purifying filter against deleterious mtDNA mutations in oocytes.(AU)

Evaluation of the therapeutic effect of methylene blue on the liver of rats submitted to ischemia and reperfusion

Purpose:To analyze the effect of methylene blue (MB) therapy during the liver ischemia-reperfusion injury (I/R) process.Methods:Thirty-five male Wistar rats were used, (70%) submitted to partial ischemia (IR) or not (NIR) (30%) were obtained from the same animal. These animals were divided into six groups: 1) Sham (SH), 2) Sham with MB (SH-MB); 3) I/R, submitted to 60 minutes of partial ischemia and 15 minutes of reperfusion; 4) NI/R, without I/R obtained from the same animal of group I/R; 5) I/R-MB submitted to I/R and MB and 6) NI/R-MB, without I/R. Mitochondrial function was evaluated. Osmotic swelling of mitochondria as well as the determination of malondialdehyde (MDA) was evaluated. Serum (ALT/AST) dosages were also performed. MB was used at the concentration of 15mg/kg, 15 minutes before hepatic reperfusion. Statistical analysis was done by the Mann Whitney test at 5%.Results:State 3 shows inhibition in all ischemic groups. State 4 was increased in all groups, except the I/R-MB and NI/R-MB groups. RCR showed a decrease in all I/R and NI/R groups. Mitochondrial osmotic swelling showed an increase in all I/R NI/R groups in the presence or absence of MB. About MDA, there was a decrease in SH values in the presence of MB and this decrease was maintained in the I/R group. AST levels were increased in all ischemic with or without MB.Conclusions:The methylene blue was not able to restore the mitochondrial parameters studied. Also, it was able to decrease lipid peroxidation, preventing the formation of reactive oxygen species.(AU)

Peça intermediária de espermatozoides bovinos: funções e anormalidades / Midpiece of bovine spermatozoa: functions and abnormalities

O reprodutor tem grande importância na cadeia produtiva por possuir maior potencial de difusão do seu material genético dentro do rebanho do que as fêmeas que produzem apenas uma cria por ano, em manejo de monta natural. Entretanto, este potencial somente se torna real quando os machos apresentam boa qualidade seminal, resultando em menores taxas de retorno ao estro. No entanto, apesar das características reprodutivas apresentarem baixa herdabilidade, é consensual que algumas patologias espermáticas possam apresentar características hereditárias e neste caso podem permanecer disseminadas nos rebanhos. Entre as anomalias espermáticas identificadas nos ejaculados de bovinos, as patologias de peça intermediária são classificadas como defeitos espermáticos maiores, comprometendo a fertilidade dos reprodutores, porém estas têm recebido pouca atenção. O objetivo deste trabalho de revisão é apresentar os principais estudos sobre a formação da peça intermediária, sua importância e como esta pode sofrer anormalidades, visto que ejaculados com presença destas anomalias apresentam baixa motilidade espermática e fertilidade.(AU)

Bulls are of immense importance in the productive chain because they have a greater potential of diffusion of genetic material within the herd than females that produce only one calf per year in natural mating management. However, this potential is only achieved when males present good seminal quality, resulting in lower rates of return to estrus. Although the reproductive traits have low heritability, it is consensual that some sperm pathologies may present hereditary characteristics and in this case they can remain disseminated in the herds. Among the sperm abnormalities identified in bovine ejaculates, the midpiece pathologies are classified as a major sperm defect, compromising bull fertility, but these pathologies have received little attention. The objective of this review is to present the main studies on the midpiece formation, its importance and how it can undergo abnormalities, since ejaculates with the presence of this pathology present low sperm motility and fertility.(AU)

Peça intermediária de espermatozoides bovinos: funções e anormalidades / Midpiece of bovine spermatozoa: functions and abnormalities

O reprodutor tem grande importância na cadeia produtiva por possuir maior potencial de difusão do seu material genético dentro do rebanho do que as fêmeas que produzem apenas uma cria por ano, em manejo de monta natural. Entretanto, este potencial somente se torna real quando os machos apresentam boa qualidade seminal, resultando em menores taxas de retorno ao estro. No entanto, apesar das características reprodutivas apresentarem baixa herdabilidade, é consensual que algumas patologias espermáticas possam apresentar características hereditárias e neste caso podem permanecer disseminadas nos rebanhos. Entre as anomalias espermáticas identificadas nos ejaculados de bovinos, as patologias de peça intermediária são classificadas como defeitos espermáticos maiores, comprometendo a fertilidade dos reprodutores, porém estas têm recebido pouca atenção. O objetivo deste trabalho de revisão é apresentar os principais estudos sobre a formação da peça intermediária, sua importância e como esta pode sofrer anormalidades, visto que ejaculados com presença destas anomalias apresentam baixa motilidade espermática e fertilidade.

Bulls are of immense importance in the productive chain because they have a greater potential of diffusion of genetic material within the herd than females that produce only one calf per year in natural mating management. However, this potential is only achieved when males present good seminal quality, resulting in lower rates of return to estrus. Although the reproductive traits have low heritability, it is consensual that some sperm pathologies may present hereditary characteristics and in this case they can remain disseminated in the herds. Among the sperm abnormalities identified in bovine ejaculates, the midpiece pathologies are classified as a major sperm defect, compromising bull fertility, but these pathologies have received little attention. The objective of this review is to present the main studies on the midpiece formation, its importance and how it can undergo abnormalities, since ejaculates with the presence of this pathology present low sperm motility and fertility.

Effect of irradiation with different laser wavelengths on oxidative stress of non-hepatectomized rats

PURPOSE: To assess the effect of two laser wavelengths, either separate or combined, on intact rat livers. METHOD: Nineteen male Wistar rats (200-300 g) were submitted to laser irradiation at 5 different sites on the liver surface.Wavelengths 660 and 780 nm were used, with a dose of irradiation of 60 J/cm2/site.The animals were divided into the groups:control (C) and animals irradiated with 660 nm laser (L1), with 780 nm laser (L2) or withboth wavelengths (L3).Mitochondrial function, mitochondrial swelling, and hepatocellular malondialdehyde (MDA) levels were determined.Data were analyzed by the Mann-Whitney test, with the level of significance set at 5%. RESULTS: There was a reduction of ADP-activated respiration (state 3) in group L1 compared to group C (p=0.0016), whereas the values of group L2 were similar to control.Group L3 also showed a reduction of state 3 (p=0.0159).There was a reduction of RCR in group L1 compared to control (p=0.0001) and to group L2 (p=0.0040).Mitochondrial swelling only differed between group L3 and control (p=0.0286).There was a increase in MDA levels in group L3 compared to control (p=0.0476) and to group L2 (p=0.0286) and in group L1 compared to group L2 (p=0.0132). CONCLUSION: Although laser irradiation reduced mitochondrial function,it did not interfere with the hepatocellular energy status.(AU)

RESPOSTAS AGUDAS HEMATOLÓGICAS, MUSCULARES E CARDÍACAS DE RATOS SOB CONDIÇÃO DE OVERREACHING

A síndrome do overtraining (SOT) é condição resultante do desequilíbrio entre intensidade de treinamento e tempo de recuperação, ocasionando redução do desempenho e fadiga. Objetivou-se investigar biomarcadores séricos musculares e perfil hematológico de ratos Wistar submetidos a um protocolo treino para provocar as condições FOR (functional overreaching) ou NFOR/SOT (nonfunctional overreaching) (PT). A amostragem inicial foi de 49 animais. Estes foram aleatoriamente distribuídos em 3 grupos iniciais: grupo sedentário (S, n=16), que não realizou nenhuma atividade física; grupo semissedentário (SSED, n=16), que realizou programa de condicionamento leve; grupo treinado (TR, n=33), que foi submetido a PT de 12 semanas. Para avaliação da aptidão física, os ratos foram submetidos a sete testes máximos de performance (TMP). No TMP-7, os animais do grupo TR foram subdivididos em dois grupos (FOR: n=19 e NFOR: n=14), identificados pela diferença da inclinação () obtida a partir da regressão linear dos desempenhos individuais nos TMP-2, 3, 4, 5, 6 e 7. Após o TMP-7, 8 ratos de cada grupo foram submetidos à eutanásia para obtenção de amostras. Realizou-se hemograma e citometria de fluxo de hemácias. Quantificaram-se as atividades séricas da creatina fosfoquinase (CK), aspartato aminotransferase (AST), lactato desidrogenase (LHD) e mioglobina. Avaliou-se as atividades da troponina cardíaca I (cTnI), isoenzima MB da creatinaquinase (CK-MB), lactato desidrogenase (LHD) e mioglobina. Foi determinada a relação miocardiossomática. Foram quantificados nos miocárdios a respiração mitocondrial máxima, atividade da citrato sintase (CS), e proteína indicadora de estresse (HSP70). Aplicou-se análise de variância de uma via (ANOVA One Way). Constatou-se valores superiores nos grupos SED e SSED em relação ao FOR para contagem de hemácias (RBC), hemoglobina (Hb) e hematócrito (Hct). O volume corpuscular médio (VCM) foi superior no grupo NFOR em comparação a SED e SSED. A contagem total de linfócitos foi maior no grupo NFOR em relação ao grupo FOR. A citometria de eritrócitos, a população 2 (P2) de RBC do grupo NFOR foi superior ao FOR. Não houve diferença entre os grupos para de CK, LDH e mioglobina. A AST do grupo SED foi menor que os demais grupos. Não houve diferença nas atividades séricas dos biomarcadores cardíacos entre os grupos. A relação miocardiossomática de NFOR foi superior aos demais grupos. O ganho de peso corporal foi semelhante entre FOR e NFOR, sendo este menor em relação a SSED. Não houve diferença da respiração máxima mitocondrial entre os grupos. A CS foi maior grupo NFOR em relação ao SED. A HSP70 foi maior no grupo SED, semelhante entre SSED e NFOR e menor no grupo FOR. A redução das variáveis eritrocitárias nos grupos FOR e NFOR foram associadas à expansão do volume plasmático e hemólise provocada por traumas inerentes ao desafio esportivo. A resposta leucocitária do grupo FOR pode estar relacionada à diminuição da resposta do eixo hipotálamo-hipófise-adrenal a agentes estressantes. A citometria de fluxo das hemácias mostrou-se promissora na diferenciação de FOR e NFOR. O condicionamento prévio pode ter atenuado a resposta dos biomarcadores musculares nos grupos treinados. À bioquímica sérica cardíaca evidenciou uma possível atenuação dos biomarcadores, como mecanismo adaptativo. O aumento da massa cardíaca em NFOR pode estar associado à hipertrofia deste órgão ou ao menor ganho de peso corporal. A avaliação da função mitocondrial evidenciou alterações adaptativas na célula muscular cardíaca em resposta ao protocolo de endurance, encorajando estudos complementares.

Overtraining syndrome (SOT) is a condition resulting from the imbalance between training intensity and recovery time, causing reduced performance and fatigue. The objective was to investigate serum muscle biomarkers and hematological profile of Wistar rats submitted to a training protocol to provoke the FOR (functional overreaching) or NFOR / SOT (nonfunctional overreaching) (PT) conditions. The initial sample was 49 animals. These were randomly assigned into 3 initial groups: sedentary group (S, n = 16), who did not perform any physical activity; semi-sedentary group (SSED, n = 16), which carried out a light conditioning program; trained group (TR, n = 33), who underwent PT for 12 weeks. To assess physical fitness, the rats were subjected to seven maximum performance tests (TMP). In the TMP-7, the animals in the TR group were subdivided into two groups (FOR: n = 19 and NFOR: n = 14), identified by the difference in the slope () obtained from the linear regression of individual performances in the TMP-2 , 3, 4, 5, 6 and 7. After TMP-7, 8 rats from each group were euthanized to obtain samples. CBC and erytrocyte flow cytometry were performed. The serum activities of creatine phosphokinase (CK), aspartate aminotransferase (AST), lactate dehydrogenase (LHD) and myoglobin were quantified. The activities of cardiac troponin I (cTnI), MB isoenzyme creatine kinase (CK-MB), lactate dehydrogenase (LHD) and myoglobin were evaluated. The myocardiosomatic relationship was determined. Maximal mitochondrial respiration, citrate synthase activity (CS), and stress indicator protein (HSP70) were quantified in the myocardium. One-way analysis of variance (ANOVA One Way) was applied. Higher values were found in the SED and SSED groups in relation to the FOR for red blood cell count (RBC), hemoglobin (Hb) and hematocrit (Hct). The mean corpuscular volume (VCM) was higher in the NFOR group compared to SED and SSED. The total lymphocyte count was higher in the NFOR group compared to the FOR group. The erythrocyte cytometry, population 2 (P2) of RBC of the NFOR group was higher than the FOR. There was no difference between groups for CK, LDH and myoglobin. The AST of the SED group was lower than the other groups. There was no difference in the serum activities of cardiac biomarkers between the groups. The myocardiosomatic ratio of NFOR was higher than the other groups. The body weight gain was similar between FOR and NFOR, which was lower in relation to SSED. There was no difference in maximum mitochondrial breathing between groups. CS was the largest NFOR group in relation to SED. HSP70 was higher in the SED group, similar between SSED and NFOR and lower in the FOR group. The reduction of erythrocyte variables in the FOR and NFOR groups was associated with the expansion of plasma volume and hemolysis caused by traumas inherent to the sports challenge. The leukocyte response of the FOR group may be related to the decreased response of the hypothalamic-pituitary-adrenal axis to stressors. RBC flow cytometry has shown promise in differentiating FOR and NFOR. The previous conditioning may have attenuated the response of the muscle biomarkers in the trained groups. Serum cardiac biochemistry showed a possible attenuation of biomarkers as an adaptive mechanism. The increase in cardiac mass in NFOR may be associated with hypertrophy of this organ or with lower body weight gain. The assessment of mitochondrial function showed adaptive changes in the cardiac muscle cell in response to the endurance protocol, encouraging further studies.

EFEITOS DO DESMAME NA FUNÇÃO MITOCONDRIAL, ESTRESSE OXIDATIVO, INFLAMAÇÃO E APOPTOSE EM LEITÕES DE DIFERENTES PESOS AO NASCER

O período pós-desmame é caracterizado por uma alta incidência de deficiência energética celular, distúrbios intestinais e infecções bacterianas que podem levar a doenças graves. Evidências indicam que a função mitocondrial e o estresse oxidativo estão intimamente relacionados no controle da eficiência energética celular, resposta imune e bactérias patogênicas. Portanto, estratégias eficazes que visam melhorar a saúde metabólica dos leitões e aumentar vigor durante o período pós-desmame precisam ser desenvolvidas. No entanto, mesmo que tal conceito seja altamente promissor, poucas informações estão disponíveis atualmente em relação à intensidade e duração desse estresse em leitões com normal e baixo peso ao nascimento. Portanto, objetivou-se caracterizar marcadores do metabolismo energético e estresse oxidativo ao longo do período peri-desmame. Um total de 30 (trinta) porcas multíparas foram inseminadas e as leitegadas foram padronizadas em 10 leitões. Os leitões foram distribuídos para um dos dois grupos experimentais: leitões de peso normal ao nascimento (NPN, 1,73 ± 0,01 kg, n = 60) e leitões de baixo peso ao nascimento (BPN, 1,01 ± 0,01 kg, n = 60). Em seguida, 10 leitões de cada grupo foram selecionados aos 14, 21, 23, 25, 29 e 35 dias de idade para a coleta de amostras de plasma e órgãos (fígado, jejuno e rins). Observou-se aumento significativo (P < 0,05) do dano oxidativo ao DNA (8-OHdG) e das proteínas plasmáticas (grupo carbonila). Uma diminuição (P < 0,05) no nível de energia celular na forma de adenosina trifosfato (ATP) foi observada no fígado após o desmame. Essa diminuição na produção de energia mitocondrial está relacionada à atividade mitocondrial maior (P<0,05) da enzima antioxidante glutationa peroxidase (GPx) e superóxido dismutase (SOD) no fígado, mucosa intestinal e rins de leitões após o desmame. Além disso, os níveis de dano oxidativo ao DNA (8-OHdG) mensurados no plasma e protéina hépatica foram maiores (P < 0,05) nos leitões BPN. Na sequência foram analisados ensaio de RT2 ProfilerTM PCR de genes relacionadas à função mitocondrial, estresse oxidativo, processos de inflamação e apoptose em leitões NPN (23d, 35d vs. 14d). A expressão dos genes selecionados foi avaliada por RT- PCR quantitativo. Essas análises revelaram que a expressão de RNAm das moléculas inflamatórias (IL-8 e CCL19) aumentou após o desmame na mucosa intestinal, enquanto a expressão de genes que codificam subunidades da cadeia respiratória mitocondrial (NDUFA2 e NDUFA5) foi reduzida no fígado de ambos os grupos (P < 0,05). Os principais moduladores da apoptose mitocondrial (BCL2A1 e BNIP3) e do sistema de defesa antioxidante (TXNRD2, GPx3, HMOX1) foram altamente expressos em leitões BPN comparados a NPN (P < 0,05). Além disso, os níveis sistêmicos de moléculas inflamatórias (TNF- e IL1-) aumentaram logo após o desmame e foram maiores nos leitões NPN (P < 0,05). Esses resultados indicaram que o desmame causa diminuição da energia mitocondrial no fígado, intestino e rim, em associação com condições de estresse oxidativo. Além disso, esses resultados sugerem que defesas antioxidantes, resposta inflamatória e apoptose estão comprometidas em leitões BPN.

The period following weaning is characterised by a high incidence of energetic deficiencies, intestinal disturbances and bacterial infections that led to serious diseases. Evidence indicates that mitochondrial function and oxidative stress are closely related to controlling cellular energetic efficiency, immune response and bacterial pathogenesis. Thus, effective strategies aiming to improve metabolic health of piglets and increased their robustness during the post-weaning period need to be developed. However,little information is currently available regarding the intensity and duration of this stress in both normal and low weight piglets. The objective was to characterize markers of energy metabolism and oxidative status throughout the peri- weaning period. Thirty (30) multiparous sows were inseminated, and litters were standardized to 10 piglets. Piglets were designed to one of two experimental groups: normal birth weight piglets (NBW, 1.73 ± 0.01 kg, n = 60) and low birth weight piglets (LBW, 1.01 ± 0. 01 kg, n = 60). Then, 10 piglets from each group were selected at 14, 21, 23, 25, 29 and 35 days of age to collect plasma and organ (liver, intestinal mucosa and kidney) samples. Analysis revealed a significant increase (P < 0.05) in plasma levels of both oxidative damage to DNA (8-OHdG) and proteins (protein carbonyls) measured in plasma increased after weaning. A decrease (P < 0.05) in cellular energy level in the form of adenosine triphosphate (ATP) content, known to be produced in mitochondria, was observed in the liver after weaning. This decrease in mitochondrial energy production was found to be linked to significantly higher (P < 0.05) mitochondrial activity major antioxidant glutathione peroxidase (GPx) and superoxide dismutase (SOD) in liver, intestinal mucosa and kidney of piglets after weaning. Additionally, the levels of oxidative damage to DNA (8-OHdG) measured in plasma and of liver protein carbonyls were found to be higher (P < 0.05) in LBW piglets. RT2 Profiler TM PCR arrays were assessed for genes related to mitochondrial function, oxidative stress, inflammation and apoptosis in NBW piglets (d 23 and 35 vs. d 14). The expression of selected genes was assessed by quantitative PCR. These analyzes revealed that the mRNA expression of inflammatory molecules (IL-8 and CCL19) increased after weaning in the intestinal mucosa, while the expression of genes encoding subunits of the mitochondrial respiratory chain (NDUFA2 and NDUFA5) was reduced in the liver of both groups (P <0.05). The main modulators of mitochondrial apoptosis (BCL2A1 and BNIP3) and the antioxidant defense system (TXNRD2, GPx3, HMOX1) were highly expressed in LBW piglets compared to NBW (P <0.05). In addition, the systemic levels of inflammatory molecules (TNF- and IL1- ) increased in the days after weaning and are higher in NBW piglets (P <0.05). These results indicated that weaning causes decrease in mitochondrial energy in liver, intestine and kidney after associated with oxidative stress conditions. Further, these findings suggest that antioxidant defenses, inflammatory response and apoptosis, are compromised in LBW piglets.

The role of dimethyl sulfoxide in the cryopreservation of Curimba (Prochilodus lineatus) semen / O papel do dimetilsulfóxido na criopreservação de sêmen de Curimba (Prochilodus lineatus)

Cryopreservation of Curimba semen (Prochilodus lineatus) is ecological and commercial importance. The objective of this study was to evaluate the effect of different concentrations (2, 5, 8 and 11%) of dimethyl sulfoxide (DMSO) diluted in Betsville Thawing Solution (BTS) on the quality of post-thaw semen Curimba. We analyzed the rate and period motility, sperm viability, membrane integrity and DNA, mitochondrial functionality, and fertilization and hatching rate. The plasma membrane and DNA integrity of a DMSO concentration of 11% obtained better results than the concentration of 5% (p<0.05). However, treatment of 5% DMSO resulted in a longer latency and a higher fertilization rate and hatching, in other sperm quality equal to that of fresh semen. The results of this study indicate that 5% DMSO is ideal for cryopreservation of semen Curimba.(AU)

A criopreservação de sêmen de Curimba (Prochilodus lineatus) é de importância ecológica e comercial. O objetivo deste estudo foi avaliar o efeito de diferentes concentrações (2, 5, 8 e 11%) de dimetilsulfóxido (DMSO) diluído em Betsville Thawing Solution (BTS) sobre a qualidade do sêmen de Curimba pós-descongelamento. Foram analisadas a taxa e período de motilidade, viabilidade espermática, integridade da membrana plasmática e do DNA, funcionalidade mitocondrial, e taxa de fertilização e eclosão. A membrana plasmática e a integridade do DNA a uma concentração de DMSO de 11% obtiveram melhores resultados que na concentração de 5% (p<0,05). Contudo, o tratamento de 5% de DMSO resultou em um aumento da latência e uma maior taxa de fertilização e de eclosão, com as demais qualidades do esperma sendo semelhante ao de sêmen fresco. Os resultados deste estudo indicam que 5% de DMSO é ideal para a criopreservação do sêmen Curimba.(AU)

Efeitos da suplementação de diferentes níveis dietéticos de INOSINA-5-MONOFOSFATO (5-IMP) para suínos em terminação (75 aos 100Kg PV)

Nos suínos com até 1 mês de vida as enzimas hexoquinases I e III convertem a glicose em glicose-6-fosfato, processo em que o ATP é hidrolisado em ADP para fornecer o grupo fosfato para glicose. Após um mês de vida, as atividades dessas enzimas são reduzidas e não convertem a glicose em glicose-6-fosfato em taxa suficiente para atender a demanda energética das atividades celulares. Diversas pesquisas apontam que a inosina5-monofosfato (5-IMP) é um nucleosídeo que possivelmente seja a principal fonte energética alternativa, devido sua conversão a adenosina monofosfato (AMP) e guanosina monofosfato (GMP). Estes nucleosídeos, posteriormente, são convertidos nos NTs (NTs) adenosina trifosfato (ATP) e guanosina trifosfato (GTP), principais moléculas energéticas empregadas nas atividades e manutenção das células. Diversas pesquisas também evidenciaram efeitos benéficos da suplementação da 5-IMP sobre o desempenho produtivo, qualidade de carne, sistema antioxidante, sistema imune e funcionalidade do metabolismo de leitões na fase de matermidade e no período pós-desmame. Diante disso, foram conduzidos três experimentos com o objetivo de avaliar a suple

In pigs up to 1-month-old as hexokinase enzymes, I and III convert glucose into glucose6-phosphate, a process in which ATP is hydrolyzed to ADP to provide the phosphate group for glucose. After a month of life, as activities of these enzymes are reduced and do not convert glucose into glucose-6-phosphate at a rate sufficient to meet the energy demand of cellular activities. Several kinds of research show that inosine-5'-monophosphate (5'-IMP) is a nucleoside that can be an alternative energy source, due to its conversion to adenosine monophosphate (AMP) and guanosine monophosphate (GMP). These nucleosides are later converted into the nucleotides adenosine triphosphate (ATP) and guanosine triphosphate (GTP), knowing as the main sources of energy used by cells, in their activities and maintenance. Several studies also show beneficial effects of 5'-IMP supplementation on productive performance, meat quality, antioxidant system, immune system, and piglet metabolism functionality in the weaning phase and postweaning period. Therefore, three experiments were conducted to evaluate the supplementation of different dietary levels of 5'-IMP in the diet of m