58 Reduced nutrient availability during invitro culture improves embryo production and morphological quality and alters metabolic status of bovine embryos

2021 ◽  
Vol 33 (2) ◽  
pp. 136
Author(s):  
É. C. dos Santos ◽  
A. M. da Fonseca Júnior ◽  
C. B. de Lima ◽  
J. Ispada ◽  
J. V. A. da Silva ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Amino Acids ◽  
Nutrient Availability ◽  
Metabolic Status ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Embryo Production ◽  
Energy Substrates ◽  
Trophectoderm Cells ◽  
Reactive Oxygen Species Content

Invitro production (IVP) of embryos is designed to reproduce an environment that resembles the female reproductive tract. However, the system does not perform optimally in terms of quality and embryo production. A major setback lies in the loss of dynamics observed in a static invitro system, which might affect the availability of substrates that reach the embryo. A reduction in the amount of nutrients in media has been used as an approach to improve IVP (Ermisch et al. 2020 Sci. Rep. 10, 9263; https://doi.org/10.1038/s41598-020-66019-4). The present study aimed at describing a defined sequential medium (embryonic culture supplementation, ECS) and to investigate the effect of reducing nutrient availability on embryo production, quality, and metabolism. ECS was developed in our laboratory and is a serum-free, salt-based culture medium supplemented with the amount of energy substrates and amino acids found in bovine oviduct (Ov) and uterus (Ut) fluids as previously described (Hugentobler et al. 2007Mol. Reprod. Dev. 74, 445–454; https://doi.org/10.1002/mrd.20607; Hugentobler et al. 2008Mol. Reprod. Dev. 75, 496–503; https://doi.org/10.1002/mrd.20760). Embryos were cultured according to the following ECS supplementation: ECS100 (supplemented with 8mg mL−1 bovine serum albumin and 100% of the energy substrates and amino acids concentrations of the Ov and Ut fluids) and ECS50 (half dilution of ECS100). Bovine oocytes from abattoir ovaries were submitted to IVP using standard protocols. On Day 0 of invitro culture, presumptive zygotes were randomly divided into groups ECS100-Ov or ECS50-Ov. On Day 4, embryos were respectively transferred to ECS100-Ut and ECS50-Ut. Expanded blastocysts were collected on Day 7 to assess embryo production, morphology (total cell number by Hoescht 33342 staining; inner cell mass and trophectoderm cells by CDX2 immunostaining), and metabolic status (mitochondrial activity and reactive oxygen species content by MitotrackerTM RedCMXRos and CellROXTM Green staining, ThermoFisher Scientific; NADH and FAD+ by autofluorescence). Data were analysed by Student’s t-test (a=4%). Although cleavage rates were similar between ECS50 and ECS100 (78.13±3.73 vs. 79.70±4.18; P=0.788), blastocyst rates were positively influenced by the reduction in concentration (28.88±1.74 vs. 16.73±2.41; P=0.004). This difference likely comes from a blockage at the morula stage in group ECS100, because the conversion from morula to blastocyst was 20% lower in this group (57.73±3.81 vs. 38.15±3.45; P=0.008). In terms of morphology, blastocysts produced in ECS50 had a higher number of cells (152.4±9.61 vs. 118.3±7.22; P=0.036), which is explained by the higher number of trophectoderm cells. Finally, metabolic status was affected by nutrient reduction: embryos from ECS50 had higher mitochondrial activity, reactive oxygen species content (P<0.0001), and lower NADH (P=0.01), suggesting higher oxidative phosphorylation to produce energy, as expected at this stage. In conclusion, ECS is a functional medium, and a reduced nutrient concentration (ECS50) improves embryo production, morphological quality, and metabolic status of blastocysts, suggesting that culture conditions must be adapted to the invitro system rather than resembling invivo conditions. This research was funded by FAPESP (2016/00350-0, 2017/18384-0).

scholarly journals Methionine and Glycine Stabilize Mitochondrial Activity in Sake Yeast During Ethanol Fermentation

2019 ◽  
Vol 57 (4) ◽  
pp. 535-543
Author(s):  
Ferdouse Jannatul ◽  
Yuki Kusaba ◽  
Yuki Fujimaru ◽  
Yuki Yamamoto ◽  
Hiroshi Kitagaki
Keyword(s):  
Reactive Oxygen Species ◽  
Amino Acids ◽  
Ethanol Fermentation ◽  
Reactive Oxygen ◽  
Yeast Cells ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Brewer’S Yeasts ◽  
Fermentation Media

Addition of amino acids to fermentation media affects the growth and brewing profiles of yeast. In addition, retaining mitochondrial activity during fermentation is critical for the fermentation profiles of brewer’s yeasts. However, a concrete mechanism linking amino acids in fermentation media with mitochondrial activity during fermentation of brewer’s yeasts is yet unknown. Here, we report that amino acids in fermentation media, especially methionine (Met) and glycine (Gly), stabilize mitochondrial activity during fermentation of sake yeast. By utilizing atg32Δ mutant sake yeast, which shows deteriorated mitochondrial activity, we screened candidate amino acids that strengthened the mitochondrial activity of sake yeast during fermentation. We identified Met and Gly as candidate amino acids that fortify mitochondrial activity in sake yeast during fermentation. To confirm this biochemically, we measured reactive oxygen species (ROS) levels in sake yeast fermented with Met and Gly. Yeast cells supplemented with Met and Gly retained high ROS levels relative to the non-supplemented sake yeast. Moreover, Met-supplemented cells showed a metabolome distinct from that of non-supplemented cells. These results indicate that specific amino acids such as Met and Gly stabilize the mitochondrial activity of sake yeast during fermentation and thus manipulate brewing profiles of yeast.

scholarly journals MitoQ Is Able to Modulate Apoptosis and Inflammation

2021 ◽  
Vol 22 (9) ◽  
pp. 4753
Author(s):  
Elisa Piscianz ◽  
Alessandra Tesser ◽  
Erika Rimondi ◽  
Elisabetta Melloni ◽  
Claudio Celeghini ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Mevalonate Pathway ◽  
Neuronal Cell ◽  
Reactive Oxygen ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Short Period ◽  
Inflammatory Stimuli ◽  
Apoptosis And Inflammation ◽  
The Impact

Mitoquinone (MitoQ) is a mitochondrial reactive oxygen species scavenger that is characterized by high bioavailability. Prior studies have demonstrated its neuroprotective potential. Indeed, the release of reactive oxygen species due to damage to mitochondrial components plays a pivotal role in the pathogenesis of several neurodegenerative diseases. The present study aimed to examine the impact of the inflammation platform activation on the neuronal cell line (DAOY) treated with specific inflammatory stimuli and whether MitoQ addition can modulate these deregulations. DAOY cells were pre-treated with MitoQ and then stimulated by a blockade of the cholesterol pathway, also called mevalonate pathway, using a statin, mimicking cholesterol deregulation, a common parameter present in some neurodegenerative and autoinflammatory diseases. To verify the role played by MitoQ, we examined the expression of genes involved in the inflammation mechanism and the mitochondrial activity at different time points. In this experimental design, MitoQ showed a protective effect against the blockade of the mevalonate pathway in a short period (12 h) but did not persist for a long time (24 and 48 h). The results obtained highlight the anti-inflammatory properties of MitoQ and open the question about its application as an effective adjuvant for the treatment of the autoinflammatory disease characterized by a cholesterol deregulation pathway that involves mitochondrial homeostasis.

scholarly journals Plant Extracts and Reactive Oxygen Species as Two Counteracting Agents with Anti- and Pro-Obesity Properties

2019 ◽  
Vol 20 (18) ◽  
pp. 4556 ◽  
Author(s):  
Hanna Zielinska-Blizniewska ◽  
Przemyslaw Sitarek ◽  
Anna Merecz-Sadowska ◽  
Katarzyna Malinowska ◽  
Karolina Zajdel ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Plant Extracts ◽  
Complex Disease ◽  
Reactive Oxygen ◽  
Complementary Therapy ◽  
Biologically Active ◽  
Mitochondrial Activity ◽  
Oxygen Species

Obesity is a complex disease of great public health significance worldwide: It entails several complications including diabetes mellitus type 2, cardiovascular dysfunction and hypertension, and its prevalence is increasing around the world. The pathogenesis of obesity is closely related to reactive oxygen species. The role of reactive oxygen species as regulatory factors in mitochondrial activity in obese subjects, molecules taking part in inflammation processes linked to excessive size and number of adipocytes, and as agents governing the energy balance in hypothalamus neurons has been examined. Phytotherapy is the traditional form of treating health problems using plant-derived medications. Some plant extracts are known to act as anti-obesity agents and have been screened in in vitro models based on the inhibition of lipid accumulation in 3T3-L1 cells and activity of pancreatic lipase methods and in in vivo high-fat diet-induced obesity rat/mouse models and human models. Plant products may be a good natural alternative for weight management and a source of numerous biologically-active chemicals, including antioxidant polyphenols that can counteract the oxidative stress associated with obesity. This review presents polyphenols as natural complementary therapy, and a good nutritional strategy, for treating obesity without serious side effects.

scholarly journals Piceatannol Increases Antioxidant Defense and Reduces Cell Death in Human Periodontal Ligament Fibroblast under Oxidative Stress

Antioxidants ◽  
2019 ◽  
Vol 9 (1) ◽  
pp. 16 ◽  
Author(s):  
Flávia Póvoa da Costa ◽  
Bruna Puty ◽  
Lygia S. Nogueira ◽  
Geovanni Pereira Mitre ◽  
Sávio Monteiro dos Santos ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Reactive Oxygen Species ◽  
Cell Death ◽  
Cell Viability ◽  
Antioxidant Capacity ◽  
Periodontal Ligament ◽  
Cellular Metabolism ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Atp Production

Piceatannol is a resveratrol metabolite that is considered a potent antioxidant and cytoprotector because of its high capacity to chelate/sequester reactive oxygen species. In pathogenesis of periodontal diseases, the imbalance of reactive oxygen species is closely related to the disorder in the cells and may cause changes in cellular metabolism and mitochondrial activity, which is implicated in oxidative stress status or even in cell death. In this way, this study aimed to evaluate piceatannol as cytoprotector in culture of human periodontal ligament fibroblasts through in vitro analyses of cell viability and oxidative stress parameters after oxidative stress induced as an injury simulator. Fibroblasts were seeded and divided into the following study groups: control, vehicle, control piceatannol, H2O2 exposure, and H2O2 exposure combined with the maintenance in piceatannol ranging from 0.1 to 20 μM. The parameters analyzed following exposure were cell viability by trypan blue exclusion test, general metabolism status by the 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide (MTT) method, mitochondrial activity through the ATP production, total antioxidant capacity, and reduced gluthatione. Piceatannol was shown to be cytoprotective due the maintenance of cell viability between 1 and 10 μM even in the presence of H2O2. In a concentration of 0.1 μM piceatannol decreased significantly cell viability but increased cellular metabolism and antioxidant capacity of the fibroblasts. On the other hand, the fibroblasts treated with piceatannol at 1 μM presented low metabolism and antioxidant capacity. However, piceatannol did not protect cells from mitochondrial damage as measured by ATP production. In summary, piceatannol is a potent antioxidant in low concentrations with cytoprotective capacity, but it does not prevent all damage caused by hydrogen peroxide.

scholarly journals Redox Regulation and Oxidative Stress: The Particular Case of the Stallion Spermatozoa

Antioxidants ◽  
2019 ◽  
Vol 8 (11) ◽  
pp. 567 ◽  
Author(s):  
Fernando J. Peña ◽  
Cristian O’Flaherty ◽  
José M. Ortiz Rodríguez ◽  
Francisco E. Martín Cano ◽  
Gemma L. Gaitskell-Phillips ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Reactive Oxygen Species ◽  
Redox Regulation ◽  
Redox Homeostasis ◽  
Reactive Oxygen ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Redox Biology ◽  
Major Interest ◽  
And Oxidative Stress

Redox regulation and oxidative stress have become areas of major interest in spermatology. Alteration of redox homeostasis is recognized as a significant cause of male factor infertility and is behind the damage that spermatozoa experience after freezing and thawing or conservation in a liquid state. While for a long time, oxidative stress was just considered an overproduction of reactive oxygen species, nowadays it is considered as a consequence of redox deregulation. Many essential aspects of spermatozoa functionality are redox regulated, with reversible oxidation of thiols in cysteine residues of key proteins acting as an “on–off” switch controlling sperm function. However, if deregulation occurs, these residues may experience irreversible oxidation and oxidative stress, leading to malfunction and ultimately death of the spermatozoa. Stallion spermatozoa are “professional producers” of reactive oxygen species due to their intense mitochondrial activity, and thus sophisticated systems to control redox homeostasis are also characteristic of the spermatozoa in the horse. As a result, and combined with the fact that embryos can easily be collected in this species, horses are a good model for the study of redox biology in the spermatozoa and its impact on the embryo.

129 EFFECT OF SERUM SUPPLEMENTATION ON AMP-ACTIVATED PROTEIN KINASE (AMPK) ACTIVITY AND LIPID METABOLISM OF IN VITRO-CULTURED BOVINE EMBRYOS

2015 ◽  
Vol 27 (1) ◽  
pp. 156
Author(s):  
S. Prastowo ◽  
F. Rings ◽  
D. S. Wondim ◽  
E. Tholen ◽  
C. Looft ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Mitochondrial Biogenesis ◽  
Reactive Oxygen ◽  
Rna Isolation ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Bovine Embryos ◽  
Fluorescent Intensity ◽  
Ampk Activity

A major problem of embryos cultured in vitro with serum is cytoplasmic lipid accumulation resulting in lower cryotolerance compared with those derived from in vivo or in the absence of serum. AMPK is known as a master regulator of lipid, glucose, and protein metabolism in mammalian cells. Moreover, it has been reported as controller of acetyl-CoA carboxylase α (ACC), the gene responsible for lipid synthesis, and associated with mitochondrial biogenesis and activities in response to oxidative stress. In the present study we aimed to investigate the regulation of AMPK during serum supplementation in vitro. For this, bovine embryos were produced in vitro in SOF media supplemented with oestrous cow serum or fatty acid–free BSA as a system without serum. Triplicate pools (each 10 blastocysts) from each group were used for RNA isolation using Arcturus®PicoPure®RNA Isolation Kit (Life Technologies, USA). Reverse transcription was performed using a combination of Oligo(dT)23 and random primers. Quantification of AMPK catalytic α1 (AMPKA1), ACC, peroxisome proliferator-activated receptor gamma coactivator 1 α (PGC1A), and sterol regulatory element binding transcription factor 2 (SREBP2) transcripts were performed using ABI PRISM® 7000 SDS system (Applied Biosystems, Foster City, CA, USA) using GAPDH as internal control. Normalized log-transformed transcript amount data were statistically analysed using t-test. In addition, AMPK protein was detected by immunofluorescence, mitochondrial activity by MitoTracker® Red (Invitrogen, Carlsbad, CA, USA), and reactive oxygen species by H2DCFDA molecular probe (Life Technologies, USA), and fluorescent intensity signals were visualised under confocal laser scanning microscopy LSM 710 (Carl Zeiss, Germany). Results showed that the expression of AMPKA1, PGC1A, a mitochondrial biogenesis protein, and SREBP2, a regulator of lipid oxidation, were found to be lower (0.4-, 0.2-, and 0.7-fold, respectively; P < 0.05) in blastocysts derived from cultured with serum compared to without serum. By contrast, ACC was up-regulated in blastocysts cultured with serum by 1.8-fold (P < 0.05) compared to without serum. In comparison to blastocyst cultured without serum, a reduced fluorescent intensity was observed in AMPKA1 protein and mitochondrial activity in blastocyst cultured with serum. The presence of serum was also found to be involved in increasing reactive oxygen species accumulation in embryos cultured with serum. The reduced level of AMPK leads to increased ACC and subsequently enhanced conversion of fatty acids into lipid, which is associated with reduced mitochondrial biogenesis protein, elevated reactive oxygen species level, and reduced lipid oxidation by suppression of SREBP2. In conclusion, the presence of serum in in vitro culture environment affected the AMPK activity and thereby genes associated with lipid metabolism in early bovine embryos.

Mitochondrial dysfunction in sepsis

1999 ◽  
Vol 66 ◽  
pp. 149-166 ◽  
Author(s):  
Mervyn Singer ◽  
David Brealey
Keyword(s):  
Nitric Oxide ◽  
Reactive Oxygen Species ◽  
Animal Models ◽  
Mitochondrial Dysfunction ◽  
Duration Models ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Vascular Control ◽  
In Vivo Animal Models

The current mainstream view of organ failure induced by sepsis revolves around inflammation and loss of vascular control. However, there has been a resurgence in interest in bioenergetic failure due to mitochondrial dysfunction. This concept is not new--studies date back 30 years; however, the data have been highly conflicting with findings of either decreased, increased or unchanged mitochondrial activity and/or nucleotide levels. These studies are virtually all based on non-human cells, isolated perfused organs or in vivo animal models that have received a variety of insults ranging from mild to severe, and monitored for different durations ranging from minutes to weeks. As a generalization, there does appear to be depression of mitochondrial function with longer-duration models of greater severity. This is confirmed by the scanty human data currently available. This chapter provides an overview, and attempts to relate the biochemical changes to the clinical condition. The potential roles of nitric oxide, intracellular calcium and reactive oxygen species are highlighted.

scholarly journals Functional inactivation of the oestrogen receptor by the antioestrogen, ZM 182780, sensitises tumour cells to reactive oxygen species

1999 ◽  
Vol 161 (2) ◽  
pp. 199-210 ◽  
Author(s):  
CJ Newton ◽  
N Drummond ◽  
CH Burgoyne ◽  
V Speirs ◽  
GK Stalla ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Cell Death ◽  
Oestrogen Receptor ◽  
Reactive Oxygen ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Rat Pituitary ◽  
Functional Inactivation ◽  
Mediate Cell

Reactive oxygen species (ROS) play a fundamental role in both apoptotic and necrotic cell death. Their importance is highlighted by studies showing that they mediate cell death in response to radiotherapy and to some forms of chemotherapy. Here we provide the first evidence for a role of ROS in response to an antiendocrine agent currently undergoing clinical trials. Using the oestrogen receptor (ER) containing rat pituitary GH3 cell line, we show that cell death is induced by the pure steroidal antioestrogen, ZM 182780, and that this is blocked by the antioxidant, N-acetyl cysteine (NAC). By flow cytometry, we show that, prior to the onset of DNA breakdown measured by ELISA, ZM 182780 exposure has no significant effect on intracellular oxidant concentrations. In contrast, ZM 182780 exposure greatly increases sensitivity to oxidants generated by blocking cellular antioxidant pathways and from exogenous administration of hydrogen peroxide (H2O2). As both necrosis and apoptosis are controlled by mitochondrial function, further experiments conducted to determine mitochondrial membrane potential (Delta|gWm) have indicated that the ZM 182780-induced loss of ER function increases the ease with which oxidants collapse mitochondrial activity and, as a consequence, cell death.

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