178 Cellular and extracellular expression of stress response transcription factors in male and female bovine pre-implantation embryos under oxidative stress conditions

2019 ◽  
Vol 31 (1) ◽  
pp. 213
Author(s):  
M. O. Taqi ◽  
S. Gebremedhn ◽  
D. Salilew-Wondim ◽  
F. Rings ◽  
C. Neuhoff ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Transcription Factors ◽  
Stress Response ◽  
Cell Count ◽  
Expression Pattern ◽  
Total Cell ◽  
Expression Level ◽  
Total Cell Count ◽  
Oxygen Level ◽  
Male And Female

Pre-implantation embryo development is a critical stage, in which several development and stress-response related transcription factors (TF) are involved. Exposing embryos to environmental insults alter some of these stress response-related TF. However, their expression pattern in male and female embryos and their release via exosomes is still unclear. Here, we aimed to investigate the effect of culture-induced oxidative stress on development and expression pattern of stress-related TF in male and female embryos and in respective spent media coupled with exosomes. For this, bovine male and female zygotes were in vitro produced using sexed semen and cultured under 5% and 20% oxygen in exosome-depleted SOFaa medium (SOF with amino acids). Blastocysts were subjected to total RNA isolation followed by quantitative RT-PCR analysis of the selected TF (Nrf2, KLF4, NOTCH1, SREBF2, E2F1, CAT1, SOD1, and OCT4), as well as protein abundance analysis using immunofluorescence and related phenotypes analysis, including reactive oxygen species (ROS) level and total cell count. Furthermore, the spent embryo culture media were collected for exosomes isolation and expression analysis of candidate TF. The data were statistically analysed using one-way ANOVA followed by multiple pair-wise comparisons using the Tukey post hoc test. Results showed that the blastocyst rates of both male (29.9% v. 34.9%) and female (16.7% v. 26.5%) bovine embryos were significantly lower in 20% than in 5% oxygen level. Female blastocysts subjected to the higher oxygen level showed increased ROS level (37.66±1.70v. 45.32±2.05 in male and 29.42±1.44v. 45.51±2.06 in female) and significantly reduced total cell count compared with the male embryo counterpart (136.55±7.8v. 112.75±2.9 in male and 138.75±2.0v. 88.25±4.3 in female cultured in 5% and 20% oxygen levels, respectively). Consequently, the expression levels of Nrf2, KLF4, SREBF2, CAT1, SOD1, and OCT4 were significantly increased in male embryos exposed to oxidative stress compared with those cultured under the lower oxygen level. However, NOTCH1 and E2F1 were significantly increased in female embryos exposed to oxidative stress compared with the male counterparts. The mRNA level of SREBF2 was significantly increased in male embryos cultured under both 5% and 20% O2 compared with female embryos. The protein expression level of Nrf2 and KLF4 was higher in embryos cultured at 20% v. 5% O2 with greater Nrf2 abundance in male embryos. Consequently, the male embryos produced at 20% O2 released a higher number of exosomes enriched with Nrf2, SOD1, and NOTCH1 mRNA than the other groups. Interestingly, the exosomal mRNA expression level of E2F1 tended to be higher in female embryos exposed to oxidative stress than their male counterparts. Taken together, the male embryos were more tolerant to oxidative stress than female embryos via the activation Nrf2-mediated oxidative stress response and development related TF. The release of these TF via exosomes could enhance cellular homeostasis maintenance under oxidative stress.

Clade III cytokinin response factors share common roles in response to oxidative stress responses linked to cytokinin synthesis

2021 ◽  
Vol 72 (8) ◽  
pp. 3294-3306
Author(s):  
Ariel M Hughes ◽  
H Tucker Hallmark ◽  
Lenka Plačková ◽  
Ondrej Novák ◽  
Aaron M Rashotte
Keyword(s):  
Oxidative Stress ◽  
Transcription Factors ◽  
Stress Response ◽  
Stress Responses ◽  
Oxidative Stress Response ◽  
Response Factors ◽  
Ontology Term ◽  
Regulated Expression ◽  
Cytokinin Response ◽  
Oxidative Stress Responses

Abstract Cytokinin response factors (CRFs) are transcription factors that are involved in cytokinin (CK) response, as well as being linked to abiotic stress tolerance. In particular, oxidative stress responses are activated by Clade III CRF members, such as AtCRF6. Here we explored the relationships between Clade III CRFs and oxidative stress. Transcriptomic responses to oxidative stress were determined in two Clade III transcription factors, Arabidopsis AtCRF5 and tomato SlCRF5. AtCRF5 was required for regulated expression of >240 genes that are involved in oxidative stress response. Similarly, SlCRF5 was involved in the regulated expression of nearly 420 oxidative stress response genes. Similarities in gene regulation by these Clade III members in response to oxidative stress were observed between Arabidopsis and tomato, as indicated by Gene Ontology term enrichment. CK levels were also changed in response to oxidative stress in both species. These changes were regulated by Clade III CRFs. Taken together, these findings suggest that Clade III CRFs play a role in oxidative stress response as well as having roles in CK signaling.

scholarly journals Increasing Oxygen Partial Pressures Induce a Distinct Transcriptional Response in Human PBMC: A Pilot Study on the “Normobaric Oxygen Paradox”

2021 ◽  
Vol 22 (1) ◽  
pp. 458
Author(s):  
Deborah Fratantonio ◽  
Fabio Virgili ◽  
Alessandro Zucchi ◽  
Kate Lambrechts ◽  
Tiziana Latronico ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Transcription Factors ◽  
Stress Response ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Oxidative Stress Response ◽  
Molecular Characteristics ◽  
Nuclear Factor ◽  
Normobaric Oxygen ◽  
Oxygen Paradox

The term “normobaric oxygen paradox” (NOP), describes the response to the return to normoxia after a hyperoxic event, sensed by tissues as oxygen shortage, and resulting in up-regulation of the Hypoxia-inducible factor 1α (HIF-1α) transcription factor activity. The molecular characteristics of this response have not been yet fully characterized. Herein, we report the activation time trend of oxygen-sensitive transcription factors in human peripheral blood mononuclear cells (PBMCs) obtained from healthy subjects after one hour of exposure to mild (MH), high (HH) and very high (VHH) hyperoxia, corresponding to 30%, 100%, 140% O2, respectively. Our observations confirm that MH is perceived as a hypoxic stress, characterized by the activation of HIF-1α and Nuclear factor (erythroid-derived 2)-like 2 (NRF2), but not Nuclear Factor kappa-light-chain-enhancer of activated B cells (NF-κB). Conversely, HH is associated to a progressive loss of NOP response and to an increase in oxidative stress leading to NRF2 and NF-kB activation, accompanied by the synthesis of glutathione (GSH). After VHH, HIF-1α activation is totally absent and oxidative stress response, accompanied by NF-κB activation, is prevalent. Intracellular GSH and Matrix metallopeptidase 9 (MMP-9) plasma levels parallel the transcription factors activation pattern and remain elevated throughout the observation time. In conclusion, our study confirms that, in vivo, the return to normoxia after MH is sensed as a hypoxic trigger characterized by HIF-1α activation. On the contrary, HH and VHH induce a shift toward an oxidative stress response, characterized by NRF2 and NF-κB activation in the first 24 h post exposure.

scholarly journals Faster Evaluation of Contaminated Surfaces for Mould Inspections by Tape Sampling

2021 ◽  
pp. 516-522
Author(s):  
Meider J ◽  
Messal C
Keyword(s):  
Cell Count ◽  
Standard Test ◽  
Total Cell ◽  
Test Method ◽  
Indoor Environmental Quality ◽  
Total Cell Count ◽  
Step Method ◽  
Innovative Method ◽  
Quantitative Results ◽  
The Common

Taking a tape-lift sample is one of the main practices used by indoor environmental quality investigators for detecting whether mould structures (for example, spores and hyphae) have either settled onto or colonized the surface. Despite the popularity of the method, there can be significant inconsistency in how tape lifts are collected and evaluated. The common ASTM standard D7910-14: Practice for the Collection of Fungal Material from Surfaces by Tape Lift, describes the correct way to collect a tape-lift sample. Using ASTM D7658-17: Standard Test Method for Direct Microscopy of Fungal Structures from Tape, semi-quantitative results in percentage of infested area in a scale from 0 up to 5 are available only. In case histories or for mould removal control, the total cell count is needed. This cannot be realized by the ASTM method. Therefore, an innovative method is asked to combine the quickness of taping and the precision of total cell count. Our research team developed two methods to quickly and fully quantify the tape samples. Regarding the assessment criteria, the user can decide to operate with the 3-LINE method to achieve the highest precision or use the faster 3-STEP method for even better results. Therefore, an innovative method is asked to combine the quickness of taping and the precision of total cell count. The aim of the work is to develop two strategies to quickly and comprehensive quantify the tape samples.

scholarly journals Persistent Sputum Cellularity and Neutrophils May Predict Bronchiectasis

2011 ◽  
Vol 18 (4) ◽  
pp. 221-224 ◽  
Author(s):  
Nicole Drost ◽  
Liesel D’silva ◽  
Ryan Rebello ◽  
Ann Efthimiadis ◽  
Frederick E Hargreave ◽  
...  
Keyword(s):  
Cell Count ◽  
Roc Curves ◽  
Total Cell ◽  
Total Cell Count ◽  
High Resolution Computed Tomography ◽  
Predictive Values ◽  
Retrospective Survey ◽  
Cell Counts ◽  
Airway Diseases ◽  
Computed Tomography Scans

BACKGROUND: Quantitative cell counts in sputum provide an accurate assessment of the type and severity of bronchitis.OBJECTIVE: To examine whether sputum cell counts could identify bronchiectasis in patients with recurrent bronchitis.METHODS: A retrospective survey of a clinical database (January 2004 to January 2005) of quantitative cell counts from sputum selected from expectorate in patients with obstructive airways diseases was used to identify predictors of bronchiectasis using ROC curves. This was prospectively evaluated (February 2005 to April 2008) using high-resolution computed tomography scans of thorax that were independently scored by a radiologist who was blinded to the clinical details.RESULTS: The retrospective survey identified 41 patients with bronchiectasis among 490 patients with airway diseases. Total cell count of 60×106/g or greater of the selected sputum with predominant neutrophils on two occasions had a sensitivity of 86.7%, a specificity of 87.5%, and positive and negative predictive values of 93% and 78%, respectively, to identify bronchiectasis. In the prospective study, 10 of 14 (71%) patients who met these criteria were identified to have bronchiectasis. Both total cell count and the percentage of neutrophils correlated with radiographic bronchiectasis severity.CONCLUSIONS: Persistent or recurrent intense sputum cellularity with neutrophilia is suggestive of bronchiectasis.

Successful Expansion of Umbilical Cord Blood Cells as a Source of Hematopoietic Progenitors for Adult Transplantation.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2857-2857
Author(s):  
Rowayda E. Peters ◽  
Roger Strair ◽  
Arnold D. Rubin ◽  
Lauri Goodell ◽  
Roger R. Mrowiec ◽  
...  
Keyword(s):  
Cord Blood ◽  
Cell Count ◽  
Umbilical Cord Blood ◽  
Human Cell ◽  
Liquid Culture ◽  
Ex Vivo ◽  
Total Cell ◽  
Total Cell Count ◽  
Cell Engraftment ◽  
Cd34 Cells

Abstract The low yield of progenitor CD34+ cells recovered from umbilical cord blood (UCB) limits the utility of this source for transplantation in adults. This limitation has triggered investigations into how ex vivo expansion of hematopoietic stem cells (HSC) could be achieved to allow for transplantation in larger recipients. In the present study, the incubation of MNC and not selected CD34+cells in the presence of SCF 25ng/ml+MGDF 10ng/ml+FLt-3 25ng/ml+IL-6 20ng/ml, and 10% human serum in stroma-free liquid culture generated long-term expansion of transplantable UCB HSC. In vitro, HSC expansion from 13 UCB lasted >7 months giving 39, 1.3x104, and 4.7 x109 fold increase in total cell count after 14, 70 and 217 d of expansion as compared to d0 (105/ml). Similarly, CD34+ and CD34+/CD38− cell populations increased reaching 229 and 2.2x105 and 91 and 2.2x104 fold after 14 and 70d of expansion. Examination of cell morphology and analysis by flow cytometry showed the presence of primitive and mature cells belonging to all hematopoietic cell lineages. Similarly, multilineage colonies with recloning capacity were generated in culture. Erythroid, myeloid and mixed colonies increased by 116 and 1.8x104 fold and megakaryocytic colonies by 8 and 527 fold after 14 and 70d. Expanded cells were karyotypically normal and lacked the most common chromosome translocations seen in AML and CML t (15,17) and t (9, 22). HSC expanded for 6 and 13 weeks and cropreserved for 6–11 months were able to re-expand in liquid culture and generate colonies capable of recloning and multi-lineage differentiation. We estimated the frequency of SCID repopulating cells (SRC) in UCB samples expanded for 2 and 12 w using 1000,500,250 and 125 unselected CD34+ cells injected intravenously into sublethally irradiated NOD/SCID mice. Mice were sacrificed 20 weeks after transplantation. Human cell engraftment measured as CD45+ (HuCD45+) was detected in all mice (x3mice/dilution) (0.1–9.8%). In addition, HuCD45+ cells with multilineage phenotype were present, (CD19 (lymphoid), CD33 (myeloid), CD71and Glycophorin-A (erythroid) as well as CD34+/CD38− cells). SRC increased by 90 fold after 2 weeks of expansion and by 187 fold after 12 weeks compared to unexpanded CD34+cells. Additional proof of human cell engraftment was documented using semisolid culture (MethoCultTM GF H4434 Stem Cell Technologies). Human myeloid and erythroid colonies were generated from all dilutions, and counts ranged between 63–271/500,000 MNC. Initial studies to test the relative magnitude of UCB HSC expansion from 24-well plates to culture bags (OptiCyte TM, Baxter) using one UCB, the total cell count increased by (6.3 and 20 (bags) Vs 2.3 and 3.3 fold (wells) after 7 and 14d) and CD34+ subpopulations including CD34+/CD38−. Based on these ongoing results, a phase II clinical trial using ex vivo expanded UCB for 14d in a setting of sub ablative Conditioning is planned.

scholarly journals Sputum Neutrophilia Can Mask Eosinophilic Bronchitis During Exacerbations

2007 ◽  
Vol 14 (5) ◽  
pp. 281-284 ◽  
Author(s):  
Liesel D’silva ◽  
Christopher J Allen ◽  
Frederick E Hargreave ◽  
Krishnan Parameswaran
Keyword(s):  
Cell Count ◽  
Corticosteroid Treatment ◽  
Airway Disease ◽  
Primary Objective ◽  
Total Cell ◽  
Total Cell Count ◽  
Sputum Eosinophilia ◽  
Eosinophilic Bronchitis ◽  
Cell Counts ◽  
Before And After

BACKGROUND: Exacerbations of airway disease are eosinophilic, neutrophilic, both or neither. The primary objective of the present study was to identify whether the treatment of a neutrophilic bronchitis can unmask an associated eosinophilia.METHODS: A retrospective survey of 2160 consecutive sputum cell counts from 1343 patients with airway disease was conducted to identify patients with an isolated neutrophilic bronchitis, which was defined as a sputum total cell count of greater than or equal to 12×106cells/g of sputum and a proportion of neutrophils of 80% or greater. The characteristics of the patients who subsequently demonstrated sputum eosinophilia (3% or greater) within eight weeks of resolving the neutrophilia were compared with the patients who subsequently did not have sputum eosinophilia.RESULTS: Two hundred thirty-seven patients had 273 neutrophilic exacerbations. The sputum was re-examined within eight weeks in 65 patients (27.4%), of whom 38 (58.5%) had resolution of the neutrophilic bronchitis after treatment with an antibiotic. Of these 38 patients, 13 (34%) showed eosinophilia.CONCLUSIONS: A neutrophilic exacerbation of airway disease was observed to mask sputum eosinophilia in one-third of patients who had sputum cell counts available before and after antibiotic therapy. Hence, the absence of sputum eosinophilia during an infective exacerbation should not be used as an indication to reduce the dose of corticosteroids. To optimize therapy, repeat sputum cell count measurements are recommended after antibiotic treatment before changing corticosteroid treatment.

Freezability and total cell count of OPU-IVF embryos derived from rBST-treated donor heifers: Preliminary results

1998 ◽  
Vol 49 (1) ◽  
pp. 163
Author(s):  
P.E.J. Bols ◽  
A. Van de Velde ◽  
A. Van Soom ◽  
A. de Kruif
Keyword(s):  
Cell Count ◽  
Total Cell ◽  
Total Cell Count ◽  
Preliminary Results

Somatic cell types in goats milk in relation to total cell count, stage and number of lactation

1993 ◽  
Vol 12 (1) ◽  
pp. 89-98 ◽  
Author(s):  
A.M. Rota ◽  
C. Gonzalo ◽  
P.L. Rodriguez ◽  
A.I. Rojas ◽  
L. Martin ◽  
...  
Keyword(s):  
Somatic Cell ◽  
Cell Count ◽  
Cell Types ◽  
Total Cell ◽  
Total Cell Count

scholarly journals Kynurenine Metabolism Lifespan Extension Mediated by Oxidative Stress Response and Hypoxic Response in C. elegans

2021 ◽  
Vol 5 (Supplement_1) ◽  
pp. 684-684
Author(s):  
Raul Castro-Portuguez ◽  
Jeremy Meyers ◽  
Sam Freitas ◽  
Hope Dang ◽  
Emily Turner ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Transcription Factors ◽  
Stress Response ◽  
Aging Process ◽  
De Novo ◽  
Kynurenine Pathway ◽  
Oxidative Stress Response ◽  
Lifespan Extension ◽  
Hypoxic Stress ◽  
C Elegans

Abstract Aging is characterized by a progressive decline in the normal physiological functions of an organism, ultimately leading to mortality. Metabolic changes throughout the aging process disrupt the balance and homeostasis of the cell. The kynurenine metabolic pathway is the sole de novo biosynthetic pathway for producing NAD+ from ingested tryptophan. Altered kynurenine pathway activity is associated with both aging and a variety of age-associated diseases, and kynurenine-based interventions can extend lifespan in Caenorhabditis elegans. Our laboratory recently demonstrated knockdown of the kynurenine pathway enzymes kynureninase (KYNU) or 3-hydroxyanthranilic acid dioxygenase (HAAO) increases lifespan by 20-30% in C elegans. However, the mechanism of how these interventions may modulate response against different stressors during the aging process has yet to be explored. Fluorescent reporter strains show the stress-responsive transcription factors skn-1 (ortholog of NRF2/NFE2L2; oxidative stress response) and hif-1 (ortholog of HIF1A; hypoxic stress response) to be highly upregulated when the kynurenine pathway is inhibited. We also demonstrated the increase expression of gst-4 and gcs-1 (transcriptional targets skn-1), which are involved in production of the antioxidant glutathione (GSH), as well as upregulation of cysl-2 (transcriptional target of hif-1), a regulator of cysteine biosynthesis from serine. We hypothesize that lifespan extension resulting from kynurenine pathway inhibition is mediated, at least in part, by upregulation of these transcription factors, providing elevated defense against oxidative stress and hypoxic stress.

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