81 Linoleic acid required for reduction of apoptosis through nuclear transcription factor-kappa B during pig embryo development

2020 ◽  
Vol 32 (2) ◽  
pp. 167
Author(s):  
D. Lee ◽  
K. Choi ◽  
J. Oh ◽  
S. Kim ◽  
M. Lee ◽  
...  
Keyword(s):  
Gene Expression ◽  
Fatty Acids ◽  
Transcription Factor ◽  
Linoleic Acid ◽  
Embryo Development ◽  
Mrna Level ◽  
Metabolic Energy ◽  
Apoptotic Gene ◽  
Nuclear Transcription Factor ◽  
Apoptotic Genes

Recent studies suggest that endogenous and exogenous free fatty acids play various important roles in mammalian oocyte and pre-implantation embryo development. Among fatty acids, linoleic acid (LA) has been reported to affect the apoptosis pathway via nuclear transcription factor-kappa B (NF-κB). The transcription factor NF-κB is a key modulator of apoptosis in a variety of cell types, but to date, this specific function of NF-κB has not been demonstrated in porcine pre-implantation embryos. To examine the effect of linoleic acid on invitro-produced parthenogenetic pig embryos, we treated LA by concentration (0, 10, 25, 50, and 100 µM) to identify developmental rate, NF-κB expression, and mRNA level of apoptotic-related genes. In addition, the mechanism was confirmed by examining the protein and mRNA expression of NF-kb and c-jun by immunostaining and quantitative PCR at the blastocyst stage. Linoleic acid had a positive effect on embryo development without toxicity at a certain concentration (25 µM), but toxicity was confirmed at higher (50-100μM) concentrations. Furthermore, it was confirmed that the concentration of NF-κB increased as the treatment concentration of LA increased, which was found to increase even at the concentration at which embryo development decreased. Previous studies have shown that the NF-κB pathway is involved in regulating anti- and pro-apoptotic gene expression. We also investigated the effects of LA on anti- (Bcl-xL, Mcl-1) and pro- (BAX1, TP53, Caspase3) apoptotic genes and NF-κB activation-related genes (RelA, JNK1, JNK2, IL-6) in porcine embryos. We have found that down-regulation of pro-apoptotic gene expression occurs in the LA-treated group. It was also found that Bcl-xL, one of the anti-apoptotic genes, was not affected by LA, which appears to be an effect of IL-6. In contrast, Mcl-1, an anti-apoptotic gene known not to be affected by IL-6, was found to have increased expression mRNA level in LA-treated pig embryos. Furthermore, through double-staining of apoptosis and immunocytochemistry, as the concentration of NF-kB level increases, the nuclear translocation of c-jun, the protein of which was also related with apoptosis, increased gradually depending on the LA concentration. These data could support that porcine embryo can use exogenous LA as a metabolic energy source. The data also demonstrate the important role of NF-kB in porcine early embryo development. Support was provided by the Korea Institute of Planning and Evaluation for Technology in Food, Agriculture, Forestry and Fisheries (IPET) through the Development of High Value-Added Food Technology program funded by the Ministry of Agriculture, Food and Rural Affairs (MAFRA, 118042-03-1-HD020).

scholarly journals Linoleic Acid Reduces Apoptosis via NF-κB During the in Vitro Development of Porcine Embryos.

2020 ◽  
Author(s):  
Chang-Kyu Lee ◽  
Dong-Kyung Lee ◽  
Kwang-Hwan Choi ◽  
Jong-Nam Oh ◽  
Seung-Hun Kim ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Transcription Factor ◽  
Linoleic Acid ◽  
Protein Expression ◽  
Embryo Development ◽  
Metabolic Energy ◽  
Mrna Levels ◽  
Apoptosis Pathway ◽  
Apoptotic Gene

Abstract Background: Recent studies suggest that endogenous and exogenous free fatty acids play many important roles in mammalian oocyte and preimplantation embryo development. Among these fatty acids, linoleic acid has been reported to affect the apoptosis pathway via nuclear transcription factor-kappa B. The transcription factor NF-κB is a key modulator of apoptosis in a variety of cell types, but to date, this specific function of NF-κB has not been demonstrated in porcine preimplantation embryos. To examine the effect of linoleic acid on parthenogenetic pig embryos produced in vitro, we treated these embryos with linoleic acid at various concentrations to examine the developmental rate, NF‐κB expression, IL-6 expression and apoptosis-related gene mRNA levels. Results: Linoleic acid had a positive effect on embryo development and was not toxic at a certain concentration, but toxicity was observed at higher concentrations. Furthermore, it was confirmed that the concentration of NF‐κB increased, unlike that of IL-6, as the concentration of linoleic acid increased, and the concentration of NF‐κB was found to increase even at the concentration of linoleic acid at which embryo development decreased. We found that pro-apoptotic gene expression was downregulated in the linoleic acid-treated group. It was also found that BCL-XL, an anti-apoptotic gene, was not affected by linoleic acid, which appears to be an effect of IL-6. In contrast, MCL-1, an anti-apoptotic gene known to be unaffected by IL-6, was found to be increased at the mRNA level in linoleic acid-treated pig embryos. Furthermore, based on both apoptosis and immunocytochemistry staining, as the concentration of NF-kB increased, the nuclear translocation of C-JUN, which is also related to apoptosis, gradually increased, which was dependent on the linoleic acid concentration. It was confirmed that NF-κB is an important factor in the development of porcine embryos by confirming that treatment with a very low concentration of ammonium pyrrolidinedithiocarbamate (APDC, inhibitor of NF-κB) affected NF-κB protein expression, IL-6 protein expression and blastocyst production. Conclusion: These datas could suggest that porcine embryos can use exogenous linoleic acid as a metabolic energy source via NF-κB. The data also demonstrate the important role of NF-kB in porcine early embryo development.

scholarly journals Hypothyroidism alters the expression of Bcl-2 family genes to induce enhanced apoptosis in the developing cerebellum

2003 ◽  
Vol 176 (1) ◽  
pp. 39-46 ◽  
Author(s):  
R Singh ◽  
G Upadhyay ◽  
S Kumar ◽  
A Kapoor ◽  
A Kumar ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Fragmentation ◽  
Granule Cells ◽  
Cerebellar Granule Cells ◽  
Cell Loss ◽  
Apoptotic Gene ◽  
Euthyroid State ◽  
Family Gene ◽  
Apoptotic Genes ◽  
High Level

Thyroid hormone (TH) deficiency results in delayed proliferation and migration of cerebellar granule cells. Although extensive cell loss during the development of the cerebellum under hypothyroid conditions is known, its nature and its mechanism are poorly understood. Bcl-2 family gene expression is known to determine the fate of cells to undergo apoptosis. We evaluated the effect of hypothyroidism on Bcl-2 family gene expression in the developing rat cerebellum. Electrophoresis and Western blotting were used to analyze DNA fragmentation and expression of DNA fragmentation factor (DFF-45), Bcl-2, Bcl-xL and Bax genes respectively. In the hypothyroid condition, extensive DNA fragmentation and enhanced cleavage of DFF-45 were seen throughout development (postnatal day 0 to day 24) and adulthood whereas they were absent in the euthyroid state. The anti-apoptotic genes Bcl-2 and Bcl-xL were down-regulated and the pro-apoptotic gene Bax was expressed at higher levels compared with the euthyroid state. These results suggest that normal levels of TH prevent cerebellar apoptosis to a large extent, whereas hypothyroidism not only increases the extent but also the duration of apoptosis by down-regulating the anti-apoptotic genes and maintaining a high level of the pro-apoptotic gene Bax.

scholarly journals MicroRNA-193b Controls Adiponectin Production in Human White Adipose Tissue

2015 ◽  
Vol 100 (8) ◽  
pp. E1084-E1088 ◽  
Author(s):  
Yasmina Belarbi ◽  
Niklas Mejhert ◽  
Silvia Lorente-Cebrián ◽  
Ingrid Dahlman ◽  
Peter Arner ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Adipose Tissue ◽  
Nuclear Receptor ◽  
White Adipose Tissue ◽  
Mrna Levels ◽  
Model Assessment ◽  
Human Adipocytes ◽  
Interacting Protein ◽  
Nuclear Transcription Factor

Context: MicroRNAs (miRNAs) are posttranscriptional regulators of gene expression. In white adipose tissue (WAT), recent studies suggest that miRNA levels are altered in various metabolic diseases, including obesity. Objective: The objective of the study was to determine whether adipocyte-expressed miRNAs altered by obesity can regulate adiponectin expression/secretion in fat cells. Design: Eleven miRNAs previously shown to be altered in obese human WAT were overexpressed in human in vitro-differentiated adipocytes followed by assessments of adiponectin levels in conditioned media. Setting: This was cohort study (n = 56) in an academic hospital. Patients: Subcutaneous WAT was obtained from nonobese and obese individuals. Interventions: There were no interventions in this study. Main Outcome Measure(s): Protein and mRNA levels of adiponectin were measured. Results: Of the 11 investigated miRNAs, three (miR-193b/-126/-26a) increased adiponectin secretion when overexpressed in human adipocytes. However, in human WAT only miR-193b expression correlated with adiponectin gene expression and homeostasis model assessment of insulin resistance. Moreover, quantitative PCR of miR-193b in both WAT and isolated adipocytes showed a significant association with serum adiponectin levels. Overexpression of miR-193b altered the gene expression of seven known adiponectin regulators. 3′-untranslated region reporter assays confirmed binding to cAMP-responsive element binding protein 5, nuclear receptor interacting protein 1, and nuclear transcription factor Yα. The effects of miR-193b on nuclear transcription factor Yα expression were confirmed at the protein level. Transfection with individual miRNA target protectors selective for nuclear transcription factor Yα and nuclear receptor interacting protein 1 abolished the stimulatory effect of miR-193b on adiponectin secretion. Conclusions: In human adipocytes, miR-193b controls adiponectin production via pathways involving nuclear transcription factor Yα and possibly nuclear receptor interacting protein 1.

scholarly journals Cyclic regulation of apoptotic gene expression in the mouse oviduct

2011 ◽  
Vol 23 (5) ◽  
pp. 638 ◽  
Author(s):  
Myoungkun Jeoung ◽  
Phillip J. Bridges
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Dynamic Structure ◽  
Oestrous Cycle ◽  
Sexually Transmitted ◽  
Apoptotic Gene ◽  
Fundamental Information ◽  
Apoptotic Genes ◽  
Ectopic Pregnancies ◽  
Cyclic Changes

The oviduct is a dynamic structure whose function relies upon cyclic changes in the morphology of both ciliated and secretory luminal epithelial cells. Unfortunately, infection of these epithelial cells by sexually transmitted pathogens can lead to pelvic inflammatory disease, ectopic pregnancies and infertility. The disruption of normal, cyclic apoptosis in the oviducal epithelium appears to be a causal factor of oviducal pathology and therefore, these pathways represent a potential target for diagnosis and therapeutic intervention. The objective of this study was to determine the pattern of expression for apoptotic genes in the oviduct of the naturally cycling mouse, generating fundamental information that can be applied to the development of animal models for research and the identification of targets for disease intervention. Whole oviducts were collected from regular cycling mice killed at 1 p.m. on each day of the oestrous cycle and the expression of 84 apoptotic genes determined by targeted PCR super-array. Intact and cleaved caspases were then evaluated by western blotting. The expression of mRNA for genes classified as pro-apoptotic (Bad, Bak1 and Bok) and anti-apoptotic (Bag3, Bnip2 and Xiap) was regulated by day (P < 0.05). Differences in the temporal expression of several p53-related genes (Trp53bp2, Trp53inp1 and Trp73), those specific to the TNF superfamily (Tnfrsf10 and Tnfsf10b) and one caspase (Casp14) were also observed (P < 0.05). The cleaved forms of Caspases-3, -6 and -12 were all detected throughout the oestrous cycle. These results represent the first pathway-wide analysis of apoptotic gene expression in the murine oviduct.

scholarly journals Fatty acids in component of milk enhance the expression of the cAMP-response-element-binding-protein-binding protein (CBP)/p300 gene in developing rats

2008 ◽  
Vol 99 (3) ◽  
pp. 481-486 ◽  
Author(s):  
Kazuki Mochizuki ◽  
Hiromi Kawai ◽  
Hiroko Mochizuki ◽  
Masaya Shimada ◽  
Sachiko Takase ◽  
...  
Keyword(s):  
Gene Expression ◽  
Fatty Acids ◽  
Small Intestine ◽  
Protein Binding ◽  
Binding Protein ◽  
Mrna Level ◽  
Camp Response Element Binding ◽  
Camp Response Element ◽  
Response Element ◽  
Element Binding Protein

Fatty acids in milk are thought to play an important role in intestinal maturation and gene expression in the rat small intestine during the suckling–weaning period. In the present study, we determined the jejunal mRNA level of the cAMP-response-element-binding-protein-binding protein (CBP)/p300, which is one of the chromatin remodelling factors and regulates histone acetylation, during the postnatal period in rats. The mRNA level of CBP/p300 was high during the suckling and middle of the weaning period (day 5 to 20) and then declined sharply to a low level at the end of the weaning period and after weaning. In situ hybridisation also showed that CBP/p300 mRNA levels in the villus as well as the basal membrane clearly decreased after weaning. Rat pups at age 17 d, weaned to a high-fat diet, showed higher levels of CBP/p300 mRNA than those weaned to a low-fat diet. Oral administration of caprylic acid, oleic acid and linoleic acid, which are major fatty acid components in milk, induced jejunal CBP/p300 gene expression. The present results suggest that fatty acids in components of milk enhance expression of the CBP/p300 genes in the small intestine.

Induction of Fetal Hemoglobin by Transcriptional Co-Activators MTF-1 and TSPYL1

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 353-353 ◽  
Author(s):  
Kenneth R Peterson ◽  
Flavia C Costa ◽  
Halyna Fedosyuk ◽  
Renee Neades ◽  
Johana Bravo de los Rios ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Protein Complex ◽  
Globin Gene ◽  
Mrna Level ◽  
Fetal Hemoglobin ◽  
K562 Cells ◽  
Zinc Metabolism ◽  
Globin Promoter ◽  
Globin Gene Expression

Abstract Abstract 353 Sickle cell disease (SCD) impacts one of 400 African-Americans born each year. Augmentation of fetal hemoglobin (HbF) levels is widely accepted as the most effective method for treating SCD, but hydroxyurea (HU) is currently the only approved drug that increases HbF. Thus, there is a need for the development of new therapies for this disease, including the identification of transcriptional activators that specifically up-regulate γ-globin (HbF). Developmental regulation of human β-like globin gene switching is controlled by several parameters, including cis- and trans-acting transcriptional determinants. Understanding the mechanisms underlying control of globin gene expression, particularly those involved in activation of γ-globin expression (HbF) is important for developing new treatments for SCD. Metal-responsive transcription factor-1 (MTF-1) is a key regulator of zinc metabolism in higher eukaryotes that controls the metal-inducible expression of metallothioneins and a number of other genes directly involved in the intracellular sequestration and efflux transport of zinc. Previous studies demonstrated that MTF-1 plays an essential role in liver development and that MTF-1-deficient mice display an anemic phenotype, suggesting a role for MTF-1 in hematopoiesis. In our study, when murine MTF-1 was expression was enforced, we observed a 5-fold increase in γ-globin expression in K562 cells. We also demonstrated increased γ-globin expression in adult blood from MTF-1 human β-globin locus yeast artificial chromosome (β-YAC) bi-transgenic (bigenic) mouse lines at the mRNA level by quantitative real-time RT-PCR (qPCR) and at the protein level by FACS analysis. Lastly, γ-globin gene expression was induced 12-fold in bone marrow cells (BMCs) derived from these bigenic mice compared to BMCs derived from β-YAC-only mice, and 3-fold after 6 hours of zinc treatment in β-YAC-only BMCs. Corroborative studies including zinc-deficient and zinc replete diets in β-YAC mice and erythroid-specific MTF-1 loss-of-function in loxP-flanked-MTF-1 LCR-β-globin promoter-Cre β-YAC mice further support a role for MTF-1 in g-globin gene expression. Chromatin immunoprecipitation (ChIP) analysis did not show recruitment of MTF-1 to any γ-globin gene-proximal metal response elements (MREs), the DNA motif that MTF-1 binds to control zinc metabolism genes. However, GATA-2 co-immunoprecipitated with MTF-1 in MTF-1 β-YAC BMCs, but not in β-YAC-only BMCs, suggesting that reactivation of γ-globin expression by MTF-1 might be mediated by a MTF-1-GATA-2 protein complex. ChIP experiments indicated that MTF-1 and GATA-2 co-occupy the same sites in the γ-globin promoter. Two of the stronger co-recruitment regions contain not only GATA sites, but also non-canonical MREs that vary by 1 or 2 bp from the canonical 7 bp MRE core. Interestingly, GATA-2 was induced 2-fold in adult blood of MTF-1 β-YAC mice, and also 3.5-fold in MTF-1 β-YAC BMCs treated with zinc for 6 hours. Our data suggest that activation of γ-globin by MTF-1 is mediated by protein-protein interaction with GATA-2 and that this multi-protein complex is targeted to GATA sites located in the γ-globin gene-promoters via binding of the GATA-2 protein. In a previous study we identified testis-specific protein Y-like 1 (TSPYL1) as a candidate gene involved in activation of γ-globin (de Andrade et al., 2006, Blood Cells, Mol. & Dis. 37:82). TSPYL1 mRNA level was increased 2–5 fold in deletional hereditary persistence of fetal hemoglobin (HPFH-2) subjects and decreased in a carrier of the Sicilian δβ-thalassemia trait. TSPYL1 is a transcription factor that is a member of the nucleosome assembly protein (NAP) family. TSPYL1 is not a DNA-binding protein; thus it exerts its effect through protein-protein interactions. When we enforced expression of human TSPYL1 in K562 cells an 11-fold induction of γ-globin expression was obtained. A reduction of γ-globin expression was observed following TSPYL1 knockdown in K562 cells. qPCR analysis of blood from TSPYL1 β-YAC bigenic mice showed that γ-globin expression was increased 4–12-fold. Taken together, our data strongly support the evidence that MTF-1 and TSPYL1 reactivate γ-globin expression in adult erythropoiesis. These two proteins represent potential new targets in strategies to reactivate γ-globin in hemoglobinopathies where higher levels of HbF would have beneficial effects. Disclosures: No relevant conflicts of interest to declare.

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