Glucose metabolism of in vitro-produced bovine embryos in cell-free and co-culture systems

2001 ◽  
Vol 2001 ◽  
pp. 216-216
Author(s):  
N.M. Orsi ◽  
J.B. Reischl ◽  
H.J. Leese
Keyword(s):  
Formation Rate ◽  
Lactate Production ◽  
Glucose Consumption ◽  
Bovine Embryos ◽  
Cell Monolayers ◽  
Glycolytic Activity ◽  
Wide Range ◽  
Oviduct Fluid

In vitro-produced (IVP) bovine embryos are used in a wide range of biotechnologies but develop less well than their in vivo counterparts and can give rise to foetal/neonatal anomalies after embryo transfer. The quality of bovine IVP embryos and the systems in which they are produced are traditionally assessed in terms of morphological and developmental criteria; namely, embryo grade and blastocyst formation rate. Lane and Gardner (1996) showed that mouse embryos selected for transfer on the basis of a low glycolytic activity (conversion of glucose to lactate), measured non-invasively, were 4 times more likely to implant than those selected randomly. Comparable data are not available for bovine embryos. The aim of this study was to assess linear glycolytic index of cattle blastocysts in vitro as a marker of viability. We have measured glucose consumption and lactate production by individual bovine IVP embryos grown in cell-free conditions and in a novel co-culture system (Orsi et al., 2000) involving confluent bovine oviduct epithelial cell monolayers on permeable supports. This preparation allows the epithelial cells to be fed by a nutritionally-rich medium via the physiological, basal, route, while the apical medium, containing the embryos, is more dilute, mimicking oviduct fluid.

Effects of bovine oviduct epithelial cells, fetal calf serum and bovine serum albumin on gene expression in single bovine embryos produced in the synthetic oviduct fluid culture system

2005 ◽  
Vol 17 (8) ◽  
pp. 751 ◽  
Author(s):  
Mona E. Pedersen ◽  
Øzen Banu Øzdas ◽  
Wenche Farstad ◽  
Aage Tverdal ◽  
Ingrid Olsaker
Keyword(s):  
Gene Expression ◽  
Fetal Calf Serum ◽  
Calf Serum ◽  
Developmental Competence ◽  
Bovine Embryos ◽  
Glut 1 ◽  
Significant Difference ◽  
Oviduct Fluid

In this study the synthetic oviduct fluid (SOF) system with bovine oviduct epithelial cell (BOEC) co-culture is compared with an SOF system with common protein supplements. One thousand six hundred bovine embryos were cultured in SOF media supplemented with BOEC, fetal calf serum (FCS) and bovine serum albumin (BSA). Eight different culture groups were assigned according to the different supplementation factors. Developmental competence and the expression levels of five genes, namely glucose transporter-1 (Glut-1), heat shock protein 70 (HSP), connexin43 (Cx43), β-actin (ACTB) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), analysed as mRNA by using reverse transcription–polymerase chain reaction, were measured on bovine embryos cultured for 9 days. Gene expression of these in vitro-produced embryos was compared with the gene expression of in vivo-produced embryos. There was no significant difference found in embryo developmental competence between the Day 9 embryos in BOEC co-culture, FCS and BSA supplements in SOF media. However, differences in gene expression were observed. With respect to gene expression in in vivo and in vitro embryos, BOEC co-culture affected the same genes as did supplementation with FCS and BSA. HSP was the only gene that differed significantly between in vitro and in vivo embryos. When the different in vitro groups were compared, a significant difference between the BOEC co-culture and the FCS supplementation groups due to Glut-1 expression was observed.

scholarly journals FOXE1 represses cell proliferation and Warburg effect by inhibiting HK2 in colorectal cancer

2020 ◽  
Vol 18 (1) ◽  
Author(s):  
Weixing Dai ◽  
Xianke Meng ◽  
Shaobo Mo ◽  
Wenqiang Xiang ◽  
Ye Xu ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cell Proliferation ◽  
Poor Prognosis ◽  
Aerobic Glycolysis ◽  
Lactate Production ◽  
Glucose Consumption ◽  
Underlying Mechanism ◽  
Low Expression

Abstract Background Low expression of FOXE1, a member of Forkhead box (FOX) transcription factor family that plays vital roles in cancers, contributes to poor prognosis of colorectal cancer (CRC) patients. However, the underlying mechanism remains unclear. Materials and methods The effects of FOXE1 on the growth of colon cancer cells and the expression of glycolytic enzymes were investigated in vitro and in vivo. Molecular biological experiments were used to reveal the underlying mechanisms of altered aerobic glycolysis. CRC tissue specimens were used to determine the clinical association of ectopic metabolism caused by dysregulated FOXE1. Results FOXE1 is highly expressed in normal colon tissues compared with cancer tissues and low expression of FOXE1 is significantly associated with poor prognosis of CRC patients. Silencing FOXE1 in CRC cell lines dramatically enhanced cell proliferation and colony formation and promoted glucose consumption and lactate production, while enforced expression of FOXE1 manifested the opposite effects. Mechanistically, FOXE1 bound directly to the promoter region of HK2 and negatively regulated its transcription. Furthermore, the expression of FOXE1 in CRC tissues was negatively correlated with that of HK2. Conclusion FOXE1 functions as a critical tumor suppressor in regulating tumor growth and glycolysis via suppressing HK2 in CRC.

The FOXC1/FBP1 signaling axispromotes tumorigenicity by enhancing the Warburg effect in colorectal cancer.

2017 ◽  
Vol 35 (15_suppl) ◽  
pp. e15123-e15123
Author(s):  
Dawei Li ◽  
Qingguo Li ◽  
Sanjun Cai ◽  
Keping Xie
Keyword(s):  
Colorectal Cancer ◽  
Lactate Production ◽  
Glucose Consumption ◽  
Glycolytic Enzyme ◽  
Prognostic Information ◽  
Aberrant Expression ◽  
Forkhead Box ◽  
Mechanistic Investigation

e15123 Background: Aberrant expression of Forkhead Box transcription factors plays vital roles in the oncogenesis and metastasis of many types of cancer. The purpose of this study is to elucidate the function of Forkhead Box C1(FOXC1) in colorectal cancer (CRC)malignancy maintenance. Methods: FOXC1 expression in CRC specimens was analyzed in the TCGA database and validated by immunohistochemistry using a tissue microarray (TMA). The effect of FOXC1 expression on cancer proliferation and glycolysis was assessed in cells by altering the expression of FOXC1 in vitro and in vivo. Mechanistic investigation was carried out by using cell and molecular biology approaches. Results: FOXC1 was found to be overexpressed in CRC specimens compared with that in the adjacent benign tissues. Univariate survival analyses of the TCGA and validated cohorts showed that high expression of FOXC1 was significantly correlated with shortened patient survival ( P< 0.05). Attenuation of FOXC1 expression inhibited proliferation, clone formation and decreased glucose consumption and lactate production. By contrast, overexpression of FOXC1 had the opposite effect. Furthermore, increased FOXC1 expression downregulated the expression of a key glycolytic enzyme,fructose-1, 6-Bisphosphatase 1 (FBP1). Mechanistically, FOXC1 bound directly to the promoter regions of the FBP1 gene and negatively regulated its transcriptional activity. Aberrant FBP1 expression contributes to CRC tumorigenicity, and decreased FBP1 coupled with increased FOXC1 provided better prognostic information than FOXC1 did alone. Conclusions: The FOXC1/FBP1 axis induces cell proliferation, reprograms the metabolic process in CRC and provides potential prognostic predictors and therapeutic targets for patients with CRC.

scholarly journals Carbohydrate metabolism by murine ovarian follicles and oocytes grown in vitro

Reproduction ◽  
2007 ◽  
Vol 134 (3) ◽  
pp. 415-424 ◽  
Author(s):  
Sarah E Harris ◽  
Iris Adriaens ◽  
Henry J Leese ◽  
Roger G Gosden ◽  
Helen M Picton
Keyword(s):  
Carbohydrate Metabolism ◽  
Culture Media ◽  
Lactate Production ◽  
Glucose Consumption ◽  
Similar Rate ◽  
Metabolic Markers ◽  
Developmental Progression ◽  
Lactate And Pyruvate

Metabolic markers are potentially valuable for assessment of follicle development in vitro. Carbohydrate metabolism of murine preantral follicles grown to maturityover 13 days in vitro has been measured, and metabolism of resulting oocyte–cumulus complexes (OCCs) and denuded oocytes has been compared with in vivo ovulated control counterparts. Spent follicle culture media were analysed for glucose, lactate and pyruvate concentrations. During follicle in vitro growth, glycolysis accounted for a rise from ∼24 to 60% of all glucose consumed. Ovulation induction caused a significant increase in glucose uptake and lactate production by in vitro-grown follicles to 71.7±1.2 and 96.6±4.8 nmoles/day respectively. OCCs grown in vitro had significantly higher rates of glucose consumption and lactate and pyruvate production (110.1± 3.5, 191.8± 8.9 and 31.7± 1.7 pmoles/h respectively) than in vivo ovulated controls (67.4± 8.1, 113.9± 17.1 and 20.2± 4.0 pmoles/h respectively), but a reduced capacity for pyruvate consumption (1.13± 0.06 vs 1.49± 0.06 pmoles/h by in vivo ovulated oocytes). Metabolism of OCCs was affected by the quality of the original follicle. In vitro-grown oocytes had a reduced cytoplasmic volume when compared with controls (168.3± 2.0 vs 199.0± 3.2 proportionately respectively) but a similar rate of metabolism per unit volume. Meiotic status influenced metabolism of both OCCs and denuded oocytes. In conclusion, glucose consumption and lactate production by cultured follicles increased in tandem with developmental progression and were stimulated prior to ovulation. Additionally, the metabolic profiles of in vitro produced OCCs and the oocytes within them are affected by long-term exposure to the culture environment.

scholarly journals Circ_0029803 serves as the sponge of miR-216b-5p to promote the progression of colorectal cancer by regulating SKIL expression

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Linfei Huang ◽  
Lei Zhu ◽  
Sheng Pan ◽  
Jing Xu ◽  
Miao Xie ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Lactate Production ◽  
Glucose Consumption ◽  
Circular Rna ◽  
Inhibitory Effect ◽  
Tumor Xenograft ◽  
Luciferase Reporter ◽  
Migration And Invasion

Abstract Background Circular RNA 0029803 (circ_0029803) was found to be upregulated in colorectal cancer (CRC) tissues, but its function and underlying molecular mechanism are not studied in CRC. Methods The expression levels of circ_0029803, microRNA-216b-5p (miR-216b-5p), and ski-oncogene-like (SKIL) were measured by quantitative real-time polymerase chain reaction (qRT-PCR). RNase R treatment was used to affirm the existence of circ_0029803. Cell proliferation, apoptosis, migration, and invasion were assessed by colony formation, flow cytometry, and Transwell assays, respectively. A glucose and lactate assay kit was used to detect glucose consumption and lactate production. Western blot was applied to analyze the levels of all proteins. Dual-luciferase reporter assay was performed to assess the relationship between miR-216b-5p and circ_0029803 or SKIL. Tumor xenograft models were established to elucidate the effect of circ_0029803 in vivo. Results Circ_0029803 expression was enhanced in CRC tissues and cells, and the 5-year overall survival rate of patients with high circ_0029803 expression was substantially reduced. Circ_0029803 depletion retarded proliferation, migration, invasion, EMT and glycolysis of CRC cells in vitro as well as the tumor growth in vivo. Mechanically, circ_0029803 could serve as miR-216b-5p sponge to regulate its expression, and miR-216b-5p knockdown reversed the inhibition of si-circ_0029803 on the malignant behaviors of CRC cells. Additionally, as the target mRNA of miR-216b-5p, SKIL could counteract the inhibitory effect of miR-216b-5p on the development of CRC cells. Importantly, silencing circ_0029803 reduced SKIL expression via sponging miR-216b-5p. Conclusion Circ_0029803 knockdown hindered proliferation, migration, invasion, EMT, and glycolysis and promoted apoptosis in CRC cells by modulating the miR-216b-5p/SKIL axis.

The mode of action of suramin on the filarial worm Brugia pahangi

Parasitology ◽  
1983 ◽  
Vol 87 (1) ◽  
pp. 29-48 ◽  
Author(s):  
R. E. Howells ◽  
A. M. Mendis ◽  
P. G. Bray
Keyword(s):  
Intestinal Epithelium ◽  
Mode Of Action ◽  
Lactate Production ◽  
Intestinal Lumen ◽  
Ultrastructural Changes ◽  
Brugia Pahangi ◽  
Glycolytic Activity ◽  
Progressive Accumulation

SUMMARYThe mode of action of suramin upon Brugia pahangi has been investigated in vivo and in vitro. The drug was without effect on the glycolytic activity of worms in vitro at 2 × 10−4 M. The lack of effect was correlated with the failure of [14C]suramin to penetrate the worms in vitro. Suramin bound to the surface of worms in vitro presumably by virtue of its polyanionic nature. B. pahangi adults ingested [14C]suramin in vivo but no reduction in the rate of lactate production, of glucose utilization or in the rates of uptake of [14C]glucose, [14C]leucine or [14C]adenosine was observed in worms recovered from jirds between weeks 1 and 5 following 4 daily doses of suramin at 50 mg/kg given intraperitoneally. Worm death occurred between weeks 5 and 7 but this delayed drug effect was not the result of a progressive accumulation of suramin in the worms. Ultrastructural changes were observed in the intestinal epithelium of worms from suramin-treated jirds and parallel observations on worms exposed to Trypan blue in vivo suggest that both polyanionic compounds are restricted to the intestinal lumen of the worms. The evidence presented is consistent with the concept that, in B. pahangi, suramin acts at the surface of the intestinal epithelium and not by primarily inhibiting glucose catabolism or inhibiting phagosome and lysosome fusion as previously demonstrated for bloodstream trypanosomes and mammalian macrophages, respectively

scholarly journals Silencing circSLAMF6 represses cell glycolysis, migration, and invasion by regulating the miR-204-5p/MYH9 axis in gastric cancer under hypoxia

2020 ◽  
Vol 40 (6) ◽  
Author(s):  
Xinhui Fang ◽  
Yangqiu Bai ◽  
Lida Zhang ◽  
Songze Ding
Keyword(s):  
Gastric Cancer ◽  
Lactate Production ◽  
Xenograft Model ◽  
Glucose Consumption ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Glucose Assay

Abstract Background: Gastric cancer (GC) is a malignant tumor of the digestive tract. Hypoxia plays an important role in the development of cancer, including GC. The present study aimed to investigate the role of circular RNA SLAMF6 (circSLAMF6) in the progression of GC under hypoxia. Methods: The expression of circSLAMF6, microRNA-204-5p (miR-204-5p) and myosin heavy chain 9 (MYH9) was measured by quantitative real-time polymerase chain reaction (qRT-PCR). GC cells were maintained under hypoxia (1% O2) for experiments in vitro. Glucose consumption and lactate production were determined by a Glucose Assay Kit and a Lactate Assay Kit, respectively. Levels of all protein were detected by Western blot. Cell migration and invasion were examined by Transwell assay. The interaction between miR-204-5p and circSLAMF6 or MYH9 was analyzed by dual-luciferase reporter and RNA immunoprecipitation (RIP) assays. Murine xenograft model was established to explore the role of circSLAMF6 in vivo. Results: CircSLAMF6 expression was increased in GC cells under hypoxia. Hypoxia promoted glycolysis, migration, and invasion in GC cells, which were reversed by circSLAMF6 knockdown. CircSLAMF6 was validated as a miR-204-5p sponge, and MYH9 was a target of miR-204-5p. Functionally, miR-204-5p inhibitor weakened the inhibition of circSLAMF6 knockdown on GC cell progression under hypoxia. Besides, MYH9 depletion suppressed glycolysis, migration, and invasion in GC cells under hypoxia. Importantly, circSLAMF6 deficiency inhibited tumor growth in vivo by regulating the miR-204-5p/MYH9 axis. Conclusion: CircSLAMF6 was involved in glycolysis, migration, and invasion by regulating the miR-204-5p/MYH9 axis in GC cells under hypoxia.

scholarly journals Regulation of the Elevated T Cell Glycolysis May Alleviate Acute Graft-Versus-Host Disease Post-Allotransplant

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 600-600
Author(s):  
Qi Wen ◽  
Song Yang ◽  
Zhong-Shi Lyu ◽  
Wei-Li Yao ◽  
Yu-Hong Chen ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Metabolic Profile ◽  
Lactate Production ◽  
Glucose Consumption ◽  
Metabolic State ◽  
Glycolytic Activity ◽  
Abnormal Metabolism ◽  
Glycolysis Inhibitor

Background: Acute graft-versus-host disease(aGVHD) remains a major complication following allogeneic hematopoietic stem cell transplantation(allo-HSCT). The pathogenesis of aGVHD is commonly considered to be caused by exaggerated and undesirable immune responses. Metabolism not only provide energy and substrates for T cell growth and survival, but also instruct effector functions, differentiation, and gene expression of T cells. In this regard, the metabolic profile of T cells was reported to play a critical role in the occurrence and development of many immunological disorders such as systemic lupus erythematosus and rheumatoid arthritis. Murine studies found that alloreactive T cells use aerobic glycolysis as the predominant metabolic process to meet activation and proliferation demand after allo-HSCT. However, the metabolic profile of T cells and the approach for regulating T cell metabolism in aGVHD patients remains to be elucidated. Aims: To determine the metabolic state in T cells of patients with aGVHD. Moreover, to investigate the effect of the novel approach targeting the abnormal metabolism in T cells of aGVHD patients, which may provide a potential therapeutic target for aGVHD patients after allo-HSCT. Methods: In this prospective case-control study, a total of 25 patients with aGVHD and 25 matched patients without aGVHD(non-aGVHD) after allo-HSCT were enrolled. T cell subsets were analyzed in aGVHD and non-aGVHD patients by flow cytometry. Th1, Th2, Th17, and Treg cells were identified as CD4+IFN-γ+, CD4+IL-4+, CD4+IL17A+, and CD4+CD25+Foxp3+, respectively. Tc1 and Tc2 cells were identified as CD8+IFN-γ+ and CD8+IL-4+, respectively. In order to determine the metabolic state in T cells of patients with aGVHD and non-aGVHD, the metabolic profile was determined using a Seahorse XF96 Analyzer. The glucose consumption and lactate production rates were detected by glucose assay kit and lactate assay kit. The mitochondrial mass, the mitochondrial membrane potential, the protein expressions for the lipid metabolism enzyme CTP1a and the glycolytic activator PFKFB3 were measured by flow cytometry. To further understand the metabolic state of T cells in aGVHD and non-aGVHD patients and investigate its mechanism, RNA sequencing (RNA-Seq) was performed to analyze the gene expression profiles of T cells. Subsequently, to explore the potential way of targeting the abnormal metabolism in T cells, the glycolysis inhibitor 3-PO was administrated to T cells from aGVHD patients. Results: When compared with T cells in non-aGVHD patients, T cells in aGVHD patients were polarized towards pro-inflammatory T cells, characterized by an elevated proportion of Tc1, Th1 and Th17. Furthermore, T cells isolated from aGVHD patients exhibited higher extracellular acidification rate, as well as the increased glucose consumption rate and lactate production rate compared to those in non-aGVHD patients. Moreover, elevated expression of PFKFB3 was observed in T cells, especially in naïve T cells of aGVHD patients, but oxygen consumption rate, CPT1A, mitochondrial mass or membrane potential showed no significant differences in T cells between aGVHD and non-aGVHD patients. These results implied higher glycolytic activity of T cells in aGVHD patients when compared with those in non-aGVHD patients. Consistent with the increased glycolytic activity observed in T cells from aGVHD patients, the mRNA levels of genes involved in the glycolytic pathway were substantially elevated in T cells of aGVHD patients compared to those in non-aGVHD patients. Importantly, in vitro treatment with glycolysis inhibitor 3-PO improved the activity of T cells derived from aGVHD patients through down-regulating glycolytic activity of T cells. Summary/Conclusion: The current study demonstrated that T cells in aGVHD patients preferentially depend on glycolysis to meet activation and proliferation demands. Furthermore, the activity of T cells from aGVHD patients could be ameliorated by glycolysis inhibitor 3-PO in vitro. Although further validation is required, T cell glycolysis promises to be a novel therapeutic target for aGVHD patients after allo-HSCT. Disclosures No relevant conflicts of interest to declare.

scholarly journals NH4+ triggers the release of astrocytic lactate via mitochondrial pyruvate shunting

2015 ◽  
Vol 112 (35) ◽  
pp. 11090-11095 ◽  
Author(s):  
Rodrigo Lerchundi ◽  
Ignacio Fernández-Moncada ◽  
Yasna Contreras-Baeza ◽  
Tamara Sotelo-Hitschfeld ◽  
Philipp Mächler ◽  
...  
Keyword(s):  
Aerobic Glycolysis ◽  
Hippocampal Slices ◽  
Lactate Production ◽  
Glucose Consumption ◽  
Oxygen Metabolism ◽  
Acute Increase ◽  
Excitatory Neurotransmission ◽  
Acute Hippocampal Slices

Neural activity is accompanied by a transient mismatch between local glucose and oxygen metabolism, a phenomenon of physiological and pathophysiological importance termed aerobic glycolysis. Previous studies have proposed glutamate and K+ as the neuronal signals that trigger aerobic glycolysis in astrocytes. Here we used a panel of genetically encoded FRET sensors in vitro and in vivo to investigate the participation of NH4+, a by-product of catabolism that is also released by active neurons. Astrocytes in mixed cortical cultures responded to physiological levels of NH4+ with an acute rise in cytosolic lactate followed by lactate release into the extracellular space, as detected by a lactate-sniffer. An acute increase in astrocytic lactate was also observed in acute hippocampal slices exposed to NH4+ and in the somatosensory cortex of anesthetized mice in response to i.v. NH4+. Unexpectedly, NH4+ had no effect on astrocytic glucose consumption. Parallel measurements showed simultaneous cytosolic pyruvate accumulation and NADH depletion, suggesting the involvement of mitochondria. An inhibitor-stop technique confirmed a strong inhibition of mitochondrial pyruvate uptake that can be explained by mitochondrial matrix acidification. These results show that physiological NH4+ diverts the flux of pyruvate from mitochondria to lactate production and release. Considering that NH4+ is produced stoichiometrically with glutamate during excitatory neurotransmission, we propose that NH4+ behaves as an intercellular signal and that pyruvate shunting contributes to aerobic lactate production by astrocytes.

Evaluation of 99mTc-Label led Vitamin K4 as an Agent for Testicular Imaging

1991 ◽  
Vol 30 (01) ◽  
pp. 35-39 ◽  
Author(s):  
H. S. Durak ◽  
M. Kitapgi ◽  
B. E. Caner ◽  
R. Senekowitsch ◽  
M. T. Ercan
Keyword(s):  
Soft Tissue ◽  
Room Temperature ◽  
Normal Subjects ◽  
Left Testis ◽  
Wide Range ◽  
Activity Ratios ◽  
The Right ◽  
Radioactivity Levels

Vitamin K4 was labelled with 99mTc with an efficiency higher than 97%. The compound was stable up to 24 h at room temperature, and its biodistribution in NMRI mice indicated its in vivo stability. Blood radioactivity levels were high over a wide range. 10% of the injected activity remained in blood after 24 h. Excretion was mostly via kidneys. Only the liver and kidneys concentrated appreciable amounts of radioactivity. Testis/soft tissue ratios were 1.4 and 1.57 at 6 and 24 h, respectively. Testis/blood ratios were lower than 1. In vitro studies with mouse blood indicated that 33.9 ±9.6% of the radioactivity was associated with RBCs; it was washed out almost completely with saline. Protein binding was 28.7 ±6.3% as determined by TCA precipitation. Blood clearance of 99mTc-l<4 in normal subjects showed a slow decrease of radioactivity, reaching a plateau after 16 h at 20% of the injected activity. In scintigraphic images in men the testes could be well visualized. The right/left testis ratio was 1.08 ±0.13. Testis/soft tissue and testis/blood activity ratios were highest at 3 h. These ratios were higher than those obtained with pertechnetate at 20 min post injection.99mTc-l<4 appears to be a promising radiopharmaceutical for the scintigraphic visualization of testes.

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