scholarly journals Glycolytic flux-signaling controls mouse embryo mesoderm development

2021 ◽  
Author(s):  
Hidenobu Miyazawa ◽  
Marteinn T. Snaebjornsson ◽  
Nicole Prior ◽  
Eleni Kafkia ◽  
Henrik M Hammarén ◽  
...  
Keyword(s):  
Soluble Fraction ◽  
Protein Localization ◽  
Proteome Analysis ◽  
Glycolytic Flux ◽  
Mesoderm Development ◽  
Subcellular Protein Localization ◽  
Fructose 1,6 Bisphosphate ◽  
Post Implantation ◽  
Medium Supplementation

How cellular metabolic state impacts cellular programs is a fundamental, unresolved question. Here we investigated how glycolytic flux impacts embryonic development, using presomitic mesoderm (PSM) patterning as the experimental model. First, we identified fructose 1,6-bisphosphate (FBP) as an in vivo sentinel metabolite that mirrors glycolytic flux within PSM cells of post-implantation mouse embryos. We found that medium-supplementation with FBP, but not with other glycolytic metabolites, such as fructose 6-phosphate and 3-phosphoglycerate, impaired mesoderm segmentation. To genetically manipulate glycolytic flux and FBP levels, we generated a mouse model enabling the conditional overexpression of dominant active, cytoplasmic Pfkfb3 (cytoPfkfb3). Overexpression of cytoPfkfb3 indeed led to increased glycolytic flux/FBP levels and caused an impairment of mesoderm segmentation, paralleled by the downregulation of Wnt-signaling, reminiscent of the effects seen upon FBP-supplementation. To probe for mechanisms underlying glycolytic flux-signaling, we performed subcellular proteome analysis and revealed that cytoPfkfb3 overexpression altered subcellular localization of certain proteins, including glycolytic enzymes, in PSM cells. Specifically, we revealed that FBP supplementation caused depletion of Pfkl and Aldoa from the nuclear-soluble fraction. Combined, we propose that FBP functions as a flux-signaling metabolite connecting glycolysis and PSM patterning, potentially through modulating subcellular protein localization.

Studies with GIP/Ins cells indicate secretion by gut K cells is KATPchannel independent

2003 ◽  
Vol 284 (5) ◽  
pp. E988-E1000 ◽  
Author(s):  
Song Yan Wang ◽  
Maggie M.-Y. Chi ◽  
Lin Li ◽  
Kelle H. Moley ◽  
Burton M. Wice
Keyword(s):  
Tricarboxylic Acid Cycle ◽  
Calcium Mobilization ◽  
Enteroendocrine Cells ◽  
Glycolytic Flux ◽  
K Cells ◽  
Antibody Staining ◽  
Methyl Pyruvate ◽  
Fructose 1,6 Bisphosphate ◽  
Insulinoma Cells

K cells are a subpopulation of enteroendocrine cells that secrete glucose-dependent insulinotropic polypeptide (GIP), a hormone that promotes glucose homeostasis and obesity. Therefore, it is important to understand how GIP secretion is regulated. GIP-producing (GIP/Ins) cell lines secreted hormones in response to many GIP secretagogues except glucose. In contrast, glyceraldehyde and methyl pyruvate stimulated hormone release. Measurements of intracellular glucose 6-phosphate, fructose 1,6-bisphosphate, and pyruvate levels, as well as glycolytic flux, in glucose-stimulated GIP/Ins cells indicated that glycolysis was not impaired. Analogous results were obtained using glucose-responsive MIN6 insulinoma cells. Citrate levels increased similarly in glucose-treated MIN6 and GIP/Ins cells. Thus pyruvate entered the tricarboxylic acid cycle. Glucose and methyl pyruvate stimulated 1.4- and 1.6-fold increases, respectively, in the ATP-to-ADP ratio in GIP/Ins cells. Glyceraldehyde profoundly reduced, rather than increased, ATP/ADP. Thus nutrient-regulated secretion is independent of the ATP-dependent potassium (KATP) channel. Antibody staining of mouse intestine demonstrated that enteroendocrine cells producing GIP, glucagon-like peptide-1, CCK, or somatostatin do not express detectable levels of inwardly rectifying potassium (Kir) 6.1 or Kir 6.2, indicating that release of these hormones in vivo may also be KATPchannel independent. Conversely, nearly all cells expressing chromogranin A or substance P and ∼50% of the cells expressing secretin or serotonin exhibited Kir 6.2 staining. Compounds that activate calcium mobilization were potent secretagogues for GIP/Ins cells. Secretion was only partially inhibited by verapamil, suggesting that calcium mobilization from intracellular and extracellular sources, independent from KATPchannels, regulates secretion from some, but not all, subpopulations of enteroendocrine cells.

scholarly journals The regulatory principles of glycolysis in erythrocytes in vivo and in vitro. A minimal comprehensive model describing steady states, quasi-steady states and time-dependent processes

1976 ◽  
Vol 154 (2) ◽  
pp. 449-469 ◽  
Author(s):  
T A. Rapoport ◽  
R Heinrich ◽  
S M. Rapoport
Keyword(s):  
Steady State ◽  
Atp Synthesis ◽  
Steady States ◽  
Time Dependent ◽  
Glycolytic Flux ◽  
Stable Steady State ◽  
Time Interval ◽  
Fructose 1,6 Bisphosphate

A simple mathematical model for glycolysis in erythrocytes is presented which takes into account ATP synthesis and consumption. The system is described by four ordinary differential equations. Conditions in vivo are described by a stable steady state. The model predicts correctly the metabolite concentrations found in vivo. The parameters involved are in agreement with data on the separate steps. The metabolite changes found in pyruvate kinase-deficient erythrocytes and the species variations among erythrocytes from different animals are described satisfactorily. The roles of the enzymes in the control of metabolites and glycolytic flux are expressed in the form of a control matrix and control strengths [R. Heinrich & T.A. Rapoport (1974) Eur. J. Biochem. 42, 89-95] respectively. Erythrocytes from various species are shown to be adapted to a maximal ATP-consumption rate. The calculated eigenvalues reveal the pronounced time-hierarchy of the glycolytic reactions. Owing to the slowness of the 2,3-bisphospho-glycerate phosphatase reaction, quasi-steady states occur during the time-interval of about 0.5-2h incubation, which are defined by perturbed 2,3-bisphosphoglycerate concentrations. The theoretical predictions agree with experimental data. In the quasi-steady state the flux control is exerted almost entirely by the hexokinase-phosphofructokinase system. The model describes satisfactorily the time-dependent changes after addition of glucose to starved erythrocytes. The theoretical consequences are discussed of the conditions in vitro with lactate accumulation and the existence of a time-independent conservation quantity for the oxidized metabolites. Even in this closed system quasi-steady states occur which are characterized by approximately constant concentrations of all glycolytic metabolites except for the accumulation of lactate, fructose 1,6-bisphosphate and triose phosphate.

scholarly journals A synthetic RNA-based biosensor for fructose-1,6-bisphosphate that reports glycolytic flux

2020 ◽  
Author(s):  
Alvaro D. Ortega ◽  
Vakil Takhaveev ◽  
Silke Bonsing-Vedelaar ◽  
Yi Long ◽  
Neus Mestre-Farràs ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
Hammerhead Ribozyme ◽  
Single Cells ◽  
Glycolytic Flux ◽  
Metabolic Heterogeneity ◽  
Fructose 1,6 Bisphosphate ◽  
Glycolytic Rate ◽  
Selection Of

ABSTRACTMetabolic heterogeneity, the occurrence of different metabolic phenotypes among cells, represents a key challenge in health and biotechnology. To unravel its molecular basis, tools probing metabolism of single cells are needed. While RNA devices harbor huge potential for the development of such tools, until today, it is challenging to create in vivo-functional sensors for any given metabolite. Here, we developed from scratch an RNA-based sensor for fructose-1,6-bisphosphate (FBP), a doubly phosphorylated intermediate of glycolysis. Starting from in vitro selection of an RNA aptamer and its structural analyses, we developed libraries of RNA-based regulatory devices with this aptamer and the hammerhead ribozyme as an actuator. Through FACS-seq-based high-throughput screening in yeast, we identified in vivo-functional FBP-sensing devices that generate fluorescent readout dependent on intracellular FBP concentration. As FBP reports the flux through glycolysis, the developed RNA device can be used to sense the glycolytic rate in single cells, offering unprecedented possibilities to investigate the causes of metabolic heterogeneity.

scholarly journals Influence of calcium ion-modified implant surfaces in protein adsorption and implant integration

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Eduardo Anitua ◽  
Andreia Cerqueira ◽  
Francisco Romero-Gavilán ◽  
Iñaki García-Arnáez ◽  
Cristina Martinez-Ramos ◽  
...  
Keyword(s):  
Protein Adsorption ◽  
Titanium Implant ◽  
Volume Density ◽  
Bone Implant ◽  
The Mean ◽  
Intervention Costs ◽  
Medial Section ◽  
Post Implantation

Abstract Background Calcium (Ca) is a well-known element in bone metabolism and blood coagulation. Here, we investigate the link between the protein adsorption pattern and the in vivo responses of surfaces modified with calcium ions (Ca-ion) as compared to standard titanium implant surfaces (control). We used LC–MS/MS to identify the proteins adhered to the surfaces after incubation with human serum and performed bilateral surgeries in the medial section of the femoral condyles of 18 New Zealand white rabbits to test osseointegration at 2 and 8 weeks post-implantation (n=9). Results Ca-ion surfaces adsorbed 181.42 times more FA10 and 3.85 times less FA12 (p<0.001), which are factors of the common and the intrinsic coagulation pathways respectively. We also detected differences in A1AT, PLMN, FA12, KNG1, HEP2, LYSC, PIP, SAMP, VTNC, SAA4, and CFAH (p<0.01). At 2 and 8 weeks post-implantation, the mean bone implant contact (BIC) with Ca-ion surfaces was respectively 1.52 and 1.25 times higher, and the mean bone volume density (BVD) was respectively 1.35 and 1.13 times higher. Differences were statistically significant for BIC at 2 and 8 weeks and for BVD at 2 weeks (p<0.05). Conclusions The strong thrombogenic protein adsorption pattern at Ca-ion surfaces correlated with significantly higher levels of implant osseointegration. More effective implant surfaces combined with smaller implants enable less invasive surgeries, shorter healing times, and overall lower intervention costs, especially in cases of low quantity or quality of bone.

scholarly journals A synthetic RNA-based biosensor for fructose-1,6-bisphosphate that reports glycolytic flux

2021 ◽  
Author(s):  
Alvaro Darío Ortega ◽  
Vakil Takhaveev ◽  
Silke Roelie Vedelaar ◽  
Yi Long ◽  
Neus Mestre-Farràs ◽  
...  
Keyword(s):  
Glycolytic Flux ◽  
Fructose 1,6 Bisphosphate

An Interlaboratory Evaluation of Five Pairs of Teratogens and Non-teratogens in Post-implantation Rat Embryo Culture

1996 ◽  
Vol 24 (2) ◽  
pp. 201-209
Author(s):  
Aldert H. Piersma ◽  
Rudolf Bechter ◽  
Nathalie Krafft ◽  
Beat P. Schmid ◽  
Jeanne Stadler ◽  
...  
Keyword(s):  
Embryo Culture ◽  
Developmental Toxicity ◽  
Culture Method ◽  
Rat Embryo ◽  
Good Correspondence ◽  
Substituted Pyridines ◽  
Phthalic Ester ◽  
Rat Embryo Culture ◽  
Post Implantation

The usefulness of the post-implantation rat embryo culture method in screening xenobiotic compounds for developmental toxicity was validated in four laboratories with five pairs of compounds. This approach was chosen to provide information on the interlaboratory reproducibility of the results and to compare the effects of chemical analogues in embryo culture. By testing analogous compounds which are known to have different embryotoxic potencies in vivo, the discriminating power of the embryo culture method for the compound classes under study could be optimally assessed. The classes selected for testing were triazole antifungals, phthalic ester metabolites, substituted pyridines, sulphonamides and methylated xanthines. In summary, it was possible to distinguish between the compounds in three of the pairs, it was not possible to discriminate between the compounds of one pair, and it was possible to discriminate between the compounds of the other pair at two out of the four laboratories. The embryo culture results generally show a good correspondence with the embryotoxic properties of the compounds tested in vivo, although the embryo culture method appeared to be able to discriminate between only some of the pairs of chemical analogues. Some discrepancies may have arisen among the laboratories, because of methodological differences. These results suggest that the post-implantation rat embryo culture method may be a useful tool for screening xenobiotics within classes of compounds known to interfere with embryogenesis during the period of development represented in culture.

scholarly journals Essential role of glucose transporter GLUT3 for post-implantation embryonic development

2008 ◽  
Vol 200 (1) ◽  
pp. 23-33 ◽  
Author(s):  
S Schmidt ◽  
A Hommel ◽  
V Gawlik ◽  
R Augustin ◽  
N Junicke ◽  
...  
Keyword(s):  
Essential Role ◽  
Glucose Transporter ◽  
Growth Arrest ◽  
Complete Loss ◽  
Transporter Gene ◽  
Wild Type ◽  
Post Implantation ◽  
Time Point

Deletion of glucose transporter geneSlc2a3(GLUT3) has previously been reported to result in embryonic lethality. Here, we define the exact time point of growth arrest and subsequent death of the embryo.Slc2a3−/−morulae and blastocysts developed normally, implantedin vivo, and formed egg-cylinder-stage embryos that appeared normal until day 6.0. At day 6.5, apoptosis was detected in the ectodermal cells ofSlc2a3−/−embryos resulting in severe disorganization and growth retardation at day 7.5 and complete loss of embryos at day 12.5. GLUT3 was detected in placental cone, in the visceral ectoderm and in the mesoderm of 7.5-day-old wild-type embryos. Our data indicate that GLUT3 is essential for the development of early post-implanted embryos.

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