149 research outputs found
Behavior of a Metabolic Cycling Population at the Single Cell Level as Visualized by Fluorescent Gene Expression Reporters
BACKGROUND: During continuous growth in specific chemostat cultures, budding yeast undergo robust oscillations in oxygen consumption that are accompanied by highly periodic changes in transcript abundance of a majority of genes, in a phenomenon called the Yeast Metabolic Cycle (YMC). This study uses fluorescent reporters of genes specific to different YMC phases in order to visualize this phenomenon and understand the temporal regulation of gene expression at the level of individual cells within the cycling population. METHODOLOGY: Fluorescent gene expression reporters for different phases of the YMC were constructed and stably integrated into the yeast genome. Subsequently, these reporter-expressing yeast were used to visualize YMC dynamics at the individual cell level in cultures grown in a chemostat or in a microfluidics platform under varying glucose concentrations, using fluorescence microscopy and quantitative Western blots. CONCLUSIONS: The behavior of single cells within a metabolic cycling population was visualized using phase-specific fluorescent reporters. The reporters largely recapitulated genome-specified mRNA expression profiles. A significant fraction of the cell population appeared to exhibit basal expression of the reporters, supporting the hypothesis that there are at least two distinct subpopulations of cells within the cycling population. Although approximately half of the cycling population initiated cell division in each permissive window of the YMC, metabolic synchrony of the population was maintained. Using a microfluidics platform we observed that low glucose concentrations appear to be necessary for metabolic cycling. Lastly, we propose that there is a temporal window in the oxidative growth phase of the YMC where the cycling population segregates into at least two subpopulations, one which will enter the cell cycle and one which does not
Changes in normal appearing spinal cord in multiple sclerosis: another brick in the wall
A conscious mouse model of gastric ileus using clinically relevant endpoints
BACKGROUND: Gastric ileus is an unsolved clinical problem and current treatment is limited to supportive measures. Models of ileus using anesthetized animals, muscle strips or isolated smooth muscle cells do not adequately reproduce the clinical situation. Thus, previous studies using these techniques have not led to a clear understanding of the pathophysiology of ileus. The feasibility of using food intake and fecal output as simple, clinically relevant endpoints for monitoring ileus in a conscious mouse model was evaluated by assessing the severity and time course of various insults known to cause ileus. METHODS: Delayed food intake and fecal output associated with ileus was monitored after intraperitoneal injection of endotoxin, laparotomy with bowel manipulation, thermal injury or cerulein induced acute pancreatitis. The correlation of decreased food intake after endotoxin injection with gastric ileus was validated by measuring gastric emptying. The effect of endotoxin on general activity level and feeding behavior was also determined. Small bowel transit was measured using a phenol red marker. RESULTS: Each insult resulted in a transient and comparable decrease in food intake and fecal output consistent with the clinical picture of ileus. The endpoints were highly sensitive to small changes in low doses of endotoxin, the extent of bowel manipulation, and cerulein dose. The delay in food intake directly correlated with delayed gastric emptying. Changes in general activity and feeding behavior were insufficient to explain decreased food intake. Intestinal transit remained unchanged at the times measured. CONCLUSION: Food intake and fecal output are sensitive markers of gastric dysfunction in four experimental models of ileus. In the mouse, delayed gastric emptying appears to be the major cause of the anorexic effect associated with ileus. Gastric dysfunction is more important than small bowel dysfunction in this model. Recovery of stomach function appears to be simultaneous to colonic recovery
Genome-Scale Reconstruction and Analysis of the Pseudomonas putida KT2440 Metabolic Network Facilitates Applications in Biotechnology
A cornerstone of biotechnology is the use of microorganisms for the efficient
production of chemicals and the elimination of harmful waste.
Pseudomonas putida is an archetype of such microbes due to
its metabolic versatility, stress resistance, amenability to genetic
modifications, and vast potential for environmental and industrial applications.
To address both the elucidation of the metabolic wiring in P.
putida and its uses in biocatalysis, in particular for the production
of non-growth-related biochemicals, we developed and present here a genome-scale
constraint-based model of the metabolism of P. putida KT2440.
Network reconstruction and flux balance analysis (FBA) enabled definition of the
structure of the metabolic network, identification of knowledge gaps, and
pin-pointing of essential metabolic functions, facilitating thereby the
refinement of gene annotations. FBA and flux variability analysis were used to
analyze the properties, potential, and limits of the model. These analyses
allowed identification, under various conditions, of key features of metabolism
such as growth yield, resource distribution, network robustness, and gene
essentiality. The model was validated with data from continuous cell cultures,
high-throughput phenotyping data, 13C-measurement of internal flux
distributions, and specifically generated knock-out mutants. Auxotrophy was
correctly predicted in 75% of the cases. These systematic analyses
revealed that the metabolic network structure is the main factor determining the
accuracy of predictions, whereas biomass composition has negligible influence.
Finally, we drew on the model to devise metabolic engineering strategies to
improve production of polyhydroxyalkanoates, a class of biotechnologically
useful compounds whose synthesis is not coupled to cell survival. The solidly
validated model yields valuable insights into genotype–phenotype
relationships and provides a sound framework to explore this versatile bacterium
and to capitalize on its vast biotechnological potential
Developmental stage-dependent metabolic regulation during meiotic differentiation in budding yeast
A case of late onset leukoencephalopathy with cerebral calcifications and cysts in a 59-year-old woman
Background: Leukoencephalopathy with cerebral calcifications and cysts (LCC) is a recently described, very rare entity, clinically characterized by progressive neurological deficits such as cognitive decline, epileptic seizures, pyramidal, extrapyramidal and cerebellar symptoms/signs. With the exception of two patients with adult onset, in all previously described cases symptoms onset occurred between early infancy and adolescence.
Results: We report a case of late onset LCC in a 59-year-old woman presenting with urinary and fecal incontinence and behavioural changes, then rapid progression with hemianopia, hemiparesis, ataxia and cognitive decline. Extensive work-up was performed, including brain magnetic resonance imaging, magnetic resonance spectroscopy, cyst fluid analysis and brain biopsy, confirming the final diagnosis of LCC.
Conclusion: Our case supports the existence of a late onset adult form of LCC
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