Administration of Escherichia coli endotoxin to rat increases liver mass and hepatocyte volume in vivo.
Ontology highlight
ABSTRACT:
SUBMITTER: Qian D
PROVIDER: S-EPMC1216932 | biostudies-other | 1996 Jan
REPOSITORIES: biostudies-other
Similar Datasets
Project description:BackgroundLipopolysaccharide (LPS), also referred to as endotoxin, is the major constituent of the outer leaflet of the outer membrane of virtually all Gram-negative bacteria. The lipid A moiety, which anchors the LPS molecule to the outer membrane, acts as a potent agonist for Toll-like receptor 4/myeloid differentiation factor 2-mediated pro-inflammatory activity in mammals and, thus, represents the endotoxic principle of LPS. Recombinant proteins, commonly manufactured in Escherichia coli, are generally contaminated with endotoxin. Removal of bacterial endotoxin from recombinant therapeutic proteins is a challenging and expensive process that has been necessary to ensure the safety of the final product.ResultsAs an alternative strategy for common endotoxin removal methods, we have developed a series of E. coli strains that are able to grow and express recombinant proteins with the endotoxin precursor lipid IVA as the only LPS-related molecule in their outer membranes. Lipid IVA does not trigger an endotoxic response in humans typical of bacterial LPS chemotypes. Hence the engineered cells themselves, and the purified proteins expressed within these cells display extremely low endotoxin levels.ConclusionsThis paper describes the preparation and characterization of endotoxin-free E. coli strains, and demonstrates the direct production of recombinant proteins with negligible endotoxin contamination.
| S-EPMC4404585 | biostudies-literature
Project description:Despite years of positive animal data, hepatocyte growth factor (HGF) has never been developed into a useful pharmaceutical, primarily due to its poor pharmacological properties. CM1021 is a fusion protein containing the K1 loop of HGF and the human IgG1 Fc region. The experiments described here demonstrate that CM1021 has the biological properties of HGF and the pharmacological properties of a monoclonal antibody. CM1021 stimulates scattering and branching morphogenesis in MDCK cells and stimulates liver growth in vivo. Unlike HGF, it is available via intraperitoneal injection and has an estimated half-life similar to an antibody.
| S-EPMC8683692 | biostudies-literature
Project description:In Salmonella enterica, PmrD is a connector protein that links the two-component systems PhoP-PhoQ and PmrA-PmrB. While Escherichia coli encodes a PmrD homolog, it is thought to be incapable of connecting PhoPQ and PmrAB in this organism due to functional divergence from the S. enterica protein. However, our laboratory previously observed that low concentrations of Mg(2+), a PhoPQ-activating signal, leads to the induction of PmrAB-dependent lipid A modifications in wild-type E. coli (C. M. Herrera, J. V. Hankins, and M. S. Trent, Mol Microbiol 76:1444-1460, 2010, http://dx.doi.org/10.1111/j.1365-2958.2010.07150.x). These modifications include phosphoethanolamine (pEtN) and 4-amino-4-deoxy-l-arabinose (l-Ara4N), which promote bacterial resistance to cationic antimicrobial peptides (CAMPs) when affixed to lipid A. Here, we demonstrate that pmrD is required for modification of the lipid A domain of E. coli lipopolysaccharide (LPS) under low-Mg(2+) growth conditions. Further, RNA sequencing shows that E. coli pmrD influences the expression of pmrA and its downstream targets, including genes coding for the modification enzymes that transfer pEtN and l-Ara4N to the lipid A molecule. In line with these findings, a pmrD mutant is dramatically impaired in survival compared with the wild-type strain when exposed to the CAMP polymyxin B. Notably, we also reveal the presence of an unknown factor or system capable of activating pmrD to promote lipid A modification in the absence of the PhoPQ system. These results illuminate a more complex network of protein interactions surrounding activation of PhoPQ and PmrAB in E. coli than previously understood.
| S-EPMC4356761 | biostudies-literature
Project description:A primary role of the liver is to regulate whole body glucose homeostasis. Glucokinase (GCK) is the main hexokinase (HK) expressed in hepatocytes and functions to phosphorylate the glucose that enters via GLUT transporters to become glucose-6-phosphate (G6P), which subsequently commits glucose to enter downstream anabolic and catabolic pathways. In the recent years, hexokinase domain-containing-1 (HKDC1), a novel 5th HK, has been characterized by our group and others. Its expression profile varies but has been identified to have low basal expression in normal liver but increases during states of stress including pregnancy, nonalcoholic fatty liver disease (NAFLD), and liver cancer. Here, we have developed a stable overexpression model of hepatic HKDC1 in mice to examine its effect on metabolic regulation. We found that HKDC1 overexpression, over time, causes impaired glucose homeostasis in male mice and shifts glucose metabolism towards anabolic pathways with an increase in nucleotide synthesis. Furthermore, we observed these mice to have larger liver sizes due to greater hepatocyte proliferative potential and cell size, which in part, is mediated via yes-associated protein (YAP) signaling.
| S-EPMC10192376 | biostudies-literature
Project description:All living cells employ an array of different mechanisms to help them survive changes in extra cellular osmotic pressure. The difference in the concentration of chemicals in a bacterium's cytoplasm and the external environment generates an osmotic pressure that inflates the cell. It is thought that the bacterium Escherichia coli use a number of interconnected systems to adapt to changes in external pressure, allowing them to maintain turgor and live in surroundings that range more than two-hundred-fold in external osmolality. Here, we use fluorescence imaging to make the first measurements of cell volume changes over time during hyperosmotic shock and subsequent adaptation on a single cell level in vivo with a time resolution on the order of seconds. We directly observe two previously unseen phases of the cytoplasmic water efflux upon hyperosmotic shock. Furthermore, we monitor cell volume changes during the post-shock recovery and observe a two-phase response that depends on the shock magnitude. The initial phase of recovery is fast, on the order of 15-20 min and shows little cell-to-cell variation. For large sucrose shocks, a secondary phase that lasts several hours adds to the recovery. We find that cells are able to recover fully from shocks as high as 1 Osmol/kg using existing systems, but that for larger shocks, protein synthesis is required for full recovery.
| S-EPMC3325977 | biostudies-literature
Project description:Escherichia coli is a widely used platform for metabolic engineering due to its fast growth and well-established engineering techniques. However, there has been a demand for faster-growing E. coli for higher production of desired substances. Here, to increase the growth of E. coli cells, we optimized the expression level of Hfq protein, which plays an essential role in stress responses. Six variants of the hfq gene with a different ribosome binding site sequence and thereby a different expression level were constructed. When the Hfq expression level was optimized in DH5α, its growth rate was increased by 12.1% and its cell density was also increased by 4.5%. RNA-seq and network analyses revealed the upregulation of stress response genes and metabolic genes, which increases the tolerance against pH changes. When the same strategy was applied to five other E. coli strains (BL21 (DE3), JM109, TOP10, W3110, and MG1655), all their growth rates were increased by 18-94% but not all their densities were increased (- 12 - + 32%). In conclusion, the Hfq expression optimization can increase cell growth rate and probably their cell densities as well. Since the hfq gene is highly conserved across bacterial species, the same strategy could be applied to other bacterial species to construct faster-growing strains.
| S-EPMC7890833 | biostudies-literature
Project description:Background.Other than numerous experimental data assessing phage therapy efficacy, questions regarding safety of this approach are not sufficiently addressed. In particular, as phages can kill bacterial cells within <10 minutes, the associated endotoxin release (ER) in severe infections caused by gram-negative bacteria could be a matter of concern.Methods.Two therapeutic virulent phages and 4 reference antibiotics were studied in vitro for their ability to kill 2 pathogenic strains of Escherichia coli and generate an ER. The early interaction (first 3 hours) between these actors was assessed over time by studying the instantaneous cell viability, the colony-forming unit count, the concentration of free endotoxin released, and the cell morphology under light microscope.Results.While β-lactams have a relatively slow effect, both tested phages, as well as amikacin, were able to rapidly abolish the bacterial growth. Even when considering the fastest phage (cell lysis in 9 minutes), the concentrations of phage-induced ER never reached the highest values, which were recorded with antibiotic treatments. Cumulative concentrations of endotoxin over time in phage-treated conditions were lower than those observed with β-lactams and close to those observed with amikacin. Whereas β-lactams were responsible for strong cell morphology changes (spheroplast with imipenem, filamentous cells with cefoxitin and ceftriaxone), amikacin and phages did not modify cell shape but produced intracellular inclusion bodies.Conclusions.This work provides important and comforting data regarding the safety of phage therapy. Therapeutically relevant phages, with their low endotoxin release profile and fast bactericidal effect, are not inferior to β-lactams.
| S-EPMC5434335 | biostudies-literature
Project description:Changes in the milk microbiota during the course of mastitis are due to the nature of a sporadic occurring disease difficult to study. In this study we experimentally induced mastitis by infusion of Escherichia coli endotoxins in one udder quarter each of nine healthy lactating dairy cows and assessed the bacteriological dynamics and the milk microbiota at four time points before and eight time points after infusion. As control, saline was infused in one udder quarter each of additionally nine healthy cows that followed the same sampling protocol. The milk microbiota was assessed by sequencing of the 16 S rRNA gene and a range of positive and negative controls were included for methodological evaluation. Two different data filtration models were used to identify and cure data from contaminating taxa. Endotoxin infused quarters responded with transient clinical signs of inflammation and increased SCC while no response was observed in the control cows. In the milk microbiota data no response to inflammation was identified. The data analysis of the milk microbiota was largely hampered by laboratory and reagent contamination. Application of the filtration models caused a marked reduction in data but did not reveal any associations with the inflammatory reaction. Our results indicate that the microbiota in milk from healthy cows is unaffected by inflammation.
| S-EPMC10251687 | biostudies-literature
Project description:In Gram-negative bacteria, lipopolysaccharide (LPS) contributes to the robust permeability barrier of the outer membrane, preventing entry of toxic molecules such as antibiotics. Mutations in lptD, the beta-barrel component of the LPS transport and assembly machinery, compromise LPS assembly and result in increased antibiotic sensitivity. Here, we report rare vancomycin-resistant suppressors that improve barrier function of a subset of lptD mutations. We find that all seven suppressors analyzed mapped to the essential gene cdsA, which is responsible for the conversion of phosphatidic acid to CDP-diacylglycerol in phospholipid biosynthesis. These cdsA mutations cause a partial loss of function and, as expected, accumulate phosphatidic acid. We show that this suppression is not confined to mutations that cause defects in outer membrane biogenesis but rather that these cdsA mutations confer a general increase in vancomycin resistance, even in a wild-type cell. We use genetics and quadrupole time of flight (Q-TOF) liquid chromatography-mass spectrometry (LC-MS) to show that accumulation of phosphatidic acid by means other than cdsA mutations also increases resistance to vancomycin. We suggest that increased levels of phosphatidic acid change the physical properties of the outer membrane to impede entry of vancomycin into the periplasm, hindering access to its target, an intermediate required for the synthesis of the peptidoglycan cell wall.
| S-EPMC4135694 | biostudies-literature
Project description:Mistranslation is typically deleterious for cells, although specific mistranslated proteins can confer a short-term benefit in a particular environment. However, given its large overall cost, the prevalence of high global mistranslation rates remains puzzling. Altering basal mistranslation levels of Escherichia coli in several ways, we show that generalized mistranslation enhances early survival under DNA damage, by rapidly activating the SOS response. Mistranslating cells maintain larger populations after exposure to DNA damage, and thus have a higher probability of sampling critical beneficial mutations. Both basal and artificially increased mistranslation increase the number of cells that are phenotypically tolerant and genetically resistant under DNA damage; they also enhance survival at high temperature. In contrast, decreasing the normal basal mistranslation rate reduces cell survival. This wide-ranging stress resistance relies on Lon protease, which is revealed as a key effector that induces the SOS response in addition to alleviating proteotoxic stress. The new links between error-prone protein synthesis, DNA damage, and generalised stress resistance indicate surprising coordination between intracellular stress responses and suggest a novel hypothesis to explain high global mistranslation rates.
| S-EPMC7082066 | biostudies-literature