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At least 271 records · Page 15Linked to original sources
PubMed1996

DNA ligase I is required for fetal liver erythropoiesis but is not essential for mammalian cell viability.

Four distinct DNA ligase activities (I-IV) have been identified within mammalian cells. Evidence has indicated that DNA ligase I is central to DNA replication, as well as being involved in DNA repair processes. A patient with altered DNA ligase I displayed a phenotype similar to Bloom's syndrome, being immunodeficient, growth retarded and predisposed to cancer. Fibroblasts isolated from this patient (46BR) exhibited abnormal lagging strand synthesis and repair deficiency. It has been reported that DNA ligase I is essential for cell viability, but here we show that cells lacking DNA ligase I are in fact viable. Using gene targeting in embryonic stem (ES) cells, we have produced DNA ligase I-deficient mice. Embryos develop normally to mid-term when haematopoiesis usually switches to the fetal liver. Thereupon acute anaemia develops, despite the presence of erythroid-committed progenitor cells in the liver. Thus DNA ligase I is required for normal development, but is not essential for replication. Hence a previously unsuspected redundancy must exist between mammalian DNA ligases.

Animals↗
PubMed2002

Changes in macrophage morphology and prolonged cell viability following exposure to polyethylene particulate in vitro.

The interaction of macrophages and ultra-high molecular weight polyethylene (PE) wear plays an important role in perpetuating chronic inflammation at the bone implant interface, leading to peri-implant osteolysis and mechanical failure of the implant. A model to study the interaction of human mature macrophages with orthopaedic biomaterial wear has been previously developed. With the use of the model, in this study, the mature human monocyte-derived macrophages (MDMs) were observed with light, fluorescent, and scanning electron microscopy (SEM), as well as transmission electron microscopy (TEM). The cell viability was investigated using calcein and ethidium staining. Following exposure to PE particulate, the morphology of the human MDMs was heterogeneous: rounded, flattened, and elongated. There was no morphological evidence of cytotoxicity or apoptosis. The MDM viability was not influenced by phagocytosis of PE particulate in a negative fashion. In fact, more prolonged cell viability was observed in the human MDMs exposed to PE particulate when compared to controls.

Arthroplasty, Replacement↗
PubMed1997

When does the lung die? Kfc, cell viability, and adenine nucleotide changes in the circulation-arrested rat lung.

Lungs harvested from cadaveric circulation-arrested donors may increase the donor pool for lung transplantation. To determine the degree and time course of ischemia-reperfusion injury, we evaluated the effect of O2 ventilation on capillary permeability [capillary filtration coefficient (Kfc)], cell viability, and total adenine nucleotide (TAN) levels in in situ circulation-arrested rat lungs. Kfc increased with increasing postmortem ischemic time (r = 0.88). Lungs ventilated with O2 1 h postmortem had similar Kfc and wet-to-dry ratios as controls. Nonventilated lungs had threefold (P < 0.05) and sevenfold (P < 0.0001) increases in Kfc at 30 and 60 min postmortem compared with controls. Cell viability decreased in all groups except for 30-min postmortem O2-ventilated lungs. TAN levels decreased with increasing ischemic time, particularly in nonventilated lungs. Loss of adenine nucleotides correlated with increasing Kfc values (r = 0.76). This study indicates that lungs retrieved 1 h postmortem may have normal Kfc with preharvest O2 ventilation. The relationship between Kfc and TAN suggests that vascular permeability may be related to lung TAN levels.

Adenine Nucleotides↗
PubMed2004

Impact of ethanol and acetaldehyde on DNA and cell viability of cultured neurones.

Ethanol consumption has long been associated with brain damage. However, the mechanism underlying this deleterious effect remains unclear. Among different hypotheses, acetaldehyde is regarded by certain authors as playing a major role in the expression of ethanol toxicity, but there are still some uncertainties about the exact nature of its implication. We therefore tried to characterize the profile of the alterations of neuronal viability and DNA integrity obtained after either a direct exposure to ethanol or to acetaldehyde. Ethanol at concentrations within the range of blood alcohol levels in intoxicated humans (< or = 100 mmol/L) induced DNA alterations without any apparent effect on cell viability. Acetaldehyde (< or = 1000 micromol/L) can also induce DNA alterations but with a different profile of the DNA cellular alterations. The comparison between the distributions of the comet tail DNA indicated that ethanol induced strong breaks (tail DNA > or = 60 a.u.) generation whereas acetaldehyde rather induced lower breaks (20 < or = tail DNA < or = 50 a.u.) formation but affecting a greater number of neurones. Acetaldehyde had thus a different genotoxic potential which may suggest a different mode of action or a different cellular target. Furthermore, when a single 100 mmol/L ethanol exposure did not lead to any loss of cell viability, the addition of an inhibitor of aldehyde dehydrogenase was followed by a significant loss in viability. In contrast, the inhibition of catalase, which suppresses acetaldehyde synthesis, led to no reduced viability in the same exposure conditions. ROS also reduced viability, but this was observed only after both cytochrome P450 stimulation and catalase inhibition. These combined results could suggest that acetaldehyde may play a significant role in the expression of ethanol toxicity in brain.

Acetaldehyde↗
PubMed2006

Effect of rACLF, a recombinant snake venom metallopeptidase on cell viability, chemokine expression and degradation of extracellular matrix proteins.

Snake venom metallopeptidases (SVMPs) comprise a family of zinc-dependent enzymes, which display many different biological activities. ACLF is a 23kDa fibrinolytic non-hemorrhagic metallopeptidase from the venom of the snake Agkistrodon contortrix laticinctus. We have previously developed an expression system for production of recombinant ACLF (rACLF) in bacteria. To achieve a better understanding of the role of such enzyme in envenoming cases, we have studied the biological properties of rACLF, including the ability of enzyme to degrade extracellular proteins, as well its cytotoxic effect in human fibroblasts and HeLa cells. Our results showed that rACLF hydrolyzed laminin, fibronectin, type IV collagen and thrombospondin. rACLF decreased HeLa cell viability, changed cell morphology and induced detachment, while for human fibroblasts no cytotoxic effects were observed after treatment with rACLF. In addition, growth-related oncogene (GRO) and monocyte chemoattractant protein 1 (MCP-1/CCL2) were chemokines detected in the culture supernatant of human fibroblasts incubated with rACLF for 48h. These chemokines could contribute to the severe local lesion induced by Agkistrodon contortrix lacticinctus venom. These findings suggest a relevant role for ACLF in envenomation.

Agkistrodon↗
PubMed2003

Quantitative assessment of cell viability and apoptosis in cultured epidermal autografts: application to burn therapy.

Cultured epidermal autografts (CEA) have been used in the treatment of burns for almost two decades but the clinical results are still inconsistent. In a group of 37 patients with extensive burn wounds admitted to the University Hospital of Lausanne, CEA take ranged between 10 and 100% with a mean of 65%. To investigate CEA efficacy in burns, twelve CEA preparations were tested for their biological properties with particular emphasis on the balance between cell viability and apoptosis. Apoptosis was evaluated by in situ end-labeling (TUNEL), detection of DNA fragments in CEA extracts and analysis of caspase-3 activity. All CEA samples displayed a high cell viability (> 90%) and a low apoptosis rate (< 6%). However, several biological parameters including the activity of transglutaminase showed wide interindividual variability suggesting that CEA therapeutic efficacy could be partly determined by intrinsic biological factors.

Adolescent↗
PubMed2002

Genetic code ambiguity. Cell viability related to the severity of editing defects in mutant tRNA synthetases.

The rules of the genetic code are established in reactions that aminoacylate tRNAs with specific amino acids. Ambiguity in the code is prevented by editing activities whereby incorrect aminoacylations are cleared by specialized hydrolytic reactions of aminoacyl tRNA synthetases. Whereas editing reactions have long been known, their significance for cell viability is still poorly understood. Here we investigated in vitro and in vivo four different mutations in the center for editing that diminish the proofreading activity of valyl-tRNA synthetase (ValRS). The four mutant enzymes were shown to differ quantitatively in the severity of the defect in their ability to clear mischarged tRNA in vitro. Strikingly, in the presence of excess concentrations of alpha-aminobutyrate, one of the amino acids that is misactivated by ValRS, growth of bacterial strains bearing these mutant alleles is arrested. The concentration of misactivated amino acid required for growth arrest correlates inversely in a rank order with the degree of the editing defect seen in vitro. Thus, cell viability depends directly on the suppression of genetic code ambiguity by these specific editing reactions and is finely tuned to any perturbation of these reactions.

Amino Acid Sequence↗
PubMed2006

Cyclic tensile stretch load and oxidized low density lipoprotein synergistically induce lectin-like oxidized ldl receptor-1 in cultured bovine chondrocytes, resulting in decreased cell viability and proteoglycan synthesis.

Mechanical stimulation is known to be an essential factor in the regulation of cartilage metabolism. We tested the hypothesis that expression of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) can be modulated by cyclic tensile stretch load in chondrocytes. Cyclic loading of repeated stretch stress at 10 cycles per minute with 10 kPa of stress for 6 h induced expression of LOX-1 to 2.6 times control in cultured bovine articular chondrocytes, equivalent to the addition of 10 microg/mL oxidized low density lipoprotein (ox-LDL) (2.4 times control). Application of the cyclic load to the chondrocytes along with 10 microg/mL ox-LDL resulted in synergistically increased LOX-1 expression to 6.3 times control. Individual application of cyclic loading and 10 microg/mL ox-LDL significantly suppressed chondrocytes viability (84.6% +/- 3.4% and 80.9% +/- 3.2% of control at 24 h, respectively; n = 3; p < 0.05) and proteoglycan synthesis [81.0% +/- 7.1% and 85.7% +/- 5.2% of control at 24 h, respectively; p < 0.05 when compared with 94.6% +/- 4.6% for native-LDL (n = 3)]. Cyclic loading and 10 microg/mL ox-LDL synergistically affected cell viability and proteoglycan synthesis, which were significantly suppressed to 45.6% +/- 4.9% and 48.7% +/- 6.7% of control at 24 h, respectively (n = 3; p < 0.01 when compared with individual application of cyclic loading or 10 microg/mL ox-LDL). In this study, we demonstrated synergistic effects of cyclic tensile stretch load and ox-LDL on cell viability and proteoglycan synthesis in chondrocytes, which may be mediated through enhanced expression of LOX-1 and which has important implications in the progression of cartilage degeneration in osteoarthritis.

Animals↗
PubMed1984

Photodynamic inactivation of selected intracellular enzymes by hematoporphyrin derivative and their relationship to tumor cell viability in vitro.

The photosensitivity of freshly dissociated R3230AC mammary adenocarcinoma cells was examined by measurement of the activities of selected intracellular enzymes after treatment with hematoporphyrin derivative (Hpd) and exposure to light in vitro. Enzymes selected as representative of the cytosolic cell compartment showed no loss in activity, whereas malate dehydrogenase, located in the mitochondrial matrix, displayed a modest decrease (approximately 15%) in activity. In contrast, cytochrome c oxidase and succinate dehydrogenase, enzymes associated with the mitochondrial membrane, demonstrated a very rapid and more marked inhibition of activities, approximately 45% and 25%, respectively. The time-course of inhibition of these mitochondrial membrane enzymes preceded the loss of cell viability, which displayed slower kinetics as seen by a more gradual and progressive pattern of loss in viability. These data suggest that the mitochondria are an early-affected and important intracellular site for Hpd photosensitization.

Adenocarcinoma↗
PubMedDate not supplied

Co-overexpression of prlF increases cell viability and enzyme yields in recombinant Escherichia coli expressing Bacillus stearothermophilus alpha-amylase.

The effects on cloned amylase production of co-overexpression of prlF, a gene that appears to interact with the sec protein export machinery in Escherichia coli, was investigated by comparing three expression systems: (i) a high copy number plasmid with the Bacillus stearothermophilus alpha-amylase gene (amyS) cloned with its promoter downstream of the lac promoter; (ii) a pBR322-based vector with amyS under control of the indigenous Bacillus promoter; and (iii) a temperature-inducible vector with runaway replicon and lambda pL promoter-controlled gene expression. In addition, protease mutants (lon-) of E. coli C600 were used to evaluate the influence of the Lon protease on net enzyme formation and activity degradation during batch fermentations. Our results show that alpha-amylase synthesis occurred during exponential growth and ceased in the stationary phase. While strong promoters on high copy number plasmids severely impaired cell viability, resulting in culture lysis at mid-log phase, co-overexpression of prlF greatly improved cell viability, as well as the yield and specific production of alpha-amylase for the expression constructs considered. lon deficiency slightly increased amylase stability during the late stationary phase. However, the specific productivity of lon- strains was only about 40-60% that of the isogenic E. coli C600 equivalent.

Endopeptidases↗
PubMedDate not supplied

Gestational age and cell viability determine the effect of frozen storage on human fetal hematopoietic progenitor cell preparations.

We have assessed hematopoietic progenitor cell preparations (including CD34-positive stem cells) obtained from human fetal liver at 6-17 weeks of gestational age for total cell numbers, viability and the ability to tolerate (frozen) storage. Hematopoietic cell preparations obtained from the 16-to 17-week gestational age range had the highest total cell count and the greatest proportion of CD34-positive stem cells. However, these preparations were insufficiently robust to withstand the freeze/thaw protocol required. Cells obtained from the 13- to 15-week gestational age range showed optimal post-thaw viability and it is suggested that these cell preparations are the most applicable for in utero transplantation.

Abortifacient Agents, Steroidal↗
PubMed1999

The Rpb4 subunit of fission yeast Schizosaccharomyces pombe RNA polymerase II is essential for cell viability and similar in structure to the corresponding subunits of higher eukaryotes.

Both the gene and the cDNA encoding the Rpb4 subunit of RNA polymerase II were cloned from the fission yeast Schizosaccharomyces pombe. The cDNA sequence indicates that Rpb4 consists of 135 amino acid residues with a molecular weight of 15,362. As in the case of the corresponding subunits from higher eukaryotes such as humans and the plant Arabidopsis thaliana, Rpb4 is smaller than RPB4 from the budding yeast Saccharomyces cerevisiae and lacks several segments, which are present in the S. cerevisiae RPB4 subunit, including the highly charged sequence in the central portion. The RPB4 subunit of S. cerevisiae is not essential for normal cell growth but is required for cell viability under stress conditions. In contrast, S. pombe Rpb4 was found to be essential even under normal growth conditions. The fraction of RNA polymerase II containing RPB4 in exponentially growing cells of S. cerevisiae is about 20%, but S. pombe RNA polymerase II contains the stoichiometric amount of Rpb4 even at the exponential growth phase. In contrast to the RPB4 homologues from higher eukaryotes, however, S. pombe Rpb4 formed stable hybrid heterodimers with S. cerevisiae RPB7, suggesting that S. pombe Rpb4 is similar, in its structure and essential role in cell viability, to the corresponding subunits from higher eukaryotes. However, S. pombe Rpb4 is closer in certain molecular functions to S. cerevisiae RPB4 than the eukaryotic RPB4 homologues.

Amino Acid Sequence↗
PubMed2003

Killing activity of micafungin against Aspergillus fumigatus hyphae assessed by specific fluorescent staining for cell viability.

We studied the anti-Aspergillus activity of micafungin by using two fluorescent dyes to detect cell viability. Micafungin induced flattened hyphae, caused by the bursting of cells, which had lost their viability. Micafungin has killing activity against actively growing hyphae, even though it is not fungicidal against the whole burden of Aspergillus fumigatus.

Antifungal Agents↗
PubMed1984

Effects of reduced calcium on proliferation and cell viability in tumorigenic and nontumorigenic rat tracheal epithelial cell lines.

We have compared the in vitro growth and viability of tumorigenic and nontumorigenic rat tracheal epithelial cell lines over a range of calcium concentrations from 0.003 to 0.85 mM. A greater dependence on calcium for proliferation was seen in the nontumorigenic line as compared to the tumorigenic line at both the colony formation level and in mass cultures. In the latter culture condition, a marked differential effect on cell survival was also demonstrated. These differences in calcium dependence were seen in media containing fetal bovine serum or low concentrations of newborn calf serum and in a serum-free medium developed for these cells. The effect was also independent of the method used for calcium removal i.e., either by chelex treatment or the inclusion of EGTA. Therefore, loss of calcium dependence may be associated with tumorigenicity in rat tracheal epithelial cells offering a selectable marker for neoplastic cells in carcinogen-exposed preneoplastic cell populations.

Animals↗
PubMed2006

Functions of multiple exonucleases are essential for cell viability, DNA repair and homologous recombination in recD mutants of Escherichia coli.

Heterotrimeric RecBCD enzyme unwinds and resects a DNA duplex containing blunt double-stranded ends and directs loading of the strand-exchange protein RecA onto the unwound 3'-ending strand, thereby initiating the majority of recombination in wild-type Escherichia coli. When the enzyme lacks its RecD subunit, the resulting RecBC enzyme, active in recD mutants, is recombination proficient although it has only helicase and RecA loading activity and is not a nuclease. However, E. coli encodes for several other exonucleases that digest double-stranded and single-stranded DNA and thus might act in consort with the RecBC enzyme to efficiently promote recombination reactions. To test this hypothesis, I inactivated multiple exonucleases (i.e., exonuclease I, exonuclease X, exonuclease VII, RecJ, and SbcCD) in recD derivatives of the wild-type and nuclease-deficient recB1067 strain and assessed the ability of the resultant mutants to maintain cell viability and to promote DNA repair and homologous recombination. A complex pattern of overlapping and sometimes competing activities of multiple exonucleases in recD mutants was thus revealed. These exonucleases were shown to be essential for cell viability, DNA repair (of UV- and gamma-induced lesions), and homologous recombination (during Hfr conjugation and P1 transduction), which are dependent on the RecBC enzyme. A model for donor DNA processing in recD transconjugants and transductants was proposed.

Cell Survival↗
PubMed2006

Measurement of cell viability in in vitro cultures.

An overview of the methods for assessing cell viability in in vitro cultures is presented. The protocols of four of the most commonly used assays are described in detail, so the readers may be able to determine which assay is suitable for their own projects using plant cell cultures.

Cell Culture Techniques↗
PubMed1998

The nonactive site cysteine residues of yeast protein disulfide isomerase are not required for cell viability.

Protein disulfide isomerase (PDI), the product of the essential PDI1 gene of Saccharomyces cerevisiae catalyzes oxidization of thiols, reduction of disulfide bonds, and isomerization of disulfides. It can also act as a chaperone to facilitate folding of denatured proteins. The protein has 6 cysteine (Cys) residues. Four of these Cys are part of the 2 thioredoxin-like catalytic sites (-CGHC-), one of which is located near the N- and the other near the C-terminus. In addition, it has 2 non-active site Cys near the N-terminus. The function of these non-active site Cys of yeast PDI is poorly understood. Whereas in yeast PDI, these Cys residues are in the vicinity of the N-terminal-most active site, in mammalian PDI their position is closer to the C-terminal-most active site. We have examined their role and that of the active site cysteines by constructing an extensive set of mutants in which the Cys were systematically replaced by Ser. As reported earlier, the N-terminal Cys of the two active sites sequences of yeast PDI were found to be required for cell viability, but mutation of the C-terminal Cys to Ser in the two active sites was not lethal. We found that replacement of the two non-active site Cys with Ser did not affect cell viability, but in the case of the double mutant in which both Cys were replaced by Ser the processing and secretion of CPY was impaired.

Amino Acid Sequence↗
PubMed1992

Characterization of cspB, a Bacillus subtilis inducible cold shock gene affecting cell viability at low temperatures.

A new class of cold shock-induced proteins that may be involved in an adaptive process required for cell viability at low temperatures or may function as antifreeze proteins in Escherichia coli and Saccharomyces cerevisiae has been identified. We purified a small Bacillus subtilis cold shock protein (CspB) and determined its amino-terminal sequence. By using mixed degenerate oligonucleotides, the corresponding gene (cspB) was cloned on two overlapping fragments of 5 and 6 kb. The gene encodes an acidic 67-amino-acid protein (pI 4.31) with a predicted molecular mass of 7,365 Da. Nucleotide and deduced amino acid sequence comparisons revealed 61% identity to the major cold shock protein of E. coli and 43% identity to a family of eukaryotic DNA binding proteins. Northern RNA blot and primer extension studies indicated the presence of one cspB transcript that was initiated 119 bp upstream of the initiation codon and was found to be induced severalfold when exponentially growing B. subtilis cell cultures were transferred from 37 degrees C to 10 degrees C. Consistent with this cold shock induction of cspB mRNA, a six- to eightfold induction of a cspB-directed beta-galactosidase synthesis was observed upon downshift in temperature. To investigate the function of CspB, we inactivated the cold shock protein by replacing the cspB gene in the B. subtilis chromosome with a cat-interrupted copy (cspB::cat) by marker replacement recombination. The viability of cells of this mutant strain, GW1, at freezing temperatures was strongly affected. However, the effect of having no CspB in GW1 could be slightly compensated for when cells were preincubated at 10 degrees C before freezing. These results indicate that CspB belongs to a new type of stress-inducible proteins that might be able to protect B. subtilis cells from damage caused by ice crystal formation during freezing.

Amino Acid Sequence↗