Determination of Intrachromosomal Recombination Rates in Cultured Mammalian Cells

Arsenic and lead can induce genetic injuries and epigenetic signaling pathways in cultured mammalian cells. To test whether signaling pathways affect the extent of genetic injuries, we explored the impacts of extracellular signal-regulated kinase 1 and 2 (ERK) on nucleotide excision repair (NER), cytotoxicity, and genotoxicity following sodium arsenite [As(III)] and lead acetate [Pb(II)]. Sustained ERK activation was observed in human cells exposed to As(III) and Pb(II). As(III) inhibited the cellular NER synthesis capability; conversely, Pb(II) stimulated it. ERK activation contributed to the As(III)-induced NER inhibition and micronucleus formation. In contrast, this signal was required for inducing cellular NER activity and preventing mutagenesis following Pb(II). ERK activation by Pb(II) was dependent on protein kinase C (PKCα) that also exhibited anti-mutagenicity. Enforced expression of ERK signaling markedly elevated the cellular NER activity, which was suppressed by As(III). Nonetheless, ERK activation could counteract the cytotoxicity caused by these two metals. Together, the results indicate that pro-survival ERK signaling exhibits dual and opposing impacts on NER process following As(III) and Pb(II) exposures. The findings also suggest that ERK is an important epigenetic signaling in the determination of metal genotoxicity.

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Chromatid Interactions during Intrachromosomal Recombination in Mammalian Cells

Mechanisms of Eukaryotic DNA Recombination ◽

10.1016/b978-0-12-293445-2.50004-1 ◽

1992 ◽

pp. 3-13 ◽

Cited By ~ 1

Author(s):

RONI J. BOLLAG ◽

R. MICHAEL LISKAY

Keyword(s):

Mammalian Cells ◽

Intrachromosomal Recombination

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Multi-scale modeling of cell survival and death mediated by the p53 network: a systems pharmacology framework

Molecular BioSystems ◽

10.1039/c5mb00304k ◽

2015 ◽

Vol 11 (11) ◽

pp. 3011-3021 ◽

Cited By ~ 3

Author(s):

Yuan Wang ◽

Zihu Guo ◽

Xuetong Chen ◽

Wenjuan Zhang ◽

Aiping Lu ◽

...

Keyword(s):

Cell Survival ◽

Cell Fate ◽

Mammalian Cells ◽

Regulatory Process ◽

Systems Pharmacology ◽

Multi Scale ◽

Scale Modeling ◽

Multi Scale Modeling ◽

P53 Network

The determination of cell fate is a key regulatory process for the development of complex organisms that are controlled by distinct genes in mammalian cells.

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Intermolecular homologous recombination between transfected sequences in mammalian cells is primarily nonconservative.

Molecular and Cellular Biology ◽

10.1128/mcb.7.10.3561 ◽

1987 ◽

Vol 7 (10) ◽

pp. 3561-3565 ◽

Cited By ~ 25

Author(s):

M M Seidman

Keyword(s):

Resistance Gene ◽

Mammalian Cells ◽

Restriction Analysis ◽

Shuttle Vector ◽

Recombination Reaction ◽

Neomycin Resistance Gene ◽

African Green Monkey Cells ◽

The Individual ◽

Neomycin Resistance

Intermolecular recombination in mammalian cells was studied by coinfecting African green monkey cells in culture with two shuttle vector plasmids, each carrying an incomplete but overlapping portion of the gene for neomycin resistance. The region of homology between the two plasmids was about 0.6 kilobases. Recombination between the homology regions could reconstruct the neomycin resistance gene, which was monitored by analysis of progeny plasmids in bacteria. The individual plasmids carried additional markers which, in combination with restriction analysis, allowed the determination of the frequency of formation of the heterodimeric plasmid which would be formed in a conservative recombination reaction between the homologous sequences. Reconstruction of the neomycin resistance gene was readily observed, but only 1 to 2% of the neomycin resistance plasmids had the structure of the conservative heterodimer. Treatment of the plasmids which enhanced the frequency of the neomycin resistance gene reconstruction reaction did not significantly increase the relative frequency of conservative product plasmids. The results support nonconservative models for recombination of these sequences.

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[12] Determination of guanine nucleotides bound to Ras in mammalian cells

Methods in Enzymology - Small GTPases and Their Regulators, Part A: RAS Family ◽

10.1016/s0076-6879(95)55014-3 ◽

1995 ◽

pp. 118-125 ◽

Cited By ~ 12

Author(s):

Jackson B. Gibbs

Keyword(s):

Mammalian Cells ◽

Guanine Nucleotides

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Determination of Peroxisomal pH in Living Mammalian Cells Using pHRed

Methods in Molecular Biology - Peroxisomes ◽

10.1007/978-1-4939-6937-1_16 ◽

2017 ◽

pp. 181-189 ◽

Cited By ~ 1

Author(s):

Luis F. Godinho ◽

Michael Schrader

Keyword(s):

Mammalian Cells

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Conservative intrachromosomal recombination between inverted repeats in mouse cells: association between reciprocal exchange and gene conversion.

Genetics ◽

10.1093/genetics/119.1.161 ◽

1988 ◽

Vol 119 (1) ◽

pp. 161-169

Author(s):

R J Bollag ◽

R M Liskay

Keyword(s):

Gene Conversion ◽

Mammalian Cells ◽

Inverted Repeats ◽

Intrachromosomal Recombination ◽

Reciprocal Exchange ◽

Sequence Inversion ◽

Mouse Cells ◽

Simplex Virus ◽

The Relationship ◽

Recombination Gene

Abstract Recombination in mammalian cells is thought to involve both reciprocal and nonreciprocal modes of exchange, although rigorous proof is lacking due to the inability to recover all products of an exchange. To investigate further the relationship between these modes of exchange, we have analyzed intrachromosomal recombination between duplicated herpes simplex virus thymidine kinase (HSV tk) mutant alleles arranged as inverted repeats in cultured mouse L cells. In crosses between inverted repeats, a single intrachromatid reciprocal exchange leads to inversion of the sequence between the crossover sites and recovery of both genes involved in the event. The majority of recombinant products do not display such inversion and are thus consistent with a nonreciprocal mode of recombination (gene conversion). The remaining products display the sequence inversion predicted for intrachromatid reciprocal exchange. In light of the fact that intrachromatid exchanges occur, the rarity of intrachromatid double reciprocal exchanges strengthens the interpretation that the majority of events in this and previous investigations involve gene conversion. Furthermore, in accord with prediction, one-third of the reciprocal recombinants (inversions) display associated gene conversion. This association suggests that reciprocal and nonreciprocal modes of exchange are mechanistically related in mammalian cells. Finally, the occurrence of inversion recombinants suggests that intrachromosomal recombination can be a conservative (nondestructive) process.

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CHITOSAN NANOPARTICLES AS DRUG DELIVERY SYSTEM FOR CEPHALEXIN AND ITS ANTIMICROBIAL ACTIVITY AGAINST MULTIIDRUG RESISTENT BACTERIA

International Journal of Pharmacy and Pharmaceutical Sciences ◽

10.22159/ijpps.2019v11i7.33375 ◽

2019 ◽

pp. 14-27 ◽

Cited By ~ 2

Author(s):

Mona Elassal ◽

Nagwan El-Manofy

Keyword(s):

Antimicrobial Activity ◽

Electron Microscope ◽

Mammalian Cells ◽

Synergistic Effects ◽

Drug Loading ◽

Chitosan Nanoparticles ◽

Clinical Isolates ◽

Antibiofilm Activity ◽

Diffusion Kinetic

Objective: The evolution of antimicrobial resistance is a universal obstacle that necessities the innovation of more effective and safe antimicrobial alternatives with synergistic properties. The purpose of this study was to investigate the possible improvement of cephalexin antimicrobial treatments by loading into chitosan-based nanoparticles, then evaluate their antibacterial and antibiofilm activities as well as determination of its cytotoxicity. Methods: Chitosan nanoparticles (CSNPs) were prepared by ionic gelation method. Parameters were studied to optimize the particle size of CSNPs including pH, stirring rate, homogenization and ultra-sonication time. Size was measured by transmission electron microscope (TEM) and Zeta sizer, morphology seen by scanning electron microscope (SEM). Entrapment efficiency, drug loading and drug content were calculated. Stability of both plain and loaded chitosan Nano-carriers, Drug release and Kinetics also compatibilities were studied. Antimicrobial activity of CSNPs and cephalexin loaded CSNPs were evaluated against 4 Gram-positive and 4 Gram-negative standard and clinical isolates by microdilution method, also assessment of antibiofilm activity of both formulas was investigated against two biofilm producers clinical isolates by tube assay in addition to determination of their cytotoxicity by MTT(3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Results: Chitosan nanoparticles and its loaded antibiotics proved compatible combination with small Zeta size, suitable Zeta potential, maximum EE% and drug-loading capacity, sustained controlled release properties followed diffusion kinetic model and six month stability studies. Cephalexin loaded CSNPs showed better antimicrobial activity than plain CSNPs. Synergistic effects were found against S. aureus (ATCC 25923), B. subtilis (ATCC 9372), S. epidermidis, E. faecalis, P. aeruginosa (ATCC 29853) in addition to two carbapenem resistant isolates k. pneumoniae and E. coli. Also cephalexin loaded CSNPs exhibited antibiofilm activity against E. faecalis clinical isolate. Even though, cephalexin loaded CSNPs exhibited significant antibacterial activity, it showed less toxicity against mammalian cells, it had IC50 equal to 231.893 and did not exhibit any cytotoxicity against the WI-38 fibroblast cells at concentration 23.4 µg/ml. Conclusion: Cephalexin loaded CSNPs possessed good stability and sustained release effect in addition to its antimicrobial, antibiofilm activities and reduced cytotoxicity.

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