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Results for “genetic modifiers”

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At least 649 records · Page 36Linked to original sources
PubMed1996

Bioprocessing with genetically modified and other organisms: case studies in processing constraints.

Whereas the gene stability related considerations are important in bioprocessing with recombinant cultures, bioreactor design and scale-up require attention to the often reduced shear tolerance of the genetically altered biocatalysts relative to the corresponding wild strains. In addition, the peculiarities of expression of the rDNA product impact the downstream recovery methods. As a consequence, a bioprocessing scheme and the process machinery designed for a naturally occurring organism may need significant modifications for use with a genetically modified variety of the same organism. The case studies described highlight some of the processing constraints and consideration of general relevance.

Animals↗
PubMed2005

Use of genetically modified mouse models to assess pathways of benzene-induced bone marrow cytotoxicity and genotoxicity.

Benzene induces bone marrow cytotoxicity and chromosomal breaks as a primary mode of action for the induction of bone marrow toxicity. Our research group has used genetically modified mouse models to examine metabolic and genomic response pathways involved in benzene induced cytotoxicity and genotoxicity in bone marrow and in hematopoietic stem cells (HSC). We review our studies using NQO1-/- mice and mEH-/- mice to examine the roles of these enzymes, NAD(P)H:quinone oxidoreductase-1 (NQO1) and microsomal epoxide hydrolase (mEH) in mediating benzene-induced toxicity. NQO1 catalyzes the detoxication of benzene quinone metabolites and mEH catalyzes the hydrolysis of benzene oxide. Our studies using gene expression profiling of bone marrow and enriched HSC populations isolated from the bone marrow of benzene-exposed mice demonstrate differential gene expression responses of key genes induced by inhaled benzene. These studies show that benzene toxicity is regulated by a number of genetic pathways that affect the production of reactive metabolites and DNA damage response pathways in a target tissue.

Animals↗
PubMed2002

Systemic delivery of human growth hormone using genetically modified tissue-engineered microvascular networks: prolonged delivery and endothelial survival with inclusion of nonendothelial cells.

Endothelial cells have the potential to provide efficient long-term delivery of therapeutic proteins to the circulation if a sufficient number of genetically modified endothelial cells can be incorporated into the host vasculature and if these cells persist for an adequate period of time. Here we describe the ability of nonendothelial cells to modulate the survival of implanted endothelial cells and their incorporation into host vasculature. Bovine aortic endothelial cells (BAECs) suspended in Matrigel and cultured in vitro remained spherical and decreased in number over time. Subcutaneous implantation of gels containing BAECs secreting human growth hormone (hGH) in mice initially resulted in detectable plasma hGH levels, which were undetectable after 2 weeks. When mixed with fibroblasts and suspended in Matrigel, hGH-secreting BAECs formed microvascular networks in vitro. Implantation of these gels resulted in plasma hGH levels that decreased slightly over 2 weeks and then remained stable for at least 6 weeks. BAECs incorporated into blood vessels within both the implant and fibrous capsule that surrounded and invaded implants. Within implants containing BAECs and fibroblasts, viable BAECs were present for at least 6 weeks at a higher density than in implants containing BAECs alone at 3 weeks. These results indicate that implanted BAECs can incorporate into host blood vessels and that inclusion of fibroblasts in this system prolongs BAEC survival and hGH delivery.

Animals↗
PubMed2005

Effects of Na+-Ca2+ exchange expression on excitation-contraction coupling in genetically modified mice.

We have created genetically altered mice to investigate how expression of the Na(+)-Ca(2+) exchange protein alters excitation-contraction (E-C) coupling. Whereas low levels of exchanger overexpression have minimal effects on E-C coupling properties, high levels of overexpression in homozygous animals results in susceptibility to hypertrophy and heart failure, along with a significant reduction in E-C coupling gain. While global knockout of the exchanger in mice is embryonic-lethal, conditional knockout mice live to adulthood. Cardiac function is surprisingly normal in seven-week-old mice, but E-C coupling gain is apparently increased. Thus, genetic modification of exchanger expression has a major influence on E-C coupling.

Animals↗
PubMed2003

Cadmium stimulates transcription of the cytochrome p450 side chain cleavage gene in genetically modified stable porcine granulosa cells.

We investigated the effects of cadmium (Cd2+) on transcription of the cytochrome p450 side chain cleavage (p450scc) gene and on progesterone synthesis in stable granulosa cells. We used the stable porcine granulosa cell line, JC-410, genetically modified to express a luciferase genomic construct carrying 2320 base pairs (bp) of the p450scc gene promoter (p450scc-2320-LUC). A construct containing only the luciferase gene, pOLUC, was used as a promoterless control. At 1 microM, cadmium chloride (CdCl2) increased transient expression of p450scc-2320-LUC in JC-410 cells by 2.6-fold after 24-h incubation. A similar pattern of stimulation by CdCl2 was observed in cells transiently transfected with a luciferase genomic construct carrying 100 bp of the p450scc gene promoter p450scc-100-LUC, whereas no stimulation by CdCl2 was observed in cells transfected with pOLUC. At 0.6, 1, and 2 microM, CdCl2 stimulated the activity of the p450scc-2320-LUC promoter in a dose-related fashion by 1.58-, 3.19-, and 2.67-fold, respectively, after 24-h incubation. Northern blot analysis showed that CdCl2 at 0.1, 1, 2, and 3 microM increased p450scc mRNA levels by 3.13-, 1.38-, 1.61-, and 1.57-fold, respectively, after 24-h incubation. After 48-h incubation, CdCl2 at 0.6, 1, and 2 microM further increased p450scc mRNA levels by 3.43-, 2.08-, and 2.4-fold, respectively. At 1, 2, and 3 microM, CdCl2 inhibited progesterone synthesis to 0.48-, 0.38-, and 0.29-fold, respectively. After 48-h incubation, CdCl2 at 0.1 microM stimulated progesterone synthesis by 1.6-fold. We conclude that Cd2+ has a dual action in stable porcine granulosa cells: Low concentrations activate, whereas high concentrations inhibit, expression of the p450scc gene and progesterone synthesis. The stimulatory effect of Cd2+ appears to be mediated via a cis-acting element located 100 bp upstream of the p450scc gene transcription start site.

Animals↗
PubMed2005

Two polyene amides produced by genetically modified Streptomyces diastaticus var. 108.

Streptomyces diastaticus var. 108, a newly isolated strain, was recently characterized as a producer of two polyene macrolide antibiotics (rimocidin and CE-108), and the biosynthetic gene cluster was partially characterized. When the producer strain was genetically modified by transformation with some engineered SCP2*-derived vectors carrying the ermE gene, two previously uncharacterized macrolides were detected in the fermentation broth of the recombinant strain and chemically characterized as the amides of the parental polyene carboxylic acids. The biological activity and some in vitro toxicity assays showed that this chemical modification resulted in pharmaceuticals with improved biological properties compared with the parental products.

Antifungal Agents↗
PubMed1999

Adoptive transfer of genetically modified macrophages elucidated TGF-beta-mediated 'self-defence' of the glomerulus against local action of macrophages.

TGF-beta has several anti-inflammatory properties which may be relevant to prevention of or recovery from acute glomerular inflammation. Using genetically modified mesangial cells and a technique for in vivo macrophage transfer, this article provides evidence for TGF-beta-mediated 'self-defence' of the glomerulus against macrophages. Rat mesangial cells stably transfected with TGF-beta1 showed a blunted response to the macrophage-derived, proinflammatory cytokine IL-1beta. In contrast, mesangial cells expressing the dominant-interfering TGF-beta receptor showed an enhanced response to IL-1. Similarly, externally added TGF-beta1 inhibited the cytokine response of normal glomeruli, and isolated nephritic glomeruli producing active TGF-beta1 showed a depressed response to IL-1beta, compared to normal glomeruli. Consistent with these in vitro results, in vivo transfer of activated macrophages revealed that the TGF-beta-producing glomeruli are insensitive to the effector action of macrophages. These results indicate that TGF-beta1 functions as an endogenous 'defender' that counteracts local action of activated macrophages in the glomerulus.

Adoptive Transfer↗
PubMed2005

Genetic modifiers affecting severity of epilepsy caused by mutation of sodium channel Scn2a.

Mutations in the voltage-gated sodium channels SCN 1 A and SCN 2 A are responsible for several types of human epilepsy. Variable expressivity among family members is a common feature of these inherited epilepsies, suggesting that genetic modifiers may influence the clinical manifestation of epilepsy. The transgenic mouse model Scn 2 a(Q 54) has an epilepsy phenotype as a result of a mutation in Scn 2 a that slows channel inactivation. The mice display progressive epilepsy that begins with short-duration partial seizures that appear to originate in the hippocampus. The partial seizures become more frequent and of longer duration with age and often induce secondary generalized seizures. Clinical severity of the Scn 2 a(Q 54) phenotype is influenced by genetic background. Congenic C57BL/6J.Q 54 mice exhibit decreased incidence of spontaneous seizures, delayed seizure onset, and longer survival in comparison with [C57BL/6J x SJL/J]F(1).Q 54 mice. This observation indicates that strain SJL/J carries dominant modifier alleles at one or more loci that determine the severity of the epilepsy phenotype. Genome-wide interval mapping in an N(2) backcross revealed two modifier loci on Chromosomes 11 and 19 that influence the clinical severity of of this sodium channel-induced epilepsy. Modifier genes affecting clinical severity in the Scn 2 a(Q 54) mouse model may contribute to the variable expressivity seen in epilepsy patients with sodium channel mutations.

Alleles↗
PubMed1992

Grafting of genetically modified cells: effects of acetylcholine release in vivo.

In this study, microdialysis was used to investigate functional recovery of central cholinergic neurons in the forebrain of rats with cortical devascularizing lesions. Mature male rats were unilaterally lesioned by disruption of the pia arachnoid vessels and genetically modified fibroblasts secreting nerve growth factor (NGF) were placed at the site of the lesion. One month following surgery, microdialysis probes were installed in the remaining cortex and were perfused with artificial cerebrospinal fluid (csf) containing neostigmine (5 nM) and/or KCl (100 mM). The basal (non-stimulated) release of acetylcholine (ACh) in the cortex was similar in all experimental groups, whereas KCl stimulated release of ACh was significantly augmented (P < 0.05) in the ipsilateral remaining cortex in lesioned animals that have been implanted with fibroblasts secreting NGF. These results suggest that NGF secreted by genetically engineered fibroblasts modulates neuroplasticity in the adult mammalian CNS and may favour recovery of cortical function following injury.

Acetylcholine↗
PubMed1996

Using genetically modified mice to study apolipoprotein B.

The B apolipoproteins, apo-B48 and apo-B100, are key proteins in mammalian lipoprotein metabolism and are components of all classes of lipoproteins considered to be atherogenic. Our laboratory has generated an array of genetically modified mice for studying apo-B biology. Using gene targeting in mouse embryonic stem cells, we have generated apo-B-deficient mice. Heterozygotes had low plasma levels of apo-B and cholesterol; homozygotes died early in embryonic development, most likely because the absence of lipoprotein secretion by the yolk sac interfered with the delivery of lipid nutrients to the developing embryo. We have also generated human apo-B transgenic mice with an 80-kb genomic DNA fragment spanning the entire human apo-B gene; those mice had markedly increased plasma levels of low density lipoprotein cholesterol and exhibited increased susceptibility to atherosclerosis. The human apo-B transgenic mice have also yielded insights regarding the regulation of apo-B expression in different tissues. Although the 80-kb transgene contained nearly 20 kb of 5' and 3' flanking sequences and was expressed at high levels in the liver, no transgene expression was detectable in the intestine. Subsequent transgenic mouse studies have demonstrated that the expression of the apo-B gene in the intestine is controlled by DNA sequences that are very distant from the structural gene. Transgenic mice have also proved useful for studying apo-B structure/function relationships. By expressing mutant forms of human apo B in transgenic mice, we have examined the structural features of the apo-B molecule that are required for lipoprotein (a) formation. We have demonstrated that the carboxyl terminal cystine residue of apo-B100, cysteine-4326, is required for apo-B100's disulfide linkage with apo(a) to form lipoprotein (a). Finally, we have used gene targeting techniques to generate mice that synthesize exclusively apo-B48 (apo B48-only mice) and mice that synthesize exclusively apo-B100 (apo-B100 only mice): These mice have helped to clarify the unique metabolic roles of the two apo-B proteins.

Animals↗
PubMedDate not supplied

[The use of genetically modified microorganisms in agriculture to reduce or eliminate pesticide exposure].

P. graminea is the casual agent of barley leaf stripe. An early selection method of resistant types of barley leaf stripe was realized and validated in this research. This new method, based on Microrganism Genetically Modified (MOGM) GUS2 construction, obtains results comparable with those of classical method, reduces the work time, the use of chemicals (pesticides) and productive plants as greenhouses. Moreover, the use of MOGM GUS2 is restricted in laboratory ambient, therefore the risk of environmental spread is reduced. The early selection method has allowed to estimate the reaction to P. graminea agent in 12 several barley types usually farmed in Italy. The results were compared both, with the classical method data based on artificial clone Dg2 inoculum, and with natural inoculum data obtained in field. At all times we observed a ranking likeness.

Agriculture↗
PubMedDate not supplied

[Effect of natural and genetically modified rhizospheric Pseudomonas aureofaciens bacteria on accumulation of arsenic by plants].

Gene constructions rendering bacteria resistant to arsenic and capable of dissolving phosphates and/or arsenates were created by cloning ars operon and the gene of citrate synthase from a chromosome of the strain Pseudomonas aeruginosa PAO1. Genetically modified variants of the strain Pseudomonas aureofaciens BS1393 have been constructed, which are resistant to high concentrations of arsenic and dissolve poorly soluble phosphates and/or arsenates. Recombinant strains P. aureofaciens BS1393(pUCP22::arsRBC) and P. aureofaciens BS1393(pUCP22::gltA) exerted positive effects on the survival of sorgo (Sorghum saccharatum L.) and its ability to accumulate arsenic.

Arsenic↗
PubMed2007

Retinal capillary basement membrane thickness in diabetic mice genetically modified at the haptoglobin locus.

BACKGROUND: Individuals with diabetes mellitus (DM) homozygous for the haptoglobin (Hp) 1 allele are at decreased risk of retinopathy as compared to DM individuals with the Hp 2 allele. We sought to recapitulate these findings in DM mice genetically modified at the Hp locus. METHODS: An early morphological characteristic of the microangiopathy seen in diabetic retinal disease is retinal capillary basement membrane (RCBM) thickening. RCBM thickness as assessed by electron microscopy was performed on a total of 12 eyes taken from three mice in each of the four study groups (three eyes from C57Bl/6 Hp 1 and C57Bl/6 Hp 2 mice with and without streptozotocin-induced diabetes). RESULTS: The non-parametric Kruskal-Wallis ANOVA test demonstrated that there was a highly significant difference between the four groups of mice (P < 0.0001). Mann-Whitney tests for specific pair-wise comparisons demonstrated that there was no significant difference in the RCBM thickness between Hp 1 and Hp 2 mice (p = 0.70) or between DM Hp 1 and non-DM Hp 1 mice (p = 0.42). However, induction of diabetes resulted in a marked increase in RCBM thickness in Hp 2 mice compared to non-DM Hp 2 mice (p = 0.0004) and compared to DM Hp 1 mice (p = 0.0005). CONCLUSIONS: A highly significant increase in RCBM thickness was observed in DM mice with the Hp 2 genotype. These data provide important support for association studies done in humans showing an increased prevalence of diabetic retinopathy in individuals with the Hp 2 genotype.

Animals↗
PubMed2004

A novel genetic modifier of p53, mop1, results in embryonic lethality.

The heterogeneity that occurs in the tumor spectrum and latency in Li-Fraumeni syndrome (LFS) patients with inherited mutations in p53 suggest risk modifiers at loci other than the major gene. We developed a mouse model to investigate these risk modifiers. Inbred CE/J mice, which succumb to multiple types of tumors similar to those found in LFS, were crossed with the p53-null 129/Sv (129-Trp53(tm1Tyj)) mouse. In this cross, we uncovered evidence for a genetic modifier of p53, mop1, based on an unexpected mix of genotypes in the F2 progeny from Mendelian expectations. A model in which a recessive CE/J allele in combination with p53 heterozygosity or homozygosity results in lethality most closely fits the data. Using simple-sequence length polymorphism analysis of the entire genome, we identified a putative chromosomal region for this modifier of p53 on mouse chromosome 11 centromeric to p53.

Animals↗
PubMed1997

[Intracerebral transplantation of genetically modified cells for Parkinson's disease].

The clinical application of brain transplantation for treating Parkinson's disease (PD) is limited because it is difficult to obtain ideal donor cells. In this study, the tyrosine hydroxylase (TH) gene was introduced into NIH-3T3 cell line and the genetically modified cells reacted positively with TH antiserum and released L-dopa. These cells were implanted into the striatum of PD rats. 3,6,9,12 days after implantation, the average rate of the abnormal rotational behavior recovery was 52.6%, 68.4%, 63.2%, 44.7%, respectively. This result is considered to be potential for further clinical application.

3T3 Cells↗
PubMed2003

Stable lines of genetically modified dendritic cells from mouse embryonic stem cells.

BACKGROUND: The capacity to activate naïve T cells sets dendritic cells (DCs) apart from other antigen-presenting cells, making them attractive targets for immune intervention during deleterious immune responses. The inherent resistance of terminally differentiated DCs to conventional strategies for genetic modification has, however, greatly limited our understanding of the molecular mechanisms underlying their function. METHODS AND RESULTS: We report the derivation of long-term cultures of untransformed DCs, uniformly expressing a defined mutant phenotype by the directed differentiation of cloned embryonic stem cells, stably transfected with a reporter gene. Introduction of the gene encoding enhanced green fluorescent protein into pluripotent stem cells demonstrated no observable impact on the phenotype, immunogenicity, or capacity for maturation of DCs differentiated from them. CONCLUSIONS: The production of unlimited numbers of mutant DCs from genetically modified embryonic stem cells paves the way for the systematic elucidation of gene function in this cell type and the rational design of DCs for use in immunotherapy.

Animals↗
PubMed2003

Targeting exogenous genes to tumor angiogenesis by transplantation of genetically modified hematopoietic stem cells.

Angiogenic tumor vessels are promising targets for the activity and the selective delivery of cancer therapeutics. The bone marrow contributes different cell types to the tumor stroma, including hematopoietic cells and, as recently suggested, vascular endothelial cells (ECs). Thus, transplantation of genetically modified bone marrow progenitors may represent a vehicle for the transport of gene therapy to tumors. We transduced bone marrow progenitors with lentiviral vectors expressing genes from transcription-regulatory elements of Tie2/Tek gene. When tumors were grown in the transplanted mice, the new vector marked a distinct hematopoietic population that 'homed' to the tumor and closely interacted with vascular ECs at the tumor periphery. These Tie2-expressing mononuclear (TEM) cells had a distinguishable phenotype and were present selectively at angiogenic sites. Unexpectedly, we did not find bone marrow-derived ECs in tumor vessels when we transplanted bone marrow progenitors constitutively expressing a marker gene from the Tie2 or ubiquitously active promoters. By delivering a 'suicide' gene, we selectively eliminated the TEM cells and achieved substantial inhibition of angiogenesis and slower tumor growth without systemic toxicity. Thus, TEM cells may account for the proangiogenic activity of bone marrow-derived cells in tumors, may represent a new target for drug development and may provide the means for selective gene delivery and targeted inhibition of tumor angiogenesis.

Animals↗
PubMed2003

Genetic modifiers of cardiac arrhythmias.

Rhythmic contraction of a four-chambered mammalian heart is a highly coordinated process, requiring a functional conduction system. Both acquired and inherited forms of arrhythmia can be life threatening, and are major causes of mortality and morbidity in developed nations. Knowledge derived from human genetics and from studies of mouse genetic models has led to the discovery of multiple molecular defects responsible for arrhythmogenesis, including mutations in ion channels, cytoplasmic ion-channel-interacting proteins, gap-junction proteins, transcription factors and, most recently, a kinase subunit. However, phenotypic expression of a given mutation does not always appear to be uniform in human patients, implying a contribution from environmental factors and/or the presence of other genetic modifiers. Accumulating evidence suggests that 'multiple hits' affecting the interaction and integrity of multiple pathways might be responsible for many forms of arrhythmia.

AMP-Activated Protein Kinase Kinases↗