Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.5.1.3 (dihydrofolate reductase)
 5,819 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Delivery of neurotrophic molecules to the central nervous system has gained considerable attention as a potential strategy for the treatment of neurological disorders. In the present study, a DHFR-based expression vector containing the human nerve growth factor gene (hNGF) was transfected into a baby hamster fibroblast cell line (BHK). Using an immunoisolatory polymeric device, encapsulated BHK-control cells and those secreting hNGF (BHK-hNGF) were transplanted unilaterally into rat lateral ventricles. Three days later, the same animals received unilateral injections of quinolinic acid (QA, 225 nmol) or the saline vehicle into the ipsilateral striatum. Approximately 2 weeks following surgery, animals were tested for apomorphine-induced rotation behavior. Animals which received BHK-hNGF cells rotated significantly less than those animals receiving BHK-control cells or QA alone. Histological analysis 29-30 days following capsule implantation demonstrated that BHK-hNGF cells attenuated the extent of host neural damage produced by QA as assessed by a sparing of ChAT- and NADPH-d-positive neurons. Moreover, a lessened GFAP reaction was apparent within the striatum of animals receiving BHK-hNGF cells. As measured by ELISA, hNGF was released by the encapsulated BHK-hNGF cells prior to implantation and following removal. Morphology of retrieved capsules revealed numerous viable and mitotically active BHK cells. These results suggest that implantation of polymer-encapsulated hNGF-releasing cells can be used to protect neurons from excitotoxin damage.
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PMID:Implantation of polymer-encapsulated human nerve growth factor-secreting fibroblasts attenuates the behavioral and neuropathological consequences of quinolinic acid injections into rodent striatum. 782 89

The effect of recombinant human nerve growth factor (hNGF) and mouse NGF on cultured rat cortical neurons was examined. The DNA fragment coding the human NGF gene was isolated and inserted downstream from the SV40 promoter in a plasmid containing the dihydrofolate reductase cDNA, and this plasmid was introduced into Chinese hamster ovary (CHO) cells to establish cells producing recombinant hNGF. The recombinant hNGF protein secreted by CHO cells was confirmed to be biologically active in an assay using PC12 cells. Brief exposure of cortical cells to glutamate followed by incubation with glutamate-free medium reduced cell viability by 60-70% when compared with the control culture. Simultaneous addition of recombinant hNGF or mouse NGF to rat cortical cultures with glutamate did not affect this reduction of cell viability. However, 24 h pretreatment of rat cortical cultures with recombinant hNGF or mouse NGF resulted in a significant reduction of glutamate-induced neuronal damage. Mouse NGF also protected cortical neurons against N-methyl-D-aspartate (NMDA)- and kainate-induced neuronal damage. These findings suggest that NGF can protect cortical neurons against glutamate-induced neurotoxicity.
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PMID:Protective effect of nerve growth factor against glutamate-induced neurotoxicity in cultured cortical neurons. 790 99

The long-term delivery of growth factors and other proteins into the CNS at putatively therapeutic yet safe levels continues to be technically constrained. In the present studies, the gene encoding human nerve growth factor (hNGF), introduced into a dihydrofolate reductase-based pNUT expression vector system, was engineered into a clonal baby hamster kidney (BHK) cell line. BHK-hNGF23 and mock-transfected cells were encapsulated in an immunoisolating polymeric device and transplanted into the lateral ventricles of healthy young adult rats for 13.5 months. As measured by ELISA, nanogram quantities of hNGF were released by encapsulated cells both prior to implantation (3.6 +/- 0.8 ng/device/24 h) and upon removal from rat lateral ventricles after 13.5 months in vivo (2.2 +/- 0.4 ng/ device/24 h). In addition, the hNGF released into the tissue culture medium was biologically active. Long-term encapsulated cell survival was confirmed by histologic analysis. The presence of genomic DNAs (hNGF transgene), as determined by PCR analyses, revealed that the transgene copy number from the recovered BHK-hNGF23 cells after 13.5 months in vivo was equivalent to preimplant levels. No deleterious effects from hNGF were detectable on body weight, mortality rate, motor/ambulatory function, or cognitive function as assessed with the Morris water maze and delayed matching to position in healthy young adult rats. In addition, there was no evidence that hNGF from these encapsulated cells produced hyperalgesia. Only tests of somatosensory thresholds revealed statistically significant effects related to the hNGF delivered in the present study, and that effect was limited to a decrease in the number of trials to asymptote. Animals receiving BHK-hNGF23 implants exhibited a marked hypertrophy of cholinergic neurons within the striatum (22% increase) and nucleus basalis (7% increase) but not the medial septum ipsilateral to the capsule. Moreover a robust sprouting of cholinergic fibers was observed within the frontal cortex and lateral septum proximal to the implant. These results indicate that encapsulated xenogeneic cells provide a safe and effective method for the long-term delivery of hNGF and potentially other neurotrophic factors within the CNS.
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PMID:Polymer-encapsulated genetically modified cells continue to secrete human nerve growth factor for over one year in rat ventricles: behavioral and anatomical consequences. 869 56

Sympathetic neurons undergo protein synthesis-dependent apoptosis when deprived of nerve growth factor (NGF). Expression of SM-20 is up-regulated in NGF-deprived sympathetic neurons, and ectopic SM-20 is sufficient to promote neuronal death in the presence of NGF. We now report that SM-20 is a mitochondrial protein that promotes cell death through a caspase-dependent mechanism. SM-20 immunofluorescence was present in the cytoplasm in a punctate pattern that colocalized with cytochrome oxidase I and with mitochondria-selective dyes. Analysis of SM-20/dihydrofolate reductase fusion proteins revealed that the first 25 amino acids of SM-20 contain a functional mitochondrial targeting sequence. An amino-terminal truncated form of SM-20 was not restricted to mitochondria but instead localized throughout the cytosol and nucleus. Nevertheless, the truncated SM-20 retained the ability to induce neuronal death, similar to the wild type protein. SM-20-induced death was accompanied by caspase-3 activation and was blocked by a general caspase inhibitor. Additionally, overexpression of SM-20, under conditions where cell death is blocked by a general caspase inhibitor, did not result in widespread release of cytochrome c from mitochondria. These results indicate that SM-20 is a novel mitochondrial protein that may be an important mediator of neurotrophin-withdrawal-mediated cell death.
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PMID:SM-20 is a novel mitochondrial protein that causes caspase-dependent cell death in nerve growth factor-dependent neurons. 1106 Mar 9