Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have previously described the isolation of a cDNA clone corresponding to an mRNA rapidly induced to high levels in PC12 cells by treatment with NGF. We report here the complete amino acid sequence of the protein (named VGF8a) as deduced by nucleotide sequencing of overlapping cDNA clones. VGF8a is particularly rich in proline residues and has a conspicuous number of short stretches of basic amino acid residues which may represent potential targets for proteolytic cleavage. Antibodies directed against recombinant VGF8a-beta-galactosidase fusion proteins were used for immunofluorescent staining of the protein in PC12 cells as well as for its localization, by Western blot analysis, in subfractions of cell homogenates. We demonstrate that in PC12 cells, VGF8a protein is stored in secretory vesicles and is released in response to a variety of stimuli that are known to induce the regulated secretion of neurotransmitters.
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PMID:A protein induced by NGF in PC12 cells is stored in secretory vesicles and released through the regulated pathway. 267 16

Although adrenal medullary chromaffin cells have been used extensively for intracerebral grafting, their survival has generally been poor. Improved survival of the implanted cells has been achieved by exposing the chromaffin cells to NGF in vivo. Culture studies have shown, however, that chromaffin cells are converted into sympathetic neurons when NGF is included in the medium. The degree to which such a transdifferentiation may occur in vivo has not been determined. We assessed the effects of cografting chromaffin cells with primary fibroblasts genetically engineered to express NGF. Chromaffin cells from 10 d old rats were implanted with NGF-producing or beta-galactosidase-producing primary fibroblasts (control fibroblasts) into the striatum of 6-hydroxydopamine treated adult rats of the same strain. Eight weeks postgrafting, chromaffin cells cografted with NGF-producing fibroblasts displayed many of the features of mature sympathetic neurons such as large somata, long processes, transmitter vesicles similar to those found in neurons, and positive immunolabeling for the neuronal markers neurofilament, MAP2 and SCG10. Chromaffin-derived neuron number was also significantly enhanced in the presence of NGF-producing fibroblasts. While control fibroblasts were also found to increase chromaffin cell number above that of chromaffin cells grafted alone, the control fibroblasts did not induce neuronal transdifferentiation. These results demonstrate that chromaffin cells cografted with NGF-producing fibroblasts undergo transdifferentiation in vivo and express many characteristics of mature sympathetic neurons. The consequences of this transdifferentiation on the long term survival and function of the transplanted cells in vivo remain to be clarified.
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PMID:Enhanced survival and neuronal differentiation of adrenal chromaffin cells cografted into the striatum with NGF-producing fibroblasts. 786 93

Adult rat chromaffin cells in vitro show a large proliferative response to NGF, followed by neuronal differentiation. Infection of replicating chromaffin cells with a retrovirus carrying the Escherichia coli beta-galactosidase (beta-gal) gene demonstrates beta-gal expression in cells that continue to multiply, that differentiate into neurons, and that become static. The effects of NGF on proliferation and differentiation are abolished by the protein kinase inhibitors K252a and staurosporine, and by cholera toxin, an activator of adenylate cyclase. They are diminished, but not abolished, by high concentrations of dexamethasone. Both cholera toxin alone and phorbol myristate acetate (PMA), an activator of protein kinase C, elicit small and inconsistent mitogenic responses. The responses to PMA cannot be shown to be additive with the effects of NGF. NGF is a known mitogen and neuritogen for chromaffin cells from neonatal rats, but has not previously been believed to affect similarly chromaffin cells from adults. The present findings suggest that portions of NGF signaling pathways might continue to be involved in regulating proliferation of adult rat chromaffin cells in vivo, and might be constitutively activated in PC12 cells and other adrenal medullary tumors. They further suggest that rat chromaffin cells might be propagated in vitro to obtain large numbers of sympathetic neurons expressing normal or exogenous genes.
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PMID:Nerve growth factor is a potent inducer of proliferation and neuronal differentiation for adult rat chromaffin cells in vitro. 846 33

In earlier studies we found that treatment with interferon-gamma (IFN-gamma) produced an 8- to 11-fold increase in choline acetyltransferase (ChAT) in cultured cells taken from Embryonic Day 16 (E16) septal nuclei with adjacent basal forebrain (SN/BF). Since younger cultures responded even more profoundly to IFN treatment, we have tested the possibility that the action of IFN (or its intermediate; see below) is to prompt the cholinergic differentiation of neuronal precursors. SN/BF cultures of various ages were labeled with a retrovirus engineered to express beta-galactosidase (Lac-Z), and ChAT-positive descendants of the retrovirally labeled precursors were counted. IFN-gamma treatment of cultures caused as much as an 8.8-fold increase in the proportion of ChAT-positive cells present in Lac-Z-positive clones, suggesting that IFN promoted cholinergic differentiation in precursor populations. By contrast, bFGF increased clone size but did not change the proportion of ChAT-positive cells. NGF affected neither. Only ameboid microglia present in the cultures responded to IFN with characteristic nuclear translocation of the signal transducing molecule p91, suggesting that a microglial-derived molecule may mediate the action of IFN. Consistent with this hypothesis, conditioned media from cultures of enriched, activated microglia also increased ChAT activity in a dose-dependent fashion. Conditioned media from an unstimulated macrophage/monocyte cell line (RAW 264.7) also proved extremely efficacious in raising ChAT activity. In addition, conditioned media from both activated microglia and RAW 264.7 cells increased the proportion of ChAT-positive cells in retrovirally labeled clones to the same extent as IFN itself, suggesting the possibility that they contain the molecule(s) that mediates the action of IFN. Preliminary characterization of this molecule suggests that it is a very stable and large protein. Together these data suggest that a molecule promoting cholinergic differentiation is produced by activated microglia and other macrophage-like cells. The identity of this molecule and its precise role in normal development await its further purification.
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PMID:Conditioned medium from activated microglia promotes cholinergic differentiation in the basal forebrain in vitro. 866 Aug 79

Focal molecular genetic alteration of the intact mammalian brain will be required to elucidate gene product function in cells comprising synaptic networks. To this end, a somatic mosaic approach has been developed for the mouse whereby a dormant germline transgene is activated by the somatic delivery and expression of cre recombinase. Transgenic mice harboring a recombinational substrate, the germline-transmitted nerve growth factor excision activation transgene (NGF-XAT) were generated. Somatic delivery of virus vectors expressing cre recombinase into the brain of NGF-XAT mice resulted in regional recombination and activation of the transgene as demonstrated at the DNA level by PCR and at the protein level by both immunocytochemistry and ELISA. This approach has been used to evaluate a behavioral correlate of unilateral NGF mosaicism within the dorsal hippocampal formation. NGF-XAT mice activated by expression of cre recombinase manifest increased locomotor activity compared with NGF-XAT mice transduced by a control virus expressing Escherichia coli beta-galactosidase. These data indicate that focally increased expression of NGF in one part of a synaptic network can elicit changes in behavior presumably by altering the overall function of NGF-responsive neural circuitry. This approach should have broad application to other gene products and promises to provide the unprecedented ability to create and study discrete genetic modifications in the context of an intact adult mammal.
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PMID:Nerve growth factor somatic mosaicism produced by herpes virus-directed expression of cre recombinase. 903 7

Muscle injuries are a challenging problem in traumatology, and the most frequent occurrence in sports medicine. Muscle contusions are among the most common muscle injuries. Although this injury is capable of healing, an incomplete functional recovery often occurs, depending on the severity of the blunt trauma. We have developed an animal model of muscle contusion in mice (high energy blunt trauma) and characterized the muscle's ability to heal following this injury using histology and immunohistochemistry to determine the level of muscle regeneration and the development of scar tissue. We have observed a massive muscle regeneration occurring in the first 2 wk postinjury that is subsequently followed by the development of muscle fibrosis. Based on these observations, we propose that the enhancement of muscle growth and regeneration, as well as the prevention of fibrotic development, could be used as approach(es) to improve the healing of muscle injuries. In fact, we have identified three growth factors (bFGF, IGF-1, and NGF) capable of enhancing myoblast proliferation and differentiation in vitro and improving the healing of the injured muscle in vivo. Furthermore, the ability of adenovirus to mediate direct and ex vivo gene transfer of beta-galactosidase into the injured site opens possibilities of delivering an efficient and persistent expression of these growth factors in the injured muscle. These studies should help in the development of strategies to promote efficient muscle healing with complete functional recovery following muscle contusion.
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PMID:Development of approaches to improve the healing following muscle contusion. 985 87

Nerve growth factor therapy has been proposed as a potential means of preventing degeneration of basal forebrain cholinergic neurons in Alzheimer's disease and thereby improving cognition. However, NGF has been reported to upregulate expression of the beta-amyloid precursor protein, which in turn could accelerate deposition of "mature" beta-amyloid in the brain. To address this possibility, the brains of 16 adult and aged rhesus monkeys were examined for beta-amyloid plaque deposition in the presence or absence of NGF treatment. Six aged monkeys received intraparenchymal grafts into the cholinergic basal forebrain of autologous cells genetically modified to secrete NGF, six aged monkeys received intraparenchymal grafts of autologous control cells expressing the reporter gene beta-galactosidase, and four adult nonoperated monkeys served as additional controls. All brains were examined for expression of mature beta-amyloid using an antibody recognizing amino acids 1-40 of the beta-amyloid peptide. Amyloid plaques were systematically quantified in representative sections of the temporal, frontal, cingulate, insular, and parietal cortices and in the amygdala and hippocampus. Results disclosed that aging resulted in an increase in amyloid plaque formation: no plaques at all were detected in nonaged monkeys, whereas a mean of 20 +/- 13 plaques per section were present in control-aged monkeys. Aged subjects with intraparenchymal NGF-secreting grafts for 3 months contained a mean of 29 +/- 14 plaques per section, an amount that did not differ significantly from control-aged monkeys (P = 0.66). Thus, 3 months of intraparenchymal NGF delivery did not significantly increase beta-amyloid deposition.
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PMID:Targeted intraparenchymal delivery of human NGF by gene transfer to the primate basal forebrain for 3 months does not accelerate beta-amyloid plaque deposition. 987 92