Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.24.3 (collagenase)
18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In the present study, we sought to identify the T cell-replacing factor which selectively induces IgG2b antibody formation in lipopolysaccharide-activated mouse spleen cells in vitro and in vivo, and which is present in the synovial fluid (SF) of rheumatoid arthritis (RA) patients. The protein A plaque assay was used to measure IgM, IgG1, IgG2b, and IgG3 plaque-forming cells. An enzyme-linked immunosorbent assay was used to measure interleukin-6 (IL-6) levels in RA SF. We found that IgG2b induction by RA SF is not caused by IL-6, IL-1, or any other inflammatory cytokines or mediators, such as transforming growth factor beta, platelet-derived growth factor, nerve growth factor, fibroblast growth factor, epidermal growth factor, elastase, collagenase, and phospholipase A2. IgG2b-inducing factor in RA SF has unique biological properties compared with those of the interleukins and inflammatory mediators known to be present in RA SF.
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PMID:Relationship between IgG2b-inducing activity in rheumatoid arthritis synovial fluid and other well-known cytokines and inflammatory mediators. 195 23

In order to maintain a chronic supply of growth factor for medulla cells in vitro, chromaffin cells from rat, African green monkeys and man were co-cultured with C6 glioma cells, which secrete growth factors that sustain sympathetic neurons in vitro. The response of chromaffin cells to coculture was compared to treatment of medullary cells with nerve growth factor (NGF) alone. Dispersed chromaffin cell preparations were obtained by a trypsin-collagenase procedure, and subjected to differential plating on collagen-coated surfaces. With both human and monkey tissue, non-chromaffin cells did attach to the culture plates and an enriched chromaffin cell population could be replated. Rat adrenal medulla cells survived very poorly in vitro and were not enriched in this procedure. Cultured human and monkey chromaffin cells survived as epithelial cells (50%) and showed neuritic outgrowth on 55 to 66% of the cells after eight days when treated with nerve growth factor (NGF). These cells showed strong catecholamine histofluorescence, tyrosine hydroxylase (TH) and dopamine beta hydroxylase (DBH) immunoreactivity. In contrast, only ten percent of adult rat chromaffin cells survived in culture, although NGF treatment rescued an additional 20% of the cells and induced neuritic outgrowth after one week in vitro. C6 glioma cells were treated with mitomycin C bromodeoxyuridine to inhibit mitosis and were plated with the various medulla cells in a one to one ratio. Both human and monkey chromaffin cells expressed extensive and enhanced neuritic arborization within eight days of co-culture, (64-82% respectively) and exhibited intimate contact with the glioma cells as seen at the ultrastructural level. Importantly, survival of adult rat adrenal medulla cells was enhanced to 50% or more with 40% of the cells extending neurites when co-cultured with glioma cells for seven days. Chromaffin cells from all three species reacted for TH, DBH and PNMT in co-culture and were histo-fluorescent. The majority of these cells were also immunoreactive for serotonin and enkephalin, while only 37% of chromaffin cells indicated the presence of NPY. These data indicate that adrenal medulla can be maintained in vitro as the neuronal phenotype when co-cultured with growth factor producing cells and that this strategy may be useful for in vivo transplantation studies.
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PMID:Rodent and primate adrenal medullary cells in vitro: phenotypic plasticity in response to coculture with C6 glioma cells or NGF. 256 44

Chromaffin cells from the monkey adrenal medulla were maintained in vitro in the presence of nerve growth factor (NGF) and the neuronal properties of these cells were assessed. Single-cell preparations were obtained by collagenase-trypsin treatment of the minced adrenal medulla tissue. Cells assumed a glandular to epithelioid morphology after twenty-four hours of culture. Twelve percent of these cells were shown to extend neurites spontaneously after five days. NGF-stimulated neuritic outgrowth from most cells after five days of culture and these neurites remained for at least three weeks. Cells exhibited intense histofluorescence for catecholamines even after three weeks in vitro in the presence of NGF and positive staining for tyrosine hydroxylase and dopamine beta hydroxylase could be detected by immunocytochemistry. Moreover, the chromaffin cells were shown to bind tetanus toxin, which is a specific marker for neurons. Tetanus toxin labelling was not dependent upon the presence of neurites on these cells. Transmission electron microscopy indicated that cultured cells contained numerous dense-core vesicles similar to non-cultured medulla cells. Many of the neurites possessed the morphological features of axons; long varicose processes resembling noradrenergic fibers were identified by catecholamine histofluorescence and tyrosine hydroxylase immunocytochemistry. Microtubular arrays, in an axonal-like organization pattern, were seen ultrastructurally along with the presence of many dense-core vesicles. These data support the potential of adult primate chromaffin cells as a source of sympathetic neuronal tissue for neural transplantation.
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PMID:Neuronal properties of monkey adrenal medulla in vitro. 287 Aug 11

Adult rat muscle fibres were dissociated by using collagenase and maintained in culture. One to nine days later, neurons obtained from stages 22-30 Xenopus laevis embryos, or neonatal spinal cord, or pheochromocytoma (PC12) cells treated with nerve growth factor were added. Subsequently, the co-cultures were maintained for up to eight days. Functional synapses were formed with variable efficiency: 12% in rat-Xenopus nerve-muscle co-cultures, 23% in rat-rat and 33% in PC12 co-cultures. Miniature endplate potentials (MEPPs) and currents (MEPCs) were recorded, at frequencies ranging from 0.01 to 0.9 Hz. Their mean amplitude was smaller than in normal mammalian muscles. The rise time and time-constant of decay of MEPCs was about seven to ten times longer than that found in the original muscle, resembling immature synapses. (+)-Tubocurarine abolished the MEPPs in the rat-PC12 neuromuscular junctions. It is concluded that dissociated adult rat muscle fibres retain their ability of being reinnervated, and can form functional synapses with foreign neurons and transformed chromaffin cells.
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PMID:In vitro reinnervation of adult rat muscle fibres by foreign neurons and transformed chromaffin PC12 cells. 290 Nov 8

We studied the distribution of the molecular forms of acetylcholinesterase (AChE) in a stable variant (F3) of the rat pheochromocytoma cell line, PC12, that lacks a heparan sulfate proteoglycan on the cell surface. After treatment with nerve growth factor F3 cells synthesize less 4S enzyme, and more 10S and 16S enzyme than normal PC12 cells. This distribution is similar to that seen in normal cells after incubation with beta-D-xylosides, molecules that interfere with proteoglycan assembly. Using collagenase treatment and membrane-permeable and -impermeable inhibitors of AChE, we determined the cellular location of the AChE forms. Although in normal cells greater than 90% of the 16S AChE is on the cell surface, approximately 60% is present in an internal pool in the variant. Following irreversible inhibition of all forms of AChE in the variant, the newly synthesized 16S AChE appears in the internal pool after a 1-h lag, but is not detected on the cell surface until after 2.5 h. Our results thus show that 16S AChE is assembled internally within neuronal cells and that alterations in the synthesis and distribution of proteoglycans affect the total amount and cellular localization of the 16S AChE form.
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PMID:Atypical distribution of asymmetric acetylcholinesterase in mutant PC12 pheochromocytoma cells lacking a cell surface heparan sulfate proteoglycan. 315 21

Rat sympathetic neurons, plated onto extracellular matrix produced by cultured bovine corneal endothelial cells, rapidly extended neurites in the absence of nerve growth factor (NGF). The response was unaffected by antiserum to NGF. Rapid outgrowth also occurred when sympathetic neurons were plated onto polylysine-coated surfaces that had been exposed to serum-free medium conditioned by corneal endothelial cells (CMSF). A response was seen even when the neurons were cultured without serum. When plated onto a polylysine-coated dish treated with CMSF over half its surface, only the neurons on the treated half extended neurites. The active factor in CMSF was destroyed by trypsin, acid (pH 1.6), base (pH 12.7), or heating to 80 degrees C; it was stable to heating to 60 degrees C, collagenase, deoxyribonuclease, and neuraminidase. The factor elutes just after the void volume of a Sepharose 6B column. In associative cesium chloride gradients, it sediments as a peak centered at a density of 1.36-1.37, corresponding to a peak of material that can be biosynthetically labeled with [35S]sulfate or [3H]leucine. Material from this fraction was inactivated by heparinase, but not chondroitinase ABC, implying that a heparin sulfate proteoglycan is essential for the factor's activity. Inactivation by contaminants in the heparinase preparation was ruled out. Further purification indicated that the active factor may exist as an aggregate containing a heparin sulfate proteoglycan and other molecules. CMSF also promoted neurite outgrowth by other types of neurons. Furthermore, a variety of cell types were shown to produce factors similar to that in CMSF.
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PMID:Characterization of a factor that promotes neurite outgrowth: evidence linking activity to a heparan sulfate proteoglycan. 621 11

Cultured corneal epithelial cells release a factor(s) that stimulates trigeminal neurons to form neurites in vitro. To characterize this trophic effect, conditioned media (serum free, supplemented) from cultures for corneal epithelium, stromal fibroblasts, and endothelium were studied further. Only epithelial conditioned medium (PCM) prolonged neuronal survival and induced neurite outgrowth. This trophic influence peaked after 2 to 3 days and gradually declined thereafter during a week when the medium was not renewed. Using a bioassay to score the percentage of initially viable neurons that extended neurites, it was found that the trophic effect of PCM was proportional to the conditioned medium concentration and to the cell density of the epithelial culture used for the conditioning. Maximum activity in PCM was correlated with confluency of the epithelial culture. Experiments using antiserum to nerve growth factor (NGF) and purified antibody to cold-insoluble globulin (CIG) indicated that the tropic effect of PCM was not derived from NGF or CIG. The trophic activity of PCM was abolished totally by heat or trypsin treatment but was not affected by collagenase. Although a fraction of the trophic activity was associated with the substratum after adsorption of PCM, this and other evidence did not suggest that the primary action of PCM was to enhance neuronal adhesion.
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PMID:Action of a trophic factor(s) from rabbit corneal epithelial culture on dissociated trigeminal neurons. 728 78

In rat pheochromocytoma (PC12) cells treated with nerve growth factor (NGF), there are several molecular forms of the enzyme acetylcholinesterase (AChE) which sediment on sucrose density gradients at 4 to 6, 10, and 16 S, respectively. We have investigated the cellular localization of these forms in PC12 cells. In order to determine which forms are soluble and which are membrane bound, we extracted PC12 cells in buffers of various ionic strengths and detergent compositions. To distinguish internal from external forms of the enzyme, we examined the effect of di-isopropyl fluorophosphate and BW284c51 dibromide, membrane-permeable and -impermeable inhibitors of AChE, respectively, AChE forms in intact cells. We also determined the susceptibility of the forms in intact cells to collagenase treatment. Based on these studies, we conclude that the globular G1 and G2 (4 to 6 S) forms are internal and consist of both soluble and membrane-associated species. Thirty percent of the G4 (10 S) form is bound to cytoplasmic membrane structures, while the remainder occurs as an integral component of the plasma membrane. The asymmetric A12 (16 S) form is also a surface protein but is extracted by high salt without detergent and is released from intact cells by collagenase. This form thus contains a collagenous domain and is located outside of the plasma membrane, where it may be associated with an extracellular matrix.
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PMID:Cellular localization of the molecular forms of acetylcholinesterase in rat pheochromocytoma PC12 cells treated with nerve growth factor. 731 Apr 86

Newborn rat brain astrocytes cultured in vitro in a chemically defined medium are shown to secrete enhanced levels of nerve growth factor (NGF) when they are exposed to various types of proteases. Proteolytic enzymes such as alpha-thrombin or collagenase induce a continuous, dose-dependent enhancement of the levels of cell-secreted NGF. Incubation of astrocytes for a 24-h period with 300 ng/ml of alpha-thrombin (approximately 9 nM, or 1 U/ml) results in an increase of the levels of cell-secreted NGF by a factor of three- to fourfold, and at doses 10 times higher, stimulation by a factor of up to four- to fivefold was observed. This phenomenon reflects an enhancement of the cellular pool of NGF mRNA, already noticeable after 3 h of treatment, which is preceded by a temporary activation of protooncogenes encoding transcription factors of the AP-1 family, such as c-fos, c-jun or junB. Trypsin, plasmin, alpha-chymotrypsin, or elastase also enhanced, to different extents, the levels of cell-secreted NGF. However, unlike alpha-thrombin or collagenase, these enzymes cause, above a critical concentration, an extensive cell detachment from the solid support, and this is accompanied by a decrease of their activity on the production of NGF, so that their dose-response curves are bell shaped. Stimulation was maximal at those concentrations that cause a limited loosening of the cell-substratum interactions, as evidenced by a retraction of some cell processes after 24 h of treatment. Studies of the effect of alpha-thrombin indicate that the proteolytic activity itself is required to enhance the production of NGF by astrocytes. Inactivation of alpha-thrombin with D-phenyl-alanyl-L-propyl-L-arginine chloromethyl ketone, phenylmethylsulfonyl fluoride, antithrombin III, or hirudin results in a marked decrease of the stimulatory effect. Furthermore, the prolonged presence of alpha-thrombin is required to elicit a maximal effect on the levels of extracellular NGF, which was observed after 48 h of treatment. It is known that some effects of alpha-thrombin require binding to the cell surface. We found that gamma-thrombin, which still has some proteolytic activity but has lost its ability to bind to the cell surface, is almost as potent as alpha-thrombin in promoting the release of NGF. It is concluded that the effect of thrombin on NGF synthesis is essentially mediated by its proteolytic activity.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Enhancement of the synthesis and secretion of nerve growth factor in primary cultures of glial cells by proteases: a possible involvement of thrombin. 843 76

The IGFBP proteases were first described in pregnancy serum as a proteolytic activity against IGFBP-3. Since then, IGFBP proteases have been described in many other clinical situations, in various body fluids, and have been shown to cleave IGFBP-2 to -6 with varying specificity. The molecular nature of some of these proteases is being unraveled and three classes of IGFBP proteases have been recognized. These include kallikreins, cathepsins and matrix metalloproteinases (MMPs). We utilized two cellular systems to demonstrate the significance of IGFBP proteases in cellular growth regulation. In primary cultures of prostatic cells, we have shown that prostate-specific antigen (PSA) has the ability to enhance IGF mitogenic action by reducing the effects of IGFBPs. Similar kallikreins such as gamma nerve growth factor (NGF) share this activity. Within the prostatic milieu, we have also demonstrated epithelial production of the acid-activated IGFBP protease, cathepsin D, and its secretion into seminal plasma, as well as the serum of patients with prostate malignancy. We have also identified MMPs in prostatic cells and fluids. Using cultured airway smooth muscle (ASM) cells, we have demonstrated the synergism between IGFs and inflammatory agents in mediating ASM cell proliferation. Examination of this phenomenon revealed that these agents (e.g. leukotriene D4 and interleukin1-beta) induce the secretion of an IGFBP protease which cleaves the IGFBPs secreted by ASM cells, allowing IGFs to stimulate proliferation. Using several methods, including immunoblotting and immunodepletion techniques, we have identified this protease as MMP-1. These two pathophysiological systems demonstrate the importance of IGFBP proteases as autocrine paracrine growth regulators. Furthermore, IGFBP proteases may be critical elements in malignant and benign proliferative diseases, including prostate cancer and the ASM hyperplasia of long-standing asthma.
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PMID:Insulin-like growth factor binding protein (IGFBP) proteases: functional regulators of cell growth. 881 70


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