Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.3 (collagenase)
 18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have detected the presence of nuclear 3,5,3'-triiodothyronine (T3) receptors in primary cultures of chick embryo hepatocytes and neurons. Hepatocytes were isolated from livers of embryos of 12, 16 and 19 days by treatment with 0.2% collagenase and hyaluronidase. They were plated at a density of 3-4 x 10(5)/35-mm petri dish in Ham's F-10 medium containing fetal calf serum, tryptose phosphate, and antibiotics. Cells were used for the binding assay at Day 3 of culture. Neurons from 8-day-old embryo brains were cultured in a serum-free medium at a density of 1.2 x 10(6) cells/35-mm petri dish and used for the binding assay after 7 days of culture. Biological activity of hepatocytes was determined by measuring insulin binding, inositol phosphate formation, and 5'-monodeiodinase activity. Neurons or glial cells in culture were identified by immunostaining with anti-neurofilaments and anti-glial fibrillary acidic protein antisera. Binding assay was performed with isolated nuclei and 0.4 M NaCl nuclear extracts. With the latter preparation, the Scatchard analysis showed, in both cells, a single, high-affinity, low-capacity T3 receptor. In the hepatocytes of 12-, 16-, and 19-day-old embryos association constants (Ka) were, respectively, 0.93 +/- 0.02, 0.74 +/- 0.03, and 0.56 +/- 0.04 nM-1, whereas the maximal binding capacities (MBC) were 2.26 +/- 0.2, 2.72 +/- 0.33, and 1.83 +/- 0.19 fmol/microgram DNA (mean +/- SE, n = 3). In neurons Ka was 1.25 +/- 0.53 nM-1 and MBC 0.59 +/- 0.14 fmol/microgram DNA (n = 3). The receptor had a sedimentation coefficient of 3.4 S, an estimated Mr of 59 kDa, and the following relative affinity for thyroid hormone analogues: TRIAC greater than L-T3 greater than L-T4. These data indicate that cultured hepatocytes and neurons of chick embryo contained T3 receptors with properties similar to those described in intact tissues from this and other species. Only the MBC of neurons was 50% lower than that observed in whole brain of embryo, but was comparable to values observed in cultured neurons from other species.
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PMID:Characterization of 3,5,3'-triiodothyronine receptors in primary cultures of hepatocytes and neurons from chick embryo. 160 Dec 52

A new cell line MGM-1 was established from a primary tumor of the left temporal lobe with histological diagnosis of glioblastoma multiforme, removed from a 64-year-old Japanese male. The patient died of recurrence and unusual extracranial metastases of the tumor 7 months after the surgery. The cultured MGM-1 cells are spindle or polygonal in shape. After serial passages, glial fibrillary acidic protein became negative immunocytochemically in vitro. The modal chromosome number was 61-64. Doubling time and soft agar colony forming efficiency were 42.9h and 0.4%, respectively (at 25th passage). MGM-1 is a highly motile cell line in vitro and its serum-free conditioned medium is chemotactic and chemokinetic for other glioma cells. Secretion of gelatinases (probably MMP-2/72-kDa type i.v. collagenase) and MMP-9/92-kDa type i.v. collagenase) and urokinase-type plasminogen activator were also investigated. MGM-1 would therefore be useful for studying the mechanisms regulating glioma-cell motility and invasion. The MGM-1 cell line has been propagated continuously by serial passages (more than 100 passages) during the past 4 years.
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PMID:Establishment and characterization of a new human glioblastoma cell line (MGM-1) with highly motile phenotype. 923 71

The overall hypothesis that cell death after intracerebral hemorrhage is mediated in part by apoptotic mechanisms was tested. Intracerebral hemorrhage was induced in rats using stereotactic infusions of 0.5 U of collagenase (1-microL volume) into the striatum. After 24 hours, large numbers of TUNEL-positive stained cells with morphologies suggestive of apoptosis were present in the center and periphery of the hemorrhage. Double staining with Nissl and immunocytochemical labeling with antibodies against neuronal nuclei and glial fibrillary acidic protein suggested that these TUNEL-positive cells were mostly neurons and astrocytes. Electrophoresis of hemorrhagic brain extracts showed evidence of DNA laddering into approximately 200-bp fragments. Western blots showed cleavage of the cytosolic caspase substrate gelsolin. The density of TUNEL-positive cells at 24 and 48 hours after hemorrhage was significantly reduced by treatment with the broad-spectrum caspase inhibitor zVADfmk. It was unlikely that apoptotic changes were due to neurotoxicity of injected collagenase because TUNEL-positive cells and DNA laddering were also obtained in an alternative model of hemorrhage where autologous blood was infused into the striatum. Furthermore, equivalent doses of collagenase did not induce cell death in primary neuronal cultures. These results provide initial evidence that apoptotic mechanisms may mediate some of the injury in brain after intracerebral hemorrhage.
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PMID:Evidence for apoptosis after intercerebral hemorrhage in rat striatum. 1069 78

We have investigated the adhesion of the human fibrosarcoma cell line, HT-1080, transfected with glial fibrillary acidic protein (GFAP) to a variety of extracellular matrix macromolecules (ECM) including collagen type IV, laminin, and fibronectin. The GFAP-transfectants demonstrated altered adhesiveness to extracellular matrix substrates when compared to controls. GFAP-positive, heavy metal-induced fibrosarcoma cells were more adherent to plastic and collagen type IV than were the parental or uninduced cells. In contrast, GFAP-positive fibrosarcoma cells were less adherent to laminin- or fibronectin-coated dishes than controls. Time course adhesion studies over 9 days showed that the heavy metal-induced fibrosarcoma cells progressively became more adherent to collagen type IV and less adherent to laminin- or fibronectin-coated dishes than did uninduced cells. However, with the removal of heavy metal from the medium, the HT-1080 fibrosarcoma cells were restored to their original adhesive potential. By phase microscopy, uninduced and induced HT-1080 cells demonstrated different morphological features and remained viable in an anchorage-dependent fashion on collagen type IV as a substrate. By way of contrast, GFAP-induced HT-1080 cells were not particularly viable in monolayer culture and readily detached from laminin as a substrate. The expression of beta1 integrin in GFAP-positive fibrosarcoma cells was decreased following heavy metal induction by Western blot analyses. In contrast, the expression of alpha2 integrin was increased whereas alpha5 integrin was unchanged in HT-1080 cells following the induction of GFAP. Gelatin zymography showed that 72 kDa collagenase was less expressed in GFAP-induced clones than in controls. Our data suggest that the forced expression of the intermediate filament, GFAP, in HT-1080 cells may modulate cell adhesion to different ECM substrates through alterations in expression of integrins.
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PMID:Cytoskeletal-induced alterations in the adhesion of HT-1080 fibrosarcoma cells to extracellular matrix. 1129 52

Induction of EAE can be inhibited or repressed by administration of soluble metalloproteinase inhibitors. We studied the matrix metalloproteinase (MMP) and their tissue inhibitor (TIMP) expression pattern in experimental autoimmune encephalomyelitis (EAE) of the resistant Th2 prone BALB/c mouse, where the disease can be induced with ultrasound-emulsified antigen/adjuvant (son-ag), but not with conventional technique (syr-ag). We found highly elevated expression of MMP-8 (neutrophil collagenase) mRNA and protein in diseased son-ag challenged mice, colocalizing to neutrophil infiltrates found in brain and extensively in the spinal cord submeningeal space. MMP-8 expression has not been found previously in sensitive mouse strains. The infiltrates stained positive also for MMP-9 protein, and brain homogenates from corresponding mice showed MMP-9 activity during overt disease (days 12-16 post-immunization). TIMP-1 gene expression could be detected in CNS samples from diseased son-ag challenged mice but not in syr-ag or control mice, and the TIMP-1 protein colocalized with GFAP-staining. In contrast, in syr-ag mice both TIMP-2 and TIMP-3 gene expression in the spinal cords was elevated. The results show that sonication, but not extrusion, creates an adjuvant formula potent in activating the matrix metalloproteinase cascade similar to sensitive mouse strains, strongly implicating their role in EAE induction in this Th2 prone strain. The study provides the basis for establishment of MMP-specific therapy in this model.
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PMID:Up-regulation of MMP-8 and MMP-9 activity in the BALB/c mouse spinal cord correlates with the severity of experimental autoimmune encephalomyelitis. 1198 14

Hepatic stellate cells (HSCs), also referred to as Ito cells, perisinusiodal cells and fat-storing cells, have numerous vital functions. They are the main extracellular matrix-producing cells within the liver and are involved in the storage of retinol. HSCs are also known to secrete a number of liver mitogens. Current isolation techniques are cumbersome and most require a pronase digestion step, which destroys any hepatocytes present. We present a simple method for isolation and culture of hepatic stellate cells from the normally discarded washings from a two-step collagenase hepatocyte isolation, which has shown a yield of more than 1.5 x 10(6) viable HSCs after 5 days in culture. The cells were positively identified as HSCs by staining for two intermediate filaments (desmin and GFAP) and observing their distinct morphology from other liver cell types. This efficient method allows rapid and consistent isolation of stellate cells to give a culture that may be passaged several times.
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PMID:A simple method for the simultaneous isolation of stellate cells and hepatocytes from rat liver tissue. 1287 Jun 60

Dexamethasone was evaluated for the treatment of intracerebral hemorrhage using a rat model of cerebral hematoma induced by intracerebral injection of collagenase. The treatment group consisted of hematoma rats receiving dexamethasone 1 mg/kg intraperitoneal (i.p.) at 1 and 24 h following surgery. Controls included hematoma rats receiving saline i.p. and sham-operated animals receiving saline i.p. Each animal was evaluated neurologically prior to, as well as 24 and 48 h following surgery. After the last neurological evaluation, animals were deeply anesthetized and the brain was removed following perfusion for microscopic examination and glial fibrillary acidic protein immunohistochemistry. Behavioral scores were significantly improved in the treated group (P < 0.0001). The hematoma volume was significantly smaller (P < 0.02). Neutrophils and astrocytes were less numerous in the hematoma of dexamethasone-treated animals (P < 0.001), however the number of necrotic neurons in the penumbra was not changed by the treatment. The number of necrotic neurons in the cerebral cortex was less in treated than in nontreated animals (P < 0.01). Controls had many vascular changes including necrotic endothelium and fibrin deposits compared with treated animals. In conclusion, dexamethasone administered shortly after an intracerebral hematoma appears beneficial for the treatment of this condition.
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PMID:Evaluation of dexamethasone for the treatment of intracerebral hemorrhage using a collagenase-induced intracerebral hematoma model in rats. 1550 May 70

The goal of studies of autoimmune disease biomarkers is to identity markers that fluctuate with disease development and severity, but then normalize following successful therapy. The perfect marker could thus serve as a diagnostic tool, as well as a monitoring device for therapeutic drug efficacy. Current biomarker discovery efforts are focused on three groups of proteins reflective of the autoimmune disease process: (1) degradation products arising from destruction of affected tissues, (2) enzymes that play a role in tissue degradation and (3) cytokines and other proteins associated with immune activation. Potential biomarkers for two autoimmune diseases, rheumatoid arthritis and multiple sclerosis, have been described in recent publications. For rheumatoid arthritis, these markers (by group) include (1) aggrecan fragments, C-propeptide of type II collagen and cartilage oligomeric matrix protein, (2) matrix metalloprotease (MMP)-1, MMP-3 and MMP-1/inhibitor complexes and (3) thioredoxin, IL-16 and tumour necrosis factor (TNF)-alpha. For multiple sclerosis, they include (1) neurofilament light protein and glial fibrillary acidic protein, (2) MMP-2 and MMP-9 and (3) TNF-alpha and soluble vascular adhesion molecule-1. The utility of most of these markers is limited by their restriction to relatively inaccessible anatomic sites (synovial or cerebrospinal fluid). Thus, from a practical standpoint, the most useful autoimmune biomarkers will be those measurable in serum or plasma.
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PMID:Biomarkers for diagnosing and monitoring autoimmune diseases. 1629 11

Hepatic stellate cells (HSC) are located in Disse spaces of normal rat liver. In their quiescent state they serve as a storage site for vitamin A. In fibrotic liver they become activated, proliferate and they undergo transdifferentiation into myofibroblast-like cells. Changes in the cell phenotype are accompanied by changes in the cellular cytoskeleton. We have studied the expression of alpha-smooth muscle actin and intermediate filament proteins vimentin, desmin and glial fibrillary acidic protein (GFAP) by immunocytochemistry in HSC cultured for 2 or 7 days after isolation. Normal or cirrhotic rat liver was perfused with solutions of pronase and collagenase and HSC were isolated by density gradient centrifugation of the resulting cell suspension. Liver cirrhosis was produced in rats by repeated carbon tetrachloride administration. Vimentin was detected in all cells from normal and cirrhotic liver. The concentration of desmin in the cells from cirrhotic liver was slightly higher than that in normal cells and it increased with time in culture. GFAP could be detected only in normal cells 2 days after their isolation. In contrast, alpha smooth muscle actin (alpha-SMA) was absent from normal cells at this time but its expression was pronouced later. In most cells from cirrhotic liver this antigen was already present on the second day of culture and its expression further increased.
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PMID:Expression of cytoskeletal proteins in hepatic stellate cells isolated from normal and cirrhotic rat liver. 1664 26

Human processed lipoaspirate is a source of multipotent adult stem cells that are able to differentiate between mesenchymal and neurogenic lineage. We characterized PLA cells by cytometry and then they were cultured to induce differentiation into myogenic and neurogenic lineage. Lipoaspirates were digested with collagenase to obtain the pellet, which was labelled with anti-CD44, anti-CD45, and anti-CD90. We used BD FACS Calibur flow cytometer to acquire cellular events. Some cells were cultured at 37 degrees C and 5% CO(2) in neurogenic or myogenic medium and analysed by immunocytochemistry, using Neuron specific enolase, Vimentin, Glial fibrillary acidic protein, Tau, MAP2 to confirm neurogenic differentiation, MyoD1, Myosin heavy chain, Actin smooth muscle, vimentin to confirm myogenic differentiation. The cytometry results suggest that a part of the cells are of a mesenchymal origin, among which there are progenitor endothelial cells and leucocytes. Microscopy observation already reveals neuronal morphology and longitudinal multinucleated cells compared to control cells. Neurogenic cells only express NSE (early neuronal progenitor marker), but not GFAP, Tau, MAP2 (mature neuron and glial markers); myogenic cells are positive for MyoD1 and Myosin heavy chain. This demonstrates that lipoaspirate cells are capable of differentiating in vitro over a short period of time, and could be employed in biological and clinical research, like mesenchymal adult stem cells.
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PMID:Characterization and induction of human pre-adipocytes. 1711 45


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