Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.6.4 (chondroitinase)
 2,039 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Transfer of radioactive materials to fixed cells from an overlying layer of living cells has been examined to determine whether fixed cells can act as acceptors of glycosyltransferases of living cells. After the incubation of living cells were removed by EDTA treatment, and the radioactivity associated with the fixed cells was determined. Lipids, proteins and carbohydrates were found to be transfered from the living cells to the fixed cells. The amount of radioactivity transferred to the fixed cells was dependent on the number of both fixed and living cells and increased with the time of incubation. When fixed cells were treated with chloroform-methanol before the addition of living cells, the transfer of both lipids and proteins to the fixed cells decreased drastically, but only a slight decrease incarbohydrate transfer was observed. Most of the radioactive materials transferred from living cells labeled with glucosamine or fucose to chloroform-methanol-treated fixed cells were solubilized by trypsin but not by the detergents tested. Approximately 55% of the materials transferred from the cells labeled with glucosamine could be solubilized by hyaluronidase and chondroitinase, and the rest was solubilized by neuraminidase and a glycosidase mixture. The treatment of chloroform-methanol-extracted fixed cells with trypsin caused a significant decrease in the transfer from cells labeled with glucosamine. When nucleotide sugars were used as the radioactive precursor, no significant amount of radioactivity was transferred to the fixed cells.
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PMID:Cellular interaction between fixed and living cells. Transfer of radioactive materials from living cells to fixed cells. 37 19

Constituents of the bone marrow microenvironment have the capacity to influence both normal and malignant hematopoietic cell behavior. For example, HL-60 human promyelocytic leukemia cells in vitro display a more mature phenotype when grown on a bone marrow stroma-derived matrix. To elucidate which component(s) of the stromal matrix is capable of modulating HL-60 cell phenotype, matrices were treated with a variety of chemicals and enzymes prior to being used in the differentiation assay. Treatment of matrices with collagenase, pronase, chondroitinase, or chloroform:methanol:ether could not abolish the differentiation-promoting activity of bone marrow stroma. In contrast, the activity was destroyed by alkali treatment (0.5 M NaOH for 18 h) or heparinase/heparitinase enzymes. Heparin added to cultures increased maturation of HL-60 cells as determined by esterase production, Fc rosette formation, and morphological appearance. Other stromal components such as laminin, fibronectin, collagen I, collagen IV, or chondroitin sulfate did not alter the HL-60 leukemia cell phenotype. Stroma-derived matrix material which labeled with [35S]sulfate and eluted on a DEAE ion-exchange column as a high ionic fraction in 1.5 M LiCl and 7.5% sodium dodecyl sulfate contained the active fraction. A heparan sulfate proteoglycan component isolated by polyacrylamide-agarose gel electrophoresis induced a more mature HL-60 phenotype, and digestion with heparinase/heparitinase in the presence of protease inhibitors abrogated the effects on HL-60 phenotype. We conclude that a heparan sulfate-associated fraction of the bone marrow matrix plays a key role in the regulation of leukemic cell maturation.
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PMID:A heparan sulfate-containing fraction of bone marrow stroma induces maturation of HL-60 cells in vitro. 214 Feb 91

The mineral deposits in rabbit articular cartilage induced by intra-articular injections of glucocorticoid were studied by light and electron microscopy, using histochemical techniques and x-ray-probe microanalysis. This study demonstrated that the mineral deposits consisted of hydroxyapatite crystals. The initial deposition of hydroxyapatite crystals was seen around degenerating chondrocytes, where a halo-like pericellular space contained a large amount of electron-dense amorphous material. The initial precipitation of the crystals with a low ratio of calcium to phosphorus and the subsequent growth of crystals were seen only on or within the electron-dense amorphous material until the crystals formed mature, calcified nodules. The electron-dense amorphous material frequently coexisted with proteoglycans and degenerated collagen fibers. Digestion studies using chondroitinase ABC, papain, or chloroform and methanol suggested that the electron-dense amorphous material consisted of some protein and a small amount of lipid. Matrix vesicles were rarely seen in the calcifying areas. In addition, there was a correlation between sulphur, calcium, and phosphorus in the calcifying areas, where the relative element concentrations were: S (estimation counts of sulphur) = -0.862 X (calcium counts) + 1.472 X (phosphorus counts) + 102.146. This study demonstrated that electron-dense amorphous material, proteoglycans, and degenerated collagen fibers are present in loci where the hydroxyapatite crystals are formed in articular cartilage.
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PMID:Hydroxyapatite deposition in articular cartilage by intra-articular injections of methylprednisolone. A histological, ultrastructural, and x-ray-microprobe analysis in rabbits. 300 26

A human follicular fluid (HFF) fraction prepared by Sephadex G-75 column chromatography has been previously shown by this laboratory to initiate the human sperm acrosome reaction (AR) in vitro. In the present report, the apparent molecular weight (MW) of this AR activity determined by a longer G-75 column than was used in the previous work was 50,000 +/- 5,106. The G-75 Sephadex void volume fractions of some but not all HFF samples were also found to contain some AR-initiating activity. The occasional void volume activity was less potent than that of the 50,000 MW fraction and was not studied further. Further characterization of the 50,000 MW fraction was carried out. A time-course study demonstrated that maximum AR were obtained within 5 min following the addition of the 50,000 MW fraction. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by silver staining revealed that the 50,000 MW fraction was still a relatively crude preparation. Treatment of the 50,000 MW fraction with chloroform:methanol did not extract the AR-initiating activity into the lipid phase. The AR-initiating activity of the untreated 50,000 MW fraction was precipitated when it was boiled, but the activity was partially resistant to boiling after overnight incubation. Treatment of the 50,000 MW fraction with pronase E or with several glycosaminoglycan hydrolases did not destroy the activity. Pronase treatment resulted in a higher amount of boiling-resistant AR-initiating activity. The AR-initiating activity of the untreated 50,000 MW fraction was partially dialyzable, but the activity of an undialyzed fraction did not pass through an ultrafiltration membrane with a 10,000 MW cut-off. However, treatment of the 50,000 MW fraction with protease, peptide:N-glycosidase F, and to a lesser extent chondroitinase ABC yielded an active lower MW activity which could pass through such an ultrafiltration membrane. The lower MW activity released by peptide:N-glycosidase F eluted in the included volume (5,000-1,000) of a Sephadex G-25 column. Neutral hexose but not protein or peptide was detected in the G-25 peak of AR-initiating activity. These results suggest that the AR-initiating activity present in the 50,000 MW fraction of HFF: 1) is present either as two different AR factors (a high-MW factor and a low-MW, noncovalently bound factor) or as a single factor responsible for both the nondialyzable and dialyzable AR-initiating activities (the latter being enzymatically released from the former), and 2) may be at least partially associated with N-linked oligosaccharides of a glycoprotein or proteoglycan.
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PMID:Partial characterization of a fraction from human follicular fluid that initiates the human sperm acrosome reaction in vitro. 323 26

This report describes the detection and partial characterization of preovulatory human cumulus oophorus and mural granulosa cell-associated activity capable of initiating the human sperm acrosome reaction (AR) in vitro. Fragments of preovulatory human cumulus (cells plus extracellular matrix) were washed 3 times, incubated for 24 hr and the spent media and washes assayed for their ability to initiate the human sperm acrosome reaction (AR) in vitro. AR activity was present in the first two washes but not the third wash; however, AR activity was recovered in the spent medium after 3 X-washed fragments were incubated for 24 hr under conditions which maintained the viability of the cumulus cells. The spent media of preovulatory human mural granulosa cells contained AR-initiating activity after 1-3, 3-6, and 6-9 days of culture. The properties of the AR activity present in spent media of human cumulus fragments included resistance to loss of activity during treatment with pronase; resistance to loss of activity during treatment with chondroitinase ABC or bacterial hyaluronidase; heat stability after overnight incubation; lack of extraction by chloroform-methanol; an apparent molecular weight (MW) of 50,000, as determined by Sephadex G-75 column chromatography; conversion to a lower apparent MW activity by incubation with pronase. These properties are also characteristic of a fraction derived by Sephadex G-75 chromatography of preovulatory human follicular fluid which also has been shown to stimulate the human sperm acrosome reaction in vitro. The AR activity from spent media of human mural granulosa cells is also found in a 50,000 MW Sephadex G-75 fraction. We propose that the sources of the 50,000 MW human follicular fluid AR activity are the cumulus oophorus and the mural granulosa cells.
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PMID:Human sperm acrosome reaction-initiating activity associated with the human cumulus oophorus and mural granulosa cells. 338 73

Polysaccharides and other complex carbohydrates were released by proteolysis of the chloroform-methanol insoluble residue of 10 day-old worms and eggs of Hymenolepis diminuta. Gas-liquid chromatographic analysis of alditol acetate derivatives of monosaccharides released from the polysaccharides by hydrolysis revealed that in the 10 day-old worm, glucose was the most abundant sugar, followed by galactose, glucosamine, galactosamine, fucose and possibly rhamnose. Mannose was least abundant and xylose was absent. In the egg, glucose and galactose were equally abundant, followed by the same sugars found in 10 day-old worms, and xylose was present. Uronic acid was detected in both fractions by specific chemical tests. None of the saccharide material from eggs and worms was susceptible to degradation by Streptomyces hyaluronidase, chondroitinase AC, and slightly susceptible to chondroitinase ABC, as shown by electrophoretic analysis on composite 2.2% acrylamide-agarose slab gels and 4.5/12.5% polyacrylamide gels before and after enzymatic treatment. One of the gel-separable bands, however, was degradable by both nitrous acid and Flavobacterium heparinase. Both bands from eggs were degradable by nitrous acid. These results suggest that eggs contain heparin and/or heparan sulfate and perhaps dermatan sulfate and that 10 day-old worms also have these polyglycans but possibly not chondroitin sulfate or hyaluronic acid.
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PMID:Characterization of polysaccharides of the eggs and adults of Hymenolepis diminuta. 653 86

This study deals with the ultrastructure of the chondroitin sulphate proteoglycans of the Kurloff body, a large lysosomal organelle that stains metachromatically with Toluidine Blue and which is present in Kurloff cells (a blood cell unique to the guinea pig). Splenic tissues were fixed with 1% cetylpyridinium chloride (CPC) added to 4% paraformaldehyde and examined either after Spicer's high-iron diamine staining for sulphated anionic sites followed by post-fixation with ferrocyanide-osmium tetroxide or after a simple post-fixation with ferrocyanide-osmium tetroxide. CPC-precipitated sulphated sites were preferentially located at the periphery of the Kurloff body but, unexpectedly, were absent in the central matrix. Although their electron opacity was lower, these anionic sites were readily observable in the absence of HID-staining after sole post-fixation by ferrocyanide-reduced osmium. CPC-precipitated sulphated anionic sites were either associated with the myelin figures or constituted unexpected structures. They contained (i) tightly-stacked lamellae, with a very regular 4 nm periodicity, and (ii) groups of 2, 3, 4 short dense lines with a 3-5 nm periodicity. By taking into account the susceptibility of these HID-reactive structures towards chondroitinase ABC, these different sulphated components were assumed to be related to the proteochondroitin-4-sulphate previously characterized as the only major sulphated glycoconjugate of the Kurloff cell. Their colocalization with phospholipidic structures was suggested following observation of sections treated by a chloroform-methanol mixture.
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PMID:Kurloff cell ultrastructure after combined formaldehyde-cetylpyridinium chloride fixation and high-iron diamine staining. 767 81

Cytochemical and immunocytochemical approaches have been applied to the study of the surface of articular cartilage in humans, bovine and rats. Specimens were fixed in situ or soon after bioptic sampling with chemicals able to preserve and visualize proteins (glutaraldehyde, tannic acid), lipids (osmium tetroxide, malachite green, uranyl acetate) and proteoglycans (toluidine blue O, cuprolinic blue, cetyl pyridinium chloride). Mixtures of reagents were also used. Oriented serial thin sections were observed as such or after treatment with chemicals (chloroform-methanol, Triton X 100) or enzymes (chondroitinases, hyaluronidases, trypsin). Hyaluronan was detected by the use of glial-hyaluronate-binding-protein and antibodies against it. High concentration of osmium tetroxide or fixatives containing markers for lipid or for proteoglycans revealed that the surface of the articular cartilage, in all animal species examined, was covered by mono-multilayered discontinuous three-laminar sheets, which could be partly removed by chloroform-methanol and Triton X 100, were sensitive to hyaluronidase, chondroitinase and trypsin, and were immunopositive for hyaluronan. Each three-laminar sheet was 12-14 nm thick, was always separated from the cartilage itself and could be easily displaced. It is proposed that the surface of normal articular cartilage is covered by a discontinuous mono/multilayered pseudo-membrane, that can be better preserved by fixatives injected into the joint cavity and seems to consist of phospholipids, glycosaminoglycans and proteins. This membrane-like structure might have a protecting role in preventing direct contacts between the articular cartilage and toxic agents present in the synovial fluid and/or exert a lubricating effect within the articular joint.
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PMID:Ultrastructural identification of a membrane-like structure on the surface of normal articular cartilage. 876 81

We performed the quantitative analysis of chondroitin sulfate (CS) obtained from raw materials and various pharmaceutical preparations. To quantify CS content in raw materials and in an ophthalmic solution, each test sample and the authentic CS were first digested by chondroitinase ABC. The CS disaccharides produced were analyzed by strong anion-exchange high-performance liquid chromatography (SAX-HPLC) and CS content was quantified by calculating the total peak areas of the disaccharides derived from a CS calibration curve. In the case of soft capsules, CS was first extracted with hexane followed by phenol-chloroform to remove oil and protein ingredients. The extracted CS samples were depolymerized by chondroitinase ABC and CS content was determined. Quantitative analysis of the disaccharides derived from raw materials and an ophthalmic solution showed the CS contents (%) were 39.5+/-0.1 to 105.6+/-0.1 and 103.3+/-1.2, respectively. In case of CS analysis in soft capsules and liquid preparations, the overall recovery (%) of the spiked CS was 96.79+/-0.53-103.54+/-1.78 and 97.10+/-1.82 to 103.17+/-2.34, respectively. In conclusion, the quantitative analysis of the disaccharides produced by enzymatic digestion can be used in the direct quantitation of CS containing pharmaceutical formulations.
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PMID:Quantitative analysis of chondroitin sulfate in raw materials, ophthalmic solutions, soft capsules and liquid preparations. 1573 52