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Philip Stashenko - One of the best experts on this subject based on the ideXlab platform.
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expression of 92 kd type iv collagenase gelatinase b in human osteoclasts
Journal of Bone and Mineral Research, 2009Co-Authors: Anne L Wucherpfennig, William G Stetlerstevenson, Andrew E. Rosenberg, Yiping Li, Philip StashenkoAbstract:The digestion of type I collagen is an essential step in bone resorption. It is well established that osteoclasts solubilize the mineral phase of bone during the resorptive process, but the mechanism by which they degrade type I collagen, the major proteinaceous component of bone, is controversial. Differential screening of a human Osteoclastoma cDNA library was performed to characterize genes specifically expressed in osteoclasts. A large number of cDNA clones obtained by this procedure were found to represent 92 kD type IV collagenase (gelatinase B; MMP-9, EC 3.4.24.35), as well as tartrate-resistant acid phosphatase. In situ hybridization localized mRNA for gelatinase B to multinucleated giant cells in human Osteoclastomas. Gelatinase B immunoreactivity was demonstrated in giant cells from eight of eight Osteoclastomas, osteoclasts in normal bone, and osteoclasts of Paget's disease by use of a polyclonal antiserum raised against a synthetic gelatinase B peptide. In contrast, no immunoreactivity for 72 kD type IV collagenase (gelatinase A; MMP-2, EC 3.4.24.24), which is the product of a separate gene, was detected in Osteoclastomas or normal osteoclasts. We propose that the 92 kD type IV collagenase/gelatinase B plays an important role in the resorption of collagen during bone remodeling.
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Expression of 92 kD type IV collagenase/gelatinase B in human osteoclasts
Journal of Bone and Mineral Research, 2009Co-Authors: Anne L Wucherpfennig, William G. Stetler-stevenson, Andrew E. Rosenberg, Yiping Li, Philip StashenkoAbstract:The digestion of type I collagen is an essential step in bone resorption. It is well established that osteoclasts solubilize the mineral phase of bone during the resorptive process, but the mechanism by which they degrade type I collagen, the major proteinaceous component of bone, is controversial. Differential screening of a human Osteoclastoma cDNA library was performed to characterize genes specifically expressed in osteoclasts. A large number of cDNA clones obtained by this procedure were found to represent 92 kD type IV collagenase (gelatinase B; MMP-9, EC 3.4.24.35), as well as tartrate-resistant acid phosphatase. In situ hybridization localized mRNA for gelatinase B to multinucleated giant cells in human Osteoclastomas. Gelatinase B immunoreactivity was demonstrated in giant cells from eight of eight Osteoclastomas, osteoclasts in normal bone, and osteoclasts of Paget's disease by use of a polyclonal antiserum raised against a synthetic gelatinase B peptide. In contrast, no immunoreactivity for 72 kD type IV collagenase (gelatinase A; MMP-2, EC 3.4.24.24), which is the product of a separate gene, was detected in Osteoclastomas or normal osteoclasts. We propose that the 92 kD type IV collagenase/gelatinase B plays an important role in the resorption of collagen during bone remodeling.
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Molecular cloning and characterization of a putative novel human osteoclast-specific 116-kDa vacuolar proton pump subunit.
Biochemical and biophysical research communications, 1996Co-Authors: Wei Chen, Philip StashenkoAbstract:Abstract A cDNA encoding a possible novel human 116-kDa polypeptide subunit of the osteoclastic proton pump (OC-116KDa) has been identified by differential screening of a human Osteoclastoma cDNA library. The predicted sequence of OC-116KDa consists of 822 amino acids and is 46.9% and 47.2% identical at the amino acid level to the 116-KDa polypeptide of the vacuolar proton pump of rat and bovine brain respectively. OC-116KDa mRNA was found at high levels in Osteoclastomas by Northern analysis but was not detected in tumor stromal cells or in other tissues including kidney, liver, skeletal muscle and brain. OC-116KDa mRNA was localized to multinucleated giant cells within the Osteoclastoma tumor byin situhybridization.
Anne L Wucherpfennig - One of the best experts on this subject based on the ideXlab platform.
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expression of 92 kd type iv collagenase gelatinase b in human osteoclasts
Journal of Bone and Mineral Research, 2009Co-Authors: Anne L Wucherpfennig, William G Stetlerstevenson, Andrew E. Rosenberg, Yiping Li, Philip StashenkoAbstract:The digestion of type I collagen is an essential step in bone resorption. It is well established that osteoclasts solubilize the mineral phase of bone during the resorptive process, but the mechanism by which they degrade type I collagen, the major proteinaceous component of bone, is controversial. Differential screening of a human Osteoclastoma cDNA library was performed to characterize genes specifically expressed in osteoclasts. A large number of cDNA clones obtained by this procedure were found to represent 92 kD type IV collagenase (gelatinase B; MMP-9, EC 3.4.24.35), as well as tartrate-resistant acid phosphatase. In situ hybridization localized mRNA for gelatinase B to multinucleated giant cells in human Osteoclastomas. Gelatinase B immunoreactivity was demonstrated in giant cells from eight of eight Osteoclastomas, osteoclasts in normal bone, and osteoclasts of Paget's disease by use of a polyclonal antiserum raised against a synthetic gelatinase B peptide. In contrast, no immunoreactivity for 72 kD type IV collagenase (gelatinase A; MMP-2, EC 3.4.24.24), which is the product of a separate gene, was detected in Osteoclastomas or normal osteoclasts. We propose that the 92 kD type IV collagenase/gelatinase B plays an important role in the resorption of collagen during bone remodeling.
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Expression of 92 kD type IV collagenase/gelatinase B in human osteoclasts
Journal of Bone and Mineral Research, 2009Co-Authors: Anne L Wucherpfennig, William G. Stetler-stevenson, Andrew E. Rosenberg, Yiping Li, Philip StashenkoAbstract:The digestion of type I collagen is an essential step in bone resorption. It is well established that osteoclasts solubilize the mineral phase of bone during the resorptive process, but the mechanism by which they degrade type I collagen, the major proteinaceous component of bone, is controversial. Differential screening of a human Osteoclastoma cDNA library was performed to characterize genes specifically expressed in osteoclasts. A large number of cDNA clones obtained by this procedure were found to represent 92 kD type IV collagenase (gelatinase B; MMP-9, EC 3.4.24.35), as well as tartrate-resistant acid phosphatase. In situ hybridization localized mRNA for gelatinase B to multinucleated giant cells in human Osteoclastomas. Gelatinase B immunoreactivity was demonstrated in giant cells from eight of eight Osteoclastomas, osteoclasts in normal bone, and osteoclasts of Paget's disease by use of a polyclonal antiserum raised against a synthetic gelatinase B peptide. In contrast, no immunoreactivity for 72 kD type IV collagenase (gelatinase A; MMP-2, EC 3.4.24.24), which is the product of a separate gene, was detected in Osteoclastomas or normal osteoclasts. We propose that the 92 kD type IV collagenase/gelatinase B plays an important role in the resorption of collagen during bone remodeling.
Yiping Li - One of the best experts on this subject based on the ideXlab platform.
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expression of 92 kd type iv collagenase gelatinase b in human osteoclasts
Journal of Bone and Mineral Research, 2009Co-Authors: Anne L Wucherpfennig, William G Stetlerstevenson, Andrew E. Rosenberg, Yiping Li, Philip StashenkoAbstract:The digestion of type I collagen is an essential step in bone resorption. It is well established that osteoclasts solubilize the mineral phase of bone during the resorptive process, but the mechanism by which they degrade type I collagen, the major proteinaceous component of bone, is controversial. Differential screening of a human Osteoclastoma cDNA library was performed to characterize genes specifically expressed in osteoclasts. A large number of cDNA clones obtained by this procedure were found to represent 92 kD type IV collagenase (gelatinase B; MMP-9, EC 3.4.24.35), as well as tartrate-resistant acid phosphatase. In situ hybridization localized mRNA for gelatinase B to multinucleated giant cells in human Osteoclastomas. Gelatinase B immunoreactivity was demonstrated in giant cells from eight of eight Osteoclastomas, osteoclasts in normal bone, and osteoclasts of Paget's disease by use of a polyclonal antiserum raised against a synthetic gelatinase B peptide. In contrast, no immunoreactivity for 72 kD type IV collagenase (gelatinase A; MMP-2, EC 3.4.24.24), which is the product of a separate gene, was detected in Osteoclastomas or normal osteoclasts. We propose that the 92 kD type IV collagenase/gelatinase B plays an important role in the resorption of collagen during bone remodeling.
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Expression of 92 kD type IV collagenase/gelatinase B in human osteoclasts
Journal of Bone and Mineral Research, 2009Co-Authors: Anne L Wucherpfennig, William G. Stetler-stevenson, Andrew E. Rosenberg, Yiping Li, Philip StashenkoAbstract:The digestion of type I collagen is an essential step in bone resorption. It is well established that osteoclasts solubilize the mineral phase of bone during the resorptive process, but the mechanism by which they degrade type I collagen, the major proteinaceous component of bone, is controversial. Differential screening of a human Osteoclastoma cDNA library was performed to characterize genes specifically expressed in osteoclasts. A large number of cDNA clones obtained by this procedure were found to represent 92 kD type IV collagenase (gelatinase B; MMP-9, EC 3.4.24.35), as well as tartrate-resistant acid phosphatase. In situ hybridization localized mRNA for gelatinase B to multinucleated giant cells in human Osteoclastomas. Gelatinase B immunoreactivity was demonstrated in giant cells from eight of eight Osteoclastomas, osteoclasts in normal bone, and osteoclasts of Paget's disease by use of a polyclonal antiserum raised against a synthetic gelatinase B peptide. In contrast, no immunoreactivity for 72 kD type IV collagenase (gelatinase A; MMP-2, EC 3.4.24.24), which is the product of a separate gene, was detected in Osteoclastomas or normal osteoclasts. We propose that the 92 kD type IV collagenase/gelatinase B plays an important role in the resorption of collagen during bone remodeling.
Andrew E. Rosenberg - One of the best experts on this subject based on the ideXlab platform.
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expression of 92 kd type iv collagenase gelatinase b in human osteoclasts
Journal of Bone and Mineral Research, 2009Co-Authors: Anne L Wucherpfennig, William G Stetlerstevenson, Andrew E. Rosenberg, Yiping Li, Philip StashenkoAbstract:The digestion of type I collagen is an essential step in bone resorption. It is well established that osteoclasts solubilize the mineral phase of bone during the resorptive process, but the mechanism by which they degrade type I collagen, the major proteinaceous component of bone, is controversial. Differential screening of a human Osteoclastoma cDNA library was performed to characterize genes specifically expressed in osteoclasts. A large number of cDNA clones obtained by this procedure were found to represent 92 kD type IV collagenase (gelatinase B; MMP-9, EC 3.4.24.35), as well as tartrate-resistant acid phosphatase. In situ hybridization localized mRNA for gelatinase B to multinucleated giant cells in human Osteoclastomas. Gelatinase B immunoreactivity was demonstrated in giant cells from eight of eight Osteoclastomas, osteoclasts in normal bone, and osteoclasts of Paget's disease by use of a polyclonal antiserum raised against a synthetic gelatinase B peptide. In contrast, no immunoreactivity for 72 kD type IV collagenase (gelatinase A; MMP-2, EC 3.4.24.24), which is the product of a separate gene, was detected in Osteoclastomas or normal osteoclasts. We propose that the 92 kD type IV collagenase/gelatinase B plays an important role in the resorption of collagen during bone remodeling.
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Expression of 92 kD type IV collagenase/gelatinase B in human osteoclasts
Journal of Bone and Mineral Research, 2009Co-Authors: Anne L Wucherpfennig, William G. Stetler-stevenson, Andrew E. Rosenberg, Yiping Li, Philip StashenkoAbstract:The digestion of type I collagen is an essential step in bone resorption. It is well established that osteoclasts solubilize the mineral phase of bone during the resorptive process, but the mechanism by which they degrade type I collagen, the major proteinaceous component of bone, is controversial. Differential screening of a human Osteoclastoma cDNA library was performed to characterize genes specifically expressed in osteoclasts. A large number of cDNA clones obtained by this procedure were found to represent 92 kD type IV collagenase (gelatinase B; MMP-9, EC 3.4.24.35), as well as tartrate-resistant acid phosphatase. In situ hybridization localized mRNA for gelatinase B to multinucleated giant cells in human Osteoclastomas. Gelatinase B immunoreactivity was demonstrated in giant cells from eight of eight Osteoclastomas, osteoclasts in normal bone, and osteoclasts of Paget's disease by use of a polyclonal antiserum raised against a synthetic gelatinase B peptide. In contrast, no immunoreactivity for 72 kD type IV collagenase (gelatinase A; MMP-2, EC 3.4.24.24), which is the product of a separate gene, was detected in Osteoclastomas or normal osteoclasts. We propose that the 92 kD type IV collagenase/gelatinase B plays an important role in the resorption of collagen during bone remodeling.
Michael J. Warburton - One of the best experts on this subject based on the ideXlab platform.
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Degradation of bone matrix proteins by osteoclast cathepsins
The International journal of biochemistry, 1993Co-Authors: Alison E. Page, T.j. Chambers, Alison R. Hayman, L.m.b. Andersson, Michael J. WarburtonAbstract:Abstract 1. 1. The degradation of the bone matrix proteins osteocalcin, osteonectin and α2HS-glycoprotein by human cathepsins B and L and human Osteoclastoma cathepsins has been investigated. 2. 2. Intermediate degradation products (Mr > 12kDa) were not observed during the digestion of α2HS-glycoprotein and osteonectin by cathepsins B and L although they were observed with some of the Osteoclastoma cathepsins. Most of the Osteoclastoma cathepsins were capable of degrading these two proteins to small peptides at comparable rates. 3. 3. Each cathepsin produced a different pattern of osteocalcin degradation products. 4. 4. The extensive range of non-collagenous proteins in bone matrix may necessitate the production by osteoclasts of cathepsins with different specificities during bone resorption.
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Tartrate-resistant acid phosphatase from human Osteoclastomas is translated as a single polypeptide.
Biochemical Journal, 1991Co-Authors: Alison R. Hayman, T.j. Chambers, A J Dryden, Michael J. WarburtonAbstract:Tartrate-resistant acid phosphatases have been isolated from a number of sources. These enzymes consist of one subunit (Mr 30,000-40,000) or two dissimilar subunits (Mr 15,000-20,000). Previously we isolated the enzyme from human Osteoclastomas, as a two-subunit protein. By Northern blotting and hybridization with radiolabelled oligonucleotides corresponding to the N-terminal sequences of the two subunits, we demonstrate here that the enzyme is transcribed as one mRNA which is translated in vitro to produce a single polypeptide of approx. Mr 33,000. Transcription as a single mRNA species is also the case in other tissues. These results suggest that the Osteoclastoma enzyme undergoes post-translational modification in the form of cleavage of a single peptide bond to give a disulphide-bonded two-subunit protein.
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Purification and characterisation of cysteine proteinases from human Osteoclastomas
Biochemical Society transactions, 1991Co-Authors: Alison E. Page, Michael J. Warburton, Timothy J. Chambers, Alison R. HaymanAbstract:The process of normal bone remodeling requires the continual resorption and replacement of bone mineral and bone matrix proteins. Excessive bone resorption is a feature of a number of pathological conditions. These include osteoporosis, rheumatoid arthritis and Paget's disease [l]. Bone resorption is accomplished by the osteoclast. Osteoclasts attached to bone are highly polarised cells [2]. The osteoclast attaches to bone through a peripheral attachment zone within which the surface (apical) membrane is thrown into complex ruffles. The space enclosed by the osteoclast apical surface and the bone surface is acidified, probably by the action of a proton pump, resulting in the dissolution of bone mineral. Lysosomal enzymes are released into the acidified subosteoclastic space where they are responsible for the degradation of bone matrix components. Bone matrix is largely composed of type I collagen with small amounts of types 111, V and VI collagens [31. In addition, bone contains a number of glycoproteins (e.g. osteopontin, osteonectin and bone sialoproteins) and proteoglycans [4]. In vitro studies using disaggregated osteoclasts have suggested that cysteine proteinases released by osteoclasts are responsible for the degradation of bone proteins 151. Investigations of the properties of osteoclast cysteine proteinases are hampered by the scarcity of osteoclasts in normal bone. As our starting material we have used Osteoclastomas. These are primary bone tumours that contain large numbers of apparently normal osteoclasts. Using the synthetic peptide Z-PheArg-NHMec as substrate, we have separated six cysteine proteinase activities from Osteoclastoma extracts by sequential chromatography on S-Sepharose, phenyl-Sepharose, heparin-Sepharose and Sephacryl S200HR. The proteinases have molecular weights ranging from 20,000 to 42,000 (determined by gel filtration). Z-Phe-Arg-NHMec was the preferred synthetic substrate. Four of the enzymes showed some activity towards Bz-Phe-Val-Arg-NHMec but only two demonstrated low activity against Z-Arg-Arg-NHMec. The kinetics of hydrolysis of these substrates gave values within the range expected for cathepsin B [61. All six activities were capable of degrading soluble and insoluble collagen. The pH optima for the hydrolysis of Z-Phe-Arg-NHMec and type I collagen by the six enzymes are shown in table 1. A variety of inhibitors of cysteine proteinases (eg. leupeptin, E-64, cystatin) were potent inhibitors of the Osteoclastoma enzymes. The kinetics of inhibition by cystatin and the rate constants of inactivation by Z-PheTyr-(O-t-Bu)CHNz were similar to published values for cathepsin B [7,81. Immunolocalisation of cathepsin B in sections of Osteoclastomas revealed that osteoclasts stained strongly with an antibody to cathepsin B whereas the neoplastic osteoblasts were only weakly stained. The Table 1 pH-Optima of Osteoclastoma cysteiiie protciilases
