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Y Yoshihara - One of the best experts on this subject based on the ideXlab platform.

  • superficial zone chondrocytes in normal and osteoarthritic human articular cartilages synthesize novel truncated forms of inter alpha trypsin Inhibitor heavy chains which are attached to a chondroitin sulfate proteoglycan other than bikunin
    Osteoarthritis and Cartilage, 2008
    Co-Authors: Y Yoshihara, Barbara Osborn, Arkady Margulis, F Nelson, Matthew C Stewart, Caroline M Milner, Marylin S Rugg, Anna Plaas, K Nemoto
    Abstract:

    OBJECTIVE: We have examined the occurrence of the inflammation-associated Inter-Alpha-Trypsin Inhibitor (IalphaI) components, bikunin, heavy chain (HC)1 and HC2 in normal cartilage and osteoarthritis (OA) cartilage and synovial fluids. DESIGN/METHODS: Cartilage extracts from normal donors and late-stage OA patients, and synovial fluids from OA patients were studied by Western blot with multiple antibodies to bikunin, HC1 and HC2. Cell and matrix localization was determined by immunohistochemistry and mRNA by RT-PCR. RESULTS: Bikunin.chondroitin sulfate (CS) and IalphaI were abundant in OA cartilages, but virtually undetectable in normal. In both OA and normal cartilages, HCs were largely present in a novel C-terminally truncated 50-kDa form, with most, if not all of these being attached to CS on a proteoglycan other than bikunin. Synovial fluids from OA patients contained bikunin.CS and full-length (approximately 90 kDa) HCs linked to hyaluronan (HA) as HC.HA (SHAP.HA). Immunohistochemistry showed intracellular and cell-associated staining for bikunin and HCs, consistent with their synthesis by superficial zone chondrocytes. PCR on multiple human normal and OA cartilage samples detected transcripts for HC1 and HC2 but not for bikunin. In OA cartilages, immunostaining was predominantly matrix-associated, being most intense in regions with a pannus-like fibrotic overgrowth. CONCLUSION: The truncated structure of HCs, their attachment to a proteoglycan other than bikunin, PCR data and intracellular staining are all consistent with synthesis of HC1 and HC2 by human articular chondrocytes. The presence of bikunin.CS and IalphaI in OA cartilage, but not in normal, appears to be due to diffusional uptake and retention through fibrillated (but not deeply fissured) cartilage surfaces.

  • superficial zone chondrocytes in normal and osteoarthritic human articular cartilages synthesize novel truncated forms of inter alpha trypsin Inhibitor heavy chains which are attached to a chondroitin sulfate proteoglycan other than bikunin
    Osteoarthritis and Cartilage, 2008
    Co-Authors: Y Yoshihara, Barbara Osborn, Arkady Margulis, F Nelson, Matthew C Stewart, Caroline M Milner, Marylin S Rugg, Anna Plaas, Anthony J Day
    Abstract:

    Summary Objective We have examined the occurrence of the inflammation-associated Inter-Alpha-Trypsin Inhibitor (IαI) components, bikunin, heavy chain (HC)1 and HC2 in normal cartilage and osteoarthritis (OA) cartilage and synovial fluids. Design/methods Cartilage extracts from normal donors and late-stage OA patients, and synovial fluids from OA patients were studied by Western blot with multiple antibodies to bikunin, HC1 and HC2. Cell and matrix localization was determined by immunohistochemistry and mRNA by RT-PCR. Results Bikunin·chondroitin sulfate (CS) and IαI were abundant in OA cartilages, but virtually undetectable in normal. In both OA and normal cartilages, HCs were largely present in a novel C-terminally truncated 50-kDa form, with most, if not all of these being attached to CS on a proteoglycan other than bikunin. Synovial fluids from OA patients contained bikunin·CS and full-length (∼90kDa) HCs linked to hyaluronan (HA) as HC·HA (SHAP·HA). Immunohistochemistry showed intracellular and cell-associated staining for bikunin and HCs, consistent with their synthesis by superficial zone chondrocytes. PCR on multiple human normal and OA cartilage samples detected transcripts for HC1 and HC2 but not for bikunin. In OA cartilages, immunostaining was predominantly matrix-associated, being most intense in regions with a pannus-like fibrotic overgrowth. Conclusion The truncated structure of HCs, their attachment to a proteoglycan other than bikunin, PCR data and intracellular staining are all consistent with synthesis of HC1 and HC2 by human articular chondrocytes. The presence of bikunin·CS and IαI in OA cartilage, but not in normal, appears to be due to diffusional uptake and retention through fibrillated (but not deeply fissured) cartilage surfaces.

Antonietta Salustri - One of the best experts on this subject based on the ideXlab platform.

  • impaired cumulus mucification and female sterility in tumor necrosis factor induced protein 6 deficient mice
    Development, 2003
    Co-Authors: Csaba Fulop, Sandor Szanto, Durba Mukhopadhyay, Tamas Bardos, Rajesh V Kamath, Marylin S Rugg, Antonietta Salustri, Vincent C Hascall, Tibor T Glant, Katalin Mikecz
    Abstract:

    Mucification of the cumulus layer around the oocyte is an obligatory process for female fertility. Tumor necrosis factor-induced protein-6 (TNFIP6 or TSG6) has been shown to be specifically expressed during this process. We have generated TNFIP6-deficient mice and tested the ability of their cumulus cells to undergo mucification. Cumulus cell-oocyte complexes fail to expand in TNFIP6-deficient female mice because of the inability of the cumulus cells to assemble their hyaluronan-rich extracellular matrix. The impaired cumulus matrix formation is due to the lack of covalent complexes between hyaluronan and the heavy chains of the inter-α-trypsin Inhibitor family. As a consequence, TNFIP6-deficient females are sterile. Cultured TNFIP6-deficient cumulus cell-oocyte complexes also fail to expand when stimulated with dibutyryl cyclic AMP or epidermal growth factor. Recombinant TNFIP6 is able to catalyze the covalent transfer of heavy chains to hyaluronan in a cell-free system, restore the expansion of Tnfip6 -null cumulus cell-oocyte complexes in vitro, and rescue the fertility in Tnfip6 -null females. These results provide clear evidence that TNFIP6 is a key catalyst in the formation of the cumulus extracellular matrix and indispensable for female fertility.

  • two distinct populations of tumor necrosis factor stimulated gene 6 protein in the extracellular matrix of expanded mouse cumulus cell oocyte complexes
    Archives of Biochemistry and Biophysics, 2001
    Co-Authors: Durba Mukhopadhyay, Antonietta Salustri, Vincent C Hascall, Anthony J Day, Csaba Fulop
    Abstract:

    After the luteinizing hormone surge, the cumulus cell-oocyte complexes (COCs) in the preovulatory follicles produce a viscoelastic extracellular matrix, a process that requires the synthesis of hyaluronan as well as the incorporation of some components of the Inter-Alpha-Trypsin Inhibitor (IalphaI) family. In this study we report, that a hyaluronan-binding protein, the translated product of tumor necrosis factor-stimulated gene-6 (TSG-6), is also specifically accumulated in this matrix. TSG-6 mRNA expression is quickly upregulated and peaks at approximately 1500 copies/cell 4 h after the ovulatory stimuli as assessed by quantitative reverse transcription-polymerase chain reaction. Immunohistochemistry reveals the colocalization of the TSG-6 protein and hyaluronan around the cumulus and granulosa cells. The TSG-6 protein exists in two distinct populations in the COC matrix as demonstrated by Western-blot analysis. One population is a monomer that is anchored to the matrix by a noncovalent interaction. The second population is a covalent complex with either of the heavy chains of IalphaI and is bound to hyaluronan through a strong interaction that is resistant to denaturing conditions. The specific incorporation of the TSG-6 protein into the COC matrix suggests a structural role for this molecule.

  • Coding sequence, exon-intron structure and chromosomal localization of murine TNF-stimulated gene 6 that is specifically expressed by expanding cumulus cell-oocyte complexes.
    Gene, 1997
    Co-Authors: C Fülöp, Antonietta Salustri, Tibor T Glant, R V Kamath, J M Otto, B R Olsen, Vincent C Hascall
    Abstract:

    Tumor necrosis factor stimulated gene-6 (TSG-6) has been previously shown to be induced in vitro in several cell types by proinflammatory cytokines, and in vivo in pathological conditions such as rheumatoid arthritis. In this study, we report the complete coding sequence for the mouse TSG-6 protein, and the exon intron structure and the chromosomal localization of the gene. We have identified a 1605 nt cDNA sequence from mouse cumulus cell oocyte complexes (COCs) induced to expand in vivo. The sequence contains an open reading frame of 825 nt that codes for the 275 amino acid TSG-6 protein. The gene contains six exons separated by 1.1-5.8 kb introns and has been localized to the murine chromosome 2 by linkage analysis. Comparative reverse transcription-polymerase chain reaction studies have revealed that TSG-6 mRNA is specifically expressed after COC expansion induced in vivo, identifying the first non-pathological process in which TSG-6 may play an important role. Since TSG-6 binds to hyaluronan and interacts with Inter-Alpha-Trypsin Inhibitor (IalphaI), molecules that are essential for matrix formation by COCs, this protein may have a structural role in the matrix or may enhance the antiproteolytic effect of IalphaI to protect the matrix from degradation.

  • effects of exogenous hyaluronic acid and serum on matrix organization and stability in the mouse cumulus cell oocyte complex
    Journal of Biological Chemistry, 1993
    Co-Authors: Antonella Camaioni, Vincent C Hascall, Masaki Yanagishita, Antonietta Salustri
    Abstract:

    Compact cumulus cell-oocyte complexes (COCs) isolated from preovulatory mouse follicles undergo expansion in vitro when high levels of hyaluronic acid (HA) are synthesized and organized into an extracellular matrix. We studied the effects of fetal bovine serum (FBS) and of exogenous HA and HA-oligomers on the expansion process. Maximum retention of HA in the COC matrix, and hence complete COC expansion, occurs when 1% FBS is continuously present during the first 18 h of culture. Irrespective of the culture time, HA synthesized when serum is absent is primarily in the medium, whereas HA synthesized when serum is present is primarily in the cell matrix. These findings support the hypothesis that the serum factor, identified as an Inter-Alpha-Trypsin Inhibitor by Chen et al. (Chen, L., Mao, S. J., and Larsen, W. J. (1992) J. Biol. Chem. 267, 12380-12386), is a structural component of the matrix. Addition of exogenous HA or of HA oligomers of decasaccharide size (GlcUA-GlcNAc)5 or larger effectively displaces endogenously synthesized HA from the matrix into the medium, thereby preventing COC expansion. Addition of exogenous chondroitin sulfate affects neither matrix organization nor COC expansion, thus indicating specificity of the binding of some structural component(s) to HA. Fully expanded COCs disassemble when cultured longer than 18 h, a process which occurs also in vivo and which correlates with loss of oocyte fertilizability both in vivo and in vitro. This process involves release of macromolecular HA from the matrix into the medium, with loss of 50% of the HA in the first 8 h of incubation after full expansion. The release is not facilitated when HA oligomers, long enough to prevent matrix formation, are added to the culture medium after the COCs are fully expanded. This suggests that cooperative binding to HA of either the serum factor, an endogenously synthesized factor(s), or both is required to stabilize the fully expanded COC matrix.

Anthony J Day - One of the best experts on this subject based on the ideXlab platform.

  • superficial zone chondrocytes in normal and osteoarthritic human articular cartilages synthesize novel truncated forms of inter alpha trypsin Inhibitor heavy chains which are attached to a chondroitin sulfate proteoglycan other than bikunin
    Osteoarthritis and Cartilage, 2008
    Co-Authors: Y Yoshihara, Barbara Osborn, Arkady Margulis, F Nelson, Matthew C Stewart, Caroline M Milner, Marylin S Rugg, Anna Plaas, Anthony J Day
    Abstract:

    Summary Objective We have examined the occurrence of the inflammation-associated Inter-Alpha-Trypsin Inhibitor (IαI) components, bikunin, heavy chain (HC)1 and HC2 in normal cartilage and osteoarthritis (OA) cartilage and synovial fluids. Design/methods Cartilage extracts from normal donors and late-stage OA patients, and synovial fluids from OA patients were studied by Western blot with multiple antibodies to bikunin, HC1 and HC2. Cell and matrix localization was determined by immunohistochemistry and mRNA by RT-PCR. Results Bikunin·chondroitin sulfate (CS) and IαI were abundant in OA cartilages, but virtually undetectable in normal. In both OA and normal cartilages, HCs were largely present in a novel C-terminally truncated 50-kDa form, with most, if not all of these being attached to CS on a proteoglycan other than bikunin. Synovial fluids from OA patients contained bikunin·CS and full-length (∼90kDa) HCs linked to hyaluronan (HA) as HC·HA (SHAP·HA). Immunohistochemistry showed intracellular and cell-associated staining for bikunin and HCs, consistent with their synthesis by superficial zone chondrocytes. PCR on multiple human normal and OA cartilage samples detected transcripts for HC1 and HC2 but not for bikunin. In OA cartilages, immunostaining was predominantly matrix-associated, being most intense in regions with a pannus-like fibrotic overgrowth. Conclusion The truncated structure of HCs, their attachment to a proteoglycan other than bikunin, PCR data and intracellular staining are all consistent with synthesis of HC1 and HC2 by human articular chondrocytes. The presence of bikunin·CS and IαI in OA cartilage, but not in normal, appears to be due to diffusional uptake and retention through fibrillated (but not deeply fissured) cartilage surfaces.

  • two distinct populations of tumor necrosis factor stimulated gene 6 protein in the extracellular matrix of expanded mouse cumulus cell oocyte complexes
    Archives of Biochemistry and Biophysics, 2001
    Co-Authors: Durba Mukhopadhyay, Antonietta Salustri, Vincent C Hascall, Anthony J Day, Csaba Fulop
    Abstract:

    After the luteinizing hormone surge, the cumulus cell-oocyte complexes (COCs) in the preovulatory follicles produce a viscoelastic extracellular matrix, a process that requires the synthesis of hyaluronan as well as the incorporation of some components of the Inter-Alpha-Trypsin Inhibitor (IalphaI) family. In this study we report, that a hyaluronan-binding protein, the translated product of tumor necrosis factor-stimulated gene-6 (TSG-6), is also specifically accumulated in this matrix. TSG-6 mRNA expression is quickly upregulated and peaks at approximately 1500 copies/cell 4 h after the ovulatory stimuli as assessed by quantitative reverse transcription-polymerase chain reaction. Immunohistochemistry reveals the colocalization of the TSG-6 protein and hyaluronan around the cumulus and granulosa cells. The TSG-6 protein exists in two distinct populations in the COC matrix as demonstrated by Western-blot analysis. One population is a monomer that is anchored to the matrix by a noncovalent interaction. The second population is a covalent complex with either of the heavy chains of IalphaI and is bound to hyaluronan through a strong interaction that is resistant to denaturing conditions. The specific incorporation of the TSG-6 protein into the COC matrix suggests a structural role for this molecule.

Marylin S Rugg - One of the best experts on this subject based on the ideXlab platform.

  • superficial zone chondrocytes in normal and osteoarthritic human articular cartilages synthesize novel truncated forms of inter alpha trypsin Inhibitor heavy chains which are attached to a chondroitin sulfate proteoglycan other than bikunin
    Osteoarthritis and Cartilage, 2008
    Co-Authors: Y Yoshihara, Barbara Osborn, Arkady Margulis, F Nelson, Matthew C Stewart, Caroline M Milner, Marylin S Rugg, Anna Plaas, K Nemoto
    Abstract:

    OBJECTIVE: We have examined the occurrence of the inflammation-associated Inter-Alpha-Trypsin Inhibitor (IalphaI) components, bikunin, heavy chain (HC)1 and HC2 in normal cartilage and osteoarthritis (OA) cartilage and synovial fluids. DESIGN/METHODS: Cartilage extracts from normal donors and late-stage OA patients, and synovial fluids from OA patients were studied by Western blot with multiple antibodies to bikunin, HC1 and HC2. Cell and matrix localization was determined by immunohistochemistry and mRNA by RT-PCR. RESULTS: Bikunin.chondroitin sulfate (CS) and IalphaI were abundant in OA cartilages, but virtually undetectable in normal. In both OA and normal cartilages, HCs were largely present in a novel C-terminally truncated 50-kDa form, with most, if not all of these being attached to CS on a proteoglycan other than bikunin. Synovial fluids from OA patients contained bikunin.CS and full-length (approximately 90 kDa) HCs linked to hyaluronan (HA) as HC.HA (SHAP.HA). Immunohistochemistry showed intracellular and cell-associated staining for bikunin and HCs, consistent with their synthesis by superficial zone chondrocytes. PCR on multiple human normal and OA cartilage samples detected transcripts for HC1 and HC2 but not for bikunin. In OA cartilages, immunostaining was predominantly matrix-associated, being most intense in regions with a pannus-like fibrotic overgrowth. CONCLUSION: The truncated structure of HCs, their attachment to a proteoglycan other than bikunin, PCR data and intracellular staining are all consistent with synthesis of HC1 and HC2 by human articular chondrocytes. The presence of bikunin.CS and IalphaI in OA cartilage, but not in normal, appears to be due to diffusional uptake and retention through fibrillated (but not deeply fissured) cartilage surfaces.

  • superficial zone chondrocytes in normal and osteoarthritic human articular cartilages synthesize novel truncated forms of inter alpha trypsin Inhibitor heavy chains which are attached to a chondroitin sulfate proteoglycan other than bikunin
    Osteoarthritis and Cartilage, 2008
    Co-Authors: Y Yoshihara, Barbara Osborn, Arkady Margulis, F Nelson, Matthew C Stewart, Caroline M Milner, Marylin S Rugg, Anna Plaas, Anthony J Day
    Abstract:

    Summary Objective We have examined the occurrence of the inflammation-associated Inter-Alpha-Trypsin Inhibitor (IαI) components, bikunin, heavy chain (HC)1 and HC2 in normal cartilage and osteoarthritis (OA) cartilage and synovial fluids. Design/methods Cartilage extracts from normal donors and late-stage OA patients, and synovial fluids from OA patients were studied by Western blot with multiple antibodies to bikunin, HC1 and HC2. Cell and matrix localization was determined by immunohistochemistry and mRNA by RT-PCR. Results Bikunin·chondroitin sulfate (CS) and IαI were abundant in OA cartilages, but virtually undetectable in normal. In both OA and normal cartilages, HCs were largely present in a novel C-terminally truncated 50-kDa form, with most, if not all of these being attached to CS on a proteoglycan other than bikunin. Synovial fluids from OA patients contained bikunin·CS and full-length (∼90kDa) HCs linked to hyaluronan (HA) as HC·HA (SHAP·HA). Immunohistochemistry showed intracellular and cell-associated staining for bikunin and HCs, consistent with their synthesis by superficial zone chondrocytes. PCR on multiple human normal and OA cartilage samples detected transcripts for HC1 and HC2 but not for bikunin. In OA cartilages, immunostaining was predominantly matrix-associated, being most intense in regions with a pannus-like fibrotic overgrowth. Conclusion The truncated structure of HCs, their attachment to a proteoglycan other than bikunin, PCR data and intracellular staining are all consistent with synthesis of HC1 and HC2 by human articular chondrocytes. The presence of bikunin·CS and IαI in OA cartilage, but not in normal, appears to be due to diffusional uptake and retention through fibrillated (but not deeply fissured) cartilage surfaces.

  • impaired cumulus mucification and female sterility in tumor necrosis factor induced protein 6 deficient mice
    Development, 2003
    Co-Authors: Csaba Fulop, Sandor Szanto, Durba Mukhopadhyay, Tamas Bardos, Rajesh V Kamath, Marylin S Rugg, Antonietta Salustri, Vincent C Hascall, Tibor T Glant, Katalin Mikecz
    Abstract:

    Mucification of the cumulus layer around the oocyte is an obligatory process for female fertility. Tumor necrosis factor-induced protein-6 (TNFIP6 or TSG6) has been shown to be specifically expressed during this process. We have generated TNFIP6-deficient mice and tested the ability of their cumulus cells to undergo mucification. Cumulus cell-oocyte complexes fail to expand in TNFIP6-deficient female mice because of the inability of the cumulus cells to assemble their hyaluronan-rich extracellular matrix. The impaired cumulus matrix formation is due to the lack of covalent complexes between hyaluronan and the heavy chains of the inter-α-trypsin Inhibitor family. As a consequence, TNFIP6-deficient females are sterile. Cultured TNFIP6-deficient cumulus cell-oocyte complexes also fail to expand when stimulated with dibutyryl cyclic AMP or epidermal growth factor. Recombinant TNFIP6 is able to catalyze the covalent transfer of heavy chains to hyaluronan in a cell-free system, restore the expansion of Tnfip6 -null cumulus cell-oocyte complexes in vitro, and rescue the fertility in Tnfip6 -null females. These results provide clear evidence that TNFIP6 is a key catalyst in the formation of the cumulus extracellular matrix and indispensable for female fertility.

Csaba Fulop - One of the best experts on this subject based on the ideXlab platform.

  • impaired cumulus mucification and female sterility in tumor necrosis factor induced protein 6 deficient mice
    Development, 2003
    Co-Authors: Csaba Fulop, Sandor Szanto, Durba Mukhopadhyay, Tamas Bardos, Rajesh V Kamath, Marylin S Rugg, Antonietta Salustri, Vincent C Hascall, Tibor T Glant, Katalin Mikecz
    Abstract:

    Mucification of the cumulus layer around the oocyte is an obligatory process for female fertility. Tumor necrosis factor-induced protein-6 (TNFIP6 or TSG6) has been shown to be specifically expressed during this process. We have generated TNFIP6-deficient mice and tested the ability of their cumulus cells to undergo mucification. Cumulus cell-oocyte complexes fail to expand in TNFIP6-deficient female mice because of the inability of the cumulus cells to assemble their hyaluronan-rich extracellular matrix. The impaired cumulus matrix formation is due to the lack of covalent complexes between hyaluronan and the heavy chains of the inter-α-trypsin Inhibitor family. As a consequence, TNFIP6-deficient females are sterile. Cultured TNFIP6-deficient cumulus cell-oocyte complexes also fail to expand when stimulated with dibutyryl cyclic AMP or epidermal growth factor. Recombinant TNFIP6 is able to catalyze the covalent transfer of heavy chains to hyaluronan in a cell-free system, restore the expansion of Tnfip6 -null cumulus cell-oocyte complexes in vitro, and rescue the fertility in Tnfip6 -null females. These results provide clear evidence that TNFIP6 is a key catalyst in the formation of the cumulus extracellular matrix and indispensable for female fertility.

  • two distinct populations of tumor necrosis factor stimulated gene 6 protein in the extracellular matrix of expanded mouse cumulus cell oocyte complexes
    Archives of Biochemistry and Biophysics, 2001
    Co-Authors: Durba Mukhopadhyay, Antonietta Salustri, Vincent C Hascall, Anthony J Day, Csaba Fulop
    Abstract:

    After the luteinizing hormone surge, the cumulus cell-oocyte complexes (COCs) in the preovulatory follicles produce a viscoelastic extracellular matrix, a process that requires the synthesis of hyaluronan as well as the incorporation of some components of the Inter-Alpha-Trypsin Inhibitor (IalphaI) family. In this study we report, that a hyaluronan-binding protein, the translated product of tumor necrosis factor-stimulated gene-6 (TSG-6), is also specifically accumulated in this matrix. TSG-6 mRNA expression is quickly upregulated and peaks at approximately 1500 copies/cell 4 h after the ovulatory stimuli as assessed by quantitative reverse transcription-polymerase chain reaction. Immunohistochemistry reveals the colocalization of the TSG-6 protein and hyaluronan around the cumulus and granulosa cells. The TSG-6 protein exists in two distinct populations in the COC matrix as demonstrated by Western-blot analysis. One population is a monomer that is anchored to the matrix by a noncovalent interaction. The second population is a covalent complex with either of the heavy chains of IalphaI and is bound to hyaluronan through a strong interaction that is resistant to denaturing conditions. The specific incorporation of the TSG-6 protein into the COC matrix suggests a structural role for this molecule.