Biomedical subjects

J Goding

Publications and source records attributed to J Goding.

10 recordsLinked to original sources

PubMed2001

Up-regulated expression of the phosphodiesterase nucleotide pyrophosphatase family member PC-1 is a marker and pathogenic factor for knee meniscal cartilage matrix calcification.

OBJECTIVE: Elevated cartilage inorganic pyrophosphate (PPi) production and PPi-generating nucleoside triphosphate pyrophosphohydrolase (NTPPPH) activity are strongly linked with aging-related cartilage calcification in meniscal and articular cartilages. We hypothesized that there were divergent relationships of 3 NTPPPH isozymes with cartilage matrix calcification and sought to identify them. METHODS: We studied knee medial meniscal expression in situ of 3 NTPPPH isozymes of the phosphodiesterase nucleotide pyrophosphatase (PDNP) family: plasma cell membrane glycoprotein 1 (PC-1, or PDNP1), autotaxin (ATX, or PDNP2), and B10/PDNP3. We also used complementary DNA transfection to assess differential functions in matrix calcification of each NTPPPH isozyme in vitro in meniscal cells. RESULTS: We observed diffuse cell-associated ATX and B10/PDNP3 expression in central (chondrocytic) and, to a lesser degree, peripheral (fibroblastic) regions of normal, degenerative uncalcified, and degenerative calcified menisci. In contrast, PC-1 expression was only robust at sites of apoptotic cells and calcification in central regions of degenerative menisci. Only PC-1 was abundant at the perimeter of meniscal cells and in association with meniscal cell-derived matrix vesicles (MVs). Because each PDNP-family isozyme was expressed by cells near calcifications, we transfected the isozymes in nonadherent knee meniscal cells cultured with ascorbic acid, beta-glycerophosphate, and dexamethasone supplementation to stimulate them to calcify the matrix. PC-1, but not ATX or B10/PDNP3, consistently promoted increased MV NTPPPH, MV-associated PPi, and extracellular PPi. PC-1 also increased matrix calcification (with hydroxyapatite crystals) by meniscal cells. ATX uniquely induced alkaline phosphatase activity, but promoted only moderately increased matrix calcification. CONCLUSION: We identified divergent effects of 3 PDNP-family NTPPPH isozymes on meniscal cell matrix calcification. Increased expression of PC-1 is both a marker and a potential pathogenic factor for knee meniscal cartilage matrix calcification.

Aging↗

PubMed2001

The nucleoside triphosphate pyrophosphohydrolase isozyme PC-1 directly promotes cartilage calcification through chondrocyte apoptosis and increased calcium precipitation by mineralizing vesicles.

OBJECTIVE: Aging associated elevations of cartilage extracellular inorganic pyrophosphate (PPi) and PPi-generating nucleoside triphosphate pyrophosphohydrolase (NTPPPH) are linked with degenerative arthritis in chondrocalcinosis. Increased chondrocyte apoptosis and expression of annexin V occur at sites of matrix calcification in degenerative arthritis, and membrane limited chondrocyte apoptotic bodies containing NTPPPH may promote chondrocalcinosis by acting as mineralizing matrix vesicles (MV). Because chondrocytes express 3 related NTPPPH isozymes [PC-1, autotaxin (ATX), and B10/PDNP3], we evaluated the effects on apoptosis and MV mediated calcium precipitation of direct expression of NTPPPH isozymes. METHODS: To achieve "gain of function" of NTPPPH isozymes, we expressed the isozymes in cultured chondrocytic cells. RESULTS: Plasmid cDNA transfection of PC-1, but not ATX or B10/PDNP3, markedly increased apoptosis of cultured chondrocytic knee meniscal cells and increased calcium precipitation by MV fractions. The capacity of PC-1 to increase chondrocyte and meniscal cell apoptosis, and calcium precipitation by MV, further analyzed using adenoviral gene transfer in cultured meniscal cells and articular chondrocytes, was shown to be dependent on integrity of the PC-I NTPPPH catalytic site. The MV-containing fraction released from meniscal cells and chondrocytes that overexpressed wild-type PC-1 had increased annexin V. Use of antibodies to annexin V and PC-1 revealed that both annexin V and PC-1 directly mediated the elevated calcium-precipitating capacity of MV. The increased ability of MV to precipitate calcium from PC-1-overexpressing cells did not require exogenous ATP. CONCLUSION: Upregulated expression of enzymatically active PC-1 directly promotes apoptosis, increased MV annexin V, and an increased capacity of meniscal cell and articular chondrocyte MV to precipitate calcium. These results suggest a direct link between increased PC-1 expression and the pathogenesis of chondrocalcinosis.

Apoptosis↗

PubMed1999

Differential mechanisms of inorganic pyrophosphate production by plasma cell membrane glycoprotein-1 and B10 in chondrocytes.

OBJECTIVE: Increased nucleoside triphosphate pyrophosphohydrolase (NTPPPH) activity in chondrocytes is associated with cartilage matrix inorganic pyrophosphate (PPi) supersaturation in chondrocalcinosis. This study compared the roles of the transforming growth factor beta (TGFbeta)-inducible plasma cell membrane glycoprotein-1 (PC-1) and the closely related B10 NTPPPH activities in chondrocyte PPi metabolism. METHODS: NTPPPH expression was studied using reverse transcriptase-polymerase chain reaction and Western blotting. Transmembrane PC-1 (tmPC-1), water-soluble secretory PC-1 (secPC-1), and transmembrane B10 were expressed by adenoviral gene transfer or plasmid transfection, and expression of PPi was assessed in cultured articular chondrocytes and immortalized NTPPPH-deficient costal chondrocytes (TC28 cells). RESULTS: PC-1 and B10 messenger RNA were demonstrated in articular cartilages in situ, in untreated cultured normal articular chondrocytes, and in TC28 cells. Expression of tmPC-1 and secPC-1, but not B10, rendered the NTPPPH-deficient TC28 cells able to increase expression of extracellular PPi, with or without addition of TGFbeta (10 ng/ml) to the media. More plasma membrane NTPPPH activity was detected in cells transfected with tmPC-1 than in cells transfected with B10. Furthermore, confocal microscopy with immunofluorescent staining of articular chondrocytes confirmed preferential plasma membrane localization of PC-1, relative to B10. Finally, both PC-1 and B10 increased the levels of intracellular PPi, but PC-1 and B10 appeared to act principally in different intracellular compartments (Golgi and post-Golgi versus pre-Golgi, respectively). CONCLUSION: PC-1 and B10 NTPPPH activities were not redundant in chondrocytes. Although increased PC-1 and B10 expression caused elevations in intracellular PPi, the major effects of PC-1 and B10 were exerted in distinct subcellular compartments. Moreover, PC-1 (transmembrane and secreted), but not B10, increased the levels of extracellular PPi. Differential expression of PC-1 and B10 could modulate cartilage mineralization in degenerative joint diseases.

Brefeldin A↗

PubMed1999

Matrix vesicle plasma cell membrane glycoprotein-1 regulates mineralization by murine osteoblastic MC3T3 cells.

A naturally occurring nonsense truncation mutation of the inorganic pyrophosphate (PPi)-generating nucleoside triphosphate pyrophosphohydrolase (NTPPPH) PC-1 is associated with spinal and periarticular ligament hyperostosis and cartilage calcification in "tiptoe walking" (ttw) mice. Thus, we tested the hypothesis that PC-1 acts directly in the extracellular matrix to restrain mineralization. Cultured osteoblastic MC3T3 cells expressed PC-1 mRNA and produced hydroxyapatite deposits at 12-14 days. NTPPPH activity increased steadily over 14 days. Transforming growth factor-beta and 1,25-dihydroxyvitamin D3 increased PC-1 and NTPPPH in matrix vesicles (MVs). Because PC-1/NTPPPH was regulated in mineralizing MC3T3 cells, we stably transfected or infected cells with recombinant adenovirus, in order to express 2- to 6-fold more PC-1. PC-1/NTPPPH and PPi content increased severalfold in MVs derived from cells transfected with PC-1. Furthermore, MC3T3 cells transfected with PC-1 deposited approximately 80-90% less hydroxyapatite (by weight) than cells transfected with empty plasmid or enzymatically inactive PC-1. ATP-dependent 45Ca precipitation by MVs from cells overexpressing active PC-1 was comparably diminished. Thus, regulation of PC-1 controls the PPi content and function of osteoblast-derived MVs and matrix hydroxyapatite deposition. PC-1 may provide a novel therapeutic target in certain disorders of bone mineralization.

Adenoviridae↗

PubMed1996

Coordinated regulation in human T cells of nucleotide-hydrolyzing ecto-enzymatic activities, including CD38 and PC-1. Possible role in the recycling of nicotinamide adenine dinucleotide metabolites.

The human leukocyte surface Ag CD38 was recently identified as a nicotinamide adenine dinucleotide (NAD)(+)-glycohydrolase ecto-enzyme, degrading NAD into nicotinamide and ADP-ribose. We show here that expression of CD38 is increased in the Jurkat T cell line after treatment with agents that augment intracellular cAMP, with the permeant cAMP analogue dibutyryl-cAMP (db-cAMP), and also with PMA, which activates protein kinase C. Treatment of human PBL T cells with db-cAMP or submitogenic doses of PMA also increased CD38 expression. Two other nucleotide-hydrolyzing activities were induced on the T cell surface concomitantly with CD38: the human PC-1 molecule, a nucleotide phosphodiesterase/pyrophosphatase that produces AMP from NAD or ADP-ribose, and a nucleotidase that produces adenosine from AMP, but which may be distinct from the CD73 5'-nucleotidase. All three enzymes were up-regulated after stimulation of human peripheral blood T cells with PHA. The coordinated regulation of these ecto-enzymes suggested that, besides a possible signaling function, they may recycle extracellular NAD by degrading it to adenosine and nicotinamide, which can be taken up by cells. In support of this hypothesis, db-cAMP-treated Jurkat cells could degrade extracellular NAD for de novo synthesis of purines, while untreated cells could not. Activated lymphocytes are often located in tissues in which cell death is common. It is suggested that the coordinated expression of these enzymes may allow activated T cells to re-use NAD and nucleotides from dead cells.

ADP-ribosyl Cyclase↗

PubMed1995

Interleukin 1 beta suppresses transforming growth factor-induced inorganic pyrophosphate (PPi) production and expression of the PPi-generating enzyme PC-1 in human chondrocytes.

Articular cartilage chondrocytes have the unique ability to elaborate large amounts of extracellular pyrophosphate (PPi), and transforming growth factor beta (TGF beta) appears singular among cartilage regulatory factors in stimulating PPi production. TGF beta caused a time and dose-dependent increase in intracellular and extracellular PPi in human articular chondrocyte cultures. TGF beta and interleukin 1 beta (IL-1 beta) antagonistically regulate certain chondrocyte functions. IL-1 beta profoundly inhibited basal and TGF beta-induced PPi elaboration. To address mechanisms involved with the regulation of PPi synthesis by IL-1 beta and TGF beta, we analyzed the activity of the PPi-generating enzyme NTP pyrophosphohydrolase (NTPPPH) and the PPi-hydrolyzing enzyme alkaline phosphatase. Human chondrocyte NTPPPH activity was largely attributable to plasma cell membrane glycoprotein 1, PC-1. Furthermore, TGF beta induced comparable increases in the activity of extracellular PPi, intracellular PPi, and cellular NTPPPH and in the levels of PC-1 protein and mRNA in chondrocytes as well as a decrease in alkaline phosphatase. All of these TGF beta-induced responses were completely blocked by IL-1 beta. Thus, IL-1 beta may be an important regulator of mineralization in chondrocytes by inhibiting TGF beta-induced PPi production and PC-1 expression.

Adult↗

PubMed1994

Causal link between nucleotide pyrophosphohydrolase overactivity and increased intracellular inorganic pyrophosphate generation demonstrated by transfection of cultured fibroblasts and osteoblasts with plasma cell membrane glycoprotein-1. Relevance to calcium pyrophosphate dihydrate deposition disease.

OBJECTIVE: In subjects with idiopathic calcium pyrophosphate dihydrate (CPPD) deposition disease, cartilage chondrocytes elaborate increased amounts of PPi. The mechanism of the intracellular PPi elevation is not known. Plasma membrane 5'-nucleotide phosphodiesterase I/nucleotide pyrophosphohydrolase (NTPPPH) activity also is elevated in chondrocytes and dermal fibroblasts of patients with idiopathic CPPD deposition disease. NTPPPH, as an ecto-enzyme, could act within certain intracellular compartments. Thus, we hypothesized a potential causal link between increased NTPPPH activity and increased intracellular PPi. METHODS: Transformed simian fibroblasts (COS cells) and human osteoblasts (U2OS cells) were transfected with the 5'-nucleotide phosphodiesterase I ecto-enzyme plasma cell membrane glycoprotein-1 (PC-1), recently shown to be expressed in cartilage, osteoblasts, and fibroblasts. RESULTS: Transfection with PC-1 markedly up-regulated 5'-nucleotode phosphodiesterase I activity and increased intracellular PPi concentrations by increasing the capacity of cells to generate PPi. Importantly, this did not require supplementation with exogenous nucleotides. CONCLUSION: Cellular overexpression of PC-1 produces NTPPPH overactivity and increased intracellular PPi generation in vitro. These findings support the potential importance of NTPPPH overactivity in PPi generation, both inside and outside the cell, in some subjects with CPPD deposition disease.

Animals↗

PubMed1994

Expression of the murine plasma cell nucleotide pyrophosphohydrolase PC-1 is shared by human liver, bone, and cartilage cells. Regulation of PC-1 expression in osteosarcoma cells by transforming growth factor-beta.

A bone and cartilage enzyme with both 5'-nucleotide phosphodiesterase I and nucleotide pyrophosphohydrolase (NTPPPH) activity modulates physiologic mineralization and pathologic chondrocalcinosis by generating inorganic pyrophosphate. We hypothesized that, as for alkaline phosphatase, expression of an NTPPPH gene can be shared by cells from bone, cartilage, and liver and by certain leukocytes. Recently, we demonstrated the hepatocyte and murine plasma cell membrane glycoprotein PC-1 to have both 5'-nucleotide phosphodiesterase I and NTPPPH activity. We detected polypeptides cross-reactive with PC-1 in human U20S osteosarcoma cells, articular chondrocytes, homogenized human knee cartilages, human knee synovial fluids, hepatoma cells, and murine plasmacytoma cells. Constitutive low abundance PC-1 mRNA expression was detected in U20S cells and chondrocytes by a nested RNA-PCR assay and by Northern blotting. TGF beta is known to substantially increase NTPPPH activity in primary osteoblast cultures. We demonstrated that TGF beta 1 increased NTPPPH activity and the level of PC-1 mRNA and immunoprecipitable [35S]-methionine-labeled PC-1 polypeptides in U20S cells. The identification of PC-1 as an NTPPPH expressed in cells derived from bone and cartilage may prove useful in furthering the understanding of the role of NTPPPH i n physiologic and pathologic mineralization.

Animals↗

PubMed1980

Ir gene testing in H-2 mutants.

Four different C57BL/6 mutants, each with a mutation presumably in the H-2Kb locus, were studied to determine whether the mutations had affected Ir gene function. When tested with the antigens, (TG)-A-L, ovalbumin (OA), and lactic dehydrogenase (LDHB), there was no difference in the antibody response of the mutants when compared to the parental C57BL/6 strain and there was also no difference in the response to the histocompatibility H-Y antigen. The responses to these antigens have been mapped, wholly or partially in the K-IA, IA or IB subregions and therefore the identical response of parent and mutants may provisionally exclude H-2Kb as a site of Ir gene function for the four antigens tested. In addition, the BALB/c-dm2 mutant, a mutation in the H-2Ld locus, gave identical responses to the parental BALB/cKh strain.

Animals↗

PubMedDate not supplied

Membrane proteins stimulated by interferon-alpha in growth-inhibited, but not in resistant, Daudi cells.

Interferon-alpha (alpha-IFN) stimulates the expression of a number of plasma membrane proteins in wild-type Daudi cells. These proteins are not induced in a variant subline of Daudi cells, which is insensitive to growth inhibition, nor in another which is partially growth-inhibited by alpha-IFN. Two-dimensional polyacrylamide gel electrophoresis of [125I]-surface labelled Daudi membrane proteins indicates that the major protein induced de novo has a molecular weight of 16 kD (p16), and proteins of 20 kD (p20), 28 kD (p28) and 106 kD (p106) which are present in untreated cells are also induced by alpha-IFN. A monoclonal antibody was produced against p106 by immunizing mice with cells treated for 16 h with alpha-IFN followed by differential screening against treated and untreated cells. The antibody (A96/G8) immunoprecipitated p106 from cell lysates and was used to show that p106 is induced approximately two-fold by alpha-IFN in wild-type Daudi cells, but not at all in Daudi cells which show a complete or partial resistance to growth inhibition by alpha-IFN; gamma-IFN does not induce p16 or p106 and also does not inhibit cell growth. The induction of p16 and p106 precedes the observed inhibition of growth by IFN. The levels of p16 and p106 are elevated by 8.5 h and in the case of p16 continue to rise for 16 h after the addition of IFN, whereas p106 reaches a peak by 8.5 h. In contrast, growth inhibition is not observed by 8 h and does not become clear-cut until 24 h after the addition of IFN.

Animals↗