Abstract | OBJECTIVE: 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.
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Authors | K Johnson, S Vaingankar, Y Chen, A Moffa, M B Goldring, K Sano, P Jin-Hua, A Sali, J Goding, R Terkeltaub |
Journal | Arthritis and rheumatism
(Arthritis Rheum)
Vol. 42
Issue 9
Pg. 1986-97
(Sep 1999)
ISSN: 0004-3591 [Print] United States |
PMID | 10513816
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, Non-P.H.S., Research Support, U.S. Gov't, P.H.S.)
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Chemical References |
- Diphosphates
- Membrane Glycoproteins
- Protein Synthesis Inhibitors
- Brefeldin A
- Phosphoric Diester Hydrolases
- Enpp3 protein, rat
- ectonucleotide pyrophosphatase phosphodiesterase 1
- Pyrophosphatases
- nucleoside triphosphate pyrophosphatase
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Topics |
- Brefeldin A
(pharmacology)
- Cartilage, Articular
(cytology)
- Cell Line
- Chondrocytes
(chemistry)
- Diphosphates
(metabolism)
- Dose-Response Relationship, Drug
- Humans
- Membrane Glycoproteins
(biosynthesis, pharmacology)
- Phosphoric Diester Hydrolases
- Protein Synthesis Inhibitors
(pharmacology)
- Pyrophosphatases
(metabolism, pharmacology)
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