Probenecid inhibits transforming growth factor-beta 1 induced pyrophosphate elaboration by chondrocytes.
OBJECTIVE
The elaboration of excess extracellular inorganic pyrophosphate (ePPi) by cartilage contributes to calcium pyrophosphate dihydrate (CPPD) crystal deposition disease. Transforming growth factor-beta 1 (TGF beta 1) is the only defined physiologic stimulant of cartilage ePPi elaboration. The mechanism of ePPi generation by chondrocytes is unknown, but current evidence suggests that TGF beta 1 induced ePPi is made intracellularly. An active transport mechanism such as an anion transporter would then be necessary to export ePPi to the matrix where crystals form. We determined the effect of probenecid (PB), an anion transport inhibitor, on TGF beta 1 induced ePPi elaboration.


METHODS
Porcine hyaline articular chondrocytes in high density monolayer cultures were exposed to serum-free media with and without TGF beta 1 and/or PB. ePPi was measured in the media after 48-96 h of exposure. Cell injury was measured by examining the release of 3H-deoxyglucose from chondrocytes. The activity of the ePPi generating ectoenzyme nucleoside triphosphate pyrophosphohydrolase (NTPPPH) and media lactate concentrations were measured with standard colorimetric assays. As PB may inhibit phosphodiesterase (PDE), its effects on ePPi generation were compared with isobutylmethylxanthine (IBMX), a specific PDE inhibitor.


RESULTS
PB inhibited TGF beta 1 induced ePPi elaboration by chondrocytes. PB did not cause membrane injury or decrease NTPPPH activity. Lactate production was decreased by PB but did not correlate with the effects of PB on ePPi elaboration. IBMX did not inhibit TGF beta 1 effect on ePPi elaboration.


CONCLUSIONS
PB blocks TGF beta 1 induced ePPi elaboration. This effect is independent of cell membrane injury, decreased NTPPPH activity, or PDE inhibition. Our data implicate a role for anion transport in TGF beta 1 induced ePPi elaboration, and suggest a potential therapy for CPPD disease.
Rosenthal
A K
AK
Department of Medicine, Medical College of Wisconsin, Milwaukee.
Ryan
L M
LM
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000818:Animals; D002358:Cartilage, Articular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D002805:Chondrocalcinosis / Q000628:therapy; D011756:Diphosphates / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D007719:Knee Joint; D007773:Lactates / Q000378:metabolism; D019344:Lactic Acid; D011339:Probenecid / Q000494:pharmacology*; D011755:Pyrophosphatases / Q000187:drug effects / Q000378:metabolism; D013552:Swine; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors*
D011756:Diphosphates; D007773:Lactates; D016212:Transforming Growth Factor beta; D019344:Lactic Acid; D011755:Pyrophosphatases; C021895:nucleoside triphosphate pyrophosphatase; D011339:Probenecid; D015056:1-Methyl-3-isobutylxanthine