Inorganic phosphate (Pi) modulates the expression of key regulatory proteins of the inorganic pyrophosphate (PPi) metabolism in TGF-β1-stimulated chondrocytes

The balance between extracellular inorganic phosphate (ePi) and extracellular inorganic pyrophosphate (ePPi) is controlled by four membrane proteins: the transporters ANK (exporting PPi outside the cells) and PiT-1 (importing ePi into the cells), and the enzymes PC-1 (generating ePPi from nucleotides) and Tissue Non-specific Alkaline Phosphatase (TNAP, hydrolyzing ePPi into ePi). TGF-beta 1 was shown to stimulate ANK and PC-1 expression in articular chondrocytes, and subsequent ePPi level, as well as to increase ePi uptake by inducing PiT-1 expression in a chondrogenic cell line. Thus, we investigated the ability of ePi to modulate the effect of TGF-beta 1 on the regulatory proteins of the ePi/ePPi balance in chondrocytes. In the pathophysiological range of 0.01-1 mM, ePi was inactive by itself but potentiated the stimulatory effects of TGF-beta 1 on ANK, PC-1 or PiT-1 mRNA (RT-qPCR) and protein (Western blot) levels. PC-1 activity was also increased by TGF-beta 1 and further potentiated by ePi supplementation. TNAP mRNA and activity became undetectable in response to TGF-beta 1. These data suggest that ePi could increase ePPi level by changing the control of ANK and PC-1 expression by TGF-beta 1, further highlighting an adaptative regulation of the Pi/PPi balance to prevent basic calcium phosphate deposition into the joints.