Expression of HIF prolyl hydroxylase isozymes in growth plate chondrocytes: Relationship between maturation and apoptotic sensitivity

The overall goal of the current study was to examine the functional activity of the prolyl hydroxylases (PHDs) in maturing chondrocytes. Herein, we show for the first time that the PHDs are expressed in the maturing zone of the growth plate, and by a chondrocytic cell line. We determined if this protein and its substrate, hypoxia inducible factor (HIF)-1 alpha, modulated the induction of apoptosis. Using a chondrocyte cell line that matured in culture, we inhibited HIF-1 alpha expression using siRNA technology and pharmacologically blocked PHD activity. We noted that PHD suppression sensitized the cells to an apoptotic challenge with H2O2. We next examined the interplay between the PHDs and HIF-1 alpha by suppressing HIF-1 alpha and blocking PHD activity. We noted reduced killing when the mature HIF-silenced cells were challenged with H2O2. In contrast, there was limited change in the viability of immature cells. Based on these differences in chondrocyte susceptibility, it is concluded that HIF-1 alpha sensitizes maturing cells to H2O2-mediated killing. We next determined if this change in the viability of the PHD-inhibited cells was linked to changes in activation of caspase-3. It was noted that there was a minimal change in enzyme activity of the PHD-inhibited HIF-1 alpha suppressed cells. Finally, we found that as the chondrocytes mature, the activities of catalase and SOD were significantly reduced and that there was a decrease in the levels of Bcl-2 and Bcl(XL). This loss of protective activity together with the changes mediated by HIF would be expected to generate conditions that would favor the induction of chondrocyte apoptosis.