Protein tyrosine kinase inhibitors block the stimulatory actions of phosphotyrosine phosphatase inhibitors to increase cell proliferation, alkaline phosphatase activity, and collagen synthesis in normal human bone cells

The present study sought to test whether inhibition of phosphotyrosine phos phatases (PTPs) would stimulate proliferation and differentiation of normal bone cells, and whether the PTP inhibitor-mediated effects would be blocke d by protein tyrosine kinase (PTK) inhibitors. Three inhibitors [phenylarsi ne oxide (PAO), orthovanadate (VO4), and molybdate (MoO4)] and two normal h uman bone cells with different basal differentiation status (i.e., mandible - and vertebra-derived bone cells) were used. Cell proliferation was determ ined with [H-3]thymidine incorporation, and confirmed by cell counting. Bon e cell differentiation was assessed by increases in alkaline phosphatase (A LP) specific activity and collagen synthesis. The th ree test PTP inhibitor s each stimulated [H-3]thymidine incorporation in both human bone cell type s in a biphasic, dose-dependent manner with optimal doses of 20 nM PAO, 1 m u M VO4 and 2 mu M MoO4, respectively. These PTP inhibitors at mitogenic do ses each significantly and reproducibly increased ALP specific activity and collagen synthesis. To determine whether the stimulatory effects of PTP in hibitors could be blocked by PTK inhibitors, the effects of tyrphostin A51 and erbstatin, two potent PTK inhibitors, on the actions of PTK inhibitors on [H-3]thymidine incorporation and ALP specific activity were evaluated. B oth tyrphostin A51 and erbstatin, which by themselves alone significantly i nhibited human bone cell proliferation and increased ALP specific activity, completely abolished the stimulatory effects of each of the th ree test PT P inhibitors on bone cell proliferation and ALP specific activity. In concl usion, these findings confirm the premise that inhibition of PTP activities in normal human bone cells could lead to increases in cell proliferation a nd differentiation, effects that are independent of basal differentiation s tatus of the cells. More importantly, this study demonstrates for the first time that the stimulatory actions of the PTP inhibitor on bone cell prolif eration and ALP could be blocked by a PTK inhibitor, suggesting that the os teogenic effects of PTP inhibitors may depend on PTK activities, presumably to increase basal tyrosyl phosphorylation level. Accordingly, one should i nterpret results of studies using PTK inhibitors with caution in that an in hibition by a PTK inhibitor does not necessarily indicate the requirement o f PTK activities, as it could also suggest involvement of an inhibition of PTPs. Copyright (C) 2000 S. Karger AG. Basel.