MODULATION OF THE PLASMA MEMBRANE Ca2+ -ATPase IN CELL LINES OF RAT THYROID

Previous studies performed in cell lines mimicking the thyroid thumorogenic processes have demonstrated that the functional expression of the plasma-membrane Ca2+-ATPase (PMCA) is down regulated following neoplastic transformation. In the present study possible modifications in the regulation of PMCA, potentially involved in the disturbance of the thyroid cell calcium homeostasis, were investigated. In particular the effects of different extracellular signalling molecules as TSH, insulin, ATP and nitric oxide (NO) on PMCA activity were evaluated both in normal (PC Cl3) and transformed (PC E1APy, PC E1ARaf) rat thyroid cells. The specific activity of PMCA was measured by a spectrophotometric method that monitors the appearance of Pi in the presence or in the absence of the eosin, a PMCA inhibitor. Normal and transformed rat thyroid cells lines, previously incubated with increasing TSH concentrations, showed a dose and time dependent activation of the PMCA activity, although TSH stimulation is more effective in normal PC Cl3 than in transformed cell lines PC E1APy. Similar effects, also if at a lesser extent, were also obtained by using insulin. The incubation with ATP evoked a rapid and significant increase of PMCA activity in normal PC Cl3 and in transformed cells PC E1Apy; on the other hand, ATP didn’t produce any effect on PC E1Araf cell lines. The presence of UTP induced an activation of PMCA comparable to that elicited by ATP. The activator effects on PMCA activity induced by ATP/UTP were blocked by staurosporin, a potent inhibitor of protein kinases. Furthermore the incubation of cell lysate with sodium nitroprusside (SNP) a NO donor, was able to produce a significant increase of PMCA activity in normal PC Cl3. These results suggest that the regulatory pathways of cell calcium homeostasis involving the functional expression of plasma membrane Ca2+-ATPase, can be maintained during the neoplastic transformation of thyroid.