The Effect of Cadmium on Cytosolic Free Calcium, Protein Kinase C, and Collagen Synthesis in Rat Osteosarcoma (ROS 17/2.8) Cells

Abstract

Cadmium affects normal bone growth but the mechanisms of Cd2+toxicity are not fully understood. Calcium is an integral component of bone growth and a second messenger necessary for the actions of calciotropic hormones. Ca2+activates protein kinase C (PKC), and PKC is a mediator of [Ca2+]iand mediator of collagen synthesis in osteoblastic cells. Therefore, PKC is a possible loci of Cd2+effects on Ca2+metabolism and Ca2+-regulated processes. This work was conducted to determine the effect of Cd2+on cytosolic free Ca2+([Ca2+]i) levels, characterize the activation and/or inhibition of PKC by Cd2+and Ca2+, and measure the effect of Cd2+on collagen synthesis in ROS 17/2.8 cells. Cells were treated for 120 min with Cd2+(0 to 30 μm) and [Ca2+]iwas measured. Basal [Ca2+]iwas 132 nmand the maximal increase to 268 nmoccurred in the presence of 5 μmCd2+. Treatment with 1 or 5 μmCd2+caused an increase in [Ca2+]iat 40 min with return to basal levels at 120 min of treatment. Pretreatment (24 hr) with 0.1 μmcalphostin C (CC), a PKC inhibitor, produced no change in [Ca2+]iand prevented any rise in [Ca2+]iin response to Cd2+. Free Cd2+activates PKC with an activation constant of 7.5 × 10−11m, while Ca2+activates PKC with an activation constant of 3.6 × 10−7m. Cd2+also caused a dose-dependent decrease in collagen synthesis, a PKC-mediated process. These data suggest that Cd2+affects Ca2+metabolism and Ca2+-mediated processes via unwarranted PKC activation as demonstrated by Cd2+perturbation of collagen synthesis.