The effect of cadmium on cytosolic free calcium, protein kinase C, and collagen synthesis in rat osteosarcoma (ROS 17/2.8) cells.

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+]1 and mediator of collagen synthesis in osteoblastic cells. Therefore, PKC is a possible loci of Cd2+ effects on Ca2+ metabolism and Ca(2+)-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 microM) and [Ca2+]i was measured. Basal [Ca2+]i was 132 nM and the maximal increase to 268 nM occurred in the presence of 5 microM Cd2+. Treatment with 1 or 5 microM Cd2+ caused an increase in [Ca2+]i at 40 min with return to basal levels at 120 min of treatment. Pretreatment (24 hr) with 0.1 microM calphostin C (CC), a PKC inhibitor, produced no change in [Ca2+]i and prevented any rise in [Ca2+]i in response to Cd2+. Free Cd2+ activates PKC with an activation constant of 7.5 X 10(-11) M, while Ca2+ activates PKC with an activation constant of 3.6 X 10(-7) M. Cd2+ also caused a dose-dependent decrease in collagen synthesis, a PKC-mediated process. These data suggest that Cd2+ affects Ca2+ metabolism and Ca(2+)-mediated processes via unwarranted PKC activation as demonstrated by Cd2+ perturbation of collagen synthesis.