Redox-regulation of intrinsic prion expression in multicellular prostate tumor spheroids

Abstract

The cellular function of the intrinsic prion protein (PrP c) remains largely unknown. In the present study PrP c expression was investigated in multicellular prostate tumor spheroids and was correlated to the intracellular redox state as evaluated using the fluorescent dye 2′7′-dichlorodihydrofluorescein diacetate (H 2DCFDA). In small tumor spheroids (diameter 100 ± 20 μm) reactive oxygen species (ROS) levels were increased as compared with large (diameter 250 ± 50 μm) spheroids. ROS generation was mediated by the mitochondrial respiratory chain and a NADPH oxidaselike enzyme, because carbonylcyanide- m-chlorophenylhydrazone (CCCP), rotenone, and diphenylene iodonium chloride (DPI) significantly reduced ROS levels. The elevated ROS were correlated to an increased expression of PrP c, Cu/Zn superoxide dismutase (SOD-1), and catalase in small as compared with large spheroids. In large tumor spheroids, PrP c was predominantly expressed in the peripheral cell layers and colocalized with SOD-1 and catalase. Raising intracellular ROS in large tumor spheroids by hydrogen peroxide, menadione, buthionine sulfoximine (BSO), and incubation in glutamine-reduced medium increased PrP c expression. In small spheroids PrP c was downregulated after incubation with the radical scavengers dehydroascorbate (DHA) and vitamin E. Our data indicate that PrP c expression in tumor spheroids is related to the intracellular redox state and may participate in antioxidative defense.