In vitro studies in neuroblastoma cell lines demonstrate co-regulation of PSA/NPEPPS and SOD1. A. Western blot analysis shows decrease of endogenous SOD1 in response to PSA/NPEPPS overexpression (PSA↑) and accumulation of SOD1 in response to PSA/NPEPPS inhibition (PSA↓) with Stealth-RNAi in human SH-SY5Y neuroblastoma cell line. B. Representative immunocytochemistry demonstrates a notable decrease (top) and a rapid accumulation (bottom) of SOD1 protein (green) in response to PSA/NPEPPS overexpression (PSA↑; top; red) and PSA/NPEPPS inhibition (PSA↓; bottom), respectively, in SH-SY5Y cells. The middle image represents normal control. PSA/NPEPPS is red, SOD1 is green, and the nucleus is blue as stained with DAPI nuclear marker. Scale bar 20 μm. C. PSA/NPEPPS expression demonstrated by Western blot is up-regulated in response to SOD1 overexpression in mouse N1E-115 neuroblastoma cell line. Because mouse SOD1 overexpression construct was used, we utilized mouse neuroblastoma cell line to avoid possible side effects of heterologous expression of a mouse protein in a human cell line. For each comparison, three independent experiments were performed (n = 3) with at least three independent replicates in each experiment. Representative Western blot and immunocytochemistry images are shown. Error bars represent standard deviations.