The oxidoreductase, biliverdin reductase, is induced in human renal carcinoma - pH and cofactor-specific increase in activity

Purpose: Biliverdin reductase is an oxidoreductase unique among all enzymes characterized to date in having dual pH/dual cofactor requirement - NADH a nd NADPH at 6.7 and 8.7, respectively. The protein shows extensive microhet erogeneity that is caused by post-translational modification. The reductase converts the heme degradation product, biliverdin, to bilirubin. Bilirubin has been shown to inhibit responses of human lymphocytes, including phytoh emagglutinin-induced proliferation, interleukin-2 production, and antibody dependent and independent cell mediated cytotoxicity. In addition to acting as an antioxidant, it inhibits protein phosphorylation and activity of enz ymes such as protein kinase C and NADPH oxidase. This research was to evalu ate whether renal cell carcinoma differs from normal tissue in regard to th e expression and activity of the reductase. Materials and Methods: kidney tissue with or without visible renal carcinom a and normal kidney tissue from a brain dead patient were frozen at -80C sh ortly after removal. Ten mu m. tissue sections were used for immunostaining of biliverdin reductase, pooled isolated tumors and surrounding tissue tha t did not contain visible tumor were used for Northern blot analysis of mRN A and Western blot analysis of protein. Enzyme activity was also measured i n these preparations at pH 6.7 with NADH, and at pH 8.7 with NADPH. Ten add itional formalin fixed specimens of renal cell carcinoma were also used for immunostaining. Results: There was a striking increase in the reductase protein levels, as visualized by immunostaining in tumor tissue cells. The increase was also e vident by Western blotting, and involved in increased transcription of bili verdin reductase as suggested by Northern blot analysis. The protein would also be detected in the infiltrating monocytes, macrophages, T cells and ne utrophils as well as in circulating lymphocytes. The enzyme activity was ne arly doubled in the tumor tissue, but selectively with NADH as the cofactor . Conclusion: Increases in biliverdin reductase expression and activity only with NADH is found in renal cell carcinoma. The net effects of this change are uncertain at present but several pathways, which could be affected by t he reductase, may alter local physiology. Biliverdin reductase as a zinc me talloprotein may directly interact with other regulatory proteins, generati on of increased bilirubin may alter immune function and increased enzyme ac tivity may deplete NADH with contrasting consequence of blocking free radic al formation and depleting cellular ATP. To the benefit of the host, the la tter could culminate in tumor cell death.