Altering Two Major Redox Enzymes Affected Expression and Function of Murine REG Family Proteins

Abstract

Abstract Objectives We previously revealed a substantially-suppressed expression of regenerating islets-derived protein 2 (Reg2) in murine pancreatic islets by overexpression of glutathione peroxidase-1 (Gpx1-OE). Two experiments were conducted to determine: 1) how knockouts of Gpx1 and superoxide dismutase-1 (Sod1) alone and together (dKO) affected the expression profiles of the full Reg family genes in three tissues; and 2) how GPX1 and SOD1 activities regulated effects of REG2 on islet proliferation. Methods In Expt. 1, six groups of mice (Gpx1−/−, Gpx1-OE, and their wild-type, WT1; Sod1−/−, dKO, and their WT2) (male, 2–4 months old (n = 6 – 8) were fed an Se-adequate diet and killed to collect islets, liver, and intestine samples. In Expt. 2, islets isolated from the 6 groups of mice were cultured in RPMI 1640 medium and treated with recombinant REG2 and REG2 mutant proteins (1 μg/mL), GPX mimic (ebselen, 50 μM) and SOD mimic (CuDIP, 10 μM) for 48 h. The proliferation of islets was estimated using the bromodeoxyuridine (Sigma) assay. Results Compared with the WT1 mice, the Gpx1−/− mice had greater (P < 0.05) and the Gpx1-OE mice had lower (P < 0.05) mRNA levels of all 7 assayed Reg genes with a few exceptions of Reg3ẞ,Reg3δ, or Reg4 in the three tissues. Similar inductions of the 7 Reg mRNA levels were also seen in the islets of Sod1−/− and dKO mice over their controls. However, responses of these 7 genes in the liver and intestine of these two genotypes in comparison with the controls were less consistent. The incubation of islets with REG2, but not the REG2 mutant, inhibited islet proliferation in the Gpx1−/−, Sod1−/−, dKO, and Gpx1-OE groups, where no such effect was seen in the two WT groups. Co-incubations of islets with CuDIP and ebselen partially blocked the REG2 inhibitory effect on the islet proliferation. Conclusions Altering GPX1 and SOD1 expression and activity affected the Reg family gene expression in the islets, liver, and intestine of mice as well as the function of REG2 protein in the islets. Our results reveal a novel dependence of the REG family protein expression and function on cellular redox status. Funding Sources This research was supported in an NIH grant DK 53,018.