Superoxide dismutase (SOD) is an enzyme that converts superoxide radicals into hydrogen peroxide and molecular oxygen and has been shown to contribute to the virulence of many human-pathogenic bacteria through its ability to neutralize toxic levels of reactive oxygen species generated by the host. SOD has also been speculated to be important in the pathogenesis of fungal infections, but the role of this enzyme has not been rigorously investigated. To examine the contribution of SOD to the pathogenesis of fungal infections, we cloned the Cu,Zn SOD-encoding gene (SOD1) from the human-pathogenic yeast Cryptococcus neoformans and made mutants via targeted disruption. The sod1 mutant strains had marked decreases in SOD activity and were strikingly more susceptible to reactive oxygen species in vitro. A sod1 mutant was significantly less virulent than the wild-type strain and two independent reconstituted strains, as measured by cumulative survival in the mouse inhalational model. In vitro studies established that the sod1 strain had attenuated growth compared to the growth of the wild type and a reconstituted strain inside macrophages producing reduced amounts of nitric oxide. These findings demonstrate that (i) the Cu,Zn SOD contributes to virulence but is not required for pathogenicity in C. neoformans; (ii) the decreased virulence of the sod1 strain may be due to increased susceptibility to oxygen radicals within macrophages; and (iii) other antioxidant defense systems in C. neoformans can compensate for the loss of the Cu,Zn SOD in vivo.Invasive fungal infections in humans are increasing in prevalence in parallel with the growing population of immunocompromised patients. There is a need for new antifungal drugs to treat these infections since the drugs currently available are either excessively toxic or lack broad fungicidal properties. Studies on the pathogenesis of fungal infections should provide insights that can help with the diagnosis and treatment of these important human diseases. Cryptococcus neoformans is a basidiomycetous yeast that has been used successfully as a model pathogenic fungus in a variety of molecular pathogenesis studies. We used C. neoformans to evaluate the contribution of superoxide dismutase (SOD) to the pathogenesis of fungal infections.SODs are metalloenzymes that detoxify oxygen radicals through the conversion of superoxide to hydrogen peroxide and oxygen (20). These enzymes are present in virtually all cells, and this very high degree of conservation is testament to their importance in cellular homeostasis. The primary role of SODs is to protect cells from endogenously generated superoxide anion, which is a by-product of normal aerobic respiration. SODs can be complexed with iron, manganese, and copper plus zinc. The iron and manganese SODs are genetically similar to each other, whereas the Cu,Zn SOD exhibits no significant homology with the other two enzymes (16,20,21,32). Eukaryotic cells generally contain an Mn SOD in the mitochondrial matrix and a Cu,Zn SOD which is located pre...

OBJECTIVE-Liver-specific inactivation of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) by a dominant-negative transgene (L-SACC1 mice) impaired insulin clearance, caused insulin resistance, and increased hepatic lipogenesis. To discern whether this phenotype reflects a physiological function of CEACAM1 rather than the effect of the dominant-negative transgene, we characterized the metabolic phenotype of mice with null mutation of the Ceacam1 gene (Cc1RESEARCH DESIGN AND METHODS-Mice were originally generated on a mixed C57BL/6x129sv genetic background and then backcrossed 12 times onto the C57BL/6 background. More than 70 male mice of each of the Cc1 Ϫ/Ϫ and wild-type Cc1 ϩ/ϩ groups were subjected to metabolic analyses, including insulin tolerance, hyperinsulinemic-euglycemic clamp studies, insulin secretion in response to glucose, and determination of fasting serum insulin, C-peptide, triglyceride, and free fatty acid levels. RESULTS-Like L-SACC1, Cc1Ϫ/Ϫ mice exhibited impairment of insulin clearance and hyperinsulinemia, which caused insulin resistance beginning at 2 months of age, when the mutation was maintained on a mixed C57BL/6x129sv background, but not until 5-6 months of age on a homogeneous inbred C57BL/6 genetic background. Hyperinsulinemic-euglycemic clamp studies revealed that the inbred Cc1 Ϫ/Ϫ mice developed insulin resistance primarily in liver. Despite substantial expression of CEACAM1 in pancreatic ␤-cells, insulin secretion in response to glucose in vivo and in isolated islets was normal in Cc1 Ϫ/Ϫ mice (inbred and outbred strains). CONCLUSIONS-Intact insulin secretion in response to glucose and impairment of insulin clearance in L-SACC1 and Cc1Ϫ/Ϫ mice suggest that the principal role of CEACAM1 in insulin action is to mediate insulin clearance in liver.

Glucocorticoid hormones control diverse physiological processes, including metabolism and immunity, by activating the major glucocorticoid receptor (GR) isoform, GRalpha. However, humans express an alternative isoform, human (h)GRbeta, that acts as an inhibitor of hGRalpha to produce a state of glucocorticoid resistance. Indeed, evidence exists that hGRbeta contributes to many diseases and resistance to glucocorticoid hormone therapy. However, rigorous testing of the GRbeta contribution has not been possible, because rodents, especially mice, are not thought to express the beta-isoform. Here, we report expression of GRbeta mRNA and protein in the mouse. The mGRbeta isoform arises from a distinct alternative splicing mechanism utilizing intron 8, rather than exon 9 as in humans. The splicing event produces a form of beta that is similar in structure and functionality to hGRbeta. Mouse (m)GRbeta has a degenerate C-terminal region that is the same size as hGRbeta. Using a variety of newly developed tools, such as a mGRbeta-specific antibody and constructs for overexpression and short hairpin RNA knockdown, we demonstrate that mGRbeta cannot bind dexamethasone agonist, is inhibitory of mGRalpha, and is up-regulated by inflammatory signals. These properties are the same as reported for hGRbeta. Additionally, novel data is presented that mGRbeta is involved in metabolism. When murine tissue culture cells are treated with insulin, no effect on mGRalpha expression was observed, but GRbeta was elevated. In mice subjected to fasting-refeeding, a large increase of GRbeta was seen in the liver, whereas mGRalpha was unchanged. This work uncovers the much-needed rodent model of GRbeta for investigations of physiology and disease.

No abstract