John E. Donello

Differential Effects of PPARγ Ligands on Oxidative Stress–Induced Death of Retinal Pigmented Epithelial Cells

PURPOSEnTo investigate the role of the peroxisome proliferator-activated receptor (PPAR)-γ in modulating retinal pigmented epithelium (RPE) responses to oxidative stress.nnnMETHODSnARPE-19 cells were treated with the oxidant, t-butylhydroperoxide (tBH) to induce apoptosis. Cells pretreated with synthetic PPARγ agonists of the antidiabetic thiazolidinediones class before tBH challenge were assessed for viability and, by microarray analysis, for effects on gene expression.nnnRESULTSnTreatment of ARPE-19 cells with tBH resulted in a loss of viability and global changes in the pattern of gene expression. PPARγ ligands were found to have differential modulatory effects on tBH-induced apoptosis of RPE cells. Whereas rosiglitazone and pioglitazone potentiated cell death, troglitazone acted as a potent cytoprotective agent. Downregulation of PPARγ expression by an siRNA resulted in enhanced cell death in response to tBH treatment and blocked the cytoprotective effect of troglitazone consistent with a role of PPARγ in mediating this response. Microarray analysis revealed that while rosiglitazone and pioglitazone had little effect on gene changes induced by tBH treatment, troglitazone dramatically reduced the number of changes caused by oxidative stress. A unique subset of genes that were deregulated by tBH and selectively normalized by troglitazone were identified.nnnCONCLUSIONSnThese findings demonstrate that PPARγ agonists can have differential effects on RPE survival in response to oxidative stress. Oxidative stress leads to deregulation of a large set of genes in ARPE-19 cells. A specific subset of these genes can be selectively modulated by troglitazone and represent potential novel targets for cytoprotective therapies.

Function of brain α2B-adrenergic receptor characterized with subtype-selective α2B antagonist and KO mice

Noradrenergic signaling, through the α2A and α2C adrenergic receptors modulates the cognitive and behavioral symptoms of disorders such as schizophrenia, attention deficit hyperactivity disorder (ADHD), and addiction. However, it is unknown whether the α2B receptor has any significant role in CNS function. The present study elucidates the potential role of the α2B receptor in CNS function via the discovery and use of the first subtype-selective α2B antagonist (AGN-209419), and behavioral analyses of α-receptor knockout (KO) mice. Using AGN-209419 as radioligand, α2B receptor binding sites were identified within the olfactory bulb, cortex, thalamus, cerebellum, and striatum. Based on the observed expression patterns of α2 subtypes in the brain, we compared α2B KO, α2A KO and α2C KO mice behavioral phenotypes with their respective wild-type lines in anxiety (plus maze), compulsive (marble burying), and sensorimotor (prepulse inhibition) tasks. α2B KO mice exhibited increased marble burying and α2C KO mice exhibited an increased startle response to a pulse stimulus, but otherwise intact prepulse inhibition. To further explore compulsive behavior, we evaluated novelty-induced locomotor hyperactivity and found that α2B KO and α2C KO mice exhibited increased locomotion in the open field. Interestingly, when challenged with amphetamine, α2C KO mice increased activity at lower doses relative to either α2A KO or WT mice. However, α2B KO mice exhibited stereotypy at doses of amphetamine that were only locomotor stimulatory to all other genotypes. Following co-administration of AGN-209419 with low-dose amphetamine in WT mice, stereotypy was observed, mimicking the α2B KO phenotype. These findings suggest that the α2B receptor is involved in CNS behaviors associated with sensorimotor gating and compulsivity, and may be therapeutically relevant for disorders such as schizophrenia, ADHD, post-traumatic stress disorder, addiction, and obsessive compulsive disorder.