Sequential combined Treatment of Pifithrin-α and Posiphen Enhances Neurogenesis and Functional Recovery After Stroke.

OBJECTIVE: Although cerebral ischemia can activate endogenous reparative processes, such as proliferation of endogenous neural stem cells (NSCs) in the subventricular zone (SVZ) and subgranular zone (SGZ), the majority of these new cells die shortly after injury and do not appropriately differentiate into neurons, or migrate and functionally integrate into the brain. The purpose of this study was to examine a novel strategy for treatment of stroke after injury by optimizing the survival of ischemia-induced endogenous NSCs in the SVZ and SGZ. METHODS: Adult SVZ and SGZ NSCs were grown as neurospheres in culture and treated with a p53 inactivator, pifithrin-α (PFT-α), and an amyloid precursor protein (APP)-lowering drug, posiphen, and effects on neurosphere number, size and neuronal differentiation were evaluated. This combined sequential treatment approach was then evaluated in mice challenged with middle cerebral artery occlusion (MCAo). Locomotor behavior and cognition were evaluated at 4 weeks, and the number of new surviving neurons was quantified in nestin creERT2-YFP mice. RESULTS: PFT-α and posiphen enhanced the self-renewal, proliferation rate and neuronal differentiation of adult SVZ and SGZ NSCs in culture. Their sequential combination in mice challenged with MCAo-induced stroke mitigated locomotor and cognitive impairments and increased the survival of SVZ and SGZ NSCs cells. PFT-α and the combined posiphen+PFT-α treatment similarly improved locomotion behavior in stroke challenged mice. Notably, however, the combined treatment provided significantly more potent cognitive function enhancement in stroke mice, as compared with PFT-α single treatment. INTERPRETATION: Delayed combined sequential treatment with an inhibitor of p53 dependent apoptosis (PFT-α) and APP synthesis (posiphen) proved able to enhance stroke-induced endogenous neurogenesis and improve the functional recovery in stroke animals. Whereas the combined sequential treatment provided no further improvement in locomotor function, as compared with PFT-α alone treatment, suggesting a potential ceiling in the locomotion behavioral outcome in stroke animals, combined treatment more potently augmented cognitive function recovery after stroke.

Identification of 2-piperidone as a biomarker of CYP2E1 activity through metabolomic phenotyping.

Cytochrome P450 2E1 (CYP2E1) is a key enzyme in the metabolic activation of many low molecular weight toxicants and also an important contributor to oxidative stress. A noninvasive method to monitor CYP2E1 activity in vivo would be of great value for studying the role of CYP2E1 in chemical-induced toxicities and stress-related diseases. In this study, a mass spectrometry-based metabolomic approach was used to identify a metabolite biomarker of CYP2E1 through comparing the urine metabolomes of wild-type (WT), Cyp2e1-null, and CYP2E1-humanized mice. Metabolomic analysis with multivariate models of urine metabolites revealed a clear separation of Cyp2e1-null mice from WT and CYP2E1-humanized mice in the multivariate models of urine metabolomes. Subsequently, 2-piperidone was identified as a urinary metabolite that inversely correlated to the CYP2E1 activity in the three mouse lines. Backcrossing of WT and Cyp2e1-null mice, together with targeted analysis of 2-piperidone in mouse serum, confirmed the genotype dependency of 2-piperidone. The accumulation of 2-piperidone in the Cyp2e1-null mice was mainly caused by the changes in the biosynthesis and degradation of 2-piperidone because compared with the WT mice, the conversion of cadaverine to 2-piperidone was higher, whereas the metabolism of 2-piperidone to 6-hydroxy-2-piperidone was lower in the Cyp2e1-null mice. Overall, untargeted metabolomic analysis identified a correlation between 2-piperidone concentrations in urine and the expression and activity of CYP2E1, thus providing a noninvasive metabolite biomarker that can be potentially used in to monitor CYP2E1 activity.