Neuroprotective Effects Of Catalpol And Pre-B-Cell Colony Enhancing Factor

In this study, the protective effects and mechanisms of catalpol and PBEF on neurodegenerative disease were investigated with Parkinson's disease (PD) and Cerebral ischemia models in cortical astrocyte-enriched cultures, cortical or mesencephalic neuron-enriched cultures and mesencephalic neuron-astrocyte cultures, the primary coverage as follows:1. Neuroprotective Effects of CatalpolNeuroprotective effects of catalpol in astrocytes primary cultures. On one hand, we treated astrocytes with H2O2to induce oxidant stress. The results showed that catalpol could significantly increase the cell viability and reduce the intracellular ROS formation. Furthermore, catalpol attenuated H2O2-induced oxidative stress via preventing the decrease in the activities of antioxidant enzymes in glutathione redox cycling such as glutathione reductase (GR), glutathione peroxidase (GSH-Px), and glutathione content. However, the catalase (CAT) activity did not appear to be elevated by catalpol adequately. Together, the main mechanism underlying the protective effects of catalpol in H2O2-injured astrocytes might be related to the maintenance of glutathione metabolism balance and the decrease of ROS formation. On the other hand, astrocytes were pretreated with catalpol prior to lipopolysaccharide (LPS) plus interferon-γ (IFN-γ) stimulation which induced a inflammatory response. Biochemical analyses showed that nitric oxide (NO), ROS production and the inducible nitric oxide synthase (iNOS) activity were significantly reduced by catalpol. The data at transcriptional level also demonstrated that catalpol potently attenuated gene expressions which involved in inflammation, such as iNOS, cyclooxygenase-2(COX-2) and toll-like receptor4(TLR4). In addition, our exploration further revealed that the suppressive action of catalpol on inflammation was mediated via inhibiting nuclear factor-KB (NF-κB) activation. Collectively, these results adequately suggested that catalpol could exert inhibitory effects on inflammatory reaction in astrocytes and the inactivation of NF-κB could be the major determinant for its anti-inflammatory mechanism.Neuroprotective effects of catalpol in mesencephalic neurons. In current study, we stimulated primary mesencephalic neurons by rotenone to simulate PD occurring, and the results indicated that catalpol inhibited primary mesencephalic neurons from apoptosis by morphological assay, immunocytochemistry and flow cytometric evaluation. Moreover, the Extracellular regulated protein kinases (ERK) signaling pathway plays an important role in NO-mediated degeneration of neuron.Neuroprotective effects of catalpol in mesencephalic neuron-astrocyte cultures. MPTP is well known to produce clinical, biochemical and neurochemical changes similar to those which occur in PD. Furthermore, the accumulated evidence suggests that MPP+, conversed by monoamine oxidase type B (MAO-B) in astrocytes principally, is the active metabolite of MPTP and the major cause to PD associated with mitochondrial dysfunction. In this study, we treated mesencephalic neuron-astrocyte with MPTP respectively to investigate the neuroprotective effects of catalpol and the underlying protective mechanisms. Our results showed that pre-treatment with catalpol prior to MPTP treatment attenuated mitochondrial dysfunction by reversing the activity of mitochondrial complex I, mitochondrial membrane potential (MMP), intracellular Ca+level, and ROS accumulation as well as mitochondrial permeability transition pore (MPTp) opening.2. Neuroprotective Effects of PBEFPre-B-Cell Colony-Enhancing Factor (PBEF) is a rate-limiting enzyme to convert nicotinamide to nicotinamide mononucleotide (NMN) in the salvage pathway of nicotinamide adenine dinucleotide (NAD+) biosynthesis. Previously we found PBEF is exclusively expressed in neurons in mouse brain and heterozygous PBEF knockout (Pbef+/-) mice have larger ischemic lesion than wild type mice using photothrombosis-induced ischemia model. For mechanistic study of neuronal protective role of PBEF, we used in vitro oxygen-glucose deprivation (OGD) and glutamate excitotoxicity models of primary cultured neurons. Our results show that the treatments of neurons with nicotinamide and NAD+, the substrate and product of PBEF respectively, reduce neuronal death after OGD and glutamate excitotoxicity. Neurons with treatment of FK866, a PBEF inhibitor, have reduced cell viability and NAD+level after OGD as compared with neurons without treatment. Furthermore, overexpression of PBEF reduced glutamate excitotoxicity. We further tested whether PBEF affect mitochondrial function and biogenesis. Inhibition of PBEF reduces mitochondrial biogenesis, while addition of NAD+and NAM increase mitochondrial biogenesis. We further show overexpression of PBEF in neurons reduces MMP depolarization following glutamate stimulation using fluorescent live cell imaging. We conclude that PBEF exerts neuroprotection in ischemia through its enzymatic activity for NAD+production that can ameliorate mitochondrial dysfunction.