A highly efficient recombinant canarypox virus-based vaccine against canine distemper virus constructed using the CRISPR/Cas9 gene editing method.

Canine distemper virus (CDV) is the causative agent of canine distemper (CD), which is one of the most important infectious diseases affecting wild and domestic carnivores. Vaccination represents an effective approach to prevent CDV infection among domestic carnivores. Canarypox-vectored recombinant CD vaccines (such as Recombitek CDV, PureVax Ferret Distemper, and Merial) with the CDV hemagglutinin (H) and fusion (F) genes can induce a potent immune response in dogs and ferrets. However, the vaccine's effectiveness varies with the species. In the current study, we developed a highly efficient recombinant canarypox virus termed as "ALVAC-CDV-M-F-H/C5-" that contained CDV virus-like particles (VLPs) by using the CRISPR/Cas9 gene editing method, which enabled concurrent expression of the matrix (M), H, and F genes. The recombinant strain provided faster seroconversion than the parent strain among minks as well as provided higher rates of antibody positivity than the parent strain among foxes and minks even before the administration of a second booster vaccination. We demonstrated, for the first time, that the CRISPR/Cas9 system can be applied for the rapid and efficient modification of the ALVAC-CDV-F-H genome and also that a high-dose new recombinant strain that produces CDV VLPs may present good outcomes in the prevention of CD among foxes and minks.

Paliperidone increases spontaneous and evoked firing of mesocortical dopaminergic neurons by activating a hyperpolarization-activated inward current.

Mesocortical dopaminergic (DA) subtype neurons specifically project to the prefrontal cortex, which is closely related with schizophrenia. Mesocortical DA neurons have unique physiological characteristics that are different from those of mesostriatal and mesolimbic DA neurons. Paliperidone, an atypical antipsychotic, is currently used to treat schizophrenia and has better therapeutic effects than typical antipsychotics. However, the underlying physiological mechanism remains unclear. To explore the effects of paliperidone on mesocortical DA neuron activity, here, we retrogradely labeled these cells with fluorescent microsphere retrobeads, and the electrophysiological changes were recorded in whole-cell recordings in rat midbrain slices with or without paliperidone. The data showed that paliperidone (20µmol/L) increased the spontaneous firing rates of labeled mesocortical neurons (P<0.05). Moreover, paliperidone also increased the frequency of evoked action potentials by current injection stimulation (P<0.05), whereas the accompanying amplitude decreased. Furthermore, to explore the mechanisms of paliperidone's effect, Ih currents were detected, and the results showed that hyperpolarizing voltage pulses evoked instantaneous Ih inward currents and paliperidone increased the maximum Ih current. In addition, paliperidone decreased the spontaneous inhibitory postsynaptic currents. Thus, paliperidone increased the spontaneous and evoked firing of mesocortical neurons, possibly by activating the Ih inward current and reducing the inhibitory synaptic transmission, which provides an underlying mechanism of paliperidone's application in schizophrenia.

Resveratrol inhibits ß-amyloid-induced neuronal apoptosis through regulation of SIRT1-ROCK1 signaling pathway.

Alzheimer's disease (AD) is characterized by the accumulation of ß-amyloid peptide (Aß) and loss of neurons. Recently, a growing body of evidences have indicated that as a herbal compound naturally derived from grapes, resveratrol modulates the pathophysiology of AD, however, with a largely unclear mechanism. Therefore, we aimed to investigate the protection of resveratrol against the neurotoxicity of ß-amyloid peptide 25-35 (Aß(25-35)) and further explore its underlying mechanism in the present study. PC12 cells were injuried by Aß(25-35), and resveratrol at different concentrations was added into the culture medium. We observed that resveratrol increased cell viability through the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) colorimetric assays. Flow cytometry indicated the reduction of cell apoptosis by resveratrol. Moreover, resveratrol also stabilized the intercellular Ca(2+) homeostasis and attenuated Aß(25-35) neurotoxicity. Additionally, Aß(25-35)-suppressed silent information regulator 1 (SIRT1) activity was significantly reversed by resveratrol, resulting in the downregulation of Rho-associated kinase 1 (ROCK1). Our results clearly revealed that resveratrol significantly protected PC12 cells and inhibited the ß-amyloid-induced cell apoptosis through the upregulation of SIRT1. Moreover, as a downstream signal molecule, ROCK1 was negatively regulated by SIRT1. Taken together, our study demonstrated that SIRT1-ROCK1 pathway played a critical role in the pathomechanism of AD.

Paliperidone protects prefrontal cortical neurons from damages caused by MK-801 via Akt1/GSK3ß signaling pathway.

Recent studies have suggested that neurodegeneration is involved in the pathogenesis of schizophrenia, and some atypical antipsychotics appear to prevent or retard progressive morphological brain changes. However, the underlying molecular mechanisms are largely unknown. Whether changes in intracellular signaling pathways are related to their neuroprotective effects remains undefined. In the present study, we used mouse embryonic prefrontal cortical neurons to examine the neuroprotection of paliperidone against the neuronal damage induced by exposure to the NMDA receptor antagonist, MK-801. Paliperidone inhibited MK-801 induced neurotoxicity both in MTT metabolism assay (p<0.01) and in lactate dehydrogenase (LDH) activity assay (p<0.01). Time course studies revealed that paliperidone effectively attenuated the elevation of intracellular free calcium concentration ([Ca(2+)]i) induced by exposure to MK-801 (p<0.01). Moreover, paliperidone could significantly retard MK-801-mediated inhibition of neurite outgrowth (p<0.01) and reverse MK-801-induced decreases of gene expression and phosphorylation of Akt1 and GSK3ß (both p<0.01). Furthermore, these protective effects of paliperidone were blocked by pretreatment with a PI3K inhibitor LY294002. Taking together, our results demonstrated that paliperidone could protect prefrontal cortical neurons from MK-801-induced damages via Akt1/GSK3ß signaling pathway.

Neuroprotective effects of Buyang Huanwu decoction against hydrogen peroxide induced oxidative injury in Schwann cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Buyang Huanwu decoction (BYHWD) is a traditional Chinese medicine and can be used to promote peripheral nerve regeneration. However the regenerative mechanism of BYHWD remains unclear. The objective of this study was to investigate the protective mechanisms of BYHWD in Schwann cells damaged by hydrogen peroxide (H(2)O(2)). MATERIALS AND METHODS: Schwann cells which were derived from neonatal sciatic nerves of rats were used in subsequent experiments. Schwann cells were injured by various concentrations of H(2)O(2) (0.25, 0.5 and 1mM final concentration). BYHWD (600 µg/ml final concentration) was added to the medium either simultaneously or 1h later after the addition of H(2)O(2). Subsequently, methyl thiazolyl tetrazolium (MTT) assay was performed. Superoxide dismutase (SOD) activity and malondialdehyde (MDA) levels were also examined after 12h. The expression of Caspase 3 and the concentration of intercellular Ca(2+) ([Ca(2+)]i) were also determined. RESULTS: Among three concentrations of H(2)O(2), 0.5mM H(2)O(2) induced Schwann cells swelled and neuritis disappeared after 12h. In the presence of BYHWD, MTT assay showed that more cells were viable in comparison with the H(2)O(2) injury group. Moreover, the addition of BYHWD has also increased the SOD activity with decreased in MDA level. Furthermore, the concentration of [Ca(2+)]i and expression of Caspase 3 were decreased with the addition of BYHWD in culture. CONCLUSIONS: Our results revealed that BYHWD protected Schwann cells from oxidative injury. The mechanism of BYHWD promoting neural regeneration possibly associated with its anti-oxidative activity.