We developed a method for the efficient generation of functional dopaminergic (DA) neurons from human embryonic stem cells (hESCs) on a large scale. The most unique feature of this method is the generation of homogeneous spherical neural masses (SNMs) from the hESC-derived neural precursors. These SNMs provide several advantages: (i) they can be passaged for a long time without losing their differentiation capability into DA neurons; (ii) they can be coaxed into DA neurons at much higher efficiency than that from previous reports (86% tyrosine hydroxylase-positive neurons/total neurons); (iii) the induction of DA neurons from SNMs only takes 14 days; and (iv) no feeder cells are required during differentiation. These advantages allowed us to obtain a large number of DA neurons within a short time period and minimized potential contamination of unwanted cells or pathogens coming from the feeder layer. The highly efficient differentiation may not only enhance the efficacy of the cell therapy but also reduce the potential tumor formation from the undifferentiated residual hESCs. In line with this effect, we have never observed any tumor formation from the transplanted animals used in our study. When grafted into a parkinsonian rat model, the hESC-derived DA neurons elicited clear behavioral recovery in three behavioral tests. In summary, our study paves the way for the large-scale generation of purer and functional DA neurons for future clinical applications.is a neurodegenerative disorder characterized by progressive and selective loss of dopaminergic (DA) neurons in the midbrain substantia nigra (1). Currently, the prevailing strategy for the treatment of PD is pharmacological. However, pharmacological treatment with L-DOPA works initially, but over time, the effectiveness of L-DOPA wanes and side effects develop (2). An alternative approach may be the transplantation of DA-synthesizing cells. One source of DA-synthesizing cells is embryonic stem cells (ESCs). ESCs are pluripotent and capable of self-renewal (3-5). For the purpose of applying the ESCs to PD, many researchers have tried to develop protocols by which ESCs from some species can differentiate into DA neuronal phenotypes (6-11). Although some progress has been made in the generation of DA neurons from human ESCs (hESCs) (12-22), there are still many technical improvements to be made before the application of hESCs to treat PD. Examples include increasing the purity of DA neurons, supplying a sufficient quantity of DA neurons for clinical applications, decreasing tumor formation after transplantation, and clearly demonstrating the functionality of hESC-derived DA neurons in a parkinsonian animal model.Here, we introduce a method that allows us to differentiate hESCs into functional tyrosine hydroxylase-positive (TH ϩ ) neurons up to near 86% of the total hESC-derived neurons, which is the highest purity ever reported. Achieving high efficiency of DA neuronal derivation is an important issue in cell therapy, because it would not only increase the effica...

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Extensive evidence has indicated that a high rate of cholesterol biogenesis and abnormal neuronal energy metabolism play key roles in Alzheimer's disease (AD) pathogenesis. Here, for we believe the first time, we used osmotin, a plant protein homolog of mammalian adiponectin, to determine its therapeutic efficacy in different AD models. Our results reveal that osmotin treatment modulated adiponectin receptor 1 (AdipoR1), significantly induced AMP-activated protein kinase (AMPK)/Sirtuin 1 (SIRT1) activation and reduced SREBP2 (sterol regulatory element-binding protein 2) expression in both in vitro and in vivo AD models and in Adipo−/− mice. Via the AdipoR1/AMPK/SIRT1/SREBP2 signaling pathway, osmotin significantly diminished amyloidogenic Aβ production, abundance and aggregation, accompanied by improved pre- and post-synaptic dysfunction, cognitive impairment, memory deficits and, most importantly, reversed the suppression of long-term potentiation in AD mice. Interestingly, AdipoR1, AMPK and SIRT1 silencing not only abolished osmotin capability but also further enhanced AD pathology by increasing SREBP2, amyloid precursor protein (APP) and β-secretase (BACE1) expression and the levels of toxic Aβ production. However, the opposite was true for SREBP2 when silenced using small interfering RNA in APPswe/ind-transfected SH-SY5Y cells. Similarly, osmotin treatment also enhanced the non-amyloidogenic pathway by activating the α-secretase gene that is, ADAM10, in an AMPK/SIRT1-dependent manner. These results suggest that osmotin or osmotin-based therapeutic agents might be potential candidates for AD treatment.

Summary Experience-dependent plasticity in the adult brain has clinical potential for functional rehabilitation following central and peripheral nerve injuries. Here, plasticity induced by unilateral infraorbital (IO) nerve resection in four week-old rats was mapped using MRI and synaptic mechanisms were elucidated by slice electrophysiology. Functional MRI demonstrates a cortical potentiation compared to thalamus two weeks after IO nerve resection. Tracing thalamocortical (TC) projections with manganese-enhanced MRI revealed circuit changes in the spared layer 4 (L4) barrel cortex. Brain slice electrophysiology revealed TC input strengthening onto L4 stellate cells due to an increase in postsynaptic strength and the number of functional synapses. This work shows that the TC input is a site for robust plasticity after the end of the previously defined critical period for this input. Thus, TC inputs may represent a major site for adult plasticity in contrast to the consensus that adult plasticity mainly occurs at cortico-cortical connections.