Erceg S.,Cabimer Centro Andaluz Of Biologia Molecular Y Medicina Regenerativa |
Lukovic D.,Cabimer Centro Andaluz Of Biologia Molecular Y Medicina Regenerativa |
Moreno-Manzano V.,Neural Regeneration Laboratory |
Stojkovic M.,Spebo Medical |
And 2 more authors.
Current Protocols in Stem Cell Biology | Year: 2012
Here we provide a protocol for differentiation of human embryonic stem cells (hESC) into cerebellar neurons using a novel defined culture method. This protocol is based on the application of inductive signaling factors involved in the early patterning of the cerebellar region of the neural tube, followed by the application of factors responsible for cerebellar neuron specification. Human pluripotent stem cells are induced to form spherical embryonic-like structures called embryoid bodies (EBs) and neuroepithelial tube-like rosettes using defined chemical conditions. In the presence of FGF, Wnt, and RA signaling factors the rosettes were specified to OTX2-expressing cells. Further specification of derived cells involves application of BMP factors involved in early development of granule cell progenitors, followed by mitogens and neurotrophins. It typically takes 5 weeks to generate the functional cerebellar granule neurons. This protocol is feeder-free, applies human recombinant factors, and produces high yield of desired neurons. © 2012 by John Wiley & Sons, Inc. Source
Tsai M.-J.,Neural Regeneration Laboratory |
Tsai M.-J.,Taipei Veterans General Hospital |
Chen Y.-M.A.,National Yang Ming University |
Weng C.-F.,National Dong Hwa University |
And 22 more authors.
Annals of the New York Academy of Sciences | Year: 2010
Glycine N-methyltransferase (GNMT) is the most abundant hepatic methyltransferase and plays important roles in regulating methyl group metabolism. In the central nervous system, GNMT expression is low and its function has not been revealed. The present study examines the effect of GNMT overexpression by adenovirus-mediated transfer in cortical mixed neuron-glial cultures. Infection of adenovirus encoding green fluorescence protein to cultures demonstrates high preference for non-neuronal cells. Optimal GNMT overexpression in cultures by adenoviral GNMT (Ad-GNMT) infection not only induces protein kinase C phosphorylation, but also increases neuronaloligodendroglial survival. Furthermore, these Ad-GNMT-infected cultures are significantly resistant to H2O2 toxicity and lipopolysaccharide stimulation. Conditioned media from Ad-GNMT-infected microglia also significantly enhance neuronal survival. Taken together, enhanced GNMT expression in mixed neuronal-glial cultures is neuroprotective, most likely mediated through non-neuronal cells. © 2010 New York Academy of Sciences. Source
Bennett S.A.L.,Ottawa Institute of Systems Biology |
Bennett S.A.L.,Neural Regeneration Laboratory |
Bennett S.A.L.,University of Ottawa |
Valenzuela N.,Neural Regeneration Laboratory |
And 12 more authors.
Frontiers in Physiology | Year: 2013
Not all of the mysteries of life lie in our genetic code. Some can be found buried in our membranes. These shells of fat, sculpted in the central nervous system into the cellular (and subcellular) boundaries of neurons and glia, are themselves complex systems of information. The diversity of neural phospholipids, coupled with their chameleon-like capacity to transmute into bioactive molecules, provides a vast repertoire of immediate response second messengers. The effects of compositional changes on synaptic function have only begun to be appreciated. Here, we mined 29 neurolipidomic datasets for changes in neuronal membrane phospholipid metabolism in Alzheimer's Disease (AD). Three overarching metabolic disturbances were detected. We found that an increase in the hydrolysis of platelet activating factor precursors and ethanolamine-containing plasmalogens, coupled with a failure to regenerate relatively rare alkyl-acyl and alkenyl-acyl structural phospholipids, correlated with disease severity. Accumulation of specific bioactive metabolites [i.e., PC(O-16:0/2:0) and PE(P-16:0/0:0)] was associated with aggravating tau pathology, enhancing vesicular release, and signaling neuronal loss. Finally, depletion of PI(16:0/20:4), PI(16:0/22:6), and PI(18:0/22:6) was implicated in accelerating Aβ42 biogenesis. Our analysis further suggested that converging disruptions in platelet activating factor, plasmalogen, phosphoinositol, phosphoethanolamine (PE), and docosahexaenoic acid metabolism may contribute mechanistically to catastrophic vesicular depletion, impaired receptor trafficking, and morphological dendritic deformation. Together, this analysis supports an emerging hypothesis that aberrant phospholipid metabolism may be one of multiple critical determinants required for Alzheimer disease conversion. © 2013 Bennett, Valenzuela, Xu, Franko, Fai and Figeys. Source