Purifications of small molecules that regulate neuronal cell-lineage specification

There are certain important factors that are essential for specification, development and maintenance vertebrate nervous system. During development these factors regulate axonal growth, dendrite pruning, cell fate decision and different other neuronal functions. In adult nervous system, they modulate synaptic activity and regulate neuronal connectivity. Cultivation of mammalian cells for production of recombinant protein has wide scientific and clinical applications because it offers many advantages especially the posttranslational modification. Human embryonic kidney (HEK) 293 cell lines expressing gene of interests will be constructed using calcium phosphate transfection of expression vectors. I have constructed stable cell lines for expressing large quantity of stable neurotrophic and neurological fate determinant factors including: Brain derived neurotrophic factor (BDNF),Glial derived neurotrophic factor (GDNF), Neurotrophin 3(NT3), Noggin and Sonic the hedgehog (SHH).This recombinant proteins were successfully purified by affinity chromatography and are tested on the conversion of induced pluripotent stem (iPS) cells into specific type of neurons, we found that some of the purified proteins are capable of support neuronal differentiation. These neurons have complex morphology, mature synaptic marker such as synapsin, and exhibit mature membrane properties and synaptic activity. We conclude that mammalian cell produced recombinant factors can be used to regulate human neuronal maturation and cell type specification.M.S.Includes bibliographical referencesby Aula Hamo