Bakhshalizadeh, S. H., Esmaeili, F., Shirzad, H., Houshmand, F., Ebrahimi-Hafshejani, M., and Ghasemi, S.
Journal of Shahrekord University of Medical Sciences. Dec2014/Jan2015, Vol. 16 Issue 5, p32-43. 12p.
Background and aims: P19 cells are a line of pluripotent embryonal carcinoma stem cells, that are able to grow continuously in serum-supplemented media and can be induced to differentiate into all three lineages, mesoderm, endoderm and ectoderm. The sequence of foreign DNA can be introduced into these cells using of standard methods of transfection, through which to examine the cell function and differentiation. This study aimed to induce neuronal phenotype in P19 cells transfected with green fluorescent protein (GFP) gene, using an antiparkinson drug, deprenyl. Methods: In this laboratory descriptive study, calcium phosphate precipitation method was used to introduce pML8 plasmid, encoding GFP and puromycin resistance gene, into these cells. The cells were cultured using MEM-A medium, supplemented with 15% fetal bovine serum. Here 108 M concentrations of deprenyl was used to induce embryoid body differentiation to neuronal lineage. Cresyl violet staining was used to evaluate the morphology of differentiated P19 cells. In addition, immunofluorescence techniques was used to evaluate neuron specific proteins, synaptophysin and β-III tubulin. Results: In addition to production of stable transfected P19 cells, neuronal differentiation of these cells was carried out by the effect of deprenyl induction. GFP-positive neurons derived from embryonic carcinoma cells expressed neuron specific markers. Conclusions: GFP-positive differentiated cells can be used in transplantation studies and basic researches in association with cell therapy and neurodegenerative diseases. [ABSTRACT FROM AUTHOR]