Abstract: Objective Establish a culture method and a differentiation method for the cynomolgus monkey ES cells, to prepare for future experiments of transplantating neural progenitors derived from cynomolgus monkey ES cells into monkey disease model. Methods Prepare mouse embryonic fibroblasts (MEF) as feeder cells, culture the cynomolgus monkey ES cells for a long period, then characterize the cells by marker staining. The cynomolgus ES cells were differentiated into neural progenitors by adding noggin into differentiation medium, and the cells were stained in each stage during differentiation by immno-histochemistry. Results The cynomolgus monkey ES cells form colonies, and express cell markers of undifferentiated primate ES cells for more than 20 passage doublings. In neural differentiaion assay, the ES cells formed rosette and neural tube after 14 days neural induction. The staining showed large numbers of Nestin positive and Tuj-1 positive neural progenitors. If induction period extended to 21 days, the number of Nestin positive and Tuj-1 positive cells was still numerous. After 35 days of differentiation, many GFAP positive cells started to emerge, and Tuj-1 positive cells decreased. Conclusions The culture system of cynomolgus monkey ES cells was successfully established. The induction of ES cells toward neural lineages produced a large number of neural progenitors, and non-mature neurons expressing Tuj-1 marker.

Key words: cynomolgus monkey, embryonic stem cells, cell culture, neural differentiation