Author/Authors :
Zandi، Mohammad نويسنده Department of Animal and Poultry Science and Fisheries, Agricultural Institute, Iranian Research Organisation for Science and Technology (IROST), Tehran, Iran , , Muzaffar، Musharifa نويسنده Embryo Biotechnology Laboratory, Animal Biotechnology Centre, National Dairy Research Institute (NDRI), Karnal, India , , Shah، Syed Mohmad نويسنده Embryo Biotechnology Laboratory, Animal Biotechnology Centre, National Dairy Research Institute (NDRI), Karnal, India , , Singh، Manoj Kumar 1972- نويسنده , , Palta، Prabhat نويسنده Embryo Biotechnology Laboratory, Animal Biotechnology Centre, National Dairy Research Institute (NDRI), Karnal, India , , Singla، Suresh Kumar نويسنده Embryo Biotechnology Laboratory, Animal Biotechnology Centre, National Dairy Research Institute (NDRI), Karnal, India , , Manik، Radheysham نويسنده Embryo Biotechnology Laboratory, Animal Biotechnology Centre, National Dairy Research Institute (NDRI), Karnal, India , , Singh Chauhan، Manmohan نويسنده Embryo Biotechnology Laboratory, Animal Biotechnology Centre, National Dairy Research Institute (NDRI), Karnal, India ,
Abstract :
Objective: In order to retain an undifferentiated pluripotent state, embryonic stem (ES)
cells have to be cultured on feeder cell layers. However, use of feeder layers limits stem
cell research, since experimental data may result from a combined ES cell and feeder cell
response to various stimuli.
Materials and Methods: In this experimental study, a buffalo ES cell line was established
from in vitro derived blastocysts and characterized by the Alkaline phosphatase (AP) and
immunoflourescence staining of various pluripotency markers. We examined the effect of
various factors like fibroblast growth factor 2 (FGF-2), leukemia inhibitory factor (LIF) and
Y-27632 to support the growth and maintenance of bubaline ES cells on gelatin coated
dishes, in order to establish feeder free culture systems. We also analyzed the effect of
feeder-conditioned media on stem cell growth in gelatin based cultures both in the presence
as well as in the absence of the growth factors.
Results: The results showed that Y-27632, in the presence of FGF-2 and LIF, resulted
in higher colony growth and increased expression of Nanog gene. Feeder-Conditioned
Medium resulted in a significant increase in growth of buffalo ES cells on gelatin coated
plates, however, feeder layer based cultures produced better results than gelatin based
cultures. Feeder layers from buffalo fetal fibroblast cells can support buffalo ES cells for
more than two years.
Conclusion: We developed a feeder free culture system that can maintain buffalo ES
cells in the short term, as well as feeder layer based culture that can support the long
term maintenance of buffalo ES cells.
