miRNA expression profiling in mouse embryonic stem cell conditioned media

To identify novel putative factors that the anti-senescence effects of mouse embryonic stem cell conditioned media (E-CM) against celllular senescence, we conduvted a miRNA array analysis on E-CM and the mouse embryonic fibroblast (MEF)-CM The miRNA array was performed using the affymetrix GeneChip® miRNA 3.0 Array (Affymetrix, Santa Clara, California, United States). CM was collected from ESCs in OPTI-MEM on gelatin without feeder, FBS or other supplements. We have used CM to define ESC-specific secretory factors excluding potential factors from FBS and MEFs Gelatin coating was used to feeder free conditions, but some feeder cells may remain in culture. M-CM was used as a control to exclude remaining feeder cell effects. We collected conditioned media from MEF (mouse embryonic fibroblast) cells cultured in OPTI-MEM. We isolated the total RNA from these 3 samples and performed the miRNA array (Affymetrix platform) to identify the differentially expressed miRNAs in E-CM and MEF-CM