309 PROTEOMIC ANALYSIS OF THE BLASTOCOEL FLUID AND REMAINING CELLS OF BOVINE BLASTOCYSTS

Human embryonic stem cells (hESC) are derived from the human blastocyst and possess the potential to differentiate into any cell type present in the adult human body. Human ESC are considered to have great potential in regenerative medicine for the future treatment of severe diseases and conditions such as Parkinson’s disease, diabetes, and spinal cord injury. One of today’s challenges in regenerative medicine is to define proper culture conditions for hESC. The natural milieu in the blastocyst may provide clues on how to improve culture conditions, and the aim of the present study was to determine the proteome of the blastocoel fluid and the remaining cells of bovine blastocysts. Bovine blastocysts were produced by in vitro fertilization of oocytes retrieved from slaughterhouse ovaries. The blastocoel from 195 blastocysts (1–8 nL per blastocyst) were isolated by micromanipulation and analysed by nano-HPLC tandem mass spectrometry along with the remaining cells of the blastocyst. Searching the mass spectrometry data against a combined bovine database (SwissProt/TrEMBL), we identified 263 proteins in the blastocoel fluid and 1606 proteins in the cellular compartment of the blastocyst. A Venn diagram showed 124 proteins in overlap between the two compartments of the blastocyst. Several heat shock proteins and specific antioxidants were identified in both the blastocoel and cell material. A selection of proteins identified in the blastocoel fluid is to be tested on hESC in cell culture experiments, with proliferation of undifferentiated cells as the primary endpoint. The results from this study provide new knowledge about early mammalian preimplantation development, and the data can be used in the continued pursue of improving culture conditions for hESC, which further facilitates the clinical application of these cells.