Downregulation of Caspase-2 Expression in Somitic Cells following Coculture with Chicken Notochord

Abstract

Somites are spherical aggregations of mesodermal cells located on either sides of neural tube and are differentiated into sclerotome and dermomyotome. Notochord as an axial mesoderm has a major role in somitic cell survival and differentiation in vivo. Despite secreting the survival factors, how to notochord inhibits somitic cells apoptosis remains to be elusive. So, this study was aimed to investigate downregulation of caspase-2 expression in somitic cells upon coculturing with notochord. By using alginate system to encapsulate the isolated notochord in Somite + Notochord group, the embryonic somites were cocultured with the notochord on different days. Concurrently in somite group, the somites were cultured alone. Survival assay with MTT showed that the rate of viability in somitic cells cocultured with notochord increased from 59% on day 2 to 89.7% on day 6 but decreased to 38.5% on day 10 after coculturing. Reverse transcriptase-polymerase chain reaction and spectrophotometry analysis also confirmed these findings and showed low caspase-2 and high Bcl-2 expressions and low caspase-2 enzyme activity in somitic cells cocultured with notochord, respectively. These results clearly show that the notochord enhances survival of somitic cells in vitro through downregulating of caspase-2 expression along with triggering differentiation of somitic cells to Pax-1 expressing mesenchymal cells.