tailieunhanh - Stem Cells in Endocrinology - part 5

Đại diện thử nghiệm so sánh sản lượng từ các giao thức bảo quản lạnh. Phân tích số lượng và kích thước của HES3 (A) và HSF6 (B) thuộc địa sau khi bảo quản lạnh bằng hai phương pháp khác nhau. Tế bào gốc phôi đã được rã đông và nuôi cấy trong 1 tuần, sau đó qua khảo nghiệm cho số lượng và kích thước của các thuộc địa. * p | 102 Beattie et al. Fig. 6. Representative experiment of comparison of yields from cryopreservation protocols. Analysis of number and size of HES3 A and HSF6 B colonies after cryopreservation by two different methods. Embryonic stem cells were thawed and cultured for 1 week then assayed for number and size of colonies. p compared with treatment with dimethyl sulfoxide fetal bovine serum. the FBS Fig. 6A . However yield of HSF6 cells after thawing was always higher than that for hES3 and the recovery was similar using either trehalose or FBS Fig. 6B again demonstrating an underlying difference between the two cell lines. The reasons for this discrepancy are unknown but may relate to the differences in growth characteristics. hES3 colonies are much more compact with cells very close together see Fig. 5 and they survive better when transferred in discrete chunks rather than in loose cell clusters. Because hES3 remain in larger tighter clusters than HSF6 during the cryoprotection process it is necessary to get the cryoprotectant medium into the cells inside the clusters. The trehalose strategy used here allows the cells to become transiently porous by exposing them to a thermotropic lipid-phase transition with a high concentration of trehalose outside the cells during the cooling process enabling the cryoprotectant to enter the cells efficiently thus stabilizing the cell membranes from the inside. This method facilitates transfer of the cryoprotectant medium into the cells inside the clusters as well as on the periphery as we have described for human adult pancreatic islets and fetal islet-like cell clusters 36 . From the discussions here it is evident that there are no uniform culture conditions for ES cells and that different conditions may lead to different morphologies and phenotypes. It follows that not only the mechanism for selfrenewal may be different for each cell type but also the ability to differentiate into certain cell lineages. Chapter 5 Murine .