Importance of culture conditions during the morula-to-blastocyst period on capacity of inner cell-mass cells of bovine blastocysts for establishment of self-renewing pluripotent cells.

The hypothesis was tested that the pluripotency of the inner cell mass (ICM) of the bovine embryo is enhanced by the glycogen synthase kinase-3ß inhibitor CHIR99021 and the MAPK1 and MAPK3 inhibitor PD032591. Treatment with the two inhibitors from Days 6 to 8 after insemination increased blastocyst steady state concentrations of mRNA for NANOG (P < 0.05) and SOX2 (P = 0.055) and tended to decrease (P = 0.09) expression of GATA6. To evaluate pluripotency, the inner cell mass was isolated by immunosurgery at Day 8, seeded on a feeder layer of bovine embryonic fibroblasts, and cultured in the presence of the inhibitors. Ten of 52 (19%) ICM from control embryos had primary outgrowth formation vs. 23 of 50 (46%) of the ICM from embryos cultured with inhibitors (P < 0.01). For ICM outgrowths from embryos cultured without inhibitors, colonies either did not persist through Passage 2 or became differentiated. In contrast, for the inhibitor group, four colonies survived beyond Passage 2, and one line persisted for 19 passages. This cell line possessed alkaline phosphatase activity, expressed several genes characteristically expressed in pluripotent cells, and differentiated into embryoid bodies when cultured in the absence of the signal transduction inhibitors and the feeder layer. Propagation of the cells was difficult due to slow growth and inefficiency in survival through each passage. In conclusion, exposure to inhibitors during the morula-blastocyst transition facilitated formation of self-renewing pluripotent cell lines from bovine blastocysts.

Alternative birth rooms and birth options.

The hospital-based alternative birth room has become a popular alternative to the conventional labor and delivery room setting. Responses to a mailed questionnaire from 78 of 82 (95%) Washington State hospitals with obstetric services were used to relate available birth options to alternative birth room status and obstetric volume. Alternative birth rooms were in existence, or planned, in 63% of responding hospitals. The presence of an alternative birth room was directly related to obstetric volume (P less than .05) and perceived local incidence of home deliveries (P less than .025). Hospitals with an alternative birth room had more postpartum options regardless of obstetric volume. Hospitals with an alternative birth room and less than 1000 annual deliveries had more delivery and bonding options. The results of this analysis, plus the fact that 47 of 78 (60%) responding hospitals offered more than half of the options surveyed, suggest that Washington State hospitals have responded to new consumer desires more than previously appreciated.