Properties of mouse inner cell masses isolated by Immunosurgery, exposure to the calcium ionophore A23187 or low osmolarity

Abstract

Mouse inner cell masses remained intact when exposed to extreme osmotic stress (distilled water) for short periods, but the trophectoderm was lysed in one-third of blastocysts. However, these inner cell masses were not viable as they could not fluoresce after incubation in fluorescence diacetate nor continue development in vitro. It was concluded that cells of the inner cell mass are not more tolerant of osmotic stresses than those of the trophectoderm. Inner cell masses were isolated from only 50% of blastocysts when exposed to the calcium ionophore A23187. Some lysing trophectoderm cells are probably able to restore their normal calcium levels after transfer to fresh culture medium allowing normal blastocyst-like development in vitro. Further evidence which suggests that not all trophectoderm cells are removed after incubation in ionophore is that these inner cell masses produced more trophoblast-like outgrowths in vitro and were able to induce the decidual cell reaction in vivo more often than their immunosurgically isolated counterpart. Immunosurgically isolated inner cell masses were viable as they fluoresced brightly after incubation in fluorescein diacetate and developed normally in vitro. There was little or no contamination of these inner cell masses with trophectoderm cells as they formed considerably smaller outgrowths than control blastocysts and were rarely able to induce the decidual cell reaction in vivo. Trophoblast-like giant cells were found less frequently from inner cell masses isolated from 142-h, post-HCG blastocysts and cultured in vitro, than from 118-h, post-HCG blastocysts. These results are discussed in relation to a current theory on the time of inner cell mass determination which states that apparently differentiated inner cell mass cells may reverse their direction of development in response to altered environmental conditions.