Development of mouse morulae after encapsulation in alginate microgels or poly-l-lysine microcapsule
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Three experiments were conducted to evaluate in vitro and in vivo development of zona pellucid a-intact (ZPI) and zona pellucida-free (ZPF) mouse embryos after encapsulation in either 2% sodium alginate or 0.1% poly-L-Iysine (PLL). In Experiment 1, rate of development of ZPI embryos (n = 150) from morulae to hatched blastocysts was measured after encapsulation in alginate or PLL and as unencapsulated controls. Following encapsulation, developmental stages were recorded every 24 h for 120 h. Percentage of encapsulated embryos completely hatched from the zona pellucid a were not different from each other but were lower than unencapsulated controls at 48, 72, 96 and 120 h.
Development of ZPI and ZPF mouse embryos after encapsulation in either alginate or PLL was examined in Experiment 2. Developmental stages and diameters were recorded every 24 h for 72 h. At 72 h, embryos were stained and fixed on slides to examine nuclei. Percentage of ZPI embryos developing to expanded blastocysts, their diameters and nuclear counts were not different from each other or from ZPF embryos. Percentage of ZPI embryos initiating hatching or completely hatched from the zona pellucida, their diameters and nuclear cell numbers were also similar.
In the final experiment, ZPI mouse morulae were unencapsulated or encapsulated in either alginate or PLL and transferred into recipients to examine in vivo development. Recipients were allowed to develop fetuses to term. Recipients receiving encapsulated embryos failed to deliver pups. However, five of six recipients of unencapsulated embryos (n = 71) delivered a total of 16 live pups. Additional transfers were performed to examine viable fetuses and resorption sites on day 10 of gestation. Pregnancy rates, diagnosed by the presence of via bIe fetuses or resorption sites, were similar for all treatments: unencapsulated (71.4%), a1ginate (87.5%) and PLL (87.5%). However, the total number of viable fetuses present was higher for unencapsulated embryos (42.1 %) when compared to embryos in alginate microgels (17%) and embryos in PLL microcapsules (14.60/0). Additionally, recipients of alginate and PLL encapsulated embryos had more resorption sites (4 0/0 and 13.4%) when compared to recipients of unencapsulated embryos (0%).
These investigations demonstrated that development of encapsulated ZPI mouse morulae is impaired at the hatched blastocyst stage; however, encapsulated ZPI and ZPF mouse morulae develop similarly in size and nuclear counts. In vivo development of ZPI morulae was also impaired due to an asynchronous condition between the uterine environment and the developing embryos.
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