Figure 4. EVs from ESCs Help Maintain the Ability of ESCs to Incorporate into a Developing Embryo and Generate a Chimeric Animal.

(A) v6.5 mouse ESCs cultured in 2i+LIF medium, or N2B27 medium supplemented without (untreated) or with MVs and exosomes from ESCs for 30 h were immunoblotted for Nanog and vinculin as the loading control.*

(B) Diagram depicting the experiment used to show whether the ESCs treated with EVs were capable of generating chimeric mice. v6.5 mouse ESCs grown in 2i+LIF medium or N2B27 medium supplemented with MVs and exosomes from ESCs were injected into embryonic day 3.5 (E3.5) blastocysts and surgically implanted into the uteri of surrogate mice. The ability of the cells to successfully incorporate into the embryos was evaluated by a change in coat color.

(C) Image showing an example of a chimeric mouse (i.e., ESCs successfully integrated) and a mouse with a white coat (i.e., ESCs failed to integrate).

(D) Table showing the number of embryos injected and transferred, the number of mice born, the number of chimeric mice born, and the integration rates of ESCs grown in 2i+LIF versus N2B27 medium supplemented with MVs and exosomes isolated from pluripotent ESCs.

*The data shown in (A) are presented as mean ± SD. The experiment was performed at least three independent times, and statistical significance was determined using Student’s t test; ***p < 0.001 and **p < 0.01.