Figure 2. Neonatal mouse uterine mesenchyme induces MIXL1 expression in MIXL1^GFP/w EBs.

(A) nMUM induced GFP expression in MIXL1 ^GFP/w EB recombinants after three days of co-culture (3^rd panel), representative example of n>100. The duration of reporter activation is comparable to that observed in growth factor treated EBs (2^nd panel, days 3,5), while little to no reporter activity was detected in the control (1^st panel) (n>100) (×40 magnification, first 3 panels are fluorescent images, images in 4^th panel represent phase contrast images of recombinants in 3^rd panel). (B) Co-culture of two nMUM pieces (0.5 mm) with a larger EB (>4000 cells) activated reporter activity locally (dotted area is the EB) (×20 magnification). (C) No reporter activity was detected in embryoid bodies co-cultured with neonatal uterine epithelium (0.5 mm) (×40 magnification). Areas marked as mesenchyme in recombinants are based on morphological evidence and recombination experiments using ENVY hESC (refer to Figure S2A–C). Abbreviations: EB, embryoid body; E, neonatal mouse uterine epithelium; GF, growth factors; M, neonatal mouse uterine mesenchyme.