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. 2022 Sep 19;23(11):e55209. doi: 10.15252/embr.202255209

Figure 3. Epithelial plasticity does not correlate with cell age.

Figure 3

  • A
    Diagram depicting the use of TRE‐H2B‐GFP mice for label retention studies. TRE‐H2B‐GFP mice were given doxycycline continually for at least 4 weeks prior to variable withdrawal periods to generate label retaining cell populations of different ages. The GFP+ LRCs were then FACS‐purified, embedded in Matrigel, and exposed to niche growth factors (EGF, R‐Spondin, Noggin) for in vitro organoid formation assays.
  • B
    Immunofluorescence images of small intestines of TRE‐H2B‐GFP mice after the dox withdrawal periods outlined in panel (A). Crypts are outlined with dashed lines. Scale bar = 15 μm.
  • C
    Immunofluorescent image of a TRE‐H2B‐GFP Chga CreER‐2A‐tdTomato mouse after a 10 day dox withdrawal period, demonstrating that EECs and LRCs represent an overlapping population, with half of EECs retaining an H2B‐GFP label. Scale bar = 100 μm. Inset scale bar = 10 μm.
  • D
    Organoid formation efficiency of different aged LRC populations. N = 3 mice/group, N = 3 wells quantified per mouse. Data are expressed as mean ± SEM. N = 3 mice/group, P‐value generated by Ordinary one‐way ANOVA.
  • E
    Images of organoids quantified in (D). Scale bar = 500 μm.