Extended Data Figure 9 : Long-term corneal restoration by donor human ABCB5+ cell transplants 13 months after transplantation.

From: ABCB5 is a limbal stem cell gene required for corneal development and repair

Extended Data Figure 9

a, Representative H&E composite corneal cross-sections of recipient immunodeficient NSG mice 13 months after LSCD induction followed by transplantation of donor fibrin gel grafts containing: (1) no cells (negative control), (2) ABCB5+ cells, (3) ABCB5 cells, and (4) unsegregated cells. A normal untreated NSG cornea (no LSCD) served as a positive control. The positive control displays the typical stratified corneal epithelium. Mice that received transplants with no cells displayed evidence of conjunctivalization that occurs following a LSC deficiency; that is, unstratified conjunctival epithelium covers the cornea with extensive neovascularization and synechia (anterior segment inflammation is muted in NSG mice due to their immunodeficiency). In contrast, mice that received transplants containing ABCB5+ cells displayed restored stratified epithelium, whereas recipients receiving ABCB5 cell grafts did not. b, Representative immunofluorescent KRT12 staining (green) of corneas derived from NSG mice 13 months after LSCD induction followed by transplantation of donor fibrin gel grafts containing the following human limbal epithelial cell subpopulations: (1) no cells (negative control), (2) ABCB5+ cells, (3) ABCB5 cells, or (4) unsegregated cells. Normal untreated murine cornea (no LSCD), shown here as a positive control, displayed a high intensity of KRT12 staining. As expected, recipients of grafts containing no cells displayed no KRT12 expression. Mice transplanted with ABCB5+ cells exhibited significant KRT12 expression, also enhanced compared to mice transplanted with unsegregated limbal epithelial cells. No KRT12 expression was detected in mice transplanted with ABCB5 cells. The white arrow depicts the area shown at ×40 magnification. Montage images are shown at ×10 magnification. c, Representative immunofluorescent KRT12 staining (red) of human and mouse cornea confirms specific antibody reactivity with human KRT12 (top left), and no cross-reactivity with murine Krt12 (bottom left). Isotype control antibody staining is shown for the respective tissues in the right panels. Nuclei are stained with DAPI (blue). Bar graph (bottom) demonstrates aggregate antibody staining data of either human cornea (pixel intensity 142.3 ± 2.4 pixels µm−2, mean ± s.e.m.) or mouse cornea (pixel intensity 1.3 ± 0.7 pixels µm−2, mean ± s.e.m.). Aggregate human KRT12 antibody staining data of either human or mouse cornea was derived from the analyses on n = 2 corneas per group. Within a standardized area in (b), all corneal epithelial cells were counted in at least n = 3 consecutive cross-sections. Data were analysed using the unpaired t-test. Error bars show s.e.m. ***P < 0.001. NS, not significant.