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Fig. 5
Divisions of Unpigmented mitfa-Expressing Cells during Melanocyte Regeneration
(A–C) Lineage tracing of unpigmented mitfa-expressing cells on the flank of adult Tg(miniCoopR-mitfa:nlsEGFP) zebrafish after neocuproine treatment.
(A) Symmetric division of an unpigmented mitfa-expressing cell (yellow arrowheads, days 0, 1, 2, 6). Neither daughter cell differentiated (yellow arrowheads, days 7, 21). Fish were treated with epinephrine only on day 0 prior to imaging. mC, miniCoopR; BF, brightfield, neo; neocuproine. Scale bar, 25 µM.
(B) Asymmetric division of an unpigmented mitfa-expressing cell (yellow arrowheads, days 0, 1, 3, 5) in which one daughter differentiated (white arrowheads, days 7, 18) and the other daughter remained undifferentiated (yellow arrowheads, days 7, 18). Fish were treated with epinephrine only on day 0 prior to imaging. mC, miniCoopR; BF, brightfield, neo; neocuproine. Scale bar, 25 µM.
(C) Schematic summarizing lineages of unpigmented mitfa-expressing cells that differentiated or divided following neocuproine-mediated ablation (dashed line). Percentages of each lineage are indicated.
(D) Timeline of the experiment. Adult wild-type zebrafish were treated with neocuproine for 24 hr to ablate differentiated melanocytes and were injected with EdU every other day until day 8 (arrowheads). After melanocyte regeneration fish were treated with neocuproine again at day 30 for 24 hr. Control animals were injected with EdU but were not treated with neocuproine. Fish were sacrificed at day 40 and were scored for EdU incorporation. neo, neocuproine.
(E) Representative images of control (top row) or neocuproine-treated (bottom row) Mitfa-positive melanocytes after melanocyte regeneration. neo, neocuproine; BF, brightfield. BF scale bar, 15 µM; Mitfa, EdU scale bar, 10 µM.
(F) Quantification of percent EdU-positive, Mitfa-positive melanocytes. n = 97, 256 and 365 cells from each of three fish, respectively, for neo; n = 43, 53, and 85 cells from each of three fish, respectively, for w/o neo. Data are shown as mean percent positive per fish ± SEM; p value calculated by Student’s t test. neo, neocuproine.
See also Table S1.
Reprinted from Developmental Cell, 33(6), Iyengar, S., Kasheta, M., Ceol, C.J., Poised Regeneration of Zebrafish Melanocytes Involves Direct Differentiation and Concurrent Replenishment of Tissue-Resident Progenitor Cells, 631-43, Copyright (2015) with permission from Elsevier. Full text @ Dev. Cell