Figure 6.

Role of mitochondria in stem cell differentiation

(A and A′) Planarian cells stained with MTG and MTO were analyzed through flow cytometry. X1 (A) and X2 (A′) were gated for MTG^Low and ^−High and analyzed for MTO intensity. The bar graph indicates the median MTO intensity normalized to FCCP. Mean ± SEM, over three independent replicates. a.u., arbitrary units.

(B) Representative images of undergoing regeneration in the presence of 30 nM FCCP or DMSO (vehicle control). Scale bar, 200 μm.

(C) Representative confocal images of piwi-1 RNA FISH at 3 dpa in trunk fragments regenerating in 30 nM FCCP or DMSO (control). Scale bar, 100 μm.

(D) Quantitation of piwi-1⁺ cells from the 3dpa anterior blastema region from (C). Mean ± SD, n = 7 animals.

(E) Quantitation of piwi-1 mRNA levels through qPCR from 3 dpa regenerating animals. Mean ± SD, n = 30 worms over three independent replicates.

(F) Schematic representing the in vitro assay for perturbing mitochondrial activity using FCCP.

(G) PIWI-1^High quantitation through flow cytometry in 4N-MTG^Low cells treated with the indicated amount of FCCP for 48 h in culture. Mean ± SEM, over three independent replicates.

(H and I) Representative piwi-1 RNA FISH (H) and quantification of piwi-1⁺ cells (I) per transplant in lethally irradiated sexual worms 8 dpt of ~2500 cells from 4N-MTG^Low cells cultured in vitro for 48 h with or without FCCP. Scale bar, 200 μm. Mean ± SEM, n = 9 worms.

(J) Percentage of worms rescued following injection of ~2500 4N-MTG^Low cells cultured in vitro for 48 h with or without FCCP. Mean ± SEM, n = 24 worms performed over three independent replicates.

^∗, ^∗∗, n.s. represents p < 0.05, p < 0.01, and nonsignificant, respectively.