Skip to main content
. 2024 Jul 3;19(8):1107–1121. doi: 10.1016/j.stemcr.2024.06.002

Figure 6.

Figure 6

Lipidomic analysis of CORD21 and isogenic control ROs

(A) Heatmaps demonstrate statistically significant changes in lipid species under positive and negative ion modes of data acquisition based on t test and p value <0.05 cutoff (n = 4 different differentiation experiments each consisting of 48 ROs/sample).

(B) Recoverin (RCVRN) and ceramide double staining in day 220 ROs shows ceramide accumulation basally to photoreceptor ISs. In WT and isogenic controls, ceramide expression is mostly limited to the IS of the photoreceptor layer indicated by RCVRN staining. By contrast, in CORD21-Ros, ceramide accumulation extends beyond the ISs into the photoreceptor cell bodies, and further basally into the reaches of secondary neurons at the apical edge. These are representative examples from 15 ROs imaged from three different differentiation experiments. Scale bars represent 20 μm.

(C) Ceramide expression (green) pertains to the RCVRN+ photoreceptor layer (red, right image) where ceramide is co-expressed with DRAM2 within the IS region of wild-type ROs (red, left image). These are representative examples from 15 ROs imaged from three different differentiation experiments/sample. Scale bars top images 20 μm; bottom magnified images scale bars 10 μm.

(D) POS treatment leads to the accumulation of lipid-containing organelles (LCOs, red arrowheads) and lamellar bodies (LBs, black arrowheads) in CORD21-P1 POS (+) relative to untreated CORD21-P1 POS (−) or CORD21-P1c POS (+) RPE cells (scale bar, 1 μm).

(E) TEM analysis showed accumulation of LCOs, stage II abnormal mitochondria, and LBs in POS-treated CORD21-P1 RPE cells. LCO (scale bar, 0.5 μm), LBs (scale bar, 0.25 μm). Stage II aberrant mitochondria (SIIMito) (scale bar, 0.5 μm). Plots show the significant accumulation of LCOs, SIIMito, and LBs in CORD21-P1 POS (+) relative to CORD21-P1 POS (−) and CORD21-P1c POS (+) RPE cells (Kruskal-Wallis test). Data are presented as mean +SEM (n = 10–12 images of RPE cells from three different differentiation experiments/sample). Statistical comparisons for CORD21-P1 POS (−) vs. CORD21-P1 POS (+), CORD21-P1c POS (−) vs. CORD21-P1c POS (+) and CORD21-P1 POS (−) vs. CORD21-P1c POS (+) are denoted by ∗p <0 .05, ∗∗p <0 .01, ∗∗∗p <0 .001.