Figure 7.
APOE4 promotes large LDs with highly unsaturated triglyceride and impaired turnover. (A) Representative confocal slices of TRAE3-H or TRAE4-H cells labeled for LDs with BODIPY 493/503 and imaged live at each timepoint of the OA pulse-chase assay described in Fig. 5 A. Scale bar, 10 µm. (B–D) Quantification of LD parameters in TRAE3-H or TRAE4-H cells at each timepoint of the OA pulse-chase assay. (B) Total LD area was measured as the area of the entire LD mask per cell in µm2. (C) Average LD size was calculated as the mean LD area per cell in µm2. (D) Number of LDs per cell. Each data point represents one cell, and each color represents data collected from a separate, independent experiment. N = 90 cells per genotype, timepoint, and independent experiment. ns P > 0.05, **** P < 0.0001. P values were calculated using a clustered Wilcox rank sum test via the Rosner-Glynn-Lee method and Bonferonni-corrected for multiple comparisons. (E) Comparison of total triglyceride between TRAE3-H and TRAE4-H cells. Lipidomics data were collected from two independently performed experiments, which each used three separate plates of cells as technical replicates. Each data point denotes a single technical replicate, and the dot colors (purple and yellow) indicate data collected from the same independently performed experiment. There is significantly more total triglyceride in E4 cells at the pulse-chase timepoint. * P < 0.05, ns P > 0.05. P values were calculated using the Wilcox rank sum test and Bonferonni-corrected for multiple comparisons. (F) Heatmap showing the relative abundance of measured lipid classes at each timepoint of the assay. Heatmap values are derived from the means of three technical replicates from two independently performed experiments (shown as separate columns) for each condition. Means were then grouped by lipid class and Z-score normalized. The green box frame lipid species that are enriched in E4, while the red box frames lipid species enriched in E3. PG, phosphatidylglycerol; ChE, cholesterol ester; PI, phosphatidylinositol; PC, phosphatidyl choline; PE, phosphatidyl ethanolamine; DG, diacylglycerol; TG, triacylglycerol. (G) Heatmap of the abundance of triglyceride species separated by their degree of unsaturation at each timepoint of the assay. Heatmap values were derived from the means of three technical replicates from two independently performed experiments (shown as separate columns) for each condition. Means were grouped by lipid class and Z-score normalized. The green box frames lipid species enriched in E4.

APOE4 promotes large LDs with highly unsaturated triglyceride and impaired turnover. (A) Representative confocal slices of TRAE3-H or TRAE4-H cells labeled for LDs with BODIPY 493/503 and imaged live at each timepoint of the OA pulse-chase assay described in Fig. 5 A. Scale bar, 10 µm. (B–D) Quantification of LD parameters in TRAE3-H or TRAE4-H cells at each timepoint of the OA pulse-chase assay. (B) Total LD area was measured as the area of the entire LD mask per cell in µm2. (C) Average LD size was calculated as the mean LD area per cell in µm2. (D) Number of LDs per cell. Each data point represents one cell, and each color represents data collected from a separate, independent experiment. N = 90 cells per genotype, timepoint, and independent experiment. ns P > 0.05, **** P < 0.0001. P values were calculated using a clustered Wilcox rank sum test via the Rosner-Glynn-Lee method and Bonferonni-corrected for multiple comparisons. (E) Comparison of total triglyceride between TRAE3-H and TRAE4-H cells. Lipidomics data were collected from two independently performed experiments, which each used three separate plates of cells as technical replicates. Each data point denotes a single technical replicate, and the dot colors (purple and yellow) indicate data collected from the same independently performed experiment. There is significantly more total triglyceride in E4 cells at the pulse-chase timepoint. * P < 0.05, ns P > 0.05. P values were calculated using the Wilcox rank sum test and Bonferonni-corrected for multiple comparisons. (F) Heatmap showing the relative abundance of measured lipid classes at each timepoint of the assay. Heatmap values are derived from the means of three technical replicates from two independently performed experiments (shown as separate columns) for each condition. Means were then grouped by lipid class and Z-score normalized. The green box frame lipid species that are enriched in E4, while the red box frames lipid species enriched in E3. PG, phosphatidylglycerol; ChE, cholesterol ester; PI, phosphatidylinositol; PC, phosphatidyl choline; PE, phosphatidyl ethanolamine; DG, diacylglycerol; TG, triacylglycerol. (G) Heatmap of the abundance of triglyceride species separated by their degree of unsaturation at each timepoint of the assay. Heatmap values were derived from the means of three technical replicates from two independently performed experiments (shown as separate columns) for each condition. Means were grouped by lipid class and Z-score normalized. The green box frames lipid species enriched in E4.

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