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. 2022 May 25;29(11):2316–2331. doi: 10.1038/s41418-022-01018-8

Fig. 4. Loss of Plin2 induces lipidomic remodeling associated with impaired mitochondrial cristae and FAO.

Fig. 4

a Principal component analysis (PCA) of lipidomic data from WT and Plin2-/- mESCs (KO1 and KO2). Each dot represents a biological replicate. b Volcano plot of differential lipid molecules between WT and Plin2-/- mESCs (KO1 and KO2). Dot size reports P value, while the color indicates fold change (FC). Lipid molecules with P < 0.05 and FC > 1.5 are considered statistically significant. The top 10 lipid molecules with FC > 2.5 are indicated. c CL, PE, and PC contents in mitochondria of WT and Plin2-/- mESCs (KO1 and KO2). Data are mean ± SD, n = 3 biological replicates. Two-tailed unpaired t-tests. ***P < 0.001, **P < 0.005, *P < 0.05. d Oxygen consumption rates (OCR) and maximal mitochondrial respiration in WT and Plin2-/- mESCs (KO1 and KO2). Maximal mitochondrial respiration was determined by the increase of OCR after FCCP treatment. Data are mean ± SEM, n = 2 biological replicates with three technical replicates each. Two-tailed unpaired t-tests. ***P < 0.001. e Endogenous fatty acid oxidation (FAO) rates and maximal endogenous FAO in WT and Plin2-/- mESCs (KO1 and KO2). Maximal endogenous FAO was determined by the increase of OCR after Eto treatment in a substrate-limited assay medium. Data are mean ± SEM, n = 3 biological replicates with three technical replicates each. Two-tailed unpaired t-tests. ***P < 0.001, *P < 0.05. f Exogenous FAO rates and maximal exogenous FAO in WT and Plin2-/- mESCs (KO1 and KO2). Maximal exogenous FAO was determined by the increase of OCR after PA treatment in a substrate-limited assay medium. Data are mean ± SEM, n = 3 biological replicates with three technical replicates each. Two-tailed unpaired t-tests. ***P < 0.001. g Representative TEM images of mitochondria and quantification of mitochondria with disorganized cristae in WT and Plin2-/- mESCs (KO1 and KO2). Representative mitochondria with normal or disorganized cristae are shown below. Scale bar, top: 1 μm, below: 200 nm. Data are mean ± SD, n = 3 biological replicates (each >50 mitochondria). Two-tailed unpaired t-tests. **P < 0.005, *P < 0.05. h Pie charts of mitochondrial proteomic data showing proportion of mitochondrial proteins changed significantly (P < 0.05 and FC > 1.5) in Plin2-/- mESCs (KO1 and KO2) compared with WT mESCs. Mitochondrial proteins are divided into four categories according to their distribution (inner membrane, outer membrane, matrix or intermembrane space). The experiments were repeated twice with similar results, and one replicate was used for this analysis. i Quantification of mitochondria with disorganized cristae in WT and Plin2-/- mESCs (KO1 and KO2) transfected with EV or MT-Plin2. Data are mean ± SD, n = 3 biological replicates (each >50 mitochondria). Two-tailed unpaired t-tests. **P < 0.005. j Maximal mitochondrial respiration in WT and Plin2-/- mESCs (KO1 and KO2) transfected with EV or MT-Plin2. Data are mean ± SEM, n = 3 independent experiments. Two-tailed unpaired t-tests. ***P < 0.001, **P < 0.005.