Fig. 2: SLCs establish fewer mitochondria-endoplasmic reticulum contacts during ageing. | Nature Communications

Fig. 2: SLCs establish fewer mitochondria-endoplasmic reticulum contacts during ageing.

From: Nestin-dependent mitochondria-ER contacts define stem Leydig cell differentiation to attenuate male reproductive ageing

Fig. 2

a Representative immunostaining pictures of colocalization between mitochondria and ER in primary Nestin-GFP + SLCs from different age groups. Mitochondria and ER are marked with Mitotracker and ER tracker, respectively. Scale bar, 10 μm. b, c Quantification of the levels of colocalization in (a) (Manders’ (of mitochondria) and Pearson’s coefficients are shown for each condition, n = 49, 55 and 51 cells for 2 m, 12 m, and 24 m SLCs, respectively; One-way ANOVA). d Representative immunostaining pictures and 3D reconstruction of mitochondria and ER in primary Nestin-GFP + SLCs from different age groups. Scale bar, 5 μm. e Representative transmission electron microscope (TEM) images of primary Nestin-GFP + SLCs from different age groups. Scale bar, 250 nm for original pictures and 100 nm for enlarged pictures. f Quantification of the extent of MERCs (<100 nm) from (e) in primary Nestin-GFP + SLCs from different age groups. The relative amount of ER-OMM apposition in percentage of OMM refers to the interaction length of ER tubules within 100 nm of the OMM covered in the percentage of total OMM per mitochondria (n = 26 to 48 mitochondria in 5 fields per condition; One-way ANOVA). g Quantification of the mito-ER proximity from e in primary Nestin-GFP + SLCs from different age groups (n = 26 to 48 mitochondria in 5 fields per condition; One-way ANOVA). h Representative immunostaining pictures, 3D reconstruction and surface-surface contact site area coverage of VDAC (mitochondria) or IP3R1(ER) in primary Nestin-GFP + SLCs from different age groups. Scale bar, 10 μm. i Quantification of the surface-surface contact site area (percentage of mitochondria) in (h) (n = 5 biological repeats for each group; One-way ANOVA). j Quantification of surface-surface contact site area (percentage of ER) in (h) (n = 5 biological repeats for each group; One-way ANOVA). Two-sided comparison; All data are mean ± SD; Error bars represent SDs. **p < 0.01, ***p < 0.001; Source data are provided as a Source Data file. See also Supplementary Fig. 2.

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