Extended Data Fig. 10: VCAM1 is an adhesive receptor for tMac-EVs.
From: An extracellular vesicle-mediated mitochondrial transfer network critical for testosterone synthesis
a, Experimental strategy to label LCs by intratesticular injection of AAV-CAG-GFP-U6-sgScramble (AAV-GFP-sgScr) into the testes of Cyp17a1Cre; R26Cas9-tdTomato mice. b, Representative confocal images of the testes from AAV-GFP-sgScr injected Cyp17a1Cre; R26Cas9-tdTomato mice. The testes sections were counterstained with DAPI (n = 5 mice). Scale bar, 20 μm. c, Percentage of LCs infected with AAV-GFP-sgScr (n = 5 mice). d, The gating strategy for sorting tMac-EVs containing mitochondria by FACS in the 3,000g pellets from TAM -induced Cx3cr1CreER R26mitoD2 mice. e, Representative images of tMac-EVs containing mitochondria after FACS (n = 5 biological replicates). Scale bar, 5 μm. f, Schematic diagram depicts experimental design. Testes tissues from Cx3cr1CreER; R26mitoD2 mice were cultured with IgG or VCAM1 mono-antibody (anti-VCAM1) for 48 h before analysis. g, Representative confocal images of testes tissues cultured with IgG or anti-VCAM1. Testes sections were immunostained with CYP17A1 and counterstained with nuclear dye TWO-PRO-3 (white) (n = 5 biological replicates per group). Scale bars, 10 μm. h, Percentage of LCs containing mitoD2+ signals (n = 5 biological replicates per group). i, The concentrations of testosterone in the supernatant of cultured testes tissue treated with IgG or anti-VCAM1 (n = 5 biological replicates per group). j, Schematic diagram depicts experimental design. Testes tissues from Cx3cr1GFP mice were cultured with IgG or ITGβ1 mono-antibody (anti-ITGβ1) for 48 h before analysis. k, Representative confocal images of testes tissues cultured with IgG or anti-ITGβ1. Testes sections were immunostained with CYP17A1 and counterstained with DAPI (n = 5 biological replicates per group). Scale bars, 2 μm. l, Percentage of LCs containing GFP+ signals (n = 5 biological replicates per group). m, The concentrations of testosterone in the supernatant of cultured testes tissue treated with IgG or anti-ITGβ1 (n = 5 biological replicates per group). Data are represented by mean ± SEM (c) or box plots (h,i,l,m). Box plots: the box bounds depict the first quartile to the third quartile, with the middle line indicating the median and the whiskers extending to the minimum and maximum values. Significances were determined using Two-tailed Student’s t-test (h,i,l,m). Schematics in a,f,j created in BioRender. Xia, K. (2026) https://biorender.com/mdov2hb. Source numerical data are provided.
