Fig. 1: Abnormally elevated exosomal dsDNA was positively correlated with the disease activity in murine colitis and active human CD.

A–C Classification of EVs in colon lavage of murine colitis. A Western blot of specific EVs markers: The simultaneous expression of three classic positive markers of EVs including Alix, CD63, and CD81, along with the absence of Calnexin, together identified EVs. The whole-cell lysate of CT26, a murine cell line, was used as a positive control. B Nanoparticle-tracking analysis of diluted EVs fractions was determined by ZetaView PMX 110 (Particle Metrix, Meerbusch, Germany). The distribution of EVs particle concentration (y axis) by size (x axis) was shown and 126.2-nm particles accounted for the highest proportion (98.5%). C Representative transmission electron microscope images of EVs fractions. The yellow arrow pointed to examples of EVs, which were cup-shape membrane-enclosed particles with diameters of 30–150 nm. D Absolute quantification of nDNA and mtDNA within EVs extracted from plasma and colon lavage of murine colitis. Hist1h3F gene and mtCOI gene were applied to represent nDNA and mtDNA respectively. Exosomal mtDNA and nDNA were significantly higher in the plasma and colon lavage of murine colitis. Murine colitis treated with GW4869 were also examined. n = 5–10/group. In the illustrations, healthy wild-type model, murine colitis, and murine colitis treated with GW4869 were abbreviated as WT, WT + DSS, and WT + DSS (GW4869 IP) respectively. Colon lavage was abbreviated as CL. E Classification of EVs isolated from plasma of active human CD, including western blot, nanoparticle-tracking analysis, and representative transmission electron microscope images. In the nanoparticle-tracking analysis, 117.7-nm particles accounted for the highest proportion (98.6%). In the representative transmission electron microscope image, the yellow arrow pointed to EVs. F Levels of exosomal nDNA and mtDNA from the plasma of CD patients were measured (n = 5/group). Both were significantly higher in patients with active CD. H3 clustered histone 7 gene and mtCOI gene were applied to represent nDNA and mtDNA respectively. G Correlations between the disease activity and levels of exosomal dsDNA in plasma in murine colitis and CD patients (Spearman’s rank correlation analysis). Disease activity index, Crohn’s Disease Activity Index, exosomal nDNA, and exosomal mtDNA were abbreviated as DAI, CDAI, exo-nDNA, and exo-mtDNA in the illustrations respectively. Data were displayed as mean values ± SD at least three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001.