Fig. 2

ARDS subjects show increased counts of circulatory EV_ARDS immunoreactive to TβRI/Alk5. a The mean count for EVs isolated from the blood of the ARDS patients (EV_ARDS) was 13,680 ± 1811, with a range 11,000–17,500; Nine EV_ARDS preparations from nine different ARDS patients were used. The EV_Ctrl mean count indicated 8615 ± 768, with a range 7700–9200; three preparations from the blood of three different control subjects were used. Data are normalized per 1 μl blood. Three independent experiments were performed. b Amount of total protein in the EV_ARDS pellet and EV_Ctrl pellet, normalized to 1 ml blood. Data are plotted as mean ± SD; *p < 0.05 vs. controls. c, d WB of Alk5 and TβRII protein expression in two EV_ARDS and two EV_Ctrl lysates, normalized to equivalent μl blood; n = 9. Actin cannot be used as loading control, as there is no evidence that the EVs contain actin. e Time course analysis of Alk5 in EV_ARDS in one of the above ARDS cases; n = 3. Day 1 is first day in ICU. f Densitometric analysis of Alk5 protein expression in the EV_{Ctrl and} EV_ARDS lysates. Data are plotted as mean ± SD; *p < 0.05 vs. controls. g Size distribution and concentration of exosomes and smaller than 300 nm diameter microparticles in the isolated EV_Ctrl and EV_ARDS preparations, measured by NTA; n = 3. h Representative WB analyses of EV_ARDS fractions of three different patients using Abs specific for CD9, CD81, syntenin-1, CD63, actinin-1 and mitofilin. Actinin-1 and mitofilin, considered specific markers for the non-exosomal microvesicles are enriched in the EV_ARDS preparations; n = 3