Fig 2: Effects of miR-142-3p over-expression(A) Western blot for TGFBR1 levels in DOK and Cal27 with miR-142 OE or OE Control vectors, Percent change values were calculated in ImageJ with levels normalized to GAPDH, and show a decrease in TGFBR1 expression of 70.1% in DOK and 40.0% in Cal27. Additionally Cal27 and DOK miR-142 OE cells were infected with TGFBR1 and control ORF rescue vectors and shown at a lower exposure time. The growth of (B, D) Cal27 and C,E: DOK by MTT proliferation assay, (B and C) demonstrating the effect of miR-142-3p over-expression and (D, E) demonstrating phenotypic rescue by the addition of TGFBR1 ORF vector. P values were determined by Student's t-test on the final day, error bars represent standard deviation.

Fig 3: Effects of SEV transfer(A) White light and fluorescent images of HMEC1 cells with and without the addition of CD63-GFP stained SEVs under phase contrast. All tube formation assay images were taken 16 hours after seeding. (B) Tube formation assay on HMEC1 cells with the addition of SEVs from Cal27 OE Control and Cal27 142 OE. (C, D) Average length of formed tubes in SEV addition HMEC1 cells (C) or over-expression lines (D) in three fields of view across three wells; error bars represent standard deviation and p values were determined by Student's t-test. (E) Tube formation assay in HMEC1 cells over-expressing miR-142-3p or a control vector. (F) Western blot for TGFBR1 and GAPDH comparing the levels in HMEC1 cells after the addition of SEVs extracted from Cal27 OE Control or Cal27 miR-142 OE cell lines and HMEC1 cells expressing miR-142 OE or OE Control vectors. Percent change values were calculated in ImageJ with levels normalized to GAPDH, there was a decrease of 36% and 79% respectively in the levels of TGFBR1 between cells given OE Control SEVs and cells given miR-142 OE SEVs when compared to cells given no SEVs. There was a decrease of 78% TGFBR1 levels between HMEC1 OE Control cells and HMEC1 miR-142 OE cells.

Fig 4: GDF1 acted mainly through the ALK7–SMAD2/3 signalling cascade.a PLC-8024 cells transfected with or without GDF1 were transduced with lentivirus-mediated shRNAs specifically targeting ALK4, ALK5, and ALK7, respectively. Knockdown efficiency was confirmed at protein level by western blot (n = 3 independent experiments). b Cell migration assay was performed in subgroups targeting different ALKs (NS, not significant, two-tailed independent Student’s t test, data are presented as mean values ± SEM, n = 3 independent experiments). c Sphere-formation assay was performed in subgroups targeting different ALKs (NS, not significant, two-tailed independent Student’s t-test, data are presented as mean values ± SEM, n = 3 independent experiments). d Recombinant GDF1 protein (50 ng/mL) was added in PLC-8024 cells, the phosphorylated and total SMAD2/3 were detected by western blot at indicated time points (n = 3 independent experiments). e, f The liver progenitor markers and hepatic differentiation markers were examined by western blot in PLC-8024-CTR and PLC-8024-GDF1 cells transduced with lentivirus-mediated shRNAs specifically targeting SMAD2 (e) or SMAD3 (f) (n = 3 independent experiments). Source data are provided as a Source Data file.