Fig 2: Blockage of MFG-E8 on the expression of AMPK and Src in microglia treated with propofol. BV2 cells were treated with or without an MFG-E8 antibody (5 μg/ml)/IgG antibody (5 μg/ml) for 45 min after treatment with propofol (12.5 μM) for 4 h. The expression of p-AMPK, AMPK, p-Src, Src, and GAPDH was measured by western blotting (a). The bands were quantitatively analyzed (b, c, d, e). The data are presented as the mean ± SD. Pro, propofol; M8 Ab, MFG-E8-neutralizing antibody. *P < 0.05 versus the control

Fig 3: Metformin increases beclin1 expression in vivo. (A) The MGC803 cells were injected subcutaneously into the flanks of male nude mice (8 × 106 cells per mouse) to establish a xenograft tumour model. The metformin treatment group was given 300 mg per day of metformin via intraperitoneal injection for 4 weeks, while the control group was given normal saline (100 μL). At 30 days, the mice were humanely euthanised, and the tumour tissue was removed and photographed. (B) Once the tumour were palpable, tumour sizes were measured every 3 days. (C) The volume of the isolated tumours were measured by calipers as shown. (D) The weight of the isolated tumours were measured as shown. (E) Representative images of immunohistochemical staining for beclin1 in each group. (F) Model for metformin activates autophagy to inhibit gastric cancer by activating AMPK and inhibiting the mTOR signalling pathway (**p < 0.01, ***p < 0.001 vs control).

Fig 4: Effects of metformin (1 mM) on the expression of AMPK, p-AMPK, and mitochondrial biogenesis genes under high glucose conditions. Cells were divided into four groups: i) Con, normal medium; ii) Man, 33.3 mM Man; iii) HG, 33.3 mM glucose; and iv) HG + Met, 33.3 mM glucose pretreated with 1 mM Met. (A) Protein expression levels of p-AMPK and AMPK were (A) determined by western blotting and (B) the ratio of p-AMPK/AMPK was semi-quantified. mRNA expression levels of (C) NDUFA1, NDUFA2, NDUFA13, Mn-SOD, (D) PGC-1α, NRF1 and NRF2 were detected via reverse transcription-quantitative PCR. *P<0.05 vs. Con; #P<0.05 vs. HG. AMPK, AMP-activated protein kinase; p, phosphorylated; Man, mannitol; HG, high glucose; Met, metformin; NDUF, NADH: Ubiquinone oxidoreductase subunit; Mn-SOD, manganese superoxide dismutase; PGC-1α, peroxisome proliferator-activated receptor-γ coactivator-1α; NRF, nuclear respiratory factor; Con, control.

Fig 5: A proposed working model for aberrant SMAD4-HNF4G in PDAC invasiveness and the effect of Metformin. In PDAC cells where SMAD4 is sufficient, the expression of the downstream oncogene HNF4G that promotes PDAC invasiveness is physiologically inhibited by the SMAD complex. In PDAC cells where SMAD4 is deficient, the expression of HNF4G is over-activated, which evokes cancer cell invasion and metastasis. Metformin may act as a target drug repressing PDAC invasion and metastasis by activating AMPK that induces phosphorylation-ubiquitination coupled HNF4G degradation