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. 2017 Dec 27;12(12):e0189514. doi: 10.1371/journal.pone.0189514

Fig 6. Effect of A2M* on expression of PTEN and miR-21 in vitro and in vivo.

Fig 6

(a) Various tumour cells (A549, HT29, LOX, MDA-MB 231) were cultured in the presence of A2M* (0, 10, 30, 100 nM) and cell lysates were probed for PTEN by immunoblotting. Protein amounts loaded were normalized to GAPDH levels, (b) 1321N1 cells mutated for PTEN were treated with A2M* and expression of SNAIL and E-Cadherin compared to GAPDH was studied by immunoblotting, (c) Separation of PCR products of SNAIL and GAPDH in 1.5% agarose gels (d) A549 cell were cultured in the presence of A2M* (0, 10, 30, 100 nM) for 8h followed by qRT-PCR analysis of pri-miR-21 and mature miR-21 transcripts (n = 3), error bars are mean ± sem. (e) A549 tumours from mice were homogenized and analysed by immunoblotting for AKT/pAKT, GSK3B/pGSK3B, GAPDH and followed by semi-quantitative pixel analysis (n = 3). Error bars are mean ± sem (t-test), (f) Analysis of PTEN expression in A549 tumours. Data are means ± sem of n = 2 technical replicates. (g) Tumour slices obtained from A2M*-treated and non-treated mice were immunostained for Ki-67 and quantitatively analysed. Data represent mean ± sem (n = 5; t-test) (*P < 0.05, **P < 0.01).