Figure 1
Induction of autophagy in THP1 cells and MDMs, following treatment with ATP, is both calcium and P2X 7 – dependent. A. THP1 cells undergo autophagy (lane 1) following 30 minutes exposure to ATP (3 mM), as revealed by the presence of LC3-II (16 kDa) by Western blot. Lane 2 represents untreated control THP-1 cells. B. ATP (3 mM) treated MDMs also express LC3-II (lane 2), which is absent in non-treated control cells (lane 1). C. THP1 cells treated with 3 mM ATP for 30 minutes cultured in calcium-free medium (lane 1) and calcium-replete medium (lane 2). LC3-II expression was observed in ATP-treated cells cultured in calcium-replete medium (lane 2) but was absent in cells cultured in calcium-free media (lane 1). D. THP1 cells were treated with different P2 agonists and antagonists. Lane 1 represents untreated control cells, while lanes 2 and 3 represent cells pre-treated with oATP (0.3 mM) and anti-P2X7 antibody (3 μg/ml) for 2 hours and 1 hour respectively, prior to treatment with ATP (3 mM for 30 minutes). Lanes 4, 5 and 6 represent cells treated with ATP (3 mM), UTP (3 mM) and bzATP (3 mM) for 30 minutes. The presence of an LC3-II band, indicating cell autophagy, was observed only in lanes 4 and 6.