(−)-Epigallocatechin-3-Gallate Induces Non-Apoptotic Cell Death in Human Cancer Cells via ROS-Mediated Lysosomal Membrane Permeabilization

Abstract

(−)-Epigallocatechin-3-gallate (EGCG) is the most extensive studied tea polyphenol for its anti-cancer function. In this study, we report a novel mechanism of action for EGCG-mediated cell death by identifying the critical role of lysosomal membrane permeabilization (LMP). First, EGCG-induced cell death in human cancer cells (both HepG2 and HeLa) was found to be caspase-independent and accompanied by evident cytosolic vacuolization, only observable when cells were treated in serum-free medium. The cytosolic vacuolization observed in EGCG-treated cells was most probably caused by lysosomal dilation. Interestingly, EGCG was able to disrupt autophagic flux at the degradation stage by impairment of lysosomal function, and EGCG-induced cell death was independent of Atg5 or autophagy. The key finding of this study is that EGCG is able to trigger LMP, as evidenced by Lyso-Tracker Red staining, cathepsin D cytosolic translocation and cytosolic acidification. Consistently, a lysosomotropic agent, chloroquine, effectively rescues the cell death via suppressing LMP-caused cytosolic acidification. Lastly, we found that EGCG promotes production of intracellular ROS upstream of LMP and cell death, as evidenced by increased level of ROS in cells treated with EGCG and the protective effects of antioxidant N-acetylcysteine (NAC) against EGCG-mediated LMP and cell death. Taken together, data from our study reveal a novel mechanism underlying EGCG-induced cell death involving ROS and LMP. Therefore, understanding this lysosome-associated cell death pathway shed new lights on the anti-cancer effects of EGCG.

Suppression of Lysosome Function Induces Autophagy via a Feedback Down-regulation of MTOR Complex 1 (MTORC1) Activity

“…3). This had been noted in another study (34), and we had used up to 100 M CQ with no obvious differences in AO staining (data not shown). Nevertheless, CQ does alter lysosomal pH (25), which would be responsible for its inhibitory effect on proteolytic activities as it has no direct effect on enzymes (Fig.…”

Epigallocatechin Gallate (EGCG) Stimulates Autophagy in Vascular Endothelial Cells

“…Previously, it has been shown that a high concentration (50 -100 M) of EGCG stimulates autophagy leading to cell death in cancer cells (44,45). Another study reported that EGCG stimulates autophagy, which leads to inhibition of endotoxin-induced septic shock through EGCG-induced degradation of HMGB1, a late lethal inflammatory factor (46).…”

FBS or BSA Inhibits EGCG Induced Cell Death through Covalent Binding and the Reduction of Intracellular ROS Production

“…To date, results from most of the studies which examined EGCG-induced cell death suggested that caspase-dependent apoptosis was responsible [3–5], although nonapoptotic cell death was also reported in several studies [6, 7]. We have also investigated the cancer cell-killing effects of EGCG in a cell model, and interestingly, it was found that although EGCG induced cell death in both HepG 2 and HeLa cells, it can only do so under serum-free condition [8]. Furthermore, we have also shown that the cells died of a nonapoptotic cell death via ROS-mediated lysosomal membrane permeabilization (LMP).…”

“…Cell viability was quantified using PI exclusion test as previously described [8]. Briefly, cells were seeded into a 96-well plate at 5 × 10 3 per well.…”