Impact of Autophagy Inhibition at Different Stages on Cytotoxic Effect of Autophagy Inducer in Glioblastoma Cells

Abstract

Background/Aims: Glioblastoma multiforme (GBM) is the most malignant primary brain tumor with a poor prognosis. Combination treatment of autophagy inducer and autophagy inhibitor may be a feasible solution to improve the therapeutic effects. However, the correlation between them is unclear. The purpose of this study was to investigate the effect of autophagy inhibition at different stages on cytotoxicity of autophagy inducers in glioblastoma cells. Methods: Autophagy inhibition at early stage was achieved by 3-methyladenine (3-MA) or Beclin 1 shRNA. Autophagy inhibition at late stage was achieved by chloroquine (CQ) or Rab7 shRNA. Cell viability was assessed by MTT assay. Autophagy was measured using transmission electron microscopy and western blot. Apoptosis was measured using western blot and flow-cytometry. Results: Inhibition of early steps of autophagy by 3-MA or Beclin 1 knockdown decreased the toxic effect of arsenic trioxide (ATO) in GBM cell lines. In contrast, blockade of autophagy flux at late stage by CQ or Rab7 knockdown enhanced the cytotoxicity of ATO, and caused accumulation of degradative autophagic vacuoles and robust apoptosis. Moreover, depletion of Beclin 1 abolished the synergistic effect of ATO and CQ by reducing autophagy and apoptosis. Combination of CQ with other autophagy inducers also induced synergistic apoptotic cell death. Conclusion: These results suggest that inhibition of late process of autophagy, not initial step, increases the cytotoxic effect of autophagy inducers via autophagy and apoptosis, which may contribute to GBM chemotherapy.

(-)-Epigallocatechin-3-Gallate Inhibits Arsenic-Induced Inflammation and Apoptosis through Suppression of Oxidative Stress in Mice

“…Arsenic has been shown to be a strong inducer of apoptosis in various models through excessive production of ROS, decreased mitochondrial membrane potential, activation of caspase, increased fragmentation of DNA and impairment of T-cell activity via up-regulation of immune and stress response genes associated with increased production of ROS, which might be involved in the apoptosis and necrosis that it induces in immune cells [10, 33-35]. Earlier reports demonstrated a strong link between immunotoxicity and formation of reactive oxygen species (ROS), which is related to arsenic exposure [36].…”

Fstl1 Promotes Glioma Growth Through the BMP4/Smad1/5/8 Signaling Pathway

“…Several genetic alterations in GBMs have been described, but the prognosis of glioma patients remains poor due to the fast proliferation rate of glioma cells, short course of the disease, difficulty of early diagnosis, unique position of the lesions, incomplete resection and low sensitivity to chemotherapy and radiotherapy [33-36]. …”

Raddeanin a Suppresses Glioblastoma Growth by Inducing ROS Generation and Subsequent JNK Activation to Promote Cell Apoptosis

“…Transmission electron microscopy observation Autophagosome formation was visualized by transmission electron microscopy (TEM) as previously described [31]. Cells (1×10 5 cells/well) were seeded in 6-well plates and allowed to attach.…”

“…Overwhelming evidence suggests that autophagy is a double-edged sword in carcinogenesis, either preventing tumor initiation or sustaining tumor metabolism [38]. To confirm the exact role of RA-induced autophagy in GBM cells, we examined cell viability in the presence or absence of an inhibitor of class III PI3K, 3-MA [31,35], which is a commonly used autophagy inhibitor. Interestingly, inhibition of autophagy exacerbated glioma cell death (Fig.…”