Browsing by Author "Ristic, Biljana (7006688884)"

Purpose To investigate the ability of chloroquine, a lysosomotropic autophagy inhibitor, to enhance the anticancer effect of nutrient deprivation. Methods Serum-deprived U251 glioma, B16 melanoma and L929 fibrosarcoma cells were treated with chloroquine in vitro. Cell viability was measured by crystal violet and MTT assay. Oxidative stress, apoptosis/necrosis and intracellular acidification were analyzed by flow cytometry. Cell morphology was examined by light and electron microscopy. Activation of AMP-activated protein kinase (AMPK) and autophagy were monitored by immunoblotting. RNA interference was used for AMPK and LC3b knockdown. The anticancer efficiency of intraperitoneal chloroquine in calorierestricted mice was assessed using a B16mouse melanoma model. Results Chloroquine rapidly killed serum-starved cancer cells in vitro. This effect was not mimicked by autophagy inhibitors or LC3b shRNA, indicating autophagy-independent mechanism. Chloroquine-induced lysosomal accumulation and oxidative stress, leading tomitochondrial depolarization, caspase activation and mixed apoptotic/necrotic cell death, were prevented by lysosomal acidification inhibitor bafilomycin. AMPK downregulation participated in chloroquine action, as AMPK activation reduced, and AMPK shRNA mimicked chloroquine toxicity. Chloroquine inhibited melanoma growth in calorie-restricted mice, causing lysosomal accumulation, mitochondrial disintegration and selective necrosis of tumor cells. Conclusion Combined treatment with chloroquine and calorie restriction might be useful in cancer therapy. © Springer Science+Business Media, LLC 2012.

Purpose To investigate the ability of chloroquine, a lysosomotropic autophagy inhibitor, to enhance the anticancer effect of nutrient deprivation. Methods Serum-deprived U251 glioma, B16 melanoma and L929 fibrosarcoma cells were treated with chloroquine in vitro. Cell viability was measured by crystal violet and MTT assay. Oxidative stress, apoptosis/necrosis and intracellular acidification were analyzed by flow cytometry. Cell morphology was examined by light and electron microscopy. Activation of AMP-activated protein kinase (AMPK) and autophagy were monitored by immunoblotting. RNA interference was used for AMPK and LC3b knockdown. The anticancer efficiency of intraperitoneal chloroquine in calorierestricted mice was assessed using a B16mouse melanoma model. Results Chloroquine rapidly killed serum-starved cancer cells in vitro. This effect was not mimicked by autophagy inhibitors or LC3b shRNA, indicating autophagy-independent mechanism. Chloroquine-induced lysosomal accumulation and oxidative stress, leading tomitochondrial depolarization, caspase activation and mixed apoptotic/necrotic cell death, were prevented by lysosomal acidification inhibitor bafilomycin. AMPK downregulation participated in chloroquine action, as AMPK activation reduced, and AMPK shRNA mimicked chloroquine toxicity. Chloroquine inhibited melanoma growth in calorie-restricted mice, causing lysosomal accumulation, mitochondrial disintegration and selective necrosis of tumor cells. Conclusion Combined treatment with chloroquine and calorie restriction might be useful in cancer therapy. © Springer Science+Business Media, LLC 2012.

Graphene-based nanomaterials (GNM) are plausible candidates for cancer therapeutics and drug delivery systems. Pure graphene and graphene oxide nanoparticles, as well as graphene quantum dots and graphene nanofibers, were all able to trigger autophagy in cancer cells through both transcriptional and post-transcriptional mechanisms involving oxidative/endoplasmic reticulum stress, AMP-activated protein kinase, mechanistic target of rapamycin, mitogen-activated protein kinase, and Toll-like receptor signaling. This was often coupled with lysosomal dysfunction and subsequent blockade of autophagic flux, which additionally increased the accumulation of autophagy mediators that participated in apoptotic, necrotic, or necroptotic death of cancer cells and influenced the immune response against the tumor. In this review, we analyze molecular mechanisms and structure–activity relationships of GNM-mediated autophagy modulation, its consequences for cancer cell survival/death and anti-tumor immune response, and the possible implications for the use of GNM in cancer therapy. © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

Graphene-based nanomaterials (GNM) are plausible candidates for cancer therapeutics and drug delivery systems. Pure graphene and graphene oxide nanoparticles, as well as graphene quantum dots and graphene nanofibers, were all able to trigger autophagy in cancer cells through both transcriptional and post-transcriptional mechanisms involving oxidative/endoplasmic reticulum stress, AMP-activated protein kinase, mechanistic target of rapamycin, mitogen-activated protein kinase, and Toll-like receptor signaling. This was often coupled with lysosomal dysfunction and subsequent blockade of autophagic flux, which additionally increased the accumulation of autophagy mediators that participated in apoptotic, necrotic, or necroptotic death of cancer cells and influenced the immune response against the tumor. In this review, we analyze molecular mechanisms and structure–activity relationships of GNM-mediated autophagy modulation, its consequences for cancer cell survival/death and anti-tumor immune response, and the possible implications for the use of GNM in cancer therapy. © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

We investigated the role of autophagy, a stress-inducible lysosomal self-digestion of cellular components, in modulation of herpes simplex virus type 1 (HSV-1)-triggered death of U251 human glioma cells. HSV-1 caused apoptotic death in U251 cells, characterized by phosphatidylserine externalization, caspase activation and DNA fragmentation. HSV-1-induced apoptosis was associated with the induction of autophagic response, as confirmed by the conversion of cytosolic LC3-I to autophagosome-associated LC3-II, increase in intracellular acidification, presence of autophagic vesicles, and increase in proteolysis of the selective autophagic target p62. HSV-1-triggered autophagy was not associated with the significant increase in the expression of proautophagic protein beclin-1 or downregulation of the major autophagy suppressor mammalian target of rapamycin (mTOR). Moreover, the phosphorylation of mTOR and its direct substrate p70 S6 kinase was augmented by HSV-1 infection, while the mTOR stimulator Akt and inhibitor AMPK-activated protein kinase (AMPK) were accordingly activated and suppressed, respectively. An shRNA-mediated knockdown of the autophagy-essential LC3β, as well as pharmacological inhibition of autophagy with bafilomycin A1 or 3-methyladenine, markedly accelerated apoptotic changes and ensuing cell death in HSV-1-infected glioma cells. These data indicate that AMPK/Akt/mTOR-independent autophagy could prolong survival of HSV-1-infected U251 glioma cells by counteracting the coinciding apoptotic response. © 2013 Institut Pasteur.

We investigated the role of autophagy, a stress-inducible lysosomal self-digestion of cellular components, in modulation of herpes simplex virus type 1 (HSV-1)-triggered death of U251 human glioma cells. HSV-1 caused apoptotic death in U251 cells, characterized by phosphatidylserine externalization, caspase activation and DNA fragmentation. HSV-1-induced apoptosis was associated with the induction of autophagic response, as confirmed by the conversion of cytosolic LC3-I to autophagosome-associated LC3-II, increase in intracellular acidification, presence of autophagic vesicles, and increase in proteolysis of the selective autophagic target p62. HSV-1-triggered autophagy was not associated with the significant increase in the expression of proautophagic protein beclin-1 or downregulation of the major autophagy suppressor mammalian target of rapamycin (mTOR). Moreover, the phosphorylation of mTOR and its direct substrate p70 S6 kinase was augmented by HSV-1 infection, while the mTOR stimulator Akt and inhibitor AMPK-activated protein kinase (AMPK) were accordingly activated and suppressed, respectively. An shRNA-mediated knockdown of the autophagy-essential LC3β, as well as pharmacological inhibition of autophagy with bafilomycin A1 or 3-methyladenine, markedly accelerated apoptotic changes and ensuing cell death in HSV-1-infected glioma cells. These data indicate that AMPK/Akt/mTOR-independent autophagy could prolong survival of HSV-1-infected U251 glioma cells by counteracting the coinciding apoptotic response. © 2013 Institut Pasteur.