Inhibition of Autophagy by Captopril Attenuates Prion Peptide-Mediated Neuronal Apoptosis via AMPK Activation.

Accumulation of prion protein (PrPc) into a protease-resistant form (PrPsc) in the brains of humans and animals affects the central nervous system. PrPsc occurs only in mammals with transmissible prion diseases. Prion protein refers to either the infectious pathogen itself or the main component of the pathogen. Recent studies suggest that autophagy is one of the major functions that keep cells alive and which has a protective effect against neurodegeneration. In this study, we investigated whether the anti-hypertensive drug, captopril, could attenuate prion peptide PrP (106-126)-induced calcium alteration-mediated neurotoxicity. Treatment with captopril increased both LC3-II (microtubule-associated protein 1A/1B-light chain 3-II) and p62 protein levels, indicating autophagy flux inhibition. Electron microscopy confirmed the occurrence of autophagic flux inhibition in neuronal cells treated with captopril. Captopril attenuated PrP (106-126)-induced neuronal cell death via AMPK activation and autophagy inhibition. Compound C suppressed AMPK activation as well as the neuroprotective effects of captopril. Thus, these data showed that an anti-hypertensive drug has a protective effect against prion-mediated neuronal cell death via autophagy inhibition and AMPK activation, and also suggest that anti-hypertensive drugs may be effective therapeutic agents against neurodegenerative disorders, including prion diseases.

Cellular prion protein regulates the differentiation and function of adipocytes through autophagy flux.

The role of autophagy modulation in adipogenic differentiation and the possible autophagy modulators targeting adipogenesis remain unclear. In this study, we investigated whether normal cellular prion protein (PrP) is involved in the modulation of autophagy and affects adipogenic differentiation in vivo and in vitro. Surprisingly, autophagy flux signals were activated in the adipose tissue of prion protein-deficient mice and PrP-deleted 3T3-L1 adipocytes. The activation of autophagy flux mediated by PrP deletion was confirmed in the adipose tissue via transmission electron microscopy. Adipocyte differentiation factors were highly induced in prion protein-deficient adipose tissue and 3T3-L1 adipocytes. In addition, deletion of prion protein significantly increased visceral fat volume, body fat weight, adipocyte cell size, and body weight gain in Prnp-knockout mice and increased lipid accumulation in PrP siRNA-transfected 3T3-L1 cells. However, the overexpression of prion protein using adenovirus inhibited the autophagic flux signals, lipid accumulation, and the PPAR-γ and C/EBP-α mRNA and protein expression levels in comparison to those in the control cells. Our results demonstrated that deletion of normal prion protein accelerated adipogenic differentiation and lipid accumulation mediated via autophagy flux activation.

AMPK Activation Mediated by Hinokitiol Inhibits Adipogenic Differentiation of Mesenchymal Stem Cells through Autophagy Flux.

BACKGROUND AND PURPOSE: Hinokitiol, a natural monopenoid present in the essential oil of Calocedrus formosana heartwood, exerts potent anticancer, anti-inflammatory, antibacterial, and neuroprotective effects on various cells. However, the antiobesity effect of hinokitiol on adipocytes is unclear. EXPERIMENTAL APPROACH: In this study, we observed that hinokitiol affected the differentiation to adipocytes in mesenchymal stem cells (MSCs). Hinokitiol was treated with 3-isobutyl-1-methylxanthine, insulin, and dexamethasone to induce differentiation and maturing adipocytes in cultured MSCs. KEY RESULTS: Hinokitiol treatment of MSCs decreased their differentiation to mature adipocytes and increased AMPK phosphorylation in a concentration-dependent manner. Moreover, we confirmed that the antiadipogenic effect of hinokitiol was associated with autophagy. The levels of LC3-II decreased and those of p62 increased in hinokitiol-treated MSCs. The treatment of hinokitiol-treated MSCs with the autophagy activator, rapamycin, restored the hinokitiol-induced decrease in the adipocyte differentiation of MSCs. The inhibition of AMPK phosphorylation also suppressed hinokitiol-mediated inhibition of autophagy and antiadipogenic effects. CONCLUSIONS AND IMPLICATIONS: Taken together, these results indicated that AMPK activation and autophagy flux inhibition mediated by hinokitiol inhibited lipid accumulation and differentiation of MSCs to adipocytes and also suggest that differentiation of mesenchymal stem cells may be regulated by using the modulator of autophagy flux and AMPK signals including hinokitiol.

Author Correction: Bisphosphonate enhances TRAIL sensitivity to human osteosarcoma cells via death receptor 5 upregulation.

After publication of this article, the authors noticed an error in the figure section.

Resveratrol sensitizes lung cancer cell to TRAIL by p53 independent and suppression of Akt/NF-κB signaling.

AIMS: TRAIL is a promising anticancer agent that has the potential to sensitize a wide variety of cancer or transformed cells by inducing apoptosis. However, resistance to TRAIL is a growing concern. Current manuscript aimed to employ combination treatment to investigate resveratrol induced TRAIL sensitization in NSCLC. METHOD: A549 and HCC-15 cells were used in an experimental design. Cell viability was determined by morphological image, crystal violet staining and MTT assay. Apoptosis was evaluated by LDH assay, Annexin V and DAPI staining. Autophagy and apoptosis indicator protein were examined by western blotting. TEM and puncta assay was carried out to evaluate the autophagy. MTP and ROS activity was evaluated by JC-1 and H2DCFDA staining. FINDINGS: Resveratrol is a polyphenolic compound capable of activation of tumor suppressor p53 and its pro-apoptotic modulator PUMA. Herein, we showed the p53-independent apoptosis by decrease the expression of phosphorylated Akt-mediated suppression of NF-κB that is also substantiated with the downregulation of anti-apoptotic factors Bcl-2 and Bcl-xl in NSCLC, resulting in an attenuation of TRAIL resistance in combined treatment. Furthermore, apoptosis was induced in TRAIL-resistant lung cancer cells with a co-treatment of resveratrol and TRAIL assessed by the loss of MMP, ROS generations which resulting the translocation of cytochrome c from the mitochondria into the cytosol due to mitochondrial dysfunction. Moreover, autophagy flux was not affected by resveratrol-induced TRAIL-mediated apoptosis in NSCLC. SIGNIFICANCE: Overall, targeting the NF-κB (p65) pathway via resveratrol attenuates TRAIL resistance and induces TRAIL-mediated apoptosis which could be the effective TRAIL-based cancer therapy regimen.

Ophiopogonin B sensitizes TRAIL-induced apoptosis through activation of autophagy flux and downregulates cellular FLICE-like inhibitory protein.

Tumor necrosis factor related apoptosis-inducing ligand (TRAIL), a type II transmembrane protein, belongs to the TNF superfamily. Compared to other family members, TRAIL is a promising anti-cancer agent that can selectively induce apoptosis of various types of transformed cells and xenografts, with negligible cytotoxicity against normal tissues. Ophiopogonin B is a bioactive ingredient of Radix Ophiopogon japonicus, which is frequently used in traditional Chinese medicine to treat cancer. In this study, we report that Cellular FLICE (FADD-like IL-1ß-converting enzyme)-inhibitory protein (c-FLIP) is the key determinant mediating TRAIL resistance in A549 cells and Ophiopogonin B downregulates c-FLIP and enhances TRAIL-induced apoptosis by activating autophagy flux. In addition, treatment with Ophiopogonin B resulted in a slight increase in the conversion of LC3-I to LC3-II and significantly decreased p62 expression levels in a dose-dependent manner. This indicates that Ophiopogonin B induces autophagy flux activation in human lung cancer cells. Inhibiting autophagy flux by applying a specific inhibitor ATG5 siRNA with Ophiopogonin B mediated enhancement of TRAIL effects. These data demonstrate that downregulation of c-FLIP by Ophiopogonin B enhances TRAIL-induced tumor cell death by activating autophagy flux in TRAIL-resistant A549 cells, and also suggests that Ophiopogonin B combined with TRAIL may be a successful therapeutic strategy for TRAIL-resistant lung cancer cells.

Autophagic flux induced by graphene oxide has a neuroprotective effect against human prion protein fragments.

Graphene oxide (GO) is a nanomaterial with newly developing biological applications. Autophagy is an intracellular degradation system that has been associated with the progression of neurodegenerative disorders. Although induction of autophagic flux by GO has been reported, the underlying signaling pathway in neurodegenerative disorders and how this is involved in neuroprotection remain obscure. We show that GO itself activates autophagic flux in neuronal cells and confers a neuroprotective effect against prion protein (PrP) (106-126)-mediated neurotoxicity. GO can be detected in SK-N-SH neuronal cells, where it triggers autophagic flux signaling. GO-induced autophagic flux prevented PrP (106-126)-induced neurotoxicity in SK-N-SH cells. Moreover, inactivation of autophagic flux blocked GO-induced neuroprotection against prion-mediated mitochondrial neurotoxicity. This is the first study to demonstrate that GO regulates autophagic flux in neuronal cells, and that activation of autophagic flux signals, induced by GO, plays a neuroprotective role against prion-mediated mitochondrial neurotoxicity. These results suggest that the nanomaterial GO may be used to activate autophagic flux and could be used in neuroprotective strategies for treatment of neurodegenerative disorders, including prion diseases.

Hypoxia-mediated autophagic flux inhibits silver nanoparticle-triggered apoptosis in human lung cancer cells.

Solid tumors are frequently associated with resistance to chemotherapy because the fraction of hypoxic tumor cells is substantial. To understand the underlying mechanism of hypoxia on silver nanoparticle (AgNPs)-induced apoptosis, the expression of hypoxia-inducible factor (HIF)-1α, a hallmark of hypoxia, was measured in the presence and absence of AgNPs. The results showed that HIF-1α expression was upregulated after AgNPs treatment under both hypoxic and normoxic conditions. Cell viability assays showed that AgNPs promoted cell death in cancer cells but not in non-cancer cells, as cancer cells are slightly more acidic than normal cells. However, reactive oxygen species generation induced by AgNPs in lung cancer cells caused high susceptibility to oxidative stress, whereas pre-exposure to hypoxia blocked AgNPs-induced oxidative stress. Notably, HIF-1α inhibited AgNPs-induced mitochondria-mediated apoptosis by regulating autophagic flux through the regulation of ATG5, LC3-II, and p62. Further, cell viability after treatment of cancer cells with AgNPs under hypoxic conditions was lower in HIF-1α siRNA-transfected cells than in control siRNA-transfected cells, indicating that HIF-1α knockdown enhances hypoxia induced decrease in cell viability. Our results suggest that hypoxia-mediated autophagy may be a mechanism for the resistance of AgNPs-induced apoptosis and that strategies targeting HIF-1α may be used for cancer therapy.

Male- and female-derived somatic and germ cell-specific toxicity of silver nanoparticles in mouse.

Silver nanoparticles (AgNPs) are widely used as an antibiotic agent in textiles, wound dressings, medical devices, and appliances such as refrigerators and washing machines. The increasing use of AgNPs has raised concerns about their potential risks to human health. Therefore, this study was aimed to determine the impact of AgNPs in germ cell specific complications in mice. The administration of AgNPs results in toxicity in mice; however, a more detailed understanding of the effects of AgNPs on germ cells remains poorly understood. Here, we demonstrate the effects of AgNPs (20 nm in diameter) in a mouse Sertoli and granulosa cells in vitro, and in male and female mice in vivo. Soluble silver ion (Ag(+))-treated cells were used as a positive control. We found that excessive AgNP-treated cells exhibited cytotoxicity, the formation of autophagosomes and autolysosomes in Sertoli cells. Furthermore, an increase in mitochondrial-mediated apoptosis by cytochrome c release from mitochondria due to translocation of Bax to mitochondria was observed. In in vivo studies, the expression of pro-inflammatory cytokines, including tumor necrosis factor α, interferon-γ, -6, -1ß, and monocyte chemoattractant protein-1 were significantly increased (p < 0.05). Histopathological analysis of AgNP-treated mice shows that a significant loss of male and female germ cells. Taken together, these data suggest that AgNPs with an average size of 20 nm have negative impact on the reproduction.

Modulation of the expression of sphingosine 1-phosphate 2 receptors regulates the differentiation of pre-adipocytes.

Sphingosine 1-phosphate (S1P) is a bioactive lipid mediator that regulates multiple signals through S1P receptors responsible for biological responses. In particular, the S1P2 receptor has distinct roles in the S1P­mediated differentiation of certain cell types. The present study was the first, to the best of our knowledge, to report the role of the S1P2 receptor in the adipocyte differentiation of 3T3­L1 pre­adipocytes. In order to investigate the influence of S1P2 receptors in the anti­adipogenic effects of S1P, S1P2 receptor silencing and overexpression of were used. S1P2 overexpression with adenoviral vectors inhibited adipogenesis and inhibited the expression of peroxisome proliferator­activated receptor γ (PPARγ), adiponectin and CCAAT/enhancer binding protein­α, which were upregulated following incubation in differentiation media. Furthermore, S1P completely lost its ability to impair adipogenic differentiation following silencing of S1P2. Silencing of the S1P2 receptor additionally blocked the downregulation of PPARγ protein and phospho­c­Jun N­terminal kinase protein induced by S1P treatment. In conclusion, the present study demonstrated that the S1P2 receptor is a key signaling molecule in the S1P­dependent inhibition of adipogenic differentiation and additionally suggested that selective targeting of S1P2 receptors may have clinical applications for the treatment of obesity.

Neuroprotective effect of cellular prion protein (PrPC) is related with activation of alpha7 nicotinic acetylcholine receptor (α7nAchR)-mediated autophagy flux.

Activation of the alpha7 nicotinic acetylcholine receptor (α7nAchR) is regulated by prion protein (PrPC) expression and has a neuroprotective effect by modulating autophagic flux. In this study, we hypothesized that PrPC may regulate α7nAchR activation and that may prevent prion-related neurodegenerative diseases by regulating autophagic flux. PrP(106-126) treatment decreased α7nAchR expression and activation of autophagic flux. In addition, the α7nAchR activator PNU-282987 enhanced autophagic flux and protected neuron cells against PrP(106-126)-induced apoptosis. However, activation of autophagy and the protective effects of PNU-282987 were inhibited in PrPC knockout hippocampal neuron cells. In addition, PrPC knockout hippocampal neuron cells showed decreased α7nAchR expression levels. Adenoviral overexpression of PrPC in PrPC knockout hippocampal neuron cells resulted in activation of autophagic flux and inhibition of prion peptide-mediated cell death via α7nAchR activation. This is the first report demonstrating that activation of α7nAchR-mediated autophagic flux is regulated by PrPC, and that activation of α7nAchR regulated by PrPC expression may play a pivotal role in protection of neuron cells against prion peptide-induced neuron cell death by autophagy. These results suggest that α7nAchR-mediated autophagic flux may be involved in the pathogenesis of prion-related diseases and may be a therapeutic target for prion-related neurodegenerative diseases.

EGCG-mediated autophagy flux has a neuroprotection effect via a class III histone deacetylase in primary neuron cells.

Prion diseases caused by aggregated misfolded prion protein (PrP) are transmissible neurodegenerative disorders that occur in both humans and animals. Epigallocatechin-3-gallate (EGCG) has preventive effects on prion disease; however, the mechanisms related to preventing prion diseases are unclear. We investigated whether EGCG, the main polyphenol in green tea, prevents neuron cell damage induced by the human prion protein. We also studied the neuroprotective mechanisms and proper signals mediated by EGCG. The results showed that EGCG protects the neuronal cells against human prion protein-induced damage through inhibiting Bax and cytochrome c translocation and autophagic pathways by increasing LC3-II and reducing and blocking p62 by using ATG5 small interfering (si) RNA and autophagy inhibitors. We further demonstrated that the neuroprotective effects of EGCG were exhibited by a class III histone deacetylase; sirt1 activation and the neuroprotective effects attenuated by sirt1 inactivation using sirt1 siRNA and sirtinol. We demonstrated that EGCG activated the autophagic pathways by inducing sirt1, and had protective effects against human prion protein-induced neuronal cell toxicity. These results suggest that EGCG may be a therapeutic agent for treatment of neurodegenerative disorders including prion diseases.

Multidimensional effects of biologically synthesized silver nanoparticles in Helicobacter pylori, Helicobacter felis, and human lung (L132) and lung carcinoma A549 cells.

Silver nanoparticles (AgNPs) are prominent group of nanomaterials and are recognized for their diverse applications in various health sectors. This study aimed to synthesize the AgNPs using the leaf extract of Artemisia princeps as a bio-reductant. Furthermore, we evaluated the multidimensional effect of the biologically synthesized AgNPs in Helicobacter pylori, Helicobacter felis, and human lung (L132) and lung carcinoma (A549) cells. UV-visible (UV-vis) spectroscopy confirmed the synthesis of AgNPs. X-ray diffraction (XRD) indicated that the AgNPs are specifically indexed to a crystal structure. The results from Fourier transform infrared spectroscopy (FTIR) indicate that biomolecules are involved in the synthesis and stabilization of AgNPs. Dynamic light scattering (DLS) studies showed the average size distribution of the particle between 10 and 40 nm, and transmission electron microscopy (TEM) confirmed that the AgNPs were significantly well separated and spherical with an average size of 20 nm. AgNPs caused dose-dependent decrease in cell viability and biofilm formation and increase in reactive oxygen species (ROS) generation and DNA fragmentation in H. pylori and H. felis. Furthermore, AgNPs induced mitochondrial-mediated apoptosis in A549 cells; conversely, AgNPs had no significant effects on L132 cells. The results from this study suggest that AgNPs could cause cell-specific apoptosis in mammalian cells. Our findings demonstrate that this environmentally friendly method for the synthesis of AgNPs and that the prepared AgNPs have multidimensional effects such as anti-bacterial and anti-biofilm activity against H. pylori and H. felis and also cytotoxic effects against human cancer cells. This report describes comprehensively the effects of AgNPs on bacteria and mammalian cells. We believe that biologically synthesized AgNPs will open a new avenue towards various biotechnological and biomedical applications in the near future.

Melatonin regulates the autophagic flux via activation of alpha-7 nicotinic acetylcholine receptors.

Our previous study suggested that melatonin-mediated neuroprotective effects are related with the activation of autophagy. However, the mechanism of melatonin-mediated autophagic activation in prion-mediated mitochondrial damage is not reported. Alpha-7 nicotinic acetylcholine receptors (α7nAchR) is a member of nicotinic acetylcholine receptors, and α7nAchR activation regulates via melatonin. Thus, we hypothesized that melatonin-mediated neuroprotective effect related with to autophagy pathway as a result of α7nAchR regulation. Inactivation of α7nAchR inhibited melatonin-mediated autophagic activation and protective effect against prion-mediated mitochondrial neurotoxicity. Also, knockdown of ATG5 blocked the melatonin-mediated neuroprotection and did not influence to the activation of α7nAchR caused by melatonin. This report is the first study demonstrating that melatonin-mediated autophagic activation regulates via modulation of α7nAchR signals, and upregulation of α7nAchR signals induced by melatonin plays a pivotal role in neuroprotection of prion-mediated mitochondrial neurotoxicity. Our results suggested that regulator of α7 nAChR signals including melatonin may have used for neuroprotective strategies for the neurodegenerative disorders including prion diseases.

Inhibition of the autophagy flux by gingerol enhances TRAIL-induced tumor cell death.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a primary anticancer agent and a member of the tumor necrosis factor family that selectively induces apoptosis in various tumor cells, but not in normal cells. Gingerol is a major ginger component with anti-inflammatory and anti­tumorigenic activities. Autophagy flux is the complete process of autophagy, in which the autophagosomes are lysed by lysosomes. The role of autophagy in cell death or cell survival is controversial. A549 adenocarcinoma cells are TRAIL-resistant. In the present study, we showed that treatment with TRAIL slightly induced cell death, but gingerol treatment enhanced the TRAIL-induced cell death in human lung cancer cells. The combination of gingerol and TRAIL increased accumulation of microtubule-associated protein light chain 3-II and p62, confirming the inhibited autophagy flux. Collectively, our results suggest that gingerol sensitizes human lung cancer cells to TRAIL-induced apoptosis by inhibiting the autophagy flux.

Induction of cellular prion protein (PrPc) under hypoxia inhibits apoptosis caused by TRAIL treatment.

Hypoxia decreases cytotoxic responses to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) protein. Cellular prion protein (PrPc) is regulated by HIF-1α in neurons. We hypothesized that PrPc is involved in hypoxia-mediated resistance to TRAIL-induced apoptosis. We found that hypoxia induced PrPc protein and inhibited TRAIL-induced apoptosis. Thus silencing of PrPc increased TRAIL-induced apoptosis under hypoxia. Overexpression of PrPc protein using an adenoviral vector inhibited TRAIL-induced apoptosis. In xenograft model in vivo, shPrPc transfected cells were more sensitive to TRAIL-induced apoptosis than in shMock transfected cells. Molecular chemo-therapy approaches based on the regulation of PrPc expression need to address anti-tumor function of TRAIL under hypoxia. Molecular chemo-therapy approaches based on the regulation of PrPc expression need to address anti-tumor function of TRAIL under hypoxia.

Activation of S1P2 receptor, a possible mechanism of inhibition of adipogenic differentiation by sphingosine 1­phosphate.

Sphingosine 1­phosphate (S1P) belongs to a significant group of signaling sphingolipids and exerts most of its activity as a ligand of G­protein­coupled receptors. In our previous study, S1P demonstrated a novel biological activity with the anti­adipogenesis of 3T3­L1 preadipocytes. In the present study, we identified a possible mechanism of S1P­mediated anti­adipogenic effects, particularly in target pathways of the S1P receptors, including S1P1 and S1P2. The mRNA levels of S1P1 and S1P2 receptors were increased by MDI media treatment, whereas S1P treatment highly induced S1P2 but not S1P1 receptor protein in adipocytes. Triglyceride accumulation assay using an agonist and antagonist of S1P receptors revealed that S1P2 receptor was only involved in S1P­mediated anti­adipogenic effects. Furthermore, pharmacological inhibition of S1P2 signals completely retrieved S1P­mediated downregulation of the transcriptional levels of peroxisome proliferator­activated receptor γ, CCAAT/enhancer binding protein α and adiponectin, which are markers of adipogenic differentiation. This study demonstrated that S1P2 receptor signals may regulate the S1P­mediated anti­adipogenic differentiation and also identifies the S1P2 receptor as a possible mechanism of anti­adipogenic differentiation.

Deferoxamine inhibits TRAIL-mediated apoptosis via regulation of autophagy in human colon cancer cells.

Deferoxamine (DFO), an iron chelator, has numerous clinical applications for patients presenting with iron overload in regards to the improvement in the quality of life and overall survival. In addition, experimental iron chelators have demonstrated potent anticancer properties. The present study investigated the effects of DFO on TNF-related apoptosis-inducing ligand (TRAIL)-induced apoptosis in colon cancer cells and and the mechanism involved. The experimental results showed that DFO treatment inhibited TRAIL-mediated cancer cell apoptosis by increasing Akt activation and decreasing caspase activation in human colon cancer cells. Furthermore, DFO treatment induced autophagy flux, and chloroquine, an autophagy inhibitor, blocked DFO-mediated inhibition of TRAIL-induced apoptosis. The present study demonstrated that DFO inhibited TRAIL-mediated tumor cell death via the autophagy pathway, and the results suggest that potent anticancer agent, DFO, can be an inhibitor against antitumor therapy including TRAIL protein.

Pifithrin-α ameliorates resveratrol-induced two-cell block in mouse preimplantation embryos in vitro.

Treatment with resveratrol at concentrations greater than 0.5 µmol/L resulted in the arrest of mouse embryo development at the two-cell stage. Resveratrol-induced cytotoxicity was investigated in embryos by evaluating morphologic features by using the bromodeoxyuridine assay and acridine orange and ethidium bromide double staining. Resveratrol was found to significantly increase the expressions of p53, p21, Atf3, smac/Diablo, Bax, Bak1, Bok, and Noxa mRNA in the embryos, whereas Cullin 3 and Cdk1 expressions were decreased. Furthermore, active p53 positive signal in embryos arrested at the two-cell stage was localized in the nucleus, whereas no active p53 signal was observed in control embryos. Pretreatment with pifithrin-α, a p53 inhibitor, downregulated active p53 in two-cell embryo nuclei and ameliorated approximately 50% of the embryonic developmental defect caused by resveratrol. The findings of the present study, therefore, suggest that pifithrin-α could be used as an effective cytoprotective agent against a reproductive toxin such as resveratrol.

Oxidative stress mediated cytotoxicity of biologically synthesized silver nanoparticles in human lung epithelial adenocarcinoma cell line.

The goal of the present study was to investigate the toxicity of biologically prepared small size of silver nanoparticles in human lung epithelial adenocarcinoma cells A549. Herein, we describe a facile method for the synthesis of silver nanoparticles by treating the supernatant from a culture of Escherichia coli with silver nitrate. The formation of silver nanoparticles was characterized using various analytical techniques. The results from UV-visible (UV-vis) spectroscopy and X-ray diffraction analysis show a characteristic strong resonance centered at 420 nm and a single crystalline nature, respectively. Fourier transform infrared spectroscopy confirmed the possible bio-molecules responsible for the reduction of silver from silver nitrate into nanoparticles. The particle size analyzer and transmission electron microscopy results suggest that silver nanoparticles are spherical in shape with an average diameter of 15 nm. The results derived from in vitro studies showed a concentration-dependent decrease in cell viability when A549 cells were exposed to silver nanoparticles. This decrease in cell viability corresponded to increased leakage of lactate dehydrogenase (LDH), increased intracellular reactive oxygen species generation (ROS), and decreased mitochondrial transmembrane potential (MTP). Furthermore, uptake and intracellular localization of silver nanoparticles were observed and were accompanied by accumulation of autophagosomes and autolysosomes in A549 cells. The results indicate that silver nanoparticles play a significant role in apoptosis. Interestingly, biologically synthesized silver nanoparticles showed more potent cytotoxicity at the concentrations tested compared to that shown by chemically synthesized silver nanoparticles. Therefore, our results demonstrated that human lung epithelial A549 cells could provide a valuable model to assess the cytotoxicity of silver nanoparticles.