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  1. Baicalein pretreatment reduces liver ischemia/reperfusion injury via induction of autophagy in rats.

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    Liu, Anding; Huang, Liang; Guo, Enshuang; Li, Renlong; Yang, Jiankun; Li, Anyi; Yang, Yan; Liu, Shenpei; Hu, Jifa; Jiang, Xiaojing; Dirsch, Olaf; Dahmen, Uta; Sun, Jian

    2016-01-01

    We previously demonstrated that baicalein could protect against liver ischemia/reperfusion (I/R) injury in mice. The exact mechanism of baicalein remains poorly understood. Autophagy plays an important role in protecting against I/R injury. This study was designed to determine whether baicalein could protect against liver I/R injury via induction of autophagy in rats. Baicalein was intraperitoneally injected 1 h before warm ischemia. Pretreatment with baicalein prior to I/R insult significantly blunted I/R-induced elevations of serum aminotransferase levels and significantly improved the histological status of livers. Electron microscopy and expression of the autophagic marker LC3B-II suggested induction of autophagy after baicalein treatment. Moreover, inhibition of the baicalein-induced autophagy using 3-methyladenine (3-MA) worsened liver injury. Furthermore, baicalein treatment increased heme oxygenase (HO)-1 expression, and pharmacological inhibition of HO-1 with tin protoporphyrin IX (SnPP) abolished the baicalein-mediated autophagy and the hepatocellular protection. In primary rat hepatocytes, baicalein-induced autophagy also protected hepatocytes from hypoxia/reoxygenation injury in vitro and the beneficial effect was abrogated by 3-MA or Atg7 siRNA, respectively. Suppression of HO-1 activity by SnPP or HO-1 siRNA prevented the baicalein-mediated autophagy and resulted in increased hepatocellular injury. Collectively, these results suggest that baicalein prevents hepatocellular injury via induction of HO-1-mediated autophagy. PMID:27150843

  2. Induction of autophagy improves embryo viability in cloned mouse embryos

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    Shen, XingHui; Zhang, Na; Wang, ZhenDong; Bai, GuangYu; Zheng, Zhong; Gu, YanLi; Wu, YanShuang; Liu, Hui; Zhou, DongJie; Lei, Lei

    2015-01-01

    Autophagy is an essential cellular mechanism that degrades cytoplasmic proteins and organelles to recycle their components. Moreover, autophagy is essential for preimplantation development in mammals. Here we show that autophagy is also important for reprogramming in somatic cell nuclear transfer (SCNT). Our data indicate that unlike fertilized oocytes, autophagy is not triggered in SCNT embryos during 6 hours of activation. Mechanistically, the inhibited autophagic induction during SCNT activation is due to the cytochalasin B (CB) caused depolymerization of actin filaments. In this study, we induced autophagy during SCNT activation by rapamycin and pp242, which could restore the expected level of autophagy and significantly enhance the development of SCNT embryos to the blastocyst stage when compared with the control (68.5% and 68.7% vs. 41.5%, P autophagy is important for development of SCNT embryos and inhibited autophagic induction during SCNT activation might be one of the serious causes of low efficiency of SCNT. PMID:26643778

  3. Induction of cytoprotective autophagy in PC-12 cells by cadmium

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    Wang, Qiwen [College of Veterinary Medicine, Yangzhou University, Yangzhou 225009 (China); Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou 225009 (China); Bijie Pilot Area Research Institute of Bijie University, Bijie 551700 (China); Zhu, Jiaqiao; Zhang, Kangbao; Jiang, Chenyang; Wang, Yi; Yuan, Yan; Bian, Jianchun; Liu, Xuezhong; Gu, Jianhong [College of Veterinary Medicine, Yangzhou University, Yangzhou 225009 (China); Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou 225009 (China); Liu, Zongping, E-mail: liuzongping@yzu.edu.cn [College of Veterinary Medicine, Yangzhou University, Yangzhou 225009 (China); Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou 225009 (China)

    2013-08-16

    Highlights: •Cadmium can promote early upregulation of autophagy in PC-12 cells. •Autophagy precedes apoptosis in cadmium-treated PC-12 cells. •Cadmium-induced autophagy is cytoprotective in PC-12 cells. •Class III PI3K/beclin-1/Bcl-2 signaling pathway plays a positive role in cadmium-triggered autophagy. -- Abstract: Laboratory data have demonstrated that cadmium (Cd) may induce neuronal apoptosis. However, little is known about the role of autophagy in neurons. In this study, cell viability decreased in a dose- and time-dependent manner after treatment with Cd in PC-12 cells. As cells were exposed to Cd, the levels of LC3-II proteins became elevated, specific punctate distribution of endogenous LC3-II increased, and numerous autophagosomes appeared, which suggest that Cd induced a high level of autophagy. In the late stages of autophagy, an increase in the apoptosis ratio was observed. Likewise, pre-treatment with chloroquine (an autophagic inhibitor) and rapamycin (an autophagic inducer) resulted in an increased and decreased percentage of apoptosis in contrast to other Cd-treated groups, respectively. The results indicate that autophagy delayed apoptosis in Cd-treated PC-12 cells. Furthermore, co-treatment of cells with chloroquine reduced autophagy and cell activity. However, rapamycin had an opposite effect on autophagy and cell activity. Moreover, class III PI3 K/beclin-1/Bcl-2 signaling pathways served a function in Cd-induced autophagy. The findings suggest that Cd can induce cytoprotective autophagy by activating class III PI3 K/beclin-1/Bcl-2 signaling pathways. In sum, this study strongly suggests that autophagy may serve a positive function in the reduction of Cd-induced cytotoxicity.

  4. Induction of cytoprotective autophagy in PC-12 cells by cadmium

    International Nuclear Information System (INIS)

    Highlights: •Cadmium can promote early upregulation of autophagy in PC-12 cells. •Autophagy precedes apoptosis in cadmium-treated PC-12 cells. •Cadmium-induced autophagy is cytoprotective in PC-12 cells. •Class III PI3K/beclin-1/Bcl-2 signaling pathway plays a positive role in cadmium-triggered autophagy. -- Abstract: Laboratory data have demonstrated that cadmium (Cd) may induce neuronal apoptosis. However, little is known about the role of autophagy in neurons. In this study, cell viability decreased in a dose- and time-dependent manner after treatment with Cd in PC-12 cells. As cells were exposed to Cd, the levels of LC3-II proteins became elevated, specific punctate distribution of endogenous LC3-II increased, and numerous autophagosomes appeared, which suggest that Cd induced a high level of autophagy. In the late stages of autophagy, an increase in the apoptosis ratio was observed. Likewise, pre-treatment with chloroquine (an autophagic inhibitor) and rapamycin (an autophagic inducer) resulted in an increased and decreased percentage of apoptosis in contrast to other Cd-treated groups, respectively. The results indicate that autophagy delayed apoptosis in Cd-treated PC-12 cells. Furthermore, co-treatment of cells with chloroquine reduced autophagy and cell activity. However, rapamycin had an opposite effect on autophagy and cell activity. Moreover, class III PI3 K/beclin-1/Bcl-2 signaling pathways served a function in Cd-induced autophagy. The findings suggest that Cd can induce cytoprotective autophagy by activating class III PI3 K/beclin-1/Bcl-2 signaling pathways. In sum, this study strongly suggests that autophagy may serve a positive function in the reduction of Cd-induced cytotoxicity

  5. Induction of autophagy by Imatinib sequesters Bcr-Abl in autophagosomes and down-regulates Bcr-Abl protein.

    LENUS (Irish Health Repository)

    Elzinga, Baukje M

    2013-06-01

    Chronic Myeloid Leukemia (CML) is a disease of hematopoietic stem cells which harbor the chimeric gene Bcr-Abl. Expression levels of this constitutively active tyrosine kinase are critical for response to tyrosine kinase inhibitor treatment and also disease progression, yet the regulation of protein stability is poorly understood. We have previously demonstrated that imatinib can induce autophagy in Bcr-Abl expressing cells. Autophagy has been associated with the clearance of large macromolecular signaling complexes and abnormal proteins, however, the contribution of autophagy to the turnover of Bcr-Abl protein in imatinib treated cells is unknown. In this study, we show that following imatinib treatment, Bcr-Abl is sequestered into vesicular structures that co-localize with the autophagy marker LC3 or GABARAP. This association is inhibited by siRNA mediated knockdown of autophagy regulators (Beclin 1\\/ATG7). Pharmacological inhibition of autophagy also reduced Bcr-Abl\\/LC3 co-localization in both K562 and CML patient cells. Bcr-Abl protein expression was reduced with imatinib treatment. Inhibition of both autophagy and proteasome activity in imatinib treated cells was required to restore Bcr-Abl protein levels to those of untreated cells. This ability to down-regulate Bcr-Abl protein levels through the induction of autophagy may be an additional and important feature of the activity of imatinib.

  6. Ghrelin Attenuated Lipotoxicity via Autophagy Induction and Nuclear Factor-κB Inhibition

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    Yuqing Mao

    2015-09-01

    Full Text Available Background/Aims: Nonalcoholic fatty liver disease (NAFLD is the most common chronic liver disease worldwide. Autophagy is associated with NAFLD. Ghrelin is a gut hormone with various functions including energy metabolism and inflammation inhibition. We investigated the therapeutic effect of ghrelin on NAFLD and its association with autophagy. Methods: C57bl/6 mice were fed a high-fat diet for 8 weeks to induce a model of chronic NAFLD, with ghrelin (10 µg/kg administrated subcutaneously twice weekly from weeks 6 to 8. LO2 cells were pretreated with ghrelin (10-8 M before stimulation with free fatty acid (palmitic and oleic acids; 1 mM. Lipid droplets were identified by hematoxylin and eosin and Red O staining and quantified by triglyceride test kits. LC3I/II, an important biomarker protein of autophagy was detected by western blotting, real-time polymerase chain reaction, immunohistochemistry and immunofluorescence. Tumor necrosis factor (TNF-a and interleukin (IL-6 were detected by ELISA and immunohistochemistry. Nuclear factor (NF-κB p65 was detected by western blotting and immunofluorescence. AMP-activated protein kinase (AMPK and mammalian target of rapamycin (mTOR were detected by western blotting. Results: Ghrelin reduced the triglyceride content in high fat diet (HFD group in vivo and free fatty acid (FFA group in vitro. TNF-a and IL-6 were significantly reduced in the ghrelin-treated mice compared with the control group. Autophagy induction was accompanied with intracellular lipid reduction in ghrelin-treated mice. Ghrelin upregulated autophagy via AMPK/mTOR restoration and inhibited translocation of NF-κB into the nucleus. Conclusions: The results indicate that ghrelin attenuates lipotoxicity by autophagy stimulation and NF-κB inhibition.

  7. Autophagy Induction Protects Against 7-Oxysterol-induced Cell Death via Lysosomal Pathway and Oxidative Stress

    OpenAIRE

    Xi-Ming Yuan; Nargis Sultana; Nabeel Siraj; Ward, Liam J.; Bijar Ghafouri; Wei Li

    2016-01-01

    7-Oxysterols are major toxic components in oxidized low-density lipoprotein and human atheroma lesions, which cause lysosomal membrane permeabilization (LMP) and cell death. Autophagy may function as a survival mechanism in this process. Here, we investigated whether 7-oxysterols mixed in an atheroma-relevant proportion induce autophagy, whether autophagy induction influences 7-oxysterol-mediated cell death, and the underlying mechanisms, by focusing on cellular lipid levels, oxidative stress...

  8. There is more to autophagy than induction: regulating the roller coaster.

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    Klionsky, Daniel J; Nemchenko, Andriy

    2011-08-01

    Considerable attention has been paid to the topic of autophagy induction. In part, this is because of the potential for modulating this process for therapeutic purposes. Of course we know that induced autophagy can also be problematic--for example, when trying to eliminate an established tumor that might be relying on autophagy for its own cytoprotective uses. Accordingly, inhibitory mechanisms have been considered; however, the corresponding studies have tended to focus on the pathways that block autophagy under non-inducing conditions, such as when nutrients are available. In contrast, relatively little is known about the mechanisms for inhibiting autophagy under inducing conditions. Yet, this type of regulation must be occurring on a routine basis. We know that dysregulation of autophagy, e.g., due to improper activation of Beclin 1 leading to excessive autophagy activity, can cause cell death. Accordingly, we assume that during starvation or other inducing conditions there must be a mechanism to modulate autophagy. That is, once you turn it on, you do not want to let it continue unchecked. But how is autophagy downregulated when the inducing conditions still exist? PMID:21636971

  9. Transforming growth factor-β1 reduces apoptosis via autophagy activation in hepatic stellate cells

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    FU, MEI-YA; HE, YA-JUN; LV, XIA; LIU, ZHI-HE; SHEN, YAN; YE, GUO-RONG; DENG, YAN-MEI; SHU, JIAN-CHANG

    2014-01-01

    Autophagy is a metabolic process that is important in fibrogenesis, in which cellular components are degraded by lysosomal machinery. Transforming growth factor β1 (TGF-β1) is a potent fibrogenic cytokine involved in liver fibrosis; however, it remains elusive whether autophagy is regulated by TGF-β1 in this process. In the present study, the function of TGF-β1-mediated autophagy in the proliferation and apoptosis of hepatic stellate cells (HSCs) was investigated. A rat HSC cell line (HSC-T6) was incubated with or without TGF-β1 followed by bafilomycin A1, and microtubule-associated proteins 1A/1B light chain 3 (LC3) small interfering (si)RNA was used to inhibit autophagy in order to assess the association between TGF-β1 and autophagy. HSC-T6 cell transient transfection was accomplished with a pLVX-AcGFP-N1-rLC3B-encoding plasmid. An MTS assay and flow cytometry were utilized to detect proliferation and apoptosis of HSC-T6 cells. Quantitative polymerase chain reaction, immunofluorescence and western blot analysis were used to detect the presence of activation markers. Proliferation was increased and apoptosis was reduced in HSC-T6 cells treated with TGF-β1 compared with cells subjected to serum deprivation. However, when HSC-T6 cells were treated with bafilomycin A1 and LC3 siRNA, increased apoptosis and reduced proliferation were observed. In addition, protein and mRNA expression levels of the autophagy marker LC3 were significantly increased. GFP-LC3 punctate markings were more prolific following TGF-β1 treatment of HSC-T6 cells, indicating that TGF-β1 may rescue HSC-T6 cells from serum deprivation and reduce apoptosis via autophagy induction. The present study elucidated the possible functions of TGF-β1-mediated autophagy in the pathological process of liver fibrosis. PMID:25059289

  10. Autophagy induction upon reactive oxygen species in Cd-stressed Arabidopsis thaliana

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    Zhang, WeiNa; Chen, WenLi

    2010-02-01

    Autophagy is a protein degradation process in which cells recycle cytoplasmic contents when subjected to environmental stress conditions or during certain stages of development. Upon the induction of autophagy, a double membrane autophagosome forms around cytoplasmic components and delivers them to the vacuole for degradation. In plants, autophagy has been shown previously to be induced during abiotic stresses including oxidative stress. Cd, as a toxicity heavy metal, resulted in the production of reactive oxygen species (ROS). In this paper, we demonstrated that ROS contributed to the induction of autophagy in Cd-stressed Arabidopsis thaliana. However, pre-incubation with ascorbic acid (AsA, antioxidant molecule) and catalase (CAT, a H2O2-specific scavenger) decreased the ROS production and the number of autolysosomal-like structures. Together our results indicated that the oxidative condition was essential for autophagy, as treatment with AsA and CAT abolished the formation of autophagosomes, and ROS may function as signal molecules to induce autophagy in abiotic stress.

  11. Autophagy induction promotes aristolochic acid-I-induced renal injury in vivo and in vitro

    International Nuclear Information System (INIS)

    Graphical abstract: - Highlights: • Aristolochic acid induced autophagy in vivo and in vitro. • Autophagy induced by aristolochic acid could promote cell apoptosis. • Inhibition autophagy by silencing ATG5 could prevent cell from programmed cell death induced by aristolochic acid. - Abstract: Studies have found that ingestion of aristolochic acid (AA) causes nephropathy first by inducing renal tubular cell apoptosis acutely. It is currently unknown whether crosstalk between autophagy and apoptosis orchestrates the fate of tubular cells in acute AA nephropathy. We tested this hypothesis by acute administration of AA in vivo and in vitro. Autophagy was first induced in vivo through enhancing Atg5 and LC3-II expressions in kidneys of AA-I-treated rats. Punctuate LC3-GFP dots and autophagosomes were detected in this acute AA-I nephropathy rat model. We subsequently utilized normal rat renal proximal tubular epithelial cells (NRK52E) to study the autophagy mechanisms involved in acute AA-I nephropathy, with 100 μM AA-I (median lethal dose 50) given in vitro. Cleavage of poly (ADP-ribose) polymerase (PARP), nuclear condensation, and fragmentation were demonstrated in the AA-I-treated NRK52E cells. Furthermore, AA-I induced Atg5 and LC3-II expressions and punctuated LC3-GFP dots. Autophagy flux by using lysosome inhibitor E64 induced the accumulation of LC3-II, which further promoted apoptosis through enhancing PARP cleavage. Inhibition of autophagy by 3-methyl adenine also led to the attenuation of AA-I-induced apoptosis, manifesting as decreased PARP cleavage, nuclei condensation, and decreased the number of cells negative for acridine orange/ethidium bromide staining. In addition, knockdown of Atg5 by short hairpin RNA attenuated LC3-II expression and PARP cleavage in NRK52E cells. Taken together, these findings suggested that the acute phase of AA-I-induced nephropathy is associated with induction of Atg5-dependent autophagy, which promotes renal tubular cell

  12. Nrf2 reduces levels of phosphorylated tau protein by inducing autophagy adaptor protein NDP52

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    Jo, Chulman; Gundemir, Soner; Pritchard, Susanne; Jin, Youngnam N.; Rahman, Irfan; Johnson, Gail V. W.

    2014-03-01

    Nuclear factor erythroid 2-related factor 2 (Nrf2) is a pivotal transcription factor in the defence against oxidative stress. Here we provide evidence that activation of the Nrf2 pathway reduces the levels of phosphorylated tau by induction of an autophagy adaptor protein NDP52 (also known as CALCOCO2) in neurons. The expression of NDP52, which we show has three antioxidant response elements (AREs) in its promoter region, is strongly induced by Nrf2, and its overexpression facilitates clearance of phosphorylated tau in the presence of an autophagy stimulator. In Nrf2-knockout mice, phosphorylated and sarkosyl-insoluble tau accumulates in the brains concurrent with decreased levels of NDP52. Moreover, NDP52 associates with phosphorylated tau from brain cortical samples of Alzheimer disease cases, and the amount of phosphorylated tau in sarkosyl-insoluble fractions is inversely proportional to that of NDP52. These results suggest that NDP52 plays a key role in autophagy-mediated degradation of phosphorylated tau in vivo.

  13. EGFR-independent autophagy induction with gefitinib and enhancement of its cytotoxic effect by targeting autophagy with clarithromycin in non-small cell lung cancer cells.

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    Sugita, Shohei; Ito, Kentaro; Yamashiro, Yutaro; Moriya, Shota; Che, Xiao-Fang; Yokoyama, Tomohisa; Hiramoto, Masaki; Miyazawa, Keisuke

    2015-05-22

    Gefitinib (GEF), an inhibitor for EGFR tyrosine kinase, potently induces autophagy in non-small cell lung cancer (NSCLC) cell lines such as PC-9 cells expressing constitutively activated EGFR kinase by EGFR gene mutation as well as A549 and H226 cells with wild-type EGFR. Unexpectedly, GEF-induced autophagy was also observed in non-NSCLC cells such as murine embryonic fibroblasts (MEF) and leukemia cell lines K562 and HL-60 without EGFR expression. Knockout of EGFR gene in A549 cells by CRISPR/Cas9 system still exhibited autophagy induction after treatment with GEF, indicating that the autophagy induction by GEF is not mediated through inhibiting EGFR kinase activity. Combined treatment with GEF and clarithromycin (CAM), a macrolide antibiotic having the effect of inhibiting autophagy flux, enhances the cytotoxic effect in NSCLC cell lines, although treatment with CAM alone exhibits no cytotoxicity. GEF treatment induced up-regulation of endoplasmic reticulum (ER)-stress related genes such as CHOP/GADD153 and GRP78. Knockdown of CHOP in PC-9 cells and Chop-knockout MEF both exhibited less sensitivity to GEF than controls. Addition of CAM in culture medium resulted in further pronounced GEF-induced ER stress loading, while CAM alone exhibited no effect. These data suggest that GEF-induced autophagy functions as cytoprotective and indicates the potential therapeutic possibility of using CAM for GEF therapy. Furthermore, it is suggested that the intracellular signaling for autophagy initiation in response to GEF can be completely dissociated from EGFR, but unknown target molecule(s) of GEF for autophagy induction might exist. PMID:25858318

  14. NF-κB p65 repression by the sesquiterpene lactone, Helenalin, contributes to the induction of autophagy cell death

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    Lim Chuan

    2012-07-01

    Full Text Available Abstract Background Numerous studies have demonstrated that autophagy plays a vital role in maintaining cellular homeostasis. Interestingly, several anticancer agents were found to exert their anticancer effects by triggering autophagy. Emerging data suggest that autophagy represents a novel mechanism that can be exploited for therapeutic benefit. Pharmacologically active natural compounds such as those from marine, terrestrial plants and animals represent a promising resource for novel anticancer drugs. There are several prominent examples from the past proving the success of natural products and derivatives exhibiting anticancer activity. Helenalin, a sesquiterpene lactone has been demonstrated to have potent anti-inflammatory and antitumor activity. Albeit previous studies demonstrating helenalin’s multi modal action on cellular proliferative and apoptosis, the mechanisms underlying its action are largely unexplained. Methods To deduce the mechanistic action of helenalin, cancer cells were treated with the drug at various concentrations and time intervals. Using western blot, FACS analysis, overexpression and knockdown studies, cellular signaling pathways were interrogated focusing on apoptosis and autophagy markers. Results We show here that helenalin induces sub-G1 arrest, apoptosis, caspase cleavage and increases the levels of the autophagic markers. Suppression of caspase cleavage by the pan caspase inhibitor, Z-VAD-fmk, suppressed induction of LC3-B and Atg12 and reduced autophagic cell death, indicating caspase activity was essential for autophagic cell death induced by helenalin. Additionally, helenalin suppressed NF-κB p65 expression in a dose and time dependent manner. Exogenous overexpression of p65 was accompanied by reduced levels of cell death whereas siRNA mediated suppression led to augmented levels of caspase cleavage, autophagic cell death markers and increased cell death. Conclusions Taken together, these results show

  15. Autophagy induction in tobacco leaves infected by potato virus Y(O) and its putative roles.

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    Choi, Dabin; Park, Jaeyoung; Oh, Seonhee; Cheong, Hyunsook

    2016-06-01

    Autophagy plays a critical role in the innate immune response of plants to pathogen infection. In the present study, we examined autophagy induced by potato virus Y ordinary strain (PVY(O)) infection in tobacco (Nicotiana benthamiana). Enzyme-linked immunosorbent assays revealed that the number of virus particles in the plant peaked at 2 weeks post-inoculation and then gradually decreased. Additionally, the amount of virus increased significantly in the 3rd and 4th leaves distal to the inoculated leaf and decreased slightly in the 5th leaf. Within 2 weeks of PVY(O) inoculation, the tobacco leaves showed typical symptoms of Potyvirus inoculation, including mottling, yellowing, a mosaic pattern, and necrotic tissue changes at the inoculated site. Based on an ultrastructural analysis of the PVY(O)-infected tobacco leaves, virus aggregates appeared as longitudinal and transverse arrays and pinwheels, which are typical of Potyvirus inoculation. Moreover, PVY(O) infection caused changes in the number, size, and shape of chloroplasts, whereas the number of plastogranules increased markedly. Furthermore, double-membrane autophagosome-like vacuoles, including electron-dense materials, laminated structures, and cellular organelles, were found. The induction of autophagy after the PVY(O) infection of tobacco leaves was further confirmed by the expression of lipidated microtubule-associated protein 1 light chain 3 (LC3)-II, an autophagy marker and p62, an autophagy adaptor protein. The LC3-II levels increased daily over the 4-week period. Although virus inoculation was performed systemically on the basal leaves of the plants, LC3-II was expressed throughout the leaves and the expression was higher in leaves distal to the inoculated leaf. Moreover, PVY(O) infection caused the activation of stress-activated protein kinases/c-Jun N-terminal kinases. Therefore, PVY(O) infection-induced autophagy was positively correlated with the virus content, suggesting that autophagy induction

  16. Suppression of mTOR pathway and induction of autophagy-dependent cell death by cabergoline.

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    Lin, Shao Jian; Leng, Zhi Gen; Guo, Yu Hang; Cai, Lin; Cai, Yu; Li, Ning; Shang, Han Bing; Le, Wei-Dong; Zhao, Wei Guo; Wu, Zhe Bao

    2015-11-17

    Cabergoline (CAB), the first-line drug for treatment of prolactinomas, is effective in suppressing prolactin hypersecretion, reducing tumor size, and restoring gonadal function. However, mechanisms for CAB-mediated tumor shrinkage are largely unknown. Here we report a novel cytotoxic mechanism for CAB. CAB induced formation of autophagosome in rat pituitary tumor MMQ and GH3 cells at the early stage through inhibiting mTOR pathway, resulting in higher conversion rates of LC3-I to LC3-II, GFP-LC3 aggregation, and increased autophagosome formation. Interestingly, CAB treatment augmented lysosome acidification and resulted in impaired proteolytic degradation within autolysosomes. This blocked the autophagic flux, leading to the accumulation of p62 aggregation and undigested autolysosomes. Knockdown of ATG7, ATG5, or Becn1, could significantly rescue the CAB-mediated cell death of MMQ cells (p < 0.05). CAB-induced autophagy and blockade of autophagy flux participated in antitumoral action in vivo. In conclusion, our study provides evidence that CAB concomitantly induces autophagy and inhibits the autophagic flux, leading to autophagy-dependent cell death. These findings elucidate novel mechanisms for CAB action. PMID:26513171

  17. Cilostazol Upregulates Autophagy via SIRT1 Activation: Reducing Amyloid-β Peptide and APP-CTFβ Levels in Neuronal Cells.

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    Hye Rin Lee

    Full Text Available Autophagy is a vital pathway for the removal of β-amyloid peptide (Aβ and the aggregated proteins that cause Alzheimer's disease (AD. We previously found that cilostazol induced SIRT1 expression and its activity in neuronal cells, and thus, we hypothesized that cilostazol might stimulate clearances of Aβ and C-terminal APP fragment β subunit (APP-CTFβ by up-regulating autophagy.When N2a cells were exposed to soluble Aβ1-42, protein levels of beclin-1, autophagy-related protein5 (Atg5, and SIRT1 decreased significantly. Pretreatment with cilostazol (10-30 μM or resveratrol (20 μM prevented these Aβ1-42 evoked suppressions. LC3-II (a marker of mammalian autophagy levels were significantly increased by cilostazol, and this increase was reduced by 3-methyladenine. To evoke endogenous Aβ overproduction, N2aSwe cells (N2a cells stably expressing human APP containing the Swedish mutation were cultured in medium with or without tetracycline (Tet+ for 48 h and then placed in Tet- condition. Aβ and APP-CTFβ expressions were increased after 12~24 h in Tet- condition, and these increased expressions were significantly reduced by pretreating cilostazol. Cilostazol-induced reductions in the expressions of Aβ and APP-CTFβ were blocked by bafilomycin A1 (a blocker of autophagosome to lysosome fusion. After knockdown of the SIRT1 gene (to ~40% in SIRT1 protein, cilostazol failed to elevate the expressions of beclin-1, Atg5, and LC3-II, indicating that cilostazol increases these expressions by up-regulating SIRT1. Further, decreased cell viability induced by Aβ was prevented by cilostazol, and this inhibition was reversed by 3-methyladenine, indicating that the protective effect of cilostazol against Aβ induced neurotoxicity is, in part, ascribable to the induction of autophagy. In conclusion, cilostazol modulates autophagy by increasing the activation of SIRT1, and thereby enhances Aβ clearance and increases cell viability.

  18. Blocking autophagy prevents bortezomib-induced NF-κB activation by reducing I-κBα degradation in lymphoma cells.

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    Li Jia

    Full Text Available Here we show that bortezomib induces effective proteasome inhibition and accumulation of poly-ubiquitinated proteins in diffuse large B-cell lymphoma (DLBCL cells. This leads to induction of endoplasmic reticulum (ER stress as demonstrated by accumulation of the protein CHOP, as well as autophagy, as demonstrated by accumulation of LC3-II proteins. Our data suggest that recruitment of both ubiquitinated proteins and LC3-II by p62 directs ubiquitinated proteins, including I-κBα, to the autophagosome. Degradation of I-κBα results in increased NF-κB nuclear translocation and transcription activity. Since bortezomib treatment promoted I-κBα phosphorylation, ubiquitination and degradation, this suggests that the route of I-κBα degradation was not via the ubiquitin-proteasome degradation system. The autophagy inhibitor chloroquine (CQ significantly inhibited bortezomib-induced I-κBα degradation, increased complex formation with NF-κB and reduced NF-κB nuclear translocation and DNA binding activity. Importantly, the combination of proteasome and autophagy inhibitors showed synergy in killing DLBCL cells. In summary, bortezomib-induced autophagy confers relative DLBCL cell drug resistance by eliminating I-κBα. Inhibition of both autophagy and the proteasome has great potential to kill apoptosis-resistant lymphoma cells.

  19. Phosphoethanolamine Modification of Neisseria gonorrhoeae Lipid A Reduces Autophagy Flux in Macrophages.

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    Susu M Zughaier

    Full Text Available Autophagy, an ancient homeostasis mechanism for macromolecule degradation, performs an important role in host defense by facilitating pathogen elimination. To counteract this host defense strategy, bacterial pathogens have evolved a variety of mechanisms to avoid or otherwise dysregulate autophagy by phagocytic cells so as to enhance their survival during infection. Neisseria gonorrhoeae is a strictly human pathogen that causes the sexually transmitted infection, gonorrhea. Phosphoethanolamine (PEA addition to the 4' position of the lipid A (PEA-lipid A moiety of the lipooligosaccharide (LOS produced by gonococci performs a critical role in this pathogen's ability to evade innate defenses by conferring decreased susceptibility to cationic antimicrobial (or host-defense peptides, complement-mediated killing by human serum and intraleukocytic killing by human neutrophils compared to strains lacking this PEA decoration. Heretofore, however, it was not known if gonococci can evade autophagy and if so, whether PEA-lipid A contributes to this ability. Accordingly, by using murine macrophages and human macrophage-like phagocytic cell lines we investigated if PEA decoration of gonococcal lipid A modulates autophagy formation. We report that infection with PEA-lipid A-producing gonococci significantly reduced autophagy flux in murine and human macrophages and enhanced gonococcal survival during their association with macrophages compared to a PEA-deficient lipid A mutant. Our results provide further evidence that PEA-lipid A produced by gonococci is a critical component in the ability of this human pathogen to evade host defenses.

  20. Phosphoethanolamine Modification of Neisseria gonorrhoeae Lipid A Reduces Autophagy Flux in Macrophages.

    Science.gov (United States)

    Zughaier, Susu M; Kandler, Justin L; Balthazar, Jacqueline T; Shafer, William M

    2015-01-01

    Autophagy, an ancient homeostasis mechanism for macromolecule degradation, performs an important role in host defense by facilitating pathogen elimination. To counteract this host defense strategy, bacterial pathogens have evolved a variety of mechanisms to avoid or otherwise dysregulate autophagy by phagocytic cells so as to enhance their survival during infection. Neisseria gonorrhoeae is a strictly human pathogen that causes the sexually transmitted infection, gonorrhea. Phosphoethanolamine (PEA) addition to the 4' position of the lipid A (PEA-lipid A) moiety of the lipooligosaccharide (LOS) produced by gonococci performs a critical role in this pathogen's ability to evade innate defenses by conferring decreased susceptibility to cationic antimicrobial (or host-defense) peptides, complement-mediated killing by human serum and intraleukocytic killing by human neutrophils compared to strains lacking this PEA decoration. Heretofore, however, it was not known if gonococci can evade autophagy and if so, whether PEA-lipid A contributes to this ability. Accordingly, by using murine macrophages and human macrophage-like phagocytic cell lines we investigated if PEA decoration of gonococcal lipid A modulates autophagy formation. We report that infection with PEA-lipid A-producing gonococci significantly reduced autophagy flux in murine and human macrophages and enhanced gonococcal survival during their association with macrophages compared to a PEA-deficient lipid A mutant. Our results provide further evidence that PEA-lipid A produced by gonococci is a critical component in the ability of this human pathogen to evade host defenses. PMID:26641098

  1. Regulation of autophagy by cytosolic acetyl-coenzyme A

    DEFF Research Database (Denmark)

    Mariño, Guillermo; Pietrocola, Federico; Eisenberg, Tobias;

    2014-01-01

    proteins, as well as the induction of autophagy, a homeostatic process of self-digestion. Multiple distinct manipulations designed to increase or reduce cytosolic AcCoA led to the suppression or induction of autophagy, respectively, both in cultured human cells and in mice. Moreover, maintenance of high Ac......CoA levels inhibited maladaptive autophagy in a model of cardiac pressure overload. Depletion of AcCoA reduced the activity of the acetyltransferase EP300, and EP300 was required for the suppression of autophagy by high AcCoA levels. Altogether, our results indicate that cytosolic AcCoA functions as a...

  2. Autophagy induction by silibinin positively contributes to its anti-metastatic capacity via AMPK/mTOR pathway in renal cell carcinoma.

    Science.gov (United States)

    Li, Feng; Ma, Zhenkun; Guan, Zhenfeng; Chen, Yule; Wu, Kaijie; Guo, Peng; Wang, Xinyang; He, Dalin; Zeng, Jin

    2015-01-01

    Silibinin, a dietary cancer chemopreventive flavonoid from the seeds of milk thistle, has been reported to exhibit anti-metastatic effects on renal cell carcinoma (RCC), but the mechanism underlying this phenomenon is not fully understood. The present study aimed at examining the potential role of autophagy in regulating silibinin-induced anti-metastatic effects on RCC cells. Using RCC ACHN and 786-O cells as a model system in vitro, we found that silibinin treatment increased the expression of LC3-II, resulted in the formation of autophagolysosome vacuoles, and caused a punctate fluorescence pattern with the monomeric red fluorescence protein-enhanced green fluorescence protein-LC3 (mRFP-EGFP-LC3) protein, which all are markers for cellular autophagy. Autophagy flux was induced by silibinin in RCC cells, as determined by LC3 turnover assay. Mechanically, the adenosine 5'-monophosphate activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) pathway was identified as involved in regulation of silibinin-induced autophagy. Furthermore, autophagy induction was demonstrated to positively contribute to silibinin-induced anti-metastatic effects on RCC cells in vitro. Activation of autophagy enhanced silibinin-induced inhibition of migration and invasion of RCC cells, while inhibition of autophagy attenuated it. These findings thus provide new information about the potential link between autophagy and metastasis inhibition induced by silibinin, and the induction of autophagy may shed some light into future treatment strategies for metastatic RCC. PMID:25884331

  3. Autophagy induction by Bcr-Abl-expressing cells facilitates their recovery from a targeted or nontargeted treatment.

    LENUS (Irish Health Repository)

    Crowley, Lisa C

    2012-01-31

    Although Imatinib has transformed the treatment of chronic myeloid leukemia (CML), it is not curative due to the persistence of resistant cells that can regenerate the disease. We have examined how Bcr-Abl-expressing cells respond to two mechanistically different therapeutic agents, etoposide and Imatinib. We also examined Bcr-Abl expression at low and high levels as elevated expression has been associated with treatment failure. Cells expressing low levels of Bcr-Abl undergo apoptosis in response to the DNA-targeting agent (etoposide), whereas high-Bcr-Abl-expressing cells primarily induce autophagy. Autophagic populations engage a delayed nonapoptotic death; however, sufficient cells evade this and repopulate following the withdrawal of the drug. Non-Bcr-Abl-expressing 32D or Ba\\/F3 cells induce both apoptosis and autophagy in response to etoposide and can recover. Imatinib treatment induces both apoptosis and autophagy in all Bcr-Abl-expressing cells and populations rapidly recover. Inhibition of autophagy with ATG7 and Beclin1 siRNA significantly reduced the recovery of Imatinib-treated K562 cells, indicating the importance of autophagy for the recovery of treated cells. Combination regimes incorporating agents that disrupt Imatinib-induced autophagy would remain primarily targeted and may improve response to the treatment in CML.

  4. Induction and adaptation of chaperone-assisted selective autophagy CASA in response to resistance exercise in human skeletal muscle

    OpenAIRE

    Ulbricht, Anna; Gehlert, Sebastian; Leciejewski, Barbara; Schiffer, Thorsten; Bloch, Wilhelm; Höhfeld, Jörg

    2015-01-01

    Chaperone-assisted selective autophagy (CASA) is a tension-induced degradation pathway essential for muscle maintenance. Impairment of CASA causes childhood muscle dystrophy and cardiomyopathy. However, the importance of CASA for muscle function in healthy individuals has remained elusive so far. Here we describe the impact of strength training on CASA in a group of healthy and moderately trained men. We show that strenuous resistance exercise causes an acute induction of CASA in affected mus...

  5. Neferine Attenuates the Protein Level and Toxicity of Mutant Huntingtin in PC-12 Cells via Induction of Autophagy

    Directory of Open Access Journals (Sweden)

    Vincent Kam Wai Wong

    2015-02-01

    Full Text Available Mutant huntingtin aggregation is highly associated with the pathogenesis of Huntington’s disease, an adult-onset autosomal dominant disorder, which leads to a loss of motor control and decline in cognitive function. Recent literature has revealed the protective role of autophagy in neurodegenerative diseases through degradation of mutant toxic proteins, including huntingtin or a-synuclein. Through the GFP-LC3 autophagy detection platform, we have  identified  neferine,  isolated  from  the  lotus  seed  embryo  of Nelumbo nucifera, which is able to induce autophagy through an AMPK-mTOR-dependent pathway. Furthermore, by overexpressing huntingtin with 74 CAG repeats (EGFP-HTT 74 in PC-12 cells, neferine reduces both the protein level and toxicity of mutant huntingtin through an autophagy-related gene 7 (Atg7-dependent mechanism. With the variety of novel active compounds present in medicinal herbs, our current study suggests the possible protective mechanism of an autophagy inducer isolated from Chinese herbal medicine, which is crucial for its further development into a potential therapeutic agent for neurodegenerative disorders in the future.

  6. Inducing autophagy

    DEFF Research Database (Denmark)

    Harder, Lea M; Bunkenborg, Jakob; Andersen, Jens S.

    2014-01-01

    Autophagy is a lysosomal-mediated catabolic process, which through degradation of different cytoplasmic components aids in maintaining cellular homeostasis and survival during exposure to extra- or intracellular stresses. Ammonia is a potential toxic and stress-inducing byproduct of glutamine...... catabolism, which has recently been found to induce autophagy in an MTOR independent way and support cancer cell survival. In this study, quantitative phosphoproteomics was applied to investigate the initial signaling events linking ammonia to the induction of autophagy. The MTOR inhibitor rapamycin was used...... as a reference treatment to emphasize the differences between an MTOR-dependent and -independent autophagy-induction. By this means 5901 phosphosites were identified of which 626 were treatment-specific regulated and 175 were coregulated. Investigation of the ammonia-specific regulated sites...

  7. Autophagy induction by leptin contributes to suppression of apoptosis in cancer cells and xenograft model: Involvement of p53/FoxO3A axis

    OpenAIRE

    Nepal, Saroj; Kim, Mi Jin; Hong, Jin Tae; Kim, Sang Hyun; Sohn, Dong-Hwan; Lee, Sung Hee; Song, Kyung; Choi, Dong Young; Lee, Eung Seok; Park, Pil-Hoon

    2015-01-01

    Leptin, a hormone mainly produced from adipose tissue, has been shown to induce proliferation of cancer cells. However, the molecular mechanisms underlying leptin-induced tumor progression have not been clearly elucidated. In the present study, we investigated the role of autophagy in leptin-induced cancer cell proliferation using human hepatoma (HepG2) and breast cancer cells (MCF-7), and tumor growth in a xenograft model. Herein, we showed that leptin treatment caused autophagy induction as...

  8. Autophagy inhibition in endogenous and nutrient-deprived conditions reduces dorsal root ganglia neuron survival and neurite growth in vitro.

    Science.gov (United States)

    Clarke, Joseph-Patrick; Mearow, Karen

    2016-07-01

    Peripheral neuropathies can result in cytoskeletal changes in axons, ultimately leading to Wallerian degeneration and cell death. Recently, autophagy has been studied as a potential target for improving axonal survival and growth during peripheral nerve damage. This study investigates the influence of autophagy on adult dorsal root ganglia (DRG) neuron survival and axonal growth under control and nutrient deprivation conditions. Constitutive autophagy was modulated with pharmacological activators (rapamycin; Rapa) and inhibitors (3-methyladenine, bafilomycin A1) in conjunction with either a nutrient-stable environment (standard culture medium) or a nutrient-deprived environment (Hank's balanced salt solution + Ca(2+) /Mg(2+) ). The results demonstrated that autophagy inhibition decreased cell viability and reduced neurite growth and branching complexity. Although autophagy was upregulated with nutrient deprivation compared with the control, it was not further activated by rapamycin, suggesting a threshold level of autophagy. Overall, both cellular and biochemical approaches combined to show the influence of autophagy on adult DRG neuron survival and growth. © 2016 Wiley Periodicals, Inc. PMID:27018986

  9. The preliminary study of autophagy induction of SA and MeSA by confocal

    Science.gov (United States)

    Yun, Lijuan; Chen, Wenli

    2010-02-01

    Autophagy appears to be a highly conserved process from unicellular to multicellular eukaryotes which contributes to the equilibrium of intracelluar environment. While it would be harmful to the cells when it is excessive by inducing programmed cell death (PCD). It is a protein degradation process in which cells recycle cytoplasmic contents when subjected to environmental stress conditions or during certain stages of development. Previous studies have demonstrated autophagy can be induced during abiotic or biotic stresses. salicylic acid (SA) and methyl salicytic (MeSA) are endogenous signal molecules. We found SA and MeSA can induce autophagy in Arabidopsis thaliana respectively. While autophagy was not induced by SA or MeSA in tobacco suspension cells under the same concentration and period. The differences in stuctures or physiological states may contribute to the results.

  10. Radiosensitization of renal cell carcinoma in vitro through the induction of autophagy

    International Nuclear Information System (INIS)

    Background and purpose: For patients diagnosed with advanced renal cell carcinoma (RCC), there are few therapeutic options. Radiation therapy is predominantly used to treat metastasis and has not proven effective in the adjuvant setting for renal cancer. Furthermore, RCC is resistant to standard cytotoxic chemotherapies. Targeted anti-angiogenics are the standard of care for RCC but are not curative. Newer agents, such as mTOR inhibitors and others that induce autophagy, have shown great promise for treating RCC. Here, we investigate the potential use of the small molecule STF-62247 to modulate radiation. Materials and methods: Using RCC cell lines, we evaluate sensitivity to radiation in addition to agents that induce autophagic cell death by clonogenic survival assays. Furthermore, these were also tested under physiological oxygen levels. Results: STF-62247 specifically induces autophagic cell death in cells that have lost VHL, an essential mutation in the development of RCC. Treatment with STF-62247 did not alter cell cycle progression but when combined with radiation increased cell killing under oxic and hypoxic/physiological conditions. Conclusions: This study highlights the possibility of combining targeted therapeutics such as STF-62247 or temsirolimus with radiation to reduce the reliance on partial or full nephrectomy and improve patient prognosis.

  11. Induction of cyto-protective autophagy by paramontroseite VO2 nanocrystals

    International Nuclear Information System (INIS)

    A variety of inorganic nanomaterials have been shown to induce autophagy, a cellular degradation process critical for the maintenance of cellular homeostasis. The overwhelming majority of autophagic responses elicited by nanomaterials were detrimental to cell fate and contributed to increased cell death. A widely held view is that the inorganic nanoparticles, when encapsulated and trapped by autophagosomes, may compromise the normal autophagic process due to the inability of the cells to degrade these materials and thus they manifest a detrimental effect on the well-being of a cell. Here we show that, contrary to this notion, nano-sized paramontroseite VO2 nanocrystals (P–VO2) induced cyto-protective, rather than death-promoting, autophagy in cultured HeLa cells. P–VO2 also caused up-regulation of heme oxygenase-1 (HO-1), a cellular protein with a demonstrated role in protecting cells against death under stress situations. The autophagy inhibitor 3-methyladenine significantly inhibited HO-1 up-regulation and increased the rate of cell death in cells treated with P–VO2, while the HO-1 inhibitor protoporphyrin IX zinc (II) (ZnPP) enhanced the occurrence of cell death in the P–VO2-treated cells while having no effect on the autophagic response induced by P–VO2. On the other hand, Y2O3 nanocrystals, a control nanomaterial, induced death-promoting autophagy without affecting the level of expression of HO-1, and the pro-death effect of the autophagy induced by Y2O3. Our results represent the first report on a novel nanomaterial-induced cyto-protective autophagy, probably through up-regulation of HO-1, and may point to new possibilities for exploiting nanomaterial-induced autophagy for therapeutic applications. (paper)

  12. Induction of cyto-protective autophagy by paramontroseite VO2 nanocrystals

    Science.gov (United States)

    Zhou, Wei; Miao, Yanyan; Zhang, Yunjiao; Liu, Liang; Lin, Jun; Yang, James Y.; Xie, Yi; Wen, Longping

    2013-04-01

    A variety of inorganic nanomaterials have been shown to induce autophagy, a cellular degradation process critical for the maintenance of cellular homeostasis. The overwhelming majority of autophagic responses elicited by nanomaterials were detrimental to cell fate and contributed to increased cell death. A widely held view is that the inorganic nanoparticles, when encapsulated and trapped by autophagosomes, may compromise the normal autophagic process due to the inability of the cells to degrade these materials and thus they manifest a detrimental effect on the well-being of a cell. Here we show that, contrary to this notion, nano-sized paramontroseite VO2 nanocrystals (P-VO2) induced cyto-protective, rather than death-promoting, autophagy in cultured HeLa cells. P-VO2 also caused up-regulation of heme oxygenase-1 (HO-1), a cellular protein with a demonstrated role in protecting cells against death under stress situations. The autophagy inhibitor 3-methyladenine significantly inhibited HO-1 up-regulation and increased the rate of cell death in cells treated with P-VO2, while the HO-1 inhibitor protoporphyrin IX zinc (II) (ZnPP) enhanced the occurrence of cell death in the P-VO2-treated cells while having no effect on the autophagic response induced by P-VO2. On the other hand, Y2O3 nanocrystals, a control nanomaterial, induced death-promoting autophagy without affecting the level of expression of HO-1, and the pro-death effect of the autophagy induced by Y2O3. Our results represent the first report on a novel nanomaterial-induced cyto-protective autophagy, probably through up-regulation of HO-1, and may point to new possibilities for exploiting nanomaterial-induced autophagy for therapeutic applications.

  13. Induction of autophagy by proteasome inhibitor is associated with proliferative arrest in colon cancer cells

    International Nuclear Information System (INIS)

    The ubiquitin-proteasome system (UPS) and lysosome-dependent macroautophagy (autophagy) are two major intracellular pathways for protein degradation. Blockade of UPS by proteasome inhibitors has been shown to activate autophagy. Recent evidence also suggests that proteasome inhibitors may inhibit cancer growth. In this study, the effect of a proteasome inhibitor MG-132 on the proliferation and autophagy of cultured colon cancer cells (HT-29) was elucidated. Results showed that MG-132 inhibited HT-29 cell proliferation and induced G2/M cell cycle arrest which was associated with the formation of LC3+ autophagic vacuoles and the accumulation of acidic vesicular organelles. MG-132 also increased the protein expression of LC3-I and -II in a time-dependent manner. In this connection, 3-methyladenine, a Class III phosphoinositide 3-kinase inhibitor, significantly abolished the formation of LC3+ autophagic vacuoles and the expression of LC3-II but not LC3-I induced by MG-132. Taken together, this study demonstrates that inhibition of proteasome in colon cancer cells lowers cell proliferation and activates autophagy. This discovery may shed a new light on the novel function of proteasome in the regulation of autophagy and proliferation in colon cancer cells

  14. MTOR-independent induction of autophagy in trabecular meshwork cells subjected to biaxial stretch.

    Science.gov (United States)

    Porter, Kristine M; Jeyabalan, Nallathambi; Liton, Paloma B

    2014-06-01

    The trabecular meshwork (TM) is part of a complex tissue that controls the exit of aqueous humor from the anterior chamber of the eye, and therefore helps maintaining intraocular pressure (IOP). Because of variations in IOP with changing pressure gradients and fluid movement, the TM and its contained cells undergo morphological deformations, resulting in distention and stretching. It is therefore essential for TM cells to continuously detect and respond to these mechanical forces and adapt their physiology to maintain proper cellular function and protect against mechanical injury. Here we demonstrate the activation of autophagy, a pro-survival pathway responsible for the degradation of long-lived proteins and organelles, in TM cells when subjected to biaxial static stretch (20% elongation), as well as in high-pressure perfused eyes (30mmHg). Morphological and biochemical markers for autophagy found in the stretched cells include elevated LC3-II levels, increased autophagic flux, and the presence of autophagic figures in electron micrographs. Furthermore, our results indicate that the stretch-induced autophagy in TM cells occurs in an MTOR- and BAG3-independent manner. We hypothesize that activation of autophagy is part of the physiological response that allows TM cells to cope and adapt to mechanical forces. PMID:24583119

  15. Autophagy contributes to resistance of tumor cells to ionizing radiation

    International Nuclear Information System (INIS)

    Background and purpose: Autophagy signaling is a novel important target to improve anticancer therapy. To study the role of autophagy on resistance of tumor cells to ionizing radiation (IR), breast cancer cell lines differing in their intrinsic radiosensitivity were used. Materials and methods: Breast cancer cell lines MDA-MB-231 and HBL-100 were examined with respect to clonogenic cell survival and induction of autophagy after radiation exposure and pharmacological interference of the autophagic process. As marker for autophagy the appearance of LC3-I and LC3-II proteins was analyzed by SDS-PAGE and Western blotting. Formation of autophagic vacuoles was monitored by immunofluorescence staining of LC3. Results: LC3-I and LC3-II formation differs markedly in radioresistant MDA-MB-231 versus radiosensitive HBL-100 cells. Western blot analyses of LC3-II/LC3-I ratio indicated marked induction of autophagy by IR in radioresistant MDA-MB-231 cells, but not in radiosensitive HBL-100 cells. Indirect immunofluorescence analysis of LC3-II positive vacuoles confirmed this differential effect. Pre-treatment with 3-methyladenine (3-MA) antagonized IR-induced autophagy. Likewise, pretreatment of radioresistant MDA-231 cells with autophagy inhibitors 3-MA or chloroquine (CQ) significantly reduced clonogenic survival of irradiated cells. Conclusion: Our data clearly indicate that radioresistant breast tumor cells show a strong post-irradiation induction of autophagy, which thus serves as a protective and pro-survival mechanism in radioresistance.

  16. Lanthanide co-doped paramagnetic spindle-like mesocrystals for imaging and autophagy induction.

    Science.gov (United States)

    Xu, Yun-Jun; Lin, Jun; Lu, Yang; Zhong, Sheng-Liang; Wang, Lei; Dong, Liang; Wu, Ya-Dong; Peng, Jun; Zhang, Li; Pan, Xiao-Feng; Zhou, Wei; Zhao, Yang; Wen, Long-Ping; Yu, Shu-Hong

    2016-07-21

    We synthesized two novel lanthanide doped spindle-like mesocrystals, YF3:Ce,Eu,Gd and YF3:Ce,Tb,Gd (abbreviated as YEG and YTG mesospindles, respectively). Both of them possess paramagnetic and fluorescent properties, and their excellent cyto-compatibility and low haemolysis are further confirmed. Therefore, they could act as dual mode contrast agents for magnetic resonance imaging (MRI) and fluorescence imaging. Furthermore, YEG and YTG mesospindles induce dose and time dependent autophagy by activating the PI3K signaling pathway. The autophagy induced by YEG and YTG mesocrystals is confirmed by enhanced autophagosome formation, normal cargo degradation, and no disruption of lysosomal function. This work is important to illustrate how rare-earth mesocrystals affect the autophagic pathway, indicating the potential of the YEG and YTG mesospindles in diagnosis and therapy. PMID:27346838

  17. Lipopolysaccharide induction of autophagy is associated with enhanced bactericidal activity in Dictyostelium discoideum

    OpenAIRE

    Pflaum, Katherine; Gerdes, Kimberly; Yovo, Kossi; Callahan, Jennifer; Snyder, Michelle L.D.

    2012-01-01

    Innate immune cells respond to microbial invaders using pattern recognition receptors that detect conserved microbial patterns. Among the cellular processes stimulated downstream of pattern recognition machinery is the initiation of autophagy, which plays protective roles against intracellular microbes. We have shown recently that Dictyostelium discoideum, which takes up bacteria for nutritive purposes, may employ pattern recognition machinery to respond to bacterial prey, as D. discoideum ce...

  18. Autophagy induction by vitamin D inhibits both Mycobacterium tuberculosis and human immunodeficiency virus type 1

    OpenAIRE

    Campbell, Grant R.; Spector, Stephen A.

    2012-01-01

    Low vitamin D levels in human immunodeficiency virus type-1 (HIV) infected persons are associated with more rapid disease progression and increased risk for Mycobacterium tuberculosis infection. We report that physiological concentrations of 1α,25-dihydroxycholecalciferol (1,25D3), the active form of vitamin D, inhibits M. tuberculosis and HIV replication in co-infected macrophages through human cathelicidin microbial peptide-dependent autophagy that requires phagosomal maturation. These find...

  19. Quercetin reduces eIF2α phosphorylation by GADD34 induction.

    Science.gov (United States)

    Hayakawa, Miki; Itoh, Masanori; Ohta, Kazunori; Li, Shimo; Ueda, Masashi; Wang, Miao-xing; Nishida, Emika; Islam, Saiful; Suzuki, Chihiro; Ohzawa, Kaori; Kobori, Masuko; Inuzuka, Takashi; Nakagawa, Toshiyuki

    2015-09-01

    The production of amyloid β (Aβ) in the brain from Aβ precursor protein (APP) through γ-secretase is important for the pathogenesis of Alzheimer's disease (AD). Our previous studies have demonstrated that autophagy impairment and endoplasmic reticulum stress increase presenilin 1 expression and enhance γ-secretase activity through the phosphorylation of eukaryotic translation initiation factor 2α (eIF2α) and the translation of activating transcription factor 4 (ATF4). However, the inhibitory molecules for γ-secretase are largely unknown. Here, we demonstrate that the levels of ATF4 expression are increased in the brain of APP23 AD model mice; furthermore, these levels enhanced in the brain of APP23 mice crossed with obese and diabetic db/db (Lepr(db/db)) mice. A polyhydroxylated flavonoid, quercetin, suppressed presenilin 1 expression and Aβ secretion in autophagy-impaired cells by the induction of growth arrest and DNA damaged-inducible gene (GADD) 34, which mediates eIF2α dephosphorylation, leading to decreased ATF4 expression. GADD34 induction was observed in the brain of wild-type mice, and APP23 mice fed quercetin in their diet. After the long-term feeding of quercetin, deterioration in memory assessed by freezing behavior was delayed in APP23 mice. These results indicate that quercetin may reduce eIF2α phosphorylation and ATF4 expression through GADD34 induction in the brain, leading to the improvement of memory in aged mice and the delay of deterioration in memory at the early stage of AD in AD model mice. PMID:26070242

  20. The Neuroprotective Effect of Erythropoietin on Rotenone-Induced Neurotoxicity in SH-SY5Y Cells Through the Induction of Autophagy.

    Science.gov (United States)

    Jang, Wooyoung; Kim, Hee Ju; Li, Huan; Jo, Kwang Deog; Lee, Moon Kyu; Yang, Hyun Ok

    2016-08-01

    Currently, the autophagy pathway is thought to be important for the pathogenesis of Parkinson's disease (PD), and the modulation of autophagy may be a novel strategy for the treatment of this disease. Erythropoietin (EPO) has been reported to have neuroprotective effects through anti-oxidative, anti-apoptotic, and anti-inflammatory mechanisms, and it has also been shown to modulate autophagy signaling in an oxygen toxicity model. Therefore, we investigated the effects of EPO on autophagy markers and evaluated its neuroprotective effect on rotenone-induced neurotoxicity. We adapted the rotenone-induced neurotoxicity model to SH-SY5Y cells as an in vitro model of PD. We measured cell viability using MTT and annexin V/propidium iodide assays and measured intracellular levels of reactive oxygen species. Immunofluorescence analysis was performed to measure the expression of LC3 and α-synuclein. Intracellular signaling proteins associated with autophagy were examined by immunoblot analysis. EPO mono-treatment increased the levels of mammalian target of rapamycin (mTOR)-independent/upstream autophagy markers, including Beclin-1, AMPK, and ULK-1. Rotenone treatment of SH-SY5Y cells reduced their viability, increased reactive oxygen species levels, and induced apoptosis and α-synuclein expression, and simultaneous exposure to EPO significantly reduced these effects. Rotenone enhanced mTOR expression and suppressed Beclin-1 expression, indicating suppression of the autophagy system. However, combined treatment with EPO restored Beclin-1 expression and decreased mTOR expression. EPO protects against rotenone-induced neurotoxicity in SH-SY5Y cells by enhancing autophagy-related signaling pathways. The experimental evidence for the EPO-induced neuroprotection against rotenone-induced dopaminergic neurotoxicity may significantly impact the development of future PD treatment strategies. PMID:26156288

  1. Autophagy sustains the replication of porcine reproductive and respiratory virus in host cells

    International Nuclear Information System (INIS)

    In this study, we confirmed the autophagy induced by porcine reproductive and respiratory syndrome virus (PRRSV) in permissive cells and investigated the role of autophagy in the replication of PRRSV. We first demonstrated that PRRSV infection significantly results in the increased double-membrane vesicles, the accumulation of LC3 fluorescence puncta, and the raised ratio of LC3-II/β-actin, in MARC-145 cells. Then we discovered that induction of autophagy by rapamycin significantly enhances the viral titers of PRRSV, while inhibition of autophagy by 3-MA and silencing of LC3 gene by siRNA reduces the yield of PRRSV. The results showed functional autolysosomes can be formed after PRRSV infection and the autophagosome–lysosome-fusion inhibitor decreases the virus titers. We also examined the induction of autophagy by PRRSV infection in pulmonary alveolar macrophages. These findings indicate that autophagy induced by PRRSV infection plays a role in sustaining the replication of PRRSV in host cells.

  2. Autophagy sustains the replication of porcine reproductive and respiratory virus in host cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Qinghao; Qin, Yixian; Zhou, Lei; Kou, Qiuwen; Guo, Xin; Ge, Xinna [Key Laboratory of Animal Epidemiology and Zoonosis of Ministry of Agriculture, College of Veterinary Medicine and State Key Laboratory of Agribiotechnology, China Agricultural University, Beijing (China); Yang, Hanchun, E-mail: yanghanchun1@cau.edu.cn [Key Laboratory of Animal Epidemiology and Zoonosis of Ministry of Agriculture, College of Veterinary Medicine and State Key Laboratory of Agribiotechnology, China Agricultural University, Beijing (China); Hu, Hongbo, E-mail: hongbo@cau.edu.cn [College of Food Science and Nutritional Engineering, China Agricultural University, Beijing (China)

    2012-08-01

    In this study, we confirmed the autophagy induced by porcine reproductive and respiratory syndrome virus (PRRSV) in permissive cells and investigated the role of autophagy in the replication of PRRSV. We first demonstrated that PRRSV infection significantly results in the increased double-membrane vesicles, the accumulation of LC3 fluorescence puncta, and the raised ratio of LC3-II/{beta}-actin, in MARC-145 cells. Then we discovered that induction of autophagy by rapamycin significantly enhances the viral titers of PRRSV, while inhibition of autophagy by 3-MA and silencing of LC3 gene by siRNA reduces the yield of PRRSV. The results showed functional autolysosomes can be formed after PRRSV infection and the autophagosome-lysosome-fusion inhibitor decreases the virus titers. We also examined the induction of autophagy by PRRSV infection in pulmonary alveolar macrophages. These findings indicate that autophagy induced by PRRSV infection plays a role in sustaining the replication of PRRSV in host cells.

  3. Adiponectin stimulates autophagy and reduces oxidative stress to enhance insulin sensitivity during high-fat diet feeding in mice.

    Science.gov (United States)

    Liu, Ying; Palanivel, Rengasamy; Rai, Esther; Park, Min; Gabor, Tim V; Scheid, Michael P; Xu, Aimin; Sweeney, Gary

    2015-01-01

    Numerous studies have characterized the antidiabetic effects of adiponectin, yet the precise cellular mechanisms in skeletal muscle, in particular, changes in autophagy, require further clarification. In the current study, we used a high-fat diet (HFD) to induce obesity and insulin resistance in wild-type (WT) or adiponectin knockout (Ad-KO) mice with and without adiponectin replenishment. Temporal analysis of glucose tolerance and insulin sensitivity using hyperinsulinemic-euglycemic clamp and muscle insulin receptor substrate and Akt phosphorylation demonstrated exaggerated and more rapid HFD-induced insulin resistance in skeletal muscle of Ad-KO mice. Superoxide dismutase activity, the reduced glutathione-to-glutathione disulfide ratio, and lipid peroxidation indicated that HFD-induced oxidative stress was corrected by adiponectin. Gene array analysis implicated several antioxidant enzymes, including Gpxs, Prdx, Sod, and Nox4, in mediating this effect. Adiponectin also attenuated palmitate-induced reactive oxygen species production in cultured myotubes and improved insulin-stimulated glucose uptake in primary muscle cells. Increased LC3-II and decreased p62 expression suggested that HFD induced autophagy in muscle of WT mice; however, these changes were not observed in Ad-KO mice. Replenishing adiponectin in Ad-KO mice increased LC3-II and Beclin1 and decreased p62 protein levels, induced fibroblast growth factor-21 expression, and corrected HFD-induced decreases in LC3, Beclin1, and ULK1 gene expression. In vitro studies examining changes in phospho-ULK1 (Ser555), LC3-II, and lysosomal enzyme activity confirmed that adiponectin directly induced autophagic flux in cultured muscle cells in an AMPK-dependent manner. We overexpressed an inactive mutant of Atg5 to create an autophagy-deficient cell model, and together with pharmacological inhibition of autophagy, demonstrated reduced insulin sensitivity under these conditions. In summary, adiponectin stimulated

  4. Induction of cellular senescence by doxorubicin is associated with upregulated miR-375 and induction of autophagy in K562 cells.

    Directory of Open Access Journals (Sweden)

    Ming-Yu Yang

    Full Text Available BACKGROUND: Cellular senescence is a specialized form of growth arrest that is generally irreversible. Upregulated p16, p53, and p21 expression and silencing of E2F target genes have been characterized to promote the establishment of senescence. It can be further aided by the transcriptional repression of proliferation-associated genes by the action of HP1γ, HMGA, and DNMT proteins to produce a repressive chromatin environment. Therefore, senescence has been suggested to functions as a natural brake for tumor development and plays a critical role in tumor suppression and aging. METHODOLOGY/PRINCIPAL FINDINGS: An in vitro senescence model has been established by using K562 cells treated with 50 nM doxorubicin (DOX. Since p53 and p16 are homozygously deleted in the K562 cells, the DOX-induced senescence in K562 cells ought to be independent of p53 and p16-pRb pathways. Indeed, no change in the expression of the typical senescence-associated premalignant cell markers in the DOX-induced senescent K562 cells was found. MicroRNA profiling revealed upregulated miR-375 in DOX-induced senescent K562 cells. Treatment with miR-375 inhibitor was able to reverse the proliferation ability suppressed by DOX (p<0.05 and overexpression of miR-375 suppressed the normal proliferation of K562 cells. Upregulated miR-375 expression was associated with downregulated expression of 14-3-3zeta and SP1 genes. Autophagy was also investigated since DOX treatment was able to induce cells entering senescence and eventually lead to cell death. Among the 24 human autophagy-related genes examined, a 12-fold increase of ATG9B at day 4 and a 20-fold increase of ATG18 at day 2 after DOX treatment were noted. CONCLUSIONS/SIGNIFICANCE: This study has demonstrated that in the absence of p53 and p16, the induction of senescence by DOX was associated with upregulation of miR-375 and autophagy initiation. The anti-proliferative function of miR-375 is possibly exerted, at least in part

  5. Autophagy induction by silver nanowires: A new aspect in the biocompatibility assessment of nanocomposite thin films

    International Nuclear Information System (INIS)

    Nanomaterials and their enabled products have increasingly been attracting global attention due to their unique physicochemical properties. Among these emerging products, silver nanowire (AgNW)-based thin films are being developed for their promising applications in next generation nanoelectronics and nanodevices. However, serious concerns remain about possible health and safety risks they may pose. Here, we employed a multi-modal systematic biocompatibility assessment of thin films incorporating AgNW. To represent the possible routes of nanomaterial entry during occupational or environmental exposure, we employed four different cell lines of epithelial, endothelial, gastric, and phagocytic origin. Utilizing a cell-based automated image acquisition and analysis procedure in combination with real-time impedance sensing, we observed a low level of cytotoxicity of AgNW, which was dependent on cell type, nanowire lengths, doses and incubation times. Similarly, no major cytotoxic effects were induced by AgNW-containing thin films, as detected by conventional cell viability and imaging assays. However, transmission electron microscopy and Western immunoblotting analysis revealed AgNW-induced autophasosome accumulation together with an upregulation of the autophagy marker protein LC3. Autophagy represents a crucial mechanism in maintaining cellular homeostasis, and our data for the first time demonstrate triggering of such mechanism by AgNW in human phagocytic cells. Finally, atomic force microscopy revealed significant changes in the topology of cells attaching and growing on these films as substrates. Our findings thus emphasize the necessity of comprehensive biohazard assessment of nanomaterials in modern applications and devices and a thorough analysis of risks associated with their possible contact with humans through occupational or environmental exposure. Highlights: ► Thin films containing nanomaterials are subject to increasing contact with humans. ► This

  6. NF-κB p65 repression by the sesquiterpene lactone, Helenalin, contributes to the induction of autophagy cell death

    OpenAIRE

    Lim Chuan; Fu Pan; Ky Nung; Zhu Hong; Feng XiaoLing; Li Jinming; Srinivasan Kandhadayar; Hamza Mohamed; Zhao Yan

    2012-01-01

    Abstract Background Numerous studies have demonstrated that autophagy plays a vital role in maintaining cellular homeostasis. Interestingly, several anticancer agents were found to exert their anticancer effects by triggering autophagy. Emerging data suggest that autophagy represents a novel mechanism that can be exploited for therapeutic benefit. Pharmacologically active natural compounds such as those from marine, terrestrial plants and animals represent a promising resource for novel antic...

  7. Enhanced autophagy as a potential mechanism for the improved physiological function by simvastatin in muscular dystrophy.

    Science.gov (United States)

    Whitehead, Nicholas P

    2016-04-01

    Autophagy has recently emerged as an important cellular process for the maintenance of skeletal muscle health and function. Excessive autophagy can trigger muscle catabolism, leading to atrophy. In contrast, reduced autophagic flux is a characteristic of several muscle diseases, including Duchenne muscular dystrophy, the most common and severe inherited muscle disorder. Recent evidence demonstrates that enhanced reactive oxygen species (ROS) production by CYBB/NOX2 impairs autophagy in muscles from the dmd/mdx mouse, a genetic model of Duchenne muscular dystrophy. Statins decrease CYBB/NOX2 expression and activity and stimulate autophagy in skeletal muscle. Therefore, we treated dmd/mdx mice with simvastatin and showed decreased CYBB/NOX2-mediated oxidative stress and enhanced autophagy induction. This was accompanied by reduced muscle damage, inflammation and fibrosis, and increased muscle force production. Our data suggest that increased autophagy may be a potential mechanism by which simvastatin improves skeletal muscle health and function in muscular dystrophy. PMID:26890413

  8. ROS-induced DNA damage and PARP-1 are required for optimal induction of starvation-induced autophagy

    Institute of Scientific and Technical Information of China (English)

    José Manuel Rodríguez-Vargas; Abelardo López Rivas; Marja J(a)(a)ttela; F Javier Oliver; María José Ruiz-Maga(n)a; Carmen Ruiz-Ruiz; Jara Majuelos-Melguizo; Andreína Peralta-Leal; María Isabel Rodríguez; José Antonio Mu(n)oz-Gámez; Mariano Ruiz de Almodóvar; Eva Siles

    2012-01-01

    In response to nutrient stress,cells start an autophagy program that can lead to adaptation or death.The mechanisms underlying the signaling from starvation to the initiation of autophagy are not fully understood.In the current study we show that the absence or inactivation of PARP-1 strongly delays starvation-induced autophagy.We have found that DNA damage is an early event of starvation-induced autophagy as measured by γ-H2AX accumulation and comet assay,with PARP-1 knockout cells displaying a reduction in both parameters.During starvation,ROS-induced DNA damage activates PARP-1,leading to ATP depletion (an early event after nutrient deprivation).The absence of PARP-1 blunted AMPK activation and prevented the complete loss of mTOR activity,leading to a delay in autophagy.PARP-1 depletion favors apoptosis in starved cells,suggesting a pro-survival role of autophagy and PARP-1 activation after nutrient deprivation.In vivo results show that neonates of PARP-1 mutant mice subjected to acute starvation,also display deficient liver autophagy,implying a physiological role for PARP-1 in starvation-induced autophagy.Thus,the PARP signaling pathway is a key regulator of the initial steps of autophagy commitment following starvation.

  9. Induction of Autophagy in the Striatum and Hypothalamus of Mice after 835 MHz Radiofrequency Exposure.

    Science.gov (United States)

    Kim, Ju Hwan; Huh, Yang Hoon; Kim, Hak Rim

    2016-01-01

    The extensive use of wireless mobile phones and associated communication devices has led to increasing public concern about potential biological health-related effects of the exposure to electromagnetic fields (EMFs). EMFs emitted by a mobile phone have been suggested to influence neuronal functions in the brain and affect behavior. However, the affects and phenotype of EMFs exposure are unclear. We applied radiofrequency (RF) of 835 MHz at a specific absorption rate (SAR) of 4.0 W/kg for 5 hours/day for 4 and 12 weeks to clarify the biological effects on mouse brain. Interestingly, microarray data indicated that a variety of autophagic related genes showed fold-change within small range after 835 MHz RF exposure. qRT-PCR revealed significant up-regulation of the autophagic genes Atg5, LC3A and LC3B in the striatum and hypothalamus after a 12-week RF. In parallel, protein expression of LC3B-II was also increased in both brain regions. Autophagosomes were observed in the striatum and hypothalamus of RF-exposed mice, based on neuronal transmission electron microscopy. Taken together, the results indicate that RF exposure of the brain can induce autophagy in neuronal tissues, providing insight into the protective mechanism or adaptation to RF stress. PMID:27073885

  10. ROS-induced DNA damage and PARP-1 are required for optimal induction of starvation-induced autophagy

    DEFF Research Database (Denmark)

    Rodríguez-Vargas, José Manuel; Ruiz-Magaña, María José; Ruiz-Ruiz, Carmen;

    2012-01-01

    delays starvation-induced autophagy. We have found that DNA damage is an early event of starvation-induced autophagy as measured by ¿-H2AX accumulation and comet assay, with PARP-1 knockout cells displaying a reduction in both parameters. During starvation, ROS-induced DNA damage activates PARP-1...

  11. Globular Adiponectin Causes Tolerance to LPS-Induced TNF-α Expression via Autophagy Induction in RAW 264.7 Macrophages: Involvement of SIRT1/FoxO3A Axis

    OpenAIRE

    Tilija Pun, Nirmala; Subedi, Amit; Kim, Mi Jin; Park, Pil-Hoon

    2015-01-01

    Adiponectin, an adipokine predominantly produced from adipose tissue, exhibited potent anti-inflammatory properties. In particular, it inhibits production of pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α), in macrophages. Autophagy, an intracellular self-digestion process, has been recently shown to regulate inflammatory responses. In the present study, we investigated the role of autophagy induction in the suppression of Lipopolysaccharide (LPS) -induced TNF-α expressi...

  12. Targeting Protective Autophagy Exacerbates UV-Triggered Apoptotic Cell Death

    Directory of Open Access Journals (Sweden)

    Shih-Hwa Chiou

    2012-01-01

    Full Text Available Autophagy is activated by various stresses, including DNA damage, and previous studies of DNA damage-induced autophagy have focused on the response to chemotherapeutic drugs, ionizing radiation, and reactive oxygen species. In this study, we investigated the biological significance of autophagic response to ultraviolet (UV irradiation in A549 and H1299 cells. Our results indicated that UV induces on-rate autophagic flux in these cells. Autophagy inhibition resulting from the knockdown of beclin-1 and Atg5 reduced cell viability and enhanced apoptosis. Moreover, we found that ATR phosphorylation was accompanied by microtubule-associated protein 1 light chain 3B II (LC3B-II expression during the early phases following UV irradiation, which is a well-established inducer of ATR. Knocking down ATR further attenuated the reduction in LC3B-II at early stages in response to UV treatment. Despite the potential role of ATR in autophagic response, reduced ATR expression does not affect autophagy induction during late phases (24 and 48 h after UV treatment. The result is consistent with the reduced ATR phosphorylation at the same time points and suggests that autophagic response at this stage is activated via a distinct pathway. In conclusion, this study demonstrated that autophagy acts as a cytoprotective mechanism against UV-induced apoptosis and that autophagy induction accompanied with apoptosis at late stages is independent of ATR activation.

  13. Carnosic Acid Prevents Beta-Amyloid-Induced Injury in Human Neuroblastoma SH-SY5Y Cells via the Induction of Autophagy.

    Science.gov (United States)

    Liu, Jie; Su, Hua; Qu, Qiu-Min

    2016-09-01

    Beta-amyloid (Aβ), the hallmark protein in Alzheimer's disease (AD), induces neurotoxicity that involves oxidative stress and mitochondrial dysfunction, leading to cell death. Carnosic acid (CA), a polyphenolic diterpene isolated from the herb rosemary (Rosemarinus officinalis), was investigated in our study to assess its neuroprotective effect and underlying mechanism against Aβ-induced injury in human neuroblastoma SH-SY5Y cells. We found that CA pretreatment alleviated the Aβ25-35-induced loss of cell viability, inhibited both Aβ1-42 accumulation and tau hyperphosphorylation, reduced reactive oxygen species generation, and maintained the mitochondrial membrane potential. Moreover, CA increased the microtubule-associated protein light chain 3 (LC3)-II/I ratio and decreased SQSTM1(p62), indicating that CA could induce autophagy. Autophagy inhibitor 3-methyladenine (3-MA) attenuated the neuroprotective effect of CA, suggesting that autophagy was involved in the neuroprotection of CA. It was also observed that CA activated AMP-activated protein kinase (AMPK) but inhibited mammalian target of rapamycin (mTOR). Furthermore, blocking AMPK with si-AMPKα successfully inhibited the upregulation of LC3-II/I, prevented the downregulation of phosphorylation of mTOR and SQSTM1(p62), indicating that CA induced autophagy in SH-SY5Y cells via the activation of AMPK. These results suggested that CA might be a potential agent for preventing AD. PMID:27168327

  14. Hepatic autophagy contributes to the metabolic response to dietary protein restriction.

    Science.gov (United States)

    Henagan, Tara M; Laeger, Thomas; Navard, Alexandra M; Albarado, Diana; Noland, Robert C; Stadler, Krisztian; Elks, Carrie M; Burk, David; Morrison, Christopher D

    2016-06-01

    Autophagy is an essential cellular response which acts to release stored cellular substrates during nutrient restriction, and particularly plays a key role in the cellular response to amino acid restriction. However, there has been limited work testing whether the induction of autophagy is required for adaptive metabolic responses to dietary protein restriction in the whole animal. Here, we found that moderate dietary protein restriction led to a series of metabolic changes in rats, including increases in food intake and energy expenditure, the downregulation of hepatic fatty acid synthesis gene expression and reduced markers of hepatic mitochondrial number. Importantly, these effects were also associated with an induction of hepatic autophagy. To determine if the induction of autophagy contributes to these metabolic effects, we tested the metabolic response to dietary protein restriction in BCL2-AAA mice, which bear a genetic mutation that impairs autophagy induction. Interestingly, BCL2-AAA mice exhibit exaggerated responses in terms of both food intake and energy expenditure, whereas the effects of protein restriction on hepatic metabolism were significantly blunted. These data demonstrate that restriction of dietary protein is sufficient to trigger hepatic autophagy, and that disruption of autophagy significantly alters both hepatic and whole animal metabolic response to dietary protein restriction. PMID:27173459

  15. Knockdown of autophagy-related protein 5, ATG5, decreases oxidative stress and has an opposing effect on camptothecin-induced cytotoxicity in osteosarcoma cells

    International Nuclear Information System (INIS)

    Autophagy induction can increase or decrease anticancer drug efficacy. Anticancer drug-induced autophagy induction is poorly characterized in osteosarcoma (OS). In this study, we investigated the impact of autophagy inhibition on camptothecin (CPT)-induced cytotoxicity in OS. Autophagy-inhibited DLM8 and K7M3 metastatic murine OS cell lines were generated by infection with lentiviral shRNA directed against the essential autophagy protein ATG5. Knockdown of ATG5 protein expression and inhibition of autophagy was confirmed by immunoblot of ATG5 and LC3II proteins, respectively. Metabolic activity was determined by MTT assay and cell viability was determined by trypan blue exclusion. Acridine orange staining and immunoblotting for LC3II protein expression were used to determine autophagy induction. Oxidative stress was assessed by staining cells with HE and DCFH-DA followed by flow cytometry analysis. Mitochondrial membrane potential was determined by staining cells with TMRE followed by flow cytometry analysis. Immunoblotting was used to detect caspase activation, Parp cleavage and p53 phosphorylation. Autophagy inhibition caused a greater deficit in metabolic activity and cell growth in K7M3 cells compared to DLM8 cells. K7M3 cells exhibited higher basal autophagy levels than DLM8 cells and non-transformed murine MCT3 osteoblasts. Autophagy inhibition did not affect CPT-induced DNA damage. Autophagy inhibition decreased CPT-induced cell death in DLM8 cells while increasing CPT-induced cell death in K7M3 cells. Autophagy inhibition reduced CPT-induced mitochondrial damage and CPT-induced caspase activation in DLM8 cells. Buthionine sulfoximine (BSO)-induced cell death was greater in autophagy-competent DLM8 cells and was reversed by antioxidant pretreatment. Camptothecin-induced and BSO-induced autophagy induction was also reversed by antioxidant pretreatment. Significantly, autophagy inhibition not only reduced CPT-induced oxidative stress but also reduced basal

  16. Lipoic acid inhibits the DNA repair protein O 6-methylguanine-DNA methyltransferase (MGMT) and triggers its depletion in colorectal cancer cells with concomitant autophagy induction.

    Science.gov (United States)

    Göder, Anja; Nagel, Georg; Kraus, Alexander; Dörsam, Bastian; Seiwert, Nina; Kaina, Bernd; Fahrer, Jörg

    2015-08-01

    Alkylating agents are present in food and tobacco smoke, but are also used in cancer chemotherapy, inducing the DNA lesion O (6)-methylguanine. This critical adduct is repaired by O (6)-methylguanine-DNA methyltransferase (MGMT), resulting in MGMT inactivation and degradation. In the present study, we analyzed the effects of the natural disulfide compound lipoic acid (LA) on MGMT in vitro and in colorectal cancer cells. We show that LA, but not its reduced form dihydrolipoic acid, potently inhibits the activity of recombinant MGMT by interfering with its catalytic Cys-145 residue, which was partially reversible by N-acetyl cysteine. Incubation of HCT116 colorectal cancer cells with LA altered their glutathione pool and caused a decline in MGMT activity. This was mirrored by LA-induced depletion of MGMT protein, which was not attributable to changes in MGMT messenger RNA levels. Loss of MGMT protein coincided with LA-induced autophagy, a process resulting in lysosomal degradation of proteins, including presumably MGMT. LA-stimulated autophagy in a p53-independent manner as revealed by the response of isogenic HCT116 cell lines. Knockdown of the crucial autophagy component beclin-1 and chemical inhibitors blocked LA-induced autophagy, but did not abrogate LA-triggered MGMT degradation. Concomitant with MGMT depletion, LA pretreatment resulted in enhanced O (6)-methylguanine levels in DNA. It also increased the cytotoxicity of the alkylating anticancer drug temozolomide in temozolomide-resistant colorectal cancer cells. Taken together, our study showed that the natural compound LA inhibits MGMT and induces autophagy. Furthermore, LA enhanced the cytotoxic effects of temozolomide, which makes it a candidate for a supplement in cancer therapy. PMID:25998848

  17. VMP1 related autophagy and apoptosis in colorectal cancer cells: VMP1 regulates cell death

    Energy Technology Data Exchange (ETDEWEB)

    Qian, Qinyi [Department of Ultrasonograph, Changshu No. 2 People’s Hospital, Changshu (China); Zhou, Hao; Chen, Yan [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China); Shen, Chenglong [Department of General Surgery, Changshu No. 2 People’s Hospital, Changshu (China); He, Songbing; Zhao, Hua; Wang, Liang [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China); Wan, Daiwei, E-mail: 372710369@qq.com [Department of Hepatobiliary Surgery, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou (China); Gu, Wen, E-mail: 505339704@qq.com [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China)

    2014-01-17

    Highlights: •This research confirmed VMP1 as a regulator of autophagy in colorectal cancer cell lines. •We proved the pro-survival role of VMP1-mediated autophagy in colorectal cancer cell lines. •We found the interaction between VMP1 and BECLIN1 also existing in colorectal cancer cell lines. -- Abstract: Vacuole membrane protein 1 (VMP1) is an autophagy-related protein and identified as a key regulator of autophagy in recent years. In pancreatic cell lines, VMP1-dependent autophagy has been linked to positive regulation of apoptosis. However, there are no published reports on the role of VMP1 in autophagy and apoptosis in colorectal cancers. Therefore, to address this gap of knowledge, we decided to interrogate regulation of autophagy and apoptosis by VMP1. We have studied the induction of autophagy by starvation and rapamycin treatment in colorectal cell lines using electron microscopy, immunofluorescence, and immunoblotting. We found that starvation-induced autophagy correlated with an increase in VMP1 expression, that VMP1 interacted with BECLIN1, and that siRNA mediated down-regulation of VMP1-reduced autophagy. Next, we examined the relationship between VMP1-dependent autophagy and apoptosis and found that VMP1 down-regulation sensitizes cells to apoptosis and that agents that induce apoptosis down-regulate VMP1. In conclusion, similar to its reported role in other cell types, VMP1 is an important regulator of autophagy in colorectal cell lines. However, in contrast to its role in pancreatic cell lines, in colorectal cancer cells, VMP1-dependent autophagy appears to be pro-survival rather than pro-cell death.

  18. VMP1 related autophagy and apoptosis in colorectal cancer cells: VMP1 regulates cell death

    International Nuclear Information System (INIS)

    Highlights: •This research confirmed VMP1 as a regulator of autophagy in colorectal cancer cell lines. •We proved the pro-survival role of VMP1-mediated autophagy in colorectal cancer cell lines. •We found the interaction between VMP1 and BECLIN1 also existing in colorectal cancer cell lines. -- Abstract: Vacuole membrane protein 1 (VMP1) is an autophagy-related protein and identified as a key regulator of autophagy in recent years. In pancreatic cell lines, VMP1-dependent autophagy has been linked to positive regulation of apoptosis. However, there are no published reports on the role of VMP1 in autophagy and apoptosis in colorectal cancers. Therefore, to address this gap of knowledge, we decided to interrogate regulation of autophagy and apoptosis by VMP1. We have studied the induction of autophagy by starvation and rapamycin treatment in colorectal cell lines using electron microscopy, immunofluorescence, and immunoblotting. We found that starvation-induced autophagy correlated with an increase in VMP1 expression, that VMP1 interacted with BECLIN1, and that siRNA mediated down-regulation of VMP1-reduced autophagy. Next, we examined the relationship between VMP1-dependent autophagy and apoptosis and found that VMP1 down-regulation sensitizes cells to apoptosis and that agents that induce apoptosis down-regulate VMP1. In conclusion, similar to its reported role in other cell types, VMP1 is an important regulator of autophagy in colorectal cell lines. However, in contrast to its role in pancreatic cell lines, in colorectal cancer cells, VMP1-dependent autophagy appears to be pro-survival rather than pro-cell death

  19. Globular Adiponectin Causes Tolerance to LPS-Induced TNF-α Expression via Autophagy Induction in RAW 264.7 Macrophages: Involvement of SIRT1/FoxO3A Axis.

    Science.gov (United States)

    Pun, Nirmala Tilija; Subedi, Amit; Kim, Mi Jin; Park, Pil-Hoon

    2015-01-01

    Adiponectin, an adipokine predominantly produced from adipose tissue, exhibited potent anti-inflammatory properties. In particular, it inhibits production of pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α), in macrophages. Autophagy, an intracellular self-digestion process, has been recently shown to regulate inflammatory responses. In the present study, we investigated the role of autophagy induction in the suppression of Lipopolysaccharide (LPS) -induced TNF-α expression by globular adiponectin (gAcrp) and its potential mechanisms. Herein, we found that gAcrp treatment increased expression of genes related with autophagy, including Atg5 and microtubule-associated protein light chain (LC3B), induced autophagosome formation and autophagy flux in RAW 264.7 macrophages. Similar results were observed in primary macrophages isolated peritoneum of mice. Interestingly, inhibition of autophagy by pretreatment with Bafilomycin A1 or knocking down of LC3B gene restored suppression of TNF-α expression, tumor necrosis factor receptor- associated factor 6 (TRAF6) expression and p38MAPK phosphorylation by gAcrp, implying a critical role of autophagy induction in the development of tolerance to LPS-induced TNF-α expression by gAcrp. We also found that knocking-down of FoxO3A, a forkhead box O member of transcription factor, blocked gAcrp-induced expression of LC3II and Atg5. Moreover, gene silencing of Silent information regulator 1 (SIRT1) blocked both gAcrp-induced nuclear translocation of FoxO3A and LC3II expression. Finally, pretreatment with ROS inhibitors, prevented gAcrp-induced SIRT1 expression and further generated inhibitory effects on gAcrp-induced autophagy, indicating a role of ROS production in gAcrp-induced SIRT1 expression and subsequent autophagy induction. Taken together, these findings indicate that globular adiponectin suppresses LPS-induced TNF-α expression, at least in part, via autophagy activation. Furthermore, SIRT1-FoxO3A

  20. Cerebrolysin reduces amyloid-β deposits, apoptosis and autophagy in the thalamus and improves functional recovery after cortical infarction.

    Science.gov (United States)

    Xing, Shihui; Zhang, Jian; Dang, Chao; Liu, Gang; Zhang, Yusheng; Li, Jingjing; Fan, Yuhua; Pei, Zhong; Zeng, Jinsheng

    2014-02-15

    Focal cerebral infarction causes amyloid-β (Aβ) deposits and secondary thalamic neuronal degeneration. The present study aimed to determine the protective effects of Cerebrolysin on Aβ deposits and secondary neuronal damage in thalamus after cerebral infarction. At 24h after distal middle cerebral artery occlusion (MCAO), Cerebrolysin (5 ml/kg) or saline as control was once daily administered for consecutive 13 days by intraperitoneal injection. Sensory function and secondary thalamic damage were assessed with adhesive-removal test, Nissl staining and immunofluorescence at 14 days after MCAO. Aβ deposits, activity of β-site amyloid precursor protein-cleaving enzyme 1 (BACE1), apoptosis and autophagy were determined by TUNEL staining, immunofluorescence and immunoblot. The results showed that Cerebrolysin significantly improved sensory deficit compared to controls (pCerebrolysin, which was accompanied by decreases in neuronal loss and astroglial activation compared to controls (all p Cerebrolysin markedly inhibited cleaved caspase-3, conversion of LC3-II, downregulation of Bcl-2 and upregulation of Bax in the ipsilateral thalamus compared to controls (all pCerebrolysin reduces Aβ deposits, apoptosis and autophagy in the ipsilateral thalamus, which may be associated with amelioration of secondary thalamic damage and functional recovery after cerebral infarction. PMID:24315581

  1. Oxygen-glucose deprivation regulates BACE1 expression through induction of autophagy in Neuro-2a/APP695 cells

    Institute of Scientific and Technical Information of China (English)

    Rong-fu Chen; Xiao-jiang Sun; Ting Zhang; Yin-yi Sun; Ya-meng Sun; Wen-qi Chen; Nan Shi; Fang Shen; Yan Zhang; Kang-yong Liu

    2015-01-01

    Our previous ifndings have demonstrated that autophagy regulation can alleviate the decline of learning and memory by eliminating deposition of extracellular beta-amyloid peptide (Aβ) in the brain after stroke, but the exact mechanism is unclear. It is presumed that the regulation of beta-site APP-cleaving enzyme 1 (BACE1), the rate-limiting enzyme in metabolism of Aβ, would be a key site. Neuro-2a/amyloid precursor protein 695 (APP695) cell models of cerebral isch-emia were established by oxygen-glucose deprivation to investigate the effects of Rapamycin (an autophagy inducer) or 3-methyladenine (an autophagy inhibitor) on the expression of BACE1. Either oxygen-glucose deprivation or Rapamycin down-regulated the expression of BACE1 while 3-methyladenine up-regulated BACE1 expression. These results confirm that oxygen-glucose deprivation down-regulates BACE1 expression in Neuro-2a/APP695 cells through the introduc-tion of autophagy.

  2. Oxygen-glucose deprivation regulates BACE1 expression through induction of autophagy in Neuro-2a/APP695 cells

    Directory of Open Access Journals (Sweden)

    Rong-fu Chen

    2015-01-01

    Full Text Available Our previous findings have demonstrated that autophagy regulation can alleviate the decline of learning and memory by eliminating deposition of extracellular beta-amyloid peptide (Aβ in the brain after stroke, but the exact mechanism is unclear. It is presumed that the regulation of beta-site APP-cleaving enzyme 1 (BACE1, the rate-limiting enzyme in metabolism of Aβ, would be a key site. Neuro-2a/amyloid precursor protein 695 (APP695 cell models of cerebral ischemia were established by oxygen-glucose deprivation to investigate the effects of Rapamycin (an autophagy inducer or 3-methyladenine (an autophagy inhibitor on the expression of BACE1. Either oxygen-glucose deprivation or Rapamycin down-regulated the expression of BACE1 while 3-methyladenine up-regulated BACE1 expression. These results confirm that oxygen-glucose deprivation down-regulates BACE1 expression in Neuro-2a/APP695 cells through the introduction of autophagy.

  3. Autophagy contributes to gefitinib-induced glioma cell growth inhibition

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Cheng-Yi [Department of Surgery, Fong-Yuan Hospital, Taichung 420, Taiwan (China); Graduate Institute of Pharmaceutical Science and Technology, Central Taiwan University of Science and Technology, Taichung 406, Taiwan (China); Kuan, Yu-Hsiang [Department of Pharmacology, School of Medicine, Chung Shan Medical University, Taichung 402, Taiwan (China); Department of Pharmacy, Chung Shan Medical University Hospital, Taichung 402, Taiwan (China); Ou, Yen-Chuan; Li, Jian-Ri [Division of Urology, Taichung Veterans General Hospital, Taichung 407, Taiwan (China); Wu, Chih-Cheng [Department of Anesthesiology, Taichung Veterans General Hospital, Taichung 407, Taiwan (China); Department of Financial and Computational Mathematics, Providence University, Taichung 433, Taiwan (China); Pan, Pin-Ho [Department of Pediatrics, Tungs’ Taichung MetroHarbor Hospital, Taichung 435, Taiwan (China); Chen, Wen-Ying [Department of Veterinary Medicine, National Chung Hsing University, Taichung 402, Taiwan (China); Huang, Hsuan-Yi [Department of Surgery, Fong-Yuan Hospital, Taichung 420, Taiwan (China); Chen, Chun-Jung, E-mail: cjchen@vghtc.gov.tw [Department of Medical Research, Taichung Veterans General Hospital, Taichung 407, Taiwan (China); Institute of Biomedical Sciences, National Chung Hsing University, Taichung 402, Taiwan (China); Rong Hsing Research Center for Translational Medicine, National Chung Hsing University, Taichung 402, Taiwan (China); Center for General Education, Tunghai University, Taichung 407, Taiwan (China); Department of Nursing, HungKuang University, Taichung 433, Taiwan (China)

    2014-09-10

    Epidermal growth factor receptor tyrosine kinase inhibitors, including gefitinib, have been evaluated in patients with malignant gliomas. However, the molecular mechanisms involved in gefitinib-mediated anticancer effects against glioma are incompletely understood. In the present study, the cytostatic potential of gefitinib was demonstrated by the inhibition of glioma cell growth, long-term clonogenic survival, and xenograft tumor growth. The cytostatic consequences were accompanied by autophagy, as evidenced by monodansylcadaverine staining of acidic vesicle formation, conversion of microtubule-associated protein-1 light chain 3-II (LC3-II), degradation of p62, punctate pattern of GFP-LC3, and conversion of GFP-LC3 to cleaved-GFP. Autophagy inhibitor 3-methyladenosine and chloroquine and genetic silencing of LC3 or Beclin 1 attenuated gefitinib-induced growth inhibition. Gefitinib-induced autophagy was not accompanied by the disruption of the Akt/mammalian target of rapamycin signaling. Instead, the activation of liver kinase-B1/AMP-activated protein kinase (AMPK) signaling correlated well with the induction of autophagy and growth inhibition caused by gefitinib. Silencing of AMPK suppressed gefitinib-induced autophagy and growth inhibition. The crucial role of AMPK activation in inducing glioma autophagy and growth inhibition was further supported by the actions of AMP mimetic AICAR. Gefitinib was shown to be capable of reducing the proliferation of glioma cells, presumably by autophagic mechanisms involving AMPK activation. - Highlights: • Gefitinib causes cytotoxic and cytostatic effect on glioma. • Gefitinib induces autophagy. • Gefitinib causes cytostatic effect through autophagy. • Gefitinib induces autophagy involving AMPK.

  4. Autophagy contributes to gefitinib-induced glioma cell growth inhibition

    International Nuclear Information System (INIS)

    Epidermal growth factor receptor tyrosine kinase inhibitors, including gefitinib, have been evaluated in patients with malignant gliomas. However, the molecular mechanisms involved in gefitinib-mediated anticancer effects against glioma are incompletely understood. In the present study, the cytostatic potential of gefitinib was demonstrated by the inhibition of glioma cell growth, long-term clonogenic survival, and xenograft tumor growth. The cytostatic consequences were accompanied by autophagy, as evidenced by monodansylcadaverine staining of acidic vesicle formation, conversion of microtubule-associated protein-1 light chain 3-II (LC3-II), degradation of p62, punctate pattern of GFP-LC3, and conversion of GFP-LC3 to cleaved-GFP. Autophagy inhibitor 3-methyladenosine and chloroquine and genetic silencing of LC3 or Beclin 1 attenuated gefitinib-induced growth inhibition. Gefitinib-induced autophagy was not accompanied by the disruption of the Akt/mammalian target of rapamycin signaling. Instead, the activation of liver kinase-B1/AMP-activated protein kinase (AMPK) signaling correlated well with the induction of autophagy and growth inhibition caused by gefitinib. Silencing of AMPK suppressed gefitinib-induced autophagy and growth inhibition. The crucial role of AMPK activation in inducing glioma autophagy and growth inhibition was further supported by the actions of AMP mimetic AICAR. Gefitinib was shown to be capable of reducing the proliferation of glioma cells, presumably by autophagic mechanisms involving AMPK activation. - Highlights: • Gefitinib causes cytotoxic and cytostatic effect on glioma. • Gefitinib induces autophagy. • Gefitinib causes cytostatic effect through autophagy. • Gefitinib induces autophagy involving AMPK

  5. Conditional Expression of Oncogenic C-RAF in Mouse Pulmonary Epithelial Cells Reveals Differential Tumorigenesis and Induction of Autophagy Leading to Tumor Regression

    Directory of Open Access Journals (Sweden)

    Fatih Ceteci

    2011-11-01

    Full Text Available Here we describe a novel conditional mouse lung tumor model for investigation of the pathogenesis of human lung cancer. On the basis of the frequent involvement of the Ras-RAF-MEK-ERK signaling pathway in human non–small cell lung carcinoma (NSCLC, we have explored the target cell availability, reversibility, and cell type specificity of transformation by oncogenic C-RAF. Targeting expression to alveolar type II cells or to Clara cells, the two likely precursors of human NSCLC, revealed differential tumorigenicity between these cells. Whereas expression of oncogenic C-RAF in alveolar type II cells readily induced multifocal macroscopic lung tumors independent of the developmental state, few tumors with type II pneumocytes features and incomplete penetrance were found when targeted to Clara cells. Induced tumors did not progress and were strictly dependent on the initiating oncogene. Deinduction of mice resulted in tumor regression due to autophagy rather than apoptosis. Induction of autophagic cell death in regressing lung tumors suggests the use of autophagy enhancers as a treatment choice for patients with NSCLC.

  6. Autophagy: Friend or Foe in Breast Cancer Development, Progression, and Treatment

    Directory of Open Access Journals (Sweden)

    Damian E. Berardi

    2011-01-01

    Although autophagy inhibition, combined with anticancer agents, could be therapeutically beneficial in some cases, autophagy induction by itself could lead to cell death in some apoptosis-resistant cancers, indicating that autophagy induction may also be used as a therapy. This paper summarizes the most important findings described in the literature about autophagy and also discusses the importance of this process in clinical settings.

  7. Disrupted cell cycle arrest and reduced proliferation in corneal fibroblasts from GCD2 patients: A potential role for altered autophagy flux

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Seung-il; Dadakhujaev, Shorafidinkhuja; Maeng, Yong-Sun; Ahn, So-yeon; Kim, Tae-im [Department of Ophthalmology, Severance Hospital, Yonsei University College of Medicine, Seoul (Korea, Republic of); Corneal Dystrophy Research Institute, Yonsei University College of Medicine, Seoul (Korea, Republic of); Kim, Eung Kweon, E-mail: eungkkim@yuhs.ac [Department of Ophthalmology, Severance Hospital, Yonsei University College of Medicine, Seoul (Korea, Republic of); Corneal Dystrophy Research Institute, Yonsei University College of Medicine, Seoul (Korea, Republic of); BK21 Plus Project for Medical Science and Severance Biomedical Science Institute, Yonsei University College of Medicine, Seoul (Korea, Republic of)

    2015-01-02

    Highlights: • Reduced cell proliferation in granular corneal dystrophy type 2. • Abnormal cell cycle arrest by defective autophagy. • Decreased Cyclin A1, B1, and D1 in Atg7 gene knockout cells. • Increase in p16 and p27 expressions were observed in Atg7 gene knockout cells. - Abstract: This study investigates the role of impaired proliferation, altered cell cycle arrest, and defective autophagy flux of corneal fibroblasts in granular corneal dystrophy type 2 (GCD2) pathogenesis. The proliferation rates of homozygous (HO) GCD2 corneal fibroblasts at 72 h, 96 h, and 120 h were significantly lower (1.102 ± 0.027, 1.397 ± 0.039, and 1.527 ± 0.056, respectively) than those observed for the wild-type (WT) controls (1.441 ± 0.029, 1.758 ± 0.043, and 2.003 ± 0.046, respectively). Flow cytometry indicated a decreased G{sub 1} cell cycle progression and the accumulation of cells in the S and G{sub 2}/M phases in GCD2 cells. These accumulations were associated with decreased levels of Cyclin A1, B1, and E1, and increased expression of p16 and p27. p21 and p53 expression was also significantly lower in GCD2 cells compared to the WT. Interestingly, treatment with the autophagy flux inhibitor, bafilomycin A{sub 1}, resulted in similarly decreased Cyclin A1, B1, D1, and p53 expression in WT fibroblasts. Furthermore, similar findings, including a decrease in Cyclin A1, B1, and D1 and an increase in p16 and p27 expression were observed in autophagy-related 7 (Atg7; known to be essential for autophagy) gene knockout cells. These data provide new insight concerning the role of autophagy in cell cycle arrest and cellular proliferation, uncovering a number of novel therapeutic possibilities for GCD2 treatment.

  8. Disrupted cell cycle arrest and reduced proliferation in corneal fibroblasts from GCD2 patients: A potential role for altered autophagy flux

    International Nuclear Information System (INIS)

    Highlights: • Reduced cell proliferation in granular corneal dystrophy type 2. • Abnormal cell cycle arrest by defective autophagy. • Decreased Cyclin A1, B1, and D1 in Atg7 gene knockout cells. • Increase in p16 and p27 expressions were observed in Atg7 gene knockout cells. - Abstract: This study investigates the role of impaired proliferation, altered cell cycle arrest, and defective autophagy flux of corneal fibroblasts in granular corneal dystrophy type 2 (GCD2) pathogenesis. The proliferation rates of homozygous (HO) GCD2 corneal fibroblasts at 72 h, 96 h, and 120 h were significantly lower (1.102 ± 0.027, 1.397 ± 0.039, and 1.527 ± 0.056, respectively) than those observed for the wild-type (WT) controls (1.441 ± 0.029, 1.758 ± 0.043, and 2.003 ± 0.046, respectively). Flow cytometry indicated a decreased G1 cell cycle progression and the accumulation of cells in the S and G2/M phases in GCD2 cells. These accumulations were associated with decreased levels of Cyclin A1, B1, and E1, and increased expression of p16 and p27. p21 and p53 expression was also significantly lower in GCD2 cells compared to the WT. Interestingly, treatment with the autophagy flux inhibitor, bafilomycin A1, resulted in similarly decreased Cyclin A1, B1, D1, and p53 expression in WT fibroblasts. Furthermore, similar findings, including a decrease in Cyclin A1, B1, and D1 and an increase in p16 and p27 expression were observed in autophagy-related 7 (Atg7; known to be essential for autophagy) gene knockout cells. These data provide new insight concerning the role of autophagy in cell cycle arrest and cellular proliferation, uncovering a number of novel therapeutic possibilities for GCD2 treatment

  9. Anticancer activity of taraxerol acetate in human glioblastoma cells and a mouse xenograft model via induction of autophagy and apoptotic cell death, cell cycle arrest and inhibition of cell migration

    Science.gov (United States)

    HONG, JING-FANG; SONG, YING-FANG; LIU, ZHENG; ZHENG, ZHAO-CONG; CHEN, HONG-JIE; WANG, SHOU-SEN

    2016-01-01

    The aim of the present study was to investigate the in vitro and in vivo anticancer and apoptotic effects of taraxerol acetate in U87 human glioblastoma cells. The effects on cell cycle phase distribution, cell cycle-associated proteins, autophagy, DNA fragmentation and cell migration were assessed. Cell viability was determined using the MTT assay, and phase contrast and fluorescence microscopy was utilized to determine the viability and apoptotic morphological features of the U87 cells. Flow cytometry using propidium iodide and Annexin V-fluorescein isothiocyanate demonstrated the effect of taraxerol acetate on the cell cycle phase distribution and apoptosis induction. Western blot analysis was performed to investigate the effect of the taraxerol acetate on cell cycle-associated proteins and autophagy-linked LC3B-II proteins. The results demonstrated that taraxerol acetate induced dose- and time-dependent cytotoxic effects in the U87 cells. Apoptotic induction following taraxerol acetate treatment was observed and the percentage of apoptotic cells increased from 7.3% in the control cells, to 16.1, 44.1 and 76.7% in the 10, 50 and 150 µM taraxerol acetate-treated cells, respectively. Furthermore, taraxerol acetate treatment led to sub-G1 cell cycle arrest with a corresponding decrease in the number of S-phase cells. DNA fragments were observed as a result of the gel electrophoresis experiment following taraxerol acetate treatment. To investigate the inhibitory effects of taraxerol acetate on the migration of U87 cell, a wound healing assay was conducted. The number of cells that migrated to the scratched area decreased significantly following treatment with taraxerol acetate. In addition, taraxerol acetate inhibited tumor growth in a mouse xenograft model. Administration of 0.25 and 0.75 µg/g taraxerol acetate reduced the tumor weight from 1.2 g in the phosphate-buffered saline (PBS)-treated group (control) to 0.81 and 0.42 g, respectively. Similarly, 0.25 and 0

  10. 7-Ketocholesterol Induces Autophagy in Vascular Smooth Muscle Cells through Nox4 and Atg4B

    OpenAIRE

    He, Chaoyong; Zhu, Huaiping; Zhang, Wencheng; Okon, Imoh; Wang, Qilong; Li, Hongliang; Le, Yun-Zheng; Xie, Zhonglin

    2013-01-01

    Oxidized lipoproteins stimulate autophagy in advanced atherosclerotic plaques. However, the mechanisms underlying autophagy induction and the role of autophagy in atherogenesis remain to be determined. This study was designed to investigate the mechanisms by which 7-ketocholesterol (7-KC), a major component of oxidized lipoproteins, induces autophagy. This study was also designed to determine the effect of autophagy induction on apoptosis, a central event in the development of atherosclerosis...

  11. IL-1 receptor blockade restores autophagy and reduces inflammation in chronic granulomatous disease in mice and in humans.

    Science.gov (United States)

    de Luca, Antonella; Smeekens, Sanne P; Casagrande, Andrea; Iannitti, Rossana; Conway, Kara L; Gresnigt, Mark S; Begun, Jakob; Plantinga, Theo S; Joosten, Leo A B; van der Meer, Jos W M; Chamilos, Georgios; Netea, Mihai G; Xavier, Ramnik J; Dinarello, Charles A; Romani, Luigina; van de Veerdonk, Frank L

    2014-03-01

    Patients with chronic granulomatous disease (CGD) have a mutated NADPH complex resulting in defective production of reactive oxygen species; these patients can develop severe colitis and are highly susceptible to invasive fungal infection. In NADPH oxidase-deficient mice, autophagy is defective but inflammasome activation is present despite lack of reactive oxygen species production. However, whether these processes are mutually regulated in CGD and whether defective autophagy is clinically relevant in patients with CGD is unknown. Here, we demonstrate that macrophages from CGD mice and blood monocytes from CGD patients display minimal recruitment of microtubule-associated protein 1 light chain 3 (LC3) to phagosomes. This defect in autophagy results in increased IL-1β release. Blocking IL-1 with the receptor antagonist (anakinra) decreases neutrophil recruitment and T helper 17 responses and protects CGD mice from colitis and also from invasive aspergillosis. In addition to decreased inflammasome activation, anakinra restored autophagy in CGD mice in vivo, with increased Aspergillus-induced LC3 recruitment and increased expression of autophagy genes. Anakinra also increased Aspergillus-induced LC3 recruitment from 23% to 51% (P CGD patients. The clinical relevance of these findings was assessed by treating CGD patients who had severe colitis with IL-1 receptor blockade using anakinra. Anakinra treatment resulted in a rapid and sustained improvement in colitis. Thus, inflammation in CGD is due to IL-1-dependent mechanisms, such as decreased autophagy and increased inflammasome activation, which are linked pathological conditions in CGD that can be restored by IL-1 receptor blockade. PMID:24550444

  12. Effects of the Total Saponins from Rosa laevigata Michx Fruit against Acetaminophen-Induced Liver Damage in Mice via Induction of Autophagy and Suppression of Inflammation and Apoptosis

    Directory of Open Access Journals (Sweden)

    Deshi Dong

    2014-05-01

    Full Text Available The effect of the total saponins from Rosa laevigata Michx fruit (RLTS against acetaminophen (APAP-induced liver damage in mice was evaluated in the present paper. The results showed that RLTS markedly improved the levels of liver SOD, CAT, GSH, GSH-Px, MDA, NO and iNOS, and the activities of serum ALT and AST caused by APAP. Further research confirmed that RLTS prevented fragmentation of DNA and mitochondrial ultrastructural alterations based on TdT-mediated dUTP nick end labeling (TUNEL and transmission electron microscopy (TEM assays. In addition, RLTS decreased the gene or protein expressions of cytochrome P450 (CYP2E1, pro-inflammatory mediators (IL-1β, IL-4, IL-6, TNF-α, iNOS, Bax, HMGB-1 and COX-2, pro-inflammatory transcription factors (NF-κB and AP-1, pro-apoptotic proteins (cytochrome C, p53, caspase-3, caspase-9, p-JNK, p-p38 and p-ERK, and increased the protein expressions of Bcl-2 and Bcl-xL. Moreover, the gene expression of IL-10, and the proteins including LC3, Beclin-1 and Atg5 induced by APAP were even more augmented by the extract. These results demonstrate that RLTS has hepatoprotective effects through antioxidative action, induction of autophagy, and suppression of inflammation and apoptosis, and could be developed as a potential candidate to treat APAP-induced liver damage in the future.

  13. Poldip2 knockout results in perinatal lethality, reduced cellular growth and increased autophagy of mouse embryonic fibroblasts.

    Directory of Open Access Journals (Sweden)

    David I Brown

    Full Text Available Polymerase-δ interacting protein 2 (Poldip2 is an understudied protein, originally described as a binding partner of polymerase delta and proliferating cell nuclear antigen (PCNA. Numerous roles for Poldip2 have been proposed, including mitochondrial elongation, DNA replication/repair and ROS production via Nox4. In this study, we have identified a novel role for Poldip2 in regulating the cell cycle. We used a Poldip2 gene-trap mouse and found that homozygous animals die around the time of birth. Poldip2-/- embryos are significantly smaller than wild type or heterozygous embryos. We found that Poldip2-/- mouse embryonic fibroblasts (MEFs exhibit reduced growth as measured by population doubling and growth curves. This effect is not due to apoptosis or senescence; however, Poldip2-/- MEFs have higher levels of the autophagy marker LC3b. Measurement of DNA content by flow cytometry revealed an increase in the percentage of Poldip2-/- cells in the G1 and G2/M phases of the cell cycle, accompanied by a decrease in the percentage of S-phase cells. Increases in p53 S20 and Sirt1 were observed in passage 2 Poldip2-/- MEFs. In passage 4/5 MEFs, Cdk1 and CyclinA2 are downregulated in Poldip2-/- cells, and these changes are reversed by transfection with SV40 large T-antigen, suggesting that Poldip2 may target the E2F pathway. In contrast, p21CIP1 is increased in passage 4/5 Poldip2-/- MEFs and its expression is unaffected by SV40 transfection. Overall, these results reveal that Poldip2 is an essential protein in development, and underline its importance in cell viability and proliferation. Because it affects the cell cycle, Poldip2 is a potential novel target for treating proliferative conditions such as cancer, atherosclerosis and restenosis.

  14. Src/STAT3-dependent heme oxygenase-1 induction mediates chemoresistance of breast cancer cells to doxorubicin by promoting autophagy

    OpenAIRE

    Tan, Qixing; Wang, Hongli; Hu, Yongliang; Hu, Meiru; Li, Xiaoguang;  , Aodengqimuge; Ma, Yuanfang; Wei, Changyuan; Song, Lun

    2015-01-01

    Chemotherapeutic resistance in breast cancer, whether acquired or intrinsic, remains a major clinical obstacle. Thus, increasing tumor cell sensitivity to chemotherapeutic agents will be helpful in improving the clinical management of breast cancer. In the present study, we found an induction of HO-1 expression in doxorubicin (DOX)-treated MDA-MB-231 human breast adenocarcinoma cells, which showed insensitivity to DOX treatment. Knockdown HO-1 expression dramatically upregulated the incidence...

  15. Reduced expression of autophagy markers correlates with high-risk human papillomavirus infection in human cervical squamous cell carcinoma

    Science.gov (United States)

    WANG, HUA-YI; YANG, GUI-FANG; HUANG, YAN-HUA; HUANG, QI-WEN; GAO, JUN; ZHAO, XIAN-DA; HUANG, LI-MING; CHEN, HONG-LEI

    2014-01-01

    Infection by an oncogenic human papillomavirus (HPV), in particular HPV16 and 18, is a high risk factor for developing cervical cancer; however, viral infection alone is not sufficient for cancer progression. Autophagy is hypothesized to be an important process during carcinogenesis. The aim of the present study was to investigate the association between autophagy and high-risk HPV (hrHPV) infection in human cervical squamous cell carcinomas (SCCs), and to analyze the clinical significance of this association. Quantum dot (QD)-based immunofluorescence histochemistry was used to detect the expression of autophagy markers, Beclin-1 and microtubule-associated proteins 1A/1B light chain 3B (LC3B) proteins, in 104 cases of cervical cancer (including 80 SCCs and 24 adenocarcinomas) and 20 normal cervical tissues. hrHPV (HPV16/18) infection was detected by QDs based fluorescence in situ hybridization in cervical cancers. The results revealed that the expression levels of Beclin-1 and LC3B were significantly lower in cervical cancer cells when compared with those of normal cervical squamous epithelial cells, and were found to negatively correlate with hrHPV infection. The expression levels of Beclin-1 and LC3B were not associated with age, tumor grade, tumor stage, tumor node metastasis stage or lymph node metastasis. However, a positive correlation was identified between Beclin-1 and LC3B protein expression. In addition, the absence of autophagy in combination with hrHPV infection may accelerate the progression of cervical SCC. In conclusion, decreased expression of Beclin-1 and LC3B may be important in cervical carcinogenesis. The hrHPV-host cell interaction may inhibit autophagy, which may aid virus duplication and infection, as well as cervical cancer development. PMID:25202355

  16. Melatonin-Mediated Intracellular Insulin during 2-Deoxy-d-glucose Treatment Is Reduced through Autophagy and EDC3 Protein in Insulinoma INS-1E Cells

    Directory of Open Access Journals (Sweden)

    Han Sung Kim

    2016-01-01

    Full Text Available 2-DG triggers glucose deprivation without altering other nutrients or metabolic pathways and then activates autophagy via activation of AMPK and endoplasmic reticulum (ER stress. We investigated whether 2-DG reduced intracellular insulin increased by melatonin via autophagy/EDC3 in insulinoma INS-1E cells. p-AMPK and GRP78/BiP level were significantly increased by 2-DG in the presence/absence of melatonin, but IRE1α level was reduced in 2-DG treatment. Levels of p85α, p110, p-Akt (Ser473, Thr308, and p-mTOR (Ser2481 were also significantly reduced by 2-DG in the presence/absence of melatonin. Mn-SOD increased with 2-DG plus melatonin compared to groups treated with/without melatonin alone. Bcl-2 was decreased and Bax increased with 2-DG plus melatonin. LC3II level increased with 2-DG treatment in the presence/absence of melatonin. Intracellular insulin production increased in melatonin plus 2-DG but reduced in treatment with 2-DG with/without melatonin. EDC3 was increased by 2-DG in the presence/absence of melatonin. Rapamycin, an mTOR inhibitor, increased GRP78/BiP and EDC3 levels in a dose-dependent manner and subsequently resulted in a decrease in intracellular production of insulin. These results suggest that melatonin-mediated insulin synthesis during 2-DG treatment involves autophagy and EDC3 protein in rat insulinoma INS-1E cells and subsequently results in a decrease in intracellular production of insulin.

  17. Tuning the autophagy-inducing activity of lanthanide-based nanocrystals through specific surface-coating peptides

    Science.gov (United States)

    Zhang, Yunjiao; Zheng, Fang; Yang, Tianlong; Zhou, Wei; Liu, Yun; Man, Na; Zhang, Li; Jin, Nan; Dou, Qingqing; Zhang, Yong; Li, Zhengquan; Wen, Long-Ping

    2012-09-01

    The induction of autophagy on exposure of cells to a variety of nanoparticles represents both a safety concern and an application niche for engineered nanomaterials. Here, we show that a short synthetic peptide, RE-1, identified by means of phage display, binds to lanthanide (LN) oxide and upconversion nanocrystals (UCN), forms a stable coating layer on the nanoparticles’ surface, and effectively abrogates their autophagy-inducing activity. Furthermore, RE-1 peptide variants exhibit a differentially reduced binding capability, and correspondingly, a varied ability to reduce the autophagic response. We also show that the addition of an arginine-glycine-aspartic acid (RGD) motif to RE-1 enhances autophagy for LN UCN through the interaction with integrins. RE-1 and its variants provide a versatile tool for tuning material-cell interactions to achieve the desired level of autophagy, and may prove useful for the various diagnostic and therapeutic applications of LN-based nanomaterials and nanodevices.

  18. Autophagy and senescence, stress responses induced by the DNA-damaging mycotoxin alternariol

    International Nuclear Information System (INIS)

    Highlights: • AOH induces autophagy, lamellar bodies and senescence in RAW264.7 macrophages. • DNA damage is suggested as a triggering signal. • The Sestrin2-AMPK-mTOR-S6K pathway is proposed to link DNA damage to autophagy. - Abstract: The mycotoxin alternariol (AOH), a frequent contaminant in fruit and grain, is known to induce cellular stress responses such as reactive oxygen production, DNA damage and cell cycle arrest. Cellular stress is often connected to autophagy, and we employed the RAW264.7 macrophage model to test the hypothesis that AOH induces autophagy. Indeed, AOH treatment led to a massive increase in acidic vacuoles often observed upon autophagy induction. Moreover, expression of the autophagy marker LC3 was markedly increased and there was a strong accumulation of LC3-positive puncta. Increased autophagic activity was verified biochemically by measuring the degradation rate of long-lived proteins. Furthermore, AOH induced expression of Sestrin2 and phosphorylation of AMPK as well as reduced phosphorylation of mTOR and S6 kinase, common mediators of signaling pathways involved in autophagy. Transmission electron microscopy analyzes of AOH treated cells not only clearly displayed structures associated with autophagy such as autophagosomes and autolysosomes, but also the appearance of lamellar bodies. Prolonged AOH treatment resulted in changed cell morphology from round into more star-shaped as well as increased β-galactosidase activity. This suggests that the cells eventually entered senescence. In conclusion, our data identify here AOH as an inducer of both autophagy and senescence. These effects are suggested to be to be linked to AOH-induced DSB (via a reported effect on topoisomerase activity), resulting in an activation of p53 and the Sestrin2-AMPK-mTOR-S6K signaling pathway

  19. Autophagy as a target for cancer therapy: new developments

    International Nuclear Information System (INIS)

    Autophagy is an evolutionarily conserved lysosomal degradation pathway that eliminates cytosolic proteins, macromolecules, organelles, and protein aggregates. Activation of autophagy may function as a tumor suppressor by degrading defective organelles and other cellular components. However, this pathway may also be exploited by cancer cells to generate nutrients and energy during periods of starvation, hypoxia, and stress induced by chemotherapy. Therefore, induction of autophagy has emerged as a drug resistance mechanism that promotes cancer cell survival via self-digestion. Numerous preclinical studies have demonstrated that inhibition of autophagy enhances the activity of a broad array of anticancer agents. Thus, targeting autophagy may be a global anticancer strategy that may improve the efficacy of many standard of care agents. These results have led to multiple clinical trials to evaluate autophagy inhibition in combination with conventional chemotherapy. In this review, we summarize the anticancer agents that have been reported to modulate autophagy and discuss new developments in autophagy inhibition as an anticancer strategy

  20. Anticancer activity of taraxerol acetate in human glioblastoma cells and a mouse xenograft model via induction of autophagy and apoptotic cell death, cell cycle arrest and inhibition of cell migration.

    Science.gov (United States)

    Hong, Jing-Fang; Song, Ying-Fang; Liu, Zheng; Zheng, Zhao-Cong; Chen, Hong-Jie; Wang, Shou-Sen

    2016-06-01

    The aim of the present study was to investigate the in vitro and in vivo anticancer and apoptotic effects of taraxerol acetate in U87 human glioblastoma cells. The effects on cell cycle phase distribution, cell cycle-associated proteins, autophagy, DNA fragmentation and cell migration were assessed. Cell viability was determined using the MTT assay, and phase contrast and fluorescence microscopy was utilized to determine the viability and apoptotic morphological features of the U87 cells. Flow cytometry using propidium iodide and Annexin V-fluorescein isothiocyanate demonstrated the effect of taraxerol acetate on the cell cycle phase distribution and apoptosis induction. Western blot analysis was performed to investigate the effect of the taraxerol acetate on cell cycle‑associated proteins and autophagy‑linked LC3B‑II proteins. The results demonstrated that taraxerol acetate induced dose‑ and time‑dependent cytotoxic effects in the U87 cells. Apoptotic induction following taraxerol acetate treatment was observed and the percentage of apoptotic cells increased from 7.3% in the control cells, to 16.1, 44.1 and 76.7% in the 10, 50 and 150 µM taraxerol acetate‑treated cells, respectively. Furthermore, taraxerol acetate treatment led to sub‑G1 cell cycle arrest with a corresponding decrease in the number of S‑phase cells. DNA fragments were observed as a result of the gel electrophoresis experiment following taraxerol acetate treatment. To investigate the inhibitory effects of taraxerol acetate on the migration of U87 cell, a wound healing assay was conducted. The number of cells that migrated to the scratched area decreased significantly following treatment with taraxerol acetate. In addition, taraxerol acetate inhibited tumor growth in a mouse xenograft model. Administration of 0.25 and 0.75 µg/g taraxerol acetate reduced the tumor weight from 1.2 g in the phosphate‑buffered saline (PBS)‑treated group (control) to 0.81 and 0.42

  1. The Proteasome Inhibitor Bortezomib Maintains Osteocyte Viability in Multiple Myeloma Patients by Reducing Both Apoptosis and Autophagy: A New Function for Proteasome Inhibitors.

    Science.gov (United States)

    Toscani, Denise; Palumbo, Carla; Dalla Palma, Benedetta; Ferretti, Marzia; Bolzoni, Marina; Marchica, Valentina; Sena, Paola; Martella, Eugenia; Mancini, Cristina; Ferri, Valentina; Costa, Federica; Accardi, Fabrizio; Craviotto, Luisa; Aversa, Franco; Giuliani, Nicola

    2016-04-01

    Multiple myeloma (MM) is characterized by severely imbalanced bone remodeling. In this study, we investigated the potential effect of proteasome inhibitors (PIs), a class of drugs known to stimulate bone formation, on the mechanisms involved in osteocyte death induced by MM cells. First, we performed a histological analysis of osteocyte viability on bone biopsies on a cohort of 37 MM patients with symptomatic disease. A significantly higher number of viable osteocytes was detected in patients treated with a bortezomib (BOR)-based regimen compared with those treated without BOR. Interestingly, both osteocyte autophagy and apoptosis were affected in vivo by BOR treatment. Thereafter, we checked the in vitro effect of BOR to understand the mechanisms whereby BOR maintains osteocyte viability in bone from MM patients. We found that osteocyte and preosteocyte autophagic death was triggered during coculturing with MM cells. Our evaluation was conducted by analyzing either autophagy markers microtubule-associated protein light chain 3 beta (LC3B) and SQSTM1/sequestome 1 (p62) levels, or the cell ultrastructure by transmission electron microscopy. PIs were found to increase the basal levels of LC3 expression in the osteocytes while blunting the myeloma-induced osteocyte death. PIs also reduced the autophagic death of osteocytes induced by high-dose dexamethasone (DEX) and potentiated the anabolic effect of PTH(1-34). Our data identify osteocyte autophagy as a new potential target in MM bone disease and support the use of PIs to maintain osteocyte viability and improve bone integrity in MM patients. PMID:26551485

  2. FGF signalling regulates bone growth through autophagy.

    Science.gov (United States)

    Cinque, Laura; Forrester, Alison; Bartolomeo, Rosa; Svelto, Maria; Venditti, Rossella; Montefusco, Sandro; Polishchuk, Elena; Nusco, Edoardo; Rossi, Antonio; Medina, Diego L; Polishchuk, Roman; De Matteis, Maria Antonietta; Settembre, Carmine

    2015-12-10

    Skeletal growth relies on both biosynthetic and catabolic processes. While the role of the former is clearly established, how the latter contributes to growth-promoting pathways is less understood. Macroautophagy, hereafter referred to as autophagy, is a catabolic process that plays a fundamental part in tissue homeostasis. We investigated the role of autophagy during bone growth, which is mediated by chondrocyte rate of proliferation, hypertrophic differentiation and extracellular matrix (ECM) deposition in growth plates. Here we show that autophagy is induced in growth-plate chondrocytes during post-natal development and regulates the secretion of type II collagen (Col2), the major component of cartilage ECM. Mice lacking the autophagy related gene 7 (Atg7) in chondrocytes experience endoplasmic reticulum storage of type II procollagen (PC2) and defective formation of the Col2 fibrillary network in the ECM. Surprisingly, post-natal induction of chondrocyte autophagy is mediated by the growth factor FGF18 through FGFR4 and JNK-dependent activation of the autophagy initiation complex VPS34-beclin-1. Autophagy is completely suppressed in growth plates from Fgf18(-/-) embryos, while Fgf18(+/-) heterozygous and Fgfr4(-/-) mice fail to induce autophagy during post-natal development and show decreased Col2 levels in the growth plate. Strikingly, the Fgf18(+/-) and Fgfr4(-/-) phenotypes can be rescued in vivo by pharmacological activation of autophagy, pointing to autophagy as a novel effector of FGF signalling in bone. These data demonstrate that autophagy is a developmentally regulated process necessary for bone growth, and identify FGF signalling as a crucial regulator of autophagy in chondrocytes. PMID:26595272

  3. Induction of C/EBP homologous protein-mediated apoptosis and autophagy by licochalcone A in non-small cell lung cancer cells.

    Science.gov (United States)

    Tang, Zheng-Hai; Chen, Xin; Wang, Zhao-Yu; Chai, Ke; Wang, Ya-Fang; Xu, Xiao-Huang; Wang, Xiao-Wen; Lu, Jia-Hong; Wang, Yi-Tao; Chen, Xiu-Ping; Lu, Jin-Jian

    2016-01-01

    Licochalcone A (LCA), a flavonoid isolated from the famous Chinese medicinal herb Glycyrrhiza uralensis Fisch, presents obvious anti-cancer effects. In this study, the anti-cancer effects and potential mechanisms of LCA in non-small cell lung cancer (NSCLC) cells were studied. LCA decreased cell viability, increased lactate dehydrogenase release, and induced apoptosis in a concentration-dependent manner in NSCLC cells while not in human embryonic lung fibroblast cells. The expression of phosphatidylethanolamine-modified microtubule-associated protein light-chain 3 (LC3-II) and formation of GFP-LC3 punta, two autophagic markers, were increased after treatment with LCA. LCA-induced LC3-II expression was increased when combined with chloroquine (CQ), while knock-down of autophagy related protein (ATG) 7 or ATG5 reversed LCA-induced LC3-II expression and GFP-LC3 punta formation, suggesting that LCA induced autophagy in NSCLC cells. Inhibition of autophagy could not reverse the LCA-induced cell viability decrease and apoptosis. In addition, LCA increased the expression of endoplasmic reticulum stress related proteins, such as binding immunoglobulin protein and C/EBP homologous protein (CHOP). Knock-down of CHOP reversed LCA-induced cell viability decrease, apoptosis, and autophagy. Taken together, LCA-induced autophagic effect is an accompanied phenomenon in NSCLC cells, and CHOP is critical for LCA-induced cell viability decrease, apoptosis, and autophagy. PMID:27184816

  4. Caspase dependence of the death of neonatal retinal ganglion cells induced by axon damage and induction of autophagy as a survival mechanism

    Directory of Open Access Journals (Sweden)

    C. Sternberg

    2010-10-01

    Full Text Available We examined the degeneration of post-mitotic ganglion cells in ex-vivo neonatal retinal explants following axon damage. Ultrastructural features of both apoptosis and autophagy were detected. Degenerating cells reacted with antibodies specific for activated caspase-3 or -9, consistent with the presence of caspase activity. Furthermore, peptidic inhibitors of caspase-9, -6 or -3 prevented cell death (100 µM Ac-LEDH-CHO, 50 µM Ac-VEID-CHO and 10 µM Z-DEVD-fmk, respectively. Interestingly, inhibition of autophagy by 7-10 mM 3-methyl-adenine increased the rate of cell death. Immunohistochemistry data, caspase activation and caspase inhibition data suggest that axotomy of neonatal retinal ganglion cells triggers the intrinsic apoptotic pathway, which, in turn, is counteracted by a pro-survival autophagic response, demonstrated by electron microscopy profiles and pharmacological autophagy inhibitor.

  5. Reduced models of doubly fed induction generator system for wind turbine simulations

    DEFF Research Database (Denmark)

    Sørensen, Poul Ejnar; Hansen, Anca Daniela; Lund, Torsten; Bindner, Henrik W.

    2005-01-01

    fed induction generator system is very well represented by the dynamics due to the generator inertia and the generator control system, whereas the electromagnetic characteristics of the generator can be represented by the steady state relations. The parameters for the proposed models are derived from......This article compares three reduced models with a detailed model of a doubly fed induction generator system for wind turbine applications. The comparisons are based on simulations only. The main idea is to provide reduced generator models which are appropriate to simulate normal wind turbine...

  6. Quantitative Proteomics Analysis of Macrophage-Derived Lipid Rafts Reveals Induction of Autophagy Pathway at the Early Time of Francisella tularensis LVS Infection

    Czech Academy of Sciences Publication Activity Database

    Hartlová, A.; Link, M.; Balounová, Jana; Benešová, Martina; Resch, U.; Strašková, A.; Sobol, Margaryta; Filimonenko, Anatolij; Hozák, Pavel; Krocová, Z.; Gekara, N.; Filipp, Dominik; Stulík, J.

    2014-01-01

    Roč. 13, č. 2 (2014), s. 796-804. ISSN 1535-3893 R&D Projects: GA MO(CZ) OVUOFVZ200808 Institutional support: RVO:68378050 Keywords : innate immune response * bacterial infection * lipid rafts * Francisella tularensis * phagocytosis * autophagy Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.245, year: 2014

  7. Autophagy in allografts rejection: A new direction?

    Science.gov (United States)

    Sun, Hukui; Cheng, Dayan; Ma, Yuanyuan; Wang, Huaiquan; Liang, Ting; Hou, Guihua

    2016-03-18

    Despite the introduction of new and effective immunosuppressive drugs, acute cellular graft rejection is still a major risk for graft survival. Modulating the dosage of immunosuppressive drugs is not a good choice for all patients, new rejection mechanisms discovery are crucial to limit the inflammatory process and preserve the function of the transplant. Autophagy, a fundamental cellular process, can be detected in all subsets of lymphocytes and freshly isolated naive T lymphocytes. It is required for the homeostasis and function of T lymphocytes, which lead to cell survival or cell death depending on the context. T cell receptor (TCR) stimulation and costimulator signals induce strong autophagy, and autophagy deficient T cells leads to rampant apoptosis upon TCR stimulation. Autophagy has been proved to be activated during ischemia-reperfusion (I/R) injury and associated with grafts dysfunction. Furthermore, Autophagy has also emerged as a key mechanism in orchestrating innate and adaptive immune response to self-antigens, which relates with negative selection and Foxp3(+) Treg induction. Although, the role of autophagy in allograft rejection is unknown, current data suggest that autophagy indeed sweeps across both in the graft organs and recipients lymphocytes after transplantation. This review presents the rationale for the hypothesis that targeting the autophagy pathway could be beneficial in promoting graft survival after transplantation. PMID:26876576

  8. Diabetes and the Brain: Oxidative Stress, Inflammation, and Autophagy

    Directory of Open Access Journals (Sweden)

    María Muriach

    2014-01-01

    Full Text Available Diabetes mellitus is a common metabolic disorder associated with chronic complications including a state of mild to moderate cognitive impairment, in particular psychomotor slowing and reduced mental flexibility, not attributable to other causes, and shares many symptoms that are best described as accelerated brain ageing. A common theory for aging and for the pathogenesis of this cerebral dysfunctioning in diabetes relates cell death to oxidative stress in strong association to inflammation, and in fact nuclear factor κB (NFκB, a master regulator of inflammation and also a sensor of oxidative stress, has a strategic position at the crossroad between oxidative stress and inflammation. Moreover, metabolic inflammation is, in turn, related to the induction of various intracellular stresses such as mitochondrial oxidative stress, endoplasmic reticulum (ER stress, and autophagy defect. In parallel, blockade of autophagy can relate to proinflammatory signaling via oxidative stress pathway and NFκB-mediated inflammation.

  9. Epigallocatechin-gallate (EGCG) regulates autophagy in human retinal pigment epithelial cells: A potential role for reducing UVB light-induced retinal damage

    Energy Technology Data Exchange (ETDEWEB)

    Li, Chao-Peng; Yao, Jin; Tao, Zhi-Fu; Li, Xiu-Miao; Jiang, Qin, E-mail: jqin710@vip.sina.com; Yan, Biao, E-mail: yanbiao1982@hotmail.com

    2013-09-06

    Highlights: •UVB irradiation induces RPE autophagy. •EGCG treatment represses UVB-mediated autophagy. •EGCG regulates UVB-mediated autophagy through mTOR signaling pathway. •EGCG sensitizes RPE cells to UVB-induced damage in an autophagy-dependent manner. -- Abstract: Autophagy is an intracellular catabolic process involved in protein and organelle degradation via the lysosomal pathway that has been linked in the pathogenesis of age-related macular degeneration (AMD). UVB irradiation-mediated degeneration of the macular retinal pigment epithelial (RPE) cells is an important hallmark of AMD, which is along with the change in RPE autophagy. Thus, pharmacological manipulation of RPE autophagy may offer an alternative therapeutic target in AMD. Here, we found that epigallocatechin-3-gallate (EGCG), a polyphenolic compound from green tea, plays a regulatory role in UVB irradiation-induced autophagy in RPE cells. UVB irradiation results in a marked increase in the amount of LC3-II protein in a dose-dependent manner. EGCG administration leads to a significant reduction in the formation of LC3-II and autophagosomes. mTOR signaling activation is required for EGCG-induced LC3-II formation, as evidenced by the fact that EGCG-induced LC3-II formation is significantly impaired by rapamycin administration. Moreover, EGCG significantly alleviates the toxic effects of UVB irradiation on RPE cells in an autophagy-dependent manner. Collectively, our study reveals a novel role of EGCG in RPE autophagy. EGCG may be exploited as a potential therapeutic reagent for the treatment of pathological conditions associated with abnormal autophagy.

  10. Epigallocatechin-gallate (EGCG) regulates autophagy in human retinal pigment epithelial cells: A potential role for reducing UVB light-induced retinal damage

    International Nuclear Information System (INIS)

    Highlights: •UVB irradiation induces RPE autophagy. •EGCG treatment represses UVB-mediated autophagy. •EGCG regulates UVB-mediated autophagy through mTOR signaling pathway. •EGCG sensitizes RPE cells to UVB-induced damage in an autophagy-dependent manner. -- Abstract: Autophagy is an intracellular catabolic process involved in protein and organelle degradation via the lysosomal pathway that has been linked in the pathogenesis of age-related macular degeneration (AMD). UVB irradiation-mediated degeneration of the macular retinal pigment epithelial (RPE) cells is an important hallmark of AMD, which is along with the change in RPE autophagy. Thus, pharmacological manipulation of RPE autophagy may offer an alternative therapeutic target in AMD. Here, we found that epigallocatechin-3-gallate (EGCG), a polyphenolic compound from green tea, plays a regulatory role in UVB irradiation-induced autophagy in RPE cells. UVB irradiation results in a marked increase in the amount of LC3-II protein in a dose-dependent manner. EGCG administration leads to a significant reduction in the formation of LC3-II and autophagosomes. mTOR signaling activation is required for EGCG-induced LC3-II formation, as evidenced by the fact that EGCG-induced LC3-II formation is significantly impaired by rapamycin administration. Moreover, EGCG significantly alleviates the toxic effects of UVB irradiation on RPE cells in an autophagy-dependent manner. Collectively, our study reveals a novel role of EGCG in RPE autophagy. EGCG may be exploited as a potential therapeutic reagent for the treatment of pathological conditions associated with abnormal autophagy

  11. In Vivo Induction of Apoptosis Correlating with Reduced Infectivity during Baculovirus Infection†

    OpenAIRE

    Clarke, Thomas E.; Clem, Rollie J.

    2003-01-01

    Spodoptera frugiperda caterpillars were infected with a mutant of Autographa californica M nucleopolyhedrovirus lacking the antiapoptotic p35 gene. Viral infectivity, replication, and spread were substantially reduced compared to that of a control revertant virus. Terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling confirmed that apoptosis occurred in mutant-infected caterpillars, thus directly correlating reduced infectivity and in vivo induction of apoptosis.

  12. Inhibition of mTOR-dependent autophagy sensitizes leukemic cells to cytarabine-induced apoptotic death.

    Directory of Open Access Journals (Sweden)

    Mihajlo Bosnjak

    Full Text Available The present study investigated the role of autophagy, a cellular self-digestion process, in the cytotoxicity of antileukemic drug cytarabine towards human leukemic cell lines (REH, HL-60, MOLT-4 and peripheral blood mononuclear cells from leukemic patients. The induction of autophagy was confirmed by acridine orange staining of intracellular acidic vesicles, electron microscopy visualization of autophagic vacuoles, as well as by the increase in autophagic proteolysis and autophagic flux, demonstrated by immunoblot analysis of p62 downregulation and LC3-I conversion to autophagosome-associated LC3-II in the presence of proteolysis inhibitors, respectively. Moreover, the expression of autophagy-related genes Atg4, Atg5 and Atg7 was stimulated by cytarabine in REH cells. Cytarabine reduced the phosphorylation of the major negative regulator of autophagy, mammalian target of rapamycin (mTOR, and its downstream target p70S6 kinase in REH cells, which was associated with downregulation of mTOR activator Akt and activation of extracellular signal- regulated kinase. Cytarabine had no effect on the activation of mTOR inhibitor AMP-activated protein kinase. Leucine, an mTOR activator, reduced both cytarabine-induced autophagy and cytotoxicity. Accordingly, pharmacological downregulation of autophagy with bafilomycin A1 and chloroquine, or RNA interference-mediated knockdown of LC3β or p62, markedly increased oxidative stress, mitochondrial depolarization, caspase activation and subsequent DNA fragmentation and apoptotic death in cytarabine-treated REH cells. Cytarabine also induced mTOR-dependent cytoprotective autophagy in HL-60 and MOLT-4 leukemic cell lines, as well as primary leukemic cells, but not normal leukocytes. These data suggest that the therapeutic efficiency of cytarabine in leukemic patients could be increased by the inhibition of the mTOR-dependent autophagic response.

  13. Mimulone-induced autophagy through p53-mediated AMPK/mTOR pathway increases caspase-mediated apoptotic cell death in A549 human lung cancer cells.

    Directory of Open Access Journals (Sweden)

    Hyun-Kyu An

    Full Text Available Anticancer properties and mechanisms of mimulone (MML, C-geranylflavonoid isolated from the Paulownia tomentosa fruits, were firstly elucidated in this study. MML prevented cell proliferation in a dose- and time-dependent way and triggered apoptosis through the extrinsic pathway in A549 human lung adenocarcinoma cells. Furthermore, MML-treated cells displayed autophagic features, such as the formation of autophagic vacuoles, a primary morphological feature of autophagy, and the accumulation of microtubule-associated protein 1 light chain 3 (LC3 puncta, another typical maker of autophagy, as determined by FITC-conjugated immunostaining and monodansylcadaverine (MDC staining, respectively. The expression levels of LC3-I and LC3-II, specific markers of autophagy, were also augmented by MML treatment. Autophagy inhibition by 3-methyladenine (3-MA, pharmacological autophagy inhibitor, and shRNA knockdown of Beclin-1 reduced apoptotic cell death induced by MML. Autophagic flux was not significantly affected by MML treatment and lysosomal inhibitor, chloroquine (CQ suppressed MML-induced autophagy and apoptosis. MML-induced autophagy was promoted by decreases in p53 and p-mTOR levels and increase of p-AMPK. Moreover, inhibition of p53 transactivation by pifithrin-α (PFT-α and knockdown of p53 enhanced induction of autophagy and finally promoted apoptotic cell death. Overall, the results demonstrate that autophagy contributes to the cytotoxicity of MML in cancer cells harboring wild-type p53. This study strongly suggests that MML is a potential candidate for an anticancer agent targeting both autophagy and apoptotic cell death in human lung cancer. Moreover, co-treatment of MML and p53 inhibitor would be more effective in human lung cancer therapy.

  14. Mimulone-induced autophagy through p53-mediated AMPK/mTOR pathway increases caspase-mediated apoptotic cell death in A549 human lung cancer cells.

    Science.gov (United States)

    An, Hyun-Kyu; Kim, Kyoung-Sook; Lee, Ji-Won; Park, Mi-Hyun; Moon, Hyung-In; Park, Shin-Ji; Baik, Ji-Sue; Kim, Cheorl-Ho; Lee, Young-Choon

    2014-01-01

    Anticancer properties and mechanisms of mimulone (MML), C-geranylflavonoid isolated from the Paulownia tomentosa fruits, were firstly elucidated in this study. MML prevented cell proliferation in a dose- and time-dependent way and triggered apoptosis through the extrinsic pathway in A549 human lung adenocarcinoma cells. Furthermore, MML-treated cells displayed autophagic features, such as the formation of autophagic vacuoles, a primary morphological feature of autophagy, and the accumulation of microtubule-associated protein 1 light chain 3 (LC3) puncta, another typical maker of autophagy, as determined by FITC-conjugated immunostaining and monodansylcadaverine (MDC) staining, respectively. The expression levels of LC3-I and LC3-II, specific markers of autophagy, were also augmented by MML treatment. Autophagy inhibition by 3-methyladenine (3-MA), pharmacological autophagy inhibitor, and shRNA knockdown of Beclin-1 reduced apoptotic cell death induced by MML. Autophagic flux was not significantly affected by MML treatment and lysosomal inhibitor, chloroquine (CQ) suppressed MML-induced autophagy and apoptosis. MML-induced autophagy was promoted by decreases in p53 and p-mTOR levels and increase of p-AMPK. Moreover, inhibition of p53 transactivation by pifithrin-α (PFT-α) and knockdown of p53 enhanced induction of autophagy and finally promoted apoptotic cell death. Overall, the results demonstrate that autophagy contributes to the cytotoxicity of MML in cancer cells harboring wild-type p53. This study strongly suggests that MML is a potential candidate for an anticancer agent targeting both autophagy and apoptotic cell death in human lung cancer. Moreover, co-treatment of MML and p53 inhibitor would be more effective in human lung cancer therapy. PMID:25490748

  15. Induction

    DEFF Research Database (Denmark)

    Sprogøe, Jonas; Elkjaer, Bente

    2010-01-01

    The purpose of this paper is to explore how induction of newcomers can be understood as both organizational renewal and the maintenance of status quo, and to develop ways of describing this in terms of learning.......The purpose of this paper is to explore how induction of newcomers can be understood as both organizational renewal and the maintenance of status quo, and to develop ways of describing this in terms of learning....

  16. Inhibition of autophagy enhances DNA damage-induced apoptosis by disrupting CHK1-dependent S phase arrest

    International Nuclear Information System (INIS)

    DNA damage has been shown to induce autophagy, but the role of autophagy in the DNA damage response and cell fate is not fully understood. BO-1012, a bifunctional alkylating derivative of 3a-aza-cyclopenta[a]indene, is a potent DNA interstrand cross-linking agent with anticancer activity. In this study, BO-1012 was found to reduce DNA synthesis, inhibit S phase progression, and induce phosphorylation of histone H2AX on serine 139 (γH2AX) exclusively in S phase cells. Both CHK1 and CHK2 were phosphorylated in response to BO-1012 treatment, but only depletion of CHK1, but not CHK2, impaired BO-1012-induced S phase arrest and facilitated the entry of γH2AX-positive cells into G2 phase. CHK1 depletion also significantly enhanced BO-1012-induced cell death and apoptosis. These results indicate that BO-1012-induced S phase arrest is a CHK1-dependent pro-survival response. BO-1012 also resulted in marked induction of acidic vesicular organelle (AVO) formation and microtubule-associated protein 1 light chain 3 (LC3) processing and redistribution, features characteristic of autophagy. Depletion of ATG7 or co-treatment of cells with BO-1012 and either 3-methyladenine or bafilomycin A1, two inhibitors of autophagy, not only reduced CHK1 phosphorylation and disrupted S phase arrest, but also increased cleavage of caspase-9 and PARP, and cell death. These results suggest that cells initiate S phase arrest and autophagy as pro-survival responses to BO-1012-induced DNA damage, and that suppression of autophagy enhances BO-1012-induced apoptosis via disruption of CHK1-dependent S phase arrest. - Highlights: • Autophagy inhibitors enhanced the cytotoxicity of a DNA alkylating agent, BO-1012. • BO-1012-induced S phase arrest was a CHK1-dependent pro-survival response. • Autophagy inhibition enhanced BO-1012 cytotoxicity via disrupting the S phase arrest

  17. Inhibition of autophagy enhances DNA damage-induced apoptosis by disrupting CHK1-dependent S phase arrest

    Energy Technology Data Exchange (ETDEWEB)

    Liou, Jong-Shian; Wu, Yi-Chen; Yen, Wen-Yen; Tang, Yu-Shuan [Institute of Cellular and Organismic Biology, Academia Sinica, Taipei 115, Taiwan, ROC (China); Kakadiya, Rajesh B.; Su, Tsann-Long [Institute of Biomedical Sciences, Academia Sinica, Taipei 115, Taiwan, ROC (China); Yih, Ling-Huei, E-mail: lhyih@gate.sinica.edu.tw [Institute of Cellular and Organismic Biology, Academia Sinica, Taipei 115, Taiwan, ROC (China)

    2014-08-01

    DNA damage has been shown to induce autophagy, but the role of autophagy in the DNA damage response and cell fate is not fully understood. BO-1012, a bifunctional alkylating derivative of 3a-aza-cyclopenta[a]indene, is a potent DNA interstrand cross-linking agent with anticancer activity. In this study, BO-1012 was found to reduce DNA synthesis, inhibit S phase progression, and induce phosphorylation of histone H2AX on serine 139 (γH2AX) exclusively in S phase cells. Both CHK1 and CHK2 were phosphorylated in response to BO-1012 treatment, but only depletion of CHK1, but not CHK2, impaired BO-1012-induced S phase arrest and facilitated the entry of γH2AX-positive cells into G2 phase. CHK1 depletion also significantly enhanced BO-1012-induced cell death and apoptosis. These results indicate that BO-1012-induced S phase arrest is a CHK1-dependent pro-survival response. BO-1012 also resulted in marked induction of acidic vesicular organelle (AVO) formation and microtubule-associated protein 1 light chain 3 (LC3) processing and redistribution, features characteristic of autophagy. Depletion of ATG7 or co-treatment of cells with BO-1012 and either 3-methyladenine or bafilomycin A1, two inhibitors of autophagy, not only reduced CHK1 phosphorylation and disrupted S phase arrest, but also increased cleavage of caspase-9 and PARP, and cell death. These results suggest that cells initiate S phase arrest and autophagy as pro-survival responses to BO-1012-induced DNA damage, and that suppression of autophagy enhances BO-1012-induced apoptosis via disruption of CHK1-dependent S phase arrest. - Highlights: • Autophagy inhibitors enhanced the cytotoxicity of a DNA alkylating agent, BO-1012. • BO-1012-induced S phase arrest was a CHK1-dependent pro-survival response. • Autophagy inhibition enhanced BO-1012 cytotoxicity via disrupting the S phase arrest.

  18. Insulin receptor substrate-1 prevents autophagy-dependent cell death caused by oxidative stress in mouse NIH/3T3 cells

    Directory of Open Access Journals (Sweden)

    Chan Shih-Hung

    2012-07-01

    Full Text Available Abstract Background Insulin receptor substrate (IRS-1 is associated with tumorigenesis; its levels are elevated in several human cancers. IRS-1 protein binds to several oncogene proteins. Oxidative stress and reactive oxygen species (ROS are involved in the initiation and progression of cancers. Cancer cells produce greater levels of ROS than normal cells do because of increased metabolic stresses. However, excessive production of ROS kills cancer cells. Autophagy usually serves as a survival mechanism in response to stress conditions, but excessive induction of autophagy results in cell death. In addition to inducing necrosis and apoptosis, ROS induces autophagic cell death. ROS inactivates IRS-1 mediated signaling and reduces intracellular IRS-1 concentrations. Thus, there is a complex relationship between IRS-1, ROS, autophagy, and cancer. It is not fully understood how cancer cells grow rapidly and survive in the presence of high ROS levels. Methods and results In this study, we established mouse NIH/3T3 cells that overexpressed IRS-1, so mimicking cancers with increased IRS-1 expression levels; we found that the IRS-1 overexpressing cells grow more rapidly than control cells do. Treatment of cells with glucose oxidase (GO provided a continuous source of ROS; low dosages of GO promoted cell growth, while high doses induced cell death. Evidence for GO induced autophagy includes increased levels of isoform B-II microtubule-associated protein 1 light chain 3 (LC3, aggregation of green fluorescence protein-tagged LC3, and increased numbers of autophagic vacuoles in cells. Overexpression of IRS-1 resulted in inhibition of basal autophagy, and reduced oxidative stress-induced autophagy and cell death. ROS decreased the mammalian target of rapamycin (mTOR/p70 ribosomal protein S6 kinase signaling, while overexpression of IRS-1 attenuated this inhibition. Knockdown of autophagy-related gene 5 inhibited basal autophagy and diminished oxidative stress

  19. Autophagy in response to photodynamic therapy: cell survival vs. cell death

    Science.gov (United States)

    Oleinick, Nancy L.; Xue, Liang-yan; Chiu, Song-mao; Joseph, Sheeba

    2009-02-01

    Autophagy (or more properly, macroautophagy) is a pathway whereby damaged organelles or other cell components are encased in a double membrane, the autophagosome, which fuses with lysosomes for digestion by lysosomal hydrolases. This process can promote cell survival by removing damaged organelles, but when damage is extensive, it can also be a mechanism of cell death. Similar to the Kessel and Agostinis laboratories, we have reported the vigorous induction of autophagy by PDT; this was found in human breast cancer MCF-7 cells whether or not they were able to efficiently induce apoptosis. One way to evaluate the role of autophagy in PDT-treated cells is to silence one of the essential genes in the pathway. Kessel and Reiners silenced the Atg7 gene of murine leukemia L1210 cells using inhibitory RNA and found sensitization to PDT-induced cell death at a low dose of PDT, implying that autophagy is protective when PDT damage is modest. We have examined the role of autophagy in an epithelium-derived cancer cell by comparing parental and Atg7-silenced MCF-7 cells to varying doses of PDT with the phthalocyanine photosensitizer Pc 4. In contrast to L1210 cells, autophagy-deficient MCF-7 cells were more resistant to the lethal effects of PDT, as judged by clonogenic assays. A possible explanation for the difference in outcome for L1210 vs. MCF-7 cells is the greatly reduced ability of the latter to undergo apoptosis, a deficiency that may convert autophagy into a cell-death process even at low PDT doses. Experiments to investigate the mechanism(s) responsible are in process.

  20. Herbivore induction of jasmonic acid and chemical defences reduce photosynthesis in Nicotiana attenuata

    OpenAIRE

    Nabity, Paul D.; Zavala, Jorge A.; DeLucia, Evan H.

    2013-01-01

    Herbivory initiates a shift in plant metabolism from growth to defence that may reduce fitness in the absence of further herbivory. However, the defence-induced changes in carbon assimilation that precede this reallocation in resources remain largely undetermined. This study characterized the response of photosynthesis to herbivore induction of jasmonic acid (JA)-related defences in Nicotiana attenuata to increase understanding of these mechanisms. It was hypothesized that JA-induced defences...

  1. Dihydroptychantol A, a macrocyclic bisbibenzyl derivative, induces autophagy and following apoptosis associated with p53 pathway in human osteosarcoma U2OS cells

    International Nuclear Information System (INIS)

    Dihydroptychantol A (DHA), a novel macrocyclic bisbibenzyl compound extracted from liverwort Asterella angusta, has antifungal and multi-drug resistance reversal properties. Here, the chemically synthesized DHA was employed to test its anti-cancer activities in human osteosarcoma U2OS cells. Our results demonstrated that DHA induced autophagy followed by apoptotic cell death accompanied with G2/M-phase cell cycle arrest in U2OS cells. DHA-induced autophagy was morphologically characterized by the formation of double membrane-bound autophagic vacuoles recognizable at the ultrastructural level. DHA also increased the levels of LC3-II, a marker of autophagy. Surprisingly, DHA-mediated apoptotic cell death was potentiated by the autophagy inhibitor 3-methyladenine, suggesting that autophagy may play a protective role that impedes the eventual cell death. Furthermore, p53 was shown to be involved in DHA-meditated autophagy and apoptosis. In this connection, DHA increased nuclear expression of p53, induced p53 phosphorylation, and upregulated p53 target gene p21Waf1/Cip1. In contrast, cytoplasmic p53 was reduced by DHA, which contributed to the stimulation of autophagy. In relation to the cell cycle, DHA decreased the expression of cyclin B1, a cyclin required for progression through the G2/M phase. Taken together, DHA induces G2/M-phase cell cycle arrest and apoptosis in U2OS cells. DHA-induced apoptosis was preceded by the induction of protective autophagy. DHA-mediated autophagy and apoptosis are associated with the cytoplasmic and nuclear functions of p53.

  2. Outcome of Membrane Sweeping in Reducing Induction Rates in Post-Date Pregnancies

    International Nuclear Information System (INIS)

    Objectives: To determine the effectiveness of membrane sweeping in reducing need for induction of labour in post-date pregnancies and to enlist types and frequencies of complications experienced with membrane sweeping. Study Design: Randomized Control trial. Setting and Duration of Study: The study was carried out at Department of Obstetrics and Gynaecology, Combined Military Hospital, Lahore from February 2007 to April 2008. Patients and Methods: One hundred primi or second gravidas with uncomplicated singleton pregnancies having cephalic presentation at 40+1-5 weeks of gestation were enrolled after informed consent, and divided randomly into two groups of fifty each. Biophysical profile of 8/8 for each case was ensured. Group A underwent membrane sweeping while group B did not. All patients not having spontaneous labour were induced at 40+5 weeks. Data regarding number of patients having spontaneous labour or induction of labour was recorded. Mode of delivery either vaginal or cesarean birth was also recorded. In group A occurence of complications i.e vaginal bleeding or leaking, discomfort, irregular pains, fever and neonatal sepsis was recorded. Results: The difference in rate of spontaneous labor, induction rate and mode of delivery was insignificant between both the groups (p>0.05). In group A, 44% felt discomfort, 4% had bleeding per vaginum, 2% had leaking per vaginum and 28% had more than one complication. There were no cases of maternal or neonatal sepsis. Twenty percent did not have any side effects. Conclusion: Sweeping of membranes is not effective in reducing induction rates in post dates pregnancies. It does not improve the spontaneous labour rate and there is no effect on the mode of delivery. Therefore, any potential benefits of this intervention must be balanced against risk of maternal discomfort and other adverse effects. (author)

  3. Dioscin strengthens the efficiency of adriamycin in MCF-7 and MCF-7/ADR cells through autophagy induction: More than just down-regulation of MDR1.

    Science.gov (United States)

    Wang, Changyuan; Huo, Xiaokui; Wang, Lijuan; Meng, Qiang; Liu, Zhihao; Liu, Qi; Sun, Huijun; Sun, Pengyuan; Peng, Jinyong; Liu, Kexin

    2016-01-01

    The purpose of present study was to investigate the effect of dioscin on activity of adriamycin (ADR) in ADR-sensitive (MCF-7) and ADR-resistant (MCF-7/ADR) human breast cancer cells and to clarify the molecular mechanisms involved. Antiproliferation effect of ADR was enhanced by dioscin in MCF-7 and MCF-7/ADR cells. Dioscin significantly inhibited MDR1 mRNA and protein expression and MDR1 promoter and nuclear factor κ-B (NF-κB) activity in MCF-7/ADR cells. Additionally, inhibitor κB-α (IκB-α) degradation was inhibited by dioscin. Moreover, dioscin induced the formation of vacuoles in the cytoplasm and protein level of LC3-II in MCF-7 and MCF-7/ADR cells. Autophagy inhibitor 3-MA abolished the effect of dioscin on ADR cytotoxicity. Dioscin inhibited phosphorylation of PI3K and Akt, resulting in upregulation of LC3-II expression. In conclusion, dioscin increased ADR chemosensitivity by down-regulating MDR1 expression through NF-κB signaling inhibition in MCF-7/ADR cells. Autophagy was induced by dioscin to ameliorate the cytotoxicity of ADR via inhibition of the PI3K/AKT pathways in MCF-7 and MCF-7/ADR cells. These findings provide evidence in support of further investigation into the clinical application of dioscin as a chemotherapy adjuvant. PMID:27329817

  4. Caloric restriction mimetics: natural/physiological pharmacological autophagy inducers.

    Science.gov (United States)

    Mariño, Guillermo; Pietrocola, Federico; Madeo, Frank; Kroemer, Guido

    2014-01-01

    Nutrient depletion, which is one of the physiological triggers of autophagy, results in the depletion of intracellular acetyl coenzyme A (AcCoA) coupled to the deacetylation of cellular proteins. We surmise that there are 3 possibilities to mimic these effects, namely (i) the depletion of cytosolic AcCoA by interfering with its biosynthesis, (ii) the inhibition of acetyltransferases, which are enzymes that transfer acetyl groups from AcCoA to other molecules, mostly leucine residues in cellular proteins, or (iii) the stimulation of deacetylases, which catalyze the removal of acetyl groups from leucine residues. There are several examples of rather nontoxic natural compounds that act as AcCoA depleting agents (e.g., hydroxycitrate), acetyltransferase inhibitors (e.g., anacardic acid, curcumin, epigallocatechin-3-gallate, garcinol, spermidine) or deacetylase activators (e.g., nicotinamide, resveratrol), and that are highly efficient inducers of autophagy in vitro and in vivo, in rodents. Another common characteristic of these agents is their capacity to reduce aging-associated diseases and to confer protective responses against ischemia-induced organ damage. Hence, we classify them as "caloric restriction mimetics" (CRM). Here, we speculate that CRM may mediate their broad health-improving effects by triggering the same molecular pathways that usually are elicited by long-term caloric restriction or short-term starvation and that imply the induction of autophagy as an obligatory event conferring organismal, organ- or cytoprotection. PMID:25484097

  5. High Ripples Reduction in DTC of Induction Motor by Using a New Reduced Switching Table

    Science.gov (United States)

    Mokhtari, Bachir; Benkhoris, Mohamed F.

    2016-05-01

    The direct torque and flux control (DTC) of electrical motors is characterized by ripples of torque and flux. Among the many solutions proposed to reduce them is to use modified switching tables which is very advantageous; because its implementation is easy and requires no additional cost compared to other solutions. This paper proposes a new reduced switching table (RST) to improve the DTC by reducing harmful ripples of torque and flux. This new switching table is smaller than the conventional one (CST) and depends principally at the flux error. This solution is studied by simulation under Matlab/Simulink and experimentally validated on a testbed with DSPACE1103. The results obtained of a DTC with RST applied to a three-phase induction motor (IM) show a good improvement and an effectiveness of proposed solution, the torque ripple decreases about 47% and 3% for the stator flux compared with a basic DTC.

  6. Autophagy: Friend or Foe in Breast Cancer Development, Progression, and Treatment

    International Nuclear Information System (INIS)

    Autophagy is a catabolic process responsible for the degradation and recycling of long-lived proteins and organelles by lysosomes. This degradative pathway sustains cell survival during nutrient deprivation, but in some circumstances, autophagy leads to cell death. Thereby, autophagy can serve as tumor suppressor, as the reduction in autophagic capacity causes malignant transformation and spontaneous tumors. On the other hand, this process also functions as a protective cell-survival mechanism against environmental stress causing resistance to antineoplastic therapies. Although autophagy inhibition, combined with anticancer agents, could be therapeutically beneficial in some cases, autophagy induction by itself could lead to cell death in some apoptosis-resistant cancers, indicating that autophagy induction may also be used as a therapy. This paper summarizes the most important findings described in the literature about autophagy and also discusses the importance of this process in clinical settings

  7. Peroxisome Proliferator-activated Receptor γ Induces Apoptosis and Inhibits Autophagy of Human Monocyte-derived Macrophages via Induction of Cathepsin L

    Science.gov (United States)

    Mahmood, Dler Faieeq Darweesh; Jguirim-Souissi, Imene; Khadija, El-Hadri; Blondeau, Nicolas; Diderot, Vimala; Amrani, Souliman; Slimane, Mohamed-Naceur; Syrovets, Tatiana; Simmet, Thomas; Rouis, Mustapha

    2011-01-01

    Macrophages play a pivotal role in the pathophysiology of atherosclerosis. These cells express cathepsin L (CatL), a cysteine protease that has been implicated in atherogenesis and the associated arterial remodeling. In addition, macrophages highly express peroxisome proliferator-activated receptor (PPAR) γ, a transcription factor that regulates numerous genes important for lipid and lipoprotein metabolism, for glucose homeostasis, and inflammation. Hence, PPARγ might affect macrophage function in the context of chronic inflammation such as atherogenesis. In the present study, we examined the effect of PPARγ activation on the expression of CatL in human monocyte-derived macrophages (HMDM). Activation of PPARγ by the specific agonist GW929 concentration-dependently increased the levels of CatL mRNA and protein in HMDM. By promoter analysis, we identified a functional PPAR response element-like sequence that positively regulates CatL expression. In addition, we found that PPARγ-induced CatL promotes the degradation of Bcl2 without affecting Bax protein levels. Consistently, degradation of Bcl2 could be prevented by a specific CatL inhibitor, confirming the causative role of CatL. PPARγ-induced CatL was found to decrease autophagy through reduction of beclin 1 and LC3 protein levels. The reduction of these proteins involved in autophagic cell death was antagonized either by the CatL inhibitor or by CatL knockdown. In conclusion, our data show that PPARγ can specifically induce CatL, a proatherogenic protease, in HMDM. In turn, CatL inhibits autophagy and induces apoptosis. Thus, the proatherogenic effect of CatL could be neutralized by apoptosis, a beneficial phenomenon, at least in the early stages of atherosclerosis. PMID:21700710

  8. Hypoxia, MTOR and autophagy

    OpenAIRE

    Blagosklonny, Mikhail V.

    2013-01-01

    Although hypoxia can cause cell cycle arrest, it may simultaneously suppress a conversion from this arrest to senescence. Furthermore, hypoxia can suppress senescence caused by diverse stimuli, maintaining reversible quiescence instead. Hypoxia activates autophagy and inhibits MTOR, thus also activating autophagy. What is the relationship between autophagy and cellular senescence? Also, can inhibition of MTOR and stimulation of autophagy explain the gerosuppressive effects of hypoxia?

  9. Induction of hemeoxygenase-1 reduces renal oxidative stress and inflammation in diabetic spontaneously hypertensive rats.

    Science.gov (United States)

    Elmarakby, Ahmed A; Faulkner, Jessica; Baban, Babak; Sullivan, Jennifer C

    2012-01-01

    The renoprotective mechanisms of hemeoxygenase-1 (HO-1) in diabetic nephropathy remain to be investigated. We hypothesize that HO-1 protects the kidney from diabetic insult via lowering renal oxidative stress and inflammation. We used control and diabetic SHR with or without HO-1 inducer cobalt protoporphyrin (CoPP) treatment for 6 weeks. Urinary albumin excretion levels were significantly elevated in diabetic SHR compared to control and CoPP significantly attenuated albumin excretion. Immuno-histochemical analysis revealed an elevation in TGF-β staining together with increased urinary collagen excretion in diabetic versus control SHR, both of which were reduced with CoPP treatment. Renal oxidative stress markers were greater in diabetic SHR and reduced with CoPP treatment. The increase in renal oxidative stress was associated with an elevation in renal inflammation in diabetic SHR. CoPP treatment also significantly attenuated the markers of renal inflammation in diabetic SHR. In vitro inhibition of HO with stannous mesoporphyrin (SnMP) increased glomerular NADPH oxidase activity and inflammation and blocked the anti-oxidant and anti-inflammatory effects of CoPP. These data suggest that the reduction of renal injury in diabetic SHR upon induction of HO-1 are associated with decreased renal oxidative stress and inflammation, implicating the role of HO-1 induction as a future treatment of diabetic nephropathy. PMID:22518298

  10. Measurement of reduced corkscrew motion on the ETA-II linear induction accelerator

    International Nuclear Information System (INIS)

    The ETA-II linear induction accelerator is used to drive a microwave free-electron laser (FEL). Corkscrew motion, which previously limited performance, has been reduced by: (1) an improved pulse distribution system which reduced energy sweep, (2) improved magnetic alignment achieved with a stretched wire alignment technique (SWAT), and (3) a unique magnetic tuning algorithm. Experiments have been carried out on a 20-cell version of ETA-II operating at 1500 A and 2.7 MeV. The measured transverse beam motion is less than 0.5 mm for 40 ns of the pulse, an improvement of a factor of 2 to 3 over previous results. Details of the computerized tuning procedure, estimates of the corkscrew phase, and relevance of these results to future FEL experiments are presented

  11. Autophagy is required for IL-2-mediated fibroblast growth

    International Nuclear Information System (INIS)

    Autophagy is an evolutionarily conserved pathway responsible for delivery of cytoplasmic material into the lysosomal degradation pathway to enable vesicular exocytosis. Interleukin (IL)-2 is produced by T-cells and its activity is important for immunoregulation. Fibroblasts are an immune competent cell type, playing a critical role in wound healing, chronic inflammation, and tumor development. Although autophagy plays an important role in each of these processes, whether it regulates IL-2 activity in fibroblasts is unknown. Here, we show that autophagy is required for IL-2-induced cell growth in fibroblasts. IL-2 significantly induced autophagy in mouse embryonic fibroblasts (MEFs) and primary lung fibroblasts. Autophagy inhibitors (e.g., 3-methylamphetamine and bafilomycin A1) or knockdown of ATG5 and beclin 1 blocked clinical grade IL-2-induced autophagy. Moreover, IL-2 induced HMGB1 cytoplasmic translocation in MEFs and promoted interaction between HMGB1 and beclin1, which is required for autophagy induction. Pharmacological and genetic inhibition of autophagy inhibited IL-2-induced cell proliferation and enhanced IL-2-induced apoptosis. These findings suggest that autophagy is an important pro-survival regulator for IL-2-induced cell growth in fibroblasts

  12. Autophagy is required for IL-2-mediated fibroblast growth

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Rui [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Tang, Daolin, E-mail: tangd2@upmc.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Lotze, Michael T., E-mail: lotzemt@upcm.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Zeh III, Herbert J., E-mail: zehh@upmc.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States)

    2013-02-15

    Autophagy is an evolutionarily conserved pathway responsible for delivery of cytoplasmic material into the lysosomal degradation pathway to enable vesicular exocytosis. Interleukin (IL)-2 is produced by T-cells and its activity is important for immunoregulation. Fibroblasts are an immune competent cell type, playing a critical role in wound healing, chronic inflammation, and tumor development. Although autophagy plays an important role in each of these processes, whether it regulates IL-2 activity in fibroblasts is unknown. Here, we show that autophagy is required for IL-2-induced cell growth in fibroblasts. IL-2 significantly induced autophagy in mouse embryonic fibroblasts (MEFs) and primary lung fibroblasts. Autophagy inhibitors (e.g., 3-methylamphetamine and bafilomycin A1) or knockdown of ATG5 and beclin 1 blocked clinical grade IL-2-induced autophagy. Moreover, IL-2 induced HMGB1 cytoplasmic translocation in MEFs and promoted interaction between HMGB1 and beclin1, which is required for autophagy induction. Pharmacological and genetic inhibition of autophagy inhibited IL-2-induced cell proliferation and enhanced IL-2-induced apoptosis. These findings suggest that autophagy is an important pro-survival regulator for IL-2-induced cell growth in fibroblasts.

  13. PML at Mitochondria-Associated Membranes Is Critical for the Repression of Autophagy and Cancer Development

    Directory of Open Access Journals (Sweden)

    Sonia Missiroli

    2016-08-01

    Full Text Available The precise molecular mechanisms that coordinate apoptosis and autophagy in cancer remain to be determined. Here, we provide evidence that the tumor suppressor promyelocytic leukemia protein (PML controls autophagosome formation at mitochondria-associated membranes (MAMs and, thus, autophagy induction. Our in vitro and in vivo results demonstrate how PML functions as a repressor of autophagy. PML loss promotes tumor development, providing a growth advantage to tumor cells that use autophagy as a cell survival strategy during stress conditions. These findings demonstrate that autophagy inhibition could be paired with a chemotherapeutic agent to develop anticancer strategies for tumors that present PML downregulation.

  14. Autophagy in Inflammatory Diseases

    Directory of Open Access Journals (Sweden)

    Alexander J. S. Choi

    2011-01-01

    Full Text Available Autophagy provides a mechanism for the turnover of cellular organelles and proteins through a lysosome-dependent degradation pathway. During starvation, autophagy exerts a homeostatic function that promotes cell survival by recycling metabolic precursors. Additionally, autophagy can interact with other vital processes such as programmed cell death, inflammation, and adaptive immune mechanisms, and thereby potentially influence disease pathogenesis. Macrophages deficient in autophagic proteins display enhanced caspase-1-dependent proinflammatory cytokine production and the activation of the inflammasome. Autophagy provides a functional role in infectious diseases and sepsis by promoting intracellular bacterial clearance. Mutations in autophagy-related genes, leading to loss of autophagic function, have been implicated in the pathogenesis of Crohn's disease. Furthermore, autophagy-dependent mechanisms have been proposed in the pathogenesis of several pulmonary diseases that involve inflammation, including cystic fibrosis and pulmonary hypertension. Strategies aimed at modulating autophagy may lead to therapeutic interventions for diseases associated with inflammation.

  15. Heme oxygenase-1 induction improves cardiac function following myocardial ischemia by reducing oxidative stress.

    Directory of Open Access Journals (Sweden)

    Yossi Issan

    Full Text Available BACKGROUND: Oxidative stress plays a key role in exacerbating diabetes and cardiovascular disease. Heme oxygenase-1 (HO-1, a stress response protein, is cytoprotective, but its role in post myocardial infarction (MI and diabetes is not fully characterized. We aimed to investigate the protection and the mechanisms of HO-1 induction in cardiomyocytes subjected to hypoxia and in diabetic mice subjected to LAD ligation. METHODS: In vitro: cultured cardiomyocytes were treated with cobalt-protoporphyrin (CoPP and tin protoporphyrin (SnPP prior to hypoxic stress. In vivo: CoPP treated streptozotocin-induced diabetic mice were subjected to LAD ligation for 2/24 h. Cardiac function, histology, biochemical damage markers and signaling pathways were measured. RESULTS: HO-1 induction lowered release of lactate dehydrogenase (LDH and creatine phospho kinase (CK, decreased propidium iodide staining, improved cell morphology and preserved mitochondrial membrane potential in cardiomyocytes. In diabetic mice, Fractional Shortening (FS was lower than non-diabetic mice (35±1%vs.41±2, respectively p<0.05. CoPP-treated diabetic animals improved cardiac function (43±2% p<0.01, reduced CK, Troponin T levels and infarct size compared to non-treated diabetic mice (P<0.01, P<0.001, P<0.01 respectively. CoPP-enhanced HO-1 protein levels and reduced oxidative stress in diabetic animals, as indicated by the decrease in superoxide levels in cardiac tissues and plasma TNFα levels (p<0.05. The increased levels of HO-1 by CoPP treatment after LAD ligation led to a shift of the Bcl-2/bax ratio towards the antiapoptotic process (p<0.05. CoPP significantly increased the expression levels of pAKT and pGSK3β (p<0.05 in cardiomyocytes and in diabetic mice with MI. SnPP abolished CoPP's cardioprotective effects. CONCLUSIONS: HO-1 induction plays a role in cardioprotection against hypoxic damage in cardiomyocytes and in reducing post ischemic cardiac damage in the diabetic heart

  16. Curcumin could reduce the monomer of TTR with Tyr114Cys mutation via autophagy in cell model of familial amyloid polyneuropathy

    Directory of Open Access Journals (Sweden)

    Li H

    2014-10-01

    Full Text Available Hui Li,1,* Yu Zhang,1,* Li Cao,1 Ran Xiong,1 Bei Zhang,1 Li Wu,1 Zongbo Zhao,1 Sheng-Di Chen1,2 1Department of Neurology and Institute of Neurology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, 2Key Laboratory of Stem Cell Biology and Laboratory of Neurodegenerative Diseases, Institute of Health Science, Shanghai Institutes of Biological Sciences, Chinese Academy of Science, and Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China *These authors contributed equally to this work Abstract: Transthyretin (TTR familial amyloid polyneuropathy (FAP is an autosomal ­dominant inherited neurodegenerative disorder caused by various mutations in the transthyretin gene. We aimed to identify the mechanisms underlying TTR FAP with Tyr114Cys (Y114C mutation. Our study showed that TTR Y114C mutation led to an increase in monomeric TTR and impaired autophagy. Treatment with curcumin resulted in a significant decrease of monomeric TTR by recovering autophagy. Our research suggests that impairment of autophagy might be involved in the pathogenesis of TTR FAP with Y114C mutation, and curcumin might be a potential therapeutic approach for TTR FAP. Keywords: curcumin, familial amyloid polyneuropathy, transthyretin, autophagy

  17. Dietary flavonoid fisetin targets caspase-3-deficient human breast cancer MCF-7 cells by induction of caspase-7-associated apoptosis and inhibition of autophagy.

    Science.gov (United States)

    Yang, Pei-Ming; Tseng, Ho-Hsing; Peng, Chih-Wen; Chen, Wen-Shu; Chiu, Shu-Jun

    2012-02-01

    The outcome of producing apoptotic defects in cancer cells is the primary obstacle that limits the therapeutic efficacy of anticancer agents, and hence the development of novel agents targeting novel non-canonical cell death pathways has become an imperative mission for clinical research. Fisetin (3,3',4',7-tetrahydroxyflavone) is a naturally occurring flavonoid commonly found in fruits and vegetables. In this study, we investigated the potential anticancer effects of fisetin on breast cancer cells. The result showed fisetin induced higher cytotoxicity in human breast cancer MCF-7 than in MDA-MB-231 cells otherwise it did not exert any detectable cytotoxicity in non-tumorigenic MCF-10A cells. We found fisetin can trigger a novel form of atypical apoptosis in caspase-3-deficient MCF-7 cells, which was characterized by several apoptotic features, including plasma membrane rupture, mitochondrial depolarization, activation of caspase-7, -8 and -9, and PARP cleavage; however, neither DNA fragmentation and phosphotidylserine (PS) externalization was observed. Although p53 was also activated by fisetin, the fisetin-induced apoptosis was not rescued by the p53 inhibitor pifithrin-α. In contrast, the fisetin-induced apoptosis was abrogated by pan-caspase inhibitor z-VAD-fmk. Furthermore, inhibition of autophagy by fisetin was shown as additional route to prompt anticancer activity in MCF-7 cells. These data allow us to propose that fisetin appears as a new potential anticancer agent which can be applied to develop a clinical protocol of human breast cancers. PMID:21922137

  18. Rcan1-1L overexpression induces mitochondrial autophagy and improves cell survival in angiotensin II-exposed cardiomyocytes

    International Nuclear Information System (INIS)

    Mitochondrial autophagy is an important adaptive stress response and can be modulated by various key molecules. A previous study found that the regulator of calcineurin 1-1L (Rcan1-1L) may regulate mitochondrial autophagy and cause mitochondria degradation in neurocytes. However, the effect of Rcan1-1L on cardiomyocytes has not been determined. In the present study, we aimed to investigate the role of Rcan1-1L in angiotensin II (Ang II)-exposed human cardiomyocytes. Above all, Human adult cardiac myocytes (HACMs) were exposed to 200 nmol/L Ang II for 4 days. Enhanced H2O2 production, cytochrome C release and mitochondrial permeability were observed in these cells, which were blocked by valsartan. Consistently, Ang II exposure significantly reduced cardiomyocyte viability. However, transfection of Rcan1-1L vector promoted cell viability and ameliorated the apoptosis caused by Ang II. Rcan1-1L clearly promoted mitochondrial autophagy in HACMs, with elevated autophagy protein (ATG) 5 and light chain 3 (LC3) expression. Transient mitochondrial biogenesis and reduced cytochrome C release was also induced by Rcan1-1L. Additionally, Rcan1-1L significantly inhibited calcineurin/nuclear factor of activated T cells (NFAT) signaling. We thus conclude that Rcan1-1L may play a protective role in Ang II-treated cardiomyocytes through the induction of mitochondrial autophagy, and may be an alternative method of cardiac protection. - Highlights: • Transfection of Rcan1-1L into HACMs promoted cell viability and reduced apoptosis. • Transfection of Rcan1-1L promoted mitochondrial autophagy in HACMs. • Rcan1-1L inhibited the calcineurin/nuclear factor of activated T cells signaling

  19. Rcan1-1L overexpression induces mitochondrial autophagy and improves cell survival in angiotensin II-exposed cardiomyocytes

    Energy Technology Data Exchange (ETDEWEB)

    Duan, Hongyan; Li, Yongqiang; Yan, Lijie; Yang, Haitao; Wu, Jintao; Qian, Peng; Li, Bing; Wang, Shanling, E-mail: shanglingwang@126.com

    2015-07-01

    Mitochondrial autophagy is an important adaptive stress response and can be modulated by various key molecules. A previous study found that the regulator of calcineurin 1-1L (Rcan1-1L) may regulate mitochondrial autophagy and cause mitochondria degradation in neurocytes. However, the effect of Rcan1-1L on cardiomyocytes has not been determined. In the present study, we aimed to investigate the role of Rcan1-1L in angiotensin II (Ang II)-exposed human cardiomyocytes. Above all, Human adult cardiac myocytes (HACMs) were exposed to 200 nmol/L Ang II for 4 days. Enhanced H{sub 2}O{sub 2} production, cytochrome C release and mitochondrial permeability were observed in these cells, which were blocked by valsartan. Consistently, Ang II exposure significantly reduced cardiomyocyte viability. However, transfection of Rcan1-1L vector promoted cell viability and ameliorated the apoptosis caused by Ang II. Rcan1-1L clearly promoted mitochondrial autophagy in HACMs, with elevated autophagy protein (ATG) 5 and light chain 3 (LC3) expression. Transient mitochondrial biogenesis and reduced cytochrome C release was also induced by Rcan1-1L. Additionally, Rcan1-1L significantly inhibited calcineurin/nuclear factor of activated T cells (NFAT) signaling. We thus conclude that Rcan1-1L may play a protective role in Ang II-treated cardiomyocytes through the induction of mitochondrial autophagy, and may be an alternative method of cardiac protection. - Highlights: • Transfection of Rcan1-1L into HACMs promoted cell viability and reduced apoptosis. • Transfection of Rcan1-1L promoted mitochondrial autophagy in HACMs. • Rcan1-1L inhibited the calcineurin/nuclear factor of activated T cells signaling.

  20. Knockdown of retinoblastoma protein may sensitize glioma cells to cisplatin through inhibition of autophagy.

    Science.gov (United States)

    Liu, Xiangyu; Sun, Kangjian; Wang, Handong; Dai, Yuyuan

    2016-05-01

    Glioblastoma multiforme (GBM) is one of the deadliest forms of cancer due to its limited sensitivity to chemotherapy and radiotherapy. Cisplatin (CCDP) is a widely used chemotherapeutic agent for tumors, but the agent often results in the development of chemo-resistance. In several cancers, cisplatin resistance is associated with autophagy induction. Here, we found that in glioma cells cisplatin treatment induced autophagy. Our data indicates that the autophagy induction plays a critical role in cisplatin resistance of glioma cells, knockdown of RB inhibited autophagy induced by cisplatin, and inhibition of autophagy improved cisplatin-induced apoptosis. It suggests that a combination of autophagy inhibitors with cisplatin may improve the therapeutic efficiency of cisplatin towards GBM with acquired resistance. PMID:27048711

  1. Autophagy: A double-edged sword in Alzheimer's disease

    Indian Academy of Sciences (India)

    Ying-Tsen Tung; Bo-Jeng Wang; Ming-Kuan Hu; Wen-Ming Hsu; Hsinyu Lee; Wei-Pang Huang; Yung-Feng Liao

    2012-03-01

    Autophagy is a major protein degradation pathway that is essential for stress-induced and constitutive protein turnover. Accumulated evidence has demonstrated that amyloid- (A) protein can be generated in autophagic vacuoles, promoting its extracellular deposition in neuritic plaques as the pathological hallmark of Alzheimer’s disease (AD). The molecular machinery for A generation, including APP, APP-C99 and -/-secretases, are all enriched in autophagic vacuoles. The induction of autophagy can be vividly observed in the brain at early stages of sporadic AD and in an AD transgenic mouse model. Accumulated evidence has also demonstrated a neuroprotective role of autophagy in mediating the degradation of aggregated proteins that are causative of various neurodegenerative diseases. Autophagy is thus widely regarded as an intracellular hub for the removal of the detrimental A peptides and Tau aggregates. Nonetheless, compelling data also reveal an unfavorable function of autophagy in facilitating the production of intracellular A. The two faces of autophagy on the homeostasis of A place it in a very unique and intriguing position in ADpathogenesis. This article briefly summarizes seminal discoveries that are shedding new light on the critical and unique roles of autophagy in AD and potential therapeutic approaches against autophagy-elicited AD.

  2. Epigenetic Regulation of Autophagy by the Methyltransferase G9a

    OpenAIRE

    Artal-Martinez de Narvajas, Amaia; Gomez, Timothy S.; Zhang, Jin-San; Mann, Alexander O.; Taoda, Yoshiyuki; Gorman, Jacquelyn A.; Herreros-Villanueva, Marta; Gress, Thomas M; Ellenrieder, Volker; Bujanda, Luis; Kim, Do-Hyung; Kozikowski, Alan P.; Koenig, Alexander; Billadeau, Daniel D.

    2013-01-01

    Macroautophagy is an evolutionarily conserved cellular process involved in the clearance of proteins and organelles. Although the cytoplasmic machinery that orchestrates autophagy induction during starvation, hypoxia, or receptor stimulation has been widely studied, the key epigenetic events that initiate and maintain the autophagy process remain unknown. Here we show that the methyltransferase G9a coordinates the transcriptional activation of key regulators of autophagosome formation by remo...

  3. Autophagy Inhibition to Increase Radiosensitization in Breast Cancer

    OpenAIRE

    Liang, Diana Hwang; El-Zein, Randa; Dave, Bhuvanesh

    2015-01-01

    Currently, many breast cancer patients with localized breast cancer undergo breast-conserving therapy, consisting of local excision followed by radiation therapy. Following radiation therapy, breast cancer cells are noted to undergo induction of autophagy, development of radioresistance, and enrichment of breast cancer stem cell subpopulations. It is hypothesized that inhibition of the cytoprotective autophagy that arises following radiation therapy increases radiosensitivity and confers long...

  4. Selective Autophagy in Drosophila

    Directory of Open Access Journals (Sweden)

    Ioannis P. Nezis

    2012-01-01

    Full Text Available Autophagy is an evolutionarily conserved process of cellular self-eating and is a major pathway for degradation of cytoplasmic material by the lysosomal machinery. Autophagy functions as a cellular response in nutrient starvation, but it is also associated with the removal of protein aggregates and damaged organelles and therefore plays an important role in the quality control of proteins and organelles. Although it was initially believed that autophagy occurs randomly in the cell, during the last years, there is growing evidence that sequestration and degradation of cytoplasmic material by autophagy can be selective. Given the important role of autophagy and selective autophagy in several disease-related processes such as neurodegeneration, infections, and tumorigenesis, it is important to understand the molecular mechanisms of selective autophagy, especially at the organismal level. Drosophila is an excellent genetically modifiable model organism exhibiting high conservation in the autophagic machinery. However, the regulation and mechanisms of selective autophagy in Drosophila have been largely unexplored. In this paper, I will present an overview of the current knowledge about selective autophagy in Drosophila.

  5. Autophagy in infection.

    Science.gov (United States)

    Deretic, Vojo

    2010-04-01

    Autophagy is a ubiquitous eukaryotic cytoplasmic quality and quantity control pathway. The role of autophagy in cytoplasmic homeostasis seamlessly extends to cell-autonomous defense against intracellular microbes. Recent studies also point to fully integrated, multitiered regulatory and effector connections between autophagy and nearly all facets of innate and adaptive immunity. Autophagy in the immune system as a whole confers measured immune responses; on the flip side, suppression of autophagy can lead to inflammation and tissue damage, as evidenced by Crohn's disease predisposition polymorphisms in autophagy basal apparatus (Atg16L) and regulatory (IRGM) genes. Polymorphisms in the IRGM gene in human populations have also been linked to predisposition to tuberculosis. There are several areas of most recent growth: first, links between autophagy regulators and infectious disease predisposition in human populations; second, demonstration of a role for autophagy in infection control in vivo in animal models; third, the definition of specific antiautophagic defenses in highly evolved pathogens; and fourth, recognition of connections between the ubiquitin system and autophagy of bacteria (and interestingly mitochondria, which are incidentally organelles of bacterial evolutionary origin) via a growing list of modifier and adapter proteins including p62/SQSTM1, NDP52, Atg32, Parkin, and Nix/BNIP3L. PMID:20116986

  6. IFNB1/interferon-ß-induced autophagy in MCF-7 breast cancer cells counteracts its proapoptotic function

    DEFF Research Database (Denmark)

    Ambjørn, Malene; Ejlerskov, Patrick; Liu, Yawei; Lees, Michael; Jäättelä, Marja; Navikas, Shohreh

    2013-01-01

    survival pathways leading to treatment resistance. Defects in autophagy, a conserved cellular degradation pathway, are implicated in numerous cancer diseases. Autophagy is induced in response to cancer therapies and can contribute to treatment resistance. While the type II IFN, IFNG, which in many aspects...... differs significantly from type I IFNs, can induce autophagy, no such function for any type I IFN has been reported. We show here that IFNB1 induces autophagy in MCF-7, MDAMB231 and SKBR3 breast cancer cells by measuring the turnover of two autophagic markers, MAP1LC3B/LC3 and SQSTM1/p62. The induction of...... autophagy in MCF-7 cells occurred upstream of the negative regulator of autophagy MTORC1, and autophagosome formation was dependent on the known core autophagy molecule ATG7 and the IFNB1 signaling molecule STAT1. Using siRNA-mediated silencing of several core autophagy molecules and STAT1, we provide...

  7. Autophagy in the control of food intake.

    Science.gov (United States)

    Singh, Rajat

    2012-04-01

    The cellular nutrient sensing apparatus detects nutritional depletion and transmits this information to downstream effectors that generate energy from alternate sources. Autophagy is a crucial catabolic pathway that turns over redundant cytoplasmic components in lysosomes to provide energy to the starved cell. Recent studies have described a role for hypothalamic autophagy in the control of food intake and energy balance. Activated autophagy in hypothalamic neurons during starvation mobilized neuron-intrinsic lipids to generate free fatty acids that increased AgRP levels. AgRP neuron-specific inhibition of autophagy decreased fasting-induced increases in AgRP levels and food intake. Deletion of autophagy in AgRP neurons led to constitutive increases in levels of proopiomelanocortin and its active processed product, α-melanocyte stimulating hormone that contributed to reduced adiposity in these rodents. The current manuscript discusses these new findings and raises additional questions that may help understand how hypothalamic autophagy controls food intake and energy balance. These studies may have implications for designing new therapies against obesity and insulin resistance. PMID:23700515

  8. Role of D-Limonene in autophagy induced by bergamot essential oil in SH-SY5Y neuroblastoma cells.

    Directory of Open Access Journals (Sweden)

    Rossella Russo

    Full Text Available Bergamot (Citrus bergamia, Risso et Poiteau essential oil (BEO is a well characterized, widely used plant extract. BEO exerts anxiolytic, analgesic and neuroprotective activities in rodents through mechanisms that are only partly known and need to be further investigated. To gain more insight into the biological effects of this essential oil, we tested the ability of BEO (0.005-0.03% to modulate autophagic pathways in human SH-SY5Y neuroblastoma cells. BEO-treated cells show increased LC3II levels and appearance of dot-like formations of endogenous LC3 protein that colocalize with the lysosome marker LAMP-1. Autophagic flux assay using bafilomycin A1 and degradation of the specific autophagy substrate p62 confirmed that the observed increase of LC3II levels in BEO-exposed cells is due to autophagy induction rather than to a decreased autophagosomal turnover. Induction of autophagy is an early and not cell-line specific response to BEO. Beside basal autophagy, BEO also enhanced autophagy triggered by serum starvation and rapamycin indicating that the underlying mechanism is mTOR independent. Accordingly, BEO did not affect the phosphorylation of ULK1 (Ser757 and p70(S6K (Thr389, two downstream targets of mTOR. Furthermore, induction of autophagy by BEO is beclin-1 independent, occurs in a concentration-dependent manner and is unrelated to the ability of BEO to induce cell death. In order to identify the active constituents responsible for these effects, the two most abundant monoterpenes found in the essential oil, d-limonene (125-750 µM and linalyl acetate (62.5-375 µM, were individually tested at concentrations comparable to those found in 0.005-0.03% BEO. The same features of stimulated autophagy elicited by BEO were reproduced by D-limonene, which rapidly increases LC3II and reduces p62 levels in a concentration-dependent manner. Linalyl acetate was ineffective in replicating BEO effects; however, it greatly enhanced LC3 lipidation

  9. Role of D-Limonene in autophagy induced by bergamot essential oil in SH-SY5Y neuroblastoma cells.

    Science.gov (United States)

    Russo, Rossella; Cassiano, Maria Gilda Valentina; Ciociaro, Antonella; Adornetto, Annagrazia; Varano, Giuseppe Pasquale; Chiappini, Carlotta; Berliocchi, Laura; Tassorelli, Cristina; Bagetta, Giacinto; Corasaniti, Maria Tiziana

    2014-01-01

    Bergamot (Citrus bergamia, Risso et Poiteau) essential oil (BEO) is a well characterized, widely used plant extract. BEO exerts anxiolytic, analgesic and neuroprotective activities in rodents through mechanisms that are only partly known and need to be further investigated. To gain more insight into the biological effects of this essential oil, we tested the ability of BEO (0.005-0.03%) to modulate autophagic pathways in human SH-SY5Y neuroblastoma cells. BEO-treated cells show increased LC3II levels and appearance of dot-like formations of endogenous LC3 protein that colocalize with the lysosome marker LAMP-1. Autophagic flux assay using bafilomycin A1 and degradation of the specific autophagy substrate p62 confirmed that the observed increase of LC3II levels in BEO-exposed cells is due to autophagy induction rather than to a decreased autophagosomal turnover. Induction of autophagy is an early and not cell-line specific response to BEO. Beside basal autophagy, BEO also enhanced autophagy triggered by serum starvation and rapamycin indicating that the underlying mechanism is mTOR independent. Accordingly, BEO did not affect the phosphorylation of ULK1 (Ser757) and p70(S6K) (Thr389), two downstream targets of mTOR. Furthermore, induction of autophagy by BEO is beclin-1 independent, occurs in a concentration-dependent manner and is unrelated to the ability of BEO to induce cell death. In order to identify the active constituents responsible for these effects, the two most abundant monoterpenes found in the essential oil, d-limonene (125-750 µM) and linalyl acetate (62.5-375 µM), were individually tested at concentrations comparable to those found in 0.005-0.03% BEO. The same features of stimulated autophagy elicited by BEO were reproduced by D-limonene, which rapidly increases LC3II and reduces p62 levels in a concentration-dependent manner. Linalyl acetate was ineffective in replicating BEO effects; however, it greatly enhanced LC3 lipidation triggered by D

  10. Induction of extracytoplasmic function sigma factors in Bacillus subtilis cells with membranes of reduced phosphatidylglycerol content.

    Science.gov (United States)

    Hashimoto, Michihiro; Takahashi, Hiroaki; Hara, Yoshinori; Hara, Hiroshi; Asai, Kei; Sadaie, Yoshito; Matsumoto, Kouji

    2009-06-01

    The Bacillus subtilis gene pgsA, which codes for the phosphatidylglycerophosphate synthase that catalyzes the committed step for the synthesis of phosphatidylglycerol (PG), is essential since Pspac-pgsA cells require IPTG for growth. Removal of the inducer caused a dramatic decrease of PG content in the membranes of cells and retarded growth. At 60 min and 120 min after removal, it was reduced to 14.1% and 8.9% of total lipid, respectively, from an initial content of 28.1%. We conjectured that the activity of some extracytoplasmic function (ECF) sigma factors, most of which are caught and regulated directly by cognate transmembrane anti-sigma factors, are affected by altered lipid composition of the membranes. Induction of the activities of ECF sigma factors (sigma(M) and sigma(V)) was observed after removal of IPTG, though that of sigma(V) was small. But other ECF sigma factors (sigma(W), sigma(X), sigma(Y), sigma(YlaC) and sigma(Z)) and the general stress sigmas sigma(B) and sigma(I) were not induced. Especially sigma(M) was activated strongly with the reduction of PG content and sustained a high level of activity, in contrast to the transient activation in PG normal cells after exposure to high salinity. This study demonstrates a new relationship between the alterations of lipid composition in the membranes and the activation of ECF sigma factors. PMID:19745567

  11. Oxazolone-induced contact hypersensitivity reduces lymphatic drainage but enhances the induction of adaptive immunity.

    Directory of Open Access Journals (Sweden)

    David Aebischer

    Full Text Available Contact hypersensitivity (CHS induced by topical application of haptens is a commonly used model to study dermal inflammatory responses in mice. Several recent studies have indicated that CHS-induced skin inflammation triggers lymphangiogenesis but may negatively impact the immune-function of lymphatic vessels, namely fluid drainage and dendritic cell (DC migration to draining lymph nodes (dLNs. On the other hand, haptens have been shown to exert immune-stimulatory activity by inducing DC maturation. In this study we investigated how the presence of pre-established CHS-induced skin inflammation affects the induction of adaptive immunity in dLNs. Using a mouse model of oxazolone-induced skin inflammation we observed that lymphatic drainage was reduced and DC migration from skin to dLNs was partially compromised. At the same time, a significantly stronger adaptive immune response towards ovalbumin (OVA was induced when immunization had occurred in CHS-inflamed skin as compared to uninflamed control skin. In fact, immunization with sterile OVA in CHS-inflamed skin evoked a delayed-type hypersensitivity (DTH response comparable to the one induced by conventional immunization with OVA and adjuvant in uninflamed skin. Striking phenotypic and functional differences were observed when comparing DCs from LNs draining uninflamed or CHS-inflamed skin. DCs from LNs draining CHS-inflamed skin expressed higher levels of co-stimulatory molecules and MHC molecules, produced higher levels of the interleukin-12/23 p40 subunit (IL-12/23-p40 and more potently induced T cell activation in vitro. Immunization experiments revealed that blockade of IL-12/23-p40 during the priming phase partially reverted the CHS-induced enhancement of the adaptive immune response. Collectively, our findings indicate that CHS-induced skin inflammation generates an overall immune-stimulatory milieu, which outweighs the potentially suppressive effect of reduced lymphatic vessel function.

  12. Reduced Progression of Cardiac Allograft Vasculopathy with Routine Use of Induction Therapy with Basiliximab

    Directory of Open Access Journals (Sweden)

    Ricardo Wang

    2015-01-01

    Full Text Available Abstract Introduction: Cardiac allograft vasculopathy (CAV is a major limitation for long-term survival of patients undergoing heart transplantation (HT. Some immunosuppressants can reduce the risk of CAV. Objectives: The primary objective was to evaluate the variation in the volumetric growth of the intimal layer measured by intracoronary ultrasound (IVUS after 1 year in patients who received basiliximab compared with that in a control group. Methods: Thirteen patients treated at a single center between 2007 and 2009 were analyzed retrospectively. Evaluations were performed with IVUS, measuring the volume of a coronary segment within the first 30 days and 1 year after HT. Vasculopathy was characterized by the volume of the intima of the vessel. Results: Thirteen patients included (7 in the basiliximab group and 6 in the control group. On IVUS assessment, the control group was found to have greater vessel volume (120–185.43 mm3 vs. 127.77–131.32 mm3; p = 0.051. Intimal layer growth (i.e., CAV was also higher in the control group (27.30–49.15 mm3 [∆80%] vs. 20.23–26.69 mm3 [∆33%]; p = 0.015. Univariate regression analysis revealed that plaque volume and prior atherosclerosis of the donor were not related to intima growth (r = 0.15, p = 0.96, whereas positive remodeling was directly proportional to the volumetric growth of the intima (r = 0.85, p < 0.001. Conclusion: Routine induction therapy with basiliximab was associated with reduced growth of the intima of the vessel during the first year after HT.

  13. The Importance of Autophagy Regulation in Breast Cancer Development and Treatment

    Directory of Open Access Journals (Sweden)

    Joanna Magdalena Zarzynska

    2014-01-01

    Full Text Available Breast cancer (BC is a potentially life-threatening malignant tumor that still causes high mortality among women. One of the mechanisms through which cancer development could be controlled is autophagy. This process exerts different effects during the stages of cancer initiation and progression due to the occurring superimposition of signaling pathways of autophagy and carcinogenesis. Chronic inhibition of autophagy or autophagy deficiency promotes cancer, due to instability of the genome and defective cell growth and as a result of cell stress. However, increased induction of autophagy can become a mechanism which allows tumor cells to survive the conditions of hypoxia, acidosis, or chemotherapy. Therefore, in the development of cancer, autophagy is regarded as a double-edged sword. Determination of the molecular mechanisms underlying autophagy regulation and its role in tumorigenesis is an essential component of modern anticancer strategies. Results of scientific studies show that inhibition of autophagy may enhance the effectiveness of currently used anticancer drugs and other therapies (like radiotherapy. However, in some cases, the promotion of autophagy can induce death and, hence, elimination of the cancer cells and reduction of tumor size. This review summarizes the current knowledge on autophagy regulation in BC and up-to-date anticancer strategies correlated with autophagy.

  14. Chemical Inhibition of Autophagy

    DEFF Research Database (Denmark)

    Baek, Eric; Lin Kim, Che; Gyeom Kim, Mi;

    2016-01-01

    Chinese hamster ovary (CHO) cells activate and undergo apoptosis and autophagy for various environmental stresses. Unlike apoptosis, studies on increasing the production of therapeutic proteins in CHO cells by targeting the autophagy pathway are limited. In order to identify the effects of chemic...

  15. Autophagy and cancer

    Institute of Scientific and Technical Information of China (English)

    Si-Zhao; Lu; Duygu; Dee; Harrison-Findik

    2013-01-01

    Autophagy is a homeostatic and evolutionarily conserved mechanism of self-digestion by which the cells degrade and recycle long-lived proteins and excess or damaged organelles.Autophagy is activated in response to both physiological and pathological stimuli including growth factor depletion,energy deficiency or the upregulation of Bcl-2 protein expression.A novel role of autophagy in various cancers has been proposed.Interestingly,evidence that supports both a positive and negative role of autophagy in the pathogenesis of cancer has been reported.As a tumor suppression mechanism,autophagy maintains genome stability,induces senescence and possibly autophagic cell death.On the other hand,autophagy participates in tumor growth and maintenance by supplying metabolic substrate,limiting oxidative stress,and maintaining cancer stem cell population.It has been proposed that the differential roles of autophagy in cancer are disease type and stage specific.In addition,substrate selectivity might be involved in carrying out the specific effect of autophagy in cancer,and represents one of the potential directions for future studies.

  16. Transient overexpression of Werner protein rescues starvation induced autophagy in Werner syndrome cells.

    Science.gov (United States)

    Maity, Jyotirindra; Bohr, Vilhelm A; Laskar, Aparna; Karmakar, Parimal

    2014-12-01

    Reduced autophagy may be associated with normal and pathological aging. Here we report a link between autophagy and Werner protein (WRNp), mutated in Werner syndrome, the human premature aging Werner syndrome (WS). WRN mutant fibroblast AG11395 and AG05229 respond weakly to starvation induced autophagy compared to normal cells. While the fusion of phagosomes with lysosome is normal, WS cells contain fewer autophagy vacuoles. Cellular starvation autophagy in WS cells is restored after transfection with full length WRN. Further, siRNA mediated silencing of WRN in the normal fibroblast cell line WI-38 results in decreased autophagy and altered expression of autophagy related proteins. Thus, our observations suggest that WRN may have a role in controlling autophagy and hereby cellular maintenance. PMID:25257404

  17. Targeting Autophagy Addiction in Cancer

    OpenAIRE

    Mancias, Joseph Douglas; Kimmelman, Alec C

    2011-01-01

    Autophagy inhibition is a novel cancer therapeutic strategy in the early stages of clinical trial testing. The initial rationale for using autophagy inhibition was generated by research revealing that autophagy is upregulated in response to external stresses, including chemotherapy and radiotherapy. Combining autophagy inhibition with agents that induce autophagy as a pro-survival response may therefore increase their therapeutic efficacy. Recent research has shown that some cancer cells, ...

  18. Osteolytic bone metastasis is hampered by impinging on the interplay among autophagy, anoikis and ossification.

    Science.gov (United States)

    Maroni, P; Bendinelli, P; Matteucci, E; Locatelli, A; Nakamura, T; Scita, G; Desiderio, M A

    2014-01-01

    Here we show that the fate of osteolytic bone metastasis depends on the balance among autophagy, anoikis resistance and ossification, and that the hepatocyte growth factor (HGF) signaling pathway seems to have an important role in orchestrating bone colonization. These findings are consistent with the pathophysiology of bone metastasis that is influenced by the cross-talk of supportive and neoplastic cells through molecular signaling networks. We adopted the strategy to target metastasis and stroma with the use of adenovirally expressed NK4 (AdNK4) and Dasatinib to block HGF/Met axis and Src activity. In human bone metastatic 1833 cells, HGF conferred anoikis resistance via Akt and Src activities and HIF-1α induction, leading to Bim isoforms degradation. When Src and Met activities were inhibited with Dasatinib, the Bim isoforms accumulated conferring anoikis sensitivity. The proviability effect of HGF, under low-nutrient stress condition, was related to a faster autophagy deactivation with respect to HGF plus Dasatinib. In the 1833 xenograft model, AdNK4 switched metastasis vasculature to blood lacunae, increasing HIF-1α in metastasis. The combination of AdNK4 plus Dasatinib gave the most relevant results for mice survival, and the following molecular and cellular changes were found to be responsible. In bone metastasis, we observed a hypoxic condition - marked by HIF-1α - and an autophagy failure - marked by p62 without Beclin-1. Then, osteolytic bone metastases were largely prevented, because of autophagy failure in metastasis and ossification in bone marrow, with osteocalcin deposition. The abnormal repair process was triggered by the dysfunctional autophagy/anoikis interplay. In conclusion, the concomitant blockade of HGF/Met axis and Src activity seemed to induce HIF-1α in metastasis, whereas the bone marrow hypoxic response was reduced. As a consequence, anoikis resistance might be hampered favoring, instead, autophagy failure and neoformation of woven

  19. Enhanced Autophagy and Reduced Expression of Cathepsin D Are Related to Autophagic Cell Death in Epstein-Barr Virus-Associated Nasal Natural Killer/T-Cell Lymphomas: An Immunohistochemical Analysis of Beclin-1, LC3, Mitochondria (AE-1), and Cathepsin D in Nasopharyngeal Lymphomas

    International Nuclear Information System (INIS)

    This study investigated autophagy in 37 cases of nasopharyngeal lymphomas including 23 nasal natural killer (NK)/T-cell lymphomas (NKTCL), 3 cytotoxic T-cell lymphomas (cytotoxic-TML) and 9 B-cell lymphomas (BML) by means of antigen-retrieval immunohistochemistry of beclin-1, LC3, mitochondria (AE-1) and cathepsin D. Peculiar necrosis was noted in EBV+ lymphomas comprising 21 NKTCL, 2 cytotoxic-TML and 1 BML. Lymphomas without peculiar necrosis showed high expression of beclin-1, macrogranular cytoplasmal stain of LC3 with sporadic nuclear stain, a hallmark of autophagic cell death (ACD), some aggregated mitochondria and high expression of cathepsin D, suggesting a state of growth with enhanced autophagy with sporadic ACD. EBV+ NKTCL with the peculiar necrosis, showed significantly low level of macrogranular staining of LC3, aggregated mitochondria and low expression of cathepsin D in the cellular areas when degenerative lymphoma cells showed decreased beclin-1, significantly advanced LC3-labeled autophagy, residual aggregated mitochondria and significantly reduced expression of cathepsin D, suggesting advanced autophagy with regional ACD. Consequently it was suggested that enhanced autophagy and reduced expression of lysosomal enzymes induced regional ACD under EBV infection in NKTCL

  20. Nanomaterials and Autophagy: New Insights in Cancer Treatment

    International Nuclear Information System (INIS)

    Autophagy represents a cell’s response to stress. It is an evolutionarily conserved process with diversified roles. Indeed, it controls intracellular homeostasis by degradation and/or recycling intracellular metabolic material, supplies energy, provides nutrients, eliminates cytotoxic materials and damaged proteins and organelles. Moreover, autophagy is involved in several diseases. Recent evidences support a relationship between several classes of nanomaterials and autophagy perturbation, both induction and blockade, in many biological models. In fact, the autophagic mechanism represents a common cellular response to nanomaterials. On the other hand, the dynamic nature of autophagy in cancer biology is an intriguing approach for cancer therapeutics, since during tumour development and therapy, autophagy has been reported to trigger both an early cell survival and a late cell death. The use of nanomaterials in cancer treatment to deliver chemotherapeutic drugs and target tumours is well known. Recently, autophagy modulation mediated by nanomaterials has become an appealing notion in nanomedicine therapeutics, since it can be exploited as adjuvant in chemotherapy or in the development of cancer vaccines or as a potential anti-cancer agent. Herein, we summarize the effects of nanomaterials on autophagic processes in cancer, also considering the therapeutic outcome of synergism between nanomaterials and autophagy to improve existing cancer therapies

  1. The regulation of autophagy during exercise in skeletal muscle.

    Science.gov (United States)

    Vainshtein, Anna; Hood, David A

    2016-03-15

    The merits of exercise on muscle health and well-being are numerous and well documented. However, the mechanisms underlying the robust adaptations induced by exercise, particularly on mitochondria, are less clear and much sought after. Recently, an evolutionary conserved cellular recycling mechanism known as autophagy has been implicated in the adaptations to acute and chronic exercise. A basal level of autophagy is constantly ongoing in cells and tissues, ensuring cellular clearance and energy homeostasis. This pathway can be further induced, as a survival mechanism, by cellular perturbations, such as energetic imbalance and oxidative stress. During exercise, a biphasic autophagy response is mobilized, leading to both an acute induction and a long-term potentiation of the process. Posttranslational modifications arising from upstream signaling cascades induce an acute autophagic response during a single bout of exercise by mobilizing core autophagy machinery. A transcriptional program involving the regulators Forkhead box O, transcription factor EB, p53, and peroxisome proliferator coactivator-1α is also induced to fuel sustained increases in autophagic capacity. Autophagy has also been documented to mediate chronic exercise-induced metabolic benefits, and animal models in which autophagy is perturbed do not adapt to exercise to the same extent. In this review, we discuss recent developments in the field of autophagy and exercise. We specifically highlight the molecular mechanisms activated during acute exercise that lead to a prolonged adaptive response. PMID:26679612

  2. Nanomaterials and Autophagy: New Insights in Cancer Treatment

    Energy Technology Data Exchange (ETDEWEB)

    Panzarini, Elisa; Inguscio, Valentina; Tenuzzo, Bernardetta Anna; Carata, Elisabetta; Dini, Luciana, E-mail: luciana.dini@unisalento.it [Department of Biological and Environmental Science and Technology (Di.S.Te.B.A.), University of Salento, Lecce 73100 (Italy)

    2013-03-21

    Autophagy represents a cell’s response to stress. It is an evolutionarily conserved process with diversified roles. Indeed, it controls intracellular homeostasis by degradation and/or recycling intracellular metabolic material, supplies energy, provides nutrients, eliminates cytotoxic materials and damaged proteins and organelles. Moreover, autophagy is involved in several diseases. Recent evidences support a relationship between several classes of nanomaterials and autophagy perturbation, both induction and blockade, in many biological models. In fact, the autophagic mechanism represents a common cellular response to nanomaterials. On the other hand, the dynamic nature of autophagy in cancer biology is an intriguing approach for cancer therapeutics, since during tumour development and therapy, autophagy has been reported to trigger both an early cell survival and a late cell death. The use of nanomaterials in cancer treatment to deliver chemotherapeutic drugs and target tumours is well known. Recently, autophagy modulation mediated by nanomaterials has become an appealing notion in nanomedicine therapeutics, since it can be exploited as adjuvant in chemotherapy or in the development of cancer vaccines or as a potential anti-cancer agent. Herein, we summarize the effects of nanomaterials on autophagic processes in cancer, also considering the therapeutic outcome of synergism between nanomaterials and autophagy to improve existing cancer therapies.

  3. Autophagy in protists.

    Science.gov (United States)

    Duszenko, Michael; Ginger, Michael L; Brennand, Ana; Gualdrón-López, Melisa; Colombo, María Isabel; Coombs, Graham H; Coppens, Isabelle; Jayabalasingham, Bamini; Langsley, Gordon; de Castro, Solange Lisboa; Menna-Barreto, Rubem; Mottram, Jeremy C; Navarro, Miguel; Rigden, Daniel J; Romano, Patricia S; Stoka, Veronika; Turk, Boris; Michels, Paul A M

    2011-02-01

    Autophagy is the degradative process by which eukaryotic cells digest their own components using acid hydrolases within the lysosome. Originally thought to function almost exclusively in providing starving cells with nutrients taken from their own cellular constituents, autophagy is in fact involved in numerous cellular events including differentiation, turnover of macromolecules and organelles, and defense against parasitic invaders. During the last 10-20 years, molecular components of the autophagic machinery have been discovered, revealing a complex interactome of proteins and lipids, which, in a concerted way, induce membrane formation to engulf cellular material and target it for lysosomal degradation. Here, our emphasis is autophagy in protists. We discuss experimental and genomic data indicating that the canonical autophagy machinery characterized in animals and fungi appeared prior to the radiation of major eukaryotic lineages. Moreover, we describe how comparative bioinformatics revealed that this canonical machinery has been subject to moderation, outright loss or elaboration on multiple occasions in protist lineages, most probably as a consequence of diverse lifestyle adaptations. We also review experimental studies illustrating how several pathogenic protists either utilize autophagy mechanisms or manipulate host-cell autophagy in order to establish or maintain infection within a host. The essentiality of autophagy for the pathogenicity of many parasites, and the unique features of some of the autophagy-related proteins involved, suggest possible new targets for drug discovery. Further studies of the molecular details of autophagy in protists will undoubtedly enhance our understanding of the diversity and complexity of this cellular phenomenon and the opportunities it offers as a drug target. PMID:20962583

  4. Autophagy as a Potential Target for Sarcopenia.

    Science.gov (United States)

    Fan, Jingjing; Kou, Xianjuan; Jia, Shaohui; Yang, Xiaoqi; Yang, Yi; Chen, Ning

    2016-07-01

    Sarcopenia is an aging-related disease with a significant reduction in mass and strength of skeletal muscle due to the imbalance between protein synthesis and protein degradation. The loss of skeletal muscle is an inevitable event during aging process, which can result in the significant impact on the quality of life, and also can increase the risk for other aging-associated diseases in the elderly. However, the underlying molecular mechanism of aging-related skeletal muscle loss is still poorly understood. Autophagy is a degradation pathway for the clearance of dysfunctional organelles and damaged macromolecules during aging process. Appropriate induction or accurate regulation of autophagic process and improved quality control of mitochondria through autophagy or other strategies are required for the maintenance of skeletal muscle mass. In this article, we have summarized the current understanding of autophagic pathways in sarcopenia, and discussed the functional status of autophagy and autophagy-associated quality control of mitochondria in the pathogenesis of sarcopenia. Moreover, this article will provide some theoretical references for the exploration of scientific and optimal intervention strategies such as exercise and caloric restriction for the prevention and treatment of sarcopenia through the regulation of autophagic pathways. PMID:26580995

  5. Spermidine and resveratrol induce autophagy by distinct pathways converging on the acetylproteome

    DEFF Research Database (Denmark)

    Morselli, Eugenia; Mariño, Guillermo; Bennetzen, Martin V; Eisenberg, Tobias; Megalou, Evgenia; Schroeder, Sabrina; Cabrera, Sandra; Bénit, Paule; Rustin, Pierre; Criollo, Alfredo; Kepp, Oliver; Galluzzi, Lorenzo; Shen, Shensi; Malik, Shoaib Ahmad; Maiuri, Maria Chiara; Horio, Yoshiyuki; López-Otín, Carlos; Andersen, Jens S.; Tavernarakis, Nektarios; Madeo, Frank; Kroemer, Guido

    2011-01-01

    Autophagy protects organelles, cells, and organisms against several stress conditions. Induction of autophagy by resveratrol requires the nicotinamide adenine dinucleotide-dependent deacetylase sirtuin 1 (SIRT1). In this paper, we show that the acetylase inhibitor spermidine stimulates autophagy...... independent of SIRT1 in human and yeast cells as well as in nematodes. Although resveratrol and spermidine ignite autophagy through distinct mechanisms, these compounds stimulate convergent pathways that culminate in concordant modifications of the acetylproteome. Both agents favor convergent deacetylation...... and acetylation reactions in the cytosol and in the nucleus, respectively. Both resveratrol and spermidine were able to induce autophagy in cytoplasts (enucleated cells). Moreover, a cytoplasm-restricted mutant of SIRT1 could stimulate autophagy, suggesting that cytoplasmic deacetylation reactions...

  6. The role of autophagy in cell survival from heavy ion irradiation in the plateau region

    International Nuclear Information System (INIS)

    To study cytotoxic effect of heavy ion irradiation in the plateau region, and investigate whether autophagy induced by heavy ion irradiation is cytoprotective, HeLa cells were irradiated with 350 MeV/u carbon ions beams, and the clonogenic survival was analyzed. The results showed that cell survival decreased with increasing doses. It was also found that G2/M-phase cells increased, and the autophagy-related activity was significantly higher than the control. When autophagy was blocked by 3-methyladenine in carbon-ion irradiated cells, G2/M phase arrest and the percentage of apoptosis cells were further elevated, and cell survival decreased significantly, indicating the induction of cytoprotective autophagy by carbon-ion irradiation. Our results demonstrated that autophagy induced by carbon ion irradiation provided a self-protective mechanism in HeLa cells, short-time inhibition of autophagy before carbon-ion irradiation could enhance radiation cytotoxicity in HeLa cells. (authors)

  7. Calcium Homeostasis and ER Stress in Control of Autophagy in Cancer Cells

    Directory of Open Access Journals (Sweden)

    Elżbieta Kania

    2015-01-01

    Full Text Available Autophagy is a basic catabolic process, serving as an internal engine during responses to various cellular stresses. As regards cancer, autophagy may play a tumor suppressive role by preserving cellular integrity during tumor development and by possible contribution to cell death. However, autophagy may also exert oncogenic effects by promoting tumor cell survival and preventing cell death, for example, upon anticancer treatment. The major factors influencing autophagy are Ca2+ homeostasis perturbation and starvation. Several Ca2+ channels like voltage-gated T- and L-type channels, IP3 receptors, or CRAC are involved in autophagy regulation. Glucose transporters, mainly from GLUT family, which are often upregulated in cancer, are also prominent targets for autophagy induction. Signals from both Ca2+ perturbations and glucose transport blockage might be integrated at UPR and ER stress activation. Molecular pathways such as IRE 1-JNK-Bcl-2, PERK-eIF2α-ATF4, or ATF6-XBP 1-ATG are related to autophagy induced through ER stress. Moreover ER molecular chaperones such as GRP78/BiP and transcription factors like CHOP participate in regulation of ER stress-mediated autophagy. Autophagy modulation might be promising in anticancer therapies; however, it is a context-dependent matter whether inhibition or activation of autophagy leads to tumor cell death.

  8. Intestinal Autophagy Improves Healthspan and Longevity in C. elegans during Dietary Restriction.

    Directory of Open Access Journals (Sweden)

    Sara Gelino

    2016-07-01

    Full Text Available Dietary restriction (DR is a dietary regimen that extends lifespan in many organisms. One mechanism contributing to the conserved effect of DR on longevity is the cellular recycling process autophagy, which is induced in response to nutrient scarcity and increases sequestration of cytosolic material into double-membrane autophagosomes for degradation in the lysosome. Although autophagy plays a direct role in DR-mediated lifespan extension in the nematode Caenorhabditis elegans, the contribution of autophagy in individual tissues remains unclear. In this study, we show a critical role for autophagy in the intestine, a major metabolic tissue, to ensure lifespan extension of dietary-restricted eat-2 mutants. The intestine of eat-2 mutants has an enlarged lysosomal compartment and flux assays indicate increased turnover of autophagosomes, consistent with an induction of autophagy in this tissue. This increase in intestinal autophagy may underlie the improved intestinal integrity we observe in eat-2 mutants, since whole-body and intestinal-specific inhibition of autophagy in eat-2 mutants greatly impairs the intestinal barrier function. Interestingly, intestinal-specific inhibition of autophagy in eat-2 mutants leads to a decrease in motility with age, alluding to a potential cell non-autonomous role for autophagy in the intestine. Collectively, these results highlight important functions for autophagy in the intestine of dietary-restricted C. elegans.

  9. PI3K/Akt/mTOR pathway inhibitors enhance radiosensitivity in radioresistant prostate cancer cells through inducing apoptosis, reducing autophagy, suppressing NHEJ and HR repair pathways.

    Science.gov (United States)

    Chang, L; Graham, P H; Hao, J; Ni, J; Bucci, J; Cozzi, P J; Kearsley, J H; Li, Y

    2014-01-01

    The PI3K/Akt/mTOR pathway has a central role in cancer metastasis and radiotherapy. To develop effective therapeutics to improve radiosensitivity, understanding the possible pathways of radioresistance involved and the effects of a combination of the PI3K/Akt/mTOR inhibitors with radiotherapy on prostate cancer (CaP) radioresistant cells is needed. We found that compared with parent CaP cells, CaP-radioresistant cells demonstrated G0/G1 and S phase arrest, activation of cell cycle check point, autophagy and DNA repair pathway proteins, and inactivation of apoptotic proteins. We also demonstrated that compared with combination of single PI3K or mTOR inhibitors (BKM120 or Rapamycin) and radiation, low-dose of dual PI3K/mTOR inhibitors (BEZ235 or PI103) combined with radiation greatly improved treatment efficacy by repressing colony formation, inducing more apoptosis, leading to the arrest of the G2/M phase, increased double-strand break levels and less inactivation of cell cycle check point, autophagy and non-homologous end joining (NHEJ)/homologous recombination (HR) repair pathway proteins in CaP-radioresistant cells. This study describes the possible pathways associated with CaP radioresistance and demonstrates the putative mechanisms of the radiosensitization effect in CaP-resistant cells in the combination treatment. The findings from this study suggest that the combination of dual PI3K/Akt/mTOR inhibitors (BEZ235 or PI103) with radiotherapy is a promising modality for the treatment of CaP to overcome radioresistance. PMID:25275598

  10. Reduced impact of induced gate noise on inductively degenerated LNAs in deep submicron CMOS technologies

    DEFF Research Database (Denmark)

    Rossi, P.; Svelto, F.; Mazzanti, A.;

    2005-01-01

    Designers of radio-frequency inductively-degenerated CMOS low-noise-amplifiers have usually not followed the guidelines for achieving minimum noise figure. Nonetheless, state-of-the- art implementations display noise figure values very close to the theoretical minimum. In this paper, we point out...

  11. Lithium and autophagy.

    Science.gov (United States)

    Motoi, Yumiko; Shimada, Kohei; Ishiguro, Koichi; Hattori, Nobutaka

    2014-06-18

    Lithium, a drug used to treat bipolar disorders, has a variety of neuroprotective mechanisms, including autophagy regulation, in various neuropsychiatric conditions. In neurodegenerative diseases, lithium enhances degradation of aggregate-prone proteins, including mutated huntingtin, phosphorylated tau, and α-synuclein, and causes damaged mitochondria to degrade, while in a mouse model of cerebral ischemia and Alzheimer's disease autophagy downregulation by lithium is observed. The signaling pathway of lithium as an autophagy enhancer might be associated with the mammalian target of rapamycin (mTOR)-independent pathway, which is involved in myo-inositol-1,4,5-trisphosphate (IP3) in Huntington's disease and Parkinson's disease. However, the mTOR-dependent pathway might be involved in inhibiting glycogen synthase kinase-3β (GSK3β) in other diseases. Lithium's autophagy-enhancing property may contribute to the therapeutic benefit of patients with neuropsychiatric disorders. PMID:24738557

  12. Autophagy Inhibition Dysregulates TBK1 Signaling and Promotes Pancreatic Inflammation.

    Science.gov (United States)

    Yang, Shenghong; Imamura, Yu; Jenkins, Russell W; Cañadas, Israel; Kitajima, Shunsuke; Aref, Amir; Brannon, Arthur; Oki, Eiji; Castoreno, Adam; Zhu, Zehua; Thai, Tran; Reibel, Jacob; Qian, Zhirong; Ogino, Shuji; Wong, Kwok K; Baba, Hideo; Kimmelman, Alec C; Pasca Di Magliano, Marina; Barbie, David A

    2016-06-01

    Autophagy promotes tumor progression downstream of oncogenic KRAS, yet also restrains inflammation and dysplasia through mechanisms that remain incompletely characterized. Understanding the basis of this paradox has important implications for the optimal targeting of autophagy in cancer. Using a mouse model of cerulein-induced pancreatitis, we found that loss of autophagy by deletion of Atg5 enhanced activation of the IκB kinase (IKK)-related kinase TBK1 in vivo, associated with increased neutrophil and T-cell infiltration and PD-L1 upregulation. Consistent with this observation, pharmacologic or genetic inhibition of autophagy in pancreatic ductal adenocarcinoma cells, including suppression of the autophagy receptors NDP52 or p62, prolonged TBK1 activation and increased expression of CCL5, IL6, and several other T-cell and neutrophil chemotactic cytokines in vitro Defective autophagy also promoted PD-L1 upregulation, which is particularly pronounced downstream of IFNγ signaling and involves JAK pathway activation. Treatment with the TBK1/IKKε/JAK inhibitor CYT387 (also known as momelotinib) not only inhibits autophagy, but also suppresses this feedback inflammation and reduces PD-L1 expression, limiting KRAS-driven pancreatic dysplasia. These findings could contribute to the dual role of autophagy in oncogenesis and have important consequences for its therapeutic targeting. Cancer Immunol Res; 4(6); 520-30. ©2016 AACR. PMID:27068336

  13. Activation of autophagy by unfolded proteins during endoplasmic reticulum stress.

    Science.gov (United States)

    Yang, Xiaochen; Srivastava, Renu; Howell, Stephen H; Bassham, Diane C

    2016-01-01

    Endoplasmic reticulum stress is defined as the accumulation of unfolded proteins in the endoplasmic reticulum, and is caused by conditions such as heat or agents that cause endoplasmic reticulum stress, including tunicamycin and dithiothreitol. Autophagy, a major pathway for degradation of macromolecules in the vacuole, is activated by these stress agents in a manner dependent on inositol-requiring enzyme 1b (IRE1b), and delivers endoplasmic reticulum fragments to the vacuole for degradation. In this study, we examined the mechanism for activation of autophagy during endoplasmic reticulum stress in Arabidopsis thaliana. The chemical chaperones sodium 4-phenylbutyrate and tauroursodeoxycholic acid were found to reduce tunicamycin- or dithiothreitol-induced autophagy, but not autophagy caused by unrelated stresses. Similarly, over-expression of BINDING IMMUNOGLOBULIN PROTEIN (BIP), encoding a heat shock protein 70 (HSP70) molecular chaperone, reduced autophagy. Autophagy activated by heat stress was also found to be partially dependent on IRE1b and to be inhibited by sodium 4-phenylbutyrate, suggesting that heat-induced autophagy is due to accumulation of unfolded proteins in the endoplasmic reticulum. Expression in Arabidopsis of the misfolded protein mimics zeolin or a mutated form of carboxypeptidase Y (CPY*) also induced autophagy in an IRE1b-dependent manner. Moreover, zeolin and CPY* partially co-localized with the autophagic body marker GFP-ATG8e, indicating delivery to the vacuole by autophagy. We conclude that accumulation of unfolded proteins in the endoplasmic reticulum is a trigger for autophagy under conditions that cause endoplasmic reticulum stress. PMID:26616142

  14. The anticancer agent di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT) overcomes prosurvival autophagy by two mechanisms: persistent induction of autophagosome synthesis and impairment of lysosomal integrity.

    Science.gov (United States)

    Gutierrez, Elaine; Richardson, Des R; Jansson, Patric J

    2014-11-28

    Autophagy functions as a survival mechanism during cellular stress and contributes to resistance against anticancer agents. The selective antitumor and antimetastatic chelator di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT) causes lysosomal membrane permeabilization and cell death. Considering the integral role of lysosomes in autophagy and cell death, it was important to assess the effect of Dp44mT on autophagy to further understand its mechanism of action. Notably, Dp44mT affected autophagy by two mechanisms. First, concurrent with its antiproliferative activity, Dp44mT increased the expression of the classical autophagic marker LC3-II as a result of induced autophagosome synthesis. Second, this effect was supplemented by a reduction in autophagosome degradation as shown by the accumulation of the autophagic substrate and receptor p62. Conversely, the classical iron chelator desferrioxamine induced autophagosome accumulation only by inhibiting autophagosome degradation. The formation of redox-active iron or copper Dp44mT complexes was critical for its dual effect on autophagy. The cytoprotective antioxidant N-acetylcysteine inhibited Dp44mT-induced autophagosome synthesis and p62 accumulation. Importantly, Dp44mT inhibited autophagosome degradation via lysosomal disruption. This effect prevented the fusion of lysosomes with autophagosomes to form autolysosomes, which is crucial for the completion of the autophagic process. The antiproliferative activity of Dp44mT was suppressed by Beclin1 and ATG5 silencing, indicating the role of persistent autophagosome synthesis in Dp44mT-induced cell death. These studies demonstrate that Dp44mT can overcome the prosurvival activity of autophagy in cancer cells by utilizing this process to potentiate cell death. PMID:25301941

  15. SIRT6 suppresses isoproterenol-induced cardiac hypertrophy through activation of autophagy.

    Science.gov (United States)

    Lu, Jing; Sun, Duanping; Liu, Zhiping; Li, Min; Hong, Huiqi; Liu, Cui; Gao, Si; Li, Hong; Cai, Yi; Chen, Shaorui; Li, Zhuoming; Ye, Jiantao; Liu, Peiqing

    2016-06-01

    Reduction in autophagy has been reported to contribute to the pathogenesis of cardiac hypertrophy. However, the molecular pathways leading to impaired autophagy at the presence of hypertrophic stimuli remain to be elucidated. The present study aimed to investigate the role of sirtuin 6 (SIRT6), a sirtuin family member, in regulating cardiomyocyte autophagy, and its implication in prevention of cardiac hypertrophy. Primary neonatal rat cardiomyocytes (NRCMs) or Sprague-Dawley (SD) rats were submitted to isoproterenol (ISO) treatment, and then the hypertrophic responses and changes in autophagy activity were measured. The influence of SIRT6 on autophagy was observed in cultured NRCMs with gain- and loss-of-function approaches to regulate SIRT6 expression, and further confirmed in vivo by intramyocardial delivery of an adenovirus vector encoding SIRT6 cDNA. In addition, the involvement of SIRT6-mediated autophagy in attenuation of cardiomyocyte hypertrophy induced by ISO was determined basing on genetic or pharmaceutical disruption of autophagy, and the underlying mechanism was preliminarily explored. ISO-caused cardiac hypertrophy accompanying with a significant decrease in autophagy activity. SIRT6 overexpression enhanced autophagy in NRCMs and in rat hearts, whereas knockdown of SIRT6 by RNA interference led to suppression of cardiomyocyte autophagy. Furthermore, the protective effect of SIRT6 against ISO-stimulated hypertrophy was associated with induction of autophagy. SIRT6 promoted nuclear retention of forkhead box O3 transcription factor possibly via attenuating Akt signaling, which was responsible for autophagy activation. Our findings revealed that SIRT6 positively regulates autophagy in cardiomyocytes, which may help to ameliorate ISO-induced cardiac hypertrophy. PMID:27016702

  16. The role of autophagy in microbial infection and immunity

    Directory of Open Access Journals (Sweden)

    Desai M

    2015-01-01

    Full Text Available Mayura Desai,1 Rong Fang,2 Jiaren Sun11Department of Microbiology and Immunology, 2Department of Pathology, University of Texas Medical Branch at Galveston, Galveston, TX, USAAbstract: The autophagy pathway represents an evolutionarily conserved cell recycling process that is activated in response to nutrient deprivation and other stress signals. Over the years, it has been linked to an array of cellular functions. Equally, a wide range of cell-intrinsic, as well as extracellular, factors have been implicated in the induction of the autophagy pathway. Microbial infections represent one such factor that can not only activate autophagy through specific mechanisms but also manipulate the response to the invading microbe's advantage. Moreover, in many cases, particularly among viruses, the pathway has been shown to be intricately involved in the replication cycle of the pathogen. Conversely, autophagy also plays a role in combating the infection process, both through direct destruction of the pathogen and as one of the key mediating factors in the host defense mechanisms of innate and adaptive immunity. Further, the pathway also plays a role in controlling the pathogenesis of infectious diseases by regulating inflammation. In this review, we discuss various interactions between pathogens and the cellular autophagic response and summarize the immunological functions of the autophagy pathway.Keywords: autophagy, xenophagy, antiviral, antibacterial

  17. Characterization of a novel autophagy-specific gene, ATG29

    International Nuclear Information System (INIS)

    Autophagy is a process whereby cytoplasmic proteins and organelles are sequestered for bulk degradation in the vacuole/lysosome. At present, 16 ATG genes have been found that are essential for autophagosome formation in the yeast Saccharomyces cerevisiae. Most of these genes are also involved in the cytoplasm to vacuole transport pathway, which shares machinery with autophagy. Most Atg proteins are colocalized at the pre-autophagosomal structure (PAS), from which the autophagosome is thought to originate, but the precise mechanism of autophagy remains poorly understood. During a genetic screen aimed to obtain novel gene(s) required for autophagy, we identified a novel ORF, ATG29/YPL166w. atg29Δ cells were sensitive to starvation and induction of autophagy was severely retarded. However, the Cvt pathway operated normally. Therefore, ATG29 is an ATG gene specifically required for autophagy. Additionally, an Atg29-GFP fusion protein was observed to localize to the PAS. From these results, we propose that Atg29 functions in autophagosome formation at the PAS in collaboration with other Atg proteins

  18. Retinoid receptor signaling and autophagy in acute promyelocytic leukemia

    Energy Technology Data Exchange (ETDEWEB)

    Orfali, Nina [Cork Cancer Research Center, University College Cork, Cork (Ireland); Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA. (United States); McKenna, Sharon L. [Cork Cancer Research Center, University College Cork, Cork (Ireland); Cahill, Mary R. [Department of Hematology, Cork University Hospital, Cork (Ireland); Gudas, Lorraine J., E-mail: ljgudas@med.cornell.edu [Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA. (United States); Mongan, Nigel P., E-mail: nigel.mongan@nottingham.ac.uk [Faculty of Medicine and Health Science, School of Veterinary Medicine and Science, University of Nottingham, LE12 5RD (United Kingdom); Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA. (United States)

    2014-05-15

    Retinoids are a family of signaling molecules derived from vitamin A with well established roles in cellular differentiation. Physiologically active retinoids mediate transcriptional effects on cells through interactions with retinoic acid (RARs) and retinoid-X (RXR) receptors. Chromosomal translocations involving the RARα gene, which lead to impaired retinoid signaling, are implicated in acute promyelocytic leukemia (APL). All-trans-retinoic acid (ATRA), alone and in combination with arsenic trioxide (ATO), restores differentiation in APL cells and promotes degradation of the abnormal oncogenic fusion protein through several proteolytic mechanisms. RARα fusion-protein elimination is emerging as critical to obtaining sustained remission and long-term cure in APL. Autophagy is a degradative cellular pathway involved in protein turnover. Both ATRA and ATO also induce autophagy in APL cells. Enhancing autophagy may therefore be of therapeutic benefit in resistant APL and could broaden the application of differentiation therapy to other cancers. Here we discuss retinoid signaling in hematopoiesis, leukemogenesis, and APL treatment. We highlight autophagy as a potential important regulator in anti-leukemic strategies. - Highlights: • Normal and aberrant retinoid signaling in hematopoiesis and leukemia is reviewed. • We suggest a novel role for RARα in the development of X-RARα gene fusions in APL. • ATRA therapy in APL activates transcription and promotes onco-protein degradation. • Autophagy may be involved in both onco-protein degradation and differentiation. • Pharmacologic autophagy induction may potentiate ATRA's therapeutic effects.

  19. Autophagy and lysosomal dysfunction as emerging mechanisms of nanomaterial toxicity

    Directory of Open Access Journals (Sweden)

    Stern Stephan T

    2012-06-01

    Full Text Available Abstract The study of the potential risks associated with the manufacture, use, and disposal of nanoscale materials, and their mechanisms of toxicity, is important for the continued advancement of nanotechnology. Currently, the most widely accepted paradigms of nanomaterial toxicity are oxidative stress and inflammation, but the underlying mechanisms are poorly defined. This review will highlight the significance of autophagy and lysosomal dysfunction as emerging mechanisms of nanomaterial toxicity. Most endocytic routes of nanomaterial cell uptake converge upon the lysosome, making the lysosomal compartment the most common intracellular site of nanoparticle sequestration and degradation. In addition to the endo-lysosomal pathway, recent evidence suggests that some nanomaterials can also induce autophagy. Among the many physiological functions, the lysosome, by way of the autophagy (macroautophagy pathway, degrades intracellular pathogens, and damaged organelles and proteins. Thus, autophagy induction by nanoparticles may be an attempt to degrade what is perceived by the cell as foreign or aberrant. While the autophagy and endo-lysosomal pathways have the potential to influence the disposition of nanomaterials, there is also a growing body of literature suggesting that biopersistent nanomaterials can, in turn, negatively impact these pathways. Indeed, there is ample evidence that biopersistent nanomaterials can cause autophagy and lysosomal dysfunctions resulting in toxicological consequences.

  20. Retinoid receptor signaling and autophagy in acute promyelocytic leukemia

    International Nuclear Information System (INIS)

    Retinoids are a family of signaling molecules derived from vitamin A with well established roles in cellular differentiation. Physiologically active retinoids mediate transcriptional effects on cells through interactions with retinoic acid (RARs) and retinoid-X (RXR) receptors. Chromosomal translocations involving the RARα gene, which lead to impaired retinoid signaling, are implicated in acute promyelocytic leukemia (APL). All-trans-retinoic acid (ATRA), alone and in combination with arsenic trioxide (ATO), restores differentiation in APL cells and promotes degradation of the abnormal oncogenic fusion protein through several proteolytic mechanisms. RARα fusion-protein elimination is emerging as critical to obtaining sustained remission and long-term cure in APL. Autophagy is a degradative cellular pathway involved in protein turnover. Both ATRA and ATO also induce autophagy in APL cells. Enhancing autophagy may therefore be of therapeutic benefit in resistant APL and could broaden the application of differentiation therapy to other cancers. Here we discuss retinoid signaling in hematopoiesis, leukemogenesis, and APL treatment. We highlight autophagy as a potential important regulator in anti-leukemic strategies. - Highlights: • Normal and aberrant retinoid signaling in hematopoiesis and leukemia is reviewed. • We suggest a novel role for RARα in the development of X-RARα gene fusions in APL. • ATRA therapy in APL activates transcription and promotes onco-protein degradation. • Autophagy may be involved in both onco-protein degradation and differentiation. • Pharmacologic autophagy induction may potentiate ATRA's therapeutic effects

  1. Piperlongumine induces autophagy by targeting p38 signaling.

    Science.gov (United States)

    Wang, Y; Wang, J-W; Xiao, X; Shan, Y; Xue, B; Jiang, G; He, Q; Chen, J; Xu, H-G; Zhao, R-X; Werle, K D; Cui, R; Liang, J; Li, Y-L; Xu, Z-X

    2013-01-01

    Piperlongumine (PL), a natural product isolated from the plant species Piper longum L., can selectively induce apoptotic cell death in cancer cells by targeting the stress response to reactive oxygen species (ROS). Here we show that PL induces cell death in the presence of benzyloxycarbonylvalyl-alanyl-aspartic acid (O-methyl)-fluoro-methylketone (zVAD-fmk), a pan-apoptotic inhibitor, and in the presence of necrostatin-1, a necrotic inhibitor. Instead PL-induced cell death can be suppressed by 3-methyladenine, an autophagy inhibitor, and substantially attenuated in cells lacking the autophagy-related 5 (Atg5) gene. We further show that PL enhances autophagy activity without blocking autophagy flux. Application of N-acetyl-cysteine, an antioxidant, markedly reduces PL-induced autophagy and cell death, suggesting an essential role for intracellular ROS in PL-induced autophagy. Furthermore, PL stimulates the activation of p38 protein kinase through ROS-induced stress response and p38 signaling is necessary for the action of PL as SB203580, a p38 inhibitor, or dominant-negative p38 can effectively reduce PL-mediated autophagy. Thus, we have characterized a new mechanism for PL-induced cell death through the ROS-p38 pathway. Our findings support the therapeutic potential of PL by triggering autophagic cell death. PMID:24091667

  2. Phosphatase and tensin homologue (PTEN)-induced putative kinase 1 reduces pancreatic β-cells apoptosis in glucotoxicity through activation of autophagy.

    Science.gov (United States)

    Zhang, Juan; Chen, Ke; Wang, Linghao; Wan, Xinxin; Shrestha, Chandrama; Zhou, Jingsong; Mo, Zhaohui

    2016-08-01

    Chronic elevated glucose is harmful to pancreatic β-cells, resulting in pancreatic β-cells dysfunction and apoptosis. Understanding the molecular mechanisms associated with β-cells survival is pivotal for the prevention of β-cells injury caused by glucotoxicity. The role of Phosphatase and tensin homologue (PTEN)-induced putative kinase 1 (PINK1) in the fate of pancreatic β-cells constantly exposed to high glucose was studied. Sustained high glucose increased PINK1 protein expression both in rat pancreatic β-cells and INS-1 β-cells, and that this increase can be inhibited by PINK1 knockdown and further enhanced by PINK1 over-expression. PINK1 deficiency aggravated glucotoxicity-induced pancreatic β-cells apoptosis and inhibition of autophagy whereas PINK1 could reverse these adverse effects. This study provides fundamental data supporting the potential protective role of PINK1 as a new therapeutic target necessary to preserve β-cells survival under non-physiological hyperglycemia conditions. PMID:27233610

  3. Energy-efficient induction motors designing with application of a modified criterion of reduced costs

    OpenAIRE

    V.S. Petrushin

    2014-01-01

    The paper introduces a modified criterion of reduced costs that employs coefficients of operation significance and priority of ohmic loss accounting to allow matching maximum efficiency with minimum reduced costs. Impact of the inflation factor on the criterion of reduced costs is analyzed.

  4. Energy-efficient induction motors designing with application of a modified criterion of reduced costs

    Directory of Open Access Journals (Sweden)

    V.S. Petrushin

    2014-03-01

    Full Text Available The paper introduces a modified criterion of reduced costs that employs coefficients of operation significance and priority of ohmic loss accounting to allow matching maximum efficiency with minimum reduced costs. Impact of the inflation factor on the criterion of reduced costs is analyzed.

  5. Are mitochondrial reactive oxygen species required for autophagy?

    Energy Technology Data Exchange (ETDEWEB)

    Jiang, Jianfei, E-mail: jjf73@pitt.edu [Center for Free Radical and Antioxidant Health, Department of Environmental and Occupational Health, University of Pittsburgh (United States); Maeda, Akihiro; Ji, Jing [Center for Free Radical and Antioxidant Health, Department of Environmental and Occupational Health, University of Pittsburgh (United States); Baty, Catherine J.; Watkins, Simon C. [Center for Biologic Imaging, Department of Cell Biology and Physiology, University of Pittsburgh (United States); Greenberger, Joel S. [Department of Radiation Oncology, University of Pittsburgh (United States); Kagan, Valerian E., E-mail: kagan@pitt.edu [Center for Free Radical and Antioxidant Health, Department of Environmental and Occupational Health, University of Pittsburgh (United States)

    2011-08-19

    Highlights: {yields} Autophageal and apoptotic pathways were dissected in cytochrome c deficient cells. {yields} Staurosporine (STS)-induced autophagy was not accompanied by ROS generation. {yields} Autophagy was detectable in mitochondrial DNA deficient {rho}{sup 0} cells. {yields} Mitochondrial ROS are not required for the STS-induced autophagy in HeLa cells. -- Abstract: Reactive oxygen species (ROS) are said to participate in the autophagy signaling. Supporting evidence is obscured by interference of autophagy and apoptosis, whereby the latter heavily relies on ROS signaling. To dissect autophagy from apoptosis we knocked down expression of cytochrome c, the key component of mitochondria-dependent apoptosis, in HeLa cells using shRNA. In cytochrome c deficient HeLa1.2 cells, electron transport was compromised due to the lack of electron shuttle between mitochondrial respiratory complexes III and IV. A rapid and robust LC3-I/II conversion and mitochondria degradation were observed in HeLa1.2 cells treated with staurosporine (STS). Neither generation of superoxide nor accumulation of H{sub 2}O{sub 2} was detected in STS-treated HeLa1.2 cells. A membrane permeable antioxidant, PEG-SOD, plus catalase exerted no effect on STS-induced LC3-I/II conversion and mitochondria degradation. Further, STS caused autophagy in mitochondria DNA-deficient {rho}{sup o} HeLa1.2 cells in which both electron transport and ROS generation were completely disrupted. Counter to the widespread view, we conclude that mitochondrial ROS are not required for the induction of autophagy.

  6. Are mitochondrial reactive oxygen species required for autophagy?

    International Nuclear Information System (INIS)

    Highlights: → Autophageal and apoptotic pathways were dissected in cytochrome c deficient cells. → Staurosporine (STS)-induced autophagy was not accompanied by ROS generation. → Autophagy was detectable in mitochondrial DNA deficient ρ0 cells. → Mitochondrial ROS are not required for the STS-induced autophagy in HeLa cells. -- Abstract: Reactive oxygen species (ROS) are said to participate in the autophagy signaling. Supporting evidence is obscured by interference of autophagy and apoptosis, whereby the latter heavily relies on ROS signaling. To dissect autophagy from apoptosis we knocked down expression of cytochrome c, the key component of mitochondria-dependent apoptosis, in HeLa cells using shRNA. In cytochrome c deficient HeLa1.2 cells, electron transport was compromised due to the lack of electron shuttle between mitochondrial respiratory complexes III and IV. A rapid and robust LC3-I/II conversion and mitochondria degradation were observed in HeLa1.2 cells treated with staurosporine (STS). Neither generation of superoxide nor accumulation of H2O2 was detected in STS-treated HeLa1.2 cells. A membrane permeable antioxidant, PEG-SOD, plus catalase exerted no effect on STS-induced LC3-I/II conversion and mitochondria degradation. Further, STS caused autophagy in mitochondria DNA-deficient ρo HeLa1.2 cells in which both electron transport and ROS generation were completely disrupted. Counter to the widespread view, we conclude that mitochondrial ROS are not required for the induction of autophagy.

  7. Receptor Proteins in Selective Autophagy

    Directory of Open Access Journals (Sweden)

    Christian Behrends

    2012-01-01

    Full Text Available Autophagy has long been thought to be an essential but unselective bulk degradation pathway. However, increasing evidence suggests selective autophagosomal turnover of a broad range of substrates. Bifunctional autophagy receptors play a key role in selective autophagy by tethering cargo to the site of autophagosomal engulfment. While the identity of molecular components involved in selective autophagy has been revealed at least to some extent, we are only beginning to understand how selectivity is achieved in this process. Here, we summarize the mechanistic and structural basis of receptor-mediated selective autophagy.

  8. Targeting autophagy in cancer management – strategies and developments

    Directory of Open Access Journals (Sweden)

    Ozpolat B

    2015-09-01

    Full Text Available Bulent Ozpolat,1 Doris M Benbrook2 1Department of Experimental Therapeutics, The University of Texas – Houston, MD Anderson Cancer Center, Houston, TX, 2Department of Obstetrics and Gynecology, University of Oklahoma HSC, Oklahoma City, OK, USA Abstract: Autophagy is a highly regulated catabolic process involving lysosomal degradation of intracellular components, damaged organelles, misfolded proteins, and toxic aggregates, reducing oxidative stress and protecting cells from damage. The process is also induced in response to various conditions, including nutrient deprivation, metabolic stress, hypoxia, anticancer therapeutics, and radiation therapy to adapt cellular conditions for survival. Autophagy can function as a tumor suppressor mechanism in normal cells and dysregulation of this process (ie, monoallelic Beclin-1 deletion may lead to malignant transformation and carcinogenesis. In tumors, autophagy is thought to promote tumor growth and progression by helping cells to adapt and survive in metabolically-challenged and harsh tumor microenvironments (ie, hypoxia and acidity. Recent in vitro and in vivo studies in preclinical models suggested that modulation of autophagy can be used as a therapeutic modality to enhance the efficacy of conventional therapies, including chemo and radiation therapy. Currently, more than 30 clinical trials are investigating the effects of autophagy inhibition in combination with cytotoxic chemotherapies and targeted agents in various cancers. In this review, we will discuss the role, molecular mechanism, and regulation of autophagy, while targeting this process as a novel therapeutic modality, in various cancers. Keywords: autophagy inhibition, chemotherapy, tumor microenvironment

  9. PUMA and Bax-induced Autophagy Contributes to Apoptosis

    OpenAIRE

    Yee, Karen S.; Wilkinson, Simon; James, John; Ryan, Kevin M.; Vousden, Karen H.

    2009-01-01

    The p53-inducible BH3-only protein PUMA is a key mediator of p53-dependent apoptosis, and PUMA has been shown to function by activating Bax and mitochondrial outer membrane permeabilization. In this study we describe an ability of PUMA to induce autophagy that leads to the selective removal of mitochondria. This function of PUMA depends on Bax/Bak and can be reproduced by overexpression of Bax. The induction of autophagy coincides with cytochrome c release, and taken together the results sugg...

  10. PUMA- and Bax-induced autophagy contributes to apoptosis

    OpenAIRE

    Yee, K S; Wilkinson, S; James, J; Ryan, K M; Vousden, K H

    2009-01-01

    The p53-inducible BH3-only protein PUMA is a key mediator of p53-dependent apoptosis, and PUMA has been shown to function by activating Bax and mitochondrial outer membrane permeabilization. In this study, we describe an ability of PUMA to induce autophagy that leads to the selective removal of mitochondria. This function of PUMA depends on Bax/Bak and can be reproduced by overexpression of Bax. The induction of autophagy coincides with cytochrome c release, and taken together the results sug...

  11. WECS Based Self-Excited Squirrel-Cage Induction Generator with Reduced Voltage Source Inverter Rating

    Directory of Open Access Journals (Sweden)

    O.Chandra sekha

    2014-07-01

    Full Text Available This paper presents the regulation of the voltage and frequency of a stand-alone fixed-pitch wind energy conversion system (WECS based on a self-excited squirrel-cage induction machine. A shunt connected voltage source inverter (VSI and a controllable dump load are used for regulation purposes. A battery bank is included in the dc side of the VSI so that it can absorb and inject active power thus increasing the efficiency and availability of the system. A control scheme for the VSI with self-governing control of real and reactive power allows the state of charge of the batteries to be kept in a safe range while maximizing the voltage regulating capabilities of the VSI. The characteristics of the wind turbine, selfexcited generator, and the ratings of the VSI are considered in order to find out the load range for which voltage and frequency can be regulated for a given wind speed range. The possibility of the proposed system is verified by MATLAB/SIMULINK. simulations.

  12. Autophagy extends lifespan via vacuolar acidification

    Directory of Open Access Journals (Sweden)

    Christoph Ruckenstuhl

    2014-05-01

    Full Text Available Methionine restriction (MetR is one of the rare regimes that prolongs lifespan across species barriers. Using a yeast model, we recently demonstrated that this lifespan extension is promoted by autophagy, which in turn requires vacuolar acidification. Our study is the first to place autophagy as one of the major players required for MetR-mediated longevity. In addition, our work identifies vacuolar acidification as a key downstream element of autophagy induction under MetR, and possibly after rapamycin treatment. Unlike other amino acids, methionine plays pleiotropic roles in many metabolism-relevant pathways. For instance, methionine (i is the N-terminal amino acid of every newly translated protein; (ii acts as the central donor of methyl groups through S-adenosyl methionine (SAM during methylation reactions of proteins, DNA or RNA; and (iii provides the sulfhydryl groups for FeS-cluster formation and redox detoxification via transsulfuration to cysteine. Intriguingly, MetR causes lifespan extension, both in yeast and in rodents. We could show that in Saccharomyces cerevisiae, chronological lifespan (CLS is increased in two specific methionine-auxotrophic strains (namely Δmet2 and Δmet15.

  13. Autophagy: a new target for nonalcoholic fatty liver disease therapy

    Directory of Open Access Journals (Sweden)

    Mao YQ

    2016-03-01

    Full Text Available Yuqing Mao,1 Fujun Yu,1 Jianbo Wang,2 Chuanyong Guo,3 Xiaoming Fan1 1Department of Gastroenterology and Hepatology, Jinshan Hospital of Fudan University, Shanghai, 2Department of Gastroenterology and Hepatology, The Central Hospital of Lishui City, Wenzhou Medical University, Zhejiang, 3Department of Gastroenterology and Hepatology, Shanghai Tenth People's Hospital, Tongji University, Shanghai, People's Republic of China Abstract: Nonalcoholic fatty liver disease (NAFLD has gained importance in recent decades due to drastic changes in diet, especially in Western countries. NAFLD occurs as a spectrum from simple hepatic steatosis, steatohepatitis to cirrhosis, and even hepatocellular carcinoma. Although the molecular mechanisms underlying the development of NAFLD have been intensively investigated, many issues remain to be resolved. Autophagy is a cell survival mechanism for disposing of excess or defective organelles, and has become a hot spot for research. Recent studies have revealed that autophagy is linked to the development of NAFLD and regulation of autophagy has therapeutic potential. Autophagy reduces intracellular lipid droplets by enclosing them and fusing with lysosomes for degradation. Furthermore, autophagy is involved in attenuating inflammation and liver injury. However, autophagy is regarded as a double-edged sword, as it may also affect adipogenesis and adipocyte differentiation. Moreover, it is unclear as to whether autophagy protects the body from injury or causes diseases and even death, and the association between autophagy and NAFLD remains controversial. This review is intended to discuss, comment, and outline the progress made in this field and establish the possible molecular mechanism involved. Keywords: nonalcoholic fatty liver disease, autophagy, steatosis, steatohepatitis, fibrosis, carcinogenesis

  14. Heavy ion irradiation induces autophagy in irradiated C2C12 myoblasts and their bystander cells

    International Nuclear Information System (INIS)

    Autophagy is one of the major processes involved in the degradation of intracellular materials. Here, we examined the potential impact of heavy ion irradiation on the induction of autophagy in irradiated C2C12 mouse myoblasts and their non-targeted bystander cells. In irradiated cells, ultrastructural analysis revealed the accumulation of autophagic structures at various stages of autophagy (id est (i.e.) phagophores, autophagosomes and autolysosomes) within 20 min after irradiation. Multivesicular bodies (MVBs) and autolysosomes containing MVBs (amphisomes) were also observed. Heavy ion irradiation increased the staining of microtubule-associated protein 1 light chain 3 and LysoTracker Red (LTR). Such enhanced staining was suppressed by an autophagy inhibitor 3-methyladenine. In addition to irradiated cells, bystander cells were also positive with LTR staining. Altogether, these results suggest that heavy ion irradiation induces autophagy not only in irradiated myoblasts but also in their bystander cells. (author)

  15. Longevity-relevant regulation of autophagy at the level of the acetylproteome

    DEFF Research Database (Denmark)

    Mariño, Guillermo; Morselli, Eugenia; Bennetzen, Martin V; Eisenberg, Tobias; Megalou, Evgenia; Schroeder, Sabrina; Cabrera, Sandra; Bénit, Paule; Rustin, Pierre; Criollo, Alfredo; Kepp, Oliver; Galluzzi, Lorenzo; Shen, Shensi; Malik, Shoaib A; Maiuri, Maria Chiara; Horio, Yoshiyuki; López-Otín, Carlos; Andersen, Jens S.; Tavernarakis, Nektarios; Madeo, Frank; Kroemer, Guido

    2011-01-01

    The acetylase inhibitor, spermidine and the deacetylase activator, resveratrol, both induce autophagy and prolong life span of the model organism Caenorhabditis elegans in an autophagydependent fashion. Based on these premises, we investigated the differences and similarities in spermidine and...... resveratrol-induced autophagy. The deacetylase sirtuin 1 (SIRT1) and its orthologs are required for the autophagy induction by resveratrol but dispensable for autophagy stimulation by spermidine in human cells, Saccharomyces cerevisiae and C. elegans. SIRT1 is also dispensable for life-span extension by...... spermidine. Mass spectrometry analysis of the human acetylproteome revealed that resveratrol and/or spermidine induce changes in the acetylation of 560 peptides corresponding to 375 different proteins. Among these, 170 proteins are part of the recently elucidated human autophagy protein network. Importantly...

  16. Application of a particle separation device to reduce inductively coupled plasma-enhanced elemental fractionation in laser ablation-inductively coupled plasma-mass spectrometry

    International Nuclear Information System (INIS)

    The particle size distribution of laser ablation aerosols are a function of the wavelength, the energy density and the pulse duration of the laser, as well as the sample matrix and the gas environment. Further the size of the particles affects the vaporization and ionization efficiency in the inductively coupled plasma (ICP). Some matrices produce large particles, which are not completely vaporized and ionized in the ICP. The previous work has shown that analytical results such as matrix-independent calibration, accuracy and precision can be significantly influenced by the particle sizes of the particles. To minimize the particle size related incomplete conversion of the sample to ions in the ICP a particle separation device was developed, which allows effective particle separation using centrifugal forces in a thin coiled tube. In this device, the particle cut-off size is varied by changing the number of turns in the coil, as well as by changing the gas flow and the tube diameter. The interaction of the laser with the different samples leads to varying particle size distributions. When carrying out quantitative analysis with non-matrix matched calibration reference materials, it was shown that different particle cut-off sizes were required depending on the ICP conditions and the instrument used for analysis. Various sample materials were investigated in this study to demonstrate the applicability of the device. For silicate matrices, the capability of the ICP to produce ions was significantly reduced for particles larger than 0.5 μm, and was dependent on the element monitored. To reduce memory effects caused by the separated particles, a washout procedure was developed, which additionally allowed the analysis of the trapped particles. These results clearly demonstrate the very important particle size dependent ICP-MS signal response and the potential of the described particle size based separator for the reduction of ICP induced elemental fractionation

  17. Enhanced autophagy as a potential mechanism for the improved physiological function by simvastatin in muscular dystrophy

    OpenAIRE

    Whitehead, Nicholas P.

    2016-01-01

    ABSTRACT Autophagy has recently emerged as an important cellular process for the maintenance of skeletal muscle health and function. Excessive autophagy can trigger muscle catabolism, leading to atrophy. In contrast, reduced autophagic flux is a characteristic of several muscle diseases, including Duchenne muscular dystrophy, the most common and severe inherited muscle disorder. Recent evidence demonstrates that enhanced reactive oxygen species (ROS) production by CYBB/NOX2 impairs autophagy ...

  18. DNA damage and autophagy

    International Nuclear Information System (INIS)

    Both exogenous and endogenous agents are a threat to DNA integrity. Exogenous environmental agents such as ultraviolet (UV) and ionizing radiation, genotoxic chemicals and endogenous byproducts of metabolism including reactive oxygen species can cause alterations in DNA structure (DNA damage). Unrepaired DNA damage has been linked to a variety of human disorders including cancer and neurodegenerative disease. Thus, efficient mechanisms to detect DNA lesions, signal their presence and promote their repair have been evolved in cells. If DNA is effectively repaired, DNA damage response is inactivated and normal cell functioning resumes. In contrast, when DNA lesions cannot be removed, chronic DNA damage triggers specific cell responses such as cell death and senescence. Recently, DNA damage has been shown to induce autophagy, a cellular catabolic process that maintains a balance between synthesis, degradation, and recycling of cellular components. But the exact mechanisms by which DNA damage triggers autophagy are unclear. More importantly, the role of autophagy in the DNA damage response and cellular fate is unknown. In this review we analyze evidence that supports a role for autophagy as an integral part of the DNA damage response.

  19. Dopamine Oxidation and Autophagy

    Directory of Open Access Journals (Sweden)

    Patricia Muñoz

    2012-01-01

    Full Text Available The molecular mechanisms involved in the neurodegenerative process of Parkinson's disease remain unclear. Currently, there is a general agreement that mitochondrial dysfunction, α-synuclein aggregation, oxidative stress, neuroinflammation, and impaired protein degradation are involved in the neurodegeneration of dopaminergic neurons containing neuromelanin in Parkinson's disease. Aminochrome has been proposed to play an essential role in the degeneration of dopaminergic neurons containing neuromelanin by inducing mitochondrial dysfunction, oxidative stress, the formation of neurotoxic α-synuclein protofibrils, and impaired protein degradation. Here, we discuss the relationship between the oxidation of dopamine to aminochrome, the precursor of neuromelanin, autophagy dysfunction in dopaminergic neurons containing neuromelanin, and the role of dopamine oxidation to aminochrome in autophagy dysfunction in dopaminergic neurons. Aminochrome induces the following: (i the formation of α-synuclein protofibrils that inactivate chaperone-mediated autophagy; (ii the formation of adducts with α- and β-tubulin, which induce the aggregation of the microtubules required for the fusion of autophagy vacuoles and lysosomes.

  20. Ribosomal trafficking is reduced in Schwann cells following induction of myelination

    Directory of Open Access Journals (Sweden)

    James M. Love

    2015-08-01

    Full Text Available Local synthesis of proteins within the Schwann cell periphery is extremely important for efficient process extension and myelination, when cells undergo dramatic changes in polarity and geometry. Still, it is unclear how ribosomal distributions are developed and maintained within Schwann cell projections to sustain local translation. In this multi-disciplinary study, we expressed a plasmid encoding a fluorescently labeled ribosomal subunit (L4-GFP in cultured primary rat Schwann cells. This enabled the generation of high-resolution, quantitative data on ribosomal distributions and trafficking dynamics within Schwann cells during early stages of myelination, induced by ascorbic acid treatment. Ribosomes were distributed throughout Schwann cell projections, with ~2-3 bright clusters along each projection. Clusters emerged within 1 day of culture and were maintained throughout early stages of myelination. Three days after induction of myelination, net ribosomal movement remained anterograde (directed away from the Schwann cell body, but ribosomal velocity decreased to about half the levels of the untreated group. Statistical and modeling analysis provided additional insight into key factors underlying ribosomal trafficking. Multiple regression analysis indicated that net transport at early time points was dependent on anterograde velocity, but shifted to dependence on anterograde duration at later time points. A simple, data-driven rate kinetics model suggested that the observed decrease in net ribosomal movement was primarily dictated by an increased conversion of anterograde particles to stationary particles, rather than changes in other directional parameters. These results reveal the strength of a combined experimental and theoretical approach in examining protein localization and transport, and provide evidence of an early establishment of ribosomal populations within Schwann cell projections with a reduction in trafficking following

  1. Autophagy studies in Bombyx mori

    Directory of Open Access Journals (Sweden)

    L Tian

    2015-03-01

    Full Text Available Autophagy, which is well conserved from yeast to mammals, plays essential roles in development and diseases. Using the domesticated silkworm, Bombyx mori, as a model insect, several reports on autophagy have been made recently. Autophagic features are observed in the midgut and fat body during the larval-pupal transition as well as the silk gland and ovarian nurse cells during the pupal stage. There are 14 autophagy related (Atg genes, including at least two transcript variants of Atg1, predicated in Bombyx. Expression of most Atg genes is consistent with the autophagy process in the fat body during the larval-pupal transition, and reduction of Atg1 expression by RNAi blocks this process. The molting hormone, 20-hydroxyecdysone (20E, and starvation induce autophagy in the fat body by upregulating Atg gene expression and blocking the PI3K-TORC1 pathway. Meanwhile, autophagy precedes apoptosis in the midgut and other larval tissues during the larval-pupal transition, while the detailed mechanism is not illustrated yet. We assume that there are at least four future directions about autophagy studies in Bombyx during the next years: (1 physiological functions of autophagy; (2 identification of new components involved in the autophagy process; (3 detailed molecular mechanism of autophagosome formation; (4 functional relationship between autophagy and apoptosis.

  2. FoxO1 antagonist suppresses autophagy and lipid droplet growth in adipocytes.

    Science.gov (United States)

    Liu, Longhua; Zheng, Louise D; Zou, Peng; Brooke, Joseph; Smith, Cayleen; Long, Yun Chau; Almeida, Fabio A; Liu, Dongmin; Cheng, Zhiyong

    2016-08-01

    Obesity and related metabolic disorders constitute one of the most pressing heath concerns worldwide. Increased adiposity is linked to autophagy upregulation in adipose tissues. However, it is unknown how autophagy is upregulated and contributes to aberrant adiposity. Here we show a FoxO1-autophagy-FSP27 axis that regulates adipogenesis and lipid droplet (LD) growth in adipocytes. Adipocyte differentiation was associated with upregulation of autophagy and fat specific protein 27 (FSP27), a key regulator of adipocyte maturation and expansion by promoting LD formation and growth. However, FoxO1 specific inhibitor AS1842856 potently suppressed autophagy, FSP27 expression, and adipocyte differentiation. In terminally differentiated adipocytes, AS1842856 significantly reduced FSP27 level and LD size, which was recapitulated by autophagy inhibitors (bafilomycin-A1 and leupeptin, BL). Similarly, AS1842856 and BL dampened autophagy activity and FSP27 expression in explant cultures of white adipose tissue. To our knowledge, this is the first study addressing FoxO1 in the regulation of adipose autophagy, shedding light on the mechanism of increased autophagy and adiposity in obese individuals. Given that adipogenesis and adipocyte expansion contribute to aberrant adiposity, targeting the FoxO1-autophagy-FSP27 axis may lead to new anti-obesity options. PMID:27260854

  3. Cocaine induces astrocytosis through ER stress-mediated activation of autophagy.

    Science.gov (United States)

    Periyasamy, Palsamy; Guo, Ming-Lei; Buch, Shilpa

    2016-08-01

    Cocaine is known to induce inflammation, thereby contributing in part, to the pathogenesis of neurodegeneration. A recent study from our lab has revealed a link between macroautophagy/autophagy and microglial activation. The current study was aimed at investigating whether cocaine could also mediate activation of astrocytes and, whether this process involved induction of autophagy. Our findings demonstrated that cocaine mediated the activation of astrocytes by altering the levels of autophagy markers, such as BECN1, ATG5, MAP1LC3B-II, and SQSTM1 in both human A172 astrocytoma cells and primary human astrocytes. Furthermore, cocaine treatment resulted in increased formation of endogenous MAP1LC3B puncta in human astrocytes. Additionally, astrocytes transfected with the GFP-MAP1LC3B plasmid also demonstrated cocaine-mediated upregulation of the green fluorescent MAP1LC3B puncta. Cocaine-mediated induction of autophagy involved upstream activation of ER stress proteins such as EIF2AK3, ERN1, ATF6 since blockage of autophagy using either pharmacological or gene-silencing approaches, had no effect on cocaine-mediated induction of ER stress. Using both pharmacological and gene-silencing approaches to block either ER stress or autophagy, our findings demonstrated that cocaine-induced activation of astrocytes (measured by increased levels of GFAP) involved sequential activation of ER stress and autophagy. Cocaine-mediated-increased upregulation of GFAP correlated with increased expression of proinflammatory mediators such as TNF, IL1B, and IL6. In conclusion, these findings reveal an association between ER stress-mediated autophagy and astrogliosis in cocaine-treated astrocytes. Intervention of ER stress and/or autophagy signaling would thus be promising therapeutic targets for abrogating cocaine-mediated neuroinflammation. PMID:27337297

  4. Endogenous antioxidant defense induction by melon superoxide dismutase reduces cardiac hypertrophy in spontaneously hypertensive rats.

    Science.gov (United States)

    Carillon, Julie; Rugale, Caroline; Rouanet, Jean-Max; Cristol, Jean-Paul; Lacan, Dominique; Jover, Bernard

    2014-08-01

    We assessed the influence of SODB, a melon superoxide dismutase (SOD), on left ventricular (LV) hypertrophy in SHR. SODB (4 or 40U SOD) was given orally for 4 or 28 days to SHR. For each treatment period, LV weight index (LVWI) and cardiomyocytes size were measured. SOD, glutathione peroxidase (GPx) and catalase expressions, and LV production and presence of superoxide anion were determined. Pro-inflammatory markers were also measured. SODB reduced LVWI and cardiomyocytes size after 4 or 28 days. Cardiac SOD and GPx increased by 30-40% with SODB. The presence but not production of superoxide anion was significantly reduced by SODB. No effect of SODB was detected on inflammatory status in any group. The beneficial effect of SODB on cardiac hypertrophy seems to be related to the stimulation of endogenous antioxidant defense, suggesting that SODB may be of interest as a dietary supplementation during conventional antihypertensive therapy. PMID:24601674

  5. Autophagy During Cardiac Stress: Joys and Frustrations of Autophagy

    Science.gov (United States)

    Gottlieb, Roberta A.; Mentzer, Robert M.

    2013-01-01

    The study of autophagy has been transformed by the cloning of most genes in the pathway and the introduction of GFP-LC3 as a reporter to allow visual assessment of autophagy. The field of cardiac biology is not alone in attempting to understand the implications of autophagy. The purpose of this review is to address some of the controversies and conundrums associated with the evolving studies of autophagy in the heart. Autophagy is a cellular process involving a complex orchestration of regulatory gene products as well as machinery for assembly, selective targeting, and degradation of autophagosomes and their contents. Our understanding of the role of autophagy in human disease is rapidly evolving as investigators examine the process in different tissues and different pathophysiological contexts. In the field of heart disease, autophagy has been examined in the settings of ischemia and reperfusion, preconditioning, cardiac hypertrophy, and heart failure. This review addresses the role of autophagy in cardioprotection, the balance of catabolism and anabolism, the concept of mitochondrial quality control, and the implications of impaired autophagic flux or frustrated autophagy. PMID:20148666

  6. Idarubicin induces mTOR-dependent cytotoxic autophagy in leukemic cells

    International Nuclear Information System (INIS)

    We investigated if the antileukemic drug idarubicin induces autophagy, a process of programmed cellular self-digestion, in leukemic cell lines and primary leukemic cells. Transmission electron microscopy and acridine orange staining demonstrated the presence of autophagic vesicles and intracellular acidification, respectively, in idarubicin-treated REH leukemic cell line. Idarubicin increased punctuation/aggregation of microtubule-associated light chain 3B (LC3B), enhanced the conversion of LC3B-I to autophagosome-associated LC3B-II in the presence of proteolysis inhibitors, and promoted the degradation of the selective autophagic target p62, thus indicating the increase in autophagic flux. Idarubicin inhibited the phosphorylation of the main autophagy repressor mammalian target of rapamycin (mTOR) and its downstream target p70S6 kinase. The treatment with the mTOR activator leucine prevented idarubicin-mediated autophagy induction. Idarubicin-induced mTOR repression was associated with the activation of the mTOR inhibitor AMP-activated protein kinase and down-regulation of the mTOR activator Akt. The suppression of autophagy by pharmacological inhibitors or LC3B and beclin-1 genetic knockdown rescued REH cells from idarubicin-mediated oxidative stress, mitochondrial depolarization, caspase activation and apoptotic DNA fragmentation. Idarubicin also caused mTOR inhibition and cytotoxic autophagy in K562 leukemic cell line and leukocytes from chronic myeloid leukemia patients, but not healthy controls. By demonstrating mTOR-dependent cytotoxic autophagy in idarubicin-treated leukemic cells, our results warrant caution when considering combining idarubicin with autophagy inhibitors in leukemia therapy. - Highlights: • Idarubicin induces autophagy in leukemic cell lines and primary leukemic cells. • Idarubicin induces autophagy by inhibiting mTOR in leukemic cells. • mTOR suppression by idarubicin is associated with AMPK activation and Akt blockade.

  7. Idarubicin induces mTOR-dependent cytotoxic autophagy in leukemic cells

    Energy Technology Data Exchange (ETDEWEB)

    Ristic, Biljana [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Bosnjak, Mihajlo [Institute of Histology and Embryology, School of Medicine, University of Belgrade, Belgrade (Serbia); Arsikin, Katarina [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Mircic, Aleksandar; Suzin-Zivkovic, Violeta [Institute of Histology and Embryology, School of Medicine, University of Belgrade, Belgrade (Serbia); Bogdanovic, Andrija [Clinic for Hematology, Clinical Centre of Serbia, School of Medicine, University of Belgrade, Belgrade (Serbia); Perovic, Vladimir [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Martinovic, Tamara; Kravic-Stevovic, Tamara; Bumbasirevic, Vladimir [Institute of Histology and Embryology, School of Medicine, University of Belgrade, Belgrade (Serbia); Trajkovic, Vladimir, E-mail: vtrajkovic@med.bg.ac.rs [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Harhaji-Trajkovic, Ljubica, E-mail: buajk@yahoo.com [Institute for Biological Research, University of Belgrade, Belgrade, Despot Stefan Blvd. 142, 11000 Belgrade (Serbia)

    2014-08-01

    We investigated if the antileukemic drug idarubicin induces autophagy, a process of programmed cellular self-digestion, in leukemic cell lines and primary leukemic cells. Transmission electron microscopy and acridine orange staining demonstrated the presence of autophagic vesicles and intracellular acidification, respectively, in idarubicin-treated REH leukemic cell line. Idarubicin increased punctuation/aggregation of microtubule-associated light chain 3B (LC3B), enhanced the conversion of LC3B-I to autophagosome-associated LC3B-II in the presence of proteolysis inhibitors, and promoted the degradation of the selective autophagic target p62, thus indicating the increase in autophagic flux. Idarubicin inhibited the phosphorylation of the main autophagy repressor mammalian target of rapamycin (mTOR) and its downstream target p70S6 kinase. The treatment with the mTOR activator leucine prevented idarubicin-mediated autophagy induction. Idarubicin-induced mTOR repression was associated with the activation of the mTOR inhibitor AMP-activated protein kinase and down-regulation of the mTOR activator Akt. The suppression of autophagy by pharmacological inhibitors or LC3B and beclin-1 genetic knockdown rescued REH cells from idarubicin-mediated oxidative stress, mitochondrial depolarization, caspase activation and apoptotic DNA fragmentation. Idarubicin also caused mTOR inhibition and cytotoxic autophagy in K562 leukemic cell line and leukocytes from chronic myeloid leukemia patients, but not healthy controls. By demonstrating mTOR-dependent cytotoxic autophagy in idarubicin-treated leukemic cells, our results warrant caution when considering combining idarubicin with autophagy inhibitors in leukemia therapy. - Highlights: • Idarubicin induces autophagy in leukemic cell lines and primary leukemic cells. • Idarubicin induces autophagy by inhibiting mTOR in leukemic cells. • mTOR suppression by idarubicin is associated with AMPK activation and Akt blockade.

  8. The thiazole derivative CPTH6 impairs autophagy.

    Science.gov (United States)

    Ragazzoni, Y; Desideri, M; Gabellini, C; De Luca, T; Carradori, S; Secci, D; Nescatelli, R; Candiloro, A; Condello, M; Meschini, S; Del Bufalo, D; Trisciuoglio, D

    2013-01-01

    We have previously demonstrated that the thiazole derivative 3-methylcyclopentylidene-[4-(4'-chlorophenyl)thiazol-2-yl]hydrazone (CPTH6) induces apoptosis and cell cycle arrest in human leukemia cells. The aim of this study was to evaluate whether CPTH6 is able to affect autophagy. By using several human tumor cell lines with different origins we demonstrated that CPTH6 treatment induced, in a dose-dependent manner, a significant increase in autophagic features, as imaged by electron microscopy, immunoblotting analysis of membrane-bound form of microtubule-associated protein 1 light chain 3 (LC3B-II) levels and by appearance of typical LC3B-II-associated autophagosomal puncta. To gain insights into the molecular mechanisms of elevated markers of autophagy induced by CPTH6 treatment, we silenced the expression of several proteins acting at different steps of autophagy. We found that the effect of CPTH6 on autophagy developed through a noncanonical mechanism that did not require beclin-1-dependent nucleation, but involved Atg-7-mediated elongation of autophagosomal membranes. Strikingly, a combined treatment of CPTH6 with late-stage autophagy inhibitors, such as chloroquine and bafilomycin A1, demonstrates that under basal condition CPTH6 reduces autophagosome turnover through an impairment of their degradation pathway, rather than enhancing autophagosome formation, as confirmed by immunofluorescence experiments. According to these results, CPTH6-induced enhancement of autophagy substrate p62 and NBR1 protein levels confirms a blockage of autophagic cargo degradation. In addition, CPTH6 inhibited autophagosome maturation and compounds having high structural similarities with CPTH6 produced similar effects on the autophagic pathway. Finally, the evidence that CPTH6 treatment decreased α-tubulin acetylation and failed to increase autophagic markers in cells in which acetyltransferase ATAT1 expression was silenced indicates a possible role of α-tubulin acetylation in

  9. Exploiting cannabinoid-induced cytotoxic autophagy to drive melanoma cell death.

    Science.gov (United States)

    Armstrong, Jane L; Hill, David S; McKee, Christopher S; Hernandez-Tiedra, Sonia; Lorente, Mar; Lopez-Valero, Israel; Eleni Anagnostou, Maria; Babatunde, Fiyinfoluwa; Corazzari, Marco; Redfern, Christopher P F; Velasco, Guillermo; Lovat, Penny E

    2015-06-01

    Although the global incidence of cutaneous melanoma is increasing, survival rates for patients with metastatic disease remain Sativex-like (a laboratory preparation comprising equal amounts of THC and cannabidiol (CBD)) to mice bearing BRAF wild-type melanoma xenografts substantially inhibited melanoma viability, proliferation, and tumor growth paralleled by an increase in autophagy and apoptosis compared with standard single-agent temozolomide. Collectively, our findings suggest that THC activates noncanonical autophagy-mediated apoptosis of melanoma cells, suggesting that cytotoxic autophagy induction with Sativex warrants clinical evaluation for metastatic disease. PMID:25674907

  10. Autophagy in cerebral ischemia and the effects of traditional Chinese medicine

    Institute of Scientific and Technical Information of China (English)

    Xiao-ping Huang; Huang Ding; Jin-dong Lu; Ying-hong Tang; Bing-xiang Deng; Chang-qing Deng

    2015-01-01

    Autophagy is a lysosome-mediated degradation process for non-essential or damaged celular constituents, playing an important homeostatic role in cel survival, differentiation and development to maintain homeostasis. Autophagy is involved in tumors as wel as neurodegenerative, cardiovascular and cerebrovascular diseases. Recently, active compounds from traditional Chinese medicine (TCM) have been found to modulate the levels of autophagy in tumor cels, nerve cels, myocardial cels and endothelial cels. Ischemic stroke is a major cause of neurological disability and places a heavy burden on family and society. Regaining function can signiifcantly reduce dependence and improve the quality of life of stroke survivors. In healthy cels, autophagy plays a key role in adapting to nutritional deprivation and eliminating aggregated proteins, however inappropriate activation of autophagy may lead to cel death in cerebral ischemia. This paper reviews the process and the molecular basis of autophagy, as wel as its roles in cerebral ischemia and the roles of TCM in modulating its activity.

  11. Protective autophagy antagonizes oxaliplatin-induced apoptosis in gastric cancer cells

    Institute of Scientific and Technical Information of China (English)

    Ling Xu; Xiu-Juan Qu; Yun-Peng Liu; Ying-Ying Xu; Jing Liu; Ke-Zuo Hou; Ye Zhang

    2011-01-01

    Oxaliplatin-based chemotherapy is used for treating gastric cancer. Autophagy has been extensively implicated in cancer cells; however, its function is not fully understood. Our study aimed to determine if oxaliplatin induce autophagy in gastric cancer MGC803 cells and to assess the effect of autophagy on apoptosis induced by oxaliplatin. MGC803 cells were cultured with oxaliplatin. Cell proliferation was measured using MTT assay, and apoptosis was determined by flow cytometry. Protein expression was detected by Western blot. Autophagy was observed using fluorescent microscopy. Our results showed that the rate of apoptosis was 9.73% and 16.36% when MGC803 cells were treated with 5 and 20 μg/mL oxaliplatin for 24 h, respectively. In addition, caspase activation and poly ADP-ribose polymerase (PARP)cleavage were detected. Furthermore, when MGC803 cells were treated with oxaliplatin for 24 h, an accumulation of punctate LC3 and an increase of LC3-Ⅱ protein were also detected, indicating the activation of autophagy. Phosphorylation of Akt and mTOR were inhibited by oxaliplatin. Compared to oxaliplatin alone, the combination of autophagy inhibitor chlorochine and oxaliplatin significantly enhanced the inhibition of cell proliferation and the induction of cell apoptosis. In conclusion, oxaliplatin-induced protective autophagy partially prevents apoptosis in gastric cancer MGC803 cells. The combination of autophagy inhibitor and oxaliplatin may be a new therapeutic option for gastric cancer.

  12. Human Immunodeficiency Virus Type 1 Nef Inhibits Autophagy through Transcription Factor EB Sequestration.

    Directory of Open Access Journals (Sweden)

    Grant R Campbell

    2015-06-01

    Full Text Available HIV Nef acts as an anti-autophagic maturation factor through interaction with beclin-1 (BECN1. We report that exposure of macrophages to infectious or non-infectious purified HIV induces toll-like receptor 8 (TLR8 and BECN1 dependent dephosphorylation and nuclear translocation of TFEB and that this correlates with an increase in autophagy markers. RNA interference for ATG13, TFEB, TLR8, or BECN1 inhibits this HIV-induced autophagy. However, once HIV establishes a productive infection, TFEB phosphorylation and cytoplasmic sequestration are increased resulting in decreased autophagy markers. Moreover, by 7 d post-infection, autophagy levels are similar to mock infected controls. Conversely, although Nef deleted HIV similarly induces TFEB dephosphorylation and nuclear localization, and increases autophagy, these levels remain elevated during continued productive infection. Thus, the interaction between HIV and TLR8 serves as a signal for autophagy induction that is dependent upon the dephosphorylation and nuclear translocation of TFEB. During permissive infection, Nef binds BECN1 resulting in mammalian target of rapamycin (MTOR activation, TFEB phosphorylation and cytosolic sequestration, and the inhibition of autophagy. To our knowledge, this is the first report of a virus modulating TFEB localization and helps to explain how HIV modulates autophagy to promote its own replication and cell survival.

  13. Regulatory coordination between two major intracellular homeostatic systems: heat shock response and autophagy.

    Science.gov (United States)

    Dokladny, Karol; Zuhl, Micah Nathaniel; Mandell, Michael; Bhattacharya, Dhruva; Schneider, Suzanne; Deretic, Vojo; Moseley, Pope Lloyd

    2013-05-24

    The eukaryotic cell depends on multitiered homeostatic systems ensuring maintenance of proteostasis, organellar integrity, function and turnover, and overall cellular viability. At the two opposite ends of the homeostatic system spectrum are heat shock response and autophagy. Here, we tested whether there are interactions between these homeostatic systems, one universally operational in all prokaryotic and eukaryotic cells, and the other one (autophagy) is limited to eukaryotes. We found that heat shock response regulates autophagy. The interaction between the two systems was demonstrated by testing the role of HSF-1, the central regulator of heat shock gene expression. Knockdown of HSF-1 increased the LC3 lipidation associated with formation of autophagosomal organelles, whereas depletion of HSF-1 potentiated both starvation- and rapamycin-induced autophagy. HSP70 expression but not expression of its ATPase mutant inhibited starvation or rapamycin-induced autophagy. We also show that exercise induces autophagy in humans. As predicted by our in vitro studies, glutamine supplementation as a conditioning stimulus prior to exercise significantly increased HSP70 protein expression and prevented the expected exercise induction of autophagy. Our data demonstrate for the first time that heat shock response, from the top of its regulatory cascade (HSF-1) down to the execution stages delivered by HSP70, controls autophagy thus connecting and coordinating the two extreme ends of the homeostatic systems in the eukaryotic cell. PMID:23576438

  14. Role of autophagy in disease resistance and hypersensitive response-associated cell death

    DEFF Research Database (Denmark)

    Hofius, Daniel; Munch, David; Bressendorff, Simon;

    2011-01-01

    documented, but how autophagy contributes to plant innate immunity and cell death is not that clear. A few research reports have appeared recently to shed light on the roles of autophagy in plant-pathogen interactions and in disease-associated host cell death. We present a first attempt to reconcile the......Ancient autophagy pathways are emerging as key defense modules in host eukaryotic cells against microbial pathogens. Apart from actively eliminating intracellular intruders, autophagy is also responsible for cell survival, for example by reducing the deleterious effects of endoplasmic reticulum...

  15. Irradiation conditions for induction of nitrite reducing activity of horse heart cytochrome c

    Energy Technology Data Exchange (ETDEWEB)

    Suruga, Kohei; Ogawa, Masahiro; Nishio, Toshiyuki; Oku, Tadatake [Nihon Univ., Fujisawa, Kanagawa (Japan); Nagasawa, Naotsugu; Kume, Tamikazu [Japan Atomic Energy Research Inst., Takasaki, Gunma (Japan). Takasaki Radiation Chemistry Research Establishment

    2000-09-01

    Denatured horse heart cytochrome (Cyt) c by gamma-irradiation showed high reducing activity of nitrite (NO{sub 2}{sup -}) to ammonia (NH{sub 4}{sup +}), in the presence of sodium dithionite and methylviologen. Effect of O{sub 2} on irradiation of the Cyt c and the detailed irradiation conditions for nitrite reducing activity of 0.03 mM Cyt c have been investigated. The Cyt c in an aqueous solution (0.03 mM) was treated by Co-60 gamma-irradiation at doses of 0.25-5.0 kGy, dose rates of 0.15-5.0 kGy/h under N{sub 2} or O{sub 2} atmosphere. The Cyt c babbled with O{sub 2} during irradiation showed much higher activity than that with N{sub 2}. The excellent irradiation conditions for increasing the activity of 0.03 mM Cyt c were total dose of 1.0 kGy under O{sub 2} atmosphere. The activity of irradiated Cyt c by these conditions was increased approximately 5 times of unirradiated one. (author)

  16. Autophagy in plants and phytopathogens.

    Science.gov (United States)

    Yoshimoto, Kohki; Takano, Yoshitaka; Sakai, Yasuyoshi

    2010-04-01

    Plants and plant-associated microorganisms including phytopathogens have to adapt to drastic changes in environmental conditions. Because of their immobility, plants must cope with various types of environmental stresses such as starvation, oxidative stress, drought stress, and invasion by phytopathogens during their differentiation, development, and aging processes. Here we briefly describe the early studies of plant autophagy, summarize recent studies on the molecular functions of ATG genes, and speculate on the role of autophagy in plants and phytopathogens. Autophagy regulates senescence and pathogen-induced cell death in plants, and autophagy and pexophagy play critical roles in differentiation and the invasion of host cells by phytopathogenic fungi. PMID:20079356

  17. Heme oxygenase-1 enhances autophagy in podocytes as a protective mechanism against high glucose-induced apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Dong, Chenglong [Department of Endocrinology, The Second Affiliated Hospital of Nanjing Medical University, Nanjing (China); Zheng, Haining [Department of Hyperbaric Oxygen, Nanjing General Hospital of Nanjing Military Command, Nanjing (China); Huang, Shanshan; You, Na; Xu, Jiarong; Ye, Xiaolong; Zhu, Qun; Feng, Yamin; You, Qiang; Miao, Heng [Department of Endocrinology, The Second Affiliated Hospital of Nanjing Medical University, Nanjing (China); Ding, Dafa, E-mail: dingdafa2004@aliyun.com [Department of Endocrinology, The Second Affiliated Hospital of Nanjing Medical University, Nanjing (China); Lu, Yibing, E-mail: luyibing2004@126.com [Department of Endocrinology, The Second Affiliated Hospital of Nanjing Medical University, Nanjing (China)

    2015-10-01

    Injury and loss of podocytes play vital roles in diabetic nephropathy progression. Emerging evidence suggests autophagy, which is induced by multiple stressors including hyperglycemia, plays a protective role. Meanwhile, heme oxygenase-1 (HO-1) possesses powerful anti-apoptotic properties. Therefore, we investigated the impact of autophagy on podocyte apoptosis under diabetic conditions and its association with HO-1. Mouse podocytes were cultured in vitro; apoptosis was detected by flow cytometry. Transmission electron microscopy and biochemical autophagic flux assays were used to measure the autophagy markers microtubule-associated protein 1 light chain 3-II (LC3-II) and beclin-1. LC3-II and beclin-1 expression peaked 12–24 h after exposing podocytes to high glucose. Inhibition of autophagy with 3-methyladenine or Beclin-1 siRNAs or Atg 5 siRNAs sensitized cells to apoptosis, suggesting autophagy is a survival mechanism. HO-1 inactivation inhibited autophagy, which aggravated podocyte injury in vitro. Hemin-induced autophagy also protected podocytes from hyperglycemia in vitro and was abrogated by HO-1 siRNA. Adenosine monophosphate-activated protein kinase phosphorylation was higher in hemin-treated and lower in HO-1 siRNA-treated podocytes. Suppression of AMPK activity reversed HO-1-mediated Beclin-1 upregulation and autophagy, indicating HO-1-mediated autophagy is AMPK dependent. These findings suggest HO-1 induction and regulation of autophagy are potential therapeutic targets for diabetic nephropathy. - Highlights: • High glucose leads to increased autophagy in podocytes at an early stage. • The early autophagic response protects against high glucose-induced apoptosis. • Heme oxygenase-1 enhances autophagy and decreases high glucose -mediated apoptosis. • Heme oxygenase-1 induces autophagy through the activation of AMPK.

  18. Heme oxygenase-1 enhances autophagy in podocytes as a protective mechanism against high glucose-induced apoptosis

    International Nuclear Information System (INIS)

    Injury and loss of podocytes play vital roles in diabetic nephropathy progression. Emerging evidence suggests autophagy, which is induced by multiple stressors including hyperglycemia, plays a protective role. Meanwhile, heme oxygenase-1 (HO-1) possesses powerful anti-apoptotic properties. Therefore, we investigated the impact of autophagy on podocyte apoptosis under diabetic conditions and its association with HO-1. Mouse podocytes were cultured in vitro; apoptosis was detected by flow cytometry. Transmission electron microscopy and biochemical autophagic flux assays were used to measure the autophagy markers microtubule-associated protein 1 light chain 3-II (LC3-II) and beclin-1. LC3-II and beclin-1 expression peaked 12–24 h after exposing podocytes to high glucose. Inhibition of autophagy with 3-methyladenine or Beclin-1 siRNAs or Atg 5 siRNAs sensitized cells to apoptosis, suggesting autophagy is a survival mechanism. HO-1 inactivation inhibited autophagy, which aggravated podocyte injury in vitro. Hemin-induced autophagy also protected podocytes from hyperglycemia in vitro and was abrogated by HO-1 siRNA. Adenosine monophosphate-activated protein kinase phosphorylation was higher in hemin-treated and lower in HO-1 siRNA-treated podocytes. Suppression of AMPK activity reversed HO-1-mediated Beclin-1 upregulation and autophagy, indicating HO-1-mediated autophagy is AMPK dependent. These findings suggest HO-1 induction and regulation of autophagy are potential therapeutic targets for diabetic nephropathy. - Highlights: • High glucose leads to increased autophagy in podocytes at an early stage. • The early autophagic response protects against high glucose-induced apoptosis. • Heme oxygenase-1 enhances autophagy and decreases high glucose -mediated apoptosis. • Heme oxygenase-1 induces autophagy through the activation of AMPK

  19. Cytotoxic Induction and Photoacoustic Imaging of Breast Cancer Cells Using Astaxanthin-Reduced Gold Nanoparticles

    Directory of Open Access Journals (Sweden)

    Subramaniyan Bharathiraja

    2016-04-01

    Full Text Available Astaxanthin, a kind of photosynthetic pigment, was employed for gold nanoparticle formation. Nanoparticles were characterized using Ulteraviolet-Visible (UV-Vis spectroscopy, transmission electron microscopy, and X-ray diffraction, and the possible presence of astaxanthin functional groups were analyzed by Fourier transform infrared spectroscopy (FTIR. The cytotoxic effect of synthesized nanoparticles was evaluated against MDA-MB-231 (human breast cancer cells using a tetrazolium-based assay, and synthesized nanoparticles exhibited dose-dependent toxicity. The morphology upon cell death was differentiated through fluorescent microscopy using different stains that predicted apoptosis. The synthesized nanoparticles were applied in ultrasound-coupled photoacoustic imaging to obtain good images of treated cells. Astaxanthin-reduced gold nanoparticle has the potential to act as a promising agent in the field of photo-based diagnosis and therapy.

  20. Autophagy in cardiovascular biology

    OpenAIRE

    Lavandero, Sergio; Chiong, Mario; Rothermel, Beverly A.; Hill, Joseph A.

    2015-01-01

    Cardiovascular disease is the leading cause of death worldwide. As such, there is great interest in identifying novel mechanisms that govern the cardiovascular response to disease-related stress. First described in failing hearts, autophagy within the cardiovascular system has been widely characterized in cardiomyocytes, cardiac fibroblasts, endothelial cells, vascular smooth muscle cells, and macrophages. In all cases, a window of optimal autophagic activity appears to be critical to the mai...

  1. AUTOPHAGY IN LUNG CANCER

    OpenAIRE

    Jaboin, Jerry J.; Hwang, Misun; Lu, Bo

    2009-01-01

    Lung cancer is the leading cause of cancer-related deaths worldwide. The relatively poor cure rate in lung cancer patients has been associated with a resistance to chemotherapy and radiation that is at least in part related to defects in cellular apoptotic machinery. Exploitation of another form of cell death, autophagy, has the capacity to improve the therapeutic gain of current therapies. In an effort to develop novel treatment strategies to enhance the therapeutic ratio for lung cancer, we...

  2. Targeting autophagy in cancer management – strategies and developments

    International Nuclear Information System (INIS)

    Autophagy is a highly regulated catabolic process involving lysosomal degradation of intracellular components, damaged organelles, misfolded proteins, and toxic aggregates, reducing oxidative stress and protecting cells from damage. The process is also induced in response to various conditions, including nutrient deprivation, metabolic stress, hypoxia, anticancer therapeutics, and radiation therapy to adapt cellular conditions for survival. Autophagy can function as a tumor suppressor mechanism in normal cells and dysregulation of this process (ie, monoallelic Beclin-1 deletion) may lead to malignant transformation and carcinogenesis. In tumors, autophagy is thought to promote tumor growth and progression by helping cells to adapt and survive in metabolically-challenged and harsh tumor microenvironments (ie, hypoxia and acidity). Recent in vitro and in vivo studies in preclinical models suggested that modulation of autophagy can be used as a therapeutic modality to enhance the efficacy of conventional therapies, including chemo and radiation therapy. Currently, more than 30 clinical trials are investigating the effects of autophagy inhibition in combination with cytotoxic chemotherapies and targeted agents in various cancers. In this review, we will discuss the role, molecular mechanism, and regulation of autophagy, while targeting this process as a novel therapeutic modality, in various cancers

  3. Autophagy in dementias.

    Science.gov (United States)

    Kragh, Christine Lund; Ubhi, Kiren; Wyss-Coray, Tony; Wyss-Corey, Tony; Masliah, Eliezer

    2012-01-01

    Dementias are a varied group of disorders typically associated with memory loss, impaired judgment and/or language and by symptoms affecting other cognitive and social abilities to a degree that interferes with daily functioning. Alzheimer's disease (AD) is the most common cause of a progressive dementia, followed by dementia with Lewy bodies (DLB), frontotemporal dementia (FTD), (VaD) and HIV-associated neurocognitive disorders (HAND). The pathogenesis of this group of disorders has been linked to the abnormal accumulation of proteins in the brains of affected individuals, which in turn has been related to deficits in protein clearance. Autophagy is a key cellular protein clearance pathway with proteolytic cleavage and degradation via the ubiquitin-proteasome pathway representing another important clearance mechanism. Alterations in the levels of autophagy and the proteins associated with the autophagocytic pathway have been reported in various types of dementias. This review will examine recent literature across these disorders and highlight a common theme of altered autophagy across the spectrum of the dementias. PMID:22150925

  4. Alcohol Dehydrogenase Protects against Endoplasmic Reticulum Stress-Induced Myocardial Contractile Dysfunction via Attenuation of Oxidative Stress and Autophagy: Role of PTEN-Akt-mTOR Signaling.

    Directory of Open Access Journals (Sweden)

    Jiaojiao Pang

    Full Text Available The endoplasmic reticulum (ER plays an essential role in ensuring proper folding of the newly synthesized proteins. Aberrant ER homeostasis triggers ER stress and development of cardiovascular diseases. ADH is involved in catalyzing ethanol to acetaldehyde although its role in cardiovascular diseases other than ethanol metabolism still remains elusive. This study was designed to examine the impact of ADH on ER stress-induced cardiac anomalies and underlying mechanisms involved using cardiac-specific overexpression of alcohol dehydrogenase (ADH.ADH and wild-type FVB mice were subjected to the ER stress inducer tunicamycin (1 mg/kg, i.p., for 48 hrs. Myocardial mechanical and intracellular Ca(2+ properties, ER stress, autophagy and associated cell signaling molecules were evaluated.ER stress compromised cardiac contractile function (evidenced as reduced fractional shortening, peak shortening, maximal velocity of shortening/relengthening, prolonged relengthening duration and impaired intracellular Ca(2+ homeostasis, oxidative stress and upregulated autophagy (increased LC3B, Atg5, Atg7 and p62, along with dephosphorylation of PTEN, Akt and mTOR, all of which were attenuated by ADH. In vitro study revealed that ER stress-induced cardiomyocyte anomaly was abrogated by ADH overexpression or autophagy inhibition using 3-MA. Interestingly, the beneficial effect of ADH was obliterated by autophagy induction, inhibition of Akt and mTOR. ER stress also promoted phosphorylation of the stress signaling ERK and JNK, the effect of which was unaffected by ADH transgene.Taken together, these findings suggested that ADH protects against ER stress-induced cardiac anomalies possibly via attenuation of oxidative stress and PTEN/Akt/mTOR pathway-regulated autophagy.

  5. SPBP is a sulforaphane induced transcriptional coactivator of NRF2 regulating expression of the autophagy receptor p62/SQSTM1.

    Directory of Open Access Journals (Sweden)

    Sagar Ramesh Darvekar

    Full Text Available Organisms exposed to oxidative stress respond by orchestrating a stress response to prevent further damage. Intracellular levels of antioxidant agents increase, and damaged components are removed by autophagy induction. The KEAP1-NRF2 signaling pathway is the main pathway responsible for cell defense against oxidative stress and for maintaining the cellular redox balance at physiological levels. Sulforaphane, an isothiocyanate derived from cruciferous vegetables, is a potent inducer of KEAP1-NRF2 signaling and antioxidant response element driven gene expression. In this study, we show that sulforaphane enhances the expression of the transcriptional coregulator SPBP. The expression curve peaks 6-8 hours post stimulation, and parallels the sulforaphane-induced expression of NRF2 and the autophagy receptor protein p62/SQSTM1. Reporter gene assays show that SPBP stimulates the expression of p62/SQSTM1 via ARE elements in the promoter region, and siRNA mediated knock down of SPBP significantly decreases the expression of p62/SQSTM1 and the formation of p62/SQSTM1 bodies in HeLa cells. Furthermore, SPBP siRNA reduces the sulforaphane induced expression of NRF2, and the expression of the autophagy marker protein LC3B. Both these proteins contain ARE-like elements in their promoter regions. Over-expressed SPBP and NRF2 acts synergistically on the p62/SQSTM1 promoter and colocalize in nuclear speckles in HeLa cells. Collectively, these results suggest that SPBP is a coactivator of NRF2, and hence may be important for securing enhanced and sustained expression of NRF2 induced genes such as proteins involved in selective autophagy.

  6. Sinomenine Hydrochloride Protects against Polymicrobial Sepsis via Autophagy

    Directory of Open Access Journals (Sweden)

    Yu Jiang

    2015-01-01

    Full Text Available Sepsis, a systemic inflammatory response to infection, is the major cause of death in intensive care units (ICUs. The mortality rate of sepsis remains high even though the treatment and understanding of sepsis both continue to improve. Sinomenine (SIN is a natural alkaloid extracted from Chinese medicinal plant Sinomenium acutum, and its hydrochloride salt (Sinomenine hydrochloride, SIN-HCl is widely used to treat rheumatoid arthritis (RA. However, its role in sepsis remains unclear. In the present study, we investigated the role of SIN-HCl in sepsis induced by cecal ligation and puncture (CLP in BALB/c mice and the corresponding mechanism. SIN-HCl treatment improved the survival of BALB/c mice that were subjected to CLP and reduced multiple organ dysfunction and the release of systemic inflammatory mediators. Autophagy activities were examined using Western blotting. The results showed that CLP-induced autophagy was elevated, and SIN-HCl treatment further strengthened the autophagy activity. Autophagy blocker 3-methyladenine (3-MA was used to investigate the mechanism of SIN-HCl in vitro. Autophagy activities were determined by examining the autophagosome formation, which was shown as microtubule-associated protein light chain 3 (LC3 puncta with green immunofluorescence. SIN-HCl reduced lipopolysaccharide (LPS-induced inflammatory cytokine release and increased autophagy in peritoneal macrophages (PM. 3-MA significantly decreased autophagosome formation induced by LPS and SIN-HCl. The decrease of inflammatory cytokines caused by SIN-HCl was partially aggravated by 3-MA treatment. Taken together, our results indicated that SIN-HCl could improve survival, reduce organ damage, and attenuate the release of inflammatory cytokines induced by CLP, at least in part through regulating autophagy activities.

  7. Burkholderia cenocepacia J2315 escapes to the cytosol and actively subverts autophagy in human macrophages.

    Science.gov (United States)

    Al-Khodor, Souhaila; Marshall-Batty, Kimberly; Nair, Vinod; Ding, Li; Greenberg, David E; Fraser, Iain D C

    2014-03-01

    Selective autophagy functions to specifically degrade cellular cargo tagged by ubiquitination, including bacteria. Strains of the Burkholderia cepacia complex (Bcc) are opportunistic pathogens that cause life-threatening infections in patients with cystic fibrosis (CF) and chronic granulomatous disease (CGD). While there is evidence that defective macrophage autophagy in a mouse model of CF can influence B. cenocepacia susceptibility, there have been no comprehensive studies on how this bacterium is sensed and targeted by the host autophagy response in human macrophages. Here, we describe the intracellular life cycle of B. cenocepacia J2315 and its interaction with the autophagy pathway in human cells. Electron and confocal microscopy analyses demonstrate that the invading bacteria interact transiently with the endocytic pathway before escaping to the cytosol. This escape triggers theselective autophagy pathway, and the recruitment of ubiquitin, the ubiquitin-binding adaptors p62 and NDP52 and the autophagosome membrane-associated protein LC3B, to the bacterial vicinity. However, despite recruitment of these key autophagy pathway effectors, B. cenocepacia blocks autophagosome completion and replicates in the host cytosol. We find that a pre-infection increase in cellular autophagy flux can significantly inhibit B. cenocepacia replication and that lower autophagy flux in macrophages from immunocompromised CGD patients could contribute to increased B. cenocepacia susceptibility, identifying autophagy manipulation as a potential therapeutic approach to reduce bacterial burden in B. cenocepacia infections. PMID:24119232

  8. Effectors of mTOR-autophagy pathway: targeting cancer, affecting the skeleton.

    Science.gov (United States)

    Chagin, Andrei S

    2016-06-01

    Although some modulators of autophagy are emerging as drugs or supplements for anti-cancer therapy, the effects of these compounds on normal tissues must be examined carefully. Here, I review the role of autophagy in skeletal tissues in this context. First, I briefly review preclinical studies indicating the role of autophagy in cancer, as well as related on-going clinical trials. Thereafter, the role of autophagy in the physiology of skeletal tissues is discussed, with a focus on recent genetic preclinical studies. Specifically, I discuss the mTOR-autophagy pathway in relationship to epiphyseal chondrocytes, articular chondrocytes, osteoblasts, osteocytes and osteoclasts and potential side effects of targeting either mTOR pathway or autophagy in general in connection with anti-cancer therapy. Current preclinical findings indicate that inhibiting autophagy will not seriously reduce bone mass and enhance osteoporosis. However, inhibition of autophagy might damage articular cartilage and cause osteoarthritis, whereas treatment with rapalogs might result in relatively beneficial effects on articular cartilage. Modulation of the mTOR pathway or autophagy during childhood may have an undesirable influence on adult height, as well as acquisition of bone mass. PMID:26921601

  9. Radiation-induced autophagy promotes esophageal squamous cell carcinoma cell survival via the LKB1 pathway.

    Science.gov (United States)

    Lu, Chi; Xie, Conghua

    2016-06-01

    Radiotherapy is an important treatment modality for esophageal cancer; however, the clinical efficacy of radiotherapy is limited by tumor radioresistance. In the present study, we explored the hypothesis that radiation induces tumor cell autophagy as a cytoprotective adaptive response, which depends on liver kinase B1 (LKB1) also known as serine/threonine kinase 11 (STK11). Radiation-induced Eca-109 cell autophagy was found to be dependent on signaling through the LKB1 pathway, and autophagy inhibitors that disrupted radiation-induced Eca-109 cell autophagy increased cell cycle arrest and cell death in vitro. Inhibition of autophagy also reduced the clonogenic survival of the Eca-109 cells. When treated with radiation alone, human esophageal carcinoma xenografts showed increased LC3B and p-LKB1 expression, which was decreased by the autophagy inhibitor chloroquine. In vivo inhibition of autophagy disrupted tumor growth and increased tumor apoptosis when combined with 6 Gy of ionizing radiation. In summary, our findings elucidate a novel mechanism of resistance to radiotherapy in which radiation-induced autophagy, via the LKB1 pathway, promotes tumor cell survival. This indicates that inhibition of autophagy can serve as an adjuvant treatment to improve the curative effect of radiotherapy. PMID:27109915

  10. Regulation of Autophagy by Kinases

    International Nuclear Information System (INIS)

    Autophagy is a process of self-degradation that maintains cellular viability during periods of metabolic stress. Although autophagy is considered a survival mechanism when faced with cellular stress, extensive autophagy can also lead to cell death. Aberrations in autophagy are associated with several diseases, including cancer. Therapeutic exploitation of this process requires a clear understanding of its regulation. Although the core molecular components involved in the execution of autophagy are well studied there is limited information on how cellular signaling pathways, particularly kinases, regulate this complex process. Protein kinases are integral to the autophagy process. Atg1, the first autophagy-related protein identified, is a serine/threonine kinase and it is regulated by another serine/threonine kinase mTOR. Emerging studies suggest the participation of many different kinases in regulating various components/steps of this catabolic process. This review focuses on the regulation of autophagy by several kinases with particular emphasis on serine/threonine protein kinases such as mTOR, AMP-activated protein kinase, Akt, mitogen-activated protein kinase (ERK, p38 and JNK) and protein kinase C that are often deregulated in cancer and are important therapeutic targets

  11. Autophagy Promotes Focal Adhesion Disassembly and Cell Motility of Metastatic Tumor Cells through the Direct Interaction of Paxillin with LC3.

    Science.gov (United States)

    Sharifi, Marina N; Mowers, Erin E; Drake, Lauren E; Collier, Chris; Chen, Hong; Zamora, Marta; Mui, Stephanie; Macleod, Kay F

    2016-05-24

    Autophagy is a conserved catabolic process that plays a housekeeping role in eliminating protein aggregates and organelles and is activated during nutrient deprivation to generate metabolites and energy. Autophagy plays a significant role in tumorigenesis, although opposing context-dependent functions of autophagy in cancer have complicated efforts to target autophagy for therapeutic purposes. We demonstrate that autophagy inhibition reduces tumor cell migration and invasion in vitro and attenuates metastasis in vivo. Numerous abnormally large focal adhesions (FAs) accumulate in autophagy-deficient tumor cells, reflecting a role for autophagy in FA disassembly through targeted degradation of paxillin. We demonstrate that paxillin interacts with processed LC3 through a conserved LIR motif in the amino-terminal end of paxillin and that this interaction is regulated by oncogenic SRC activity. Together, these data establish a function for autophagy in FA turnover, tumor cell motility, and metastasis. PMID:27184837

  12. The late stage of autophagy: cellular events and molecular regulation

    OpenAIRE

    Tong, Jingjing; Yan, Xianghua; Yu, Li

    2010-01-01

    Autophagy is an intracellular degradation system that delivers cytoplasmic contents to the lysosome for degradation. It is a “self-eating” process and plays a “house-cleaner” role in cells. The complex process consists of several sequential steps—induction, autophagosome formation, fusion of lysosome and autophagosome, degradation, efflux transportation of degradation products, and autophagic lysosome reformation. In this review, the cellular and molecular regulations of late stage of autopha...

  13. Peptide deformylase inhibitor actinonin reduces celastrol’s HSP70 induction while synergizing proliferation inhibition in tumor cells

    International Nuclear Information System (INIS)

    Celastrol is a promising anti-tumor agent, yet it also elevates heat shock proteins (HSPs), especially HSP70, this effect believed to reduce its anti-tumor effects. Concurrent use of siRNA to increase celastrol’s anti-tumor effects through HSP70 interference has been reported, but because siRNA technology is difficult to clinically apply, an alternative way to curb unwanted HSP70 elevation caused by celastrol treatment is worth exploring. In this work, we explore three alternative strategies to control HSP70 elevation: (1) Searching for cancer cell types that show no HSP70 elevation in the presence of celastrol (thus recommending themselves as suitable targets); (2) Modifying HSP70-inducing chemical groups, i.e.: the carboxyl group in celastrol; and (3) Using signaling molecule inhibitors to specifically block HSP70 elevation while protecting and/or enhancing anti-tumor effects. The first strategy was unsuccessful since celastrol treatment increased HSP70 in all 7 of the cancer cell types tested, this result related to HSF1 activation. The ubiquity of HSF1 expression in different cancer cells might explain why celastrol has no cell-type limitation for HSP70 induction. The second strategy revealed that modification of celastrol’s carboxyl group abolished its ability to elevate HSP70, but also abolished celastrol’s tumor inhibition effects. In the third strategy, 11 inhibitors for 10 signaling proteins reportedly related to celastrol action were tested, and five of these could reduce celastrol-caused HSP70 elevation. Among these, the peptide deformylase (PDF) inhibitor, actinonin, could synergize celastrol’s proliferation inhibition. Concurrent use of the chemical agent actinonin could reduce celastrol’s HSP70 elevation and also enhance proliferation inhibition by celastrol. This combination presents a novel alternative to siRNA technology and is worth further investigation for its potentially effective anti-tumor action

  14. Structure biology of selective autophagy receptors

    Science.gov (United States)

    Kim, Byeong-Won; Kwon, Do Hoon; Song, Hyun Kyu

    2016-01-01

    Autophagy is a process tightly regulated by various autophagy-related proteins. It is generally classified into non-selective and selective autophagy. Whereas non-selective autophagy is triggered when the cell is under starvation, selective autophagy is involved in eliminating dysfunctional organelles, misfolded and/or ubiquitylated proteins, and intracellular pathogens. These components are recognized by autophagy receptors and delivered to phagophores. Several selective autophagy receptors have been identified and characterized. They usually have some common domains, such as motif, a specific cargo interacting (ubiquitin-dependent or ubiquitin-independent) domain. Recently, structural data of these autophagy receptors has been described, which provides an insight of their function in the selective autophagic process. In this review, we summarize the most up-to-date findings about the structure-function of autophagy receptors that regulates selective autophagy. [BMB Reports 2016; 49(2): 73-80] PMID:26698872

  15. Activation of antibacterial autophagy by NADPH oxidases

    OpenAIRE

    Huang, Ju; Canadien, Veronica; Lam, Grace Y.; Steinberg, Benjamin E.; Mary C. Dinauer; Magalhaes, Marco A. O.; Glogauer, Michael; Grinstein, Sergio; Brumell, John H.

    2009-01-01

    Autophagy plays an important role in immunity to microbial pathogens. The autophagy system can target bacteria in phagosomes, promoting phagosome maturation and preventing pathogen escape into the cytosol. Recently, Toll-like receptor (TLR) signaling from phagosomes was found to initiate their targeting by the autophagy system, but the mechanism by which TLR signaling activates autophagy is unclear. Here we show that autophagy targeting of phagosomes is not exclusive to those containing TLR l...

  16. Improvement of low speed induction generator performances and reducing the power of excitation and voltage control system

    Energy Technology Data Exchange (ETDEWEB)

    Budisan, N. [Politechnica Univ. of Timisoara (Romania); Hentea, T.; Mahil, S. [Purdue Univ. Calumet, Hammond, IN (United States); Madescu, G. [Romanian Academy, Timisoara (Romania)

    1996-12-31

    In this paper we present the results of our investigations concerning the utilization of induction generators at very low speed. It is shown that, by proper design, it is possible to obtain high efficiency and high power factor values. The optimized induction generators require lower reactive power resulting in lower size and price of the excitation control system. 4 refs., 2 figs.

  17. A hybrid model of molecular regulation and population dynamics for yeast autophagy.

    Science.gov (United States)

    Jin, Huiqin; Lei, Jinzhi

    2016-08-01

    Autophagy is an evolutionarily conserved lysosome-based degradation process that is involved in maintaining cellular homeostasis and stress responses. Dysregulation of autophagy is known to associate with many diseases. In this paper, we establish a Hybrid model of Molecular regulation and Population dynamics (HMP model) for yeast autophagy to study how autophagy regulation at molecular level affects the cell population dynamics under the stress of starvation. The model includes interactions between amino acids, TORC1, Atg1 complex, and Atg8 lipidation at the molecular level, and cell death and division at the cell behavior level. Two feedback loops are involved in autophagy induction, in which the negative feedback of TORC1 activation has been known previously, and the positive feedback between TORC1 and Atg1 complex formation is introduced according to the similarity of Drosophila and mammalian cells. We demonstrate that the two feedback loops play distinct roles in autophagy regulation. The positive feedback is pro-survival, whereas the negative feedback has little effect on the survival of population during starvation. In addition, autophagy deficient cells can be rescued from starvation by amino acid exchanges from their neighboring wild type cells. PMID:27103581

  18. Crocin-Elicited Autophagy Rescues Myocardial Ischemia/Reperfusion Injury via Paradoxical Mechanisms.

    Science.gov (United States)

    Zeng, Chao; Li, Hu; Fan, Zhiwen; Zhong, Lei; Guo, Zhen; Guo, Yaping; Xi, Yusheng

    2016-01-01

    Crocin, the main effective component of saffron, exerts protective effects against ischemia/reperfusion injury during strokes. However, the effects of crocin in myocardial ischemia/reperfusion injury, and the mechanisms involved, remain unknown. Pretreated with crocin for 7 days, C57BL/6N mice were subjected to 30 min of myocardial ischemia followed by 12[Formula: see text]h of reperfusion (for cardiac function and infarct size, cell apoptosis and necrosis). Neonatal mouse cardiomyocytes were subjected to 2 h of hypoxia followed by 4 h of reoxygenation. NMCM's survival was assessed during hypoxia and reoxygenation in the presence or absence of the autophagy inhibitor 3-methyladenine or the inducer rapamycin. Western blotting was used to evaluate AMPK, Akt, and autophagy-related proteins. Autophagosome was observed using electron microscopy. In the in vivo experiment, crocin pretreatment significantly attenuated infarct size, myocardial apoptosis and necrosis, and improved left ventricular function following ischemia/reperfusion. In vitro data revealed that autophagy was induced during hypoxia, the levels of which were intensely elevated during reoxygenation. Crocin significantly promoted autophagy during ischemia, accompanied with the activation of AMPK. In contrast, crocin overtly inhibited autophagy during reperfusion, accompanied with Akt activation. Induction and inhibition of autophagy mitigated crocin induced protection against NMCMs injury during hypoxia and reoxygenation, respectively. Our data suggest that crocin demonstrated a myocardial protective effect via AMPK/mTOR and Akt/mTOR regulated autophagy against ischemia and reperfusion injury, respectively. PMID:27109157

  19. Autophagy Accompanied with Bisdemethoxycurcumin-induced Apoptosis in Non-small Cell Lung Cancer Cells

    Institute of Scientific and Technical Information of China (English)

    XU Jin Hong; YANG He Ping; ZHOU Xiang Dong; WANG Hai Jing; GONG Liang; TANG Chun Lan

    2015-01-01

    Objective To investigate the effects of bisdemethoxycurcumin (BDMC) on non-small cell lung cancer (NSCLC) cell line, A549, and the highly metastatic lung cancer 95D cells. Methods CCK-8 assay was used to assess the effect of BDMC on cytotoxicity. Flow cytometry was used to evaluate apoptosis. Western blot analysis, electron microscopy, and quantification of GFP-LC3 punctuates were used to test the effect of BDMC on autophagy and apoptosis of lung cancer cells. Results BDMC inhibited the viability of NSCLC cells, but had no cytotoxic effects on lung small airway epithelial cells (SAECs). The apoptotic cell death induced by BDMC was accompanied with the induction of autophagy in NSCLC cells. Blockage of autophagy by the autophagy inhibitor 3-methyladenine (3-MA) repressed the growth inhibitory effects and induction of apoptosis by BDMC. In addition, BDMC treatment significantly decreased smoothened (SMO) and the transcription factor glioma-associated oncogene 1 (Gli1) expression. Furthermore, depletion of Gli1 by siRNA and cyclopamine (a specific SMO inhibitor) induced autophagy. Conclusion Aberrant activation of Hedgehog (Hh) signaling has been implicated in several human cancers, including lung cancers. The present findings provide direct evidence that BDMC-induced autophagy plays a pro-death role in NSCLC, in part, by inhibiting Hedgehog signaling.

  20. Autophagy and neurodegenerative disorders

    Institute of Scientific and Technical Information of China (English)

    Evangelia Kesidou; Roza Lagoudaki; Olga Touloumi; Kyriaki-Nefeli Poulatsidou; Constantina Simeonidou

    2013-01-01

    Accumulation of aberrant proteins and inclusion bodies are hallmarks in most neurodegenerative diseases. Consequently, these aggregates within neurons lead to toxic effects, overproduction of reactive oxygen species and oxidative stress. Autophagy is a significant intracel ular mechanism that removes damaged organelles and misfolded proteins in order to maintain cel homeostasis. Excessive or insufficient autophagic activity in neurons leads to altered homeostasis and influences their survival rate, causing neurodegeneration. The review article provides an update of the role of autophagic process in representative chronic and acute neurodegenerative disorders.

  1. Mfn2 deficiency links age-related sarcopenia and impaired autophagy to activation of an adaptive mitophagy pathway.

    Science.gov (United States)

    Sebastián, David; Sorianello, Eleonora; Segalés, Jessica; Irazoki, Andrea; Ruiz-Bonilla, Vanessa; Sala, David; Planet, Evarist; Berenguer-Llergo, Antoni; Muñoz, Juan Pablo; Sánchez-Feutrie, Manuela; Plana, Natàlia; Hernández-Álvarez, María Isabel; Serrano, Antonio L; Palacín, Manuel; Zorzano, Antonio

    2016-08-01

    Mitochondrial dysfunction and accumulation of damaged mitochondria are considered major contributors to aging. However, the molecular mechanisms responsible for these mitochondrial alterations remain unknown. Here, we demonstrate that mitofusin 2 (Mfn2) plays a key role in the control of muscle mitochondrial damage. We show that aging is characterized by a progressive reduction in Mfn2 in mouse skeletal muscle and that skeletal muscle Mfn2 ablation in mice generates a gene signature linked to aging. Furthermore, analysis of muscle Mfn2-deficient mice revealed that aging-induced Mfn2 decrease underlies the age-related alterations in metabolic homeostasis and sarcopenia. Mfn2 deficiency reduced autophagy and impaired mitochondrial quality, which contributed to an exacerbated age-related mitochondrial dysfunction. Interestingly, aging-induced Mfn2 deficiency triggers a ROS-dependent adaptive signaling pathway through induction of HIF1α transcription factor and BNIP3. This pathway compensates for the loss of mitochondrial autophagy and minimizes mitochondrial damage. Our findings reveal that Mfn2 repression in muscle during aging is a determinant for the inhibition of mitophagy and accumulation of damaged mitochondria and triggers the induction of a mitochondrial quality control pathway. PMID:27334614

  2. Sodium Butyrate Induces Endoplasmic Reticulum Stress and Autophagy in Colorectal Cells: Implications for Apoptosis.

    Directory of Open Access Journals (Sweden)

    Jintao Zhang

    Full Text Available Butyrate, a short-chain fatty acid derived from dietary fiber, inhibits proliferation and induces cell death in colorectal cancer cells. However, clinical trials have shown mixed results regarding the anti-tumor activities of butyrate. We have previously shown that sodium butyrate increases endoplasmic reticulum stress by altering intracellular calcium levels, a well-known autophagy trigger. Here, we investigated whether sodium butyrate-induced endoplasmic reticulum stress mediated autophagy, and whether there was crosstalk between autophagy and the sodium butyrate-induced apoptotic response in human colorectal cancer cells.Human colorectal cancer cell lines (HCT-116 and HT-29 were treated with sodium butyrate at concentrations ranging from 0.5-5mM. Cell proliferation was assessed using MTT tetrazolium salt formation. Autophagy induction was confirmed through a combination of Western blotting for associated proteins, acridine orange staining for acidic vesicles, detection of autolysosomes (MDC staining, and electron microscopy. Apoptosis was quantified by flow cytometry using standard annexinV/propidium iodide staining and by assessing PARP-1 cleavage by Western blot.Sodium butyrate suppressed colorectal cancer cell proliferation, induced autophagy, and resulted in apoptotic cell death. The induction of autophagy was supported by the accumulation of acidic vesicular organelles and autolysosomes, and the expression of autophagy-associated proteins, including microtubule-associated protein II light chain 3 (LC3-II, beclin-1, and autophagocytosis-associated protein (Atg3. The autophagy inhibitors 3-methyladenine (3-MA and chloroquine inhibited sodium butyrate induced autophagy. Furthermore, sodium butyrate treatment markedly enhanced the expression of endoplasmic reticulum stress-associated proteins, including BIP, CHOP, PDI, and IRE-1a. When endoplasmic reticulum stress was inhibited by pharmacological (cycloheximide and mithramycin and genetic

  3. Degradation of paternal mitochondria by fertilization-triggered autophagy in C. elegans embryos.

    Science.gov (United States)

    Sato, Miyuki; Sato, Ken

    2011-11-25

    The mitochondrial genome is believed to be maternally inherited in many eukaryotes. Sperm-derived paternal mitochondria enter the oocyte cytoplasm upon fertilization and then normally disappear during early embryogenesis. However, the mechanism responsible for this clearance has been unknown. Here, we show that autophagy, which delivers cytosolic components to lysosomes for degradation, is required for the elimination of paternal mitochondria in Caenorhabditis elegans. Immediately after fertilization, sperm-derived components trigger the localized induction of autophagy around sperm mitochondria. Autophagosomes engulf paternal mitochondria, resulting in their lysosomal degradation during early embryogenesis. In autophagy-defective zygotes, paternal mitochondria and their genome remain even in the first larval stage. Thus, fertilization-triggered autophagy is required for selective degradation of paternal mitochondria and thereby maternal inheritance of mitochondrial DNA. PMID:21998252

  4. ER stress, autophagy, and RNA viruses

    Directory of Open Access Journals (Sweden)

    Jia-Rong eJheng

    2014-08-01

    Full Text Available Endoplasmic reticulum (ER stress is a general term for representing the pathway by which various stimuli affect ER functions. ER stress induces the evolutionarily conserved signaling pathways, called the unfolded protein response (UPR, which compromises the stimulus and then determines whether the cell survives or dies. In recent years, ongoing research has suggested that these pathways may be linked to the autophagic response, which plays a key role in the cell’s response to various stressors. Autophagy performs a self-digestion function, and its activation protects cells against certain pathogens. However, the link between the UPR and autophagy may be more complicated. These two systems may act dependently, or the induction of one system may interfere with the other. Experimental studies have found that different viruses modulate these mechanisms to allow them to escape the host immune response or, worse, to exploit the host’s defense to their advantage; thus, this topic is a critical area in antiviral research. In this review, we summarize the current knowledge about how RNA viruses, including influenza virus, poliovirus, coxsackievirus, enterovirus 71, Japanese encephalitis virus, hepatitis C virus, and dengue virus, regulate these processes. We also discuss recent discoveries and how these will produce novel strategies for antiviral treatment.

  5. Beclin1-induced autophagy abrogates radioresistance of lung cancer cells by suppressing osteopontin

    International Nuclear Information System (INIS)

    Osteopontin (OPN) serves as an indicator of resistance to radiotherapy. However, the role of OPN in the development of acquired radioresistance in human lung cancer cells has not yet been fully elucidated. Therefore, the potential importance of OPN as a marker of lung cancer with a potential significant role in the development of radioresistance against repeated radiotherapy has prompted us to define the pathways by which OPN regulates lung cancer cell growth. In addition, autophagy has been reported to play a key role in the radiosensitization of cancer cells. Here, we report that increased OPN expression through induction of nuclear p53 following irradiation was inhibited by exogenous beclin-1 (BECN1). Our results clearly show that BECN1 gene expression led to induction of autophagy and inhibition of cancer cell growth and angiogenesis. Our results suggest that the induction of autophagy abrogated the radioresistance of the cancer cells. Interestingly, we showed that knockdown of OPN by lentivirus-mediated shRNA induced the autophagy of human lung cancer cell. Taken together, these results suggest that OPN and BECN1 can be molecular targets for overcoming radioresistance by controlling autophagy. (author)

  6. Digital Simulation of Conventional Direct Torque Control of Induction Motor Drive with Reduced Flux and Torque Ripples.

    OpenAIRE

    Mr.P.Nagasekhar Reddy

    2013-01-01

    The induction motors are known as the workhorses of industry because of its simple construction and robustness. Traditionally, the induction motor has been used in constant and variable speed drive applications that do not cater for fast dynamic processes. Because of recent development of several new control technologies, such as vector control, sensor less control and direct torque control (DTC), the situation is changing rapidly. The Direct Torque Control (DTC) technique has the feature of ...

  7. Autophagy and Human Parturition: Evaluation of LC3 Expression in Placenta from Spontaneous or Medically Induced Onset of Labor

    Directory of Open Access Journals (Sweden)

    Laura Avagliano

    2013-01-01

    Full Text Available Induction of labor is one of the most used procedures in obstetrics, performed to achieve vaginal delivery through cervical ripening and stimulation of uterine contractions. We investigated the impact of induction of labor upon placental autophagy, a catabolic pathway activated in response to alteration of the physiological intracellular conditions. We collected 28 singleton placentas at the time of uncomplicated term vaginal delivery (7 spontaneous onset of labor, 21 induced labor. Autophagy was evaluated by immunohistochemistry, immunofluorescence, and immunoblotting. No significant difference in the autophagy expression was found between spontaneous or induced onset of labor. We found an inverse relationship between autophagy expression and the maternal prepregnancy body mass index, irrespective of the mode of labor onset. This result could be related to the nutritional maternal habits before and throughout pregnancy rather than rapid metabolic changes during labor.

  8. Neuronal Autophagy and Neurodevelopmental Disorders

    OpenAIRE

    Lee, Kyung-Min; Hwang, Su-Kyung; Lee, Jin-A

    2013-01-01

    Neurodevelopmental disorders include a wide range of diseases such as autism spectrum disorders and mental retardation. Mutations in several genes that regulate neural development and synapse function have been identified in neurodevelopmental disorders. Interestingly, some affected genes and pathways in these diseases are associated with the autophagy pathway. Autophagy is a complex, bulky degradative process that involves the sequestration of cellular proteins, RNA, lipids, and cellular org...

  9. Porcine Circovirus Type 2 Activates CaMMKβ to Initiate Autophagy in PK-15 Cells by Increasing Cytosolic Calcium

    OpenAIRE

    Yuanxing Gu; Baozhu Qi; Yingshan Zhou; Xiaowu Jiang; Xian Zhang; Xiaoliang Li; Weihuan Fang

    2016-01-01

    Porcine circovirus type 2 (PCV2) induces autophagy via the 5′ adenosine monophosphate-activated protein kinase (AMPK)/extracellular signal-regulated kinase (ERK)/tuberous sclerosis complex 2 (TSC2)/mammalian target of rapamycin (mTOR) pathway in pig kidney PK-15 cells. However, the underlying mechanisms of AMPK activation in autophagy induction remain unknown. With specific inhibitors and RNA interference (RNAi), we show that PCV2 infection upregulated calcium/calmodulin-dependent protein kin...

  10. Autophagy in DNA Damage Response

    Directory of Open Access Journals (Sweden)

    Piotr Czarny

    2015-01-01

    Full Text Available DNA damage response (DDR involves DNA repair, cell cycle regulation and apoptosis, but autophagy is also suggested to play a role in DDR. Autophagy can be activated in response to DNA-damaging agents, but the exact mechanism underlying this activation is not fully understood, although it is suggested that it involves the inhibition of mammalian target of rapamycin complex 1 (mTORC1. mTORC1 represses autophagy via phosphorylation of the ULK1/2–Atg13–FIP200 complex thus preventing maturation of pre-autophagosomal structures. When DNA damage occurs, it is recognized by some proteins or their complexes, such as poly(ADPribose polymerase 1 (PARP-1, Mre11–Rad50–Nbs1 (MRN complex or FOXO3, which activate repressors of mTORC1. SQSTM1/p62 is one of the proteins whose levels are regulated via autophagic degradation. Inhibition of autophagy by knockout of FIP200 results in upregulation of SQSTM1/p62, enhanced DNA damage and less efficient damage repair. Mitophagy, one form of autophagy involved in the selective degradation of mitochondria, may also play role in DDR. It degrades abnormal mitochondria and can either repress or activate apoptosis, but the exact mechanism remains unknown. There is a need to clarify the role of autophagy in DDR, as this process may possess several important biomedical applications, involving also cancer therapy.

  11. Autophagy and intestinal homeostasis.

    Science.gov (United States)

    Patel, Khushbu K; Stappenbeck, Thaddeus S

    2013-01-01

    Nutrient absorption is the basic function that drives mammalian intestinal biology. To facilitate nutrient uptake, the host's epithelial barrier is composed of a single layer of cells. This constraint is problematic, as a design of this type can be easily disrupted. The solution during the course of evolution was to add numerous host defense mechanisms that can help prevent local and systemic infection. These mechanisms include specialized epithelial cells that produce a physiochemical barrier overlying the cellular barrier, robust and organized adaptive and innate immune cells, and the ability to mount an inflammatory response that is commensurate with a specific threat level. The autophagy pathway is a critical cellular process that strongly influences all these functions. Therefore, a fundamental understanding of the components of this pathway and their influence on inflammation, immunity, and barrier function will facilitate our understanding of homeostasis in the gastrointestinal tract. PMID:23216414

  12. Neem oil limonoids induces p53-independent apoptosis and autophagy.

    Science.gov (United States)

    Srivastava, Pragya; Yadav, Neelu; Lella, Ravi; Schneider, Andrea; Jones, Anthony; Marlowe, Timothy; Lovett, Gabrielle; O'Loughlin, Kieran; Minderman, Hans; Gogada, Raghu; Chandra, Dhyan

    2012-11-01

    Azadirachta indica, commonly known as neem, has a wide range of medicinal properties. Neem extracts and its purified products have been examined for induction of apoptosis in multiple cancer cell types; however, its underlying mechanisms remain undefined. We show that neem oil (i.e., neem), which contains majority of neem limonoids including azadirachtin, induced apoptotic and autophagic cell death. Gene silencing demonstrated that caspase cascade was initiated by the activation of caspase-9, whereas caspase-8 was also activated late during neem-induced apoptosis. Pretreatment of cancer cells with pan caspase inhibitor, z-VAD inhibited activities of both initiator caspases (e.g., caspase-8 and -9) and executioner caspase-3. Neem induced the release of cytochrome c and apoptosis-inducing factor (AIF) from mitochondria, suggesting the involvement of both caspase-dependent and AIF-mediated apoptosis. p21 deficiency caused an increase in caspase activities at lower doses of neem, whereas p53 deficiency did not modulate neem-induced caspase activation. Additionally, neem treatment resulted in the accumulation of LC3-II in cancer cells, suggesting the involvement of autophagy in neem-induced cancer cell death. Low doses of autophagy inhibitors (i.e., 3-methyladenine and LY294002) did not prevent accumulation of neem-induced LC3-II in cancer cells. Silencing of ATG5 or Beclin-1 further enhanced neem-induced cell death. Phosphoinositide 3-kinase (PI3K) or autophagy inhibitors increased neem-induced caspase-3 activation and inhibition of caspases enhanced neem-induced autophagy. Together, for the first time, we demonstrate that neem induces caspase-dependent and AIF-mediated apoptosis, and autophagy in cancer cells. PMID:22915764

  13. Autophagy Protects against CYP2E1/Chronic Ethanol-Induced Hepatotoxicity

    Directory of Open Access Journals (Sweden)

    Yongke Lu

    2015-10-01

    Full Text Available Autophagy is an intracellular pathway by which lysosomes degrade and recycle long-lived proteins and cellular organelles. The effects of ethanol on autophagy are complex but recent studies have shown that autophagy serves a protective function against ethanol-induced liver injury. Autophagy was found to also be protective against CYP2E1-dependent toxicity in vitro in HepG2 cells which express CYP2E1 and in vivo in an acute alcohol/CYPE1-dependent liver injury model. The goal of the current report was to extend the previous in vitro and acute in vivo experiments to a chronic ethanol model to evaluate whether autophagy is also protective against CYP2E1-dependent liver injury in a chronic ethanol-fed mouse model. Wild type (WT, CYP2E1 knockout (KO or CYP2E1 humanized transgenic knockin (KI, mice were fed an ethanol liquid diet or control dextrose diet for four weeks. In the last week, some mice received either saline or 3-methyladenine (3-MA, an inhibitor of autophagy, or rapamycin, which stimulates autophagy. Inhibition of autophagy by 3-MA potentiated the ethanol-induced increases in serum transaminase and triglyceride levels in the WT and KI mice but not KO mice, while rapamycin prevented the ethanol liver injury. Treatment with 3-MA enhanced the ethanol-induced fat accumulation in WT mice and caused necrosis in the KI mice; little or no effect was found in the ethanol-fed KO mice or any of the dextrose-fed mice. 3-MA treatment further lowered the ethanol-decrease in hepatic GSH levels and further increased formation of TBARS in WT and KI mice, whereas rapamycin blunted these effects of ethanol. Neither 3-MA nor rapamycin treatment affected CYP2E1 catalytic activity or content or the induction CYP2E1 by ethanol. The 3-MA treatment decreased levels of Beclin-1 and Atg 7 but increased levels of p62 in the ethanol-fed WT and KI mice whereas rapamycin had the opposite effects, validating inhibition and stimulation of autophagy, respectively. These

  14. Autophagy and apoptosis: rivals or mates?

    Institute of Scientific and Technical Information of China (English)

    Yan Cheng; Jin-Ming Yang

    2013-01-01

    Autophagy,a cellular process of "self-eating" by which intracellular components are degraded within the lysosome,is an evolutionarily conserved response to various stresses.Autophagy is associated with numerous patho-physiological conditions,and dysregulation of autophagy contributes to the pathogenesis of a variety of human diseases including cancer.Depending on context,activation of autophagy may promote either cell survival or death,two major events that determine pathological process of many illnesses.Importantly,the activity of autophagy is often associated with apoptosis,another critical cellular process determining cellular fate.A better understanding of biology of autophagy and its implication in human health and disorder,as well as the relationship between autophagy and apoptosis,has the potential of facilitating the development of autophagy-based therapeutic interventions for human diseases such as cancer.

  15. Fluorescence microscopy: A tool to study autophagy

    Science.gov (United States)

    Rai, Shashank; Manjithaya, Ravi

    2015-08-01

    Autophagy is a cellular recycling process through which a cell degrades old and damaged cellular components such as organelles and proteins and the degradation products are reused to provide energy and building blocks. Dysfunctional autophagy is reported in several pathological situations. Hence, autophagy plays an important role in both cellular homeostasis and diseased conditions. Autophagy can be studied through various techniques including fluorescence based microscopy. With the advancements of newer technologies in fluorescence microscopy, several novel processes of autophagy have been discovered which makes it an essential tool for autophagy research. Moreover, ability to tag fluorescent proteins with sub cellular targets has enabled us to evaluate autophagy processes in real time under fluorescent microscope. In this article, we demonstrate different aspects of autophagy in two different model organisms i.e. yeast and mammalian cells, with the help of fluorescence microscopy.

  16. Autophagy and apoptosis: rivals or mates?

    Directory of Open Access Journals (Sweden)

    Yan Cheng

    2013-03-01

    Full Text Available Autophagy, a cellular process of "self-eating" by which intracellular components are degraded within the lysosome, is an evolutionarily conserved response to various stresses. Autophagy is associated with numerous patho-physiological conditions, and dysregulation of autophagy contributes to the pathogenesis of a variety of human diseases including cancer. Depending on context, activation of autophagy may promote either cell survival or death, two major events that determine pathological process of many illnesses. Importantly, the activity of autophagy is often associated with apoptosis, another critical cellular process determining cellular fate. A better understanding of biology of autophagy and its implication in human health and disorder, as well as the relationship between autophagy and apoptosis, has the potential of facilitating the development of autophagy-based therapeutic interventions for human diseases such as cancer.

  17. Membrane proteomics of phagosomes suggests a connection to autophagy

    Energy Technology Data Exchange (ETDEWEB)

    Shui, Wenqing; Sheu, Leslie; Liu, Jun; Smart, Brian; Petzold, Christopher J.; Hsieh, Tsung-yen; Pitcher, Austin; Keasling*, Jay D.; Bertozzi*, Carolyn R.

    2008-11-25

    Phagocytosis is the central process by which macrophage cellsinternalize and eliminate infectious microbes as well as apoptoticcells. During maturation, phagosomes containing engulfed particlesfuse with various endosomal compartments through theaction of regulatory molecules on the phagosomal membrane. Inthis study, we performed a proteomic analysis of the membranefraction from latex bead-containing (LBC) phagosomes isolatedfrom macrophages. The profile, which comprised 546 proteins,suggests diverse functions of the phagosome and potential connectionsto secretory processes, toll-like receptor signaling, andautophagy. Many identified proteins were not previously knownto reside in the phagosome. We characterized several proteins inLBC phagosomes that change in abundance on induction of autophagy,a process that has been previously implicated in the hostdefense against microbial pathogens. These observations suggestcrosstalk between autophagy and phagocytosis that may be relevantto the innate immune response of macrophages.

  18. STAT3-Mediated Autophagy Dependence Identifies Subtypes of Breast Cancer where Autophagy Inhibition can be Efficacious

    OpenAIRE

    Maycotte, Paola; Gearheart, Christy M.; Barnard, Rebecca; Aryal, Suraj; Mulcahy Levy, Jean M.; Fosmire, Susan P.; Hansen, Ryan J.; Morgan, Michael J.; Christopher C Porter; Gustafson, Daniel L.; Thorburn, Andrew

    2014-01-01

    Autophagy is a protein and organelle degradation pathway that is involved in diverse diseases including cancer. Recent evidence suggests that autophagy is a cell survival mechanism in tumor cells and that its inhibition especially in combination with other therapy could be beneficial but it remains unclear if all cancer cells behave the same way when autophagy is inhibited. We inhibited autophagy in a panel of breast cancer cell lines and found that some of them are dependent on autophagy for...

  19. Autophagy: for better or for worse

    OpenAIRE

    Wirawan, Ellen; Berghe, Tom Vanden; Lippens, Saskia; Agostinis, Patrizia; Vandenabeele, Peter

    2011-01-01

    Autophagy is a lysosomal degradation pathway that degrades damaged or superfluous cell components into basic biomolecules, which are then recycled back into the cytosol. In this respect, autophagy drives a flow of biomolecules in a continuous degradation-regeneration cycle. Autophagy is generally considered a pro-survival mechanism protecting cells under stress or poor nutrient conditions. Current research clearly shows that autophagy fulfills numerous functions in vital biological processes....

  20. Autophagy and mitophagy in cellular damage control

    Directory of Open Access Journals (Sweden)

    Jianhua Zhang

    2013-01-01

    Full Text Available Autophagy and mitophagy are important cellular processes that are responsible for breaking down cellular contents, preserving energy and safeguarding against accumulation of damaged and aggregated biomolecules. This graphic review gives a broad summary of autophagy and discusses examples where autophagy is important in controlling protein degradation. In addition we highlight how autophagy and mitophagy are involved in the cellular responses to reactive species and mitochondrial dysfunction. The key signaling pathways for mitophagy are described in the context of bioenergetic dysfunction.

  1. Possibility of reducing stray losses and parasitic torques in two-phase emergency feeding of induction motors

    Czech Academy of Sciences Publication Activity Database

    Schreier, Luděk; Bendl, Jiří; Chomát, Miroslav; Klíma, J.

    2006-01-01

    Roč. 51, č. 2 (2006), s. 109-131. ISSN 0001-7043 R&D Projects: GA ČR(CZ) GA102/04/0215 Institutional research plan: CEZ:AV0Z20570509 Keywords : induction motor * emergency operation of electrical drives * stray losses Subject RIV: JA - Electronics ; Optoelectronics, Electrical Engineering

  2. A comparison of still point induction to massage therapy in reducing pain and increasing comfort in chronic pain.

    Science.gov (United States)

    Townsend, Carolyn S; Bonham, Elizabeth; Chase, Linda; Dunscomb, Jennifer; McAlister, Susan

    2014-01-01

    A quantitative study was completed to determine whether complementary techniques provide pain relief and comfort in patients with chronic pain. Subjects participated in sessions including aromatherapy and music therapy. Massage or cranial still point induction was randomly assigned. Statistically significant improvement in pain and comfort was noted in both groups. PMID:24503744

  3. Notch1 regulated autophagy controls survival and suppressor activity of activated murine T-regulatory cells

    Science.gov (United States)

    Marcel, Nimi; Sarin, Apurva

    2016-01-01

    Cell survival is one of several processes regulated by the Notch pathway in mammalian cells. Here we report functional outcomes of non-nuclear Notch signaling to activate autophagy, a conserved cellular response to nutrient stress, regulating survival in murine natural T-regulatory cells (Tregs), an immune subset controlling tolerance and inflammation. Induction of autophagy required ligand-dependent, Notch intracellular domain (NIC) activity, which controlled mitochondrial organization and survival of activated Tregs. Consistently, NIC immune-precipitated Beclin and Atg14, constituents of the autophagy initiation complex. Further, ectopic expression of an effector of autophagy (Atg3) or recombinant NIC tagged to a nuclear export signal (NIC-NES), restored autophagy and suppressor function in Notch1-/- Tregs. Furthermore, Notch1 deficiency in the Treg lineage resulted in immune hyperactivity, implicating Notch activity in Treg homeostasis. Notch1 integration with autophagy, revealed in these experiments, holds implications for Notch regulated cell-fate decisions governing differentiation. DOI: http://dx.doi.org/10.7554/eLife.14023.001 PMID:27267497

  4. Autophagy in the Pathogenesis of Disease

    OpenAIRE

    Levine, Beth; Kroemer, Guido

    2008-01-01

    Autophagy is a lysosomal degradation pathway that is essential for survival, differentiation, development, and homeostasis. Autophagy principally serves an adaptive role to protect organisms against diverse pathologies, including infections, cancer, neurodegeneration, aging, and heart disease. However, in certain experimental disease settings, the self-cannibalistic or, paradoxically, even the prosurvival functions of autophagy may be deleterious. This Review summarizes recent advances in und...

  5. p53: The Janus of autophagy?

    OpenAIRE

    Levine, Beth; Abrams, John

    2008-01-01

    The autophagy pathway functions in adaptation to nutrient stress and tumour suppression. The p53 tumour suppressor, previously thought to positively regulate autophagy, may also inhibit it. This dual interplay between p53 and autophagy regulation is enigmatic, but may underlie key aspects of metabolism and cancer biology.

  6. Nicotinate-Curcumin Impedes Foam Cell Formation from THP-1 Cells through Restoring Autophagy Flux.

    Science.gov (United States)

    Gu, Hong-Feng; Li, Hai-Zhe; Tang, Ya-Ling; Tang, Xiao-Qing; Zheng, Xi-Long; Liao, Duan-Fang

    2016-01-01

    Our previous studies have indicated that a novel curcumin derivate nicotinate-curcumin (NC) has beneficial effects on the prevention of atherosclerosis, but the precise mechanisms are not fully understood. Given that autophagy regulates lipid metabolism, the present study was designed to investigate whether NC decreases foam cell formation through restoring autophagy flux in oxidized low-density lipoprotein (ox-LDL)-treated THP-1 cells. Our results showed that ox-LDL (100 μg/ml) was accumulated in THP-1 cells and impaired autophagy flux. Ox-LDL-induced impairment of autophagy was enhanced by treatment with the autophagy inhibitor chloroquine (CQ) and rescued by the autophagy inducer rapamycin. The aggregation of ox-LDL was increased by CQ, but decreased by rapamycin. In addition, colocalization of lipid droplets with LC3-II was remarkably reduced in ox-LDL group. In contrast, NC (10 μM) rescued the impaired autophagy flux by significantly increasing level of LC3-II, the number of autophagolysosomes, and the degradation of p62 in ox-LDL-treated THP-1 cells. Inhibition of the PI3K-Akt-mTOR signaling was required for NC-rescued autophagy flux. Notably, our results showed that NC remarkably promoted the colocalization of lipid droplets with autophagolysosomes, increased efflux of cholesterol, and reduced ox-LDL accumulation in THP-1 cells. However, treatment with 3-methyladenine (3-MA) or CQ reduced the protective effects of NC on lipid accumulation. Collectively, the findings suggest that NC decreases lipid accumulation in THP-1 cells through restoring autophagy flux, and further implicate that NC may be a potential therapeutic reagent to reverse atherosclerosis. PMID:27128486

  7. Nicotinate-Curcumin Impedes Foam Cell Formation from THP-1 Cells through Restoring Autophagy Flux

    Science.gov (United States)

    Gu, Hong-Feng; Li, Hai-Zhe; Tang, Ya-Ling; Tang, Xiao-Qing; Zheng, Xi-Long; Liao, Duan-Fang

    2016-01-01

    Our previous studies have indicated that a novel curcumin derivate nicotinate-curcumin (NC) has beneficial effects on the prevention of atherosclerosis, but the precise mechanisms are not fully understood. Given that autophagy regulates lipid metabolism, the present study was designed to investigate whether NC decreases foam cell formation through restoring autophagy flux in oxidized low-density lipoprotein (ox-LDL)-treated THP-1 cells. Our results showed that ox-LDL (100 μg/ml) was accumulated in THP-1 cells and impaired autophagy flux. Ox-LDL-induced impairment of autophagy was enhanced by treatment with the autophagy inhibitor chloroquine (CQ) and rescued by the autophagy inducer rapamycin. The aggregation of ox-LDL was increased by CQ, but decreased by rapamycin. In addition, colocalization of lipid droplets with LC3-II was remarkably reduced in ox-LDL group. In contrast, NC (10 μM) rescued the impaired autophagy flux by significantly increasing level of LC3-II, the number of autophagolysosomes, and the degradation of p62 in ox-LDL-treated THP-1 cells. Inhibition of the PI3K-Akt-mTOR signaling was required for NC-rescued autophagy flux. Notably, our results showed that NC remarkably promoted the colocalization of lipid droplets with autophagolysosomes, increased efflux of cholesterol, and reduced ox-LDL accumulation in THP-1 cells. However, treatment with 3-methyladenine (3-MA) or CQ reduced the protective effects of NC on lipid accumulation. Collectively, the findings suggest that NC decreases lipid accumulation in THP-1 cells through restoring autophagy flux, and further implicate that NC may be a potential therapeutic reagent to reverse atherosclerosis. PMID:27128486

  8. Autophagy pathway is required for IL-6 induced neuroendocrine differentiation and chemoresistance of prostate cancer LNCaP cells.

    Directory of Open Access Journals (Sweden)

    Pei-Ching Chang

    Full Text Available Prostate cancer (PCa cells undergoing neuroendocrine differentiation (NED are clinically relevant to the development of relapsed castration-resistant PCa. Increasing evidences show that autophagy involves in the development of neuroendocrine (NE tumors, including PCa. To clarify the effect of autophagy on NED, androgen-sensitive PCa LNCaP cells were examined. Treatment of LNCaP cells with IL-6 resulted in an induction of autophagy. In the absence of androgen, IL-6 caused an even stronger activation of autophagy. Similar result was identified in NED induction. Inhibition of autophagy with chloroquine (CQ markedly decreased NED. This observation was confirmed by beclin1 and Atg5 silencing experiments. Further supporting the role of autophagy in NED, we found that LC3 was up-regulated in PCa tissue that had relapsed after androgen-deprivation therapy when compared with their primary tumor counterpart. LC3 staining in relapsed PCa tissue showed punctate pattern similar to the staining of chromogranin A (CgA, a marker for NED cells. Moreover, autophagy inhibition induced the apoptosis of IL-6 induced NE differentiated PCa cells. Consistently, inhibition of autophagy by knockdown of beclin1 or Atg5 sensitized NE differentiated LNCaP cells to etoposide, a chemotherapy drug. To identify the mechanisms, phosphorylation of IL-6 downstream targets was analyzed. An increase in phospho-AMPK and a decrease in phospho-mTOR were found, which implies that IL-6 regulates autophagy through the AMPK/mTOR pathway. Most important to this study is the discovery of REST, a neuronal gene-specific transcriptional repressor that is involved in autophagy activation. REST was down-regulated in IL-6 treatment. Knockdown experiments suggest that REST is critical to NED and autophagy activation by IL-6. Together, our studies imply that autophagy is involved in PCa progression and plays a cytoprotective role when NED is induced in PCa cells by IL-6 treatment. These results

  9. Spermidine induces autophagy by inhibiting the acetyltransferase EP300.

    Science.gov (United States)

    Pietrocola, F; Lachkar, S; Enot, D P; Niso-Santano, M; Bravo-San Pedro, J M; Sica, V; Izzo, V; Maiuri, M C; Madeo, F; Mariño, G; Kroemer, G

    2015-03-01

    Several natural compounds found in health-related food items can inhibit acetyltransferases as they induce autophagy. Here we show that this applies to anacardic acid, curcumin, garcinol and spermidine, all of which reduce the acetylation level of cultured human cells as they induce signs of increased autophagic flux (such as the formation of green fluorescent protein-microtubule-associated protein 1A/1B-light chain 3 (GFP-LC3) puncta and the depletion of sequestosome-1, p62/SQSTM1) coupled to the inhibition of the mammalian target of rapamycin complex 1 (mTORC1). We performed a screen to identify the acetyltransferases whose depletion would activate autophagy and simultaneously inhibit mTORC1. The knockdown of only two acetyltransferases (among 43 candidates) had such effects: EP300 (E1A-binding protein p300), which is a lysine acetyltranferase, and NAA20 (N(α)-acetyltransferase 20, also known as NAT5), which catalyzes the N-terminal acetylation of methionine residues. Subsequent studies validated the capacity of a pharmacological EP300 inhibitor, C646, to induce autophagy in both normal and enucleated cells (cytoplasts), underscoring the capacity of EP300 to repress autophagy by cytoplasmic (non-nuclear) effects. Notably, anacardic acid, curcumin, garcinol and spermidine all inhibited the acetyltransferase activity of recombinant EP300 protein in vitro. Altogether, these results support the idea that EP300 acts as an endogenous repressor of autophagy and that potent autophagy inducers including spermidine de facto act as EP300 inhibitors. PMID:25526088

  10. Autophagy plays an essential role in cigarette smoke-induced expression of MUC5AC in airway epithelium.

    Science.gov (United States)

    Zhou, Jie-Sen; Zhao, Yun; Zhou, Hong-Bin; Wang, Yong; Wu, Yin-Fang; Li, Zhou-Yang; Xuan, Nan-Xia; Zhang, Chao; Hua, Wen; Ying, Song-Min; Li, Wen; Shen, Hua-Hao; Chen, Zhi-Hua

    2016-06-01

    Mucus hypersecretion is a common pathological feature of chronic airway inflammatory diseases including chronic obstructive pulmonary disease (COPD). However, the molecular basis for this condition remains incompletely understood. We have previously demonstrated a critical role of autophagy in COPD pathogenesis through mediating apoptosis of lung epithelial cells. In this study, we aimed to investigate the function of autophagy as well as its upstream and downstream signals in cigarette smoke-induced mucus production in human bronchial epithelial (HBE) cells and in mouse airways. Cigarette smoke extract (CSE), as well as the classical autophagy inducers starvation or Torin-1, significantly triggered MUC5AC expression, and inhibition of autophagy markedly attenuated CSE-induced mucus production. The CSE-induced autophagy was mediated by mitochondrial reactive oxygen species (mitoROS), which regulated mucin expression through the JNK and activator protein-1 pathway. Epidermal growth factor receptor (EGFR) was also required for CSE-induced MUC5AC in HBE cells, but it exerted inconsiderable effects on the autophagy-JNK signaling cascade. Airways of mice with dysfunctional autophagy-related genes displayed a markedly reduced number of goblet cells and attenuated levels of Muc5ac in response to cigarette smoke exposure. These results altogether suggest that mitoROS-dependent autophagy is essential for cigarette smoke-induced mucus hyperproduction in airway epithelial cells, and reemphasize autophagy inhibition as a novel therapeutic strategy for chronic airway diseases. PMID:27036871

  11. Egr-1 regulates autophagy in cigarette smoke-induced chronic obstructive pulmonary disease.

    Directory of Open Access Journals (Sweden)

    Zhi-Hua Chen

    Full Text Available BACKGROUND: Chronic obstructive pulmonary disease (COPD is a progressive lung disease characterized by abnormal cellular responses to cigarette smoke, resulting in tissue destruction and airflow limitation. Autophagy is a degradative process involving lysosomal turnover of cellular components, though its role in human diseases remains unclear. METHODOLOGY AND PRINCIPAL FINDINGS: Increased autophagy was observed in lung tissue from COPD patients, as indicated by electron microscopic analysis, as well as by increased activation of autophagic proteins (microtubule-associated protein-1 light chain-3B, LC3B, Atg4, Atg5/12, Atg7. Cigarette smoke extract (CSE is an established model for studying the effects of cigarette smoke exposure in vitro. In human pulmonary epithelial cells, exposure to CSE or histone deacetylase (HDAC inhibitor rapidly induced autophagy. CSE decreased HDAC activity, resulting in increased binding of early growth response-1 (Egr-1 and E2F factors to the autophagy gene LC3B promoter, and increased LC3B expression. Knockdown of E2F-4 or Egr-1 inhibited CSE-induced LC3B expression. Knockdown of Egr-1 also inhibited the expression of Atg4B, a critical factor for LC3B conversion. Inhibition of autophagy by LC3B-knockdown protected epithelial cells from CSE-induced apoptosis. Egr-1(-/- mice, which displayed basal airspace enlargement, resisted cigarette-smoke induced autophagy, apoptosis, and emphysema. CONCLUSIONS: We demonstrate a critical role for Egr-1 in promoting autophagy and apoptosis in response to cigarette smoke exposure in vitro and in vivo. The induction of autophagy at early stages of COPD progression suggests novel therapeutic targets for the treatment of cigarette smoke induced lung injury.

  12. Liraglutide prevents high glucose level induced insulinoma cells apoptosis by targeting autophagy

    Institute of Scientific and Technical Information of China (English)

    CHEN Ze-fang; LI Yan-bo; HAN Jun-yong; YIN Jia-jing; WANG Yang; ZHU Li-bo; XIE Guang-ying

    2013-01-01

    Background The pathophysiology of type 2 diabetes is progressive pancreatic beta cell failure with consequential reduced insulin secretion.Glucotoxicity results in the reduction of beta cell mass in type 2 diabetes by inducing apoptosis.Autophagy is essential for the maintenance of normal islet architecture and plays a crucial role in maintaining the intracellular insulin content by accelerating the insulin degradation rate in beta cells.Recently more attention has been paid to the effect of autophagy in type 2 diabetes.The regulatory pathway of autophagy in controlling pancreatic beta cells is still not clear.The aim of our study was to evaluate whether liraglutide can inhibit apoptosis and modulate autophagy in vitro in insulinoma cells (INS-1 cells).Methods INS-1 cells were incubated for 24 hours in the presence or absence of high levels of glucose,liraglutide (a long-acting human glucagon-like peptide-1 analogue),or 3-methyadenine (3-MA).Cell viability was measured using the Cell Counting Kit-8 (CCK8) viability assay.Autophagy of INS-1 cells was tested by monodansylcadaverine (MDC)staining,an autophagy fluorescent compound used for the labeling of autophagic vacuoles,and by Western blotting of microtubule-associated protein I light chain 3 (LC3),a biochemical markers of autophagic initiation.Results The viability of INS-1 cells was reduced after treatment with high levels of glucose.The viability of INS-1 cells was reduced and apoptosis was increased when autophagy was inhibited.The viability of INS-1 cells was significantly increased by adding liraglutide to supplement high glucose level medium compared with the cells treated with high glucose levels alone.Conclusions Apoptosis and autophagy were increased in rat INS-1 cells when treated with high level of glucose,and the viability of INS-1 cells was significantly reduced by inhibiting autophagy.Liraglutide protected INS-1 cells from high glucose level-induced apoptosis that is accompanied by a significant

  13. Oroxylin A induces autophagy in human malignant glioma cells via the mTOR-STAT3-Notch signaling pathway.

    Science.gov (United States)

    Zou, Meijuan; Hu, Chen; You, Qidong; Zhang, Aixia; Wang, Xuerong; Guo, Qinglong

    2015-11-01

    Autophagy is a tightly-regulated catabolic pathway involving degradation of cellular proteins, cytoplasm and organelles. Recent evidence suggests that autophagy plays a potential role in cell death as a tumor suppressor and that its induction especially in combination with apoptosis could be beneficial. It remains unclear if all cancer cells behave the same mechanism when autophagy is induced. Although mammalian target of rapamycin (mTOR) is well known as a negative regulator of autophagy, the relationship between signal transducer and activator of transcription 3 (STAT3) and autophagy has not yet been investigated. Oroxylin A, a natural mono-flavonoid extracted from Scutellariae radix, is a promising therapeutic agent for treating multiple cancers. Here we investigated the mechanism underlying the effect of oroxylin A on malignant glioma cells. We showed that oroxylin A inhibited the proliferation of malignant glioma cells by inducing autophagy in a dose- and time-dependent manner. Oroxylin A treatment inhibits the AKT and ERK activation and the downstream phosphorylation level of mTOR and STAT3. In addition, oroxylin A treatment decreases the expression of Notch-1 and myeloid cell leukemia-1 (Mcl-1) but upregulates Beclin 1, the key autophagy-related protein. 3-MA (autophagy inhibitor) or knockdown of Beclin 1 partially can rescue cells from oroxylin A-induced autophagic cell death. In contrast, knockdown of STAT3 aggravates oroxylin A-induced autophagic cell death. Our data reveal an important role of autophagy in enhancing cell death induced by oroxylin A and conclude that oroxylin A exerts anti-malignant glioma proficiency by inducing autophagy via the ERK/AKT-mTOR-STAT3-Notch signaling cascade. PMID:25213258

  14. Lysosomes and autophagy in aquatic animals.

    Science.gov (United States)

    Moore, Michael N; Kohler, Angela; Lowe, David; Viarengo, Aldo

    2008-01-01

    The lysosomal-autophagic system appears to be a common target for many environmental pollutants, as lysosomes accumulate many toxic metals and organic xenobiotics, which perturb normal function and damage the lysosomal membrane. In fact, autophagic reactions frequently involving reduced lysosomal membrane integrity or stability appear to be effective generic indicators of cellular well-being in eukaryotes: in social amoebae (slime mold), mollusks and fish, autophagy/membrane destabilization is correlated with many stress and toxicological responses and pathological reactions. Prognostic use of adverse lysosomal and autophagic reactions to environmental pollutants can be used for predicting cellular dysfunction and health in aquatic animals, such as shellfish and fish, which are extensively used as sensitive bioindicators in monitoring ecosystem health; and also represent a significant food resource for at least 20% of the global human population. Explanatory frameworks for prediction of pollutant impact on health have been derived encompassing a conceptual mechanistic model linking lysosomal damage and autophagic dysfunction with injury to cells and tissues. Methods are described for tracking in vivo autophagy of fluorescently labeled cytoplasmic proteins, measuring degradation of radiolabeled intracellular proteins and morphometric measurement of lysosomal/cytoplasmic volume ratio. Additional methods for the determination of lysosomal membrane stability in lower animals are also described, which can be applied to frozen tissue sections, protozoans and isolated cells in vivo. Experimental and simulated results have also indicated that nutritional deprivation (analogous in marine mussels to caloric restriction)-induced autophagy has a protective function against toxic effects mediated by reactive oxygen species (ROS). Finally, coupled measurement of lysosomal-autophagic reactions and simulation modelling is proposed as a practical toolbox for predicting toxic

  15. The cellular decision between apoptosis and autophagy

    Directory of Open Access Journals (Sweden)

    Yong-Jun Fan

    2013-03-01

    Full Text Available Apoptosis and autophagy are important molecular processes that maintain organismal and cellular homeostasis, respectively. While apoptosis fulfills its role through dismantling damaged or unwanted cells, autophagy maintains cellular homeostasis through recycling selective intracellular organelles and molecules. Yet in some conditions, autophagy can lead to cell death. Apoptosis and autophagy can be stimulated by the same stresses. Emerging evidence indicates an interplay between the core proteins in both pathways, which underlies the molecular mechanism of the crosstalk between apoptosis and autophagy. This review summarizes recent literature on molecules that regulate both the apoptotic and autophagic processes.

  16. Advances in Autophagy Regulatory Mechanisms

    Directory of Open Access Journals (Sweden)

    Laura E. Gallagher

    2016-05-01

    Full Text Available Autophagy plays a critical role in cell metabolism by degrading and recycling internal components when challenged with limited nutrients. This fundamental and conserved mechanism is based on a membrane trafficking pathway in which nascent autophagosomes engulf cytoplasmic cargo to form vesicles that transport their content to the lysosome for degradation. Based on this simple scheme, autophagy modulates cellular metabolism and cytoplasmic quality control to influence an unexpectedly wide range of normal mammalian physiology and pathophysiology. In this review, we summarise recent advancements in three broad areas of autophagy regulation. We discuss current models on how autophagosomes are initiated from endogenous membranes. We detail how the uncoordinated 51-like kinase (ULK complex becomes activated downstream of mechanistic target of rapamycin complex 1 (MTORC1. Finally, we summarise the upstream signalling mechanisms that can sense amino acid availability leading to activation of MTORC1.

  17. Advances in Autophagy Regulatory Mechanisms.

    Science.gov (United States)

    Gallagher, Laura E; Williamson, Leon E; Chan, Edmond Y W

    2016-01-01

    Autophagy plays a critical role in cell metabolism by degrading and recycling internal components when challenged with limited nutrients. This fundamental and conserved mechanism is based on a membrane trafficking pathway in which nascent autophagosomes engulf cytoplasmic cargo to form vesicles that transport their content to the lysosome for degradation. Based on this simple scheme, autophagy modulates cellular metabolism and cytoplasmic quality control to influence an unexpectedly wide range of normal mammalian physiology and pathophysiology. In this review, we summarise recent advancements in three broad areas of autophagy regulation. We discuss current models on how autophagosomes are initiated from endogenous membranes. We detail how the uncoordinated 51-like kinase (ULK) complex becomes activated downstream of mechanistic target of rapamycin complex 1 (MTORC1). Finally, we summarise the upstream signalling mechanisms that can sense amino acid availability leading to activation of MTORC1. PMID:27187479

  18. Propofol Mitigates Learning and Memory Impairment After Electroconvulsive Shock in Depressed Rats by Inhibiting Autophagy in the Hippocampus.

    Science.gov (United States)

    Li, Ping; Hao, Xue-Chao; Luo, Jie; Lv, Feng; Wei, Ke; Min, Su

    2016-01-01

    BACKGROUND The present study explored the effects of propofol on hippocampal autophagy and synaptophysin in depression-model rats undergoing electroconvulsive shock (ECS). MATERIAL AND METHODS The rat depression model was established by exposing Sprague-Dawley rats to stress for 28 consecutive days. Forty rats were assigned randomly into the depression group (group D; no treatment), the ECS group (group E), the propofol group (group P), and the propofol + ECS group (group PE). Open field tests and sucrose preference tests were applied to evaluate the depression behavior; and Morris water maze tests were used to assess the learning and memory function of the rats. Western blotting was used to detect the expression of Beclin-1 and LC3-II/I; and ELISA was applied to assess the expression of synaptophysin. RESULTS Rats in group E and group PE scored higher in the open field and sucrose preference tests compared with those in group D. Furthermore, rats in group E also had a longer escape latency, a shorter space exploration time, and increased expression of Beclin-1, LC3-II/I, and synaptophysin. Compared with group E, rats in group PE possessed a shorter escape latency, a longer space exploration time, reduced expression of Beclin-1, LC3-II/I, and synaptophysin. CONCLUSIONS Propofol could inhibit excessive ECS-induced autophagy and synaptophysin overexpression in the hippocampus, thus protecting the learning and memory functions in depressed rats after ECS. The inhibitory effects of propofol on the overexpression of synaptophysin may result from its inhibitory effects on the excessive induction of autophagy. PMID:27203836

  19. Impaired Autophagy in Adult Bone Marrow CD34+ Cells of Patients with Aplastic Anemia: Possible Pathogenic Significance

    Science.gov (United States)

    Huang, Jinbo; Ge, Meili; Lu, Shihong; Shi, Jun; Yu, Wei; Li, Xingxin; Wang, Min; Zhang, Jizhou; Feng, Sizhou; Dong, Shuxu; Cheng, Xuelian; Zheng, Yizhou

    2016-01-01

    Aplastic anemia (AA) is a bone marrow failure syndrome that is caused largely by profound quantitative and qualitative defects of hematopoietic stem and progenitor cells. However, the mechanisms underlying these defects remain unclear. Under conditions of stress, autophagy acts as a protective mechanism for cells. We therefore postulated that autophagy in CD34+ hematopoietic progenitor cells (HPCs) from AA patients might be impaired and play a role in the pathogenesis of AA. To test this hypothesis, we tested autophagy in CD34+ cells from AA samples and healthy controls and investigated the effect of autophagy on the survival of adult human bone marrow CD34+ cells. We found that the level of autophagy in CD34+ cells from AA patients was significantly lower than in age/sex-matched healthy controls, and lower in cases of severe AA than in those with non-severe AA. Autophagy in CD34+ cells improved upon amelioration of AA but, compared to healthy controls, was still significantly reduced even in AA patients who had achieved a complete, long-term response. We also showed that although the basal autophagy in CD34+ cells was low, the autophagic response of CD34+ cells to “adversity” was rapid. Finally, impaired autophagy resulted in reduced differentiation and proliferation of CD34+ cells and sensitized them to death and apoptosis. Thus, our results confirm that autophagy in CD34+ cells from AA patients is impaired, that autophagy is required for the survival of CD34+ cells, and that impaired autophagy in CD34+ HPCs may play an important role in the pathogenesis of AA. PMID:26930650

  20. Modulation of pathogen recognition by autophagy

    Directory of Open Access Journals (Sweden)

    Ji Eun eOh

    2012-03-01

    Full Text Available Autophagy is an ancient biological process for maintaining cellular homeostasis by degradation of long-lived cytosolic proteins and organelles. Recent studies demonstrated that autophagy is availed by immune cells to regulate innate immunity. On the one hand, cells exert direct effector function by degrading intracellular pathogens; on the other hand, autophagy modulates pathogen recognition and downstream signaling for innate immune responses. Pathogen recognition via pattern recognition receptors induces autophagy. The function of phagocytic cells is enhanced by recruitment of autophagy-related proteins. Moreover, autophagy acts as a delivery system for viral replication complexes to migrate to the endosomal compartments where virus sensing occurs. In another case, key molecules of the autophagic pathway have been found to negatively regulate immune signaling, thus preventing aberrant activation of cytokine production and consequent immune responses. In this review, we focus on the recent advances in the role of autophagy in pathogen recognition and modulation of innate immune responses.

  1. Historical landmarks of autophagy research

    OpenAIRE

    Ohsumi, Yoshinori

    2013-01-01

    The year of 2013 marked the 50th anniversary of C de Duve's coining of the term “autophagy” for the degradation process of cytoplasmic constituents in the lysosome/vacuole. This year we regretfully lost this great scientist, who contributed much during the early years of research to the field of autophagy. Soon after the discovery of lysosomes by de Duve, electron microscopy revealed autophagy as a means of delivering intracellular components to the lysosome. For a long time after the discove...

  2. Introduction of a methoxymethyl side chain into p-phenylenediamine attenuates its sensitizing potency and reduces the risk of allergy induction

    Energy Technology Data Exchange (ETDEWEB)

    Goebel, Carsten, E-mail: goebel.c.1@pg.com [The Procter and Gamble Co., Central Product Safety and Communications, Darmstadt (Germany); Troutman, John [The Procter and Gamble Co., Central Product Safety, Cincinnati, OH (United States); Hennen, Jenny [Dept. of Environmental Toxicology, Trier University, Trier (Germany); Rothe, Helga; Schlatter, Harald [The Procter and Gamble Co., Central Product Safety and Communications, Darmstadt (Germany); Gerberick, G. Frank [The Procter and Gamble Co., Central Product Safety, Cincinnati, OH (United States); Blömeke, Brunhilde [Dept. of Environmental Toxicology, Trier University, Trier (Germany)

    2014-02-01

    The strong sensitizing potencies of the most important primary intermediates of oxidative hair dyes, p-phenylenediamine (PPD) and p-toluylenediamine (PTD, i.e. 2-methyl-PPD) are well established. They are considered as the key sensitizers in hair dye allergic contact dermatitis. While modification of their molecular structure is expected to alter their sensitizing properties, it may also impair their color performance. With introduction of a methoxymethyl side chain we found the primary intermediate 2-methoxymethyl-p-phenylenediamine (ME-PPD) with excellent hair coloring performance but significantly reduced sensitizing properties compared to PPD and PTD: In vitro, ME-PPD showed an attenuated innate immune response when analyzed for its protein reactivity and dendritic cell activation potential. In vivo, the effective concentration of ME-PPD necessary to induce an immune response 3-fold above vehicle control (EC3 value) in the local lymph node assay (LLNA) was 4.3%, indicating a moderate skin sensitizing potency compared to values of 0.1 and 0.17% for PPD and PTD, respectively. Finally, assessing the skin sensitizing potency of ME-PPD under consumer hair dye usage conditions through a quantitative risk assessment (QRA) indicated an allergy induction risk negligible compared to PPD or PTD. - Highlights: • Methoxymethyl side chain in p-phenylenediamine reduces its strong skin sensitizing properties. • Reduced protein reactivity and dendritic cell activation. • Reduced skin sensitizing potency in local lymph node assay (LLNA). • Negligible allergy induction risk under hair dye usage conditions.

  3. Introduction of a methoxymethyl side chain into p-phenylenediamine attenuates its sensitizing potency and reduces the risk of allergy induction

    International Nuclear Information System (INIS)

    The strong sensitizing potencies of the most important primary intermediates of oxidative hair dyes, p-phenylenediamine (PPD) and p-toluylenediamine (PTD, i.e. 2-methyl-PPD) are well established. They are considered as the key sensitizers in hair dye allergic contact dermatitis. While modification of their molecular structure is expected to alter their sensitizing properties, it may also impair their color performance. With introduction of a methoxymethyl side chain we found the primary intermediate 2-methoxymethyl-p-phenylenediamine (ME-PPD) with excellent hair coloring performance but significantly reduced sensitizing properties compared to PPD and PTD: In vitro, ME-PPD showed an attenuated innate immune response when analyzed for its protein reactivity and dendritic cell activation potential. In vivo, the effective concentration of ME-PPD necessary to induce an immune response 3-fold above vehicle control (EC3 value) in the local lymph node assay (LLNA) was 4.3%, indicating a moderate skin sensitizing potency compared to values of 0.1 and 0.17% for PPD and PTD, respectively. Finally, assessing the skin sensitizing potency of ME-PPD under consumer hair dye usage conditions through a quantitative risk assessment (QRA) indicated an allergy induction risk negligible compared to PPD or PTD. - Highlights: • Methoxymethyl side chain in p-phenylenediamine reduces its strong skin sensitizing properties. • Reduced protein reactivity and dendritic cell activation. • Reduced skin sensitizing potency in local lymph node assay (LLNA). • Negligible allergy induction risk under hair dye usage conditions

  4. Wogonoside induces autophagy-related apoptosis in human glioblastoma cells.

    Science.gov (United States)

    Zhang, Li; Wang, Handong; Cong, Zixiang; Xu, Jianguo; Zhu, Jianhong; Ji, Xiangjun; Ding, Ke

    2014-09-01

    Wogonoside, a bioactive flavonoid extracted from the root of Scutellaria baicalensis Georgi, has shown preclinical anticancer efficacy in various cancer models. However, the effects of wogonoside on glioblastoma cells remain unclear. In the present study, we found that wogonoside exhibited a cytotoxic effect on human glioblastoma cells. The suppression of cell viability was due to the induction of mitochondrial apoptosis. Furthermore, the presence of autophagic hallmarks, including an increase in punctate microtubule associated protein 1 light chain 3 (LC3) dots, changes in cellular morphology and increased levels of autophagy-related proteins were observed in the wogonoside-treated cells. Wogonoside treatment also enhanced autophagic flux as reflected by the increased acidic vesicular organelle (AVO) formation, p62 degradation and LC3 turnover. Notably, blockade of autophagy by a chemical inhibitor or RNA interference decreased the anticancer effect of wogonoside. In addition, the p38 mitogen-activated protein kinase (MAPK) signaling pathway, the phosphatidylinositide 3-kinase/protein kinase B/mammalian target of rapamycin/p70S6 kinase (PI3K/AKT/mTOR/p70S6K) signaling pathway and reactive oxygen species (ROS) participated in wogonoside-induced autophagy and apoptosis. These findings support the initiation of further studies of wogonoside as a candidate for the treatment of human malignant glioma. PMID:24970553

  5. Suppression of calcium‑sensing receptor ameliorates cardiac hypertrophy through inhibition of autophagy.

    Science.gov (United States)

    Liu, Lei; Wang, Chao; Lin, Yan; Xi, Yuhui; Li, Hong; Shi, Sa; Li, Hongzhu; Zhang, Weihua; Zhao, Yajun; Tian, Ye; Xu, Changqing; Wang, Lina

    2016-07-01

    The calcium-sensing receptor (CaSR) releases intracellular calcium ([Ca2+]i) by accumulating inositol phosphate. Changes in [Ca2+]i initiate myocardial hypertrophy. Furthermore, autophagy associated with [Ca2+]i. Autophagy has previously been demonstrated to participate in the hypertrophic process. The current study investigated whether suppression of CaSR affects the hypertrophic response via modulating autophagy. Isoproterenol (ISO) was used to induce cardiac hypertrophy in Wistar rats. Hypertrophic status was determined by echocardiographic assessment, hematoxylin and eosin, and Masson's staining. The protein expression levels of CaSR and autophagy level were observed. Changes of hypertrophy and autophagy indicators were observed following intravenous injection of a CaSR inhibitor. An ISO‑induced cardiomyocyte hypertrophy model was established and used determine the involvement of GdCl3. [Ca2+]i was determined using Fluo‑4/AM dye followed by confocal microscopy. The expression levels of various active proteins were analyzed by western blotting. The size of the heart, expression levels of CaSR and autophagy level were markedly increased in hypertrophic myocardium. In addition, the present study demonstrated that the indicators of hypertrophy and autophagy were effectively suppressed by CaSR inhibitor. Furthermore, similar effects were demonstrated in neonatal rat hypertrophic cardiomyocytes treated with ISO. It was also observed that CaSR regulates the Ca2+/calmodulin‑dependent protein kinase kinase β (CaMKKβ)‑AMP‑activated protein kinase (AMPK)‑mammalian target of rapamycin (mTOR) signaling pathway induced by ISO in cardiomyocytes. Furthermore, the AMPK inhibition significantly reduced the autophagy level following CaSR stimulation (P<0.05). The results of the present demonstrated that inhibition of CaSR may ameliorate cardiac hypertrophy induced by ISO and the effect may be associated with the inhibition of autophagy and suppression of the Ca

  6. Combination treatment with fasudil and clioquinol produces synergistic anti-tumor effects in U87 glioblastoma cells by activating apoptosis and autophagy.

    Science.gov (United States)

    He, Mingliang; Luo, Ming; Liu, Qingyu; Chen, Jingkao; Li, Kaishu; Zheng, Meiguang; Weng, Yinlun; Ouyang, Leping; Liu, Anmin

    2016-04-01

    Survival of patients with glioblastoma (GBM) remains poor, and novel treatment methods are urgently needed. In this study, we tested the effects of a combination of fasudil, a ROCK inhibitor, and clioquinol, an 8-hydroxyquinoline derivative with antimicrobial properties, on human GBM U87 cells. Combination treatment synergistically inhibited the viability of glioma cells but not mouse normal neuron HT22 cells and significantly induced mitochondria-mediated apoptosis. Moreover, the combination was also found to trigger macro-autophagy (henceforth referred to as autophagy) by increasing the expression levels of several proteins involved in the induction of autophagy. Further studies showed that 3-methyladenine (3-MA) or chloroquine (CQ), two autophagy inhibitors, abrogated the cytotoxic effects of the combination treatment as well as the autophagy. Overall, we demonstrated that fasudil and clioquinol show synergistic anti-cancer effects, providing evidence for the further development of combination therapy for GBM. PMID:26725099

  7. Oxyphenisatin acetate (NSC 59687) triggers a cell starvation response leading to autophagy, mitochondrial dysfunction, and autocrine TNFα-mediated apoptosis

    International Nuclear Information System (INIS)

    Oxyphenisatin (3,3-bis(4-hydroxyphenyl)-1H-indol-2-one) and several structurally related molecules have been shown to have in vitro and in vivo antiproliferative activity. This study aims to confirm and extend mechanistic studies by focusing on oxyphenisatin acetate (OXY, NSC 59687), the pro-drug of oxyphenisatin. Results confirm that OXY inhibits the growth of the breast cancer cell lines MCF7, T47D, HS578T, and MDA-MB-468. This effect is associated with selective inhibition of translation accompanied by rapid phosphorylation of the nutrient sensing eukaryotic translation initiation factor 2α (eIF2α) kinases, GCN2 and PERK. This effect was paralleled by activation of AMP-activated protein kinase (AMPK) combined with reduced phosphorylation of the mammalian target of rapamycin (mTOR) substrates p70S6K and 4E-BP1. Microarray analysis highlighted activation of pathways involved in apoptosis induction, autophagy, RNA/protein metabolism, starvation responses, and solute transport. Pathway inhibitor combination studies suggested a role for AMPK/mTOR signaling, de novo transcription and translation, reactive oxygen species (ROS)/glutathione metabolism, calcium homeostasis and plasma membrane Na+/K+/Ca2+ transport in activity. Further examination confirmed that OXY treatment was associated with autophagy, mitochondrial dysfunction, and ROS generation. Additionally, treatment was associated with activation of both intrinsic and extrinsic apoptotic pathways. In the estrogen receptor (ER) positive MCF7 and T47D cells, OXY induced TNFα expression and TNFR1 degradation, indicating autocrine receptor-mediated apoptosis in these lines. Lastly, in an MCF7 xenograft model, OXY delivered intraperitoneally inhibited tumor growth, accompanied by phosphorylation of eIF2α and degradation of TNFR1. These data suggest that OXY induces a multifaceted cell starvation response, which ultimately induces programmed cell death. The mechanistic basis for oxyphenisatin acetate anti

  8. Glycans coated silver nanoparticles induces autophagy and necrosis in HeLa cells

    Science.gov (United States)

    Panzarini, Elisa; Mariano, Stefania; Dini, Luciana

    2015-06-01

    This study reports the induction of autophagy by two concentrations (2×103 or 2×104 NPs/cell) of 30 nm sized β-D-Glucose- and β-D-Glucose/Sucrose-coated silver NanoParticles (AgNPs-G and AgNPs-GS respectively) in HeLa cells treated for 6, 12, 24 and 48 hrs. Cell viability was assessed by Neutral Red (NR) test and morphological evaluation. In addition ROS generation (NBT test) and induction of apoptosis/necrosis (Annexin V/Propidium Iodide-Annexin V/PI staining) and autophagy (Monodansylcadaverine-MDC staining) were evaluated. Cytotoxicity, ROS generation and morphology changes depend on NPs type and amount, and incubation time. As a general result, AgNPs-G are more toxic than AgNPs-GS. Moreover, the lowest AgNPs-GS concentration is ineffective on cell viability and ROS generation. Only 10% and 25% of viable HeLa cells were found at the end of incubation time in the presence of higher amount of AgNPs - G and AgNPs-GS respectively and in parallel ROS generation is induced. To elucidate the type of cell death, Annexin V/PI and MDC staining was performed. Interestingly, irrespective of coating type and NPs amount the percentage of apoptotic cells (Annexin V+/PI-) is similar to viable HeLa cells. At contrary, we observed a NPs amount dependent autophagy and necrosis induction. In fact, the lower amount of NPs induces autophagy (MDC+/PI- cells) whereas the higher one induces necrosis (Annexin V+/PI+ cells). Our findings suggest that AgNPs-induced cytotoxicity depends on AgNPs amount and type and provide preliminary evidence of induction of autophagy in HeLa cells cultured in the presence of AgNPs.

  9. Ordered bulk degradation via autophagy

    DEFF Research Database (Denmark)

    Dengjel, Jörn; Kristensen, Anders Riis; Andersen, Jens S

    2008-01-01

    proteasomal and lysosomal degradation ample cross-talk between the two degradation pathways became evident. Degradation via autophagy appeared to be ordered and regulated at the protein complex/organelle level. This raises several important questions such as: can macroautophagy itself be specific and what is...

  10. Reduced Cost of Reactive Power in Doubly Fed Induction Generator Wind Turbine System with Optimized Grid Filter

    DEFF Research Database (Denmark)

    Zhou, Dao; Blaabjerg, Frede; Franke, Toke; Tonnes, Michael; Lau, Mogens

    The modern grid requirement has caused that the wind power system behaves more like conventional rotating generators and it is able to support certain amount of the reactive power. For a typical doubly-fed induction generator wind turbine system, the reactive power can be supported either through...... for the generator and the wind power converter in terms of the reactive power done by the rotor-side converter or the grid-side converter with various grid filters. Afterwards, the annual energy loss is also estimated based on yearly wind profile. Finally, experimental results of the loss distribution...... the rotor-side converter or the grid-side converter. This paper firstly compares the current ripples and supportive reactive power ranges between the conventional L and optimized LCL filter, if the reactive power is injected from the grid-side converter. Then, the loss distribution is evaluated both...

  11. Reduced Cost of Reactive Power in Doubly Fed Induction Generator Wind Turbine System With Optimized Grid Filter

    DEFF Research Database (Denmark)

    Zhou, Dao; Blaabjerg, Frede; Franke, Toke; Tonnes, Michael; Lau, Mogens

    2015-01-01

    The modern grid requirement has caused that the wind power system behaves more like conventional rotating generators, and it is able to support certain amount of the reactive power. For a typical doubly fed induction generator (DFIG) wind turbine system, the reactive power can be supported either...... for the generator and the wind power converter in terms of the reactive power done by the rotor-side converter or the grid-side converter with various grid filters. Afterward, the annual energy loss is also estimated based on yearly wind profile. Finally, experimental results of the loss distribution...... through the rotor-side converter or the grid-side converter. This paper first compares the current ripples and supportive reactive power ranges between the conventional L and optimized LCL filter, if the reactive power is injected from the grid-side converter. Then, the loss distribution is evaluated both...

  12. Repetitive stimulation of autophagy-lysosome machinery by intermittent fasting preconditions the myocardium to ischemia-reperfusion injury.

    Science.gov (United States)

    Godar, Rebecca J; Ma, Xiucui; Liu, Haiyan; Murphy, John T; Weinheimer, Carla J; Kovacs, Attila; Crosby, Seth D; Saftig, Paul; Diwan, Abhinav

    2015-01-01

    Autophagy, a lysosomal degradative pathway, is potently stimulated in the myocardium by fasting and is essential for maintaining cardiac function during prolonged starvation. We tested the hypothesis that intermittent fasting protects against myocardial ischemia-reperfusion injury via transcriptional stimulation of the autophagy-lysosome machinery. Adult C57BL/6 mice subjected to 24-h periods of fasting, every other day, for 6 wk were protected from in-vivo ischemia-reperfusion injury on a fed day, with marked reduction in infarct size in both sexes as compared with nonfasted controls. This protection was lost in mice heterozygous null for Lamp2 (coding for lysosomal-associated membrane protein 2), which demonstrate impaired autophagy in response to fasting with accumulation of autophagosomes and SQSTM1, an autophagy substrate, in the heart. In lamp2 null mice, intermittent fasting provoked progressive left ventricular dilation, systolic dysfunction and hypertrophy; worsening cardiomyocyte autophagosome accumulation and lack of protection to ischemia-reperfusion injury, suggesting that intact autophagy-lysosome machinery is essential for myocardial homeostasis during intermittent fasting and consequent ischemic cardioprotection. Fasting and refeeding cycles resulted in transcriptional induction followed by downregulation of autophagy-lysosome genes in the myocardium. This was coupled with fasting-induced nuclear translocation of TFEB (transcription factor EB), a master regulator of autophagy-lysosome machinery; followed by rapid decline in nuclear TFEB levels with refeeding. Endogenous TFEB was essential for attenuation of hypoxia-reoxygenation-induced cell death by repetitive starvation, in neonatal rat cardiomyocytes, in-vitro. Taken together, these data suggest that TFEB-mediated transcriptional priming of the autophagy-lysosome machinery mediates the beneficial effects of fasting-induced autophagy in myocardial ischemia-reperfusion injury. PMID:26103523

  13. Cadmium induces autophagy through ROS-dependent activation of the LKB1-AMPK signaling in skin epidermal cells

    International Nuclear Information System (INIS)

    Cadmium is a toxic heavy metal which is environmentally and occupationally relevant. The mechanisms underlying cadmium-induced autophagy are not yet completely understood. The present study shows that cadmium induces autophagy, as demonstrated by the increase of LC3-II formation and the GFP-LC3 puncta cells. The induction of autophagosomes was directly visualized by electron microscopy in cadmium-exposed skin epidermal cells. Blockage of LKB1 or AMPK by siRNA transfection suppressed cadmium-induced autophagy. Cadmium-induced autophagy was inhibited in dominant-negative AMPK-transfected cells, whereas it was accelerated in cells transfected with the constitutively active form of AMPK. mTOR signaling, a negative regulator of autophagy, was downregulated in cadmium-exposed cells. In addition, cadmium generated reactive oxygen species (ROS) at relatively low levels, and caused poly(ADP-ribose) polymerase-1 (PARP) activation and ATP depletion. Inhibition of PARP by pharmacological inhibitors or its siRNA transfection suppressed ATP reduction and autophagy in cadmium-exposed cells. Furthermore, cadmium-induced autophagy signaling was attenuated by either exogenous addition of catalase and superoxide dismutase, or by overexpression of these enzymes. Consequently, these results suggest that cadmium-mediated ROS generation causes PARP activation and energy depletion, and eventually induces autophagy through the activation of LKB1-AMPK signaling and the down-regulation of mTOR in skin epidermal cells. - Highlights: → Cadmium, a toxic heavy metal, induces autophagic cell death through ROS-dependent activation of the LKB1-AMPK signaling. → Cadmium generates intracellular ROS at low levels and this leads to severe DNA damage and PARP activation, resulting in ATP depletion, which are the upstream events of LKB1-AMPK-mediated autophagy. → This novel finding may contribute to further understanding of cadmium-mediated diseases.

  14. Autophagy in the thymic epithelium is dispensable for the development of self-tolerance in a novel mouse model.

    Directory of Open Access Journals (Sweden)

    Supawadee Sukseree

    Full Text Available The thymic epithelium plays critical roles in the positive and negative selection of T cells. Recently, it was proposed that autophagy in thymic epithelial cells is essential for the induction of T cell tolerance to self antigens and thus for the prevention of autoimmune diseases. Here we have tested this hypothesis using mouse models in which autophagy was blocked specifically in epithelial cells expressing keratin 14 (K14, including the precursor of thymic epithelial cells. While the thymic epithelial cells of mice carrying the floxed Atg7 gene (ATG7 f/f showed a high level of autophagy, as determined by LC3 Western blot analysis and fluorescence detection of the recombinant green fluorescent protein (GFP-LC3 reporter protein on autophagosomes, autophagy in the thymic epithelium was efficiently suppressed by deletion of the Atg7 gene using the Cre-loxP system (ATG7 f/f K14-Cre. Suppression of autophagy led to the massive accumulation of p62/sequestosome 1 (SQSTM1 in thymic epithelial cells. However, the structure of the thymic epithelium as well as the organization and the size of the thymus were not altered in mutant mice. The ratio of CD4 to CD8-positive T cells, as well as the frequency of activated (CD69+ CD4 T cells in lymphoid organs, did not differ between mice with autophagy-competent and autophagy-deficient thymic epithelium. Inflammatory infiltrating cells, potentially indicative of autoimmune reactions, were present in the liver, lung, and colon of a similar fraction of ATG7 f/f and ATG7 f/f K14-Cre mice. In contrast to previously reported mice, that had received an autophagy-deficient thymus transplant, ATG7 f/f K14-Cre mice did not suffer from autoimmunity-induced weight loss. In summary, the results of this study suggest that autophagy in the thymic epithelium is dispensable for negative selection of autoreactive T cells.

  15. JNK-dependent Atg4 upregulation mediates asperphenamate derivative BBP-induced autophagy in MCF-7 cells

    International Nuclear Information System (INIS)

    N-Benzoyl-O-(N′-(1-benzyloxycarbonyl-4-piperidiylcarbonyl) -D-phenylalanyl)-D-phenylalaninol (BBP), a novel synthesized asperphenamate derivative with the increased solubility, showed growth inhibitory effect on human breast carcinoma MCF-7 cells in a time- and concentration-dependent manner. The growth inhibitory effect of BBP was associated with induction of autophagy, which was demonstrated by the development of acidic vesicular organelles, cleavage of LC3 and upregulation of Atg4 in BBP-treated MCF-7 cells. Since the application of Atg4 siRNA totally blocked the cleavage of LC3, we demonstrated a central role of Atg4 in BBP-induced autophagy. The further studies showed that BBP increased the levels of reactive oxygen species (ROS), and pretreatment with NAC effectively blocked the accumulation of ROS, autophagy and growth inhibition triggered by BBP. Moreover, BBP induced the activation of JNK, and JNK inhibitor SP600125 reversed autophagy, the increase of Atg4 levels, conversion of LC3 and growth inhibition induced by BBP. Knockdown of JNK by siRNA efficiently inhibited ROS production and autophagy, but antioxidant NAC failed to block JNK activation induced by BBP, indicating that JNK activation may be a upstream signaling of ROS and should be a core component in BBP-induced autophagic signaling pathway. These results suggest that BBP produces its growth inhibitory effect through induction of the autophagic cell death in MCF-7 cells, which is modulated by a JNK-dependent Atg4 upregulation involving ROS production. -- Highlights: ► Asperphenamate derivative BBP with increased solubility was synthesized. ► BBP selectively inhibited the growth of human breast tumor cells. ► The growth inhibitory effect of BBP was associated with induction of autophagy. ► JNK-dependent Atg4 upregulation mediated BBP-induced autophagy.

  16. Contribution of autophagy inhibitor to radiation sensitization in nasopharyngeal carcinoma cells

    International Nuclear Information System (INIS)

    Objective: To investigate the role of autophagy in radiation-induced death response of human nasopharyngeal carcinoma cells. Methods: MTT method was used to detect cell viability of CNE-2 cells in different time after irradiation. Clonogenic survival assay was used to evaluate the effect of autophagy inhibitor (chloroquine phosphate) and autophagy inductor (rapamycin) on radiosensitivity of nasopharyngeal carcinoma cells.Cell apoptosis was assessed by flow cytometry. The expressions of LC3 and P62 were measured with Western blot. Cell ultrastructural analysis was performed under an electron microscope.Results Irradiation with 10 Gy induced a massive accumulation of autophagosomes accompanied with up-regulation of LC3-Ⅱ expression in CNE-2 cells. Compared with radiation alone, chloroquine phosphate (CDP) enhanced radiosensitivity significantly by decreasing cell viability (F=25.88, P<0.05), autophagic ratio (F=105.15, P<0.05), and LC3-Ⅱ protein level (F=231.68, P<0.05), while up-regulating the expression of P62 (F=117.52, P<0.05). Inhibition of autophagy increased radiation-induced apoptosis (F=143.72, P<0.05). Rapamycin (RAPA) also significantly decreased cell viability, but increased autophagic ratio and LC3-Ⅱ protein level while down-regulated the expression of P62. Induction of autophagy increased radiation-induced apoptosis (F=167.32, P<0.05). Conclusions: Blockage of autophagy with CDP could enhance radiosensitivity in human nasopharyngeal carcinoma cells, suggesting that inhibition of autophagy could be used as an adjuvant treatment to nasopharyngeal carcinoma. (authors)

  17. Autophagy regulates chlorpyrifos-induced apoptosis in SH-SY5Y cells

    International Nuclear Information System (INIS)

    Recent studies have shown that up-regulation of autophagy may be a tractable therapeutic intervention for clearing disease-causing proteins, including α-synuclein, ubiquitin, and other misfolded or aggregated proteins in pesticide-induced neurodegeneration. In a previous study, we reported that chlorpyrifos (CPF)-induced mitochondria-dependent apoptosis is mediated through reactive oxygen species in SH-SY5Y cells. In this study, we explored a novel pharmacotherapeutic approach to prevent CPF neurotoxicity involving the regulation of autophagy. We investigated the modulation of CPF-induced apoptosis according to autophagy regulation. We found that CPF induced apoptosis in SH-SY5Y cells, as demonstrated by the activation of caspase-3 and nuclear condensation. In addition, we observed that cells treated with CPF underwent autophagic cell death by monitoring the expression of LC3-II and p62. Pretreatment with the autophagy inducer rapamycin significantly enhanced the cell viability of CPF-exposed cells, and the enhancement of cell viability was partially due to alleviation of CPF-induced apoptosis via a decrease in levels of cleaved caspase-3. Specifically, rapamycin pretreatment decreased Bax and increased Bcl-2 expression in mitochondria. In addition, rapamycin significantly decreased cytochrome c release in from mitochondria into the cytosol. However, pretreatment of cells with the autophagy inhibitor, 3-methyladenine (3MA), remarkably increased CPF toxicity in these cells; this with correlated with increased expression of Bax and decreased expression of Bcl-2 in mitochondria. Our results suggest that CPF-induced cytotoxicity is modified by autophagy regulation and that rapamycin protects against CPF-induced apoptosis by enhancing autophagy. Pharmacologic induction of autophagy by rapamycin may be a useful treatment strategy in neurodegenerative disorders. - Highlights: ► Chlorpyrifos (CPF) is cytotoxic to SH-SY5Y cells ► CPF-induced cytotoxicity is mediated by

  18. Autophagy regulates chlorpyrifos-induced apoptosis in SH-SY5Y cells

    Energy Technology Data Exchange (ETDEWEB)

    Park, Jae Hyeon [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul (Korea, Republic of); Lee, Jeong Eun [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Shin, In Chul [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Koh, Hyun Chul, E-mail: hckoh@hanyang.ac.kr [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul (Korea, Republic of)

    2013-04-01

    Recent studies have shown that up-regulation of autophagy may be a tractable therapeutic intervention for clearing disease-causing proteins, including α-synuclein, ubiquitin, and other misfolded or aggregated proteins in pesticide-induced neurodegeneration. In a previous study, we reported that chlorpyrifos (CPF)-induced mitochondria-dependent apoptosis is mediated through reactive oxygen species in SH-SY5Y cells. In this study, we explored a novel pharmacotherapeutic approach to prevent CPF neurotoxicity involving the regulation of autophagy. We investigated the modulation of CPF-induced apoptosis according to autophagy regulation. We found that CPF induced apoptosis in SH-SY5Y cells, as demonstrated by the activation of caspase-3 and nuclear condensation. In addition, we observed that cells treated with CPF underwent autophagic cell death by monitoring the expression of LC3-II and p62. Pretreatment with the autophagy inducer rapamycin significantly enhanced the cell viability of CPF-exposed cells, and the enhancement of cell viability was partially due to alleviation of CPF-induced apoptosis via a decrease in levels of cleaved caspase-3. Specifically, rapamycin pretreatment decreased Bax and increased Bcl-2 expression in mitochondria. In addition, rapamycin significantly decreased cytochrome c release in from mitochondria into the cytosol. However, pretreatment of cells with the autophagy inhibitor, 3-methyladenine (3MA), remarkably increased CPF toxicity in these cells; this with correlated with increased expression of Bax and decreased expression of Bcl-2 in mitochondria. Our results suggest that CPF-induced cytotoxicity is modified by autophagy regulation and that rapamycin protects against CPF-induced apoptosis by enhancing autophagy. Pharmacologic induction of autophagy by rapamycin may be a useful treatment strategy in neurodegenerative disorders. - Highlights: ► Chlorpyrifos (CPF) is cytotoxic to SH-SY5Y cells ► CPF-induced cytotoxicity is mediated by

  19. Salvianolic acid B inhibits autophagy and protects starving cardiac myocytes

    Institute of Scientific and Technical Information of China (English)

    Xiao HAN; Jian-xun LIU; Xin-zhi LI

    2011-01-01

    Aim: To investigate the protective or lethal role of autophagy and the effects of Salvianolic acid B (Sal B) on autophagy in starving myocytes.Methods: Cardiac myocytes were incubated under starvation conditions (GD) for O, 1, 2, 3, and 6 h. Autophagic flux in starving cells was measured via chloroquine (3 μmol/L). After myocytes were treated with Sat B (50 μmol/L) in the presence or absence of chloro-quine (3 μmol/L) under GD 3 h, the amount of LC3-11, the abundance of LC3-positive fluorescent dots in cells, cell viability and cellular ATP levels were determined using immunoblotting, immunofluorescence microscopy, MTT assay and luminometer, respectively. More-over, electron microscopy (EM) and immunofluorescent duel labeling of LC3 and Caspase-8 were used to examine the characteristics of autophagy and apoptosis.Results: Immunoblot analysis showed that the amount of LC3-11 in starving cells increased in a time-dependent manner accompanied by increased LC3-positive fluorescence and decreased cell viability and ATP content. Sal B (50 μmol/L) inhibited the increase in LC3-11, reduced the abundance of LC3 immunofluorescence and intensity of Caspase-8 fluorescence, and enhanced cellular viability and ATP levels in myocytes under GD 3 h, regardless of whether chloroquine was present.Conclusion: Autophagy induced by starvation for 3 h led to cell injury. Sal B protected starving cells by blocking the early stage of autophagic flux and inhibiting apoptosis that occurred during autophagy.

  20. Molecular mechanism and regulation of autophagy

    Institute of Scientific and Technical Information of China (English)

    Ya-ping YANG; Zhong-qin LIANG; Zhen-lun GU; Zheng-hong QIN

    2005-01-01

    Autophagy is a major cellular pathway for the degradation of long-lived proteins and cytoplasmic organelles in eukaryotic cells. A large number of intracellular/extracellular stimuli, including amino acid starvation and invasion of microorganisms, are able to induce the autophagic response in cells. The discovery of the ATG genes in yeast has greatly advanced our understanding of the molecular mechanisms participating in autophagy and the genes involved in regulating the autophagic pathway. Many yeast genes have mammalian homologs,suggesting that the basic machinery for autophagy has been evolutionarily conserved along the eukaryotic phylum. The regulation of autophagy is a very complex process. Many signaling pathways, including target of rapamycin (TOR) or mammalian target of rapamycin (mTOR), phosphatidylinositol 3-kinase-I (PI3K-I)/PKB, GTPases, calcium and protein synthesis all play important roles in regulating autophagy. The molecular mechanisms and regulation of autophagy are discussed in this review.

  1. Tomato HsfA1a plays a critical role in plant drought tolerance by activating ATG genes and inducing autophagy

    OpenAIRE

    Wang, Yu; Cai, Shuyu; Yin, Lingling; Kai SHI; Xia, Xiaojian; Zhou, Yanhong; Yu, Jingquan; Zhou, Jie

    2015-01-01

    Autophagy plays critical roles in plant responses to stress. In contrast to the wealth of information concerning the core process of plant autophagosome assembly, our understanding of the regulation of autophagy is limited. In this study, we demonstrated that transcription factor HsfA1a played a critical role in tomato tolerance to drought stress, in part through its positive role in induction of autophagy under drought stress. HsfA1a expression was induced by drought stress. Virus-induced Hs...

  2. Interleukin 17A inhibits autophagy through activation of PIK3CA to interrupt the GSK3B-mediated degradation of BCL2 in lung epithelial cells

    OpenAIRE

    Liu, Hong; Mi, Su; Li, Zhe; Hua, Fang; Hu, Zhuo-Wei

    2013-01-01

    We recently found that activation of IL17A signaling promotes the development and progression of acute and chronic pulmonary fibrosis, and that the blockade of IL17A activity attenuates pulmonary fibrosis by promoting the resolution of inflammation and the activation of autophagy. Although the induction of autophagy stimulating the collagen degradation in the fibrotic lung tissue has been identified as a mechanism responsible for the antifibrotic role of targeting IL17A, it remains to be clar...

  3. Methanolic Extract of Ganoderma lucidum Induces Autophagy of AGS Human Gastric Tumor Cells

    Directory of Open Access Journals (Sweden)

    Filipa S. Reis

    2015-09-01

    Full Text Available Ganoderma lucidum is one of the most widely studied mushroom species, particularly in what concerns its medicinal properties. Previous studies (including those from some of us have shown some evidence that the methanolic extract of G. lucidum affects cellular autophagy. However, it was not known if it induces autophagy or decreases the autophagic flux. The treatment of a gastric adenocarcinoma cell line (AGS with the mushroom extract increased the formation of autophagosomes (vacuoles typical from autophagy. Moreover, the cellular levels of LC3-II were also increased, and the cellular levels of p62 decreased, confirming that the extract affects cellular autophagy. Treating the cells with the extract together with lysossomal protease inhibitors, the cellular levels of LC3-II and p62 increased. The results obtained proved that, in AGS cells, the methanolic extract of G. lucidum causes an induction of autophagy, rather than a reduction in the autophagic flux. To our knowledge, this is the first study proving that statement.

  4. Methanolic Extract of Ganoderma lucidum Induces Autophagy of AGS Human Gastric Tumor Cells.

    Science.gov (United States)

    Reis, Filipa S; Lima, Raquel T; Morales, Patricia; Ferreira, Isabel C F R; Vasconcelos, M Helena

    2015-01-01

    Ganoderma lucidum is one of the most widely studied mushroom species, particularly in what concerns its medicinal properties. Previous studies (including those from some of us) have shown some evidence that the methanolic extract of G. lucidum affects cellular autophagy. However, it was not known if it induces autophagy or decreases the autophagic flux. The treatment of a gastric adenocarcinoma cell line (AGS) with the mushroom extract increased the formation of autophagosomes (vacuoles typical from autophagy). Moreover, the cellular levels of LC3-II were also increased, and the cellular levels of p62 decreased, confirming that the extract affects cellular autophagy. Treating the cells with the extract together with lysossomal protease inhibitors, the cellular levels of LC3-II and p62 increased. The results obtained proved that, in AGS cells, the methanolic extract of G. lucidum causes an induction of autophagy, rather than a reduction in the autophagic flux. To our knowledge, this is the first study proving that statement. PMID:26426001

  5. β-Elemene-induced autophagy protects human gastric cancer cells from undergoing apoptosis

    International Nuclear Information System (INIS)

    β-Elemene, a compound found in an herb used in traditional Chinese medicine, has shown promising anti-cancer effects against a broad spectrum of tumors. The mechanism by which β-elemene kills cells remains unclear. The aim of the present study is to investigate the anti-tumor effect of β-elemene on human gastric cancer cells and the molecular mechanism involved. β-Elemene inhibited the viability of human gastric cancer MGC803 and SGC7901 cells in a dose-dependent manner. The suppression of cell viability was due to the induction of apoptosis. A robust autophagy was observed in the cells treated with β-elemene; it was characterized by the increase of punctate LC3 dots, the cellular morphology, and the increased levels of LC3-II protein. Further study showed that β-elemene treatment up-regulated Atg5-Atg12 conjugated protein but had little effect on other autophagy-related proteins. PI3K/Akt/mTOR/p70S6K1 activity was inhibited by β-elemene. Knockdown of Beclin 1 with small interfering RNA, or co-treatment with the autophagy inhibitor, 3-methyladenine or chlorochine enhanced significantly the antitumor effects of β-elemene. Our data provides the first evidence that β-elemene induces protective autophagy and prevents human gastric cancer cells from undergoing apoptosis. A combination of β-elemene with autophagy inhibitor might thus be a useful therapeutic option for advanced gastric cancer

  6. The role of autophagy in Parkinson's disease

    Institute of Scientific and Technical Information of China (English)

    Lei Zhang; Yaru Dong; Xiaoheng Xu; Zhong Xu

    2012-01-01

    Although Parkinson's disease is the most common neurodegenerative movement disorder, the mechanisms of pathogenesis remain poorly understood. Recent findings have shown that deregulation of the autophagy-lysosome pathway is involved in the pathogenesis of Parkinson's disease. This review summarizes the most recent findings and discusses the unique role of the autophagy-lysosome pathway in Parkinson's disease to highlight the possibility of Parkinson's disease treatment strategies that incorporate autophagy-lysosome pathway modulation.

  7. Oxidative Stress and Autophagy in Cardiovascular Homeostasis

    OpenAIRE

    Morales, Cyndi R.; Pedrozo, Zully; Lavandero, Sergio; Hill, Joseph A.

    2014-01-01

    Significance: Autophagy is an evolutionarily ancient process of intracellular protein and organelle recycling required to maintain cellular homeostasis in the face of a wide variety of stresses. Dysregulation of reactive oxygen species (ROS) and reactive nitrogen species (RNS) leads to oxidative damage. Both autophagy and ROS/RNS serve pathological or adaptive roles within cardiomyocytes, depending on the context. Recent Advances: ROS/RNS and autophagy communicate with each other via both tra...

  8. Autophagy and oxidative stress in cardiovascular diseases

    OpenAIRE

    Mei, Yu; Thompson, Melissa D.; Cohen, Richard A.; Tong, XiaoYong

    2014-01-01

    Autophagy is a highly conserved degradation process by which intracellular components, including soluble macromolecules (e.g. nucleic acids, proteins, carbohydrates, and lipids) and dysfunctional organelles (e.g. mitochondria, ribosomes, peroxisomes, and endoplasmic reticulum) are degraded by the lysosome. Autophagy is orchestrated by the autophagy related protein (Atg) composed protein complexes to form autophagosomes, which fuse with lysosomes to generate autolysosomes where the contents ar...

  9. Atf3 mutant mice show reduced axon regeneration and impaired regeneration-associated gene induction after peripheral nerve injury.

    Science.gov (United States)

    Gey, Manuel; Wanner, Renate; Schilling, Corinna; Pedro, Maria T; Sinske, Daniela; Knöll, Bernd

    2016-08-01

    Axon injury in the peripheral nervous system (PNS) induces a regeneration-associated gene (RAG) response. Atf3 (activating transcription factor 3) is such a RAG and ATF3's transcriptional activity might induce 'effector' RAGs (e.g. small proline rich protein 1a (Sprr1a), Galanin (Gal), growth-associated protein 43 (Gap43)) facilitating peripheral axon regeneration. We provide a first analysis of Atf3 mouse mutants in peripheral nerve regeneration. In Atf3 mutant mice, facial nerve regeneration and neurite outgrowth of adult ATF3-deficient primary dorsal root ganglia neurons was decreased. Using genome-wide transcriptomics, we identified a neuropeptide-encoding RAG cluster (vasoactive intestinal peptide (Vip), Ngf, Grp, Gal, Pacap) regulated by ATF3. Exogenous administration of neuropeptides enhanced neurite growth of Atf3 mutant mice suggesting that these molecules might be effector RAGs of ATF3's pro-regenerative function. In addition to the induction of growth-promoting molecules, we present data that ATF3 suppresses growth-inhibiting molecules such as chemokine (C-C motif) ligand 2. In summary, we show a pro-regenerative ATF3 function during PNS nerve regeneration involving transcriptional activation of a neuropeptide-encoding RAG cluster. ATF3 is a general injury-inducible factor, therefore ATF3-mediated mechanisms identified herein might apply to other cell and injury types. PMID:27581653

  10. Atf3 mutant mice show reduced axon regeneration and impaired regeneration-associated gene induction after peripheral nerve injury

    Science.gov (United States)

    Gey, Manuel; Wanner, Renate; Schilling, Corinna; Pedro, Maria T.; Sinske, Daniela

    2016-01-01

    Axon injury in the peripheral nervous system (PNS) induces a regeneration-associated gene (RAG) response. Atf3 (activating transcription factor 3) is such a RAG and ATF3's transcriptional activity might induce ‘effector’ RAGs (e.g. small proline rich protein 1a (Sprr1a), Galanin (Gal), growth-associated protein 43 (Gap43)) facilitating peripheral axon regeneration. We provide a first analysis of Atf3 mouse mutants in peripheral nerve regeneration. In Atf3 mutant mice, facial nerve regeneration and neurite outgrowth of adult ATF3-deficient primary dorsal root ganglia neurons was decreased. Using genome-wide transcriptomics, we identified a neuropeptide-encoding RAG cluster (vasoactive intestinal peptide (Vip), Ngf, Grp, Gal, Pacap) regulated by ATF3. Exogenous administration of neuropeptides enhanced neurite growth of Atf3 mutant mice suggesting that these molecules might be effector RAGs of ATF3's pro-regenerative function. In addition to the induction of growth-promoting molecules, we present data that ATF3 suppresses growth-inhibiting molecules such as chemokine (C-C motif) ligand 2. In summary, we show a pro-regenerative ATF3 function during PNS nerve regeneration involving transcriptional activation of a neuropeptide-encoding RAG cluster. ATF3 is a general injury-inducible factor, therefore ATF3-mediated mechanisms identified herein might apply to other cell and injury types. PMID:27581653

  11. Modulating autophagy: a strategy for cancer therapy

    Institute of Scientific and Technical Information of China (English)

    Jun-Lin Li; Shao-Liang Han; Xia Fan

    2011-01-01

    Autophagy is a process in which long-lived proteins,damaged cell organelles,and other cellular particles are sequestered and degraded.This process is important for maintaining the cellular microenvironment when the cell is under stress.Many studies have shown that autophagy plays a complex role in human diseases,especially in cancer,where it is known to have paradoxical effects.Namely,autophagy provides the energy for metabolism and tumor growth and leads to cell death that promotes tumor suppression.The link between autophagy and cancer is also evident in that some of the genes that regulate carcinogenesis,oncogenes and tumor suppressor genes,participate in or impact the autophagy process.Therefore,modulating autophagy will be a valuable topic for cancer therapy.Many studies have shown that autophagy can inhibit the tumor growth when autophagy modulators are combined with radiotherapy and/or chemotherapy.These findings suggest that autophagy may be a potent target for cancer therapy.

  12. Involvement of Autophagy in Coronavirus Replication

    Directory of Open Access Journals (Sweden)

    Paul Britton

    2012-11-01

    Full Text Available Coronaviruses are single stranded, positive sense RNA viruses, which induce the rearrangement of cellular membranes upon infection of a host cell. This provides the virus with a platform for the assembly of viral replication complexes, improving efficiency of RNA synthesis. The membranes observed in coronavirus infected cells include double membrane vesicles. By nature of their double membrane, these vesicles resemble cellular autophagosomes, generated during the cellular autophagy pathway. In addition, coronavirus infection has been demonstrated to induce autophagy. Here we review current knowledge of coronavirus induced membrane rearrangements and the involvement of autophagy or autophagy protein microtubule associated protein 1B light chain 3 (LC3 in coronavirus replication.

  13. The autophagy associated gene, ULK1, promotes tolerance to chronic and acute hypoxia

    International Nuclear Information System (INIS)

    Background and purpose: Tumor hypoxia is associated with therapy resistance and malignancy. Previously we demonstrated that activation of autophagy and the unfolded protein response (UPR) promote hypoxia tolerance. Here we explored the importance of ULK1 in hypoxia tolerance, autophagy induction and its prognostic value for recurrence after treatment. Material and methods: Hypoxic regulation of ULK1 mRNA and protein was assessed in vitro and in primary human head and neck squamous cell carcinoma (HNSCC) xenografts. Its importance in autophagy induction, mitochondrial homeostasis and tolerance to chronic and acute hypoxia was evaluated in ULK1 knockdown cells. The prognostic value of ULK1 mRNA expression was assessed in 82 HNSCC patients. Results: ULK1 enrichment was observed in hypoxic tumor regions. High enrichment was associated with a high hypoxic fraction. In line with these findings, high ULK1 expression in HNSCC patients appeared associated with poor local control. Exposure of cells to hypoxia induced ULK1 mRNA in a UPR and HIF1α dependent manner. ULK1 knockdown decreased autophagy activation, increased mitochondrial mass and ROS exposure and sensitized cells to acute and chronic hypoxia. Conclusions: We demonstrate that ULK1 is a hypoxia regulated gene and is associated with hypoxia tolerance and a worse clinical outcome

  14. Suppression of autophagy by CUB domain-containing protein 1 signaling is essential for anchorage-independent survival of lung cancer cells.

    Science.gov (United States)

    Uekita, Takamasa; Fujii, Satoko; Miyazawa, Yuri; Hashiguchi, Akinori; Abe, Hitosi; Sakamoto, Michiie; Sakai, Ryuichi

    2013-07-01

    CUB (C1r/C1s, urchin embryonic growth factor, BMP1) domain-containing protein 1 (CDCP1) has been implicated in promoting metastasis of cancer cells through several mechanisms, including the inhibition of anoikis, which is cell death triggered by the loss of extracellular matrix interactions. However, the mechanism inhibiting cell death regulated by CDCP1 remains elusive. Inhibition of CDCP1 expression using small interfering RNA (siRNA) induced the cell death of suspended cancer cells without cleaving caspase-3, a marker of apoptosis; cell death was not inhibited by a general caspase inhibitor, suggesting that the loss of CDCP1 induces caspase-independent cell death. In contrast, knockdown of CDCP1 as well as protein kinase Cδ (PKCδ), a downstream effector of CDCP1, in a suspension culture of lung cancer cells resulted in marked induction of membranous microtubule-associated protein 1 light chain 3 (LC3)-II protein, a hallmark of autophagy, and caused the formation of an autophagosome structure visualized using green fluorescent protein-tagged LC3-II. Expression and phosphorylation of exogenous CDCP1 by Fyn kinase reduced the formation of autophagosomes and inhibited phosphorylation of CDCP1 by PP2, a Src kinase inhibitor or inhibited PKCδ by rottlerin, stimulating autophagosome formation. Moreover, death of suspended lung cancer cells induced by CDCP1 siRNA or by PKCδ siRNA was reduced by the autophagy inhibitor 3-methyladenine. These results indicate that CDCP1-PKCδ signaling plays a critical role in inhibiting autophagy, which is responsible for anoikis resistance of lung cancer cells. PMID:23510015

  15. Chemical modification of RNA-based medicine can be used to reduce its induction of the innate immune response

    DEFF Research Database (Denmark)

    Schyth, Brian Dall; Bramsen, Jesper Bertram; Kjems, Jørgen;

    2012-01-01

    trout and VHSV to screen siRNAs containing various chemical modifications of the RNA backbone and found that was possible to modify the backbone so as to reduce the antiviral effect of the RNA. Antiviral protection was also dependent upon localisation of the modified nucleotide residues in the RNA...

  16. Coffee induces autophagy in vivo

    OpenAIRE

    Pietrocola, Federico; Malik, Shoaib Ahmad; Mariño, Guillermo; Vacchelli, Erika; Senovilla, Laura; Chaba, Kariman; Niso-Santano, Mireia; Maiuri, Maria Chiara; Madeo, Frank; Kroemer, Guido

    2014-01-01

    Epidemiological studies and clinical trials revealed that chronic consumption coffee is associated with the inhibition of several metabolic diseases as well as reduction in overall and cause-specific mortality. We show that both natural and decaffeinated brands of coffee similarly rapidly trigger autophagy in mice. One to 4 h after coffee consumption, we observed an increase in autophagic flux in all investigated organs (liver, muscle, heart) in vivo, as indicated by the increased lipidation ...

  17. Mir-30d increases intracellular survival of Helicobacter pylori through inhibition of autophagy pathway

    Science.gov (United States)

    Yang, Xiao-Jun; Si, Ruo-Huang; Liang, Yu-He; Ma, Bing-Qiang; Jiang, Ze-Bin; Wang, Bin; Gao, Peng

    2016-01-01

    AIM: To determine if mir-30d inhibits the autophagy response to Helicobacter pylori (H. pylori) invasion and increases H. pylori intracellular survival. METHODS: The expression of mir-30d was detected by quantitative polymerase chain reaction (PCR), and autophagy level was examined by transmission electron microscopy, western blot, and GFP-LC3 puncta assay in human AGS cells and GES-1 cells. Luciferase reporter assay was applied to confirm the specificity of mir-30d regulation on the expression of several core molecules involved in autophagy pathway. The expression of multiple core proteins were analyzed at both the mRNA and protein level, and the intracellular survival of H. pylori after different treatments was detected by gentamicin protection assay. RESULTS: Autophagy level was increased in AGS and GES-1 cells in response to H. pylori infection, which was accompanied by upregulation of mir-30d expression (P pylori infection). In the two gastric epithelial cell lines, mimic mir-30d was found to repress the autophagy process, whereas mir-30d inhibitor increased autophagy response to H. pylori invasion. mir-30d mimic decreased the luciferase activity of wild type reporter plasmids carrying the 3′ untranslated region (UTR) of all five tested genes (ATG2B, ATG5, ATG12, BECN1, and BNIP3L), whereas it had no effect on the mutant reporter plasmids. These five genes are core genes of autophagy pathway, and their expression was reduced significantly after mir-30d mimic transfection (P pylori in AGS cells. CONCLUSION: Mir-30d increases intracellular survival of H. pylori in gastric epithelial cells through inhibition of multiple core proteins in the autophagy pathway. PMID:27099441

  18. Posttraumatic administration of luteolin protects mice from traumatic brain injury: implication of autophagy and inflammation.

    Science.gov (United States)

    Xu, Jianguo; Wang, Handong; Lu, Xinyu; Ding, Ke; Zhang, Li; He, Jin; Wei, Wuting; Wu, Yong

    2014-09-25

    Secondary brain insult induced by traumatic brain injury (TBI), including excitotoxicity, oxidative stress, inflammatory response, and neuronal degeneration, is sensitive to therapeutic interventions; therefore, searching for neuroprotective agents represents a promising therapeutic strategy for TBI treatment. Luteolin, a member of the flavonoid family, has recently been proven to modulate autophagy. However, whether it activates autophagy after TBI thereby alleviating the secondary insult is not yet understood. Here, we aimed to evaluate the neuroprotection of luteolin against TBI and the potential role of autophagy where it is involved. For this purpose, mice were randomly divided into four groups and then subjected to TBI. The treatment mice received luteolin at a dose of 30mg/kg 30min post-TBI based on our previous study. We employed western blot, immunofluorescence and quantitative real-time PCR to determine autophagy process and inflammatory response among different groups. Autophagy was found to be enhanced after luteolin treatment according to the expressions of autophagic markers. Furthermore, luteolin decreased nuclear accumulation of p65 induced by TBI, indicating attenuation of inflammation. In line with these observations, luteolin decreased mRNA and protein expressions of pro-inflammatory factors IL-1b and TNF-a. At last, luteolin reduced neuronal degeneration, and alleviated brain edema and blood-brain barrier (BBB) disruption. In conclusion, these results implied that luteolin protected mice brain from traumatic brain injury by inhibiting inflammatory response, and luteolin-induced autophagy might play a pivotal role in its neuroprotection. PMID:25093609

  19. C60(Nd) nanoparticles enhance chemotherapeutic susceptibility of cancer cells by modulation of autophagy

    International Nuclear Information System (INIS)

    Autophagy, an evolutionally conserved intracellular process degrading cytoplasmic proteins and organelles for recycling, has become one of the most remarkable strategies applied in cancer research. The fullerene C60 nanoparticle (nC60) has been shown to induce autophagy and sensitize chemotherapeutic killing of cancer cells, but the details still remain unknown. Here we show that a water-dispersed nanoparticle solution of derivatized fullerene C60, C60(Nd) nanoparticles (nC60(Nd)), has greater potential in inducing autophagy and sensitizing chemotherapeutic killing of both normal and drug-resistant cancer cells than nC60 does in an autophagy-dependent fashion. Additionally we further demonstrated that autophagy induced by nC60/C60(Nd) and Rapamycin had completely different roles in cancer chemotherapy. Our results, for the first time, revealed a novel and more potent derivative of the C60 nanoparticle in enhancing the cytotoxicity of chemotherapeutic agents and reducing drug resistance through autophagy modulation, which may ultimately lead to novel therapeutic strategies in cancer therapy.

  20. C60(Nd) nanoparticles enhance chemotherapeutic susceptibility of cancer cells by modulation of autophagy

    Science.gov (United States)

    Wei, Pengfei; Zhang, Li; Lu, Yang; Man, Na; Wen, Longping

    2010-12-01

    Autophagy, an evolutionally conserved intracellular process degrading cytoplasmic proteins and organelles for recycling, has become one of the most remarkable strategies applied in cancer research. The fullerene C60 nanoparticle (nC60) has been shown to induce autophagy and sensitize chemotherapeutic killing of cancer cells, but the details still remain unknown. Here we show that a water-dispersed nanoparticle solution of derivatized fullerene C60, C60(Nd) nanoparticles (nC60(Nd)), has greater potential in inducing autophagy and sensitizing chemotherapeutic killing of both normal and drug-resistant cancer cells than nC60 does in an autophagy-dependent fashion. Additionally we further demonstrated that autophagy induced by nC60/C60(Nd) and Rapamycin had completely different roles in cancer chemotherapy. Our results, for the first time, revealed a novel and more potent derivative of the C60 nanoparticle in enhancing the cytotoxicity of chemotherapeutic agents and reducing drug resistance through autophagy modulation, which may ultimately lead to novel therapeutic strategies in cancer therapy.

  1. Autophagy selectivity through receptor clustering

    Science.gov (United States)

    Rutenberg, Andrew; Brown, Aidan

    Substrate selectivity in autophagy requires an all-or-none cellular response. We focus on peroxisomes, for which autophagy receptor proteins NBR1 and p62 are well characterized. Using computational models, we explore the hypothesis that physical clustering of autophagy receptor proteins on the peroxisome surface provides an appropriate all-or-none response. We find that larger peroxisomes nucleate NBR1 clusters first, and lose them due to competitive coarsening last, resulting in significant size-selectivity. We then consider a secondary hypothesis that p62 inhibits NBR1 cluster formation. We find that p62 inhibition enhances size-selectivity enough that, even if there is no change of the pexophagy rate, the volume of remaining peroxisomes can significantly decrease. We find that enhanced ubiquitin levels suppress size-selectivity, and that this effect is more pronounced for individual peroxisomes. Sufficient ubiquitin allows receptor clusters to form on even the smallest peroxisomes. We conclude that NBR1 cluster formation provides a viable physical mechanism for all-or-none substrate selectivity in pexophagy. We predict that cluster formation is associated with significant size-selectivity. Now at Simon Fraser University.

  2. Autophagy and the (prorenin receptor

    Directory of Open Access Journals (Sweden)

    KatrinaJeanBinger

    2013-10-01

    Full Text Available The (prorenin receptor (PRR is a newly reported member of the renin-angiotensin system (RAS; a hormonal cascade responsible for regulating blood pressure. Originally, the identification of PRR was heralded as the next drug target of the RAS, of which such therapies would have increased benefits against target-organ damage and hypertension. However, in the years since its discovery several conditional knockout mouse models of PRR have demonstrated an essential role for this receptor unrelated to the renin-angiotensin system and blood pressure. Deletion of PRR in podocytes or cardiomyocytes resulted in the rapid onset of organ failure, eventuating in animal mortality after only a matter of weeks. In both cases, deletion of PRR resulted in the intracellular accumulation of autophagosomes and misfolded proteins, indicating a disturbance in autophagy. In light of the fact that the majority of PRR is located intracellularly, this molecular function appears to be more relevant than its ability to bind to high, non-physiological concentrations of (prorenin. This review will focus on the role of PRR in autophagy and its importance in maintaining cellular homeostasis. Understanding the link between PRR, autophagy and how its loss results in cell death will be essential for deciphering its role in physiology and pathology.

  3. Tumor suppression in mice lacking GABARAP, an Atg8/LC3 family member implicated in autophagy, is associated with alterations in cytokine secretion and cell death.

    Science.gov (United States)

    Salah, F S; Ebbinghaus, M; Muley, V Y; Zhou, Z; Al-Saadi, K R D; Pacyna-Gengelbach, M; O'Sullivan, G A; Betz, H; König, R; Wang, Z-Q; Bräuer, R; Petersen, I

    2016-01-01

    GABARAP belongs to an evolutionary highly conserved gene family that has a fundamental role in autophagy. There is ample evidence for a crosstalk between autophagy and apoptosis as well as the immune response. However, the molecular details for these interactions are not fully characterized. Here, we report that the ablation of murine GABARAP, a member of the Atg8/LC3 family that is central to autophagosome formation, suppresses the incidence of tumor formation mediated by the carcinogen DMBA and results in an enhancement of the immune response through increased secretion of IL-1β, IL-6, IL-2 and IFN-γ from stimulated macrophages and lymphocytes. In contrast, TGF-β1 was significantly reduced in the serum of these knockout mice. Further, DMBA treatment of these GABARAP knockout mice reduced the cellularity of the spleen and the growth of mammary glands through the induction of apoptosis. Gene expression profiling of mammary glands revealed significantly elevated levels of Xaf1, an apoptotic inducer and tumor-suppressor gene, in knockout mice. Furthermore, DMBA treatment triggered the upregulation of pro-apoptotic (Bid, Apaf1, Bax), cell death (Tnfrsf10b, Ripk1) and cell cycle inhibitor (Cdkn1a, Cdkn2c) genes in the mammary glands. Finally, tumor growth of B16 melanoma cells after subcutaneous inoculation was inhibited in GABARAP-deficient mice. Together, these data provide strong evidence for the involvement of GABARAP in tumorigenesis in vivo by delaying cell death and its associated immune-related response. PMID:27124579

  4. The effect of positive mood induction on reducing reinstatement fear: Relevance for long term outcomes of exposure therapy

    OpenAIRE

    Zbozinek, Tomislav D.; Holmes, Emily A.; Craske, Michelle G.

    2015-01-01

    While exposure therapy is effective in treating anxiety, fear can return after exposure. Return of fear can be understood through mechanisms of extinction learning. One form of return of fear is reinstatement, or, the fear that results from an unsignaled unconditional stimulus (US) presentation after extinction. Though the conditional response (CR; e.g., fear) typically reduces during extinction, the excitatory conditional stimulus (CS+) valence remains negative. The more negative the CS+ val...

  5. Autophagy is involved in anti-viral activity of pentagalloylglucose (PGG) against Herpes simplex virus type 1 infection in vitro

    International Nuclear Information System (INIS)

    Research highlights: → We showed PGG has anti-viral activity against Herpes simplex virus type 1 (HSV-1) and can induce autophgy. → Autophagy may be a novel and important mechanism mediating PGG anti-viral activities. → Inhibition of mTOR pathway is an important mechanism of induction of autophagy by PGG. -- Abstract: Pentagalloylglucose (PGG) is a natural polyphenolic compound with broad-spectrum anti-viral activity, however, the mechanisms underlying anti-viral activity remain undefined. In this study, we investigated the effects of PGG on anti-viral activity against Herpes simplex virus type 1 (HSV-1) associated with autophagy. We found that the PGG anti-HSV-1 activity was impaired significantly in MEF-atg7-/- cells (autophagy-defective cells) derived from an atg7-/- knockout mouse. Transmission electron microscopy revealed that PGG-induced autophagosomes engulfed HSV-1 virions. The mTOR signaling pathway, an essential pathway for the regulation of autophagy, was found to be suppressed following PGG treatment. Data presented in this report demonstrated for the first time that autophagy induced following PGG treatment contributed to its anti-HSV activity in vitro.

  6. MARCH2 regulates autophagy by promoting CFTR ubiquitination and degradation and PIK3CA-AKT-MTOR signaling.

    Science.gov (United States)

    Xia, Dan; Qu, Liujing; Li, Ge; Hongdu, Beiqi; Xu, Chentong; Lin, Xin; Lou, Yaxin; He, Qihua; Ma, Dalong; Chen, Yingyu

    2016-09-01

    MARCH2 (membrane-associated RING-CH protein 2), an E3 ubiquitin ligase, is mainly associated with the vesicle trafficking. In the present study, for the first time, we demonstrated that MARCH2 negatively regulates autophagy. Our data indicated that overexpression of MARCH2 impaired autophagy, as evidenced by attenuated levels of LC3B-II and impaired degradation of endogenous and exogenous autophagic substrates. By contrast, loss of MARCH2 expression had the opposite effects. In vivo experiments demonstrate that MARCH2 knockout mediated autophagy results in an inhibition of tumorigenicity. Further investigation revealed that the induction of autophagy by MARCH2 deficiency was mediated through the PIK3CA-AKT-MTOR signaling pathway. Additionally, we found that MARCH2 interacts with CFTR (cystic fibrosis transmembrane conductance regulator), promotes the ubiquitination and degradation of CFTR, and inhibits CFTR-mediated autophagy in tumor cells. The functional PDZ domain of MARCH2 is required for the association with CFTR. Thus, our study identified a novel negative regulator of autophagy and suggested that the physical and functional connection between the MARCH2 and CFTR in different conditions will be elucidated in the further experiments. PMID:27308891

  7. Characterization of the autophagy marker protein Atg8 reveals atypical features of autophagy in Plasmodium falciparum.

    Directory of Open Access Journals (Sweden)

    Rahul Navale

    Full Text Available Conventional autophagy is a lysosome-dependent degradation process that has crucial homeostatic and regulatory functions in eukaryotic organisms. As malaria parasites must dispose a number of self and host cellular contents, we investigated if autophagy in malaria parasites is similar to the conventional autophagy. Genome wide analysis revealed a partial autophagy repertoire in Plasmodium, as homologs for only 15 of the 33 yeast autophagy proteins could be identified, including the autophagy marker Atg8. To gain insights into autophagy in malaria parasites, we investigated Plasmodium falciparum Atg8 (PfAtg8 employing techniques and conditions that are routinely used to study autophagy. Atg8 was similarly expressed and showed punctate localization throughout the parasite in both asexual and sexual stages; it was exclusively found in the pellet fraction as an integral membrane protein, which is in contrast to the yeast or mammalian Atg8 that is distributed among cytosolic and membrane fractions, and suggests for a constitutive autophagy. Starvation, the best known autophagy inducer, decreased PfAtg8 level by almost 3-fold compared to the normally growing parasites. Neither the Atg8-associated puncta nor the Atg8 expression level was significantly altered by treatment of parasites with routinely used autophagy inhibitors (cysteine (E64 and aspartic (pepstatin protease inhibitors, the kinase inhibitor 3-methyladenine, and the lysosomotropic agent chloroquine, indicating an atypical feature of autophagy. Furthermore, prolonged inhibition of the major food vacuole protease activity by E64 and pepstatin did not cause accumulation of the Atg8-associated puncta in the food vacuole, suggesting that autophagy is primarily not meant for degradative function in malaria parasites. Atg8 showed partial colocalization with the apicoplast; doxycycline treatment, which disrupts apicoplast, did not affect Atg8 localization, suggesting a role, but not exclusive, in

  8. Guidelines for the use and interpretation of assays for monitoring autophagy.

    Science.gov (United States)

    Klionsky, Daniel J; Abdalla, Fabio C; Abeliovich, Hagai; Abraham, Robert T; Acevedo-Arozena, Abraham; Adeli, Khosrow; Agholme, Lotta; Agnello, Maria; Agostinis, Patrizia; Aguirre-Ghiso, Julio A; Ahn, Hyung Jun; Ait-Mohamed, Ouardia; Ait-Si-Ali, Slimane; Akematsu, Takahiko; Akira, Shizuo; Al-Younes, Hesham M; Al-Zeer, Munir A; Albert, Matthew L; Albin, Roger L; Alegre-Abarrategui, Javier; Aleo, Maria Francesca; Alirezaei, Mehrdad; Almasan, Alexandru; Almonte-Becerril, Maylin; Amano, Atsuo; Amaravadi, Ravi; Amarnath, Shoba; Amer, Amal O; Andrieu-Abadie, Nathalie; Anantharam, Vellareddy; Ann, David K; Anoopkumar-Dukie, Shailendra; Aoki, Hiroshi; Apostolova, Nadezda; Arancia, Giuseppe; Aris, John P; Asanuma, Katsuhiko; Asare, Nana Y O; Ashida, Hisashi; Askanas, Valerie; Askew, David S; Auberger, Patrick; Baba, Misuzu; Backues, Steven K; Baehrecke, Eric H; Bahr, Ben A; Bai, Xue-Yuan; Bailly, Yannick; Baiocchi, Robert; Baldini, Giulia; Balduini, Walter; Ballabio, Andrea; Bamber, Bruce A; 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Pinkas-Kramarski, Ronit; Piras, Antonio; Piri, Natik; Platanias, Leonidas C; Pöggeler, Stefanie; Poirot, Marc; Poletti, Angelo; Poüs, Christian; Pozuelo-Rubio, Mercedes; Prætorius-Ibba, Mette; Prasad, Anil; Prescott, Mark; Priault, Muriel; Produit-Zengaffinen, Nathalie; Progulske-Fox, Ann; Proikas-Cezanne, Tassula; Przedborski, Serge; Przyklenk, Karin; Puertollano, Rosa; Puyal, Julien; Qian, Shu-Bing; Qin, Liang; Qin, Zheng-Hong; Quaggin, Susan E; Raben, Nina; Rabinowich, Hannah; Rabkin, Simon W; Rahman, Irfan; Rami, Abdelhaq; Ramm, Georg; Randall, Glenn; Randow, Felix; Rao, V Ashutosh; Rathmell, Jeffrey C; Ravikumar, Brinda; Ray, Swapan K; Reed, Bruce H; Reed, John C; Reggiori, Fulvio; Régnier-Vigouroux, Anne; Reichert, Andreas S; Reiners, John J; Reiter, Russel J; Ren, Jun; Revuelta, José L; Rhodes, Christopher J; Ritis, Konstantinos; Rizzo, Elizete; Robbins, Jeffrey; Roberge, Michel; Roca, Hernan; Roccheri, Maria C; Rocchi, Stephane; Rodemann, H Peter; Rodríguez de Córdoba, Santiago; Rohrer, Bärbel; Roninson, Igor B; Rosen, Kirill; Rost-Roszkowska, Magdalena M; Rouis, Mustapha; Rouschop, Kasper M A; Rovetta, Francesca; Rubin, Brian P; Rubinsztein, David C; Ruckdeschel, Klaus; Rucker, Edmund B; Rudich, Assaf; Rudolf, Emil; Ruiz-Opazo, Nelson; Russo, Rossella; Rusten, Tor Erik; Ryan, Kevin M; Ryter, Stefan W; Sabatini, David M; Sadoshima, Junichi; Saha, Tapas; Saitoh, Tatsuya; Sakagami, Hiroshi; Sakai, Yasuyoshi; Salekdeh, Ghasem Hoseini; Salomoni, Paolo; Salvaterra, Paul M; Salvesen, Guy; Salvioli, Rosa; Sanchez, Anthony M J; Sánchez-Alcázar, José A; Sánchez-Prieto, Ricardo; Sandri, Marco; Sankar, Uma; Sansanwal, Poonam; Santambrogio, Laura; Saran, Shweta; Sarkar, Sovan; Sarwal, Minnie; Sasakawa, Chihiro; Sasnauskiene, Ausra; Sass, Miklós; Sato, Ken; Sato, Miyuki; Schapira, Anthony H V; Scharl, Michael; Schätzl, Hermann M; Scheper, Wiep; Schiaffino, Stefano; Schneider, Claudio; Schneider, Marion E; Schneider-Stock, Regine; Schoenlein, Patricia V; Schorderet, Daniel F; Schüller, Christoph; Schwartz, Gary K; Scorrano, Luca; Sealy, Linda; Seglen, Per O; Segura-Aguilar, Juan; Seiliez, Iban; Seleverstov, Oleksandr; Sell, Christian; Seo, Jong Bok; Separovic, Duska; Setaluri, Vijayasaradhi; Setoguchi, Takao; Settembre, Carmine; Shacka, John J; Shanmugam, Mala; Shapiro, Irving M; Shaulian, Eitan; Shaw, Reuben J; Shelhamer, James H; Shen, Han-Ming; Shen, Wei-Chiang; Sheng, Zu-Hang; Shi, Yang; Shibuya, Kenichi; Shidoji, Yoshihiro; Shieh, Jeng-Jer; Shih, Chwen-Ming; Shimada, Yohta; Shimizu, Shigeomi; Shintani, Takahiro; Shirihai, Orian S; Shore, Gordon C; Sibirny, Andriy A; Sidhu, Stan B; Sikorska, Beata; Silva-Zacarin, Elaine C M; Simmons, Alison; Simon, Anna Katharina; Simon, Hans-Uwe; Simone, Cristiano; Simonsen, Anne; Sinclair, David A; Singh, Rajat; Sinha, Debasish; Sinicrope, Frank A; Sirko, Agnieszka; Siu, Parco M; Sivridis, Efthimios; Skop, Vojtech; Skulachev, Vladimir P; Slack, Ruth S; Smaili, Soraya S; Smith, Duncan R; Soengas, Maria S; Soldati, Thierry; 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Thumm, Michael; Tian, FengFeng; Tian, Yuan; Tocchini-Valentini, Glauco; Tolkovsky, Aviva M; Tomino, Yasuhiko; Tönges, Lars; Tooze, Sharon A; Tournier, Cathy; Tower, John; Towns, Roberto; Trajkovic, Vladimir; Travassos, Leonardo H; Tsai, Ting-Fen; Tschan, Mario P; Tsubata, Takeshi; Tsung, Allan; Turk, Boris; Turner, Lorianne S; Tyagi, Suresh C; Uchiyama, Yasuo; Ueno, Takashi; Umekawa, Midori; Umemiya-Shirafuji, Rika; Unni, Vivek K; Vaccaro, Maria I; Valente, Enza Maria; Van den Berghe, Greet; van der Klei, Ida J; van Doorn, Wouter; van Dyk, Linda F; van Egmond, Marjolein; van Grunsven, Leo A; Vandenabeele, Peter; Vandenberghe, Wim P; Vanhorebeek, Ilse; Vaquero, Eva C; Velasco, Guillermo; Vellai, Tibor; Vicencio, Jose Miguel; Vierstra, Richard D; Vila, Miquel; Vindis, Cécile; Viola, Giampietro; Viscomi, Maria Teresa; Voitsekhovskaja, Olga V; von Haefen, Clarissa; Votruba, Marcela; Wada, Keiji; Wade-Martins, Richard; Walker, Cheryl L; Walsh, Craig M; Walter, Jochen; Wan, Xiang-Bo; Wang, Aimin; 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Yang, Wannian; Yang, Wei Yuan; Yang, Zhifen; Yao, Meng-Chao; Yao, Tso-Pang; Yeganeh, Behzad; Yen, Wei-Lien; Yin, Jia-jing; Yin, Xiao-Ming; Yoo, Ook-Joon; Yoon, Gyesoon; Yoon, Seung-Yong; Yorimitsu, Tomohiro; Yoshikawa, Yuko; Yoshimori, Tamotsu; Yoshimoto, Kohki; You, Ho Jin; Youle, Richard J; Younes, Anas; Yu, Li; Yu, Long; Yu, Seong-Woon; Yu, Wai Haung; Yuan, Zhi-Min; Yue, Zhenyu; Yun, Cheol-Heui; Yuzaki, Michisuke; Zabirnyk, Olga; Silva-Zacarin, Elaine; Zacks, David; Zacksenhaus, Eldad; Zaffaroni, Nadia; Zakeri, Zahra; Zeh, Herbert J; Zeitlin, Scott O; Zhang, Hong; Zhang, Hui-Ling; Zhang, Jianhua; Zhang, Jing-Pu; Zhang, Lin; Zhang, Long; Zhang, Ming-Yong; Zhang, Xu Dong; Zhao, Mantong; Zhao, Yi-Fang; Zhao, Ying; Zhao, Zhizhuang J; Zheng, Xiaoxiang; Zhivotovsky, Boris; Zhong, Qing; Zhou, Cong-Zhao; Zhu, Changlian; Zhu, Wei-Guo; Zhu, Xiao-Feng; Zhu, Xiongwei; Zhu, Yuangang; Zoladek, Teresa; Zong, Wei-Xing; Zorzano, Antonio; Zschocke, Jürgen; Zuckerbraun, Brian

    2012-04-01

    In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused

  9. Guidelines for the use and interpretation of assays for monitoring autophagy

    Science.gov (United States)

    Klionsky, Daniel J.; Abdalla, Fabio C.; Abeliovich, Hagai; Abraham, Robert T.; Acevedo-Arozena, Abraham; Adeli, Khosrow; Agholme, Lotta; Agnello, Maria; Agostinis, Patrizia; Aguirre-Ghiso, Julio A.; Ahn, Hyung Jun; Ait-Mohamed, Ouardia; Ait-Si-Ali, Slimane; Akematsu, Takahiko; Akira, Shizuo; Al-Younes, Hesham M.; Al-Zeer, Munir A.; Albert, Matthew L.; Albin, Roger L.; Alegre-Abarrategui, Javier; Aleo, Maria Francesca; Alirezaei, Mehrdad; Almasan, Alexandru; Almonte-Becerril, Maylin; Amano, Atsuo; Amaravadi, Ravi K.; Amarnath, Shoba; Amer, Amal O.; Andrieu-Abadie, Nathalie; Anantharam, Vellareddy; Ann, David K.; Anoopkumar-Dukie, Shailendra; Aoki, Hiroshi; Apostolova, Nadezda; Arancia, Giuseppe; Aris, John P.; Asanuma, Katsuhiko; Asare, Nana Y.O.; Ashida, Hisashi; Askanas, Valerie; Askew, David S.; Auberger, Patrick; Baba, Misuzu; Backues, Steven K.; Baehrecke, Eric H.; Bahr, Ben A.; Bai, Xue-Yuan; Bailly, Yannick; Baiocchi, Robert; Baldini, Giulia; Balduini, Walter; Ballabio, Andrea; Bamber, Bruce A.; Bampton, Edward T.W.; Juhász, Gábor; Bartholomew, Clinton R.; Bassham, Diane C.; Bast, Robert C.; Batoko, Henri; Bay, Boon-Huat; Beau, Isabelle; Béchet, Daniel M.; Begley, Thomas J.; Behl, Christian; Behrends, Christian; Bekri, Soumeya; Bellaire, Bryan; Bendall, Linda J.; Benetti, Luca; Berliocchi, Laura; Bernardi, Henri; Bernassola, Francesca; Besteiro, Sébastien; Bhatia-Kissova, Ingrid; Bi, Xiaoning; Biard-Piechaczyk, Martine; Blum, Janice S.; Boise, Lawrence H.; Bonaldo, Paolo; Boone, David L.; Bornhauser, Beat C.; Bortoluci, Karina R.; Bossis, Ioannis; Bost, Frédéric; Bourquin, Jean-Pierre; Boya, Patricia; Boyer-Guittaut, Michaël; Bozhkov, Peter V.; Brady, Nathan R; Brancolini, Claudio; Brech, Andreas; Brenman, Jay E.; Brennand, Ana; Bresnick, Emery H.; Brest, Patrick; Bridges, Dave; Bristol, Molly L.; Brookes, Paul S.; Brown, Eric J.; Brumell, John H.; Brunetti-Pierri, Nicola; Brunk, Ulf T.; Bulman, Dennis E.; Bultman, Scott J.; Bultynck, Geert; Burbulla, Lena F.; Bursch, Wilfried; Butchar, Jonathan P.; Buzgariu, Wanda; Bydlowski, Sergio P.; Cadwell, Ken; Cahová, Monika; Cai, Dongsheng; Cai, Jiyang; Cai, Qian; Calabretta, Bruno; Calvo-Garrido, Javier; Camougrand, Nadine; Campanella, Michelangelo; Campos-Salinas, Jenny; Candi, Eleonora; Cao, Lizhi; Caplan, Allan B.; Carding, Simon R.; Cardoso, Sandra M.; Carew, Jennifer S.; Carlin, Cathleen R.; Carmignac, Virginie; Carneiro, Leticia A.M.; Carra, Serena; Caruso, Rosario A.; Casari, Giorgio; Casas, Caty; Castino, Roberta; Cebollero, Eduardo; Cecconi, Francesco; Celli, Jean; Chaachouay, Hassan; Chae, Han-Jung; Chai, Chee-Yin; Chan, David C.; Chan, Edmond Y.; Chang, Raymond Chuen-Chung; Che, Chi-Ming; Chen, Ching-Chow; Chen, Guang-Chao; Chen, Guo-Qiang; Chen, Min; Chen, Quan; Chen, Steve S.-L.; Chen, WenLi; Chen, Xi; Chen, Xiangmei; Chen, Xiequn; Chen, Ye-Guang; Chen, Yingyu; Chen, Yongqiang; Chen, Yu-Jen; Chen, Zhixiang; Cheng, Alan; Cheng, Christopher H.K.; Cheng, Yan; Cheong, Heesun; Cheong, Jae-Ho; Cherry, Sara; Chess-Williams, Russ; Cheung, Zelda H.; Chevet, Eric; Chiang, Hui-Ling; Chiarelli, Roberto; Chiba, Tomoki; Chin, Lih-Shen; Chiou, Shih-Hwa; Chisari, Francis V.; Cho, Chi Hin; Cho, Dong-Hyung; Choi, Augustine M.K.; Choi, DooSeok; Choi, Kyeong Sook; Choi, Mary E.; Chouaib, Salem; Choubey, Divaker; Choubey, Vinay; Chu, Charleen T.; Chuang, Tsung-Hsien; Chueh, Sheau-Huei; Chun, Taehoon; Chwae, Yong-Joon; Chye, Mee-Len; Ciarcia, Roberto; Ciriolo, Maria R.; Clague, Michael J.; Clark, Robert S.B.; Clarke, Peter G.H.; Clarke, Robert; Codogno, Patrice; Coller, Hilary A.; Colombo, María I.; Comincini, Sergio; Condello, Maria; Condorelli, Fabrizio; Cookson, Mark R.; Coombs, Graham H.; Coppens, Isabelle; Corbalan, Ramon; Cossart, Pascale; Costelli, Paola; Costes, Safia; Coto-Montes, Ana; Couve, Eduardo; Coxon, Fraser P.; Cregg, James M.; Crespo, José L.; Cronjé, Marianne J.; Cuervo, Ana Maria; Cullen, Joseph J.; Czaja, Mark J.; D'Amelio, Marcello; Darfeuille-Michaud, Arlette; Davids, Lester M.; Davies, Faith E.; De Felici, Massimo; de Groot, John F.; de Haan, Cornelis A.M.; De Martino, Luisa; De Milito, Angelo; De Tata, Vincenzo; Debnath, Jayanta; Degterev, Alexei; Dehay, Benjamin; Delbridge, Lea M.D.; Demarchi, Francesca; Deng, Yi Zhen; Dengjel, Jörn; Dent, Paul; Denton, Donna; Deretic, Vojo; Desai, Shyamal D.; Devenish, Rodney J.; Di Gioacchino, Mario; Di Paolo, Gilbert; Di Pietro, Chiara; Díaz-Araya, Guillermo; Díaz-Laviada, Inés; Diaz-Meco, Maria T.; Diaz-Nido, Javier; Dikic, Ivan; Dinesh-Kumar, Savithramma P.; Ding, Wen-Xing; Distelhorst, Clark W.; Diwan, Abhinav; Djavaheri-Mergny, Mojgan; Dokudovskaya, Svetlana; Dong, Zheng; Dorsey, Frank C.; Dosenko, Victor; Dowling, James J.; Doxsey, Stephen; Dreux, Marlène; Drew, Mark E.; Duan, Qiuhong; Duchosal, Michel A.; Duff, Karen E.; Dugail, Isabelle; Durbeej, Madeleine; Duszenko, Michael; Edelstein, Charles L.; Edinger, Aimee L.; Egea, Gustavo; Eichinger, Ludwig; Eissa, N. Tony; Ekmekcioglu, Suhendan; El-Deiry, Wafik S.; Elazar, Zvulun; Elgendy, Mohamed; Ellerby, Lisa M.; Eng, Kai Er; Engelbrecht, Anna-Mart; Engelender, Simone; Erenpreisa, Jekaterina; Escalante, Ricardo; Esclatine, Audrey; Eskelinen, Eeva-Liisa; Espert, Lucile; Espina, Virginia; Fan, Huizhou; Fan, Jia; Fan, Qi-Wen; Fan, Zhen; Fang, Shengyun; Fang, Yongqi; Fanto, Manolis; Fanzani, Alessandro; Farkas, Thomas; Farre, Jean-Claude; Faure, Mathias; Fechheimer, Marcus; Feng, Carl G.; Feng, Jian; Feng, Qili; Feng, Youji; Fésüs, László; Feuer, Ralph; Figueiredo-Pereira, Maria E.; Fimia, Gian Maria; Fingar, Diane C.; Finkbeiner, Steven; Finkel, Toren; Finley, Kim D.; Fiorito, Filomena; Fisher, Edward A.; Fisher, Paul B.; Flajolet, Marc; Florez-McClure, Maria L.; Florio, Salvatore; Fon, Edward A.; Fornai, Francesco; Fortunato, Franco; Fotedar, Rati; Fowler, Daniel H.; Fox, Howard S.; Franco, Rodrigo; Frankel, Lisa B.; Fransen, Marc; Fuentes, José M.; Fueyo, Juan; Fujii, Jun; Fujisaki, Kozo; Fujita, Eriko; Fukuda, Mitsunori; Furukawa, Ruth H.; Gaestel, Matthias; Gailly, Philippe; Gajewska, Malgorzata; Galliot, Brigitte; Galy, Vincent; Ganesh, Subramaniam; Ganetzky, Barry; Ganley, Ian G.; Gao, Fen-Biao; Gao, George F.; Gao, Jinming; Garcia, Lorena; Garcia-Manero, Guillermo; Garcia-Marcos, Mikel; Garmyn, Marjan; Gartel, Andrei L.; Gatti, Evelina; Gautel, Mathias; Gawriluk, Thomas R.; Gegg, Matthew E.; Geng, Jiefei; Germain, Marc; Gestwicki, Jason E.; Gewirtz, David A.; Ghavami, Saeid; Ghosh, Pradipta; Giammarioli, Anna M.; Giatromanolaki, Alexandra N.; Gibson, Spencer B.; Gilkerson, Robert W.; Ginger, Michael L.; Ginsberg, Henry N.; Golab, Jakub; Goligorsky, Michael S.; Golstein, Pierre; Gomez-Manzano, Candelaria; Goncu, Ebru; Gongora, Céline; Gonzalez, Claudio D.; Gonzalez, Ramon; González-Estévez, Cristina; González-Polo, Rosa Ana; Gonzalez-Rey, Elena; Gorbunov, Nikolai V.; Gorski, Sharon; Goruppi, Sandro; Gottlieb, Roberta A.; Gozuacik, Devrim; Granato, Giovanna Elvira; Grant, Gary D.; Green, Kim N.; Gregorc, Ales; Gros, Frédéric; Grose, Charles; Grunt, Thomas W.; Gual, Philippe; Guan, Jun-Lin; Guan, Kun-Liang; Guichard, Sylvie M.; Gukovskaya, Anna S.; Gukovsky, Ilya; Gunst, Jan; Gustafsson, Åsa B.; Halayko, Andrew J.; Hale, Amber N.; Halonen, Sandra K.; Hamasaki, Maho; Han, Feng; Han, Ting; Hancock, Michael K.; Hansen, Malene; Harada, Hisashi; Harada, Masaru; Hardt, Stefan E.; Harper, J. Wade; Harris, Adrian L.; Harris, James; Harris, Steven D.; Hashimoto, Makoto; Haspel, Jeffrey A.; Hayashi, Shin-ichiro; Hazelhurst, Lori A.; He, Congcong; He, You-Wen; Hébert, Marie-Josée; Heidenreich, Kim A.; Helfrich, Miep H.; Helgason, Gudmundur V.; Henske, Elizabeth P.; Herman, Brian; Herman, Paul K.; Hetz, Claudio; Hilfiker, Sabine; Hill, Joseph A.; Hocking, Lynne J.; Hofman, Paul; Hofmann, Thomas G.; Höhfeld, Jörg; Holyoake, Tessa L.; Hong, Ming-Huang; Hood, David A.; Hotamisligil, Gökhan S.; Houwerzijl, Ewout J.; Høyer-Hansen, Maria; Hu, Bingren; Hu, Chien-an A.; Hu, Hong-Ming; Hua, Ya; Huang, Canhua; Huang, Ju; Huang, Shengbing; Huang, Wei-Pang; Huber, Tobias B.; Huh, Won-Ki; Hung, Tai-Ho; Hupp, Ted R.; Hur, Gang Min; Hurley, James B.; Hussain, Sabah N.A.; Hussey, Patrick J.; Hwang, Jung Jin; Hwang, Seungmin; Ichihara, Atsuhiro; Ilkhanizadeh, Shirin; Inoki, Ken; Into, Takeshi; Iovane, Valentina; Iovanna, Juan L.; Ip, Nancy Y.; Isaka, Yoshitaka; Ishida, Hiroyuki; Isidoro, Ciro; Isobe, Ken-ichi; Iwasaki, Akiko; Izquierdo, Marta; Izumi, Yotaro; Jaakkola, Panu M.; Jäättelä, Marja; Jackson, George R.; Jackson, William T.; Janji, Bassam; Jendrach, Marina; Jeon, Ju-Hong; Jeung, Eui-Bae; Jiang, Hong; Jiang, Hongchi; Jiang, Jean X.; Jiang, Ming; Jiang, Qing; Jiang, Xuejun; Jiang, Xuejun; Jiménez, Alberto; Jin, Meiyan; Jin, Shengkan V.; Joe, Cheol O.; Johansen, Terje; Johnson, Daniel E.; Johnson, Gail V.W.; Jones, Nicola L.; Joseph, Bertrand; Joseph, Suresh K.; Joubert, Annie M.; Juhász, Gábor; Juillerat-Jeanneret, Lucienne; Jung, Chang Hwa; Jung, Yong-Keun; Kaarniranta, Kai; Kaasik, Allen; Kabuta, Tomohiro; Kadowaki, Motoni; Kågedal, Katarina; Kamada, Yoshiaki; Kaminskyy, Vitaliy O.; Kampinga, Harm H.; Kanamori, Hiromitsu; Kang, Chanhee; Kang, Khong Bee; Kang, Kwang Il; Kang, Rui; Kang, Yoon-A; Kanki, Tomotake; Kanneganti, Thirumala-Devi; Kanno, Haruo; Kanthasamy, Anumantha G.; Kanthasamy, Arthi; Karantza, Vassiliki; Kaushal, Gur P.; Kaushik, Susmita; Kawazoe, Yoshinori; Ke, Po-Yuan; Kehrl, John H.; Kelekar, Ameeta; Kerkhoff, Claus; Kessel, David H.; Khalil, Hany; Kiel, Jan A.K.W.; Kiger, Amy A.; Kihara, Akio; Kim, Deok Ryong; Kim, Do-Hyung; Kim, Dong-Hou; Kim, Eun-Kyoung; Kim, Hyung-Ryong; Kim, Jae-Sung; Kim, Jeong Hun; Kim, Jin Cheon; Kim, John K.; Kim, Peter K.; Kim, Seong Who; Kim, Yong-Sun; Kim, Yonghyun; Kimchi, Adi; Kimmelman, Alec C.; King, Jason S.; Kinsella, Timothy J.; Kirkin, Vladimir; Kirshenbaum, Lorrie A.; Kitamoto, Katsuhiko; Kitazato, Kaio; Klein, Ludger; Klimecki, Walter T.; Klucken, Jochen; Knecht, Erwin; Ko, Ben C.B.; Koch, Jan C.; Koga, Hiroshi; Koh, Jae-Young; Koh, Young Ho; Koike, Masato; Komatsu, Masaaki; Kominami, Eiki; Kong, Hee Jeong; Kong, Wei-Jia; Korolchuk, Viktor I.; Kotake, Yaichiro; Koukourakis, Michael I.; Flores, Juan B. Kouri; Kovács, Attila L.; Kraft, Claudine; Krainc, Dimitri; Krämer, Helmut; Kretz-Remy, Carole; Krichevsky, Anna M.; Kroemer, Guido; Krüger, Rejko; Krut, Oleg; Ktistakis, Nicholas T.; Kuan, Chia-Yi; Kucharczyk, Roza; Kumar, Ashok; Kumar, Raj; Kumar, Sharad; Kundu, Mondira; Kung, Hsing-Jien; Kurz, Tino; Kwon, Ho Jeong; La Spada, Albert R.; Lafont, Frank; Lamark, Trond; Landry, Jacques; Lane, Jon D.; Lapaquette, Pierre; Laporte, Jocelyn F.; László, Lajos; Lavandero, Sergio; Lavoie, Josée N.; Layfield, Robert; Lazo, Pedro A.; Le, Weidong; Le Cam, Laurent; Ledbetter, Daniel J.; Lee, Alvin J.X.; Lee, Byung-Wan; Lee, Gyun Min; Lee, Jongdae; lee, Ju-hyun; Lee, Michael; Lee, Myung-Shik; Lee, Sug Hyung; Leeuwenburgh, Christiaan; Legembre, Patrick; Legouis, Renaud; Lehmann, Michael; Lei, Huan-Yao; Lei, Qun-Ying; Leib, David A.; Leiro, José; Lemasters, John J.; Lemoine, Antoinette; Lesniak, Maciej S.; Lev, Dina; Levenson, Victor V.; Levine, Beth; Levy, Efrat; Li, Faqiang; Li, Jun-Lin; Li, Lian; Li, Sheng; Li, Weijie; Li, Xue-Jun; Li, Yan-Bo; Li, Yi-Ping; Liang, Chengyu; Liang, Qiangrong; Liao, Yung-Feng; Liberski, Pawel P.; Lieberman, Andrew; Lim, Hyunjung J.; Lim, Kah-Leong; Lim, Kyu; Lin, Chiou-Feng; Lin, Fu-Cheng; Lin, Jian; Lin, Jiandie D.; Lin, Kui; Lin, Wan-Wan; Lin, Weei-Chin; Lin, Yi-Ling; Linden, Rafael; Lingor, Paul; Lippincott-Schwartz, Jennifer; Lisanti, Michael P.; Liton, Paloma B.; Liu, Bo; Liu, Chun-Feng; Liu, Kaiyu; Liu, Leyuan; Liu, Qiong A.; Liu, Wei; Liu, Young-Chau; Liu, Yule; Lockshin, Richard A.; Lok, Chun-Nam; Lonial, Sagar; Loos, Benjamin; Lopez-Berestein, Gabriel; López-Otín, Carlos; Lossi, Laura; Lotze, Michael T.; Low, Peter; Lu, Binfeng; Lu, Bingwei; Lu, Bo; Lu, Zhen; Luciano, Fréderic; Lukacs, Nicholas W.; Lund, Anders H.; Lynch-Day, Melinda A.; Ma, Yong; Macian, Fernando; MacKeigan, Jeff P.; Macleod, Kay F.; Madeo, Frank; Maiuri, Luigi; Maiuri, Maria Chiara; Malagoli, Davide; Malicdan, May Christine V.; Malorni, Walter; Man, Na; Mandelkow, Eva-Maria; Manon, Stephen; Manov, Irena; Mao, Kai; Mao, Xiang; Mao, Zixu; Marambaud, Philippe; Marazziti, Daniela; Marcel, Yves L.; Marchbank, Katie; Marchetti, Piero; Marciniak, Stefan J.; Marcondes, Mateus; Mardi, Mohsen; Marfe, Gabriella; Mariño, Guillermo; Markaki, Maria; Marten, Mark R.; Martin, Seamus J.; Martinand-Mari, Camille; Martinet, Wim; Martinez-Vicente, Marta; Masini, Matilde; Matarrese, Paola; Matsuo, Saburo; Matteoni, Raffaele; Mayer, Andreas; Mazure, Nathalie M.; McConkey, David J.; McConnell, Melanie J.; McDermott, Catherine; McDonald, Christine; McInerney, Gerald M.; McKenna, Sharon L.; McLaughlin, BethAnn; McLean, Pamela J.; McMaster, Christopher R.; McQuibban, G. Angus; Meijer, Alfred J.; Meisler, Miriam H.; Meléndez, Alicia; Melia, Thomas J.; Melino, Gerry; Mena, Maria A.; Menendez, Javier A.; Menna-Barreto, Rubem F. S.; Menon, Manoj B.; Menzies, Fiona M.; Mercer, Carol A.; Merighi, Adalberto; Merry, Diane E.; Meschini, Stefania; Meyer, Christian G.; Meyer, Thomas F.; Miao, Chao-Yu; Miao, Jun-Ying; Michels, Paul A.M.; Michiels, Carine; Mijaljica, Dalibor; Milojkovic, Ana; Minucci, Saverio; Miracco, Clelia; Miranti, Cindy K.; Mitroulis, Ioannis; Miyazawa, Keisuke; Mizushima, Noboru; Mograbi, Baharia; Mohseni, Simin; Molero, Xavier; Mollereau, Bertrand; Mollinedo, Faustino; Momoi, Takashi; Monastyrska, Iryna; Monick, Martha M.; Monteiro, Mervyn J.; Moore, Michael N.; Mora, Rodrigo; Moreau, Kevin; Moreira, Paula I.; Moriyasu, Yuji; Moscat, Jorge; Mostowy, Serge; Mottram, Jeremy C.; Motyl, Tomasz; Moussa, Charbel E.-H.; Müller, Sylke; Muller, Sylviane; Münger, Karl; Münz, Christian; Murphy, Leon O.; Murphy, Maureen E.; Musarò, Antonio; Mysorekar, Indira; Nagata, Eiichiro; Nagata, Kazuhiro; Nahimana, Aimable; Nair, Usha; Nakagawa, Toshiyuki; Nakahira, Kiichi; Nakano, Hiroyasu; Nakatogawa, Hitoshi; Nanjundan, Meera; Naqvi, Naweed I.; Narendra, Derek P.; Narita, Masashi; Navarro, Miguel; Nawrocki, Steffan T.; Nazarko, Taras Y.; Nemchenko, Andriy; Netea, Mihai G.; Neufeld, Thomas P.; Ney, Paul A.; Nezis, Ioannis P.; Nguyen, Huu Phuc; Nie, Daotai; Nishino, Ichizo; Nislow, Corey; Nixon, Ralph A.; Noda, Takeshi; Noegel, Angelika A.; Nogalska, Anna; Noguchi, Satoru; Notterpek, Lucia; Novak, Ivana; Nozaki, Tomoyoshi; Nukina, Nobuyuki; Nürnberger, Thorsten; Nyfeler, Beat; Obara, Keisuke; Oberley, Terry D.; Oddo, Salvatore; Ogawa, Michinaga; Ohashi, Toya; Okamoto, Koji; Oleinick, Nancy L.; Oliver, F. Javier; Olsen, Laura J.; Olsson, Stefan; Opota, Onya; Osborne, Timothy F.; Ostrander, Gary K.; Otsu, Kinya; Ou, Jing-hsiung James; Ouimet, Mireille; Overholtzer, Michael; Ozpolat, Bulent; Paganetti, Paolo; Pagnini, Ugo; Pallet, Nicolas; Palmer, Glen E.; Palumbo, Camilla; Pan, Tianhong; Panaretakis, Theocharis; Pandey, Udai Bhan; Papackova, Zuzana; Papassideri, Issidora; Paris, Irmgard; Park, Junsoo; Park, Ohkmae K.; Parys, Jan B.; Parzych, Katherine R.; Patschan, Susann; Patterson, Cam; Pattingre, Sophie; Pawelek, John M.; Peng, Jianxin; Perlmutter, David H.; Perrotta, Ida; Perry, George; Pervaiz, Shazib; Peter, Matthias; Peters, Godefridus J.; Petersen, Morten; Petrovski, Goran; Phang, James M.; Piacentini, Mauro; Pierre, Philippe; Pierrefite-Carle, Valérie; Pierron, Gérard; Pinkas-Kramarski, Ronit; Piras, Antonio; Piri, Natik; Platanias, Leonidas C.; Pöggeler, Stefanie; Poirot, Marc; Poletti, Angelo; Poüs, Christian; Pozuelo-Rubio, Mercedes; Prætorius-Ibba, Mette; Prasad, Anil; Prescott, Mark; Priault, Muriel; Produit-Zengaffinen, Nathalie; Progulske-Fox, Ann; Proikas-Cezanne, Tassula; Przedborski, Serge; Przyklenk, Karin; Puertollano, Rosa; Puyal, Julien; Qian, Shu-Bing; Qin, Liang; Qin, Zheng-Hong; Quaggin, Susan E.; Raben, Nina; Rabinowich, Hannah; Rabkin, Simon W.; Rahman, Irfan; Rami, Abdelhaq; Ramm, Georg; Randall, Glenn; Randow, Felix; Rao, V. Ashutosh; Rathmell, Jeffrey C.; Ravikumar, Brinda; Ray, Swapan K.; Reed, Bruce H.; Reed, John C.; Reggiori, Fulvio; Régnier-Vigouroux, Anne; Reichert, Andreas S.; Reiners, John J.; Reiter, Russel J.; Ren, Jun; Revuelta, José L.; Rhodes, Christopher J.; Ritis, Konstantinos; Rizzo, Elizete; Robbins, Jeffrey; Roberge, Michel; Roca, Hernan; Roccheri, Maria C.; Rocchi, Stephane; Rodemann, H. 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    2012-01-01

    In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused

  10. Streptococcus pneumoniae induces autophagy through the inhibition of the PI3K-I/Akt/mTOR pathway and ROS hypergeneration in A549 cells.

    Directory of Open Access Journals (Sweden)

    Pu Li

    Full Text Available The present study focused on the action mechanism of S. pneumoniae (Sp in inducing autophagy in human alveolar epithelial cells. Sp, a gram-positive extracellular bacterium, activates autophagy with considerably increased microtuble-associated protein light chain 3 (LC3 punctation in A549 cells. The accumulation of typical autophagosomes and conjugation of LC3 to phosphatidylethanolamine were observed in Sp-infected cells as an indication of autophagy. Using the pneumolysin (PLY mutant, we successfully demonstrated that PLY is involved in initiating autophagy without affecting the expression levels of PI3K-III and Beclin1. PLY-mediated autophagy depends on the inhibition of the phosphoinositide 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR pathway. Furthermore, Sp could also lead to the reactive oxygen species (ROS hypergeneration in A549 cells. Taken together, Sp infection-induced autophagy is PLY-mediated through ROS hypergeneration and mTOR inhibition. PI3K-I and rapamycin (autophagy inducers enhanced bacterial clearance, whereas wortmannin (autophagy inhibitor and acetylcysteine (ROS inhibitor reduced intracellular bacteria clearance. Thus, Sp-induced autophagy represents a host-protective mechanism, providing new insight into the pathogenesis of respiratory tract Sp infection.

  11. Autophagy- An emerging target for melanoma therapy.

    Science.gov (United States)

    Ndoye, Abibatou; Weeraratna, Ashani T

    2016-01-01

    Melanoma accounts for only 5% of all cancers but is the leading cause of skin cancer death due to its high metastatic potential. Patients with metastatic melanoma have a 10-year survival rate of less than 10%. While the clinical landscape for melanoma is evolving rapidly, lack of response to therapies, as well as resistance to therapy remain critical obstacles for treatment of this disease. In recent years, a myriad of therapy resistance mechanisms have been unravelled, one of which is autophagy, the focus of this review. In advanced stages of malignancy, melanoma cells hijack the autophagy machinery in order to alleviate drug-induced and metabolic stress in the tumor microenvironment, thereby promoting resistance to multiple therapies, tumor cell survival, and progression.  Autophagy is an essential cellular process that maintains cellular homeostasis through the recycling of intracellular constituents. Early studies on the role of autophagy in cancer generated controversy as to whether autophagy was pro- or anti-tumorigenic. Currently, there is a consensus that autophagy is tumor-suppressive in the early stages of cancer and tumor-promoting in established tumors.  This review aims to highlight current understandings on the role of autophagy in melanoma malignancy, and specifically therapy resistance; as well as to evaluate recent strategies for therapeutic autophagy modulation. PMID:27583134

  12. Tumor Suppression and Promotion by Autophagy

    Directory of Open Access Journals (Sweden)

    Yenniffer Ávalos

    2014-01-01

    Full Text Available Autophagy is a highly regulated catabolic process that involves lysosomal degradation of proteins and organelles, mostly mitochondria, for the maintenance of cellular homeostasis and reduction of metabolic stress. Problems in the execution of this process are linked to different pathological conditions, such as neurodegeneration, aging, and cancer. Many of the proteins that regulate autophagy are either oncogenes or tumor suppressor proteins. Specifically, tumor suppressor genes that negatively regulate mTOR, such as PTEN, AMPK, LKB1, and TSC1/2 stimulate autophagy while, conversely, oncogenes that activate mTOR, such as class I PI3K, Ras, Rheb, and AKT, inhibit autophagy, suggesting that autophagy is a tumor suppressor mechanism. Consistent with this hypothesis, the inhibition of autophagy promotes oxidative stress, genomic instability, and tumorigenesis. Nevertheless, autophagy also functions as a cytoprotective mechanism under stress conditions, including hypoxia and nutrient starvation, that promotes tumor growth and resistance to chemotherapy in established tumors. Here, in this brief review, we will focus the discussion on this ambiguous role of autophagy in the development and progression of cancer.

  13. Hypothalamic corticotropin-releasing factor immunoreactivity is reduced during induction of pituitary tumors by chronic estrogen treatment

    International Nuclear Information System (INIS)

    The role that estrogen plays in the regulation of corticotropin-releasing factor (CRF) is not known. A radioimmunoassay specific for rat CRF was utilized to measure the CRF-like immunoreactivity (CRF-ir) in the hypothalamus of ovariectomized rats treated with estradiol for periods up to 12 weeks. Compared to ovariectomized controls, estradiol treatment resulted in significantly reduced CRF-ir after 3 and 12 weeks, although no significant change was seen after 8 weeks. Anterior pituitary (AP) weight was greatly increased by estradiol treatment at all time points studied. Bromocriptine treatment for the last 3 weeks of the 12-week period, or removal of estradiol for 3 weeks after 9 weeks of treatment did not reverse the changes in CRF-ir even though significant regressions of tumor size was achieved. There was no correlation between AP weight and CRF-ir in individual animals. These data show that chronic treatment with estrogen reduced hypothalamic CRF-ir content. Neither a direct estrogenic effect or an indirect effect mediated through alterations in the adenohypophysis could be ruled out. 21 references, 3 figures

  14. Stress management by autophagy: Implications for chemoresistance.

    Science.gov (United States)

    Huang, Zhao; Zhou, Li; Chen, Zhibin; Nice, Edouard C; Huang, Canhua

    2016-07-01

    Development of chemoresistance, which limits the efficiency of anticancer agents, has long been a major problem in cancer therapy and urgently needs to be solved to improve clinical outcomes. Factors contributing to chemoresistance are various, but a key factor is the cell's capability for stress management. Autophagy, a favored survival strategy that organisms employ to get over many kinds of stress, is emerging as a crucial player in drug resistance. It has been shown that autophagy facilitates the resistance of tumor cells to anticancer agents, and abrogation of autophagy could be therapeutically beneficial in some cases, suggesting autophagy could be a promising target for cancer treatments. Thus, defining the roles of autophagy in chemoresistance, and the mechanisms involved, will be critical to enhance the efficiency of chemotherapy and develop novel anticancer strategy interventions. PMID:26757106

  15. Autophagy in the pathogenesis of ankylosing spondylitis.

    Science.gov (United States)

    Ciccia, Francesco; Haroon, Nigil

    2016-06-01

    The pathogenesis of ankylosing spondylitis (AS) is not well understood, and treatment options have met with limited success. Autophagy is a highly conserved mechanism of controlled digestion of damaged organelles within a cell. It helps in the maintenance of cellular homeostasis. The process of autophagy requires the formation of an isolation membrane. They form double-membraned vesicles called "autophagosomes" that engulf a portion of the cytoplasm. Beyond the role in maintenance of cellular homeostasis, autophagy has been demonstrated as one of the most remarkable tools employed by the host cellular defense against bacteria invasion. Autophagy also affects the immune system and thus is implicated in several rheumatic disease processes. In this article, we explore the potential role of autophagy in the pathogenesis of AS. PMID:27075464

  16. Autophagy and the immune function in aging.

    Science.gov (United States)

    Cuervo, Ana Maria; Macian, Fernando

    2014-08-01

    Just when you thought that you had heard it all about autophagy-the conserved cellular process that mediates turnover of cellular constituents in the lysosomes - studies keep coming out highlighting new types of autophagy, new functions for autophagy or even new autophagy-independent roles for the proteins associated with this process. The field of immunology has been riding the autophagic wave since the beginning of its revival; first due to its role in the host defense against pathogens, and more recently through the better understanding of the unique characteristics and functions of different autophagic pathways in immune cells. Here, we describe some of these new functions that are tightening the connection between autophagy and acquired or innate immunity and their malfunctioning with age. PMID:24929664

  17. Induction of ischemic tolerance in rat liver via reduced nicotinamide adenine dinucleotide phosphate oxidase in Kupffer cells

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    AIM: To elucidate the mechanisms of hepatocyte preconditioning by H2O2 to better understand the pathophysiology of ischemic preconditioning.METHODS: The in vitro effect of H2O2 pretreatment was investigated in rat isolated hepatocytes subjected to anoxia/reoxygenation. Cell viability was assessed with propidium iodide fluorometry. In other experiments, rat livers were excised and subjected to warm ischemia/reperfusion in an isolated perfused liver system to determine leakage of liver enzymes. Preconditioning was performed by H2O2 perfusion, or by stopping the perfusion for 10 min followed by 10 min of reperfusion.To inhibit Kupffer cell function or reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase,gadolinium chloride was injected prior to liver excision, or diphenyleneiodonium, an inhibitor of NADPH oxidase, was added to the perfusate, respectively. Histological detection of o~gen radical formation in Kupffer cells was performed by perfusion with nitro blue tetrazolium.RESULTS: Anoxia/reoxygenation decreased hepatocyte viability compared to the controls. Pretreatment with H2O2 did not improve such hepatocyte injury. In liver perfusion experiments, however, H2O2 preconditioning reduced warm ischemia/reperfusion injury, which was reversed by inhibition of Kupffer cell function or NADPH oxidase. Histological examination revealed that H2O2 preconditioning induced oxygen radical formation in Kupffer cells. NADPH oxidase inhibition also reversed hepatoprotection by ischemic preconditioning.CONCLUSION: H2O2 preconditioning protects hepatocytes against warm ischemia/reperfusion injury via NADPH oxidase in Kupffer cells, and not directly. NADPH oxidase also mediates hepatoprotection by ischemic preconditioning.

  18. Oxidative Stress and Autophagy in the Regulation of Lysosome-Dependent Neuron Death

    OpenAIRE

    Pivtoraiko, Violetta N.; Stone, Sara L; Roth, Kevin A.; Shacka, John J

    2009-01-01

    Lysosomes critically regulate the pH-dependent catabolism of extracellular and intracellular macromolecules delivered from the endocytic/heterophagy and autophagy pathways, respectively. The importance of lysosomes to cell survival is underscored not only by their unique ability effectively to degrade metalloproteins and oxidatively damaged macromolecules, but also by the distinct potential for induction of both caspase-dependent and -independent cell death with a compromise in the integrity ...

  19. NOX4 mediates cytoprotective autophagy induced by the EGFR inhibitor erlotinib in head and neck cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Sobhakumari, Arya [Interdisciplinary Graduate Program in Human Toxicology, The University of Iowa, Iowa City, IA (United States); Department of Pathology, The University of Iowa, Iowa City, IA (United States); Schickling, Brandon M. [Department of Internal Medicine, The University of Iowa, Iowa City, IA (United States); Love-Homan, Laurie; Raeburn, Ayanna [Department of Pathology, The University of Iowa, Iowa City, IA (United States); Fletcher, Elise V.M. [Interdisciplinary Graduate Program in Human Toxicology, The University of Iowa, Iowa City, IA (United States); Department of Pathology, The University of Iowa, Iowa City, IA (United States); Case, Adam J. [Free Radical and Radiation Biology Program, Department of Radiation Oncology, The University of Iowa, Iowa City, IA (United States); Domann, Frederick E. [Interdisciplinary Graduate Program in Human Toxicology, The University of Iowa, Iowa City, IA (United States); Department of Pathology, The University of Iowa, Iowa City, IA (United States); Free Radical and Radiation Biology Program, Department of Radiation Oncology, The University of Iowa, Iowa City, IA (United States); Holden Comprehensive Cancer Center, University of Iowa Hospitals and Clinics (UIHC), Iowa City, IA (United States); Miller, Francis J. [Department of Internal Medicine, The University of Iowa, Iowa City, IA (United States); Free Radical and Radiation Biology Program, Department of Radiation Oncology, The University of Iowa, Iowa City, IA (United States); Holden Comprehensive Cancer Center, University of Iowa Hospitals and Clinics (UIHC), Iowa City, IA (United States); and others

    2013-11-01

    Most head and neck squamous cell carcinomas (HNSCCs) overexpress epidermal growth factor receptor (EGFR) and EGFR inhibitors are routinely used in the treatment of HNSCC. However, many HNSCC tumors do not respond or become refractory to EGFR inhibitors. Autophagy, which is a stress-induced cellular self-degradation process, has been reported to reduce the efficacy of chemotherapy in various disease models. The purpose of this study is to determine if the efficacy of the EGFR inhibitor erlotinib is reduced by activation of autophagy via NOX4-mediated oxidative stress in HNSCC cells. Erlotinib induced the expression of the autophagy marker LC3B-II and autophagosome formation in FaDu and Cal-27 cells. Inhibition of autophagy by chloroquine and knockdown of autophagy pathway genes Beclin-1 and Atg5 sensitized both cell lines to erlotinib-induced cytotoxicity, suggesting that autophagy may serve as a protective mechanism. Treatment with catalase (CAT) and diphenylene iodonium (DPI) in the presence of erlotinib suppressed the increase in LC3B-II expression in FaDu and Cal-27 cells. Erlotinib increased NOX4 mRNA and protein expression by increasing its promoter activity and mRNA stability in FaDu cells. Knockdown of NOX4 using adenoviral siNOX4 partially suppressed erlotinib-induced LC3B-II expression, while overexpression of NOX4 increased expression of LC3B-II. These studies suggest that erlotinib may activate autophagy in HNSCC cells as a pro-survival mechanism, and NOX4 may play a role in mediating this effect. - Highlights: • Erlotinib increased LC3B-II and autophagosome formation in HNSCC cells. • Inhibition of autophagy sensitized HNSCC cells to erlotinib. • Erlotinib increased NOX4 promoter and 3′UTR luciferase activity. • Manipulating NOX4 decreases or increases autophagy.

  20. NOX4 mediates cytoprotective autophagy induced by the EGFR inhibitor erlotinib in head and neck cancer cells

    International Nuclear Information System (INIS)

    Most head and neck squamous cell carcinomas (HNSCCs) overexpress epidermal growth factor receptor (EGFR) and EGFR inhibitors are routinely used in the treatment of HNSCC. However, many HNSCC tumors do not respond or become refractory to EGFR inhibitors. Autophagy, which is a stress-induced cellular self-degradation process, has been reported to reduce the efficacy of chemotherapy in various disease models. The purpose of this study is to determine if the efficacy of the EGFR inhibitor erlotinib is reduced by activation of autophagy via NOX4-mediated oxidative stress in HNSCC cells. Erlotinib induced the expression of the autophagy marker LC3B-II and autophagosome formation in FaDu and Cal-27 cells. Inhibition of autophagy by chloroquine and knockdown of autophagy pathway genes Beclin-1 and Atg5 sensitized both cell lines to erlotinib-induced cytotoxicity, suggesting that autophagy may serve as a protective mechanism. Treatment with catalase (CAT) and diphenylene iodonium (DPI) in the presence of erlotinib suppressed the increase in LC3B-II expression in FaDu and Cal-27 cells. Erlotinib increased NOX4 mRNA and protein expression by increasing its promoter activity and mRNA stability in FaDu cells. Knockdown of NOX4 using adenoviral siNOX4 partially suppressed erlotinib-induced LC3B-II expression, while overexpression of NOX4 increased expression of LC3B-II. These studies suggest that erlotinib may activate autophagy in HNSCC cells as a pro-survival mechanism, and NOX4 may play a role in mediating this effect. - Highlights: • Erlotinib increased LC3B-II and autophagosome formation in HNSCC cells. • Inhibition of autophagy sensitized HNSCC cells to erlotinib. • Erlotinib increased NOX4 promoter and 3′UTR luciferase activity. • Manipulating NOX4 decreases or increases autophagy

  1. Reduced secondary cytokine induction by BAY 50-4798, a high-affinity receptor-specific interleukin-2 analog.

    Science.gov (United States)

    Steppan, Sonja; Eckart, Michael R; Bajsarowicz, Krystyna; Sternberg, Lawrence R; Greve, Jeffrey M; Cassell, Delanie J

    2006-03-01

    Recombinant interleukin-2 (IL-2) (aldesleukin, Proleukin, Chiron, Emeryville, CA) is approved for treatment of cancer patients and under investigation in HIV-infected individuals. However, treatment with aldesleukin is associated with toxicity, which may be due to its elicitation of inflammatory mediators from cells that express the intermediate-affinity IL-2 receptor. BAY 50-4798, a novel IL-2 analog, is a selective agonist for the high-affinity receptor. It induces the proliferation of activated T cells with a potency similar to that of aldesleukin but has reduced activity on cells expressing the intermediate-affinity receptor. In the current study, we compared cytokine responses elicited in peripheral blood mononuclear cell (PBMC) cultures stimulated with BAY 50-4798 or aldesleukin. BAY 50-4798 induced approximately 5-fold lower mean levels of endogenous IL-2 than aldesleukin, and at least 50% lower levels of proinflammatory cytokines, such as tumor necrosis fctor-alpha (TNF-alpha), IL-1beta, IL-6, and interferon-gamma (IFN-gamma). Furthermore, statistically significant reductions in the levels of IL-5, IL-8, IL-10, IL-13, and granulocyte-macrophage colony-stimulating factor (GM-CSF) were observed in response to BAY 50-4798. These findings increase our understanding of the biologic action of BAY 50-4798 and suggest a mechanism by which it may exhibit better safety than aldesleukin in humans. PMID:16542139

  2. Induction of p38 mitogen-activated protein kinase reduces early endosome autoantigen 1 (EEA1) recruitment to phagosomal membranes.

    Science.gov (United States)

    Fratti, Rutilio A; Chua, Jennifer; Deretic, Vojo

    2003-11-21

    Mycobacterium tuberculosis survives in the infected host by parasitizing macrophages in which the bacillus resides in a specialized phagosome sequestered from the phagolysosomal degradative pathway. Here we report a role of the stress-induced p38 mitogen-activated protein kinase (p38 MAPK) in the component of M. tuberculosis phagosome maturation arrest that has been linked previously to the reduced recruitment of the endosomal and phagosomal membrane-tethering molecule called early endosome autoantigen 1 (EEA1; Fratti, R. A., Backer, J. M., Gruenberg, J., Corvera, S., and Deretic, V. (2001) J. Cell Biol. 154, 631-644). A pharmacological inhibition of M. tuberculosis var. bovis Bacillus Calmette-Guérin-induced p38 MAPK activity caused a marked increase in EEA1 colocalization with mycobacterial phagosomes. Consistent with the increase in EEA1 association and its role in phagosomal maturation, the pharmacological block of p38 activity caused phagosomal acidification and enrichment of the late endocytic markers lysobisphosphatidic acid and CD63 (lysosomal integral membrane protein 1) on mycobacterial phagosomes. A negative regulatory role of p38 MAPK activation in phagosome maturation was further demonstrated by converse experiments with latex bead phagosomes. Artificial activation of p38 MAPK caused a decrease in EEA1 colocalization with model latex bead phagosomes, which normally acquire EEA1 and subsequently mature into the phagolysosome. These findings show that p38 MAPK activity contributes to the arrest of M. tuberculosis phagosome maturation and demonstrate a negative regulatory role of p38 in phagolysosome biogenesis. PMID:12963735

  3. The benefits of whole-house in-duct air cleaning in reducing exposures to fine particulate matter of outdoor origin: a modeling analysis.

    Science.gov (United States)

    Macintosh, David L; Minegishi, Taeko; Kaufman, Matthew; Baker, Brian J; Allen, Joseph G; Levy, Jonathan I; Myatt, Theodore A

    2010-03-01

    Health risks of fine particle air pollution (PM(2.5)) are an important public health concern that has the potential to be mitigated in part by interventions such as air cleaning devices that reduce personal exposure to ambient PM(2.5). To characterize exposure to ambient PM(2.5) indoors as a function of residential air cleaners, a multi-zone indoor air quality model was used to integrate spatially resolved data on housing, meteorology, and ambient PM(2.5), with performance testing of residential air cleaners to estimate short-term and annual average PM(2.5) of outdoor origin inside residences of three metropolitan areas. The associated public health impacts of reduced ambient PM(2.5) exposure were estimated using a standard health impact assessment methodology. Estimated indoor levels of ambient PM(2.5) varied substantially among ventilation and air cleaning configurations. The median 24-h average indoor-outdoor ratio of ambient PM(2.5) was 0.57 for homes with natural ventilation, 0.35 for homes with central air conditioning (AC) with conventional filtration, and 0.1 for homes with central AC with high efficiency in-duct air cleaner. Median modeled 24-h average indoor concentrations of PM(2.5) of outdoor origin for those three configurations were 8.4, 5.3, and 1.5 microg/m(3), respectively. The potential public health benefits of reduced exposure to ambient PM(2.5) afforded by air cleaning systems were substantial. If the entire population of single-family homes with central AC in the modeling domain converted from conventional filtration to high-efficiency in-duct air cleaning, the change in ambient PM(2.5) exposure is estimated to result in an annual reduction of 700 premature deaths, 940 hospital and emergency room visits, and 130,000 asthma attacks in these metropolitan areas. In addition to controlling emissions from sources, high-efficiency whole-house air cleaner are expected to reduce exposure to particles of outdoor origin and are projected to be an

  4. The multifunctional autophagy pathway in the human malaria parasite, Plasmodium falciparum

    Science.gov (United States)

    Cervantes, Serena; Bunnik, Evelien M; Saraf, Anita; Conner, Christopher M; Escalante, Aster; Sardiu, Mihaela E; Ponts, Nadia; Prudhomme, Jacques; Florens, Laurence; Le Roch, Karine G

    2014-01-01

    Autophagy is a catabolic pathway typically induced by nutrient starvation to recycle amino acids, but can also function in removing damaged organelles. In addition, this pathway plays a key role in eukaryotic development. To date, not much is known about the role of autophagy in apicomplexan parasites and more specifically in the human malaria parasite Plasmodium falciparum. Comparative genomic analysis has uncovered some, but not all, orthologs of autophagy-related (ATG) genes in the malaria parasite genome. Here, using a genome-wide in silico analysis, we confirmed that ATG genes whose products are required for vesicle expansion and completion are present, while genes involved in induction of autophagy and cargo packaging are mostly absent. We subsequently focused on the molecular and cellular function of P. falciparum ATG8 (PfATG8), an autophagosome membrane marker and key component of the autophagy pathway, throughout the parasite asexual and sexual erythrocytic stages. In this context, we showed that PfATG8 has a distinct and atypical role in parasite development. PfATG8 localized in the apicoplast and in vesicles throughout the cytosol during parasite development. Immunofluorescence assays of PfATG8 in apicoplast-minus parasites suggest that PfATG8 is involved in apicoplast biogenesis. Furthermore, treatment of parasite cultures with bafilomycin A1 and chloroquine, both lysosomotropic agents that inhibit autophagosome and lysosome fusion, resulted in dramatic morphological changes of the apicoplast, and parasite death. Furthermore, deep proteomic analysis of components associated with PfATG8 indicated that it may possibly be involved in ribophagy and piecemeal microautophagy of the nucleus. Collectively, our data revealed the importance and specificity of the autophagy pathway in the malaria parasite and offer potential novel therapeutic strategies. PMID:24275162

  5. Role and Regulation of Autophagy in Heat Stress Responses of Tomato Plants

    Directory of Open Access Journals (Sweden)

    Jie eZhou

    2014-04-01

    Full Text Available As sessile organisms, plants are constantly exposed to a wide spectrum of stress conditions such as high temperature, which causes protein misfolding. Misfolded proteins are highly toxic and must be efficiently removed to reduce cellular proteotoxic stress if restoration of native conformations is unsuccessful. Although selective autophagy is known to function in protein quality control by targeting degradation of misfolded and potentially toxic proteins, its role and regulation in heat stress responses have not been analyzed in crop plants. In the present study, we found that heat stress induced expression of autophagy-related (ATG genes and accumulation of autophagosomes in tomato plants. Virus-induced gene silencing of tomato ATG5 and ATG7 genes resulted in increased sensitivity of tomato plants to heat stress based on both increased development of heat stress symptoms and compromised photosynthetic parameters of heat-stressed leaf tissues. Silencing of tomato homologs for the selective autophagy receptor NBR1, which targets ubiquitinated protein aggregates, also compromised tomato heat tolerance. To better understand the regulation of heat-induced autophagy, we found that silencing of tomato ATG5, ATG7 or NBR1 compromised heat-induced expression of not only the targeted genes but also other autophagy-related genes. Furthermore, we identified two tomato genes encoding proteins highly homologous to Arabidopsis WRKY33 transcription factor, which has been previously shown to interact physically with an autophagy protein. Silencing of tomato WRKY33 genes compromised tomato heat tolerance and reduced heat-induced ATG gene expression and autophagosome accumulation. Based on these results, we propose that heat-induced autophagy in tomato is subject to cooperative regulation by both WRKY33 and ATG proteins and plays a critical role in tomato heat tolerance, mostly likely through selective removal of heat-induced protein aggregates.

  6. Autophagy and bacterial clearance: a not so clear picture

    OpenAIRE

    Mostowy, Serge

    2012-01-01

    Autophagy, an intracellular degradation process highly conserved from yeast to humans, is viewed as an important defence mechanism to clear intracellular bacteria. However, recent work has shown that autophagy may have different roles during different bacterial infections that restrict bacterial replication (antibacterial autophagy), act in cell autonomous signalling (non-bacterial autophagy) or support bacterial replication (pro-bacterial autophagy). This review will focus on newfound intera...

  7. Newly synthesized bis-benzimidazole compound 8 induces apoptosis, autophagy and reactive oxygen species generation in HeLa cells.

    Science.gov (United States)

    Chu, Naying; Yao, Guodong; Liu, Yuan; Cheng, Maosheng; Ikejima, Takashi

    2016-09-01

    Compound 8 (C8) is a newly synthesized bis-benzimidazole derivative and exerts significant anti-tumor activity in vitro. Previous studies demonstrated that C8 induced apoptosis and autophagy in human promyelocytic leukemia HL60 cells. However, cytotoxicity study on human peripheral blood mononuclear cells (hPBMC) showed that C8 exhibited less toxicity in normal cells. In this study, the molecular mechanism of C8 on human cervical carcinoma HeLa cells was investigated. The results showed that C8 inhibited the growth of HeLa cells and triggered both apoptotic and autophagic cell death. Subsequent experiment also indicated that reactive oxygen species (ROS) generation was induced in C8-treated HeLa cells. Since ROS scavenger decreased the ratio of apoptotic and autophagic cells, ROS generation contributed to C8-induced apoptosis and autophagy. Furthermore, inhibitors of apoptosis and autophagy also reduced ROS generation, respectively. Autophagy inhibition increased cell growth compared to C8-treated group and attenuated apoptotic cell death, indicating that C8-induced autophagy promoted apoptosis for cell death. However, the percentage of autophagic cells was enhanced when limiting apoptosis process. Taken together, C8 induced ROS-mediated apoptosis and autophagy in HeLa cells, autophagy promoted apoptosis but the former was antagonized by the latter. The data also gave us a new perspective on the anti-tumor effect of C8. PMID:27497983

  8. METACASPASE9 modulates autophagy to confine cell death to the target cells during Arabidopsis vascular xylem differentiation

    Directory of Open Access Journals (Sweden)

    Sacha Escamez

    2016-02-01

    Full Text Available We uncovered that the level of autophagy in plant cells undergoing programmed cell death determines the fate of the surrounding cells. Our approach consisted of using Arabidopsis thaliana cell cultures capable of differentiating into two different cell types: vascular tracheary elements (TEs that undergo programmed cell death (PCD and protoplast autolysis, and parenchymatic non-TEs that remain alive. The TE cell type displayed higher levels of autophagy when expression of the TE-specific METACASPASE9 (MC9 was reduced using RNAi (MC9-RNAi. Misregulation of autophagy in the MC9-RNAi TEs coincided with ectopic death of the non-TEs, implying the existence of an autophagy-dependent intercellular signalling from within the TEs towards the non-TEs. Viability of the non-TEs was restored when AUTOPHAGY2 (ATG2 was downregulated specifically in MC9-RNAi TEs, demonstrating the importance of autophagy in the spatial confinement of cell death. Our results suggest that other eukaryotic cells undergoing PCD might also need to tightly regulate their level of autophagy to avoid detrimental consequences for the surrounding cells.

  9. Arsenite-induced autophagy is associated with proteotoxicity in human lymphoblastoid cells

    International Nuclear Information System (INIS)

    Epidemiological studies of arsenic-exposed populations have provided evidence that arsenic exposure in humans is associated with immunosuppression. Previously, we have reported that arsenite-induced toxicity is associated with the induction of autophagy in human lymphoblastoid cell lines (LCL). Autophagy is a cellular process that functions in the degradation of damaged cellular components, including protein aggregates formed by misfolded or damaged proteins. Accumulation of misfolded or damaged proteins in the endoplasmic reticulum (ER) lumen causes ER stress and activates the unfolded protein response (UPR). In an effort to investigate the mechanism of autophagy induction by arsenite in the LCL model, we examined the potential contribution of ER stress and activation of the UPR. LCL exposed to sodium arsenite for 8-days induced expression of UPR-activated genes, including CHOP and GRP78, at the RNA and the protein level. Evidence for activation of the three arms of the UPR was observed. The arsenite-induced activation of the UPR was associated with an accumulation of protein aggregates containing p62 and LC3, proteins with established roles in the sequestration and autophagic clearance of protein aggregates. Taken together, these data provide evidence that arsenite-induced autophagy is associated with the generation of ER stress, activation of the UPR, and formation of protein aggregates that may be targeted to the lysosome for degradation. -- Highlights: ► Arsenite induces endoplasmic reticulum stress and the unfolded protein response. ► Arsenite induces the formation of protein aggregates that contain p62 and LC3-II. ► Time-course data suggests that arsenite-induced autophagy precedes ER stress.

  10. Arsenite-induced autophagy is associated with proteotoxicity in human lymphoblastoid cells

    Energy Technology Data Exchange (ETDEWEB)

    Bolt, Alicia M.; Zhao, Fei; Pacheco, Samantha; Klimecki, Walter T., E-mail: klimecki@pharmacy.arizona.edu

    2012-10-15

    Epidemiological studies of arsenic-exposed populations have provided evidence that arsenic exposure in humans is associated with immunosuppression. Previously, we have reported that arsenite-induced toxicity is associated with the induction of autophagy in human lymphoblastoid cell lines (LCL). Autophagy is a cellular process that functions in the degradation of damaged cellular components, including protein aggregates formed by misfolded or damaged proteins. Accumulation of misfolded or damaged proteins in the endoplasmic reticulum (ER) lumen causes ER stress and activates the unfolded protein response (UPR). In an effort to investigate the mechanism of autophagy induction by arsenite in the LCL model, we examined the potential contribution of ER stress and activation of the UPR. LCL exposed to sodium arsenite for 8-days induced expression of UPR-activated genes, including CHOP and GRP78, at the RNA and the protein level. Evidence for activation of the three arms of the UPR was observed. The arsenite-induced activation of the UPR was associated with an accumulation of protein aggregates containing p62 and LC3, proteins with established roles in the sequestration and autophagic clearance of protein aggregates. Taken together, these data provide evidence that arsenite-induced autophagy is associated with the generation of ER stress, activation of the UPR, and formation of protein aggregates that may be targeted to the lysosome for degradation. -- Highlights: ► Arsenite induces endoplasmic reticulum stress and the unfolded protein response. ► Arsenite induces the formation of protein aggregates that contain p62 and LC3-II. ► Time-course data suggests that arsenite-induced autophagy precedes ER stress.

  11. Naringin induces autophagy-mediated growth inhibition by downregulating the PI3K/Akt/mTOR cascade via activation of MAPK pathways in AGS cancer cells.

    Science.gov (United States)

    Raha, Suchismita; Yumnam, Silvia; Hong, Gyeong Eun; Lee, Ho Jeong; Saralamma, Venu Venkatarame Gowda; Park, Hyeon-Soo; Heo, Jeong Doo; Lee, Sang Joon; Kim, Eun Hee; Kim, Jin-A; Kim, Gon Sup

    2015-09-01

    Naringin, one of the major bioflavonoid of Citrus, has been demonstrated as potential anticancer agent. However, the underlying anticancer mechanism still needs to be explored further. This study investigated the inhibitory effect of Naringin on human AGS cancer cells. AGS cell proliferation was inhibited by Naringin in a dose- and time-dependent manner. Naringin did not induce apoptotic cell death, determined by no DNA fragmentation and the reduced Bax/Bcl-xL ratio. Growth inhibitory role of Naringin was observed by western blot analysis demonstrating downregulation of PI3K/Akt/mTOR cascade with an upregulated p21CIPI/WAFI. Formation of cytoplasmic vacuoles and autophagosomes were observed in Naringin-treated AGS cells, further confirmed by the activation of autophagic proteins Beclin 1 and LC3B with a significant phosphorylation of mitogen activated protein kinases (MAPKs). Collectively, our observed results determined that anti-proliferative activity of Naringin in AGS cancer cells is due to suppression of PI3K/Akt/mTOR cascade via induction of autophagy with activated MAPKs. Thus, the present finding suggests that Naringin induced autophagy- mediated growth inhibition shows potential as an alternative therapeutic agent for human gastric carcinoma. PMID:26201693

  12. Probiotic Lactobacillus rhamnosus GG mono-association suppresses human rotavirus-induced autophagy in the gnotobiotic piglet intestine

    OpenAIRE

    Wu, Shaoping; Yuan, Lijuan; Zhang, Yongguo; Liu, Fangning; Li, Guohua; Wen, Ke; Kocher, Jacob; Yang, Xingdong; Sun, Jun

    2013-01-01

    Abstract Background Human rotavirus (HRV) is the most important cause of severe diarrhea in infants and young children. Probiotic Lactobacillus rhamnosus GG (LGG) reduces rotavirus infection and diarrhea. However, the molecular mechanisms of LGG-mediated protection from rotavirus infection are poorly understood. Autophagy plays an essential role in responses to microbial pathogens. However, the role of autophagy in HRV infection and LGG treatment is unknown. We hypothesize that rotavirus gast...

  13. Inhibition of HIF-1α Affects Autophagy Mediated Glycosylation in Oral Squamous Cell Carcinoma Cells

    Directory of Open Access Journals (Sweden)

    Yi-Ning Li

    2015-01-01

    Full Text Available Purpose. To validate the function of autophagy with the regulation of hypoxia inhibitor-induced glycosylation in oral squamous cell carcinoma cell. Methods. Human Tca8113 cell line was used to detect autophagy and glycosylation related protein expression by western blotting and immunofluorescence with HIF-1α inhibitor. Short interfering RNA (siRNA transfection blocked human ATG12 and ATG1. Results. HIF-1α inhibitor PX-478 reduced the amount of LC3-II and LC3-I in Tca8113 cells. PX-478 decreased the expression of O-GlcNAc and OGT and increased OGA expression. The tendency of O-GlcNAc showed a similar pattern to OGT. PX-478 gradually decreased OGT expression in Tca8113 cells. Protein level of O-GlcNAc and OGT increased in ATG12 and ATG1 depletion. The expression of OGT decreased at first and then rose slowly with the treatment of Atg12 and Atg1 siRNA and PX-478 fluctuant. Autophagy affected the stability of OGT when HIF-1α signaling was blocked. Conclusions. Autophagy reduced by hypoxic stress inhibited. HIF-1α inhibitor decreased glycosylation. OGT became unstable in the absence of autophagy when HIF-1α signaling was blocked.

  14. Autophagy-assisted glycogen catabolism regulates asexual differentiation in Magnaporthe oryzae.

    Science.gov (United States)

    Deng, Yi Zhen; Ramos-Pamplona, Marilou; Naqvi, Naweed I

    2009-01-01

    Autophagy, a conserved pathway for bulk cellular degradation and recycling in eukaryotes, regulates proper turnover of organelles, membranes and certain proteins. Such regulated degradation is important for cell growth and development particularly during environmental stress conditions, which act as key inducers of autophagy. We found that autophagy and MoATG8 were significantly induced during asexual development in Magnaporthe oryzae. An RFP-tagged MoAtg8 showed specific localization and enrichment in aerial hyphae, conidiophores and conidia. We confirmed that loss of MoATG8 results in dramatically reduced ability to form conidia, the asexual spores that propagate rice-blast disease. Exogenous supply of glucose or sucrose significantly suppressed the conidiation defects in a MoATG8-deletion mutant. Comparative proteomics based identification and characterization of Gph1, a glycogen phosphorylase that catalyzes glycogen breakdown, indicated that autophagy-assisted glycogen homeostasis is likely important for proper aerial growth and conidiation in Magnaporthe. Loss of Gph1, or addition of G6P significantly restored conidiation in the Moatg8Delta mutant. Overproduction of Gph1 led to reduced conidiation in wild-type Magnaporthe strain. We propose that glycogen autophagy actively responds to and regulates carbon utilization required for cell growth and differentiation during asexual development in Magnaporthe. PMID:19115483

  15. Autophagy in human embryonic stem cells.

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    Thien Tra

    Full Text Available Autophagy (macroautophagy is a degradative process that involves the sequestration of cytosolic material including organelles into double membrane vesicles termed autophagosomes for delivery to the lysosome. Autophagy is essential for preimplantation development of mouse embryos and cavitation of embryoid bodies. The precise roles of autophagy during early human embryonic development, remain however largely uncharacterized. Since human embryonic stem cells constitute a unique model system to study early human embryogenesis we investigated the occurrence of autophagy in human embryonic stem cells. We have, using lentiviral transduction, established multiple human embryonic stem cell lines that stably express GFP-LC3, a fluorescent marker for the autophagosome. Each cell line displays both a normal karyotype and pluripotency as indicated by the presence of cell types representative of the three germlayers in derived teratomas. GFP expression and labelling of autophagosomes is retained after differentiation. Baseline levels of autophagy detected in cultured undifferentiated hESC were increased or decreased in the presence of rapamycin and wortmannin, respectively. Interestingly, autophagy was upregulated in hESCs induced to undergo differentiation by treatment with type I TGF-beta receptor inhibitor SB431542 or removal of MEF secreted maintenance factors. In conclusion we have established hESCs capable of reporting macroautophagy and identify a novel link between autophagy and early differentiation events in hESC.

  16. Roles of autophagy in MPP+-induced neurotoxicity in vivo: the involvement of mitochondria and α-synuclein aggregation.

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    Kai-Chih Hung

    Full Text Available Macroautophagy (also known as autophagy is an intracellular self-eating mechanism and has been proposed as both neuroprotective and neurodestructive in the central nervous system (CNS neurodegenerative diseases. In the present study, the role of autophagy involving mitochondria and α-synuclein was investigated in MPP+ (1-methyl-4-phenylpyridinium-induced oxidative injury in chloral hydrate-anesthetized rats in vivo. The oxidative mechanism underlying MPP+-induced neurotoxicity was identified by elevated lipid peroxidation and heme oxygenase-1 levels, a redox-regulated protein in MPP+-infused substantia nigra (SN. At the same time, MPP+ significantly increased LC3-II levels, a hallmark protein of autophagy. To block MPP+-induced autophagy in rat brain, Atg7siRNA was intranigrally infused 4 d prior to MPP+ infusion. Western blot assay showed that in vivo Atg7siRNA transfection not only reduced Atg7 levels in the MPP+-infused SN but attenuated MPP+-induced elevation in LC3-II levels, activation of caspase 9 and reduction in tyrosine hydroxylase levels, indicating that autophagy is pro-death. The immunostaining study demonstrated co-localization of LC3 and succinate dehydrogenase (a mitochondrial complex II as well as LC3 and α-synuclein, suggesting that autophagy may engulf mitochondria and α-synuclein. Indeed, in vivo Atg7siRNA transfection mitigated MPP+-induced reduction in cytochrome c oxidase. In addition, MPP+-induced autophagy differentially altered the α-synuclein aggregates in the infused SN. In conclusion, autophagy plays a prodeath role in the MPP+-induced oxidative injury by sequestering mitochondria in the rat brain. Moreover, our data suggest that the benefits of autophagy depend on the levels of α-synuclein aggregates in the nigrostriatal dopaminergic system of the rat brain.

  17. Genipin inhibits NLRP3 and NLRC4 inflammasome activation via autophagy suppression

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    Yu, Shui-Xing; Du, Chong-Tao; Chen, Wei; Lei, Qian-Qian; Li, Ning; Qi, Shuai; Zhang, Xiao-Jing; Hu, Gui-Qiu; Deng, Xu-Ming; Han, Wen-Yu; Yang, Yong-Jun

    2015-01-01

    Inflammasomes are cytoplasmic, multiprotein complexes that trigger caspase-1 activation and IL-1β maturation in response to diverse stimuli. Although inflammasomes play important roles in host defense against microbial infection, overactive inflammasomes are deleterious and lead to various autoinflammatory diseases. In the current study, we demonstrated that genipin inhibits the induction of IL-1β production and caspase-1 activation by NLRP3 and NLRC4 inflammasomes. Furthermore, genipin specifically prevented NLRP3-mediated, but not NLRC4-mediated, ASC oligomerization. Notably, genipin inhibited autophagy, leading to NLRP3 and NLRC4 inflammasome inhibition. UCP2-ROS signaling may be involved in inflammasome suppression by genipin. In vivo, we showed that genipin inhibited NLRP3-dependent IL-1β production and neutrophil flux in LPS- and alum-induced murine peritonitis. Additionally, genipin provided protection against flagellin-induced lung inflammation by reducing IL-1β production and neutrophil recruitment. Collectively, our results revealed a novel role in inhibition of inflammatory diseases for genipin that has been used as therapeutics for centuries in herb medicine. PMID:26659006

  18. Genipin inhibits NLRP3 and NLRC4 inflammasome activation via autophagy suppression.

    Science.gov (United States)

    Yu, Shui-Xing; Du, Chong-Tao; Chen, Wei; Lei, Qian-Qian; Li, Ning; Qi, Shuai; Zhang, Xiao-Jing; Hu, Gui-Qiu; Deng, Xu-Ming; Han, Wen-Yu; Yang, Yong-Jun

    2015-01-01

    Inflammasomes are cytoplasmic, multiprotein complexes that trigger caspase-1 activation and IL-1β maturation in response to diverse stimuli. Although inflammasomes play important roles in host defense against microbial infection, overactive inflammasomes are deleterious and lead to various autoinflammatory diseases. In the current study, we demonstrated that genipin inhibits the induction of IL-1β production and caspase-1 activation by NLRP3 and NLRC4 inflammasomes. Furthermore, genipin specifically prevented NLRP3-mediated, but not NLRC4-mediated, ASC oligomerization. Notably, genipin inhibited autophagy, leading to NLRP3 and NLRC4 inflammasome inhibition. UCP2-ROS signaling may be involved in inflammasome suppression by genipin. In vivo, we showed that genipin inhibited NLRP3-dependent IL-1β production and neutrophil flux in LPS- and alum-induced murine peritonitis. Additionally, genipin provided protection against flagellin-induced lung inflammation by reducing IL-1β production and neutrophil recruitment. Collectively, our results revealed a novel role in inhibition of inflammatory diseases for genipin that has been used as therapeutics for centuries in herb medicine. PMID:26659006

  19. Noncoding RNA blockade of autophagy is therapeutic in medullary thyroid cancer

    International Nuclear Information System (INIS)

    Micro-RNAs are dysregulated in medullary thyroid carcinoma (MTC) and preliminary studies have shown that miRNAs may enact a therapeutic effect through changes in autophagic flux. Our aim was to study the in vitro effect of miR-9-3p on MTC cell viability, autophagy and to investigate the mRNA autophagy gene profile of sporadic versus hereditary MTC. The therapeutic role of miR-9-3p was investigated in vitro using human MTC cell lines (TT and MZ-CRC-1 cells), cell viability assays, and functional mechanism studies with a focus on cell cycle, apoptosis, and autophagy. Post-miR-9-3p transfection mRNA profiling of cell lines was performed using a customized, quantitative RT-PCR gene array card. This card was also run on clinical tumor samples (sporadic: n = 6; hereditary: n = 6) and correlated with clinical data. Mir-9-3p transfection resulted in reduced in vitro cell viability; an effect mediated through autophagy inhibition. This was accompanied by evidence of G2 arrest in the TT cell line and increased apoptosis in both cell lines. Atg5 was validated as a predicted miR-9-3p mRNA target in TT cells. Post-miR-9-3p transfection array studies showed a significant global decline in autophagy gene expression (most notably in PIK3C3, mTOR, and LAMP-1). Autophagy gene mRNAs were generally overexpressed in sporadic (vs. hereditary MTC) and Beclin-1 overexpression was shown to correlate with residual disease. Autophagy is a tumor cell survival mechanism in MTC that when disabled, is of therapeutic advantage. Beclin-1 expression may be a useful prognostic biomarker of aggressive disease

  20. Mevalonate cascade regulation of airway mesenchymal cell autophagy and apoptosis: a dual role for p53.

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    Saeid Ghavami

    Full Text Available Statins inhibit the proximal steps of cholesterol biosynthesis, and are linked to health benefits in various conditions, including cancer and lung disease. We have previously investigated apoptotic pathways triggered by statins in airway mesenchymal cells, and identified reduced prenylation of small GTPases as a primary effector mechanism leading to p53-mediated cell death. Here, we extend our studies of statin-induced cell death by assessing endpoints of both apoptosis and autophagy, and investigating their interplay and coincident regulation. Using primary cultured human airway smooth muscle (HASM and human airway fibroblasts (HAF, autophagy, and autophagosome formation and flux were assessed by transmission electron microscopy, cytochemistry (lysosome number and co-localization with LC3 and immunoblotting (LC3 lipidation and Atg12-5 complex formation. Chemical inhibition of autophagy increased simvastatin-induced caspase activation and cell death. Similarly, Atg5 silencing with shRNA, thus preventing Atg5-12 complex formation, increased pro-apoptotic effects of simvastatin. Simvastatin concomitantly increased p53-dependent expression of p53 up-regulated modulator of apoptosis (PUMA, NOXA, and damage-regulated autophagy modulator (DRAM. Notably both mevalonate cascade inhibition-induced autophagy and apoptosis were p53 dependent: simvastatin increased nuclear p53 accumulation, and both cyclic pifithrin-α and p53 shRNAi partially inhibited NOXA, PUMA expression and caspase-3/7 cleavage (apoptosis and DRAM expression, Atg5-12 complex formation, LC3 lipidation, and autophagosome formation (autophagy. Furthermore, the autophagy response is induced rapidly, significantly delaying apoptosis, suggesting the existence of a temporally coordinated p53 regulation network. These findings are relevant for the development of statin-based therapeutic approaches in obstructive airway disease.

  1. Carbamazepine suppresses calpain-mediated autophagy impairment after ischemia/reperfusion in mouse livers

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jae-Sung, E-mail: Jae.Kim@surgery.ufl.edu; Wang, Jin-Hee, E-mail: jin-hee.wang@surgery.ufl.edu; Biel, Thomas G., E-mail: Thomas.Biel@surgery.ufl.edu; Kim, Do-Sung, E-mail: do-sung.kim@surgery.med.ufl.edu; Flores-Toro, Joseph A., E-mail: Joseph.Flores-Toro@surgery.ufl.edu; Vijayvargiya, Richa, E-mail: rvijayvargiya@ufl.edu; Zendejas, Ivan, E-mail: ivan.zendejas@surgery.ufl.edu; Behrns, Kevin E., E-mail: Kevin.Behrns@surgery.ufl.edu

    2013-12-15

    Onset of the mitochondrial permeability transition (MPT) plays a causative role in ischemia/reperfusion (I/R) injury. Current therapeutic strategies for reducing reperfusion injury remain disappointing. Autophagy is a lysosome-mediated, catabolic process that timely eliminates abnormal or damaged cellular constituents and organelles such as dysfunctional mitochondria. I/R induces calcium overloading and calpain activation, leading to degradation of key autophagy-related proteins (Atg). Carbamazepine (CBZ), an FDA-approved anticonvulsant drug, has recently been reported to increase autophagy. We investigated the effects of CBZ on hepatic I/R injury. Hepatocytes and livers from male C57BL/6 mice were subjected to simulated in vitro, as well as in vivo I/R, respectively. Cell death, intracellular calcium, calpain activity, changes in autophagy-related proteins (Atg), autophagic flux, MPT and mitochondrial membrane potential after I/R were analyzed in the presence and absence of 20 μM CBZ. CBZ significantly increased hepatocyte viability after reperfusion. Confocal microscopy revealed that CBZ prevented calcium overloading, the onset of the MPT and mitochondrial depolarization. Immunoblotting and fluorometric analysis showed that CBZ blocked calpain activation, depletion of Atg7 and Beclin-1 and loss of autophagic flux after reperfusion. Intravital multiphoton imaging of anesthetized mice demonstrated that CBZ substantially reversed autophagic defects and mitochondrial dysfunction after I/R in vivo. In conclusion, CBZ prevents calcium overloading and calpain activation, which, in turn, suppresses Atg7 and Beclin-1 depletion, defective autophagy, onset of the MPT and cell death after I/R. - Highlights: • A mechanism of carbamazepine (CBZ)-induced cytoprotection in livers is proposed. • Impaired autophagy is a key event contributing to lethal reperfusion injury. • The importance of autophagy is extended and confirmed in an in vivo model. • CBZ is a potential

  2. Carbamazepine suppresses calpain-mediated autophagy impairment after ischemia/reperfusion in mouse livers

    International Nuclear Information System (INIS)

    Onset of the mitochondrial permeability transition (MPT) plays a causative role in ischemia/reperfusion (I/R) injury. Current therapeutic strategies for reducing reperfusion injury remain disappointing. Autophagy is a lysosome-mediated, catabolic process that timely eliminates abnormal or damaged cellular constituents and organelles such as dysfunctional mitochondria. I/R induces calcium overloading and calpain activation, leading to degradation of key autophagy-related proteins (Atg). Carbamazepine (CBZ), an FDA-approved anticonvulsant drug, has recently been reported to increase autophagy. We investigated the effects of CBZ on hepatic I/R injury. Hepatocytes and livers from male C57BL/6 mice were subjected to simulated in vitro, as well as in vivo I/R, respectively. Cell death, intracellular calcium, calpain activity, changes in autophagy-related proteins (Atg), autophagic flux, MPT and mitochondrial membrane potential after I/R were analyzed in the presence and absence of 20 μM CBZ. CBZ significantly increased hepatocyte viability after reperfusion. Confocal microscopy revealed that CBZ prevented calcium overloading, the onset of the MPT and mitochondrial depolarization. Immunoblotting and fluorometric analysis showed that CBZ blocked calpain activation, depletion of Atg7 and Beclin-1 and loss of autophagic flux after reperfusion. Intravital multiphoton imaging of anesthetized mice demonstrated that CBZ substantially reversed autophagic defects and mitochondrial dysfunction after I/R in vivo. In conclusion, CBZ prevents calcium overloading and calpain activation, which, in turn, suppresses Atg7 and Beclin-1 depletion, defective autophagy, onset of the MPT and cell death after I/R. - Highlights: • A mechanism of carbamazepine (CBZ)-induced cytoprotection in livers is proposed. • Impaired autophagy is a key event contributing to lethal reperfusion injury. • The importance of autophagy is extended and confirmed in an in vivo model. • CBZ is a potential

  3. Hydrogen sulfide lowers proliferation and induces protective autophagy in colon epithelial cells.

    Directory of Open Access Journals (Sweden)

    Ya C Wu

    Full Text Available Hydrogen sulfide (H(2S is a gaseous bacterial metabolite that reaches high levels in the large intestine. In the present study, the effect of H(2S on the proliferation of normal and cancerous colon epithelial cells was investigated. An immortalized colon epithelial cell line (YAMC and a panel of colon cancer cell lines (HT-29, SW1116, HCT116 were exposed to H(2S at concentrations similar to those found in the human colon. H(2S inhibited normal and cancerous colon epithelial cell proliferation as measured by MTT assay. The anti-mitogenic effect of H(2S was accompanied by G(1-phase cell cycle arrest and the induction of the cyclin-dependent kinase inhibitor p21(Cip. Moreover, exposure to H(2S led to features characteristic of autophagy, including increased formation of LC3B(+ autophagic vacuoles and acidic vesicular organelles as determined by immunofluorescence and acridine orange staining, respectively. Abolition of autophagy by RNA interference targeting Vps34 or Atg7 enhanced the anti-proliferative effect of H(2S. Further mechanistic investigation revealed that H(2S stimulated the phosphorylation of AMP-activated protein kinase (AMPK and inhibited the phosphorylation of mammalian target of rapamycin (mTOR and S6 kinase. Inhibition of AMPK significantly reversed H(2S-induced autophagy and inhibition of cell proliferation. Collectively, we demonstrate that H(2S inhibits colon epithelial cell proliferation and induces protective autophagy via the AMPK pathway.

  4. MCOLN1 is a ROS sensor in lysosomes that regulates autophagy

    Science.gov (United States)

    Zhang, Xiaoli; Cheng, Xiping; Yu, Lu; Yang, Junsheng; Calvo, Raul; Patnaik, Samarjit; Hu, Xin; Gao, Qiong; Yang, Meimei; Lawas, Maria; Delling, Markus; Marugan, Juan; Ferrer, Marc; Xu, Haoxing

    2016-01-01

    Cellular stresses trigger autophagy to remove damaged macromolecules and organelles. Lysosomes ‘host' multiple stress-sensing mechanisms that trigger the coordinated biogenesis of autophagosomes and lysosomes. For example, transcription factor (TF)EB, which regulates autophagy and lysosome biogenesis, is activated following the inhibition of mTOR, a lysosome-localized nutrient sensor. Here we show that reactive oxygen species (ROS) activate TFEB via a lysosomal Ca2+-dependent mechanism independent of mTOR. Exogenous oxidants or increasing mitochondrial ROS levels directly and specifically activate lysosomal TRPML1 channels, inducing lysosomal Ca2+ release. This activation triggers calcineurin-dependent TFEB-nuclear translocation, autophagy induction and lysosome biogenesis. When TRPML1 is genetically inactivated or pharmacologically inhibited, clearance of damaged mitochondria and removal of excess ROS are blocked. Furthermore, TRPML1's ROS sensitivity is specifically required for lysosome adaptation to mitochondrial damage. Hence, TRPML1 is a ROS sensor localized on the lysosomal membrane that orchestrates an autophagy-dependent negative-feedback programme to mitigate oxidative stress in the cell. PMID:27357649

  5. Down regulated lncRNA MEG3 eliminates mycobacteria in macrophages via autophagy

    Science.gov (United States)

    Pawar, Kamlesh; Hanisch, Carlos; Palma Vera, Sergio Eliseo; Einspanier, Ralf; Sharbati, Soroush

    2016-01-01

    Small non-coding RNA play a major part in host response to bacterial agents. However, the role of long non-coding RNA (lncRNA) in this context remains unknown. LncRNA regulate gene expression by acting e.g. as transcriptional coactivators, RNA decoys or microRNA sponges. They control development, differentiation and cellular processes such as autophagy in disease conditions. Here, we provide an insight into the role of lncRNA in mycobacterial infections. Human macrophages were infected with Mycobacterium bovis BCG and lncRNA expression was studied early post infection. For this purpose, lncRNA with known immune related functions were preselected and a lncRNA specific RT-qPCR protocol was established. In addition to expression-based prediction of lncRNA function, we assessed strategies for thorough normalisation of lncRNA. Arrayed quantification showed infection-dependent repression of several lncRNA including MEG3. Pathway analysis linked MEG3 to mTOR and PI3K-AKT signalling pointing to regulation of autophagy. Accordingly, IFN-γ induced autophagy in infected macrophages resulted in sustained MEG3 down regulation and lack of IFN-γ allowed for counter regulation of MEG3 by viable M. bovis BCG. Knockdown of MEG3 in macrophages resulted in induction of autophagy and enhanced eradication of intracellular M. bovis BCG. PMID:26757825

  6. Oxidative stress-induced autophagy: Role in pulmonary toxicity

    International Nuclear Information System (INIS)

    Autophagy is an evolutionarily conserved catabolic process important in regulating the turnover of essential proteins and in elimination of damaged organelles and protein aggregates. Autophagy is observed in the lung in response to oxidative stress generated as a consequence of exposure to environmental toxicants. Whether autophagy plays role in promoting cell survival or cytotoxicity is unclear. In this article recent findings on oxidative stress-induced autophagy in the lung are reviewed; potential mechanisms initiating autophagy are also discussed. A better understanding of autophagy and its role in pulmonary toxicity may lead to the development of new strategies to treat lung injury associated with oxidative stress. - Highlights: • Exposure to pulmonary toxicants is associated with oxidative stress. • Oxidative stress is known to induce autophagy. • Autophagy is upregulated in the lung following exposure to pulmonary toxicants. • Autophagy may be protective or pathogenic

  7. Oxidative stress-induced autophagy: Role in pulmonary toxicity

    Energy Technology Data Exchange (ETDEWEB)

    Malaviya, Rama [Department of Pharmacology and Toxicology, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, NJ 08854 (United States); Laskin, Jeffrey D. [Department of Environmental and Occupational Medicine, Robert Wood Johnson Medical School, Rutgers University, Piscataway, NJ 08854 (United States); Laskin, Debra L., E-mail: laskin@eohsi.rutgers.edu [Department of Pharmacology and Toxicology, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, NJ 08854 (United States)

    2014-03-01

    Autophagy is an evolutionarily conserved catabolic process important in regulating the turnover of essential proteins and in elimination of damaged organelles and protein aggregates. Autophagy is observed in the lung in response to oxidative stress generated as a consequence of exposure to environmental toxicants. Whether autophagy plays role in promoting cell survival or cytotoxicity is unclear. In this article recent findings on oxidative stress-induced autophagy in the lung are reviewed; potential mechanisms initiating autophagy are also discussed. A better understanding of autophagy and its role in pulmonary toxicity may lead to the development of new strategies to treat lung injury associated with oxidative stress. - Highlights: • Exposure to pulmonary toxicants is associated with oxidative stress. • Oxidative stress is known to induce autophagy. • Autophagy is upregulated in the lung following exposure to pulmonary toxicants. • Autophagy may be protective or pathogenic.

  8. Changes in autophagy in rats after spinal cord injury and the effect of hyperbaric oxygen on autophagy.

    Science.gov (United States)

    Sun, Yongming; Liu, Dong; Su, Peng; Lin, Fanguo; Tang, Qifeng

    2016-04-01

    The purpose of this study was to explore the effects of Hyperbaric oxygen (HBO) on the autophagic changes after induction of spinal cord injury (SCI) in rats. A total of 75 rats were randomly divided into the sham-operated group, the spinal cord injury group, and the SCI+HBO group. We found that at 7 d and 14 d after surgery, the BBB scores were higher in the SCI+HBO group in comparison to the SCI group. The expression of Beclin-1 and LC3II was upregulated in the SCI and SCI+HBO groups after SCI. Fluorescently stained Beclin-1 and LC3II proteins were barely detectable in the sham group. In contrast, Beclin-l and LC3II expression was observed in neurons and glial cells from the SCI and SCI+HBO groups. Beclin-1 and LC3II expression appeared at 6h after SCI. At each time point, Beclin-1 and LC3II expression was significantly higher in the SCI+HBO group compared to the SCI group. These results suggest that autophagy is activated in rats after SCI and sustained over a period of time. HBO treatment enhances autophagy expression in rats after SCI and accelerates cell repair rate, which may represent one of the mechanisms of action of HBO in the treatment of SCI. PMID:26949182

  9. Downregulation of the autophagy protein ATG-7 correlates with distinct sphingolipid profile in MCF-7 cells sensitized to photodamage

    Science.gov (United States)

    Separovic, Duska; Kelekar, Ameeta; Tarca, Adi L.; Bielawski, Jacek; Kessel, David

    2009-06-01

    The objective of this study was to determine the sphingolipid (SL) profile in autophagy-defective cells and overall cell death after PDT with Pc 4 (PDT). Human breast cancer MCF-7 cells with downregulated autophagy protein ATG-7 and their scrambled controls (Scr) were used. Exposure of ATG-7 knockdown cells to PDT led to defective processing of the autophagy marker LC3, and increased overall cell killing. In both cell types PDT evoked an early (2 h) increase in ceramides and dihydroceramides (DHceramides). When the two cell types were compared regarding time (2 and 24 h) and treatment conditions (with and without PDT), the levels of several ceramides and DHceramides were reduced, whereas the concentrations of C14-ceramide, C16-ceramide and C12-DHceramide were higher in ATG-7 knockdown cells. The data imply that the SL profile might be a marker of autophagy-deficiency in cells sensitized to PDT.

  10. Labor Induction

    Science.gov (United States)

    ... QUESTIONS FAQ154 LABOR, DELIVERY, AND POSTPARTUM CARE Labor Induction • What is labor induction? • Why is labor induced? • What is the Bishop ... oxytocin? • What are the risks associated with labor induction? • Is labor induction always effective? • Glossary What is ...

  11. Reduced-Capacity Inrush Current Suppressor Using a Matrix Converter in a Wind Power Generation System with Squirrel-Cage Induction Machines

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    Sho Shibata

    2016-03-01

    Full Text Available This paper describes the reduced capacity of the inrush current suppressor using a matrix converter (MC in a large-capacity wind power generation system (WPGS with two squirrel-cage induction machines (SCIMs. These SCIMs are switched over depending on the wind speed. The input side of the MC is connected to the source in parallel. The output side of the MC is connected in series with the SCIM through matching transformers. The modulation method of the MC used is direct duty ratio pulse width modulation. The reference output voltage of the MC is decided by multiplying the SCIM current with the variable control gain. Therefore, the MC performs as resistors for the inrush current. Digital computer simulation is implemented to confirm the validity and practicability of the proposed inrush current suppressor using PSCAD/EMTDC (power system computer-aided design/electromagnetic transients including DC. Furthermore, the equivalent resistance of the MC is decided by the relationship between the equivalent resistance and the capacity of the MC. Simulation results demonstrate that the proposed inrush current suppressor can suppress the inrush current perfectly.

  12. Re-expression of ARHI (DIRAS3) induces autophagy in breast cancer cells and enhances the inhibitory effect of paclitaxel

    International Nuclear Information System (INIS)

    ARHI is a Ras-related imprinted gene that inhibits cancer cell growth and motility. ARHI is downregulated in the majority of breast cancers, and loss of its expression is associated with its progression from ductal carcinoma in situ (DCIS) to invasive disease. In ovarian cancer, re-expression of ARHI induces autophagy and leads to autophagic death in cell culture; however, ARHI re-expression enables ovarian cancer cells to remain dormant when they are grown in mice as xenografts. The purpose of this study is to examine whether ARHI induces autophagy in breast cancer cells and to evaluate the effects of ARHI gene re-expression in combination with paclitaxel. Re-expression of ARHI was achieved by transfection, by treatment with trichostatin A (TSA) or by a combination of TSA and 5-aza-2'-deoxycytidine (DAC) in breast cancer cell cultures and by liposomal delivery of ARHI in breast tumor xenografts. ARHI re-expression induces autophagy in breast cancer cells, and ARHI is essential for the induction of autophagy. When ARHI was re-expressed in breast cancer cells treated with paclitaxel, the growth inhibitory effect of paclitaxel was enhanced in both the cell culture and the xenografts. Although paclitaxel alone did not induce autophagy in breast cancer cells, it enhanced ARHI-induced autophagy. Conversely, ARHI re-expression promoted paclitaxel-induced apoptosis and G2/M cell cycle arrest. ARHI re-expression induces autophagic cell death in breast cancer cells and enhances the inhibitory effects of paclitaxel by promoting autophagy, apoptosis, and G2/M cell cycle arrest

  13. Re-expression of ARHI (DIRAS3 induces autophagy in breast cancer cells and enhances the inhibitory effect of paclitaxel

    Directory of Open Access Journals (Sweden)

    Bast Robert C

    2011-01-01

    Full Text Available Abstract Background ARHI is a Ras-related imprinted gene that inhibits cancer cell growth and motility. ARHI is downregulated in the majority of breast cancers, and loss of its expression is associated with its progression from ductal carcinoma in situ (DCIS to invasive disease. In ovarian cancer, re-expression of ARHI induces autophagy and leads to autophagic death in cell culture; however, ARHI re-expression enables ovarian cancer cells to remain dormant when they are grown in mice as xenografts. The purpose of this study is to examine whether ARHI induces autophagy in breast cancer cells and to evaluate the effects of ARHI gene re-expression in combination with paclitaxel. Methods Re-expression of ARHI was achieved by transfection, by treatment with trichostatin A (TSA or by a combination of TSA and 5-aza-2'-deoxycytidine (DAC in breast cancer cell cultures and by liposomal delivery of ARHI in breast tumor xenografts. Results ARHI re-expression induces autophagy in breast cancer cells, and ARHI is essential for the induction of autophagy. When ARHI was re-expressed in breast cancer cells treated with paclitaxel, the growth inhibitory effect of paclitaxel was enhanced in both the cell culture and the xenografts. Although paclitaxel alone did not induce autophagy in breast cancer cells, it enhanced ARHI-induced autophagy. Conversely, ARHI re-expression promoted paclitaxel-induced apoptosis and G2/M cell cycle arrest. Conclusions ARHI re-expression induces autophagic cell death in breast cancer cells and enhances the inhibitory effects of paclitaxel by promoting autophagy, apoptosis, and G2/M cell cycle arrest.

  14. Mechanical stress triggers cardiomyocyte autophagy through angiotensin II type 1 receptor-mediated p38MAP kinase independently of angiotensin II.

    Directory of Open Access Journals (Sweden)

    Li Lin

    Full Text Available Angiotensin II (Ang II type 1 (AT1 receptor is known to mediate a variety of physiological actions of Ang II including autophagy. However, the role of AT1 receptor in cardiomyocyte autophagy triggered by mechanical stress still remains elusive. The aim of this study was therefore to examine whether and how AT1 receptor participates in cardiomyocyte autophagy induced by mechanical stresses. A 48-hour mechanical stretch and a 4-week transverse aorta constriction (TAC were imposed to cultured cardiomyocytes of neonatal rats and adult male C57B/L6 mice, respectively, to induce cardiomyocyte hypertrophy prior to the assessment of cardiomyocyte autophagy using LC3b-II. Losartan, an AT1 receptor blocker, but not PD123319, the AT2 inhibitor, was found to significantly reduce mechanical stretch-induced LC3b-II upregulation. Moreover, inhibition of p38MAP kinase attenuated not only mechanical stretch-induced cardiomyocyte hypertrophy but also autophagy. To the contrary, inhibition of ERK and JNK suppressed cardiac hypertrophy but not autophagy. Intriguingly, mechanical stretch-induced autophagy was significantly inhibited by Losartan in the absence of Ang II. Taken together, our results indicate that mechanical stress triggers cardiomyocyte autophagy through AT1 receptor-mediated activation of p38MAP kinase independently of Ang II.

  15. Effect of Exendin-4 on Autophagy Clearance in Beta Cell of Rats with Tacrolimus-induced Diabetes Mellitus

    Science.gov (United States)

    Lim, Sun Woo; Jin, Long; Jin, Jian; Yang, Chul Woo

    2016-01-01

    Growing evidence suggests that GLP-1 protects beta cells against various cellular injuries by modulating autophagy. In this study, we examined whether exendin-4 (Ex-4), a GLP-1 analog, had preventive effects on tacrolimus (Tac)-induced beta cell injury by improving autophagy clearance. Rats with Tac-induced diabetes mellitus exhibited increased autophagy-associated protein expression, light chain 3B levels, and autophagic vacuole numbers in pancreatic beta cells. Additionally, Tac increased autophagy in a dose- and time-dependent manner in vitro, and inhibition of autophagosome using 3-methyladenine reduced Tac-induced islet cell injury by decreasing reactive oxygen species production and apoptosis. Ex-4 treatment decreased Tac-induced hyperglycaemia, oxidative stress, and apoptosis, accompanied by decreased autophagy-associated protein expression and autophagosome numbers. In vivo and in vitro studies showed that Tac treatment impaired lysosomal function and autophagosome-lysosome fusion; these processes were improve by Ex-4 treatment. Moreover, addition of bafilomycin A1, an inhibitor of lysosomal function, abolished the protective effects of Ex-4. Our findings reveal that Tac-induced diabetes mellitus was a state of excessive burden of autophagosomes and impairment of autophagy clearance and that Ex-4 protected against Tac-induced pancreatic islet injury by reducing the burden of autophagosomes via activation of autophagosome clearance. Thus, Ex-4 had therapeutic effects on Tac-induced pancreatic beta cell injury. PMID:27436514

  16. Autophagy in lung disease pathogenesis and therapeutics

    OpenAIRE

    Ryter, Stefan W.; Augustine M K Choi

    2015-01-01

    Autophagy, a cellular pathway for the degradation of damaged organelles and proteins, has gained increasing importance in human pulmonary diseases, both as a modulator of pathogenesis and as a potential therapeutic target. In this pathway, cytosolic cargos are sequestered into autophagosomes, which are delivered to the lysosomes where they are enzymatically degraded and then recycled as metabolic precursors. Autophagy exerts an important effector function in the regulation of inflammation, an...

  17. Skeletal Muscle Autophagy: A New Metabolic Regulator

    OpenAIRE

    Neel, Brian A.; Lin, Yuxi; Pessin, Jeffrey E.

    2013-01-01

    Autophagy classically functions as a physiological process to degrade cytoplasmic components, protein aggregates, and/or organelles, as a mechanism for nutrient breakdown, and as a regulator of cellular architecture. Proper autophagic flux is vital for both functional skeletal muscle, which controls support and movement of the skeleton, and muscle metabolism. The role of autophagy as a metabolic regulator in muscle has been previously studied; however, the underlying molecular mechanisms that...

  18. The role of autophagy in Parkinson's disease☆

    OpenAIRE

    Zhang, Lei; Dong, Yaru; Xu, Xiaoheng; Xu, Zhong

    2012-01-01

    Although Parkinson's disease is the most common neurodegenerative movement disorder, the mechanisms of pathogenesis remain poorly understood. Recent findings have shown that deregulation of the autophagy-lysosome pathway is involved in the pathogenesis of Parkinson's disease. This review summarizes the most recent findings and discusses the unique role of the autophagy-lysosome pathway in Parkinson's disease to highlight the possibility of Parkinson's disease treatment strategies that incorpo...

  19. Mechanisms of mitochondria and autophagy crosstalk

    OpenAIRE

    Rambold, Angelika S.; Lippincott-Schwartz, Jennifer

    2011-01-01

    Autophagy is a cellular survival pathway that recycles intracellular components to compensate for nutrient depletion and ensures the appropriate degradation of organelles. Mitochondrial number and health are regulated by mitophagy, a process by which excessive or damaged mitochondria are subjected to autophagic degradation. Autophagy is thus a key determinant for mitochondrial health and proper cell function. Mitophagic malfunction has been recently proposed to contribute to progressive neuro...

  20. RUN and FYVE domain-containing protein 4 enhances autophagy and lysosome tethering in response to Interleukin-4.

    Science.gov (United States)

    Terawaki, Seigo; Camosseto, Voahirana; Prete, Francesca; Wenger, Till; Papadopoulos, Alexia; Rondeau, Christiane; Combes, Alexis; Rodriguez Rodrigues, Christian; Vu Manh, Thien-Phong; Fallet, Mathieu; English, Luc; Santamaria, Rodrigo; Soares, Ana R; Weil, Tobias; Hammad, Hamida; Desjardins, Michel; Gorvel, Jean-Pierre; Santos, Manuel A S; Gatti, Evelina; Pierre, Philippe

    2015-09-28

    Autophagy is a key degradative pathway coordinated by external cues, including starvation, oxidative stress, or pathogen detection. Rare are the molecules known to contribute mechanistically to the regulation of autophagy and expressed specifically in particular environmental contexts or in distinct cell types. Here, we unravel the role of RUN and FYVE domain-containing protein 4 (RUFY4) as a positive molecular regulator of macroautophagy in primary dendritic cells (DCs). We show that exposure to interleukin-4 (IL-4) during DC differentiation enhances autophagy flux through mTORC1 regulation and RUFY4 induction, which in turn actively promote LC3 degradation, Syntaxin 17-positive autophagosome formation, and lysosome tethering. Enhanced autophagy boosts endogenous antigen presentation by MHC II and allows host control of Brucella abortus replication in IL-4-treated DCs and in RUFY4-expressing cells. RUFY4 is therefore the first molecule characterized to date that promotes autophagy and influences endosome dynamics in a subset of immune cells. PMID:26416964

  1. RUFY4: Immunity piggybacking on autophagy?

    Science.gov (United States)

    Terawaki, Seigo; Camosseto, Voahirana; Pierre, Philippe; Gatti, Evelina

    2016-03-01

    Although autophagy is a highly conserved mechanism among species and cell types, few are the molecules involved with the autophagic process that display cell- or tissue- specific expression. We have unraveled the positive regulatory role on autophagy of RUFY4 (RUN and FYVE domain containing 4), which is expressed in subsets of immune cells, including dendritic cells (DCs). DCs orchestrate the eradication of pathogens by coordinating the action of the different cell types involved in microbe recognition and destruction during the immune response. To fulfill this function, DC display particular regulation of their endocytic and autophagy pathways in response to the immune environment. Autophagy flux is downmodulated in DCs upon microbe sensing, but is remarkably augmented, when cells are differentiated in the presence of the pleiotropic cytokine IL4 (interleukin 4). From gene expression studies aimed at comparing the impact of IL4 on DC differentiation, we identified RUFY4, as a novel regulator that augments autophagy flux and, when overexpressed, induces drastic membrane redistribution and strongly tethers lysosomes. RUFY4 is therefore one of the few known positive regulators of autophagy that is expressed in a cell-specific manner or under specific immunological conditions associated with IL4 expression such as allergic asthma. PMID:26760128

  2. Lexicographic Path Induction

    DEFF Research Database (Denmark)

    Schürmann, Carsten; Sarnat, Jeffrey

    2009-01-01

    Programming languages theory is full of problems that reduce to proving the consistency of a logic, such as the normalization of typed lambda-calculi, the decidability of equality in type theory, equivalence testing of traces in security, etc. Although the principle of transfinite induction is...... routinely employed by logicians in proving such theorems, it is rarely used by programming languages researchers, who often prefer alternatives such as proofs by logical relations and model theoretic constructions. In this paper we harness the well-foundedness of the lexicographic path ordering to derive an...... induction principle that combines the comfort of structural induction with the expressive strength of transfinite induction. Using lexicographic path induction, we give a consistency proof of Martin-Löf’s intuitionistic theory of inductive definitions. The consistency of Heyting arithmetic follows directly...

  3. Salinomycin induces autophagy in colon and breast cancer cells with concomitant generation of reactive oxygen species.

    Directory of Open Access Journals (Sweden)

    Berlinda Verdoodt

    Full Text Available BACKGROUND: Salinomycin is a polyether ionophore antibiotic that has recently been shown to induce cell death in human cancer cells displaying multiple mechanisms of drug resistance. The underlying mechanisms leading to cell death after salinomycin treatment have not been well characterized. We therefore investigated the role of salinomycin in caspase dependent and independent cell death in colon cancer (SW480, SW620, RKO and breast cancer cell lines (MCF-7, T47D, MDA-MB-453. METHODOLOGY/PRINCIPAL FINDINGS: We detected features of apoptosis in all cell lines tested, but the executor caspases 3 and 7 were only strongly activated in RKO and MDA-MB-453 cells. MCF-7 and SW620 cells instead presented features of autophagy such as cytoplasmic vacuolization and LC3 processing. Caspase proficient cell lines activated autophagy at lower salinomycin concentrations and before the onset of caspase activation. Salinomycin also led to the formation of reactive oxygen species (ROS eliciting JNK activation and induction of the transcription factor JUN. Salinomycin mediated cell death could be partially inhibited by the free radical scavenger N-acetyl-cysteine, implicating ROS formation in the mechanism of salinomycin toxicity. CONCLUSIONS: Our data indicate that, in addition to its previously reported induction of caspase dependent apoptosis, the initiation of autophagy is an important and early effect of salinomycin in tumor cells.

  4. Porcine Circovirus Type 2 Activates CaMMKβ to Initiate Autophagy in PK-15 Cells by Increasing Cytosolic Calcium

    Science.gov (United States)

    Gu, Yuanxing; Qi, Baozhu; Zhou, Yingshan; Jiang, Xiaowu; Zhang, Xian; Li, Xiaoliang; Fang, Weihuan

    2016-01-01

    Porcine circovirus type 2 (PCV2) induces autophagy via the 5′ adenosine monophosphate-activated protein kinase (AMPK)/extracellular signal-regulated kinase (ERK)/tuberous sclerosis complex 2 (TSC2)/mammalian target of rapamycin (mTOR) pathway in pig kidney PK-15 cells. However, the underlying mechanisms of AMPK activation in autophagy induction remain unknown. With specific inhibitors and RNA interference (RNAi), we show that PCV2 infection upregulated calcium/calmodulin-dependent protein kinase kinase-beta (CaMKKβ) by increasing cytosolic Ca2+ via inositol 1,4,5-trisphosphate receptor (IP3R). Elevation of cytosolic calcium ion (Ca2+) did not seem to involve inositol 1,4,5-trisphosphate (IP3) release from phosphatidylinositol 4,5-bisphosphate (PIP2) by phosphoinositide phospholipase C-gamma (PLC-γ). CaMKKβ then activated both AMPK and calcium/calmodulin-dependent protein kinase I (CaMKI). PCV2 employed CaMKI and Trp-Asp (WD) repeat domain phosphoinositide-interacting protein 1 (WIPI1) as another pathway additional to AMPK signaling in autophagy initiation. Our findings could help better understanding of the signaling pathways of autophagy induction as part of PCV2 pathogenesis. Further research is warranted to study if PCV2 interacts directly with IP3R or indirectly with the molecules that antagonize IP3R activity responsible for increased cytosolic Ca2+ both in PK-15 cells and PCV2-targeted primary cells from pigs. PMID:27213427

  5. Porcine Circovirus Type 2 Activates CaMMKβ to Initiate Autophagy in PK-15 Cells by Increasing Cytosolic Calcium

    Directory of Open Access Journals (Sweden)

    Yuanxing Gu

    2016-05-01

    Full Text Available Porcine circovirus type 2 (PCV2 induces autophagy via the 5′ adenosine monophosphate-activated protein kinase (AMPK/extracellular signal-regulated kinase (ERK/tuberous sclerosis complex 2 (TSC2/mammalian target of rapamycin (mTOR pathway in pig kidney PK-15 cells. However, the underlying mechanisms of AMPK activation in autophagy induction remain unknown. With specific inhibitors and RNA interference (RNAi, we show that PCV2 infection upregulated calcium/calmodulin-dependent protein kinase kinase-beta (CaMKKβ by increasing cytosolic Ca2+ via inositol 1,4,5-trisphosphate receptor (IP3R. Elevation of cytosolic calcium ion (Ca2+ did not seem to involve inositol 1,4,5-trisphosphate (IP3 release from phosphatidylinositol 4,5-bisphosphate (PIP2 by phosphoinositide phospholipase C-gamma (PLC-γ. CaMKKβ then activated both AMPK and calcium/calmodulin-dependent protein kinase I (CaMKI. PCV2 employed CaMKI and Trp-Asp (WD repeat domain phosphoinositide-interacting protein 1 (WIPI1 as another pathway additional to AMPK signaling in autophagy initiation. Our findings could help better understanding of the signaling pathways of autophagy induction as part of PCV2 pathogenesis. Further research is warranted to study if PCV2 interacts directly with IP3R or indirectly with the molecules that antagonize IP3R activity responsible for increased cytosolic Ca2+ both in PK-15 cells and PCV2-targeted primary cells from pigs.

  6. Porcine Circovirus Type 2 Activates CaMMKβ to Initiate Autophagy in PK-15 Cells by Increasing Cytosolic Calcium.

    Science.gov (United States)

    Gu, Yuanxing; Qi, Baozhu; Zhou, Yingshan; Jiang, Xiaowu; Zhang, Xian; Li, Xiaoliang; Fang, Weihuan

    2016-01-01

    Porcine circovirus type 2 (PCV2) induces autophagy via the 5' adenosine monophosphate-activated protein kinase (AMPK)/extracellular signal-regulated kinase (ERK)/tuberous sclerosis complex 2 (TSC2)/mammalian target of rapamycin (mTOR) pathway in pig kidney PK-15 cells. However, the underlying mechanisms of AMPK activation in autophagy induction remain unknown. With specific inhibitors and RNA interference (RNAi), we show that PCV2 infection upregulated calcium/calmodulin-dependent protein kinase kinase-beta (CaMKKβ) by increasing cytosolic Ca(2+) via inositol 1,4,5-trisphosphate receptor (IP3R). Elevation of cytosolic calcium ion (Ca(2+)) did not seem to involve inositol 1,4,5-trisphosphate (IP3) release from phosphatidylinositol 4,5-bisphosphate (PIP2) by phosphoinositide phospholipase C-gamma (PLC-γ). CaMKKβ then activated both AMPK and calcium/calmodulin-dependent protein kinase I (CaMKI). PCV2 employed CaMKI and Trp-Asp (WD) repeat domain phosphoinositide-interacting protein 1 (WIPI1) as another pathway additional to AMPK signaling in autophagy initiation. Our findings could help better understanding of the signaling pathways of autophagy induction as part of PCV2 pathogenesis. Further research is warranted to study if PCV2 interacts directly with IP3R or indirectly with the molecules that antagonize IP3R activity responsible for increased cytosolic Ca(2+) both in PK-15 cells and PCV2-targeted primary cells from pigs. PMID:27213427

  7. Characterization of early autophagy signaling by quantitative phosphoproteomics

    DEFF Research Database (Denmark)

    Rigbolt, Kristoffer Tg; Zarei, Mostafa; Sprenger, Adrian;

    2014-01-01

    Under conditions of nutrient shortage autophagy is the primary cellular mechanism ensuring availability of substrates for continuous biosynthesis. Subjecting cells to starvation or rapamycin efficiently induces autophagy by inhibiting the MTOR signaling pathway triggering increased autophagic flu...

  8. Plac8 Links Oncogenic Mutations to Regulation of Autophagy and Is Critical to Pancreatic Cancer Progression

    Directory of Open Access Journals (Sweden)

    Conan Kinsey

    2014-05-01

    Full Text Available Mutations in p53 and RAS potently cooperate in oncogenic transformation, and correspondingly, these genetic alterations frequently coexist in pancreatic ductal adenocarcinoma (PDA and other human cancers. Previously, we identified a set of genes synergistically activated by combined RAS and p53 mutations as frequent downstream mediators of tumorigenesis. Here, we show that the synergistically activated gene Plac8 is critical for pancreatic cancer growth. Silencing of Plac8 in cell lines suppresses tumor formation by blocking autophagy, a process essential for maintaining metabolic homeostasis in PDA, and genetic inactivation in an engineered mouse model inhibits PDA progression. We show that Plac8 is a critical regulator of the autophagic machinery, localizing to the lysosomal compartment and facilitating lysosome-autophagosome fusion. Plac8 thus provides a mechanistic link between primary oncogenic mutations and the induction of autophagy, a central mechanism of metabolic reprogramming, during PDA progression.

  9. Rapamycin requires AMPK activity and p27 expression for promoting autophagy-dependent Tsc2-null cell survival.

    Science.gov (United States)

    Campos, Tania; Ziehe, Javiera; Fuentes-Villalobos, Francisco; Riquelme, Orlando; Peña, Daniela; Troncoso, Rodrigo; Lavandero, Sergio; Morin, Violeta; Pincheira, Roxana; Castro, Ariel F

    2016-06-01

    Tuberous sclerosis complex (TSC) disease results from inactivation of the TSC1 or TSC2 gene, and is characterized by benign tumors in several organs. Because TSC tumorigenesis correlates with hyperactivation of mTORC1, current therapies focus on mTORC1 inhibition with rapamycin or its analogs. Rapamycin-induced tumor shrinkage has been reported, but tumor recurrence occurs on withdrawal from rapamycin. Autophagy has been associated with development of TSC tumors and with tumor cell survival during rapamycin treatment. mTORC1 and AMPK directly inhibit and activate autophagy, respectively. AMPK is hyperactivated in TSC cells and tumors, and drives cytoplasmic sequestration of the cell-cycle inhibitor p27KIP (p27). Whether AMPK and p27 are involved in rapamycin-induced autophagy and survival of TSC cells remain unexplored. Here, we show that inhibition of AMPK by compound C or by shRNA-mediated depletion of LKB1 reduces activation of autophagy by rapamycin in Tsc2-null cells. Similarly, shRNA-mediated depletion of p27 inhibited rapamycin-induced autophagy. In support of p27 lying downstream of AMPK on the activation of autophagy in Tsc2-null cells, a p27 mutant that preferentially localizes in the cytosol recovered the effect of rapamycin on autophagy in both p27- and LKB1-depleted cells, but a nuclear p27 mutant was inactive. Finally, we show that p27-dependent activation of autophagy is involved in Tsc2-null cell survival under rapamycin treatment. These results indicate that an AMPK/p27 axis is promoting a survival mechanism that could explain in part the relapse of TSC tumors treated with rapamycin, exposing new avenues for designing more efficient treatments for TSC patients. PMID:26975583

  10. Apoptosis and autophagy induced by pyropheophorbide-α methyl ester-mediated photodynamic therapy in human osteosarcoma MG-63 cells.

    Science.gov (United States)

    Huang, Qiu; Ou, Yun-Sheng; Tao, Yong; Yin, Hang; Tu, Ping-Hua

    2016-06-01

    Pyropheophorbide-α methyl ester (MPPa) was a second-generation photosensitizer with many potential applications. Here, we explored the impact of MPPa-mediated photodynamic therapy (MPPa-PDT) on the apoptosis and autophagy of human osteosarcoma (MG-63) cells as well as the relationships between apoptosis and autophagy of the cells, and investigated the related molecular mechanisms. We found that MPPa-PDT demonstrated the ability to inhibit MG-63 cell viability in an MPPa concentration- and light dose-dependent manner, and to induce apoptosis via the mitochondrial apoptosis pathway. Additionally, MPPa-PDT could also induce autophagy of MG-63 cell. Meanwhile, the ROS scavenger N-acetyl-L-cysteine (NAC) and the Jnk inhibitor SP600125 were found to inhibit the MPPa-PDT-induced autophagy, and NAC could also inhibit Jnk phosphorylation. Furthermore, pretreatment with the autophagy inhibitor 3-methyladenine or chloroquine showed the potential in reducing the apoptosis rate induced by MPPa-PDT in MG-63 cells. Our results indicated that the mitochondrial pathway was involved in MPPa-PDT-induced apoptosis of MG-63 cells. Meanwhile the ROS-Jnk signaling pathway was involved in MPPa-PDT-induced autophagy, which further promoted the apoptosis in MG-63 cells. PMID:27108344

  11. Apoptotic Cell Death Induced by Resveratrol Is Partially Mediated by the Autophagy Pathway in Human Ovarian Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Fangfang Lang

    Full Text Available Resveratrol (trans-3,4,5'-trihydroxystilbene is an active compound in food, such as red grapes, peanuts, and berries. Resveratrol exhibits an anticancer effect on various human cancer cells. However, the mechanism of resveratrol-induced anti-cancer effect at the molecular level remains to be elucidated. In this study, the mechanism underlying the anti-cancer effect of resveratrol in human ovarian cancer cells (OVCAR-3 and Caov-3 was investigated using various molecular biology techniques, such as flow cytometry, western blotting, and RNA interference, with a major focus on the potential role of autophagy in resveratrol-induced apoptotic cell death. We demonstrated that resveratrol induced reactive oxygen species (ROS generation, which triggers autophagy and subsequent apoptotic cell death. Resveratrol induced ATG5 expression and promoted LC3 cleavage. The apoptotic cell death induced by resveratrol was attenuated by both pharmacological and genetic inhibition of autophagy. The autophagy inhibitor chloroquine, which functions at the late stage of autophagy, significantly reduced resveratrol-induced cell death and caspase 3 activity in human ovarian cancer cells. We also demonstrated that targeting ATG5 by siRNA also suppressed resveratrol-induced apoptotic cell death. Thus, we concluded that a common pathway between autophagy and apoptosis exists in resveratrol-induced cell death in OVCAR-3 human ovarian cancer cells.

  12. Hypercapnia Inhibits Autophagy and Bacterial Killing in Human Macrophages by Increasing Expression of Bcl-2 and Bcl-xL

    Science.gov (United States)

    Casalino-Matsuda, S. Marina; Nair, Aisha; Beitel, Greg J.; Gates, Khalilah L.; Sporn, Peter H. S.

    2015-01-01

    Hypercapnia, the elevation of CO2 in blood and tissue, commonly develops in patients with advanced lung disease and severe pulmonary infections, and is associated with high mortality. We previously reported that hypercapnia alters expression of host defense genes, inhibits phagocytosis, and increases the mortality of Pseudomonas pneumonia in mice. However, the effect of hypercapnia on autophagy, a conserved process by which cells sequester and degrade proteins and damaged organelles that also plays a key role in antimicrobial host defense and pathogen clearance, has not previously been examined. In the present study we show that hypercapnia inhibits autophagy induced by starvation, rapamycin, LPS, heat-killed and live bacteria in the human macrophage. Inhibition of autophagy by elevated CO2 was not attributable to acidosis. Hypercapnia also reduced macrophage killing of Pseudomonas aeruginosa. Moreover, elevated CO2 induced the expression of Bcl-2 and Bcl-xL, anti-apoptotic factors that negatively regulate autophagy by blocking Beclin 1, an essential component of the autophagy initiation complex. Furthermore, siRNA targeting Bcl-2 and Bcl-xL and the small molecule Z36, which blocks Bcl-2 and Bcl-xL binding to Beclin 1, prevented hypercapnic inhibition of autophagy and bacterial killing. These results suggest that targeting the Bcl-2/Bcl-xL-Beclin 1 interaction may hold promise for ameliorating hypercapnia-induced immunosuppression and improving resistance to infection in patients with advanced lung disease and hypercapnia. PMID:25895534

  13. Effector caspase Dcp-1 and IAP protein Bruce regulate starvation-induced autophagy during Drosophila melanogaster oogenesis.

    Science.gov (United States)

    Hou, Ying-Chen Claire; Chittaranjan, Suganthi; Barbosa, Sharon González; McCall, Kimberly; Gorski, Sharon M

    2008-09-22

    A complex relationship exists between autophagy and apoptosis, but the regulatory mechanisms underlying their interactions are largely unknown. We conducted a systematic study of Drosophila melanogaster cell death-related genes to determine their requirement in the regulation of starvation-induced autophagy. We discovered that six cell death genes--death caspase-1 (Dcp-1), hid, Bruce, Buffy, debcl, and p53-as well as Ras-Raf-mitogen activated protein kinase signaling pathway components had a role in autophagy regulation in D. melanogaster cultured cells. During D. melanogaster oogenesis, we found that autophagy is induced at two nutrient status checkpoints: germarium and mid-oogenesis. At these two stages, the effector caspase Dcp-1 and the inhibitor of apoptosis protein Bruce function to regulate both autophagy and starvation-induced cell death. Mutations in Atg1 and Atg7 resulted in reduced DNA fragmentation in degenerating midstage egg chambers but did not appear to affect nuclear condensation, which indicates that autophagy contributes in part to cell death in the ovary. Our study provides new insights into the molecular mechanisms that coordinately regulate autophagic and apoptotic events in vivo. PMID:18794330

  14. Dual role of autophagy in HIV-1 replication and pathogenesis

    OpenAIRE

    Killian M

    2012-01-01

    Abstract Autophagy, the major mechanism for degrading long-lived intracellular proteins and organelles, is essential for eukaryotic cell homeostasis. Autophagy also defends the cell against invasion by microorganisms and has important roles in innate and adaptive immunity. Increasingly evident is that HIV-1 replication is dependent on select components of autophagy. Fittingly, HIV-1 proteins are able to modulate autophagy to maximize virus production. At the same time, HIV-1 proteins appear t...

  15. Bone Cell Autophagy Is Regulated by Environmental Factors

    OpenAIRE

    Zahm, Adam M.; Bohensky, Jolene; Adams, Christopher S.; Shapiro, Irving M.; Srinivas, Vickram

    2011-01-01

    The goal of this investigation was to ascertain whether bone cells undergo autophagy and to determine if this process is regulated by environmental factors. We showed that osteocytes in both murine and human cortical bone display a punctuate distribution of microtubule-associated protein light chain 3, indicative of autophagy. In addition, we noted a basal level of autophagy in preosteocyte-like murine long bone-derived osteocytic (MLO)-A5 cells. Autophagy was upregulated following nutrient d...

  16. Interactions between Shigella flexneri and the Autophagy Machinery

    OpenAIRE

    Krokowski, Sina; Mostowy, Serge

    2016-01-01

    Autophagy, an intracellular degradation process, is increasingly recognized as having important roles in host defense. Interactions between Shigella flexneri and the autophagy machinery were first discovered in 2005. Since then, work has shown that multiple autophagy pathways are triggered by S. flexneri, and autophagic responses can have different roles during Shigella infection. Here, we review the interactions between S. flexneri and the autophagy machinery, highlighting that studies using...

  17. Hepatitis B and C virus-induced hepatitis: Apoptosis, autophagy, and unfolded protein response

    Science.gov (United States)

    Yeganeh, Behzad; Rezaei Moghadam, Adel; Alizadeh, Javad; Wiechec, Emilia; Alavian, Seyed Moayed; Hashemi, Mohammad; Geramizadeh, Bita; Samali, Afshin; Bagheri Lankarani, Kamran; Post, Martin; Peymani, Payam; Coombs, Kevin M; Ghavami, Saeid

    2015-01-01

    AIM: To investigate the co-incidence of apoptosis, autophagy, and unfolded protein response (UPR) in hepatitis B (HBV) and C (HCV) infected hepatocytes. METHODS: We performed immunofluorescence confocal microscopy on 10 liver biopsies from HBV and HCV patients and tissue microarrays of HBV positive liver samples. We used specific antibodies for LC3β, cleaved caspase-3, BIP (GRP78), and XBP1 to detect autophagy, apoptosis and UPR, respectively. Anti-HCV NS3 and anti-HBs antibodies were also used to confirm infection. We performed triple blind counting of events to determine the co-incidence of autophagy (LC3β punctuate), apoptosis (cleaved caspase-3), and unfolded protein response (GRP78) with HBV and HCV infection in hepatocytes. All statistical analyses were performed using SPSS software for Windows (Version 16 SPSS Inc, Chicago, IL, United States). P-values autophagy, apoptosis, and UPR in HBV- and HCV-infected cells and adjacent non-infected cells. RESULTS: Our results showed that infection of hepatocytes with either HBV and HCV induces significant increase (P autophagy (LC3β punctate), and UPR (increase in GRP78 expression) in the HCV- and HBV-infected cells, as compared to non-infected cells of the same biopsy sections. Our tissue microarray immunohistochemical expression analysis of LC3β in HBVNeg and HBVPos revealed that majority of HBV-infected hepatocytes display strong positive staining for LC3β. Interestingly, although XBP splicing in HBV-infected cells was significantly higher (P 0.05). Furthermore, our evaluation of patients with HBV and HCV infection based on stage and grade of the liver diseases revealed no correlation between these pathological findings and induction of apoptosis, autophagy, and UPR. CONCLUSION: The results of this study indicate that HCV and HBV infection activates apoptosis, autophagy and UPR, but slightly differently by each virus. Further studies are warranted to elucidate the interconnections between these pathways in

  18. Streptococcus pneumoniae induces pyroptosis through the regulation of autophagy in murine microglia.

    Science.gov (United States)

    Kim, Ji-Yun; Paton, James C; Briles, David E; Rhee, Dong-Kwon; Pyo, Suhkneung

    2015-12-29

    Streptococcus pneumoniae is responsible for significant mortality and morbidity worldwide and causes invasive pneumococcal diseases including pneumococcal meningitis. Pyroptosis is caspase-1-dependent inflammatory cell death and is known to be induced by various microbial infections. In the present study, we investigated the molecular mechanisms that regulate pyroptosis induced by S. pneumoniae in microglia. Our results revealed that S. pneumoniae induced pyroptosis through caspase-1 activation and IL-1β production. We also found that the activation of caspase-1 and the maturation of IL-1β and IL-18 in the S. pneumoniae-triggered pyroptotic cell death process were mediated by NLRP3 inflammasome. In addition, pneumococcal infection increased the expression of autophagy-related genes and induced autophagosome formation. We also showed that the inhibition of autophagy promoted pneumococcus-induced pyroptosis. Furthermore, ROS was generated by pneumococcal infection and inhibited caspase-1 activation within 4 h of infection. However, in the late phase of infection, IL-1β secretion and caspase-1-dependent cell death were induced by ROS. These results suggest that autophagy induction transiently delay pyroptosis induced by S. pneumoniae in microglia. Our study also revealed that the activation of caspase-1 and the production of IL-1β were induced by pneumolysin and that pneumolysin triggered pyroptosis in microglial cells. Similar to the in vitro results, S. pneumoniae induced caspase-1 activation and caspase-1-dependent cytokine maturation in the mouse meningitis model. Thus, the present data demonstrate that S. pneumoniae induces pyroptosis in murine microglia and that NLRP3 inflammasome is critical for caspase-1 activation during the process. Furthermore, the induction of autophagy could transiently protect microglia from pyroptosis. PMID:26683708

  19. Lipocalin-2 inhibits autophagy and induces insulin resistance in H9c2 cells.

    Science.gov (United States)

    Chan, Yee Kwan; Sung, Hye Kyoung; Jahng, James Won Suk; Kim, Grace Ha Eun; Han, Meng; Sweeney, Gary

    2016-07-15

    Lipocalin-2 (Lcn2; also known as neutrophil gelatinase associated lipocalin, NGAL) levels are increased in obesity and diabetes and associate with insulin resistance. Correlations exist between Lcn2 levels and various forms or stages of heart failure. Insulin resistance and autophagy both play well-established roles in cardiomyopathy. However, little is known about the impact of Lcn2 on insulin signaling in cardiomyocytes. In this study, we treated H9c2 cells with recombinant Lcn2 for 1 h followed by dose- and time-dependent insulin treatment and found that Lcn2 attenuated insulin signaling assessed via phosphorylation of Akt and p70S6K. We used multiple assays to demonstrate that Lcn2 reduced autophagic flux. First, Lcn2 reduced pULK1 S555, increased pULK1 S757 and reduced LC3-II levels determined by Western blotting. We validated the use of DQ-BSA to assess autolysosomal protein degradation and this together with MagicRed cathepsin B assay indicated that Lcn2 reduced lysosomal degradative activity. Furthermore, we generated H9c2 cells stably expressing tandem fluorescent RFP/GFP-LC3 and this approach verified that Lcn2 decreased autophagic flux. We also created an autophagy-deficient H9c2 cell model by overexpressing a dominant-negative Atg5 mutant and found that reduced autophagy levels also induced insulin resistance. Adding rapamycin after Lcn2 could stimulate autophagy and recover insulin sensitivity. In conclusion, our study indicated that acute Lcn2 treatment caused insulin resistance and use of gain and loss of function approaches elucidated a causative link between autophagy inhibition and regulation of insulin sensitivity by Lcn2. PMID:27090568

  20. Autophagy and Retromer Components in Plant Innate Immunity

    DEFF Research Database (Denmark)

    Munch, David

    -hormone salicylic acid. Here, I present data that make it clear that NPR1 does not directly regulate autophagy, but instead control stress responses that indirectly activate autophagy. The observations presented will also clarify why autophagy has been described as being both a pro-death and pro-life pathway under...

  1. Autophagy induced by HIF1α overexpression supports trophoblast invasion by supplying cellular energy.

    Directory of Open Access Journals (Sweden)

    Mikiko Yamanaka-Tatematsu

    Full Text Available Extravillous trophoblasts (EVTs characterize the invasion of the maternal decidua under low oxygen and poor nutrition at the early feto-maternal interface to establish a successful pregnancy. We previously reported that autophagy in EVTs was activated under 2% O2 in vitro, and autophagy activation was also observed in EVTs at the early feto-maternal interface in vivo. Here, we show that autophagy is an energy source for the invasion of EVTs. Cobalt chloride (CoCl2, which induces hypoxia inducible factor 1α (HIF1α overexpression, activated autophagy in HTR8/SVneo cells, an EVT cell line. The number of invading HTR8-ATG4B(C74A cells, an autophagy-deficient EVT cell line, was markedly reduced by 81 percent with the CoCl2 treatment through the suppression of MMP9 level, although CoCl2 did not affect the cellular invasion of HTR8-mStrawberry cells, a control cell line. HTR8-ATG4B(C74A cells treated with CoCl2 showed a decrease in cellular adenosine triphosphate (ATP levels and a compensatory increase in the expression of purinergic receptor P2X ligand-gated ion channel 7 (P2RX7, which is stimulated with ATP, whereas HTR8-mStrawberry cells maintained cellular ATP levels and did not affect P2RX7 expression. Furthermore, the decreased invasiveness of HTR8-ATG4B(C74A cells treated with CoCl2 was neutralized by ATP supplementation to the level of HTR8-ATG4B(C74A cells treated without CoCl2. These results suggest that autophagy plays a role in maintaining homeostasis by countervailing HIF1α-mediated cellular energy consumption in EVTs.

  2. Role of autophagy in the bimodal stage after spinal cord ischemia reperfusion injury in rats.

    Science.gov (United States)

    Fang, Bo; Li, Xiao-Qian; Bao, Na-Ren; Tan, Wen-Fei; Chen, Feng-Shou; Pi, Xiao-Li; Zhang, Ying; Ma, Hong

    2016-07-22

    Autophagy plays an important role in spinal cord ischemia reperfusion (I/R) injury, but its neuroprotective or neurodegenerative role remains controversial. The extent and persistence of autophagy activation may be the critical factor to explain the opposing effects. In this study, the different roles and action mechanisms of autophagy in the early and later stages after I/R injury were investigated in rats. Thespinal cord I/R injury was induced by 14-min occlusion of the aortic arch, after which rats were treated with autophagic inhibitor (3-methyladenine, 3-MA) or agonist (rapamycin) immediately or 48h following the injury. Autophagy markers, microtubule-associated protein light chain 3-II (LC3-II) and Beclin 1 increased and peaked at the early stage (8h) and the later stage (72h) after spinal cord I/R injury. Beclin 1 was mostly expressed in neurons, but was also expressed to an extent in astrocytes, microglia and vascular endothelial cells. 8h after injury, rats treated with 3-MA showed a decrease in the hind-limb Basso-Beattie-Bresnahan (BBB) motor function scores, surviving motor neurons, and B-cell lymphoma-2 (Bcl-2) expression, and increase in the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end labeling (TUNEL)-positive cells, Bcl-2-associated X protein (Bax), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) expression, and activation of microglia, while those treated with rapamycin showed opposing effects. However, 72h after injury, rats treated with 3-MA improved the BBB scores, and the surviving motor neurons, and reduced the autophagic cell death, while those treated with rapamycin had adverse effects. These findings provide the first evidence that early activated autophagy alleviates spinal cord I/R injury via inhibiting apoptosis and inflammation; however later excessively elevated autophagy aggravates I/R injury through inducing autophagic cell death. PMID:27109922

  3. Protein kinase B/Akt1 inhibits autophagy by down-regulating UVRAG expression

    International Nuclear Information System (INIS)

    Autophagy, or autophagocytosis, is a selective intracellular degradative process involving the cell's own lysosomal apparatus. An essential component in cell development, homeostasis, repair and resistance to stress, autophagy may result in either cell death or survival. The targeted region of the cell is sequestered within a membrane structure, the autophagosome, for regulation of the catabolic process. A key factor in both autophagosome formation and autophagosome maturation is a protein encoded by the ultraviolet irradiation resistance-associated gene (UVRAG). Conversely, the serine/threonine-specific protein kinase B (PKB, also known as Akt), which regulates survival in various cancers, inhibits autophagy through mTOR activation. We found that Akt1 may also directly inhibit autophagy by down-regulating UVRAG both in a 293T transient transfection system and breast cancer cells stably expressing Akt1. The UVRAG with mutations at putative Akt1-phosphorylation sites were still inhibited by Akt1, and dominant-negative Akt1 also inhibited UVRAG expression, suggesting that Akt1 down-regulates UVRAG by a kinase activity-independent mechanism. We showed that Akt1 overexpression in MDA-MB-231 breast cancer cells down-regulated UVRAG transcription. Cells over-expressing Akt1 were more resistant than control cells to ultraviolet light-induced autophagy and exhibited the associated reduction in cell viability. Levels of the autophagosome indicator protein LC3B-II and mRFP-GFP-LC3 were reduced in cells that over-expressing Akt1. Inhibiting Akt1 by siRNA or reintroducing UVRAG gene rescued the level of LC3B-II in UV-irradiation. Altogether, these data suggest that Akt1 may inhibit autophagy by decreasing UVRAG expression, which also sensitizes cancer cells to UV irradiation.

  4. Autophagy activation prevents sevoflurane-induced neurotoxicity in H4 human neuroglioma cells

    Science.gov (United States)

    Zhou, You-fa; Wang, Qing-xia; Zhou, Hai-yan; Chen, Gang

    2016-01-01

    Aim: The inhaled anesthetic sevoflurane may induce cognitive impairment in both animals and humans. Previous study has shown that sevoflurane triggers ER stress and may lead to apoptosis in rat hippocampal neurons. In this study, we examined whether sevoflurane caused autophagy and its contributions to sevoflurane induced neuronal cell injury. Methods: H4 human neuroglioma cells were exposed to 4.1% sevoflurane for 6 h. Cell viability and apoptosis ratio were assessed using a CCK8 kit and flow cytometry, respectively. Autophagosomes in the cells were detected using GFP-LC3 plasmid transfection or transmission electronic microscopy. The expression of LC3B, p62/SQSTM, C/EBP homologous protein (CHOP) and glucose-related protein 78 (GRP78) was assessed with Western blotting. Results: Sevoflurane treatment induced apoptosis and markedly increased the LC3-II level and GFP-LC3 puncta number, decreased p62 expression in H4 cells. Activation of autophagy by rapamycin (1 μmol/L) significantly reduced sevoflurane-induced apoptosis and increased cell viability, whereas inhibition of autophagy with 3-MA (5 mmol/L) caused the opposite effects. Furthermore, sevoflurane treatment markedly increased the expression of CHOP and GRP78, two hallmark proteins of ER stress. Inhibition of ER stress by 4-phenylbutyrate (500 μmol/L) abrogated sevoflurane-induced autophagy and apoptosis, and improved the viability. Moreover, sevoflurane-stimulated expression of CHOP and GRP78 was inhibited by rapamycin, but further enhanced by 3-MA. Conclusion: Sevoflurane treatment induces ER stress and activates autophagy, which antagonizes sevoflurane-induced apoptosis in H4 human neuroglioma cells. The results suggest that autophagy may be a potential therapeutic target in preventing sevoflurane-induced neurotoxicity. PMID:27041458

  5. GLP-1 analogue improves hepatic lipid accumulation by inducing autophagy via AMPK/mTOR pathway.

    Science.gov (United States)

    He, Qin; Sha, Sha; Sun, Lei; Zhang, Jing; Dong, Ming

    2016-08-01

    The incidence of nonalcoholic fatty liver disease (NAFLD) keeps rising year by year, and NAFLD is rapidly becoming the most common liver disease worldwide. Clinical studies have found that glucagon-like peptide-1 (GLP-1) analogue, liraglutide (LRG), cannot only reduce glucose levels, but also improve hepatic lipase, especially in patients also with type 2 diabetes mellitus (T2DM). In addition, enhancing autophagy decreases lipid accumulation in hepatocytes. The aim of the present study is to explore the effect of LRG on hepatocyte steatosis and the possible role of autophagy. We set up an obesity mouse model with a high-fat diet (HFD) and induced hepatocyte steatosis with free fatty acids (FFA) in human L-O2 cells. LRG and two inhibitors of autophagy, Chloroquine (CQ) and bafilomycin A1 (Baf), were added into each group, respectively. The lipid profiles and morphological modifications of each group were tested. Immunohistochemistry, immunofluorescence staining and transmission electron microscopy (TEM) were used to measure autophagy in this study. The autophagy protein expression of SQSTM1 (P62), and LC3B, along with the signaling pathway proteins of mTOR, phosphorylated mTOR (p-mTOR), AMPK, phosphorylated AMPK (p-AMPK) and Beclin1, were evaluated by western blot. Our results showed that LRG improved hepatocyte steatosis by inducing autophagy, and the AMPK/mTOR pathway is involved. These findings suggest an important mechanism for the positive effects of LRG on hepatic steatosis, and provide new evidence for clinical use of LRG in NAFLD. PMID:27208776

  6. Protein kinase B/Akt1 inhibits autophagy by down-regulating UVRAG expression

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Wonseok; Ju, Ji-hyun; Lee, Kyung-min; Nam, KeeSoo; Oh, Sunhwa [Department of Life Science, College of Natural Science, Hanyang University, 17 Haengdang-dong, Seongdong-gu, Seoul 133-791 (Korea, Republic of); Shin, Incheol, E-mail: incheol@hanyang.ac.kr [Department of Life Science, College of Natural Science, Hanyang University, 17 Haengdang-dong, Seongdong-gu, Seoul 133-791 (Korea, Republic of)

    2013-02-01

    Autophagy, or autophagocytosis, is a selective intracellular degradative process involving the cell's own lysosomal apparatus. An essential component in cell development, homeostasis, repair and resistance to stress, autophagy may result in either cell death or survival. The targeted region of the cell is sequestered within a membrane structure, the autophagosome, for regulation of the catabolic process. A key factor in both autophagosome formation and autophagosome maturation is a protein encoded by the ultraviolet irradiation resistance-associated gene (UVRAG). Conversely, the serine/threonine-specific protein kinase B (PKB, also known as Akt), which regulates survival in various cancers, inhibits autophagy through mTOR activation. We found that Akt1 may also directly inhibit autophagy by down-regulating UVRAG both in a 293T transient transfection system and breast cancer cells stably expressing Akt1. The UVRAG with mutations at putative Akt1-phosphorylation sites were still inhibited by Akt1, and dominant-negative Akt1 also inhibited UVRAG expression, suggesting that Akt1 down-regulates UVRAG by a kinase activity-independent mechanism. We showed that Akt1 overexpression in MDA-MB-231 breast cancer cells down-regulated UVRAG transcription. Cells over-expressing Akt1 were more resistant than control cells to ultraviolet light-induced autophagy and exhibited the associated reduction in cell viability. Levels of the autophagosome indicator protein LC3B-II and mRFP-GFP-LC3 were reduced in cells that over-expressing Akt1. Inhibiting Akt1 by siRNA or reintroducing UVRAG gene rescued the level of LC3B-II in UV-irradiation. Altogether, these data suggest that Akt1 may inhibit autophagy by decreasing UVRAG expression, which also sensitizes cancer cells to UV irradiation.

  7. Andrographolide Analogue Induces Apoptosis and Autophagy Mediated Cell Death in U937 Cells by Inhibition of PI3K/Akt/mTOR Pathway.

    Directory of Open Access Journals (Sweden)

    Deepak Kumar

    Full Text Available Current chemotherapeutic agents based on apoptosis induction are lacking in desired efficacy. Therefore, there is continuous effort to bring about new dimension in control and gradual eradication of cancer by means of ever evolving therapeutic strategies. Various forms of PCD are being increasingly implicated in anti-cancer therapy and the complex interplay among them is vital for the ultimate fate of proliferating cells. We elaborated and illustrated the underlying mechanism of the most potent Andrographolide analogue (AG-4 mediated action that involved the induction of dual modes of cell death-apoptosis and autophagy in human leukemic U937 cells.AG-4 induced cytotoxicity was associated with redox imbalance and apoptosis which involved mitochondrial depolarisation, altered apoptotic protein expressions, activation of the caspase cascade leading to cell cycle arrest. Incubation with caspase inhibitor Z-VAD-fmk or Bax siRNA decreased cytotoxic efficacy of AG-4 emphasising critical roles of caspase and Bax. In addition, AG-4 induced autophagy as evident from LC3-II accumulation, increased Atg protein expressions and autophagosome formation. Pre-treatment with 3-MA or Atg 5 siRNA suppressed the cytotoxic effect of AG-4 implying the pro-death role of autophagy. Furthermore, incubation with Z-VAD-fmk or Bax siRNA subdued AG-4 induced autophagy and pre-treatment with 3-MA or Atg 5 siRNA curbed AG-4 induced apoptosis-implying that apoptosis and autophagy acted as partners in the context of AG-4 mediated action. AG-4 also inhibited PI3K/Akt/mTOR pathway. Inhibition of mTOR or Akt augmented AG-4 induced apoptosis and autophagy signifying its crucial role in its mechanism of action.Thus, these findings prove the dual ability of AG-4 to induce apoptosis and autophagy which provide a new perspective to it as a potential molecule targeting PCD for future cancer therapeutics.

  8. Autophagy and Autoimmunity CrossTalks

    Directory of Open Access Journals (Sweden)

    Abhisek eBhattacharya

    2013-04-01

    Full Text Available Autophagy, initially viewed as a conserved bulk-degradation mechanism, has emerged as a central player in a multitude of immune functions. Autophagy is important in host defense against intracellular and extracellular pathogens, metabolic syndromes, immune cell homeostasis, antigen processing and presentation and maintenance of tolerance. The observation that the above processes are implicated in triggering or exacerbating autoimmunity raises the possibility that the autophagy pathway is involved in mediating autoimmune processes, either directly or as a consequence of innate or adaptive functions mediated by the pathway. Genome-wide association studies have shown association between single nucleotide polymorphisms (SNPs in autophagy related gene 5 (Atg5, and Atg16l1 with susceptibility to systemic lupus erythematous (SLE and Crohn’s disease, respectively. Enhanced expression of Atg5 was also reported in blood of mice with experimental autoimmune encephalomyelitis (EAE, a mouse model of multiple sclerosis (MS, and in T cells isolated from blood or brain tissues from patients with active relapse of MS. This review explores the roles of autophagy pathway in the innate and adaptive immune systems on regulating or mediating the onset, progression or exacerbation of autoimmune processes.

  9. Autophagy and Liver Ischemia-Reperfusion Injury

    Directory of Open Access Journals (Sweden)

    Raffaele Cursio

    2015-01-01

    Full Text Available Liver ischemia-reperfusion (I-R injury occurs during liver resection, liver transplantation, and hemorrhagic shock. The main mode of liver cell death after warm and/or cold liver I-R is necrosis, but other modes of cell death, as apoptosis and autophagy, are also involved. Autophagy is an intracellular self-digesting pathway responsible for removal of long-lived proteins, damaged organelles, and malformed proteins during biosynthesis by lysosomes. Autophagy is found in normal and diseased liver. Although depending on the type of ischemia, warm and/or cold, the dynamic process of liver I-R results mainly in adenosine triphosphate depletion and in production of reactive oxygen species (ROS, leads to both, a local ischemic insult and an acute inflammatory-mediated reperfusion injury, and results finally in cell death. This process can induce liver dysfunction and can increase patient morbidity and mortality after liver surgery and hemorrhagic shock. Whether autophagy protects from or promotes liver injury following warm and/or cold I-R remains to be elucidated. The present review aims to summarize the current knowledge in liver I-R injury focusing on both the beneficial and the detrimental effects of liver autophagy following warm and/or cold liver I-R.

  10. Autophagy is required during cycling hypoxia to lower production of reactive oxygen species

    International Nuclear Information System (INIS)

    Background and purpose: Human tumors are characterized by the presence of cells that experience periodic episodes of hypoxia followed by reoxygenation. These cells are exposed to reactive oxygen species (ROS) upon reoxygenation and require adaptation to this stress by lowering ROS production or enhancing ROS-clearance for their survival. We hypothesized that autophagy, a lysosomal degradation pathway, may be involved in reducing ROS during periodic hypoxia through removal of ROS producing species. Materials and methods: Human tumor cells (MCF-7, HT29, U373) were exposed to cycles of hypoxia (O2 < 0.02%) and reoxygenation in the absence or presence of the autophagy inhibitor chloroquine (CQ). Clonogenic survival, ROS production and mitochondrial-DNA content were assessed. In addition, A549 cells overexpressing wild-type or K63-mutated ubiquitin (K63R) were analyzed for ROS production. Results: Our data indicate that CQ treatment sensitizes cells to cycling hypoxia, due to increased production of ROS, associated with an incapacity to reduce mitochondrial content. Addition of the ROS-scavenger N-acetyl-cysteine increased cell viability and neutralized CQ-effects. Additionally, genetic prevention of K63-linked ubiquitin chains that are required for the removal of toxic protein aggregates by autophagy, resulted in increased ROS production. Conclusions: Inhibition of autophagy substantially increases cell death induced by cycling hypoxia through increased ROS production, providing an opportunity to decrease the hypoxic fraction within tumors and enhance tumor therapy.

  11. Exercise induces autophagy in peripheral tissues and in the brain

    OpenAIRE

    He, Congcong; Sumpter, Jr., Rhea; Levine, Beth

    2012-01-01

    We recently identified physical exercise as a newly defined inducer of autophagy in vivo. Exercise induced autophagy in multiple organs involved in metabolic regulation, such as muscle, liver, pancreas and adipose tissue. To study the physiological role of exercise-induced autophagy, we generated mice with a knock-in nonphosphorylatable mutation in BCL2 (Thr69Ala, Ser70Ala and Ser84Ala) (BCL2 AAA) that are defective in exercise- and starvation-induced autophagy but not in basal autophagy. We ...

  12. Autophagy in the light of sphingolipid metabolism

    DEFF Research Database (Denmark)

    Harvald, Eva Bang; Olsen, Anne Sofie Braun; Færgeman, Nils J.

    2015-01-01

    Maintenance of cellular homeostasis requires tight and coordinated control of numerous metabolic pathways, which are governed by interconnected networks of signaling pathways and energy-sensing regulators. Autophagy, a lysosomal degradation pathway by which the cell self-digests its own components......, has over the past decade been recognized as an essential part of metabolism. Autophagy not only rids the cell of excessive or damaged organelles, misfolded proteins, and invading microorganisms, it also provides nutrients to maintain crucial cellular functions. Besides serving as essential structural...... moieties of biomembranes, lipids including sphingolipids are increasingly being recognized as central regulators of a number of important cellular processes, including autophagy. In the present review we describe how sphingolipids, with special emphasis on ceramides and sphingosine-1-phosphate, can act as...

  13. Heme oxygenase‑1 protects H2O2‑insulted glomerular mesangial cells from excessive autophagy.

    Science.gov (United States)

    Xu, Jia; Li, Jinshun; Wang, Jinhua; Chi, Yanchun; Zhang, Kun; Cui, Rui

    2016-06-01

    Increasing evidence has demonstrated that the activation of heme oxygenase (HO)‑1 reduces autophagy stimulated by oxidative stress injury, in which the supraphysiological production of reactive oxygen species (ROS) is detected. However, the potential mechanism underlying this effect remains unclear. The present study aimed to investigate the function of HO‑1 activation in the regulation of autophagy in glomerular mesangial cells subjected to H2O2‑induced oxidative stress injury. The results demonstrated that the HO‑1 agonist, hemin, reduces the LC3 protein level, which was enhanced by H2O2 treatment. Furthermore, hemin‑activated HO‑1 may function as a regulator of oxidative stress‑induced autophagy in a dose‑dependent manner. Pharmacological activation of c‑Jun N‑terminal kinase (JNK) inhibited the effect of hemin, indicating that the JNK signaling pathway is associated with the mechanism of HO‑1 in impeding excessive autophagy. In addition to successfully alleviating H2O2‑induced oxidative stress and cellular apoptosis, hemin‑activated HO‑1 may provide cytoprotection against rapamycin, a specific autophagy agonist. The present result suggested the inhibitory action of HO‑1 in the avoidance of a potentially enhanced linkage between autophagy and apoptosis, particularly in the setting of excessive ROS. Therefore, enhancing the intracellular activity of HO‑1 may assist the crosstalk between oxidative stress, autophagy and apoptosis, and represent a novel therapeutic strategy for renal ischemic disease. PMID:27122182

  14. What to Eat: Evidence for Selective Autophagy in Plants

    Institute of Scientific and Technical Information of China (English)

    Brice E.Floyd; Stephanie C.Morriss; Gustavo C.Maclntosh; Diane C.Bassham

    2012-01-01

    Autophagy is a macromolecular degradation pathway by which cells recycle their contents as a developmental process,house-keeping mechanism,and response to environmental stress.In plants,autophagy involves the sequestration of cargo to be degraded,transport to the cell vacuole in a double-membrane bound autophagosome,and subsequent degradation by lytic enzymes.Autophagy has generally been considered to be a non-selective mechanism of degradation.However,studies in yeast and animals have found numerous examples of selective autophagy,with cargo including proteins,protein aggregates,and organelles.Recent work has also provided evidence for several types of selective autophagy in plants.The degradation of protein aggregates was the first selective autophagy described in plants,and,more recently,a hybrid protein of the mammalian selective autophagy adaptors p62 and NBR1,which interacts with the autophagy machinery and may function in autophagy of protein aggregates,was described in plants.Other intracellular components have been suggested to be selectively targeted by autophagy in plants,but the current evidence is limited.Here,we discuss recent findings regarding the selective targeting of cell components by autophagy in plants.

  15. Dehydroandrographolide, an iNOS inhibitor, extracted from Andrographis paniculata (Burm.f.) Nees, induces autophagy in human oral cancer cells.

    Science.gov (United States)

    Hsieh, Ming-Ju; Lin, Chiao-Wen; Chiou, Hui-Ling; Yang, Shun-Fa; Chen, Mu-Kuan

    2015-10-13

    Autophagy, which is constitutively executed at the basal level in all cells, promotes cellular homeostasis by regulating the turnover of organelles and proteins. Andrographolide and dehydroandrographolide (DA) are the two principle components of Andrographis paniculata (Burm.f.) Nees. and are the main contributors to its therapeutic properties. However, the pharmacological activities of dehydroandrographolide (DA) remain unclear. In this study, DA induces oral cancer cell death by activating autophagy. Treatment with autophagy inhibitors inhibited DA-induced human oral cancer cell death. In addition, DA increased LC3-II expression and reduced p53 expression in a time- and concentration-dependent manner. Furthermore, DA induced autophagy and decreased cell viability through modulation of p53 expression. DA-induced autophagy was triggered by an activation of JNK1/2 and an inhibition of Akt and p38. In conclusion, this study demonstrated that DA induced autophagy in human oral cancer cells by modulating p53 expression, activating JNK1/2, and inhibiting Akt and p38. Finally, an administration of DA effectively suppressed the tumor formation in the oral carcinoma xenograft model in vivo. This is the first study to reveal the novel function of DA in activating autophagy, suggesting that DA could serve as a new and potential chemopreventive agent for treating human oral cancer. PMID:26356821

  16. Cisplatin-induced downregulation of miR-199a-5p increases drug resistance by activating autophagy in HCC cell

    International Nuclear Information System (INIS)

    Highlights: ► miR-199a-5p levels were significantly decreased after cisplatin treatment. ► Cisplatin treatment induced autophagy activation. ► Cisplatin-induced downregulation of miR-199a-5p increases drug resistance by activating autophagy in HCC cell. -- Abstract: Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide. Systemic chemotherapy plays an important role in the treatment of patients with advanced liver cancer. However, chemoresistance to cisplatin is a major limitation of cisplatin-based chemotherapy in the clinic, and the underlying mechanism of such resistance is not fully understood. In the study, we found that miR-199a-5p levels were significantly reduced in HCC patients treated with cisplatin-based chemotherapy. Cisplatin treatment also resulted in decreased miR-199a-5p levels in human HCC cell lines. Forced expression of miR-199a-5p promoted cisplatin-induced inhibition of cell proliferation. Cisplatin treatment activated autophagy in Huh7 and HepG2 cells, which increased cell proliferation. We further demonstrated that downregulated miR-199a-5p enhanced autophagy activation by targeting autophagy-associated gene 7 (ATG7). More important, autophagy inhibition abrogated miR-199a-5p downregulation-induced cell proliferation. These data demonstrated that miR-199a-5p/autophagy signaling represents a novel pathway regulating chemoresistance, thus offering a new target for chemotherapy of HCC.

  17. Autophagy: A double-edged sword in intervertebral disk degeneration.

    Science.gov (United States)

    Zhang, Shu-Jun; Yang, Wei; Wang, Cheng; He, Wen-Si; Deng, Hai-Yang; Yan, Yi-Guo; Zhang, Jian; Xiang, Yong-Xiao; Wang, Wen-Jun

    2016-06-01

    Autophagy is a homeostatic mechanism through which intracellular damaged organelles and proteins are degraded and recycled in response to increased metabolic demands or stresses. Although primarily cytoprotective, dysfunction of autophagy is often associated with many degenerative diseases, including intervertebral disc (IVD) degeneration (IDD). As a main contributing factor to low back pain, IDD is the pathological basis for various debilitating spinal diseases. Either higher or lower levels of autophagy are observed in degenerative IVD cells. Despite the precise role of autophagy in disc degeneration that is still controversial, with difference from protection to aggravation, targeting autophagy has shown promise for mitigating disc degeneration. In the current review, we summarize the changes of autophagy in degenerative IVD cells and mainly discuss the relationship between autophagy and IDD. With continued efforts, modulation of the autophagic process could be a potential and attractive therapeutic strategy for degenerative disc disease. PMID:27018178

  18. High glucose induces autophagy of MC3T3-E1 cells via ROS-AKT-mTOR axis.

    Science.gov (United States)

    Wang, Xiaoju; Feng, Zhengping; Li, Jiling; Chen, Lixue; Tang, Weixue

    2016-07-01

    In the present study, we investigate the function of ROS-AKT-mTOR axis on the apoptosis, proliferation and autophagy of MC3T3-E1 cells, and the proliferation of MC3T3-E1 cells after autophagy inhibition under high glucose conditions. MC3T3-E1 cells cultured in vitro were divided into the following groups: normal control group, N-acetylcysteine (NAC) group, 11.0 mM high glucose group, 11.0 mM high glucose + NAC group, 22.0 mM high glucose group, 22.0 mM high glucose + NAC group, CQ group, 22.0 mM high glucose + CQ group, 3-MA group and 3-MA + 22.0 mM high glucose group. ROS production was measured by DCFH-DA fluorescent probe. Cell proliferation was measured by MTT assay. Cells in different groups were stained with Annexin V-FITC/PI, and then apoptosis rate was detected by flow cytometry. Nucleus morphology was observed under fluorescence microscope after being incubated with Honchest33258. Protein expression was measured using Western blotting and immunofluorescence. Cell apoptosis and proliferation in high glucose group were increased and decreased, respectively, in a dose-dependent manner. Autophagy was significantly induced in high glucose group, even though different concentration of glucose induced autophagy in different stages of autophagy. ROS production in MC3T3-E1 cells was remarkably increased in high glucose group, but not in a dose-dependent manner. NAC, as an antioxidant, reduced ROS production and ameliorated cell apoptosis, proliferation abnormity and autophagy caused by high glucose. Expression of p-AKT and p-mTOR proteins were dramatically decreased in high glucose group, and NAC reversed their expression. In addition, 3-MA, an inhibitor of autophagy, significantly decreased the proliferation of MC3T3-E1 cells. When cocultured with 22.0 mM glucose that induced autophagy, proliferation of MC3T3-E1 cells was not affected compared to 22.0 mM high glucose group. Our present findings reveal that high glucose affects apoptosis

  19. Unification By Induction

    OpenAIRE

    Adewole, A. I. A.

    2001-01-01

    We show that the problem of unifying electromagnetism with gravity has an elegant solution in classical physics through the phenomenon of induction. By studying the way that induction leads to the formation of electromagnetic fields, we identify the classical field equations which the unified field must satisfy and a corresponding set of constitutive equations for the medium sustaining the field. The unification problem is then reduced to the problem of finding the exact form of these constit...

  20. LC3B is indispensable for selective autophagy of p62 but not basal autophagy

    International Nuclear Information System (INIS)

    Highlights: • Knockdown of LC3 or GABARAP families did not affect the basal autophagy. • LC3B has a higher affinity for the autophagy-specific substrate, p62, than GABARAPs. • siRNA-mediated knockdown of LC3B, but not that of GABARAPs, resulted in significant accumulation of p62. - Abstract: Autophagy is a unique intracellular protein degradation system accompanied by autophagosome formation. Besides its important role through bulk degradation in supplying nutrients, this system has an ability to degrade certain proteins, organelles, and invading bacteria selectively to maintain cellular homeostasis. In yeasts, Atg8p plays key roles in both autophagosome formation and selective autophagy based on its membrane fusion property and interaction with autophagy adaptors/specific substrates. In contrast to the single Atg8p in yeast, mammals have 6 homologs of Atg8p comprising LC3 and GABARAP families. However, it is not clear these two families have different or similar functions. The aim of this study was to determine the separate roles of LC3 and GABARAP families in basal/constitutive and/or selective autophagy. While the combined knockdown of LC3 and GABARAP families caused a defect in long-lived protein degradation through lysosomes, knockdown of each had no effect on the degradation. Meanwhile, knockdown of LC3B but not GABARAPs resulted in significant accumulation of p62/Sqstm1, one of the selective substrate for autophagy. Our results suggest that while mammalian Atg8 homologs are functionally redundant with regard to autophagosome formation, selective autophagy is regulated by specific Atg8 homologs

  1. LC3B is indispensable for selective autophagy of p62 but not basal autophagy

    Energy Technology Data Exchange (ETDEWEB)

    Maruyama, Yoko [Protein Metabolism Project, Tokyo Metropolitan Institute of Medical Science, Tokyo 156-8506 (Japan); Department of Pediatrics, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Sou, Yu-Shin; Kageyama, Shun [Protein Metabolism Project, Tokyo Metropolitan Institute of Medical Science, Tokyo 156-8506 (Japan); Takahashi, Takao [Department of Pediatrics, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Ueno, Takashi [Division of Proteomics and Biomolecular Science, Center for Biomedical Research Resources, Juntendo University Graduate School of Medicine, Tokyo 113-8421 (Japan); Tanaka, Keiji [Laboratory of Protein Metabolism, Tokyo Metropolitan Institute of Medical Science, Tokyo 156-8506 (Japan); Komatsu, Masaaki, E-mail: komatsu-ms@igakuken.or.jp [Protein Metabolism Project, Tokyo Metropolitan Institute of Medical Science, Tokyo 156-8506 (Japan); Department of Biochemistry, School of Medicine, Niigata University, Niigata 951-8510 (Japan); Ichimura, Yoshinobu, E-mail: ichimura-ys@igakuken.or.jp [Protein Metabolism Project, Tokyo Metropolitan Institute of Medical Science, Tokyo 156-8506 (Japan)

    2014-03-28

    Highlights: • Knockdown of LC3 or GABARAP families did not affect the basal autophagy. • LC3B has a higher affinity for the autophagy-specific substrate, p62, than GABARAPs. • siRNA-mediated knockdown of LC3B, but not that of GABARAPs, resulted in significant accumulation of p62. - Abstract: Autophagy is a unique intracellular protein degradation system accompanied by autophagosome formation. Besides its important role through bulk degradation in supplying nutrients, this system has an ability to degrade certain proteins, organelles, and invading bacteria selectively to maintain cellular homeostasis. In yeasts, Atg8p plays key roles in both autophagosome formation and selective autophagy based on its membrane fusion property and interaction with autophagy adaptors/specific substrates. In contrast to the single Atg8p in yeast, mammals have 6 homologs of Atg8p comprising LC3 and GABARAP families. However, it is not clear these two families have different or similar functions. The aim of this study was to determine the separate roles of LC3 and GABARAP families in basal/constitutive and/or selective autophagy. While the combined knockdown of LC3 and GABARAP families caused a defect in long-lived protein degradation through lysosomes, knockdown of each had no effect on the degradation. Meanwhile, knockdown of LC3B but not GABARAPs resulted in significant accumulation of p62/Sqstm1, one of the selective substrate for autophagy. Our results suggest that while mammalian Atg8 homologs are functionally redundant with regard to autophagosome formation, selective autophagy is regulated by specific Atg8 homologs.

  2. Loss of autophagy enhances MIF/macrophage migration inhibitory factor release by macrophages.

    Science.gov (United States)

    Lee, Jacinta P W; Foote, Andrew; Fan, Huapeng; Peral de Castro, Celia; Lang, Tali; Jones, Sarah A; Gavrilescu, Nichita; Mills, Kingston H G; Leech, Michelle; Morand, Eric F; Harris, James

    2016-06-01

    MIF (macrophage migration inhibitory factor [glycosylation-inhibiting factor]) is a pro-inflammatory cytokine expressed in multiple cells types, including macrophages. MIF plays a pathogenic role in a number of inflammatory diseases and has been linked to tumor progression in some cancers. Previous work has demonstrated that loss of autophagy in macrophages enhances secretion of IL1 family cytokines. Here, we demonstrate that loss of autophagy, by pharmacological inhibition or siRNA silencing of Atg5, enhances MIF secretion by monocytes and macrophages. We further demonstrate that this is dependent on mitochondrial reactive oxygen species (ROS). Induction of autophagy with MTOR inhibitors had no effect on MIF secretion, but amino acid starvation increased secretion. This was unaffected by Atg5 siRNA but was again dependent on mitochondrial ROS. Our data demonstrate that autophagic regulation of mitochondrial ROS plays a pivotal role in the regulation of inflammatory cytokine secretion in macrophages, with potential implications for the pathogenesis of inflammatory diseases and cancers. PMID:27163877

  3. Detection of Autophagy in Caenorhabditis elegans by Western Blotting Analysis of LGG-1.

    Science.gov (United States)

    Palmisano, Nicholas J; Meléndez, Alicia

    2016-01-01

    A common way to measure the induction of autophagy in yeast and mammalian cells is to compare the amount of Atg8/LC3-I with that of Atg8-PE/LC3-II by using western blot analysis. This is because changes in the amount of LC3-II correlate closely with changes in the number of autophagosomes present in cells. Atg8/LC3 is initially synthesized as an unprocessed form, which is proteolytically processed to form Atg8/LC3-I, and then this is modified into the phosphatidylethanolamine (PE)-conjugated Atg8-PE/LC3-II form. Atg8/LC3-II is membrane bound, whereas Atg8-PE/LC3-I is cytosolic. By associating with both the inner and outer membranes of the autophagosome, Atg8-PE/LC3-II is the only autophagy reporter that is reliably associated with completed autophagosomes. In the nematode Caenorhabditis elegans, the ortholog of Atg8/LC3 is LGG-1. Here, we discuss how changes in the levels of LGG-1-II (and the paralog LGG-2) protein can, with appropriate controls, be used to monitor autophagy activity in nematodes and present a protocol for monitoring changes in the protein levels of different forms of LGG-1 by western blotting. PMID:26832685

  4. MicroRNA-21 activation of ERK signaling via PTEN is involved in arsenite-induced autophagy in human hepatic L-02 cells.

    Science.gov (United States)

    Liu, Xinlu; Luo, Fei; Ling, Min; Lu, Lu; Shi, Le; Lu, Xiaolin; Xu, Hui; Chen, Chao; Yang, Qianlei; Xue, Junchao; Li, Jun; Zhang, Aihua; Liu, Qizhan

    2016-06-11

    Autophagy, an evolutionarily conserved cellular process, has diverse physiological and pathological roles in biological functions. Whether autophagy is induced by arsenite, a well-established human carcinogen, and the molecular mechanisms involved, remain to be established. Further, microRNAs (miRNAs) act as regulators in various cancers, but how miRNAs regulate autophagy remains largely unexplored. We have found that, in human hepatic epithelial (L-02) cells, arsenite increases levels of autophagy-related proteins in a concentration- and time-dependent manner and elevates the number of autophagic vacuoles (AVs). Arsenite also activates the ERK pathway in a dose- and time-dependent manner. In L-02 cells exposed to arsenite, microRNA-21 (miRNA-21) is over-expressed, and its target proteins, PTEN, PDCD4, and Spry1, are decreased. Moreover, inhibition of miR-21 increases levels of PTEN, and reduces levels of Beclin 1 and LC3 II/I, indicating that miR-21 is involved in arsenite-induced autophagy. In addition, ectopic expression of PTEN blocks the effect of miR-21 on the arsenite-induced autophagy and decreases p-ERK levels. Also, ERK promotes the autophagy induced by arsenite. In sum, upon exposure of cells to arsenite, over-expression of miR-21 activates ERK through PTEN, factors that participate in arsenite-induced autophagy. This link, mediated through miRNAs, establishes a mechanism for the development of autophagy that is associated with arsenic toxicity. Such information contributes to an understanding of the liver toxicity caused by arsenite. PMID:27107786

  5. Autophagy Alleviates Melamine-Induced Cell Death in PC12 Cells Via Decreasing ROS Level.

    Science.gov (United States)

    Wang, Hui; Gao, Na; Li, Zhigui; Yang, Zhuo; Zhang, Tao

    2016-04-01

    Since melamine was illegally added to raw milk for increasing the apparent protein content, such a scandal has not been quite blown out. Previous studies showed that melamine induced apoptosis and oxidative damage in both in vivo and in vitro experiments. It is well known that autophagy is closely related to oxidative stress. In the present study, we examined whether autophagy played an important role in protecting PC12 cells, which were damaged by melamine. Immunofluorescence assay showed that melamine enhanced the number of punctuate dot, indicating the increase of autophagosomes. Western blot assay presented that melamine significantly elevated the expression level of autophagy markers including LC3-II/LC3-I ratio, beclin-1, and Atg 7. Rapamycin further enhanced the effect, whereas 3-methyadenine (3-MA) inhibited it. MTT assay exhibited that rapamycin significantly enhanced the cell viability (P < 0.01), while 3-MA considerably reduced it in melamine-treated PC12 cells (P < 0.01). Furthermore, flow cytometry assay showed that rapamycin considerably reduced the reactive oxygen species (ROS) level of the cells (P < 0.01), but 3-MA increased the generation of ROS (P < 0.01). Additionally, the superoxide dismutase (SOD) activity was notably increased by rapamycin in melamine-treated PC12 cells (P < 0.01), while the activity of which was prominently decreased by 3-MA (P < 0.01). Malondialdehyde (MDA) assay showed that rapamycin remarkably decreased the MDA level of the cells (P < 0.05), while 3-MA increased it (P < 0.01). Consequently, this study demonstrated that autophagy protected PC12 cells from melamine-induced cell death via inhibiting the excessive generation of ROS. Regulating autophagy may become a new targeted therapy to relieve the damage induced by melamine. PMID:25724280

  6. Cationic poly(amidoamine) dendrimers induced cyto-protective autophagy in hepatocellular carcinoma cells

    International Nuclear Information System (INIS)

    Poly(amidoamine) (PAMAM) dendrimers are proposed as one of the most promising nanomaterials for biomedical applications because of their unique tree-like structure, monodispersity and tunable properties. In this study, we found that PAMAM dendrimers could induce the formation of autophagosomes and the conversion of microtubule-associated protein 1 light chain 3 (LC3) in hepatocellular carcinoma HepG2 cells, while the inhibition of the Akt/mTOR and activation of the Erk 1/2 signaling pathways were involved in autophagy-induced by PAMAM dendrimers. We also investigated the suppression of autophagy with the obviously enhanced cytotoxicity of PAMAM dendrimers. Moreover, the blockage of a reactive oxygen species (ROS) could enhance the growth inhibition and apoptosis of hepatocellular carcinoma cells, induced by PAMAM dendrimers through reducing autophagic effects. Taken together, these findings explored the role and mechanism of autophagy induced by PAMAM dendrimers in HepG2 cells, provided new insight into the effect of autophagy on drug delivery nanomaterials and tumor cells and contributed to the use of a drug delivery vehicle for hepatocellular carcinoma treatment. (paper)

  7. Cationic poly(amidoamine) dendrimers induced cyto-protective autophagy in hepatocellular carcinoma cells

    Science.gov (United States)

    Li, Yubin; Wang, Shaofei; Wang, Ziyu; Qian, Xiaolu; Fan, Jiajun; Zeng, Xian; Sun, Yun; Song, Ping; Feng, Meiqing; Ju, Dianwen

    2014-09-01

    Poly(amidoamine) (PAMAM) dendrimers are proposed as one of the most promising nanomaterials for biomedical applications because of their unique tree-like structure, monodispersity and tunable properties. In this study, we found that PAMAM dendrimers could induce the formation of autophagosomes and the conversion of microtubule-associated protein 1 light chain 3 (LC3) in hepatocellular carcinoma HepG2 cells, while the inhibition of the Akt/mTOR and activation of the Erk 1/2 signaling pathways were involved in autophagy-induced by PAMAM dendrimers. We also investigated the suppression of autophagy with the obviously enhanced cytotoxicity of PAMAM dendrimers. Moreover, the blockage of a reactive oxygen species (ROS) could enhance the growth inhibition and apoptosis of hepatocellular carcinoma cells, induced by PAMAM dendrimers through reducing autophagic effects. Taken together, these findings explored the role and mechanism of autophagy induced by PAMAM dendrimers in HepG2 cells, provided new insight into the effect of autophagy on drug delivery nanomaterials and tumor cells and contributed to the use of a drug delivery vehicle for hepatocellular carcinoma treatment.

  8. Nanoparticle-facilitated autophagy inhibition promotes the efficacy of chemotherapeutics against breast cancer stem cells.

    Science.gov (United States)

    Sun, Rong; Shen, Song; Zhang, Yun-Jiao; Xu, Cong-Fei; Cao, Zhi-Ting; Wen, Long-Ping; Wang, Jun

    2016-10-01

    Cancer stem cells (CSCs) have garnered increasing attention over the past decade, as they are believed to play a crucial role in tumor initiation, progression and metastasis, relapse and drug resistance. Therapeutic strategies which simultaneously exterminate both bulk tumor cells and the rare CSC subpopulation may produce striking response and result in long-term tumor remission. Accumulating evidence provides insight into the function of autophagy in maintenance, plasticity and survival of CSCs. The role of autophagy in the susceptibility of breast CSCs to chemotherapeutics was investigated in the present work, reduced 'stemness' and increased susceptibility to chemotherapy drugs (doxorubicin, DOX and docetaxel, DTXL) were observed after chloroquine (CQ)-mediated autophagy inhibition in sorted ALDH(hi) cells of breast cancer cell line MDA-MB-231. We further proved that nanoparticle-mediated autophagy inhibition promoted the efficacy of chemotherapeutics against ALDH(hi) MDA-MB-231 cells in vitro. Administration of drug delivery systems significantly prolonged the circulation half-life and augmented enrichment of two different drugs in tumor tissues and ALDH(hi) cells. More importantly, compared with single treatment, the combined delivery systems NPCQ/NPDOX and NPCQ/DOX (NPCQ/NPDTXL and NPCQ/DTXL) showed most effective and persistent tumor growth inhibitory effect by eliminating bulk tumor cells as well as CSCs (p breast cancer. PMID:27376558

  9. Regulation of autophagy in human skeletal muscle: effects of exercise, exercise training and insulin stimulation

    DEFF Research Database (Denmark)

    Fritzen, Andreas Mæchel; Madsen, Agnete Louise Bjerregaard; Kleinert, Maximilian;

    2016-01-01

    Studies in rodent muscle suggest that autophagy is regulated by acute exercise, exercise training and insulin stimulation. However, little is known about the regulation of autophagy in human skeletal muscle. Here we investigate the autophagic response to acute one-legged exercise, one-legged exer......Studies in rodent muscle suggest that autophagy is regulated by acute exercise, exercise training and insulin stimulation. However, little is known about the regulation of autophagy in human skeletal muscle. Here we investigate the autophagic response to acute one-legged exercise, one......-legged exercise training as well as in response to subsequent insulin stimulation in exercised and non-exercised human muscle. Acute one-legged exercise decreased (p<0.01) lipidation of LC3 (∼50 %) and the LC3-II/LC3-I ratio (∼60 %) indicating that content of autophagosomes decreases with exercise in human muscle....... The decrease in LC3-II/LC3-I ratio did not correlate with activation of AMPK trimer complexes in human muscle. Consistently, pharmacological AMPK activation with AICAR in mouse muscle did not affect the LC3-II/LC3-I ratio. Four hours after exercise, insulin further reduced (p<0.01) the LC3-II/LC3-I...

  10. Avermectin induced autophagy in pigeon spleen tissues.

    Science.gov (United States)

    Liu, Ci; Zhao, Yanbing; Chen, Lijie; Zhang, Ziwei; Li, Ming; Li, Shu

    2015-12-01

    The level of autophagy is considered as an indicator for monitoring the toxic impact of pesticide exposure. Avermectin (AVM), a widely used insecticide, has immunotoxic effects on the pigeon spleen. The aim of this study was to investigate the status of autophagy and the expression levels of microtubule-associated protein1 light chain 3 (LC3), beclin-1, dynein, autophagy associated gene (Atg) 4B, Atg5, target of rapamycin complex 1 (TORC1) and target of rapamycin complex 2 (TORC2) in AVM-treated pigeon spleens. Eighty two-month-old pigeons were randomly divided into four groups: a control group, a low-dose group, a medium-dose group and a high-dose group, which were fed a basal diet spiked with 0, 20, 40 and 60 mg AVM/kg diet, respectively. Microscopic cellular morphology revealed a significant increase in autophagic structures in the AVM-treated groups. The expression of LC3, beclin-1, dynein, Atg4B and Atg5 increased, while mRNA levels of TORC1 and TORC2 were decreased in the AVM-treated groups relative to the control groups at 30, 60 and 90 days in the pigeon spleen. These results indicated that AVM exposure could up-regulate the level of autophagy in a dose-time-dependent manner in the pigeon spleen. PMID:26541719

  11. MicroRNA regulation of Autophagy

    DEFF Research Database (Denmark)

    Frankel, Lisa B; Lund, Anders H

    2012-01-01

    progress has recently contributed to our understanding of the molecular mechanisms of the autophagy machinery, yet several gaps remain in our knowledge of this process. The discovery of microRNAs (miRNAs) established a new paradigm of post-transcriptional gene regulation and during the past decade these...

  12. Alpha-synuclein aggregation involves a bafilomycin A 1-sensitive autophagy pathway.

    Science.gov (United States)

    Klucken, Jochen; Poehler, Anne-Maria; Ebrahimi-Fakhari, Darius; Schneider, Jacqueline; Nuber, Silke; Rockenstein, Edward; Schlötzer-Schrehardt, Ursula; Hyman, Bradley T; McLean, Pamela J; Masliah, Eliezer; Winkler, Juergen

    2012-05-01

    Synucleinopathies like Parkinson disease and dementia with Lewy bodies (DLB) are characterized by α-synuclein aggregates within neurons (Lewy bodies) and their processes (Lewy neurites). Whereas α-synuclein has been genetically linked to the disease process, the pathological relevance of α-synuclein aggregates is still debated. Impaired degradation is considered to result in aggregation of α-synuclein. In addition to the ubiquitin-proteasome degradation, the autophagy-lysosomal pathway (ALP) is involved in intracellular degradation processes for α-synuclein. Here, we asked if modulation of ALP affects α-synuclein aggregation and toxicity. We have identified an induction of the ALP markers LAMP-2A and LC3-II in human brain tissue from DLB patients, in a transgenic mouse model of synucleinopathy, and in a cell culture model for α-synuclein aggregation. ALP inhibition using bafilomycin A 1 (BafA1) significantly potentiates toxicity of aggregated α-synuclein species in transgenic mice and in cell culture. Surprisingly, increased toxicity is paralleled by reduced aggregation in both in vivo and in vitro models. The dichotomy of effects on aggregating and nonaggregating species of α-synuclein was specifically sensitive to BafA1 and could not be reproduced by other ALP inhibitors. The present study expands on the accumulating evidence regarding the function of ALP for α-synuclein degradation by isolating an aggregation specific, BafA1-sensitive, ALP-related pathway. Our data also suggest that protein aggregation may represent a detoxifying event rather than being causal for cellular toxicity. PMID:22647715

  13. Liver autophagy in anorexia nervosa and acute liver injury.

    Science.gov (United States)

    Kheloufi, Marouane; Boulanger, Chantal M; Durand, François; Rautou, Pierre-Emmanuel

    2014-01-01

    Autophagy, a lysosomal catabolic pathway for long-lived proteins and damaged organelles, is crucial for cell homeostasis, and survival under stressful conditions. During starvation, autophagy is induced in numerous organisms ranging from yeast to mammals, and promotes survival by supplying nutrients and energy. In the early neonatal period, when transplacental nutrients supply is interrupted, starvation-induced autophagy is crucial for neonates' survival. In adult animals, autophagy provides amino acids and participates in glucose metabolism following starvation. In patients with anorexia nervosa, autophagy appears initially protective, allowing cells to copes with nutrient deprivation. However, when starvation is critically prolonged and when body mass index reaches 13 kg/m(2) or lower, acute liver insufficiency occurs with features of autophagic cell death, which can be observed by electron microscopy analysis of liver biopsy samples. In acetaminophen overdose, a classic cause of severe liver injury, autophagy is induced as a protective mechanism. Pharmacological enhancement of autophagy protects against acetaminophen-induced necrosis. Autophagy is also activated as a rescue mechanism in response to Efavirenz-induced mitochondrial dysfunction. However, Efavirenz overdose blocks autophagy leading to liver cell death. In conclusion, in acute liver injury, autophagy appears as a protective mechanism that can be however blocked or overwhelmed. PMID:25250330

  14. Impairment of autophagy: From hereditary disorder to drug intoxication

    International Nuclear Information System (INIS)

    At first, the molecular mechanism of autophagy was unveiled in a unicellular organism Saccharomyces cerevisiae (budding yeast), followed by the discovery that the basic mechanism of autophagy is conserved in multicellular organisms including mammals. Although autophagy was considered to be a non-selective bulk protein degradation system to recycle amino acids during periods of nutrient starvation, it is also believed to be an essential mechanism for the selective elimination of proteins/organelles that are damaged under pathological conditions. Research advances made using autophagy-deficient animals have revealed that impairments of autophagy often underlie the pathogenesis of hereditary disorders such as Danon, Parkinson's, Alzheimer's, and Huntington's diseases, and amyotrophic lateral sclerosis. On the other hand, there are many reports that drugs and toxicants, including arsenic, cadmium, paraquat, methamphetamine, and ethanol, induce autophagy during the development of their toxicity on many organs including heart, brain, lung, kidney, and liver. Although the question as to whether autophagic machinery is involved in the execution of cell death or not remains controversial, the current view of the role of autophagy during cell/tissue injury is that it is an important, often essential, cytoprotective reaction; disturbances in cytoprotective autophagy aggravate cell/tissue injuries. The purpose of this review is to provide (1) a gross summarization of autophagy processes, which are becoming more important in the field of toxicology, and (2) examples of important studies reporting the involvement of perturbations in autophagy in cell/tissue injuries caused by acute as well as chronic intoxication

  15. Liver Autophagy in Anorexia Nervosa and Acute Liver Injury

    Directory of Open Access Journals (Sweden)

    Marouane Kheloufi

    2014-01-01

    Full Text Available Autophagy, a lysosomal catabolic pathway for long-lived proteins and damaged organelles, is crucial for cell homeostasis, and survival under stressful conditions. During starvation, autophagy is induced in numerous organisms ranging from yeast to mammals, and promotes survival by supplying nutrients and energy. In the early neonatal period, when transplacental nutrients supply is interrupted, starvation-induced autophagy is crucial for neonates’ survival. In adult animals, autophagy provides amino acids and participates in glucose metabolism following starvation. In patients with anorexia nervosa, autophagy appears initially protective, allowing cells to copes with nutrient deprivation. However, when starvation is critically prolonged and when body mass index reaches 13 kg/m2 or lower, acute liver insufficiency occurs with features of autophagic cell death, which can be observed by electron microscopy analysis of liver biopsy samples. In acetaminophen overdose, a classic cause of severe liver injury, autophagy is induced as a protective mechanism. Pharmacological enhancement of autophagy protects against acetaminophen-induced necrosis. Autophagy is also activated as a rescue mechanism in response to Efavirenz-induced mitochondrial dysfunction. However, Efavirenz overdose blocks autophagy leading to liver cell death. In conclusion, in acute liver injury, autophagy appears as a protective mechanism that can be however blocked or overwhelmed.

  16. Fluoxetine induces cytotoxic endoplasmic reticulum stress and autophagy in triple negative breast cancer

    Science.gov (United States)

    Bowie, Michelle; Pilie, Patrick; Wulfkuhle, Julia; Lem, Siya; Hoffman, Abigail; Desai, Shraddha; Petricoin, Emanuel; Carter, Amira; Ambrose, Adrian; Seewaldt, Victoria; Yu, Dihua; Ibarra Drendall, Catherine

    2015-01-01

    AIM: To investigate the mechanism of action of lipophilic antidepressant fluoxetine (FLX) in representative molecular subtypes of breast cancer. METHODS: The anti-proliferative effects and mechanistic action of FLX in triple-negative (SUM149PT) and luminal (T47D and Au565) cancer cells and non-transformed MCF10A were investigated. Reverse phase protein microarray (RPPM) was performed with and without 10 μmol/L FLX for 24 and 48 h to determine which proteins are significantly changed. Viability and cell cycle analysis were also performed to determine drug effects on cell growth. Western blotting was used to confirm the change in protein expression examined by RPPM or pursue other signaling proteins. RESULTS: The FLX-induced cell growth inhibition in all cell lines was concentration- and time-dependent but less pronounced in early passage MCF10A. In comparison to the other lines, cell growth reduction in SUM149PT coincided with significant induction of endoplasmic reticulum (ER) stress and autophagy after 24 and 48 h of 10 μmol/L FLX, resulting in decreased translation of proteins along the receptor tyrosine kinase/Akt/mammalian target of rapamycin pathways. The increase in autophagy marker, cleaved microtubule-associated protein 1 light chain 3, in SUM149PT after 24 h of FLX was likely due to increased metabolic demands of rapidly dividing cells and ER stress. Consequently, the unfolded protein response mediated by double-stranded RNA-dependent protein kinase-like ER kinase resulted in inhibition of protein synthesis, growth arrest at the G1 phase, autophagy, and caspase-7-mediated cell death. CONCLUSION: Our study suggests a new role for FLX as an inducer of ER stress and autophagy, resulting in death of aggressive triple negative breast cancer SUM149PT. PMID:26677444

  17. Avian influenza A virus H5N1 causes autophagy-mediated cell death through suppression of mTOR signaling

    Institute of Scientific and Technical Information of China (English)

    Jianhui Ma; Qian Sun; Ruifang Mi; Hongbing Zhang

    2011-01-01

    Of the few avian influenza viruses that have crossed the species barrier to infect humans,the highly pathogenic influenza A (H5N1) strain has claimed the lives of more than half of the infected patients.With largely unknown mechanism of lung injury by H5N1 infection,acute respiratory distress syndrome (ARDS) is the major cause of death among the victims.Here we present the fact that H5N1 caused autophagic cell death through suppression of mTOR signaling.Inhibition of autophagy,either by depletion of autophagy gene Beclinl or by autophagy inhibitor 3-methyladenine (3-MA),significantly reduced H5N1 mediated cell death.We suggest that autophagic cell death may contribute to the development of ARDS in H5N1 influenza patients and inhibition of autophagy could therefore become a novel strategy for the treatment of H5N1 infection.

  18. Autophagy Has a Beneficial Role in Relieving Cigarette Smoke-Induced Apoptotic Death in Human Gingival Fibroblasts

    Science.gov (United States)

    Kim, Moon-Soo; Yun, Jeong-Won; Park, Jin-Ho; Park, Bong-Wook; Kang, Young-Hoon; Hah, Young-Sool; Hwang, Sun-Chul; Woo, Dong Kyun; Byun, June-Ho

    2016-01-01

    The deleterious role of cigarette smoke has long been documented in various human diseases including periodontal complications. In this report, we examined this adverse effect of cigarette smoke on human gingival fibroblasts (HGFs) which are critical not only in maintaining gingival tissue architecture but also in mediating immune responses. As well documented in other cell types, we also observed that cigarette smoke promoted cellular reactive oxygen species in HGFs. And we found that this cigarette smoke-induced oxidative stress reduced HGF viability through inducing apoptosis. Our results indicated that an increased Bax/Bcl-xL ratio and resulting caspase activation underlie the apoptotic death in HGFs exposed to cigarette smoke. Furthermore, we detected that cigarette smoke also triggered autophagy, an integrated cellular stress response. Interesting, a pharmacological suppression of the cigarette smoke-induced autophagy led to a further reduction in HGF viability while a pharmacological promotion of autophagy increased the viability of HGFs with cigarette smoke exposures. These findings suggest a protective role for autophagy in HGFs stressed with cigarette smoke, highlighting that modulation of autophagy can be a novel therapeutic target in periodontal complications with cigarette smoke.

  19. Galectin-4 expression is down-regulated in response to autophagy during differentiation of rat trophoblast cells.

    Science.gov (United States)

    Arikawa, Tomohiro; Liao, Shengjun; Shimada, Hiroki; Inoue, Tomoki; Sakata-Haga, Hiromi; Nakamura, Takanori; Hatta, Toshihisa; Shoji, Hiroki

    2016-01-01

    Placental development and trophoblast invasion of the maternal endometrium establish the maternal-fetal interface, which is critical for the developing embryo and fetus. Herein we show that overexpression of Galectin-4 (Gal-4) during trophoblast differentiation inhibited the enlargement of Rcho-1 cells (a model for rat trophoblast differentiation) and promoted cell-cell adhesion, whereas trophoblast specific markers and MMP-9 activity were not affected. In the rat placenta, microtubule associated protein 1 light chain 3 alpha (LC3) protein, an autophagy marker, is highly expressed on the maternal side of the decidua where Gal-4 expression is weak. In vitro assays showed that the expression of trophoblast-specific differentiation markers was reduced by 3-Methyladenine (3-MA) and Bafilomycin A1, known as autophagy inhibitors, compared to control cells. Furthermore, Gal-4 expression in Rcho-1 cells, which is normally down-regulated during differentiation, was not attenuated in the presence of autophagy inhibitors, suggesting that autophagy is upstream of Gal-4 expression. We herein describe a possible mechanism by which autophagy regulates trophoblast differentiation via regulation of Gal-4 expression in order to establish the maternal-fetal interface. PMID:27572741

  20. Autophagy protects type II alveolar epithelial cells from Mycobacterium tuberculosis infection

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Xu-Guang [Center for Clinical Laboratory Medicine of PLA, Xijing Hospital, Fourth Military Medical University, Xi' an (China); Department of Laboratory Medicine, The Third Affiliated Hospital of Guangzhou Medical University, Guangzhou (China); Ji, Tian-Xing [Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou Medical University, Guangzhou (China); Xia, Yong, E-mail: gysyxy@gmail.com [Center for Clinical Laboratory Medicine of PLA, Xijing Hospital, Fourth Military Medical University, Xi' an (China); Ma, Yue-Yun, E-mail: cmbmayy@fmmu.edu.cn [Center for Clinical Laboratory Medicine of PLA, Xijing Hospital, Fourth Military Medical University, Xi' an (China)

    2013-03-08

    Highlights: ► We investigated the protective effect of autophagy pathway against MTB infection. ► MTB-infected A549 cells had higher LDH release. ► Inhibition of autophagy signaling significantly enhanced the MTB-induced necrosis. ► Autophagy prevents apoptosis and promotes cell survival in infected cells. -- Abstract: This study was designed to investigate the protective effect of the autophagy signaling pathway against Mycobacterium tuberculosis infection in type II alveolar epithelial cells. An in vitro M. tuberculosis system was established using human A549 cells. Infection-induced changes in the expression of the autophagic marker LC3 were assessed by reverse transcription-PCR and Western blotting. Morphological changes in autophagosomes were detected by transmission electron microscopy (TEM). The function of the autophagy signaling pathway during infection was assessed by measuring the level of cell death and the amount of lactate dehydrogenase (LDH) released in the presence or absence of the inhibitor 3-methyladenine (3-MA). In addition, effects on LDH release were assessed after the siRNA-mediated knockdown of the essential autophagosomal structural membrane protein Atg5. LC3 mRNA expression was significantly reduced in M.tuberculosis-infected A549 cells (16888.76 ± 1576.34 vs. uninfected: 12744.29 ± 1089.37; P < 0.05). TEM revealed M.tuberculosis bacilli-containing compartments that were surrounded by double membranes characteristic of the autophagic process. M.tuberculosis-infected A549 cells released more LDH (1.45 ± 0.12 vs. uninfected: 0.45 ± 0.04; P < 0.05). The inhibition of autophagy signaling significantly enhanced M.tuberculosis-induced necrosis (3-MA: 75 ± 5% vs. untreated: 15 ± 1%; P < 0.05) and LDH release (3-MA: 2.50 ± 0.24 vs. untreated: 0.45 ± 0.04; Atg5 knockdown: 3.19 ± 0.29 vs. untreated: 1.28 ± 0.11; P < 0.05). Our results indicate that autophagy signaling pathway prevents apoptosis in type II alveolar epithelial cells

  1. Autophagy protects type II alveolar epithelial cells from Mycobacterium tuberculosis infection

    International Nuclear Information System (INIS)

    Highlights: ► We investigated the protective effect of autophagy pathway against MTB infection. ► MTB-infected A549 cells had higher LDH release. ► Inhibition of autophagy signaling significantly enhanced the MTB-induced necrosis. ► Autophagy prevents apoptosis and promotes cell survival in infected cells. -- Abstract: This study was designed to investigate the protective effect of the autophagy signaling pathway against Mycobacterium tuberculosis infection in type II alveolar epithelial cells. An in vitro M. tuberculosis system was established using human A549 cells. Infection-induced changes in the expression of the autophagic marker LC3 were assessed by reverse transcription-PCR and Western blotting. Morphological changes in autophagosomes were detected by transmission electron microscopy (TEM). The function of the autophagy signaling pathway during infection was assessed by measuring the level of cell death and the amount of lactate dehydrogenase (LDH) released in the presence or absence of the inhibitor 3-methyladenine (3-MA). In addition, effects on LDH release were assessed after the siRNA-mediated knockdown of the essential autophagosomal structural membrane protein Atg5. LC3 mRNA expression was significantly reduced in M.tuberculosis-infected A549 cells (16888.76 ± 1576.34 vs. uninfected: 12744.29 ± 1089.37; P < 0.05). TEM revealed M.tuberculosis bacilli-containing compartments that were surrounded by double membranes characteristic of the autophagic process. M.tuberculosis-infected A549 cells released more LDH (1.45 ± 0.12 vs. uninfected: 0.45 ± 0.04; P < 0.05). The inhibition of autophagy signaling significantly enhanced M.tuberculosis-induced necrosis (3-MA: 75 ± 5% vs. untreated: 15 ± 1%; P < 0.05) and LDH release (3-MA: 2.50 ± 0.24 vs. untreated: 0.45 ± 0.04; Atg5 knockdown: 3.19 ± 0.29 vs. untreated: 1.28 ± 0.11; P < 0.05). Our results indicate that autophagy signaling pathway prevents apoptosis in type II alveolar epithelial cells

  2. Don’t Rush Me . . . Go the Full 40: AWHONN’s Public Health Campaign Promotes Spontaneous Labor and Normal Birth to Reduce Overuse of Inductions and Cesareans

    OpenAIRE

    Bingham, Debra; Ruhl, Catherine; Cockey, Carolyn Davis

    2013-01-01

    Don’t Rush Me . . . Go the Full 40 is a grassroots public health campaign from the Association of Women’s Health, Obstetric and Neonatal Nursing (AWHONN) that educates women about the physiologic benefits of full-term pregnancy for themselves and their babies. GoTheFull40.com seeks to increase the percentage of women who complete at least 40 weeks of pregnancy, decrease the percentage of women who choose elective induction or elective cesarean surgery, and increase nurses’ and other pregnancy...

  3. Induction of DNA damage by deguelin is mediated through reducing DNA repair genes in human non-small cell lung cancer NCI-H460 cells

    OpenAIRE

    JI, BIN-CHUAN; Chien-chih YU; YANG, SU-TSO; Hsia, Te-Chun; Yang, Jai-Sing; Lai, Kuang-Chi; Ko, Yang-Ching; Lin, Jen-Jyh; Lai, Tung-Yuan; Chung, Jing-Gung

    2012-01-01

    It has been shown that deguelin, one of the compounds of rotenoids from flavonoid family, induced cytotoxic effects through induction of cell cycle arrest and apoptosis in many types of human cancer cell lines, but deguelin-affected DNA damage and repair gene expression (mRNA) are not clarified yet. We investigated the effects of deguelin on DNA damage and associated gene expression in human lung cancer NCI-H460 cells in vitro. DNA damage was assayed by using the comet assay and DNA gel elect...

  4. Age-related disruption of autophagy in dermal fibroblasts modulates extracellular matrix components

    Energy Technology Data Exchange (ETDEWEB)

    Tashiro, Kanae [Skin Research Department, POLA Chemical Industries, Inc., Yokohama (Japan); Division of Pharmaceutical Cell Biology, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka (Japan); Shishido, Mayumi [Skin Research Department, POLA Chemical Industries, Inc., Yokohama (Japan); Fujimoto, Keiko [Division of Pharmaceutical Cell Biology, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka (Japan); Organelle Homeostasis Research Center, Kyushu University, Fukuoka (Japan); Hirota, Yuko [Division of Pharmaceutical Cell Biology, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka (Japan); Yo, Kazuyuki; Gomi, Takamasa [Skin Research Department, POLA Chemical Industries, Inc., Yokohama (Japan); Tanaka, Yoshitaka, E-mail: tanakay@bioc.phar.kyushu-u.ac.jp [Division of Pharmaceutical Cell Biology, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka (Japan); Organelle Homeostasis Research Center, Kyushu University, Fukuoka (Japan)

    2014-01-03

    Highlights: •Autophagosomes accumulate in aged dermal fibroblasts. •Autophagic degradation is impaired in aged dermal fibroblasts. •Autophagy disruption affects extracellular matrix components in dermal fibroblasts. -- Abstract: Autophagy is an intracellular degradative system that is believed to be involved in the aging process. The contribution of autophagy to age-related changes in the human skin is unclear. In this study, we examined the relationship between autophagy and skin aging. Transmission electron microscopy and immunofluorescence microscopy analyses of skin tissue and cultured dermal fibroblasts derived from women of different ages revealed an increase in the number of nascent double-membrane autophagosomes with age. Western blot analysis showed that the amount of LC3-II, a form associated with autophagic vacuolar membranes, was significantly increased in aged dermal fibroblasts compared with that in young dermal fibroblasts. Aged dermal fibroblasts were minimally affected by inhibition of autophagic activity. Although lipofuscin autofluorescence was elevated in aged dermal fibroblasts, the expression of Beclin-1 and Atg5—genes essential for autophagosome formation—was similar between young and aged dermal fibroblasts, suggesting that the increase of autophagosomes in aged dermal fibroblasts was due to impaired autophagic flux rather than an increase in autophagosome formation. Treatment of young dermal fibroblasts with lysosomal protease inhibitors, which mimic the condition of aged dermal fibroblasts with reduced autophagic activity, altered the fibroblast content of type I procollagen, hyaluronan and elastin, and caused a breakdown of collagen fibrils. Collectively, these findings suggest that the autophagy pathway is impaired in aged dermal fibroblasts, which leads to deterioration of dermal integrity and skin fragility.

  5. Age-related disruption of autophagy in dermal fibroblasts modulates extracellular matrix components

    International Nuclear Information System (INIS)

    Highlights: •Autophagosomes accumulate in aged dermal fibroblasts. •Autophagic degradation is impaired in aged dermal fibroblasts. •Autophagy disruption affects extracellular matrix components in dermal fibroblasts. -- Abstract: Autophagy is an intracellular degradative system that is believed to be involved in the aging process. The contribution of autophagy to age-related changes in the human skin is unclear. In this study, we examined the relationship between autophagy and skin aging. Transmission electron microscopy and immunofluorescence microscopy analyses of skin tissue and cultured dermal fibroblasts derived from women of different ages revealed an increase in the number of nascent double-membrane autophagosomes with age. Western blot analysis showed that the amount of LC3-II, a form associated with autophagic vacuolar membranes, was significantly increased in aged dermal fibroblasts compared with that in young dermal fibroblasts. Aged dermal fibroblasts were minimally affected by inhibition of autophagic activity. Although lipofuscin autofluorescence was elevated in aged dermal fibroblasts, the expression of Beclin-1 and Atg5—genes essential for autophagosome formation—was similar between young and aged dermal fibroblasts, suggesting that the increase of autophagosomes in aged dermal fibroblasts was due to impaired autophagic flux rather than an increase in autophagosome formation. Treatment of young dermal fibroblasts with lysosomal protease inhibitors, which mimic the condition of aged dermal fibroblasts with reduced autophagic activity, altered the fibroblast content of type I procollagen, hyaluronan and elastin, and caused a breakdown of collagen fibrils. Collectively, these findings suggest that the autophagy pathway is impaired in aged dermal fibroblasts, which leads to deterioration of dermal integrity and skin fragility

  6. Research Progression of Cellular Autophagy in Liver System Diseases

    Directory of Open Access Journals (Sweden)

    Chunyun Liu

    2013-09-01

    Full Text Available Autophagy is a basic biological phenomenon widely existed in eukaryotic cells and an important mechanism for cells to adjust to the surrounding environment, prevent invasion of pathogenic micro-organisms and maintain homeostasis, whose activity changes evidently in multiple liver system diseases, suggesting that there is close association between autophagy and the generation and development of liver system diseases. It is also reported that autophagy develops and exerts an important function in many liver-related diseases, such as hepatic carcinoma, non-alcoholic fatty liver disease, alcoholic liver disease, viral liver disease and acute liver injury. Therefore, this study aimed to summarize the relationship between autophagy and multiple liver diseases, hoping to explore the effect of autophagy in liver system diseases and further study the regulative effect of autophagy so as to provide new thoughts for their treatment.

  7. Phosphorylation of the autophagy receptor optineurin restricts Salmonella growth

    DEFF Research Database (Denmark)

    Wild, Philipp; Farhan, Hesso; McEwan, David G; Wagner, Sebastian; Rogov, Vladimir V; Brady, Nathan R; Richter, Benjamin; Korac, Jelena; Waidmann, Oliver; Choudhary, Chunaram; Dötsch, Volker; Bumann, Dirk; Dikic, Ivan

    2011-01-01

    Selective autophagy can be mediated via receptor molecules that link specific cargoes to the autophagosomal membranes decorated by ubiquitin-like microtubule-associated protein light chain 3 (LC3) modifiers. Although several autophagy receptors have been identified, little is known about mechanisms...... controlling their functions in vivo. In this work, we found that phosphorylation of an autophagy receptor, optineurin, promoted selective autophagy of ubiquitin-coated cytosolic Salmonella enterica. The protein kinase TANK binding kinase 1 (TBK1) phosphorylated optineurin on serine-177, enhancing LC3 binding...... affinity and autophagic clearance of cytosolic Salmonella. Conversely, ubiquitin- or LC3-binding optineurin mutants and silencing of optineurin or TBK1 impaired Salmonella autophagy, resulting in increased intracellular bacterial proliferation. We propose that phosphorylation of autophagy receptors might...

  8. Nanomaterial-modulated autophagy: underlying mechanisms and functional consequences.

    Science.gov (United States)

    Zheng, Wei; Wei, Min; Li, Song; Le, Weidong

    2016-06-01

    Autophagy is an essential lysosome-dependent process that controls the quality of the cytoplasm and maintains cellular homeostasis, and dysfunction of this protein degradation system is correlated with various disorders. A growing body of evidence suggests that nanomaterials (NMs) have autophagy-modulating effects, thus predicting a valuable and promising application potential of NMs in the diagnosis and treatment of autophagy-related diseases. NMs exhibit unique physical, chemical and biofunctional properties, which may endow NMs with capabilities to modulate autophagy via various mechanisms. The present review highlights the impacts of various NMs on autophagy and their functional consequences. The possible underlying mechanisms for NM-modulated autophagy are also discussed. PMID:27193191

  9. Forms, Crosstalks, and the Role of Phospholipid Biosynthesis in Autophagy

    Directory of Open Access Journals (Sweden)

    Leanne Pereira

    2012-01-01

    Full Text Available Autophagy is a highly conserved cellular process occurring during periods of stress to ensure a cell's survival by recycling cytosolic constituents and making products that can be used in energy generation and other essential processes. Three major forms of autophagy exist according to the specific mechanism through which cytoplasmic material is transported to a lysosome. Chaperone-mediated autophagy is a highly selective form of autophagy that delivers specific proteins for lysosomal degradation. Microautophagy is a less selective form of autophagy that occurs through lysosomal membrane invaginations, forming tubes and directly engulfing cytoplasm. Finally, macroautophagy involves formation of new membrane bilayers (autophagosomes that engulf cytosolic material and deliver it to lysosomes. This review provides new insights on the crosstalks between different forms of autophagy and the significance of bilayer-forming phospholipid synthesis in autophagosomal membrane formation.

  10. WASH inhibits autophagy through suppression of Beclin 1 ubiquitination

    OpenAIRE

    Xia, Pengyan; Wang, Shuo; Du, Ying; Zhao, Zhenao; Shi, Lei; Sun, Lei; Huang, Guanling; Ye, Buqing; Li, Chong; Dai, Zhonghua; Hou, Ning; Cheng, Xuan; Sun, Qingyuan; Li, Lei(Beijing Institute of Petrochemical Technology, Beijing, 102617, People's Republic of China); Yang, Xiao

    2013-01-01

    Autophagy degrades cytoplasmic proteins and organelles to recycle cellular components that are required for cell survival and tissue homeostasis. However, it is not clear how autophagy is regulated in mammalian cells. WASH (Wiskott–Aldrich syndrome protein (WASP) and SCAR homologue) plays an essential role in endosomal sorting through facilitating tubule fission via Arp2/3 activation. Here, we demonstrate a novel function of WASH in modulation of autophagy. We show that WASH deficiency causes...

  11. Pyrvinium targets autophagy addiction to promote cancer cell death

    OpenAIRE

    Deng, Longfei; Lei, Yunlong; Liu, Rui; Li, Jingyi; Yuan, Kefei; Li, Yi; Chen, Yi; Liu, Yi; Lu, You; Edwards III, Carl K; Huang, Canhua; Wei, Yuquan

    2013-01-01

    Autophagy is a cellular catabolic process by which long-lived proteins and damaged organelles are degradated by lysosomes. Activation of autophagy is an important survival mechanism that protects cancer cells from various stresses, including anticancer agents. Recent studies indicate that pyrvinium pamoate, an FDA-approved antihelminthic drug, exhibits wide-ranging anticancer activity. Here we demonstrate that pyrvinium inhibits autophagy both in vitro and in vivo. We further demonstrate that...

  12. The interplay between autophagy and ROS in tumorigenesis

    Directory of Open Access Journals (Sweden)

    VassilikiKarantza

    2012-11-01

    Full Text Available Reactive oxygen species (ROS at physiological levels are important cell signaling molecules. However, aberrantly high ROS are intimately associated with disease and commonly observed in cancer. Mitochondria are primary sources of intracellular ROS, and their maintenance is essential to cellular health. Autophagy, an evolutionarily conserved process whereby cytoplasmic components are delivered to lysosomes for degradation, is responsible for mitochondrial turnover and removal of damaged mitochondria. Impaired autophagy is implicated in many pathological conditions, including neurological disorders, inflammatory bowel disease, diabetes, aging and cancer. The first reports connecting autophagy to cancer showed that allelic loss of the essential autophagy gene BECLIN1 (BECN1 is prevalent in human breast, ovarian and prostate cancers and that Becn1+/- mice develop mammary gland hyperplasias, lymphomas, and lung and liver tumors. Subsequent studies demonstrated that Atg5-/- and Atg7-/- livers give rise to adenomas, Atg4-/- mice are susceptible to chemical carcinogenesis, and Bif1-/- mice are prone to spontaneous tumors, indicating that autophagy defects promote tumorigenesis. Due to defective mitophagy, autophagy-deficient cells accumulate damaged mitochondria and deregulated ROS levels, which likely contribute to their tumor-initiating capacity. However, the role of autophagy in tumorigenesis is complex, as more recent work also revealed tumor dependence on autophagy: autophagy-competent mutant-Ras-expressing cells form tumors more efficiently than their autophagy-deficient counterparts; similarly, FIP200 deficiency suppresses PyMT-driven mammary tumorigenesis. These latter findings are attributed to the fact that tumors driven by powerful oncogenes have high metabolic demands catered to by autophagy. In this review, we discuss the relationship between ROS and autophagy and summarize our current knowledge on their functional interactions in

  13. Autophagy and mitophagy in the myocardium: therapeutic potential and concerns

    OpenAIRE

    Jimenez, Rebecca E; Kubli, Dieter A.; Gustafsson, Åsa B.

    2014-01-01

    The autophagic-lysosomal degradation pathway is critical for cardiac homeostasis, and defects in this pathway are associated with development of cardiomyopathy. Autophagy is responsible for the normal turnover of organelles and long-lived proteins. Autophagy is also rapidly up-regulated in response to stress, where it rapidly clears dysfunctional organelles and cytotoxic protein aggregates in the cell. Autophagy is also important in clearing dysfunctional mitochondria before they can cause ha...

  14. Dexamethasone-induced autophagy mediates muscle atrophy through mitochondrial clearance

    OpenAIRE

    Troncoso, Rodrigo; Paredes, Felipe; Parra, Valentina; Gatica, Damián; Vásquez-Trincado, César; Quiroga, Clara; Bravo-Sagua, Roberto; López-Crisosto, Camila; Rodriguez, Andrea E; Oyarzún, Alejandra P; Kroemer, Guido; Lavandero, Sergio

    2014-01-01

    Glucocorticoids, such as dexamethasone, enhance protein breakdown via ubiquitin–proteasome system. However, the role of autophagy in organelle and protein turnover in the glucocorticoid-dependent atrophy program remains unknown. Here, we show that dexamethasone stimulates an early activation of autophagy in L6 myotubes depending on protein kinase, AMPK, and glucocorticoid receptor activity. Dexamethasone increases expression of several autophagy genes, including ATG5, LC3, BECN1, and SQSTM1 a...

  15. Alisertib Induces Cell Cycle Arrest, Apoptosis, Autophagy and Suppresses EMT in HT29 and Caco-2 Cells

    Directory of Open Access Journals (Sweden)

    Bao-Jun Ren

    2015-12-01

    Full Text Available Colorectal cancer (CRC is one of the most common malignancies worldwide with substantial mortality and morbidity. Alisertib (ALS is a selective Aurora kinase A (AURKA inhibitor with unclear effect and molecular interactome on CRC. This study aimed to evaluate the molecular interactome and anticancer effect of ALS and explore the underlying mechanisms in HT29 and Caco-2 cells. ALS markedly arrested cells in G2/M phase in both cell lines, accompanied by remarkable alterations in the expression level of key cell cycle regulators. ALS induced apoptosis in HT29 and Caco-2 cells through mitochondrial and death receptor pathways. ALS also induced autophagy in HT29 and Caco-2 cells, with the suppression of phosphoinositide 3-kinase (PI3K/protein kinase B (Akt/mammalian target of rapamycin (mTOR, but activation of 5′ AMP-activated protein kinase (AMPK signaling pathways. There was a differential modulating effect of ALS on p38 MAPK signaling pathway in both cell lines. Moreover, induction or inhibition of autophagy modulated basal and ALS-induced apoptosis in both cell lines. ALS potently suppressed epithelial to mesenchymal transition (EMT in HT29 and Caco-2 cells. Collectively, it suggests that induction of cell cycle arrest, promotion of apoptosis and autophagy, and suppression of EMT involving mitochondrial, death receptor, PI3K/Akt/mTOR, p38 MAPK, and AMPK signaling pathways contribute to the cancer cell killing effect of ALS on CRC cells.

  16. Autophagy counteracts apoptosis in human multiple myeloma cells exposed to oridonin in vitro via regulating intracellular ROS and SIRT1

    Institute of Scientific and Technical Information of China (English)

    Rong ZENG; Yan CHEN; Shuai ZHAO; Guo-hui CUI

    2012-01-01

    To explore the mechanisms underlying the oridonin-induced apoptosis and autophagy in human multiple myeloma cells in vitro.Methods:Human multiple myeloma RPMI8266 cells were used.The cell viability was assessed using MTT assay.Morphological changes of apoptosis and autophagy were observed under transmission electron microscope.TUNEL and annexin V-FITC/PI dual staining assays were used to measure apoptosis.Autophagy was analyzed using Western blot analysis and immunofluorescence staining with a QDs605 nm-Anti-LC3 fluorescent probe.Intracellular ROS was estimated with flow cytometry using DCFH-DA fluorescent probe.Protein levels of active caspase 3,Beclin 1 and SIRT1 were determined with Western blot analysis.Results:Exposure to oridonin (1-64 μmol/L) inhibited the proliferation of RPMI8266 cells in a concentration-dependent manner with an IC50 value of 6.74 μmol/L.Exposure to oridonin (7 μmol/L) simultaneously induced caspase 3-mediated apoptosis and Beclin 1-dependent autophagy of RPMI8266 cells.Both the apoptosis and autophagy were time-dependent,and apoptosis was the main effector pathway of cell death.Exposure to oridonin (7 μmol/L) increased intracellular ROS and reduced SIRT1 nuclear protein in a time-dependent manner.The blockade of intracellular generation of ROS by NAC (5 mmol/L) abrogated apoptosis,autophagy and the decrease of SIRT1 in the cells exposed to oridonin (7 μmol/L).The inhibition of autophagy by 3-MA (5 mmol/L) sensitized the cells to oridonin-induced apoptosis,which was accompanied by increased intracellular ROS and decreased SlRT1.Conclusion:Oridonin simultaneously induces apoptosis and autophagy of human multiple myeloma RPMI8266 cells via regulation of intracellular ROS generation and SIRT1 nuclear protein.The cytotoxicity of oridonin is mainly mediated through the apoptotic pathway,whereas the autophagy protects the cells from apoptosis.

  17. Autophagy in Lepidoptera: more than old wine in new bottle

    Directory of Open Access Journals (Sweden)

    G Tettamanti

    2011-01-01

    Full Text Available Autophagy is a cellular pathway that leads to the degradation of proteins and organelles. This process is usually involved in the maintenance of cell homeostasis when the organism experiences nutrient starvation, but in holometabolous insects autophagy also intervenes in the demolition of larval tissues and organs during metamorphosis. This review summarizes the current knowledge about autophagy research in Lepidoptera and discusses the use of moths and butterflies as models for tudying the roles and regulation of autophagy. It also gives insights into the cooperation between utophagy and apoptosis in cell death events that occur in lepidopteran in vivo and in vitro systems.

  18. Tracking autophagy during proliferation and differentiation of Trypanosoma brucei

    Directory of Open Access Journals (Sweden)

    William R. Proto

    2014-01-01

    Full Text Available Autophagy is a lysosome-dependent degradation mechanism that sequesters target cargo into autophagosomal vesicles. The Trypanosoma brucei genome contains apparent orthologues of several autophagy-related proteins including an ATG8 family. These ubiquitin-like proteins are required for autophagosome membrane formation, but our studies show that ATG8.3 is atypical. To investigate the function of other ATG proteins, RNAi compatible T. brucei were modified to function as autophagy reporter lines by expressing only either YFP-ATG8.1 or YFP-ATG8.2. In the insect procyclic lifecycle stage, independent RNAi down-regulation of ATG3 or ATG7 generated autophagy-defective mutants and confirmed a pro-survival role for autophagy in the procyclic form nutrient starvation response. Similarly, RNAi depletion of ATG5 or ATG7 in the bloodstream form disrupted autophagy, but did not impede proliferation. Further characterisation showed bloodstream form autophagy mutants retain the capacity to undergo the complex cellular remodelling that occurs during differentiation to the procyclic form and are equally susceptible to dihydroxyacetone-induced cell death as wild type parasites, not supporting a role for autophagy in this cell death mechanism. The RNAi reporter system developed, which also identified TOR1 as a negative regulator controlling YFP-ATG8.2 but not YFP-ATG8.1 autophagosome formation, will enable further targeted analysis of the mechanisms and function of autophagy in the medically relevant bloodstream form of T. brucei.

  19. Emerging role of selective autophagy in human diseases.

    Directory of Open Access Journals (Sweden)

    Kenji eMizumura

    2014-11-01

    Full Text Available AbstractAutophagy was originally described as a highly conserved system for the degradation of cytosol through a lysosome-dependent pathway. In response to starvation, autophagy degrades organelles and proteins to provide metabolites and energy for its pro-survival effects. Autophagy is recognized as playing a role in the pathogenesis of disease either directly or indirectly, through the regulation of vital processes such as programmed cell death, inflammation, and adaptive immune mechanisms. Recent studies have demonstrated that autophagy is not only a simple metabolite recycling system, but also has the ability to degrade specific cellular targets, such as mitochondria, cilia, and invading bacteria. In addition, selective autophagy has also been implicated in vesicle trafficking pathways, with potential roles in secretion and other intracellular transport processes. Selective autophagy has drawn the attention of researchers because of its potential importance in clinical diseases. Therapeutic strategies to target selective autophagy rather than general autophagy may maximize clinical benefit by enhancing selectivity. In this review, we outline the principle components of selective autophagy processes and their emerging importance in human disease, with an emphasis on pulmonary diseases.

  20. Role and Regulation of Autophagy in Heat Stress Responses of Tomato Plants

    OpenAIRE

    Jie eZhou; Jian eWang; Jingquan eYu; Zhixiang eChen

    2014-01-01

    As sessile organisms, plants are constantly exposed to a wide spectrum of stress conditions such as high temperature, which causes protein misfolding. Misfolded proteins are highly toxic and must be efficiently removed to reduce cellular proteotoxic stress if restoration of native conformations is unsuccessful. Although selective autophagy is known to function in protein quality control by targeting degradation of misfolded and potentially toxic proteins, its role and regulation in heat st...

  1. The emerging role of autophagy in peroxisome dynamics and lipid metabolism of phyllosphere microorganisms.

    OpenAIRE

    Oku, Masahide; Takano, Yoshitaka; Sakai, Yasuyoshi

    2014-01-01

    Eukaryotic microorganisms resident in the phyllosphere (above-ground, plant-surface environments) undergo dynamic changes in nutrient conditions and adapt their metabolic pathways during proliferation or in the course of infection of host plants. Some of these metabolic switches are accomplished by regulation of organelle abundance. Recent studies have shown that autophagy plays a major role in reducing the organelle quantity, thereby contributing to the metabolic switch required for survival...

  2. The emerging role of autophagy in peroxisome dynamics and lipid metabolism of phyllosphere microorganisms

    OpenAIRE

    Oku, Masahide; Takano, Yoshitaka; Sakai, Yasuyoshi

    2014-01-01

    Eukaryotic microorganisms resident in the phyllosphere (above-ground, plant-surface environments) undergo dynamic changes in nutrient conditions and adapt their metabolic pathways during proliferation or in the course of infection of host plants. Some of these metabolic switches are accomplished by regulation of organelle abundance. Recent studies have shown that autophagy plays a major role in reducing the organelle quantity, thereby contributing to the metabolic switch required for survival...

  3. Moxibustion Activates Macrophage Autophagy and Protects Experimental Mice against Bacterial Infection

    OpenAIRE

    Xiaojuan Li; Guanhua Guo; Feng Shen; Lihong Kong; Fengxia Liang; Guojie Sun

    2014-01-01

    Moxibustion is one of main therapies in traditional Chinese medicine and uses heat stimulation on the body surface from the burning of moxa to release pain or treat diseases. Emerging studies have shown that moxibustion can generate therapeutic effects by activating a series of signaling pathways and neuroendocrine-immune activities. Here we show moxibustion promoted profound macrophage autophagy in experimental Kunming mice, with reduced Akt phosphorylation and activated eIF2α phosphorylatio...

  4. Practicing induction:

    DEFF Research Database (Denmark)

    Sprogøe, Jonas; Rohde, Nicolas

    2009-01-01

    We claim that induction potentially triggers both individual and organizational learning and by drawing on practice-based theory we discuss how the interplay between individual and organization, what we call a generative dance, ignites both kinds of learning....

  5. Evolving insights on metabolism, autophagy and epigenetics in liver myofibroblasts

    Directory of Open Access Journals (Sweden)

    Zeribe Chike Nwosu

    2016-06-01

    Full Text Available Liver myofibroblasts (MFB are crucial mediators of extracellular matrix (ECM deposition in liver fibrosis. They arise mainly from hepatic stellate cells (HSCs upon a process termed activation. To a lesser extent, and depending on the cause of liver damage, portal fibroblasts, mesothelial cells and fibrocytes may also contribute to the MFB population. Targeting MFB to reduce liver fibrosis is currently an area of intense research. Unfortunately, a clog in the wheel of antifibrotic therapies is the fact that although MFB are known to mediate scar formation, and participate in liver inflammatory response, many of their molecular portraits are currently unknown. In this review, we discuss recent understanding of MFB in health and diseases, focusing specifically on three evolving research fields: metabolism, autophagy and epigenetics. We have emphasized on therapeutic prospects where applicable and mentioned techniques for use in MFB studies. Subsequently, we highlighted uncharted territories in MFB research to help direct future efforts aimed at bridging gaps in current knowledge.

  6. Downregulation of protein kinase CK2 activity facilitates tumor necrosis factor-α-mediated chondrocyte death through apoptosis and autophagy.

    Directory of Open Access Journals (Sweden)

    Sung Won Lee

    Full Text Available Despite the numerous studies of protein kinase CK2, little progress has been made in understanding its function in chondrocyte death. Our previous study first demonstrated that CK2 is involved in apoptosis of rat articular chondrocytes. Recent studies have suggested that CK2 downregulation is associated with aging. Thus examining the involvement of CK2 downregulation in chondrocyte death is an urgently required task. We undertook this study to examine whether CK2 downregulation modulates chondrocyte death. We first measured CK2 activity in articular chondrocytes of 6-, 21- and 30-month-old rats. Noticeably, CK2 activity was downregulated in chondrocytes with advancing age. To build an in vitro experimental system for simulating tumor necrosis factor (TNF-α-induced cell death in aged chondrocytes with decreased CK2 activity, chondrocytes were co-treated with CK2 inhibitors and TNF-α. Viability assay demonstrated that CK2 inhibitors facilitated TNF-α-mediated chondrocyte death. Pulsed-field gel electrophoresis, nuclear staining, flow cytometry, TUNEL staining, confocal microscopy, western blot and transmission electron microscopy were conducted to assess cell death modes. The results of multiple assays showed that this cell death was mediated by apoptosis. Importantly, autophagy was also involved in this process, as supported by the appearance of a punctuate LC3 pattern and autophagic vacuoles. The inhibition of autophagy by silencing of autophage-related genes 5 and 7 as well as by 3-methyladenine treatment protected chondrocytes against cell death and caspase activation, indicating that autophagy led to the induction of apoptosis. Autophagic cells were observed in cartilage obtained from osteoarthritis (OA model rats and human OA patients. Our findings indicate that CK2 down regulation facilitates TNF-α-mediated chondrocyte death through apoptosis and autophagy. It should be clarified in the future if autophagy observed is a consequence

  7. From Inductive Learning Towards Interactive Inductive Learning

    OpenAIRE

    Birzniece, I

    2010-01-01

    Growing amount of information in the world encourage the use of automatic data processing techniques that reduce humans routine work. There is a wide range of methods used for machine learning; however inductive learning algorithms are preferable in the systems where understanding of decision making steps and further processing of results is needed, for instance the expert systems, where the rules induced by learning algorithms can be used. As the classification tasks are getting more complic...

  8. Activation of RARα induces autophagy in SKBR3 breast cancer cells and depletion of key autophagy genes enhances ATRA toxicity.

    Science.gov (United States)

    Brigger, D; Schläfli, A M; Garattini, E; Tschan, M P

    2015-01-01

    All-trans retinoic acid (ATRA), a pan-retinoic acid receptor (RAR) agonist, is, along with other retinoids, a promising therapeutic agent for the treatment of a variety of solid tumors. On the one hand, preclinical studies have shown promising anticancer effects of ATRA in breast cancer; on the other hand, resistances occurred. Autophagy is a cellular recycling process that allows the degradation of bulk cellular contents. Tumor cells may take advantage of autophagy to cope with stress caused by anticancer drugs. We therefore wondered if autophagy is activated by ATRA in mammary tumor cells and if modulation of autophagy might be a potential novel treatment strategy. Indeed, ATRA induces autophagic flux in ATRA-sensitive but not in ATRA-resistant human breast cancer cells. Moreover, using different RAR agonists as well as RARα-knockdown breast cancer cells, we demonstrate that autophagy is dependent on RARα activation. Interestingly, inhibition of autophagy in breast cancer cells by either genetic or pharmacological approaches resulted in significantly increased apoptosis under ATRA treatment and attenuated epithelial differentiation. In summary, our findings demonstrate that ATRA-induced autophagy is mediated by RARα in breast cancer cells. Furthermore, inhibition of autophagy results in enhanced apoptosis. This points to a potential novel treatment strategy for a selected group of breast cancer patients where ATRA and autophagy inhibitors are applied simultaneously. PMID:26313912

  9. Autophagy sensitivity of neuroendocrine lung tumor cells

    OpenAIRE

    HONG, SEUNG-KEUN; Kim, Jin-Hwan; Starenki, Dmytro; Park, Jong-In

    2013-01-01

    Neuroendocrine (NE) phenotypes characterize a spectrum of lung tumors, including low-grade typical and intermediate-grade atypical carcinoid, high-grade large-cell NE carcinoma and small cell lung carcinoma. Currently, no effective treatments are available to cure NE lung tumors, demanding identification of biological features specific to these tumors. Here, we report that autophagy has an important role for NE lung tumor cell proliferation and survival. We found that the expression levels of...

  10. Cardiomyocyte autophagy: metabolic profit and loss

    OpenAIRE

    Wang, Zhao V.; Ferdous, Anwarul; Hill, Joseph A.

    2013-01-01

    Cardiovascular disease remains the leading cause of morbidity and mortality worldwide, even despite recent scientific and technological advances and comprehensive preventive strategies. The cardiac myocyte is a voracious consumer of energy, and alterations in metabolic substrate availability and consumption are hallmark features of these disorders. Autophagy, an evolutionarily ancient response to metabolic insufficiency, has been implicated in the pathogenesis of a wide range of heart patholo...

  11. Autophagy in Mycobacterium tuberculosis and HIV infections

    OpenAIRE

    Espert, Lucile; Beaumelle, Bruno; Vergne, Isabelle

    2015-01-01

    Human Immunodeficiency Virus (HIV) and Mycobacterium tuberculosis (M.tb) are among the most lethal human pathogens worldwide, each being responsible for around 1.5 million deaths annually. Moreover, synergy between acquired immune deficiency syndrome (AIDS) and tuberculosis (TB) has turned HIV/M.tb co-infection into a major public health threat in developing countries. In the past decade, autophagy, a lysosomal catabolic process, has emerged as a major host immune defense mechanism against in...

  12. The Role of Autophagy in Lupus Nephritis

    OpenAIRE

    Linlin Wang; Helen Ka Wai Law

    2015-01-01

    Systemic lupus erythematosus (SLE) is a multifactorial autoimmune disease characterized by the generation of immune responses to self-antigens. Lupus nephritis is one of the most common and severe complications in SLE patients. Though the pathogenesis of lupus nephritis has been studied extensively, unresolved questions are still left and new therapeutic methods are needed for disease control. Autophagy is a conserved catabolic process through which cytoplasmic constituents can be degraded in...

  13. Autophagy protects human brain microvascular endothelial cells against methylglyoxal-induced injuries, reproducible in a cerebral ischemic model in diabetic rats.

    Science.gov (United States)

    Fang, Lili; Li, Xue; Zhong, Yinbo; Yu, Jing; Yu, Lina; Dai, Haibin; Yan, Min

    2015-10-01

    Cerebral microvascular endothelial cells (ECs) are crucial for brain vascular repair and maintenance, but their physiological function may be impaired during ischemic stroke and diabetes. Methylglyoxal (MGO), a reactive dicarbonyl produced during glucose metabolism, could exacerbate ischemia-induced EC injury and dysfunction. We investigated the protective effect of autophagy on cultured human brain microvascular endothelial cells (HBMEC) that underwent MGO treatment. A further study was conducted to explore the underlying mechanisms of the protective effect. Autophagic activity was assessed by evaluating protein levels, using western blot. 3-methyladenine (3-MA), bafilomycin A1, ammonium chloride (AC), Beclin 1 siRNA, and chloroquine (CQ) were used to cause autophagy inhibition. Alarmar blue assay and lactate dehydrogenase release assay were used to evaluate cell viability. Streptozotocin was administered to induce type I diabetes in rats and post-permanent middle cerebral artery occlusion was performed to elicit cerebral ischemia. Blood-brain barrier permeability was also assessed. Our study found that MGO reduced HBMEC cell viability in a concentration- and time-dependent manner, and triggered the responsive autophagy activation. Autophagy inhibitors bafilomycin A1, AC, 3-MA, and BECN1 siRNA exacerbated MGO-induced HBMEC injury. FAK phosphorylation inhibitor PF573228 inhibited MGO-triggered autophagy and enhanced lactate dehydrogenase release. Meanwhile, similar autophagy activation in brain vascular ECs was observed during permanent middle cerebral artery occlusion-induced cerebral ischemia in diabetic rats, while chloroquine-induced autophagy inhibition enhanced blood-brain barrier permeability. Taken together, our study indicates that autophagy triggered by MGO defends HBMEC against injuries. PMID:26251121

  14. Autophagy and ethanol-induced liver injury

    Institute of Scientific and Technical Information of China (English)

    Terrence M Donohue Jr

    2009-01-01

    The majority of ethanol metabolism occurs in the liver. Consequently, this organ sustains the greatest damage from ethanol abuse. Ethanol consumption disturbs the delicate balance of protein homeostasis in the liver, causing intracellular protein accumulation due to a disruption of hepatic protein catabolism.Evidence indicates that ethanol or its metabolism impairs trafficking events in the liver, including the process of macroautophagy, which is the engulfment and degradation of cytoplasmic constituents by the lysosomal system. Autophagy is an essential, ongoing cellular process that is highly regulated by nutrients,endocrine factors and signaling pathways. A great number of the genes and gene products that govern the autophagic response have been characterized and the major metabolic and signaling pathways that activate or suppress autophagy have been identified. This review describes the process of autophagy, its regulation and the possible mechanisms by which ethanol disrupts the process of autophagic degradation. The implications of autophagic suppression are discussed in relation to the pathogenesis of alcohol-induced liver injury.

  15. Vector control of induction machines

    CERN Document Server

    Robyns, Benoit

    2012-01-01

    After a brief introduction to the main law of physics and fundamental concepts inherent in electromechanical conversion, ""Vector Control of Induction Machines"" introduces the standard mathematical models for induction machines - whichever rotor technology is used - as well as several squirrel-cage induction machine vector-control strategies. The use of causal ordering graphs allows systematization of the design stage, as well as standardization of the structure of control devices. ""Vector Control of Induction Machines"" suggests a unique approach aimed at reducing parameter sensitivity for

  16. Quercitrin treatment protects endothelial progenitor cells from oxidative damage via inducing autophagy through extracellular signal-regulated kinase.

    Science.gov (United States)

    Zhi, Kangkang; Li, Maoquan; Bai, Jun; Wu, Yongfa; Zhou, Sili; Zhang, Xiaoping; Qu, Lefeng

    2016-07-01

    Atherosclerosis is a disease resulting from impaired endothelial function, often caused by oxidant injury or inflammation. Endothelial progenitor cells (EPCs) play a critical role in repairing damaged endothelium and protecting against atherosclerosis. Quercitrin, a plant-derived flavonoid compound, displays antioxidant and anti-inflammatory activities. In this study, we showed that quercitrin treatment reduced the apoptosis of EPCs caused by oxidized low-density lipoprotein (ox-LDL) in a dose-dependent manner. Quercitrin improved tube formation, migration and adhesion of ox-LDL-treated EPCs. To determine the effect of quercitrin in vivo, EPCs treated with or without ox-LDL and quercitrin were locally injected into the ischemic hind limb muscle of nude mice. Those injected with EPCs treated with ox-LDL and quercitrin showed significantly increased local accumulation of EPCs, blood flow recovery and capillary density compared with the control and ox-LDL only groups. Furthermore, we showed that quercitrin enhanced autophagy and upregulated mitogen-activated protein kinase and ERK phosphorylation in a dose-dependent manner in vitro. Autophagy inhibitors, chloroquine and 3-methyladenine, abrogated quercitrin-enhanced autophagy caused by ox-LDL as evidenced by decreased numbers of branch points, migratory cells and adherent cells, and increased numbers of apoptotic cells. The ERK inhibitor PD98059 abrogated quercitrin-enhanced autophagy, as identified by decreased autophagosome formation and downregulated ERK phosphorylation. The inhibition of ERK did not affect the expression of Rac1, but enhanced phosphorylation of Akt. Quercitrin treatment also increased the expression of E-cadherin, and PD98059 abrogated the upregulation of E-cadherin induced by quercitrin. Our findings suggested that autophagy is a protective mechanism in EPCs exposed to oxidative damage. Quercitrin can promote autophagy through the activation of ERK and the ERK signaling pathway is therefore

  17. The emerging role of autophagy in peroxisome dynamics and lipid metabolism of phyllosphere microorganisms