Chapter 11 Autophagy in Neurite Injury and Neurodegeneration

Chu, Charleen T.; Plowey, Edward D.; Dagda, Ruben K.; Hickey, Robert W.; Cherra, Salvatore J.; Clark, Robert S. B.

doi:10.1016/s0076-6879(08)04011-1

Cited by 108 publications

(85 citation statements)

References 91 publications

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“…The NeuronJ plug-in for NIH ImageJ software was further used to measure the length of pseudopodia according to a previous report. 55 TEM imaging MIR517C (oe), MIR517C-NC, and MIR517C (kn) U87 and U251 cells after TMZ treatment and freshly harvested tumors from mice were fixed with 2.5% glutaraldehyde, washed, and postfixed in osmium. Following dehydration by serial dilutions of ethanol, cells were stained with uranyl acetate and embedded in culture dishes.…”

Section: Rna Isolation and Real-time Qpcrmentioning

confidence: 99%

MIR517Cinhibits autophagy and the epithelial-to-mesenchymal (-like) transition phenotype in human glioblastoma through KPNA2-dependent disruption of TP53 nuclear translocation

Xiao

Liu

et al. 2015

Autophagy

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(2015) MIR517C inhibits autophagy and the epithelial-to-mesenchymal (-like) transition phenotype in human glioblastoma through KPNA2-dependent disruption of TP53 nuclear translocation, Autophagy, 11:12, 2213-2232, DOI: 10.1080/15548627.2015 LG, low glucose; MAP1LC3B/LC3B, microtubuleassociated protein 1 light chain 3B; MU, mutation; NC, negative control; NSC, neural stem cell; oe, overexpressing; OS, overall survival; PFS, progression-free survival; qRT-PCR, quantitative real-time reverse transcription polymerase chain reaction; SNAI2, Slug (snail family zinc finger 2); SNAI1, snail (snail family zinc finger 1); TEM, transmission electron microscopy; TMZ, temozolomide; VIM, vimentin; WT, wild type; ZEB1, zinc finger E-box binding homeobox 1.The epithelial-to-mesenchymal (-like) transition (EMT), a crucial embryonic development program, has been linked to the regulation of glioblastoma (GBM) progression and invasion. Here, we investigated the role of MIR517C/miR-517c, which belongs to the C19MC microRNA cluster identified in our preliminary studies, in the pathogenesis of GBM. We found that MIR517C was associated with improved prognosis in patients with GBM. Furthermore, following treatment with the autophagy inducer temozolomide (TMZ) and low glucose (LG), MIR517C degraded KPNA2 (karyopherin alpha 2 [RAG cohort 1, importin alpha 1]) and subsequently disturbed the nuclear translocation of TP53 in the GBM cell line U87 in vitro. Interestingly, this microRNA could inhibit autophagy and reduce cell migration and infiltration in U87 cells harboring wild-type (WT) TP53, but not in U251 cells harboring mutant (MU) TP53. Moreover, the expression of epithelial markers (i.e., CDH13/T-cadherin and CLDN1 [claudin 1]) increased, while the expression of mesenchymal markers (i.e., CDH2/N-cadherin, SNAI1/Snail, and VIM [vimentin]) decreased, indicating that the EMT status was blocked by MIR517C in U87 cells. Compared with MIR517C overexpression, MIR517C knockdown promoted infiltration of U87 cells to the surrounding structures in nude mice in vivo. The above phenotypic changes were also observed in TP53 +/+ and TP53 -/-HCT116 colon cancer cells. In summary, our study provided support for a link between autophagy and EMT status in WT TP53 GBM cells and provided evidence for the signaling pathway (MIR517C-KPNA2-cytoplasmic TP53) involved in attenuating autophagy and eliminating the increased migration and invasion during the EMT.

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Section: Rna Isolation and Real-time Qpcrmentioning

confidence: 99%

MIR517Cinhibits autophagy and the epithelial-to-mesenchymal (-like) transition phenotype in human glioblastoma through KPNA2-dependent disruption of TP53 nuclear translocation

Xiao

Liu

et al. 2015

Autophagy

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“…59 These findings suggest that maintaining balanced autophagy is critical for neuronal health and function. 60 In this study, we aimed to answer whether interference of autophagy could be of potential use in counteracting the progression of PD and whether melatonin, a readily available substance that is already widely in use in humans for other indications, could be potentially translated into human clinical trials for curing PD given melatonin's good safety profile and availability.…”

Section: Discussionmentioning

confidence: 99%

“…60 Therefore, a direct test of melatonin's effect on neurons in vitro and in vivo, rather than on cell lines, is essential for further defining its role in the development of PD. Consistent with a previous finding that melatonin can save dopaminergic cells in MPTP-treated mice, 62 our study showed that MPTP treatment reduced the length of axons and dendrites in primary neurons and decreased the number and dendrites of dopaminergic neurons in the SNc region of monkeys and mice with MPTP injection.…”

Section: Discussionmentioning

confidence: 99%

Melatonin attenuates MPTP-induced neurotoxicity via preventing CDK5-mediated autophagy and SNCA/α-synuclein aggregation

et al. 2015

Autophagy

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Keywords: autophagy, CDK5, melatonin, MPTP, SNCA, TH Abbreviations: ATG7, autophagy-related 7; BAFA1, bafilomycin A 1 ; CDK5, cyclin-dependent kinase 5; CDK5R1/p35/p25, cyclindependent kinase 5, regulatory subunit 1 (p35); DA, dopamine; DAPI, 4 0 ,6-diamidino-2-phenylindole; i.p, intraperitoneally; s.c, subcutaneously; ICV, intracerebroventricular; MAP1LC3B/LC3B, microtubule-associated protein 1 light chain 3 b; MAP2, microtube-associated protein 2; MPTP, 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine; PD, Parkinson disease; PEBP1, phosphatidylethanolamine binding protein 1; PI, propidium iodide; SNc, substantia nigra pars compacta; SNCA/a-synuclein, synuclein, a (non A4 component of amyloid precursor); TH, tyrosine hydroxylase.Autophagy is involved in the pathogenesis of neurodegenerative diseases including Parkinson disease (PD). However, little is known about the regulation of autophagy in neurodegenerative process. In this study, we characterized aberrant activation of autophagy induced by neurotoxin 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) and demonstrated that melatonin has a protective effect on neurotoxicity. We found an excessive activation of autophagy in monkey brain tissues and C6 cells, induced by MPTP, which is mediated by CDK5 (cyclin-dependent kinase 5). MPTP treatment significantly reduced total dendritic length and dendritic complexity of cultured primary cortical neurons and melatonin could reverse this effect. Decreased TH (tyrosine hydroxylase)-positive cells and dendrites of dopaminergic neurons in the substantia nigra pars compacta (SNc) were observed in MPTP-treated monkeys and mice. Along with decreased TH protein level, we observed an upregulation of CDK5 and enhanced autophagic activity in the striatum of mice with MPTP injection. These changes could be salvaged by melatonin treatment or knockdown of CDK5. Importantly, melatonin or knockdown of CDK5 reduced MPTP-induced SNCA/ a-synuclein aggregation in mice, which is widely thought to trigger the pathogenesis of PD. Finally, melatonin or knockdown of CDK5 counteracted the PD phenotype in mice induced by MPTP. Our findings uncover a potent role of CDK5-mediated autophagy in the pathogenesis of PD, and suggest that control of autophagic pathways may provide an important clue for exploring potential target for novel therapeutics of PD.

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“…In order to make counting of autophagosomes and autolysosomes (yellow and red fluorescent dots, respectively) reliable and basically independent of the operator, 62 the Green and Red Puncta Colocalization ImageJ Plugin (created by Daniel J. Shiwarski, University of Pittsburgh; enhanced and modified by Ruben K. Dagda, University of Nevada School of Medicine and coapproved as an image tool for autophagic flux by Charleen T. Chu, University of Pittsburgh) was used without any setting change or adjustment on a number (indicated in each relevant histogram) of single-cell images extracted from different microscopic fields (3 to 6 for each experimental condition). 63 Autophagy and lysosomal delivery of MT For detection of MT autophagy, a construct (pcDNA3-hMT2A-mCherry) encoding a human (Homo sapiens, Hs) MT2A-mCherry was generated by ligase-independent cloning of the Mt2a cDNA (PCR-amplified from pcDNA3-GFPHsMT2A; Addgene plasmid 11613, deposited by S. Johnson) into the pcDNA3-mCherry LIC cloning vector (6B) (Addgene plasmid 30125, deposited by S. Gradia). pcDNA3-HsMT2A-mCherry was cotransfected with the autophagosomal marker pEGFP-C1-LC3.…”

Section: Determination Of Autophagic Fluxmentioning

confidence: 99%

Autophagy of metallothioneins prevents TNF-induced oxidative stress and toxicity in hepatoma cells

et al. 2015

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, tandem fluorescent microtubule-associated protein 1 light chain 3 A/B (green/red fluorescent proteins chimera); TNFRSF1A/TNFR1, tumor necrosis factor receptor superfamily member 1A; TNFRSF1B/TNFR2, tumor necrosis factor receptor superfamily member 1B.Lysosomal membrane permeabilization (LMP) induced by oxidative stress has recently emerged as a prominent mechanism behind TNF cytotoxicity. This pathway relies on diffusion of hydrogen peroxide into lysosomes containing redox-active iron, accumulated by breakdown of iron-containing proteins and subcellular organelles. Upon oxidative lysosomal damage, LMP allows relocation to the cytoplasm of low mass iron and acidic hydrolases that contribute to DNA and mitochondrial damage, resulting in death by apoptosis or necrosis. Here we investigate the role of lysosomes and free iron in death of HTC cells, a rat hepatoma line, exposed to TNF following metallothionein (MT) upregulation. Iron-binding MT does not normally occur in HTC cells in significant amounts. Intracellular iron chelation attenuates TNF and cycloheximide (CHX)-induced LMP and cell death, demonstrating the critical role of this transition metal in mediating cytokine lethality. MT upregulation, combined with starvation-activated MT autophagy almost completely suppresses TNF and CHX toxicity, while impairment of both autophagy and MT upregulation by silencing of Atg7, and Mt1a and/or Mt2a, respectively, abrogates protection. Interestingly, MT upregulation by itself has little effect, while stimulated autophagy alone depresses cytokine toxicity to some degree. These results provide evidence that intralysosomal iron-catalyzed redox reactions play a key role in TNF and CHX-induced LMP and toxicity. The finding that chelation of intralysosomal iron achieved by autophagic delivery of MT, and to some degree probably of other ironbinding proteins as well, into the lysosomal compartment is highly protective provides a putative mechanism to explain autophagy-related suppression of death by TNF and CHX.

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Chapter 11 Autophagy in Neurite Injury and Neurodegeneration

Cited by 108 publications

References 91 publications

MIR517Cinhibits autophagy and the epithelial-to-mesenchymal (-like) transition phenotype in human glioblastoma through KPNA2-dependent disruption of TP53 nuclear translocation

MIR517Cinhibits autophagy and the epithelial-to-mesenchymal (-like) transition phenotype in human glioblastoma through KPNA2-dependent disruption of TP53 nuclear translocation

Melatonin attenuates MPTP-induced neurotoxicity via preventing CDK5-mediated autophagy and SNCA/α-synuclein aggregation

Autophagy of metallothioneins prevents TNF-induced oxidative stress and toxicity in hepatoma cells

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