2c0ccbf697aec9dd361b6e63f856c7d6 pmc_33530486.pdf ae4f03277217f1ccd77b06b18d59fce0c79361fc pmc_33530486.pdf 0094866c63a4eeba9e43795a8995ed850dea63adfc3f5824515972b502f43879 pmc_33530486.pdf Title: Ursolic Acid Inhibits Collective Cell Migration and Promotes JNK-Dependent Lysosomal Associated Cell Death in Glioblastoma Multiforme Cells Subject: Ursolic acid (UA) is a bioactive compound which has demonstrated therapeutic efficacy in a variety of cancer cell lines. UA activates various signalling pathways in Glioblastoma multiforme (GBM) and offers a promising starting point in drug discovery; however, understanding the relationship between cell death and migration has yet to be elucidated. UA induces a dose dependent cytotoxic response demonstrated by flow cytometry and biochemical cytotoxicity assays. Inhibitor and fluorescent probe studies demonstrate that UA induces a caspase independent, JNK dependent, mechanism of cell death. Migration studies established that UA inhibits GBM collective cell migration in a time dependent manner that is independent of the JNK signalling pathway. Cytotoxicity induced by UA results in the formation of acidic vesicle organelles (AVOs), speculating the activation of autophagy. However, inhibitor and spectrophotometric analysis demonstrated that autophagy was not responsible for the formation of the AVOs. Confocal microscopy and isosurface visualisation determined co-localisation of lysosomes with the previously identified AVOs, thus providing evidence that lysosomes are likely to be playing a role in UA induced cell death. Collectively, our data identify that UA rapidly induces a lysosomal associated mechanism of cell death in addition to UA acting as an inhibitor of GBM collective cell migration. Keywords: ursolic acid; cell death; migration; lysosomes; nutraceuticals Author: Gillian E. Conway, Deimante Zizyte, Julie Rose Mae Mondala, Zhonglei He, Lorna Lynam, Mathilde Lecourt, Carlos Barcia, Orla Howe and James F. Curtin Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Mon Feb 1 07:13:55 2021 CET ModDate: Mon Feb 1 07:19:13 2021 CET Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 16 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 3245866 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- HKVNWV+VnURWPalladioL Type 1 Custom yes yes no 10 0 FMEOLT+URWPalladioL-Ital Type 1 Custom yes yes no 15 0 KGLNWE+URWPalladioL-Bold Type 1 Custom yes yes no 20 0 VHBRID+URWPalladioL-Roma Type 1 Custom yes yes no 24 0 VWRBXB+TimesNewRomanPS-BoldItalicMT Type 1C WinAnsi yes yes no 32 0 MCBCCI+SourceSansRoman-Regular Type 1C Custom yes yes no 54 0 MCBCCM+SourceSansRoman.613wght Type 1C Custom yes yes no 58 0 MCBCCO+SourceSansRoman-Bold Type 1C Custom yes yes no 62 0 PDYJSF+EURM10 Type 1 Builtin yes yes no 74 0 IYAOHD+PazoMath Type 1 Builtin yes yes no 81 0 QMDDXF+CMSY10 Type 1 Builtin yes yes no 85 0 Jhove (Rel. 1.22.1, 2019-04-17) Date: 2021-09-11 03:50:03 CEST RepresentationInformation: pmc_33530486.pdf ReportingModule: PDF-hul, Rel. 1.12.1 (2019-04-17) LastModified: 2021-03-29 03:38:00 CEST Size: 3245866 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 381 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@13c27452 Item: Title: Results Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@262b2c86 Children: Item: Title: UA Induced Cytotoxicity Results in Rapid Mitochondrial Membrane Depolarisation Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@371a67ec Item: Title: UA Demonstrates Enhanced Cytotoxicity Compared to Conventional Chemotherapeutic Drugs Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@5ed828d Item: Title: UA Inhibits JNK-Dependent GBM Collective Cell Migration Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@50d0686 Item: Title: UA Induces Caspase-Independent, JNK-Dependent Cell Death Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@7a3d45bd Item: Title: UA Triggers the Formation of Lysosomes Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1e7c7811 Item: Title: Discussion Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@77ec78b9 Item: Title: Materials and Methods Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1a3869f4 Children: Item: Title: Cell Culture Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@a38d7a3 Item: Title: Cytotoxicity Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@77f99a05 Item: Title: Cell Viability Assays Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@63440df3 Item: Title: Flow Cytometry Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@3aeaafa6 Children: Item: Title: JC-1 Mitochondrial Membrane Potential Assay Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@76a3e297 Item: Title: Acridine Orange (AO) Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@4d3167f4 Item: Title: Scratch Assay Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@ed9d034 Item: Title: Inhibitor Studies Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@6121c9d6 Item: Title: Confocal Microscopy Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@87f383f Item: Title: Isosurface Rendering Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@4eb7f003 Item: Title: Statistical Analysis Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@eafc191 Item: Title: References Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@612fc6eb Info: Title: Ursolic Acid Inhibits Collective Cell Migration and Promotes JNK-Dependent Lysosomal Associated Cell Death in Glioblastoma Multiforme Cells Author: Gillian E. Conway, Deimante Zizyte, Julie Rose Mae Mondala, Zhonglei He, Lorna Lynam, Mathilde Lecourt, Carlos Barcia, Orla Howe and James F. Curtin Subject: Ursolic acid (UA) is a bioactive compound which has demonstrated therapeutic efficacy in a variety of cancer cell lines. UA activates various signalling pathways in Glioblastoma multiforme (GBM) and offers a promising starting point in drug discovery; however, understanding the relationship between cell death and migration has yet to be elucidated. UA induces a dose dependent cytotoxic response demonstrated by flow cytometry and biochemical cytotoxicity assays. Inhibitor and fluorescent probe studies demonstrate that UA induces a caspase independent, JNK dependent, mechanism of cell death. Migration studies established that UA inhibits GBM collective cell migration in a time dependent manner that is independent of the JNK signalling pathway. Cytotoxicity induced by UA results in the formation of acidic vesicle organelles (AVOs), speculating the activation of autophagy. However, inhibitor and spectrophotometric analysis demonstrated that autophagy was not responsible for the formation of the AVOs. Confocal microscopy and isosurface visualisation determined co-localisation of lysosomes with the previously identified AVOs, thus providing evidence that lysosomes are likely to be playing a role in UA induced cell death. Collectively, our data identify that UA rapidly induces a lysosomal associated mechanism of cell death in addition to UA acting as an inhibitor of GBM collective cell migration. 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