{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,27]],"date-time":"2026-08-27T18:20:19Z","timestamp":1787854819218,"version":"build-2784847793"},"reference-count":52,"publisher":"Public Library of Science (PLoS)","issue":"8","license":[{"start":{"date-parts":[[2026,8,27]],"date-time":"2026-08-27T00:00:00Z","timestamp":1787788800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001509","name":"Royal Society of New Zealand","doi-asserted-by":"crossref","award":["JCF-UOA2001"],"award-info":[{"award-number":["JCF-UOA2001"]}],"id":[{"id":"10.13039\/501100001509","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/501100001509","name":"Royal Society of New Zealand","doi-asserted-by":"crossref","award":["19-UOA-043-CSG"],"award-info":[{"award-number":["19-UOA-043-CSG"]}],"id":[{"id":"10.13039\/501100001509","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/501100001537","name":"The University of Auckland","doi-asserted-by":"crossref","id":[{"id":"10.13039\/501100001537","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:p>\n                    The mesenchymal subtype of the WHO grade IV Glioblastoma (GBM) is the most common and lethal form of adult brain cancer, characterised by its rapid growth, invasiveness and colonisation. Current standard of care includes surgical resection, chemotherapy and radiotherapy. Despite this, mortality remains dismal with median patient survival of less than 15 months after treatment. For these reasons, the World Health Organisation (WHO) classifies GBM as an incurable disease. To address this challenge, the GBM scientific community are investigating new approaches to gain insights into GBM function and its communication mechanisms. These efforts aim to establish alternative therapies to slow or halt the infiltration of this aggressive cancer. In this article we demonstrate the variability between two adult patient-derived mesenchymal GBM brain cancer cell-lines and show how they can be patterned onto grid networks of biocompatible parylene-C on SiO\n                    <jats:sub>2<\/jats:sub>\n                    substrates. Thus, providing a first step for GBM Ca\n                    <jats:sup>2+<\/jats:sup>\n                    communication to be repeatedly and reliably studied. We determine the node size, node spacing and track length that provides the best cytoplasmic coverage of parylene-C for GBM brain cancer cells using five patterning indices, namely: the parylene-C adhesion index (PAI), SiO\n                    <jats:sub>2<\/jats:sub>\n                    adhesion index (SAI), node nuclei index (NNI), grid quality index (GQI), and overall patterning index (OPI). In addition, we determine the best serum, cell seeding density and incubation period\n                    <jats:italic>in vitro<\/jats:italic>\n                    to encourage GBM cell patterning. We illustrate high fidelity of the patterned GBM brain cancer networks and to demonstrate cell functionality we show that the patterned GBM cells respond to ATP stimulus with Ca\n                    <jats:sup>2+<\/jats:sup>\n                    transients. The significance of this work is that we provide a biomaterial platform that enables the detailed investigation of Ca\n                    <jats:sup>2+<\/jats:sup>\n                    communication within GBM tumour networks. Such a platform may enable the GBM scientific community to test new therapeutics that target the Ca\n                    <jats:sup>2+<\/jats:sup>\n                    signalling which is involved in the rapid infiltrative tumour growth, a property which currently makes treatment ineffective.\n                  <\/jats:p>","DOI":"10.1371\/journal.pone.0352239","type":"journal-article","created":{"date-parts":[[2026,8,27]],"date-time":"2026-08-27T17:34:29Z","timestamp":1787852069000},"page":"e0352239","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":0,"title":["First cell-patterning of primary, patient-derived mesenchymal glioblastoma brain cancer cells on parylene-C\/SiO2 substrates"],"prefix":"10.1371","volume":"21","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-9995-0097","authenticated-orcid":true,"given":"Nicholas G.","family":"Mellor","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sylvia A.","family":"Chung","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Pierrette","family":"Michaux","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Josiah","family":"Firth","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Scott E.","family":"Graham","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bryan W.","family":"Day","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Charles P.","family":"Unsworth","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2026,8,27]]},"reference":[{"issue":"6","key":"pone.0352239.ref001","doi-asserted-by":"crossref","first-page":"803","DOI":"10.1007\/s00401-016-1545-1","article-title":"The 2016 World Health Organization Classification of Tumors of the Central Nervous System: a summary","volume":"131","author":"DN Louis","year":"2016","journal-title":"Acta Neuropathol"},{"key":"pone.0352239.ref002","doi-asserted-by":"crossref","first-page":"15017","DOI":"10.1038\/nrdp.2015.17","article-title":"Glioma","volume":"1","author":"M Weller","year":"2015","journal-title":"Nat Rev Dis Primers"},{"key":"pone.0352239.ref003","doi-asserted-by":"crossref","DOI":"10.5772\/intechopen.72720","article-title":"The role of astrocytes in tumor growth and progression","author":"E Gronseth","year":"2018","journal-title":"Astrocyte - Physiology and Pathology"},{"issue":"3","key":"pone.0352239.ref004","doi-asserted-by":"crossref","first-page":"162","DOI":"10.1097\/00001622-199905000-00004","article-title":"Molecular pathogenesis of malignant gliomas","volume":"11","author":"BK Rasheed","year":"1999","journal-title":"Curr Opin Oncol"},{"issue":"41","key":"pone.0352239.ref005","doi-asserted-by":"crossref","first-page":"13827","DOI":"10.1523\/JNEUROSCI.2603-15.2015","article-title":"New insights on astrocyte ion channels: critical for homeostasis and neuron-glia signaling","volume":"35","author":"ML Olsen","year":"2015","journal-title":"J Neurosci"},{"issue":"1","key":"pone.0352239.ref006","doi-asserted-by":"crossref","DOI":"10.1016\/j.ccell.2018.05.012","article-title":"Apoptotic cell-derived extracellular vesicles promote malignancy of glioblastoma via intercellular transfer of splicing factors","volume":"34","author":"MS Pavlyukov","year":"2018","journal-title":"Cancer Cell"},{"issue":"8","key":"pone.0352239.ref007","doi-asserted-by":"crossref","first-page":"482","DOI":"10.1038\/s41582-018-0025-8","article-title":"Multidimensional communication in the microenvirons of glioblastoma","volume":"14","author":"ML Broekman","year":"2018","journal-title":"Nat Rev Neurol"},{"issue":"17","key":"pone.0352239.ref008","doi-asserted-by":"crossref","first-page":"15566","DOI":"10.18632\/oncotarget.3904","article-title":"Gap junctions modulate glioma invasion by direct transfer of microRNA","volume":"6","author":"X Hong","year":"2015","journal-title":"Oncotarget"},{"issue":"7","key":"pone.0352239.ref009","doi-asserted-by":"crossref","first-page":"517","DOI":"10.1038\/nrm1155","article-title":"Calcium signalling: dynamics, homeostasis and remodelling","volume":"4","author":"MJ Berridge","year":"2003","journal-title":"Nat Rev Mol Cell Biol"},{"issue":"6","key":"pone.0352239.ref010","doi-asserted-by":"crossref","first-page":"367","DOI":"10.1038\/nrc.2017.18","article-title":"The calcium-cancer signalling nexus","volume":"17","author":"GR Monteith","year":"2017","journal-title":"Nat Rev Cancer"},{"key":"pone.0352239.ref011","doi-asserted-by":"crossref","first-page":"587","DOI":"10.1016\/j.lfs.2010.09.013","article-title":"Calcium wave signaling in cancer cells","volume":"87","author":"J Parkash","year":"2010","journal-title":"Life Sci"},{"issue":"16","key":"pone.0352239.ref012","doi-asserted-by":"crossref","first-page":"3044","DOI":"10.1016\/j.biomaterials.2005.12.024","article-title":"Surface engineering approaches to micropattern surfaces for cell-based assays","volume":"27","author":"D Falconnet","year":"2006","journal-title":"Biomaterials"},{"issue":"3","key":"pone.0352239.ref013","doi-asserted-by":"crossref","first-page":"398","DOI":"10.1109\/JPROC.2009.2039029","article-title":"Designing Neural Networks in Culture: Experiments are described for controlled growth, of nerve cells taken from rats, in predesigned geometrical patterns on laboratory culture dishes","volume":"98","author":"BC Wheeler","year":"2010","journal-title":"Proc IEEE Inst Electr Electron Eng"},{"issue":"5016","key":"pone.0352239.ref014","doi-asserted-by":"crossref","first-page":"1183","DOI":"10.1038\/2081183a0","article-title":"Principles of cell motility: the direction of cell movement and cancer invasion","volume":"208","author":"SB Carter","year":"1965","journal-title":"Nature"},{"issue":"11","key":"pone.0352239.ref015","doi-asserted-by":"crossref","first-page":"4098","DOI":"10.1523\/JNEUROSCI.08-11-04098.1988","article-title":"Controlled outgrowth of dissociated neurons on patterned substrates","volume":"8","author":"D Kleinfeld","year":"1988","journal-title":"J Neurosci"},{"key":"pone.0352239.ref016","doi-asserted-by":"crossref","first-page":"550","DOI":"10.1002\/(SICI)1521-3773(19980316)37:5<550::AID-ANIE550>3.0.CO;2-G","article-title":"Soft lithography","volume":"37","author":"Y Xia","year":"1998","journal-title":"Angew Chem Int Ed Engl"},{"issue":"2","key":"pone.0352239.ref017","doi-asserted-by":"crossref","first-page":"362","DOI":"10.1016\/0014-4827(88)90275-3","article-title":"Cytoscribing: a method for micropositioning cells and the construction of two- and three-dimensional synthetic tissues","volume":"179","author":"RJ Klebe","year":"1988","journal-title":"Exp Cell Res"},{"key":"pone.0352239.ref018","doi-asserted-by":"crossref","first-page":"1855","DOI":"10.1016\/j.msec.2009.02.016","article-title":"Patterning multiple cell types in co-cultures: a review","volume":"29","author":"CA Goubko","year":"2009","journal-title":"Materials Science and Engineering: C"},{"issue":"5","key":"pone.0352239.ref019","doi-asserted-by":"crossref","first-page":"1350","DOI":"10.1002\/jbm.a.34813","article-title":"Patterning human neuronal networks on photolithographically engineered silicon dioxide substrates functionalized with glial analogues","volume":"102","author":"MA Hughes","year":"2014","journal-title":"J Biomed Mater Res A"},{"issue":"14","key":"pone.0352239.ref020","doi-asserted-by":"crossref","first-page":"2036","DOI":"10.1039\/b820596e","article-title":"Three-dimensional micro-electrode array for recording dissociated neuronal cultures","volume":"9","author":"K Musick","year":"2009","journal-title":"Lab Chip"},{"issue":"23","key":"pone.0352239.ref021","doi-asserted-by":"crossref","first-page":"11718","DOI":"10.1021\/la7017049","article-title":"Cell and protein compatibility of parylene-C surfaces","volume":"23","author":"TY Chang","year":"2007","journal-title":"Langmuir"},{"key":"pone.0352239.ref022","doi-asserted-by":"crossref","first-page":"3027","DOI":"10.1002\/pol.1966.150041209","article-title":"A new, general synthetic method for the preparation of linear poly-p-xylylenes","volume":"4","author":"WF Gorham","year":"1966","journal-title":"J Polym Sci A1"},{"issue":"2","key":"pone.0352239.ref023","doi-asserted-by":"crossref","first-page":"195","DOI":"10.1002\/jbm.820110206","article-title":"Histological reaction to various conductive and dielectric films chronically implanted in the subdural space","volume":"11","author":"GE Loeb","year":"1977","journal-title":"J Biomed Mater Res"},{"issue":"1","key":"pone.0352239.ref024","doi-asserted-by":"crossref","first-page":"96","DOI":"10.1007\/BF02441836","article-title":"Long-term implants of Parylene-C coated microelectrodes","volume":"26","author":"EM Schmidt","year":"1988","journal-title":"Med Biol Eng Comput"},{"issue":"6","key":"pone.0352239.ref025","doi-asserted-by":"crossref","first-page":"1342","DOI":"10.1109\/JMEMS.2008.2004945","article-title":"Microfabricated implantable parylene-based wireless passive intraocular pressure sensors","volume":"17","author":"Chen Po-Jui","year":"2008","journal-title":"J Microelectromech Syst"},{"issue":"11","key":"pone.0352239.ref026","doi-asserted-by":"crossref","first-page":"2048","DOI":"10.1016\/j.biomaterials.2008.12.049","article-title":"Guided growth of neurons and glia using microfabricated patterns of parylene-C on a SiO2 background","volume":"30","author":"E Delivopoulos","year":"2009","journal-title":"Biomaterials"},{"key":"pone.0352239.ref027","doi-asserted-by":"crossref","first-page":"169","DOI":"10.1016\/j.colsurfb.2014.12.020","article-title":"Serum protein layers on parylene-C and silicon oxide: effect on cell adhesion","volume":"126","author":"E Delivopoulos","year":"2015","journal-title":"Colloids Surf B Biointerfaces"},{"issue":"1","key":"pone.0352239.ref028","doi-asserted-by":"crossref","first-page":"47","DOI":"10.1002\/jbm.a.32662","article-title":"Effects of parylene-C photooxidation on serum-assisted glial and neuronal patterning","volume":"94","author":"E Delivopoulos","year":"2010","journal-title":"J Biomed Mater Res A"},{"issue":"10","key":"pone.0352239.ref029","doi-asserted-by":"crossref","first-page":"2566","DOI":"10.1016\/j.biomaterials.2010.12.017","article-title":"Isolating single primary rat hippocampal neurons & astrocytes on ultra-thin patterned parylene-C\/silicon dioxide substrates","volume":"32","author":"CP Unsworth","year":"2011","journal-title":"Biomaterials"},{"issue":"1","key":"pone.0352239.ref030","doi-asserted-by":"crossref","first-page":"154","DOI":"10.1016\/j.jneumeth.2010.09.022","article-title":"First human hNT neurons patterned on parylene-C\/silicon dioxide substrates: combining an accessible cell line and robust patterning technology for the study of the pathological adult human brain","volume":"194","author":"CP Unsworth","year":"2010","journal-title":"J Neurosci Methods"},{"issue":"27","key":"pone.0352239.ref031","doi-asserted-by":"crossref","first-page":"6541","DOI":"10.1016\/j.biomaterials.2011.05.041","article-title":"Patterning and detailed study of human hNT astrocytes on parylene-C\/silicon dioxide substrates to the single cell level","volume":"32","author":"CP Unsworth","year":"2011","journal-title":"Biomaterials"},{"issue":"2","key":"pone.0352239.ref032","doi-asserted-by":"crossref","first-page":"349","DOI":"10.1002\/jbm.a.34329","article-title":"Modulating patterned adhesion and repulsion of HEK 293 cells on microengineered parylene-C\/SiO(2) substrates","volume":"101","author":"MA Hughes","year":"2013","journal-title":"J Biomed Mater Res A"},{"key":"pone.0352239.ref033","doi-asserted-by":"crossref","first-page":"117","DOI":"10.1016\/j.biomaterials.2016.08.006","article-title":"Human astrocytic grid networks patterned in parylene-C inlayed SiO2 trenches","volume":"105","author":"MD Jordan","year":"2016","journal-title":"Biomaterials"},{"issue":"6","key":"pone.0352239.ref034","article-title":"Evaluation of parylene derivatives for use as biomaterials for human astrocyte cell patterning","volume":"14","author":"BJ Raos","year":"2019","journal-title":"PLoS One"},{"issue":"1","key":"pone.0352239.ref035","doi-asserted-by":"crossref","first-page":"2754","DOI":"10.1038\/s41598-018-21135-0","article-title":"Selective PEGylation of parylene-C\/SiO2 substrates for improved astrocyte cell patterning","volume":"8","author":"BJ Raos","year":"2018","journal-title":"Sci Rep"},{"issue":"6","key":"pone.0352239.ref036","doi-asserted-by":"crossref","first-page":"066001","DOI":"10.1088\/1741-2552\/ab39cc","article-title":"Large 10\u2009\u2009\u00d7\u2009\u200910 single cell grid networks of human hNT astrocytes on raised parylene-C\/SiO2 substrates","volume":"16","author":"S Li","year":"2019","journal-title":"J Neural Eng"},{"issue":"3","key":"pone.0352239.ref037","article-title":"Geometric micro-shapes facilitate trackless connections between human astrocytes","volume":"18","author":"S Li","year":"2021","journal-title":"J Neural Eng"},{"issue":"12","key":"pone.0352239.ref038","doi-asserted-by":"crossref","first-page":"3174","DOI":"10.3390\/ma14123174","article-title":"Adhesion and growth of neuralized mouse embryonic stem cells on parylene-C\/SiO2 substrates","volume":"14","author":"AF Murray","year":"2021","journal-title":"Materials (Basel)"},{"issue":"10","key":"pone.0352239.ref039","doi-asserted-by":"crossref","first-page":"1272","DOI":"10.1039\/b706081e","article-title":"Generation of static and dynamic patterned co-cultures using microfabricated parylene-C stencils","volume":"7","author":"D Wright","year":"2007","journal-title":"Lab Chip"},{"issue":"2","key":"pone.0352239.ref040","doi-asserted-by":"crossref","first-page":"530","DOI":"10.1002\/jbm.a.31281","article-title":"Reusable, reversibly sealable parylene membranes for cell and protein patterning","volume":"85","author":"D Wright","year":"2008","journal-title":"J Biomed Mater Res A"},{"issue":"6","key":"pone.0352239.ref041","first-page":"30","article-title":"Biocompatible parylene neurocages. Developing a robust method for live neural network studies","volume":"24","author":"A Tooker","year":"2005","journal-title":"IEEE Eng Med Biol Mag"},{"key":"pone.0352239.ref042","article-title":"Q-cell glioblastoma resource: proteomics analysis reveals unique cell-states are maintained in 3D culture","volume":"9","author":"RCJ D\u2019Souza","year":"2020","journal-title":"Cells"},{"issue":"1","key":"pone.0352239.ref043","doi-asserted-by":"crossref","first-page":"4902","DOI":"10.1038\/s41598-019-41277-z","article-title":"A reference collection of patient-derived cell line and xenograft models of proneural, classical and mesenchymal glioblastoma","volume":"9","author":"BW Stringer","year":"2019","journal-title":"Sci Rep"},{"key":"pone.0352239.ref044","volume-title":"Digital Image Processing","author":"RC Gonzalez","year":"2008","edition":"3"},{"key":"pone.0352239.ref045","doi-asserted-by":"crossref","DOI":"10.1002\/9780470316672","author":"Y Hochberg","year":"1987","journal-title":"Multiple Comparison Procedures"},{"issue":"9","key":"pone.0352239.ref046","doi-asserted-by":"crossref","first-page":"1187","DOI":"10.1002\/jnr.23399","article-title":"Characterization of the NT2-derived neuronal and astrocytic cell lines as alternative in vitro models for primary human neurons and astrocytes","volume":"92","author":"Y Haile","year":"2014","journal-title":"J Neurosci Res"},{"key":"pone.0352239.ref047","doi-asserted-by":"crossref","first-page":"53","DOI":"10.1007\/s11010-018-3272-5","article-title":"GL261 glioma tumor cells respond to ATP with an intracellular calcium rise and glutamate release","volume":"446","author":"AD Strong","year":"2018","journal-title":"Mol Cell Biochem"},{"issue":"2","key":"pone.0352239.ref048","doi-asserted-by":"crossref","first-page":"659","DOI":"10.1152\/physrev.00043.2006","article-title":"Physiology and pathophysiology of purinergic neurotransmission","volume":"87","author":"G Burnstock","year":"2007","journal-title":"Physiol Rev"},{"issue":"6","key":"pone.0352239.ref049","doi-asserted-by":"crossref","DOI":"10.1088\/1741-2552\/ad0e7d","article-title":"Eliciting calcium transients with UV nanosecond laser stimulation in adult patient-derived glioblastoma brain cancer cellsin vitro","volume":"20","author":"NG Mellor","year":"2023","journal-title":"J Neural Eng"},{"issue":"1","key":"pone.0352239.ref050","article-title":"Tumor microenvironment in glioblastoma: current and emerging concepts","volume":"5","author":"P Sharma","year":"2023","journal-title":"Neurooncol Adv"},{"issue":"7580","key":"pone.0352239.ref051","doi-asserted-by":"crossref","first-page":"93","DOI":"10.1038\/nature16071","article-title":"Brain tumour cells interconnect to a functional and resistant network","volume":"528","author":"M Osswald","year":"2015","journal-title":"Nature"},{"issue":"7775","key":"pone.0352239.ref052","doi-asserted-by":"crossref","first-page":"532","DOI":"10.1038\/s41586-019-1564-x","article-title":"Glutamatergic synaptic input to glioma cells drives brain tumour progression","volume":"573","author":"V Venkataramani","year":"2019","journal-title":"Nature"}],"container-title":["PLOS One"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0352239","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,8,27]],"date-time":"2026-08-27T17:34:39Z","timestamp":1787852079000},"score":1,"resource":{"primary":{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0352239"}},"subtitle":[],"editor":[{"given":"Lei","family":"Zhang","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2026,8,27]]},"references-count":52,"journal-issue":{"issue":"8","published-online":{"date-parts":[[2026,8,27]]}},"URL":"https:\/\/doi.org\/10.1371\/journal.pone.0352239","relation":{},"ISSN":["1932-6203"],"issn-type":[{"value":"1932-6203","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,8,27]]}}}