{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,1]],"date-time":"2026-10-01T04:29:34Z","timestamp":1790828974687,"version":"4.1.0"},"reference-count":42,"publisher":"Public Library of Science (PLoS)","issue":"6","license":[{"start":{"date-parts":[[2021,6,4]],"date-time":"2021-06-04T00:00:00Z","timestamp":1622764800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"DARPA INTERCEPT","award":["HR0011-17-2-0023"],"award-info":[{"award-number":["HR0011-17-2-0023"]}]},{"DOI":"10.13039\/100014588","name":"Sanofi-Pasteur","doi-asserted-by":"crossref","award":["S-RD14004"],"award-info":[{"award-number":["S-RD14004"]}],"id":[{"id":"10.13039\/100014588","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS ONE"],"abstract":"<jats:p>Japanese encephalitis virus (JEV) is the major cause of viral encephalitis in South East Asia. It has been suggested that, as a consequence of the inflammatory process during JEV infection, there is disruption of the blood-brain barrier (BBB) tight junctions that in turn allows the virus access to the central nervous system (CNS). However, what happens at early times of JEV contact with the BBB is poorly understood. In the present work, we evaluated the ability of both a virulent and a vaccine strain of JEV (JEV RP9 and SA14-14-2, respectively) to cross an <jats:italic>in vitro<\/jats:italic> human BBB model. Using this system, we demonstrated that both JEV RP9 and SA14-14-2 are able to cross the BBB without disrupting it at early times post viral addition. Furthermore, we find that almost 10 times more RP9 infectious particles than SA14-14 cross the model BBB, indicating this BBB model discriminates between the virulent RP9 and the vaccine SA14-14-2 strains of JEV. Beyond contributing to the understanding of early events in JEV neuroinvasion, we demonstrate this <jats:italic>in vitro<\/jats:italic> BBB model can be used as a system to study the viral determinants of JEV neuroinvasiveness and the molecular mechanisms by which this flavivirus crosses the BBB during early times of neuroinvasion.<\/jats:p>","DOI":"10.1371\/journal.pone.0252595","type":"journal-article","created":{"date-parts":[[2021,6,4]],"date-time":"2021-06-04T18:47:33Z","timestamp":1622832453000},"page":"e0252595","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":8,"title":["Comparative analysis of neuroinvasion by Japanese encephalitis virulent and vaccine viral strains in an in vitro model of human blood-brain barrier"],"prefix":"10.1371","volume":"16","author":[{"given":"C\u00e9cile","family":"Khou","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2983-0123","authenticated-orcid":true,"given":"Marco Aurelio","family":"D\u00edaz-Salinas","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anaelle","family":"da Costa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christophe","family":"Pr\u00e9haud","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Patricia","family":"Jeannin","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Philippe V.","family":"Afonso","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marco","family":"Vignuzzi","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4423-7996","authenticated-orcid":true,"given":"Monique","family":"Lafon","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1973-1631","authenticated-orcid":true,"given":"Nathalie","family":"Pardigon","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2021,6,4]]},"reference":[{"key":"pone.0252595.ref001","unstructured":"Lindenbach BD, Thiel H-J, Rice CM. Flaviviridae: The viruses and their replication. In: Fields Virology. 5th ed. 2007. p. 1101\u201333."},{"issue":"10","key":"pone.0252595.ref002","doi-asserted-by":"crossref","first-page":"766","DOI":"10.2471\/BLT.10.085233","article-title":"Estimated global incidence of Japanese encephalitis: a systematic review","volume":"89","author":"GL Campbell","year":"2011","journal-title":"Bull World Health Organ"},{"issue":"4","key":"pone.0252595.ref003","doi-asserted-by":"crossref","first-page":"405","DOI":"10.1136\/jnnp.68.4.405","article-title":"Japanese encephalitis.","volume":"68","author":"T Solomon","year":"2000","journal-title":"J Neurol Neurosurg Psychiatry"},{"issue":"6","key":"pone.0252595.ref004","doi-asserted-by":"crossref","first-page":"513","DOI":"10.1016\/0264-410X(88)90103-X","article-title":"Japanese encephalitis virus live-attenuated vaccine, Chinese strain SA14-14-2; adaptation to primary canine kidney cell cultures and preparation of a vaccine for human use","volume":"6","author":"KH Eckels","year":"1988","journal-title":"Vaccine"},{"issue":"2","key":"pone.0252595.ref005","doi-asserted-by":"crossref","first-page":"263","DOI":"10.4161\/hv.26902","article-title":"Japanese encephalitis: the virus and vaccines.","volume":"10","author":"S-I Yun","year":"2014","journal-title":"Hum Vaccines Immunother"},{"issue":"4","key":"pone.0252595.ref006","doi-asserted-by":"crossref","first-page":"3095","DOI":"10.1128\/JVI.73.4.3095-3101.1999","article-title":"Yellow fever\/Japanese encephalitis chimeric viruses: construction and biological properties","volume":"73","author":"TJ Chambers","year":"1999","journal-title":"J Virol"},{"issue":"15","key":"pone.0252595.ref007","doi-asserted-by":"crossref","first-page":"1768","DOI":"10.1016\/j.vaccine.2013.11.107","article-title":"Safety of Japanese encephalitis live attenuated vaccination in post-marketing surveillance in Guangdong, China, 2005\u20132012.","volume":"32","author":"Y Liu","year":"2014","journal-title":"Vaccine"},{"issue":"1","key":"pone.0252595.ref008","doi-asserted-by":"crossref","first-page":"42","DOI":"10.1038\/scientificamerican0793-42","article-title":"Viral quasispecies.","volume":"269","author":"M Eigen","year":"1993","journal-title":"Sci Am"},{"issue":"2","key":"pone.0252595.ref009","doi-asserted-by":"crossref","first-page":"159","DOI":"10.1128\/MMBR.05023-11","article-title":"Viral quasispecies evolution","volume":"76","author":"E Domingo","year":"2012","journal-title":"Microbiol Mol Biol Rev MMBR"},{"issue":"8","key":"pone.0252595.ref010","doi-asserted-by":"crossref","first-page":"e2980","DOI":"10.1371\/journal.pntd.0002980","article-title":"Neuropathogenesis of Japanese encephalitis in a primate model.","volume":"8","author":"KSA Myint","year":"2014","journal-title":"PLoS Negl Trop Dis."},{"issue":"10","key":"pone.0252595.ref011","doi-asserted-by":"crossref","first-page":"5602","DOI":"10.1128\/JVI.00143-15","article-title":"Viral Infection of the Central Nervous System and Neuroinflammation Precede Blood-Brain Barrier Disruption during Japanese Encephalitis Virus Infection","volume":"89","author":"F Li","year":"2015","journal-title":"J Virol"},{"issue":"11","key":"pone.0252595.ref012","doi-asserted-by":"crossref","first-page":"1915","DOI":"10.1002\/glia.22857","article-title":"Disruption of in vitro endothelial barrier integrity by Japanese encephalitis virus-Infected astrocytes","volume":"63","author":"C-Y Chang","year":"2015","journal-title":"Glia"},{"issue":"2","key":"pone.0252595.ref013","doi-asserted-by":"crossref","first-page":"1150","DOI":"10.1128\/JVI.02738-13","article-title":"Infection of pericytes in vitro by Japanese encephalitis virus disrupts the integrity of the endothelial barrier","volume":"88","author":"C-J Chen","year":"2014","journal-title":"J Virol"},{"issue":"3","key":"pone.0252595.ref014","doi-asserted-by":"crossref","first-page":"389","DOI":"10.1007\/s004410051130","article-title":"Japanese encephalitis virus is transported across the cerebral blood vessels by endocytosis in mouse brain","volume":"293","author":"ML Liou","year":"1998","journal-title":"Cell Tissue Res"},{"issue":"3","key":"pone.0252595.ref015","doi-asserted-by":"crossref","first-page":"871","DOI":"10.1152\/physrev.00001.2003","article-title":"Transcytosis: crossing cellular barriers.","volume":"83","author":"PL Tuma","year":"2003","journal-title":"Physiol Rev"},{"issue":"10","key":"pone.0252595.ref016","doi-asserted-by":"crossref","first-page":"2575","DOI":"10.1099\/jgv.0.000574","article-title":"Comparison of the live-attenuated Japanese encephalitis vaccine SA14-14-2 strain with its pre-attenuated virulent parent SA14 strain: similarities and differences in vitro and in vivo","volume":"97","author":"S-I Yun","year":"2016","journal-title":"J Gen Virol"},{"issue":"7","key":"pone.0252595.ref017","doi-asserted-by":"crossref","first-page":"e1004290","DOI":"10.1371\/journal.ppat.1004290","article-title":"A molecularly cloned, live-attenuated japanese encephalitis vaccine SA14-14-2 virus: a conserved single amino acid in the ij Hairpin of the Viral E glycoprotein determines neurovirulence in mice.","volume":"10","author":"S-I Yun","year":"2014","journal-title":"PLoS Pathog."},{"issue":"12","key":"pone.0252595.ref018","doi-asserted-by":"crossref","first-page":"6328","DOI":"10.1128\/JVI.00219-15","article-title":"Genetic Determinants of Japanese Encephalitis Virus Vaccine Strain SA14-14-2 That Govern Attenuation of Virulence in Mice. Diamond MS, editor","volume":"89","author":"GD Gromowski","year":"2015","journal-title":"J Virol"},{"issue":"5","key":"pone.0252595.ref019","doi-asserted-by":"crossref","first-page":"2676","DOI":"10.1128\/JVI.01176-15","article-title":"A Single Amino Acid Substitution in the M Protein Attenuates Japanese Encephalitis Virus in Mammalian Hosts","volume":"90","author":"M de Wispelaere","year":"2015","journal-title":"J Virol"},{"issue":"3","key":"pone.0252595.ref020","doi-asserted-by":"crossref","first-page":"1641","DOI":"10.1128\/JVI.01979-09","article-title":"NS1\u2019 of flaviviruses in the Japanese encephalitis virus serogroup is a product of ribosomal frameshifting and plays a role in viral neuroinvasiveness","volume":"84","author":"EB Melian","year":"2010","journal-title":"J Virol"},{"issue":"Pt 9","key":"pone.0252595.ref021","doi-asserted-by":"crossref","first-page":"1959","DOI":"10.1099\/vir.0.043844-0","article-title":"A single nucleotide mutation in NS2A of Japanese encephalitis-live vaccine virus (SA14-14-2) ablates NS1\u2019 formation and contributes to attenuation.","volume":"93","author":"Q Ye","year":"2012","journal-title":"J Gen Virol"},{"issue":"3","key":"pone.0252595.ref022","doi-asserted-by":"crossref","first-page":"437","DOI":"10.1007\/s10545-013-9608-0","article-title":"Blood-brain barrier structure and function and the challenges for CNS drug delivery","volume":"36","author":"NJ Abbott","year":"2013","journal-title":"J Inherit Metab Dis"},{"issue":"5","key":"pone.0252595.ref023","doi-asserted-by":"crossref","first-page":"862","DOI":"10.1177\/0271678X16630991","article-title":"In vitro models of the blood-brain barrier: An overview of commonly used brain endothelial cell culture models and guidelines for their use","volume":"36","author":"HC Helms","year":"2016","journal-title":"J Cereb Blood Flow Metab Off J Int Soc Cereb Blood Flow Metab"},{"issue":"1","key":"pone.0252595.ref024","doi-asserted-by":"crossref","first-page":"16","DOI":"10.1186\/2045-8118-10-16","article-title":"The hCMEC\/D3 cell line as a model of the human blood brain barrier","volume":"10","author":"B Weksler","year":"2013","journal-title":"Fluids Barriers CNS"},{"issue":"146","key":"pone.0252595.ref025","article-title":"A Human Blood-Brain Interface Model to Study Barrier Crossings by Pathogens or Medicines and Their Interactions with the Brain.","volume":"09","author":"A da Costa","year":"2019","journal-title":"J Vis Exp JoVE"},{"key":"pone.0252595.ref026","first-page":"19","article-title":"Innovative in cellulo method as an alternative to in vivo neurovirulence test for the characterization and quality control of human live Yellow Fever virus vaccines: A pilot study.","volume":"53","author":"A da Costa","year":"2018","journal-title":"Biol J Int Assoc Biol Stand"},{"issue":"21","key":"pone.0252595.ref027","doi-asserted-by":"crossref","first-page":"2746","DOI":"10.1016\/j.vaccine.2009.03.007","article-title":"A Japanese encephalitis virus vaccine candidate strain is attenuated by decreasing its interferon antagonistic ability","volume":"27","author":"J-J Liang","year":"2009","journal-title":"Vaccine"},{"issue":"11","key":"pone.0252595.ref028","doi-asserted-by":"crossref","first-page":"5862","DOI":"10.1128\/JVI.00358-15","article-title":"A Japanese Encephalitis Virus Genotype 5 Molecular Clone Is Highly Neuropathogenic in a Mouse Model: Impact of the Structural Protein Region on Virulence","volume":"89","author":"M de Wispelaere","year":"2015","journal-title":"J Virol"},{"issue":"21","key":"pone.0252595.ref029","doi-asserted-by":"crossref","first-page":"10976","DOI":"10.1128\/JVI.00846-11","article-title":"Heterogeneous nuclear ribonucleoprotein A2 participates in the replication of Japanese encephalitis virus through an interaction with viral proteins and RNA","volume":"85","author":"H Katoh","year":"2011","journal-title":"J Virol"},{"issue":"1","key":"pone.0252595.ref030","doi-asserted-by":"crossref","first-page":"111","DOI":"10.1002\/jcp.1041320115","article-title":"Molecular sieving characteristics of the cultured endothelial monolayer","volume":"132","author":"A Siflinger-Birnboim","year":"1987","journal-title":"J Cell Physiol"},{"issue":"6","key":"pone.0252595.ref031","doi-asserted-by":"crossref","first-page":"e99733","DOI":"10.1371\/journal.pone.0099733","article-title":"A stable and reproducible human blood-brain barrier model derived from hematopoietic stem cells","volume":"9","author":"R Cecchelli","year":"2014","journal-title":"PloS One"},{"issue":"8","key":"pone.0252595.ref032","doi-asserted-by":"crossref","first-page":"783","DOI":"10.1038\/nbt.2247","article-title":"Derivation of blood-brain barrier endothelial cells from human pluripotent stem cells","volume":"30","author":"ES Lippmann","year":"2012","journal-title":"Nat Biotechnol"},{"issue":"4","key":"pone.0252595.ref033","first-page":"1535","article-title":"Unfractionated Heparin Selectively Modulates the Expression of CXCL8, CCL2 and CCL5 in Endometrial Carcinoma Cells","volume":"36","author":"A Doster","year":"2016","journal-title":"Anticancer Res"},{"issue":"5","key":"pone.0252595.ref034","doi-asserted-by":"crossref","first-page":"2618","DOI":"10.3390\/v7052618","article-title":"Insights into Human Astrocyte Response to H5N1 Infection by Microarray Analysis.","volume":"7","author":"X Lin","year":"2015","journal-title":"Viruses"},{"issue":"1","key":"pone.0252595.ref035","doi-asserted-by":"crossref","first-page":"706","DOI":"10.1038\/s41467-019-08641-z","article-title":"Japanese encephalitis virus neuropenetrance is driven by mast cell chymase","volume":"10","author":"JT Hsieh","year":"2019","journal-title":"Nat Commun"},{"issue":"5","key":"pone.0252595.ref036","doi-asserted-by":"crossref","first-page":"298","DOI":"10.1038\/nrneurol.2018.30","article-title":"Japanese encephalitis\u2014the prospects for new treatments.","volume":"2614","author":"L Turtle","year":"2018","journal-title":"Nat Rev Neurol"},{"key":"pone.0252595.ref037","doi-asserted-by":"crossref","first-page":"525","DOI":"10.3389\/fmicb.2019.00525","article-title":"Pathways Exploited by Flaviviruses to Counteract the Blood-Brain Barrier and Invade the Central Nervous System.","volume":"10","author":"YM Mustaf\u00e1","year":"2019","journal-title":"Front Microbiol."},{"issue":"6","key":"pone.0252595.ref038","doi-asserted-by":"crossref","first-page":"514","DOI":"10.1080\/13550280802339643","article-title":"The blood-brain barrier in the cerebrum is the initial site for the Japanese encephalitis virus entering the central nervous system","volume":"14","author":"T-H Liu","year":"2008","journal-title":"J Neurovirol"},{"issue":"7","key":"pone.0252595.ref039","doi-asserted-by":"crossref","first-page":"e69465","DOI":"10.1371\/journal.pone.0069465","article-title":"Japanese encephalitis virus disrupts cell-cell junctions and affects the epithelial permeability barrier functions.","volume":"8","author":"T Agrawal","year":"2013","journal-title":"PloS One"},{"key":"pone.0252595.ref040","doi-asserted-by":"crossref","first-page":"1148","DOI":"10.3389\/fimmu.2018.01148","article-title":"IP-10 Promotes Blood-Brain Barrier Damage by Inducing Tumor Necrosis Factor Alpha Production in Japanese Encephalitis.","volume":"9","author":"K Wang","year":"2018","journal-title":"Front Immunol."},{"issue":"2","key":"pone.0252595.ref041","doi-asserted-by":"crossref","first-page":"250","DOI":"10.1111\/j.1471-4159.2012.07889.x","article-title":"Endothelial Japanese encephalitis virus infection enhances migration and adhesion of leukocytes to brain microvascular endothelia via MEK-dependent expression of ICAM1 and the CINC and RANTES chemokines","volume":"123","author":"C-Y Lai","year":"2012","journal-title":"J Neurochem"},{"issue":"1","key":"pone.0252595.ref042","doi-asserted-by":"crossref","first-page":"461","DOI":"10.1128\/JVI.01894-07","article-title":"Effect of cell polarization on hepatitis C virus entry","volume":"82","author":"CJ Mee","year":"2008","journal-title":"J Virol"}],"container-title":["PLOS ONE"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0252595","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2021,6,4]],"date-time":"2021-06-04T18:48:22Z","timestamp":1622832502000},"score":1,"resource":{"primary":{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0252595"}},"subtitle":[],"editor":[{"given":"Slava","family":"Rom","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2021,6,4]]},"references-count":42,"journal-issue":{"issue":"6","published-online":{"date-parts":[[2021,6,4]]}},"URL":"https:\/\/doi.org\/10.1371\/journal.pone.0252595","relation":{},"ISSN":["1932-6203"],"issn-type":[{"value":"1932-6203","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,6,4]]}}}