Zika Virus Infects, Activates, and Crosses Brain Microvascular Endothelial Cells, without Barrier Disruption

Zika virus (ZIKV) has been associated to central nervous system (CNS) harm, and virus was detected in the brain and cerebrospinal fluids of microcephaly and meningoencephalitis cases. However, the mechanism by which the virus reaches the CNS is unclear. Here, we addressed the effects of ZIKV replication in human brain microvascular endothelial cells (HBMECs), as an in vitro model of blood brain barrier (BBB), and evaluated virus extravasation and BBB integrity in an in vivo mouse experimental model. HBMECs were productively infected by African and Brazilian ZIKV strains (ZIKVMR766 and ZIKVPE243), which induce increased production of type I and type III IFN, inflammatory cytokines and chemokines. Infection with ZIKVMR766 promoted earlier cellular death, in comparison to ZIKVPE243, but infection with either strain did not result in enhanced endothelial permeability. Despite the maintenance of endothelial integrity, infectious virus particles crossed the monolayer by endocytosis/exocytosis-dependent replication pathway or by transcytosis. Remarkably, both viruses' strains infected IFNAR deficient mice, with high viral load being detected in the brains, without BBB disruption, which was only detected at later time points after infection. These data suggest that ZIKV infects and activates endothelial cells, and might reach the CNS through basolateral release, transcytosis or transinfection processes. These findings further improve the current knowledge regarding ZIKV dissemination pathways.

[1]  F. Thompson,et al.  Development of standard methods for Zika virus propagation, titration, and purification. , 2017, Journal of virological methods.

[2]  J. Schwedes,et al.  Zika Virus Persistently Infects and Is Basolaterally Released from Primary Human Brain Microvascular Endothelial Cells , 2017, mBio.

[3]  K. Kim,et al.  A Three-Dimensional Cell Culture System To Model RNA Virus Infections at the Blood-Brain Barrier , 2017, mSphere.

[4]  Daniel Lucey,et al.  Congenital Zika Syndrome in 2017. , 2017, JAMA.

[5]  D. Alcendor,et al.  Zika virus infection of cellular components of the blood-retinal barriers: implications for viral associated congenital ocular disease , 2017, Journal of Neuroinflammation.

[6]  R. Albrecht,et al.  A novel Zika virus mouse model reveals strain specific differences in virus pathogenesis and host inflammatory immune responses , 2017, PLoS pathogens.

[7]  R. DeBiasi,et al.  Zika Virus Infection With Prolonged Maternal Viremia and Fetal Brain Abnormalities , 2017 .

[8]  M. Juzych,et al.  Zika virus infects cells lining the blood-retinal barrier and causes chorioretinal atrophy in mouse eyes. , 2017, JCI insight.

[9]  S. Klein,et al.  Intrauterine Zika virus infection of pregnant immunocompetent mice models transplacental transmission and adverse perinatal outcomes , 2017, Nature Communications.

[10]  P. Garcez,et al.  Chloroquine, an Endocytosis Blocking Agent, Inhibits Zika Virus Infection in Different Cell Models , 2016, Viruses.

[11]  Alex A. Pollen,et al.  Zika virus cell tropism in the developing human brain and inhibition by azithromycin , 2016, Proceedings of the National Academy of Sciences.

[12]  F. T. Moll,et al.  Zika Virus Causing Encephalomyelitis Associated With Immunoactivation , 2016, Open forum infectious diseases.

[13]  R. Lanciotti,et al.  Pathology of congenital Zika syndrome in Brazil: a case series , 2016, The Lancet.

[14]  V. Thongboonkerd,et al.  Caveolae-mediated albumin transcytosis is enhanced in dengue-infected human endothelial cells: A model of vascular leakage in dengue hemorrhagic fever , 2016, Scientific Reports.

[15]  Sallie R. Permar,et al.  A rhesus macaque model of Asian-lineage Zika virus infection , 2016, Nature Communications.

[16]  Carlos G Schrago,et al.  Detection and sequencing of Zika virus from amniotic fluid of fetuses with microcephaly in Brazil: a case study. , 2016, The Lancet. Infectious diseases.

[17]  R. Tesh,et al.  Characterization of a Novel Murine Model to Study Zika Virus , 2016, The American journal of tropical medicine and hygiene.

[18]  G. Saade,et al.  Zika virus: History, emergence, biology, and prospects for control. , 2016, Antiviral research.

[19]  M. Diamond,et al.  Zika Virus Infection during Pregnancy in Mice Causes Placental Damage and Fetal Demise , 2016, Cell.

[20]  M. Diamond,et al.  A Mouse Model of Zika Virus Pathogenesis. , 2016, Cell host & microbe.

[21]  S. Cherry,et al.  Type III Interferons Produced by Human Placental Trophoblasts Confer Protection against Zika Virus Infection. , 2016, Cell host & microbe.

[22]  Amadou A. Sall,et al.  The Brazilian Zika virus strain causes birth defects in experimental models , 2016, Nature.

[23]  L. de Noronha,et al.  Zika virus damages the human placental barrier and presents marked fetal neurotropism , 2016, Memorias do Instituto Oswaldo Cruz.

[24]  Giovanini Evelim Coelho,et al.  Zika virus in the Americas: Early epidemiological and genetic findings , 2016, Science.

[25]  S. Rasmussen,et al.  Zika Virus and Birth Defects--Reviewing the Evidence for Causality. , 2016, The New England journal of medicine.

[26]  L. Rodrigues,et al.  Initial Description of the Presumed Congenital Zika Syndrome. , 2016, American journal of public health.

[27]  Pierre Brugières,et al.  Zika Virus Associated with Meningoencephalitis. , 2016, The New England journal of medicine.

[28]  Graham Hall,et al.  A Susceptible Mouse Model for Zika Virus Infection , 2016, bioRxiv.

[29]  Marko Kolenc,et al.  Zika Virus Associated with Microcephaly. , 2016, The New England journal of medicine.

[30]  L. Schuler‐Faccini,et al.  Possible Association Between Zika Virus Infection and Microcephaly - Brazil, 2015. , 2016, MMWR. Morbidity and mortality weekly report.

[31]  F. Cabreiro.  Metformin Joins Forces with Microbes. , 2016, Cell host & microbe.

[32]  T. Bossert,et al.  Innovation and Change in the Chilean Health System. , 2016, The New England journal of medicine.

[33]  G. Malinger,et al.  Zika virus intrauterine infection causes fetal brain abnormality and microcephaly: tip of the iceberg? , 2016, Ultrasound in obstetrics & gynecology : the official journal of the International Society of Ultrasound in Obstetrics and Gynecology.

[34]  Herman Tse,et al.  Comparative genomic analysis of pre-epidemic and epidemic Zika virus strains for virological factors potentially associated with the rapidly expanding epidemic , 2016, Emerging Microbes & Infections.

[35]  M. Diamond,et al.  Mechanisms of restriction of viral neuroinvasion at the blood–brain barrier , 2015, Current Opinion in Immunology.

[36]  T. Dermody,et al.  Comparison of three neurotropic viruses reveals differences in viral dissemination to the central nervous system. , 2016, Virology.

[37]  M. Diamond,et al.  Cell Host & Microbe Resource A Mouse Model of Zika Virus Pathogenesis , 2016 .

[38]  N. Saunders,et al.  Markers for blood-brain barrier integrity: how appropriate is Evans blue in the twenty-first century and what are the alternatives? , 2015, Front. Neurosci..

[39]  Y. Ou,et al.  Disruption of in vitro endothelial barrier integrity by Japanese encephalitis virus‐Infected astrocytes , 2015, Glia.

[40]  R. Klein,et al.  Viral sensing at the blood–brain barrier: New roles for innate immunity at the CNS vasculature , 2015, Clinical pharmacology and therapeutics.

[41]  Mandy B. Esch,et al.  TEER Measurement Techniques for In Vitro Barrier Model Systems , 2015, Journal of laboratory automation.

[42]  W. T. Hancock,et al.  Zika Virus, French Polynesia, South Pacific, 2013 , 2014, Emerging infectious diseases.

[43]  V. Cao-Lormeau,et al.  Zika Virus, French Polynesia, South Pacific, 2013 , 2014, Emerging infectious diseases.

[44]  Hung-Chuan Pan,et al.  Infection of Pericytes In Vitro by Japanese Encephalitis Virus Disrupts the Integrity of the Endothelial Barrier , 2013, Journal of Virology.

[45]  J. Palefsky,et al.  Epstein-Barr Virus Transcytosis through Polarized Oral Epithelial Cells , 2013, Journal of Virology.

[46]  L. B. de Arruda,et al.  Essential role of RIG-I in the activation of endothelial cells by dengue virus. , 2013, Virology.

[47]  G. Zimmerman,et al.  Statins Decrease Neuroinflammation and Prevent Cognitive Impairment after Cerebral Malaria , 2012, PLoS pathogens.

[48]  Oscar A. Negrete,et al.  Rift Valley Fever Virus Strain MP-12 Enters Mammalian Host Cells via Caveola-Mediated Endocytosis , 2012, Journal of Virology.

[49]  J. Castellanos,et al.  In vivo infection by a neuroinvasive neurovirulent dengue virus , 2012, Journal of NeuroVirology.

[50]  S. M. Robinson,et al.  Human Immunodeficiency Virus-1 Uses the Mannose-6-Phosphate Receptor to Cross the Blood-Brain Barrier , 2012, PloS one.

[51]  Giampietro Schiavo,et al.  A hitchhiker's guide to the nervous system: the complex journey of viruses and toxins , 2010, Nature Reviews Microbiology.

[52]  Austin S Nakatsuka,et al.  West Nile virus infection modulates human brain microvascular endothelial cells tight junction proteins and cell adhesion molecules: Transmigration across the in vitro blood-brain barrier. , 2009, Virology.

[53]  R. Lanciotti,et al.  Genetic and Serologic Properties of Zika Virus Associated with an Epidemic, Yap State, Micronesia, 2007 , 2008, Emerging infectious diseases.

[54]  A. Olsen,et al.  Increased blood-brain barrier permeability is not a primary determinant for lethality of West Nile virus infection in rodents. , 2008, The Journal of general virology.

[55]  R. Diefenbach,et al.  Transport and egress of herpes simplex virus in neurons , 2008, Reviews in medical virology.

[56]  M. Diamond,et al.  Axonal transport mediates West Nile virus entry into the central nervous system and induces acute flaccid paralysis , 2007, Proceedings of the National Academy of Sciences.

[57]  J. Scharfstein,et al.  Blood-brain barrier traversal by African trypanosomes requires calcium signaling induced by parasite cysteine protease. , 2006, The Journal of clinical investigation.

[58]  V. Racaniello.  One hundred years of poliovirus pathogenesis. , 2006, Virology.

[59]  Matthias Mack,et al.  Modulating CCR2 and CCL2 at the blood-brain barrier: relevance for multiple sclerosis pathogenesis. , 2006, Brain : a journal of neurology.

[60]  S. Finke,et al.  Replication strategies of rabies virus. , 2005, Virus research.

[61]  E. Fikrig,et al.  Toll-like receptor 3 mediates West Nile virus entry into the brain causing lethal encephalitis , 2004, Nature Medicine.

[62]  M. Nedergaard,et al.  The blood–brain barrier: an overview Structure, regulation, and clinical implications , 2004, Neurobiology of Disease.

[63]  M. Liou,et al.  Japanese encephalitis virus is transported across the cerebral blood vessels by endocytosis in mouse brain , 1998, Cell and Tissue Research.

[64]  Y. Ikehara,et al.  Brefeldin A causes disassembly of the Golgi complex and accumulation of secretory proteins in the endoplasmic reticulum. , 1988, The Journal of biological chemistry.

[65]  R. Janzer,et al.  Astrocytes induce blood–brain barrier properties in endothelial cells , 1987, Nature.

[66]  G. Dick,et al.  Zika virus. I. Isolations and serological specificity. , 1952, Transactions of the Royal Society of Tropical Medicine and Hygiene.