title: Application of scanning cytometry and confocal-microscopy-based image analysis for investigation the role of cytoskeletal elements during equine herpesvirus type 1 (EHV-1) infection of primary murine neurons.
abstract: Equine herpesvirus type 1 (EHV-1), a member of Alphaherpesvirinae, has a broad host range in vitro, allowing for study of the mechanisms of productive viral infection, including intracellular transport in various cell cultures. In the current study, quantitative methods (scanning cytometry and real-time PCR) and confocal-microscopy-based image analysis were used to investigate the contribution of microtubules and neurofilaments in the transport of virus in primary murine neurons separately infected with two EHV-1 strains. Confocal-microscopy analysis revealed that viral antigen co-localized with the β-tubulin fibres within the neurites of infected cells. Alterations in β-tubulin and neurofilaments were evaluated by confocal microscopy and scanning cytometry. Real-time PCR analysis demonstrated that inhibitor-induced (nocodazole, EHNA) disruption of microtubules and dynein significantly reduced EHV-1 replication in neurons. Our results suggest that microtubules together with the motor protein - dynein, are involved in EHV-1 replication process in neurons. Moreover, the data presented here and our earlier results support the hypothesis that microtubules and actin filaments play an important role in the EHV-1 transport in primary murine neurons, and that both cytoskeletal structures complement each-other.
authors.affiliation: Division of Microbiology, Department of Preclinical Sciences, Faculty of Veterinary Medicine, Warsaw University of Life Sciences - SGGW, Ciszewskiego 8, 02-786 Warsaw, Poland; Division of Physiology, Department of Physiological Sciences, Faculty of Veterinary Medicine, Warsaw University of Life Sciences - SGGW, Nowoursynowska 159, 02-776 Warsaw, Poland. Electronic address: anex007@op.pl.
authors.forename: A
authors.initials: A
authors.lastname: Słońska
authors.affiliation: Division of Microbiology, Department of Preclinical Sciences, Faculty of Veterinary Medicine, Warsaw University of Life Sciences - SGGW, Ciszewskiego 8, 02-786 Warsaw, Poland. Electronic address: jcymerys@op.pl.
authors.forename: J
authors.initials: J
authors.lastname: Cymerys
authors.affiliation: Division of Physiology, Department of Physiological Sciences, Faculty of Veterinary Medicine, Warsaw University of Life Sciences - SGGW, Nowoursynowska 159, 02-776 Warsaw, Poland. Electronic address: mickgodl@hotmail.com.
authors.forename: M M
authors.initials: MM
authors.lastname: Godlewski
authors.affiliation: Division of Microbiology, Department of Preclinical Sciences, Faculty of Veterinary Medicine, Warsaw University of Life Sciences - SGGW, Ciszewskiego 8, 02-786 Warsaw, Poland. Electronic address: marcin_banbura@sggw.pl.
authors.forename: M W
authors.initials: MW
authors.lastname: Bańbura
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000648:ultrastructure*; D004398:Dyneins / Q000648:ultrastructure; D004861:Herpesvirus 1, Equid / Q000502:physiology*; D006736:Horses; D007091:Image Processing, Computer-Assisted / Q000379:methods; D007382:Intermediate Filaments / Q000648:ultrastructure / Q000821:virology; D047569:Laser Scanning Cytometry / Q000379:methods*; D051379:Mice; D018613:Microscopy, Confocal / Q000379:methods*; D008870:Microtubules / Q000648:ultrastructure / Q000821:virology; D009474:Neurons / Q000821:virology*; D014779:Virus Replication
chemical_list: D004398:Dyneins
keywords: Confocal microscopy; Cytoskeleton; EHV-1; Neurons; Scanning cytometry