INTRODUCTION: The genus Orthoflavivirus of the Flaviviridae family includes several notable pathogens such as mosquito-borne West-Nile virus (Orthoflavivirus nilense, WNV) and Tick-borne encephalitis virus (Orthoflavivirus encephalitidis, TBEV) that are highly neurotropic and may cause severe neurological disease leading to lifelong disabilities, coma and death. These viruses have developed mechanisms to breach the compact blood-brain barrier (BBB) and establish infection within the central nervous system (CNS). Nevertheless, neuroinvasive mechanisms of orthoflaviviruses remain poorly understood. Complex anatomy of the CNS and the organization of the BBB is a major challenge to study neuroinvasion of orthoflaviviruses in vivo. Therefore, in vitro BBB models are useful tools to study direct interaction of viruses with the endothelial barrier. METHODS: In this study, we employed an in vitro transwell BBB model comprising primary mouse brain microvascular endothelial cells and astrocytes to compare the ability of mosquito-borne and tick-borne orthoflaviviruses to cross a compact endothelial barrier and reach the basolateral compartment of the transwell system. The influence of virus inoculation on the barrier properties was determined by measuring transendothelial electrical resistance (TEER). RESULTS: The results demonstrate that while pathogenic WNV and TBEV cross the endothelial barrier the ability of low pathogenic Usutu virus (USUV) and Langat virus (LGTV) was inconsistent. All viruses tested display virus replication within the endothelial cells. Nevertheless, virus replication did not affect the barrier function of endothelial cells as demonstrated by sustained TEER and absence of leakage of high molecular weight dextran molecules through the endothelial barrier even at several hours post infection. DISCUSSION: Our findings indicate that orthoflaviviruses can infect the endothelial cells, replicate within them without affecting the cells and its barrier function. Nevertheless, only pathogenic WNV and TBEV showed the ability to cross the endothelial barrier and reach the basolateral compartment.

• Klíčová slova
• astrocytes, blood-brain barrier, endothelial cells, neuroinvasion, orthoflavivirus, transendothelial electrical resistance,
• MeSH
• astrocyty * virologie   MeSH
• Culicidae virologie   MeSH
• endoteliální buňky * virologie   MeSH
• hematoencefalická bariéra * virologie   MeSH
• kultivované buňky MeSH
• myši MeSH
• replikace viru MeSH
• viry klíšťové encefalitidy * fyziologie   patogenita   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

The aim of this review is to follow the history of studies on endemiv arboviruses and the diseases they cause which were detected in the Czech lands (Bohemia, Moravia and Silesia (i.e., the Czech Republic)). The viruses involve tick-borne encephalitis, West Nile and Usutu flaviviruses; the Sindbis alphavirus; Ťahyňa, Batai, Lednice and Sedlec bunyaviruses; the Uukuniemi phlebovirus; and the Tribeč orbivirus. Arboviruses temporarily imported from abroad to the Czech Republic have been omitted. This brief historical review includes a bibliography of all relevant papers.

• Klíčová slova
• arthropods, birds, mammals, mosquitoes, ticks,
• MeSH
• arbovirové infekce dějiny   MeSH
• arboviry fyziologie   MeSH
• dějiny 20. století MeSH
• dějiny 21. století MeSH
• lidé MeSH
• zvířata MeSH
• Check Tag
• dějiny 20. století MeSH
• dějiny 21. století MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• historické články MeSH
• přehledy MeSH
• Geografické názvy
• Česká republika epidemiologie   MeSH

BACKGROUND: Tick-borne encephalitis (TBE) is a severe neuropathological disorder caused by tick-borne encephalitis virus (TBEV). Brain TBEV infection is characterized by extensive pathological neuroinflammation. The mechanism by which TBEV causes CNS destruction remains unclear, but growing evidence suggests that it involves both direct neuronal damage by the virus infection and indirect damage caused by the immune response. Here, we aimed to examine the TBEV-infection-induced innate immune response in mice and in human neural cells. We also compared cytokine/chemokine communication between naïve and infected neuronal cells and astrocytes. METHODS: We used a multiplexed Luminex system to measure multiple cytokines/chemokines and growth factors in mouse serum samples and brain tissue, and in human neuroblastoma cells (SK-N-SH) and primary cortical astrocytes (HBCA), which were infected with the highly pathogenic TBEV strain Hypr. We also investigated changes in cytokine/chemokine production in naïve HBCA cells treated with virus-free supernatants from TBEV-infected SK-N-SH cells and in naïve SK-N-SH cells treated with virus-free supernatants from TBEV-infected HBCA cells. Additionally, a plaque assay was performed to assess how cytokine/chemokine treatment influenced viral growth following TBEV infection. RESULTS: TBEV-infected mice exhibited time-dependent increases in serum and brain tissue concentrations of multiple cytokines/chemokines (mainly CXCL10/IP-10, and also CXCL1, G-CSF, IL-6, and others). TBEV-infected SK-N-SH cells exhibited increased production of IL-8 and RANTES and downregulated MCP-1 and HGF. TBEV infection of HBCA cells activated production of a broad spectrum of pro-inflammatory cytokines, chemokines, and growth factors (mainly IL-6, IL-8, CXCL10, RANTES, and G-CSF) and downregulated the expression of VEGF. Treatment of SK-N-SH with supernatants from infected HBCA induced expression of a variety of chemokines and pro-inflammatory cytokines, reduced SK-N-SH mortality after TBEV infection, and decreased virus growth in these cells. Treatment of HBCA with supernatants from infected SK-N-SH had little effect on cytokine/chemokine/growth factor expression but reduced TBEV growth in these cells after infection. CONCLUSIONS: Our results indicated that both neurons and astrocytes are potential sources of pro-inflammatory cytokines in TBEV-infected brain tissue. Infected/activated astrocytes produce cytokines/chemokines that stimulate the innate neuronal immune response, limiting virus replication, and increasing survival of infected neurons.

• Klíčová slova
• Luminex, Neuroinflammation, Tick-borne encephalitis, Tick-borne encephalitis virus,
• MeSH
• cytokiny imunologie   metabolismus   MeSH
• klíšťová encefalitida imunologie   metabolismus   MeSH
• lidé MeSH
• mozek imunologie   metabolismus   patologie   MeSH
• myši MeSH
• neurony imunologie   metabolismus   virologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• cytokiny MeSH

Tick-borne encephalitis virus (TBEV) causes serious, potentially fatal neurological infections that affect humans in endemic regions of Europe and Asia. Neurons are the primary target for TBEV infection in the central nervous system. However, knowledge about this viral infection and virus-induced neuronal injury is fragmental. Here, we directly examined the pathology that occurs after TBEV infection in human primary neurons. We exploited the advantages of advanced high-pressure freezing and freeze-substitution techniques to achieve optimal preservation of infected cell architecture. Electron tomographic (ET) reconstructions elucidated high-resolution 3D images of the proliferating endoplasmic reticulum, and individual tubule-like structures of different diameters in the endoplasmic reticulum cisternae of single cells. ET revealed direct connections between the tubule-like structures and viral particles in the endoplasmic reticulum. Furthermore, ET showed connections between cellular microtubules and vacuoles that harbored the TBEV virions in neuronal extensions. This study was the first to characterize the 3D topographical organization of membranous whorls and autophagic vacuoles in TBEV-infected human neurons. The functional importance of autophagy during TBEV replication was studied in human neuroblastoma cells; stimulation of autophagy resulted in significantly increased dose-dependent TBEV production, whereas the inhibition of autophagy showed a profound, dose-dependent decrease of the yield of infectious virus.

• MeSH
• autofagie účinky léků   genetika   MeSH
• benzylaminy farmakologie   MeSH
• chinazoliny farmakologie   MeSH
• endoplazmatické retikulum účinky léků   ultrastruktura   virologie   MeSH
• lidé MeSH
• mikrotubuly účinky léků   ultrastruktura   virologie   MeSH
• nádorové buněčné linie MeSH
• neurony účinky léků   ultrastruktura   virologie   MeSH
• nokodazol farmakologie   MeSH
• primární buněčná kultura MeSH
• replikace viru účinky léků   MeSH
• sirolimus farmakologie   MeSH
• tomografie elektronová MeSH
• virion účinky léků   růst a vývoj   ultrastruktura   MeSH
• viry klíšťové encefalitidy účinky léků   růst a vývoj   ultrastruktura   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• benzylaminy MeSH
• chinazoliny MeSH
• nokodazol MeSH
• sirolimus MeSH
• spautin-1 MeSH Prohlížeč

BACKGROUND: The clinical course of tick-borne encephalitis (TBE), a disease caused by TBE virus, ranges from asymptomatic or mild influenza-like infection to severe debilitating encephalitis or encephalomyelitis. Despite the medical importance of this disease, some crucial steps in the development of encephalitis remain poorly understood. In particular, the basis of the disease severity is largely unknown. METHODS: TBE virus growth, neutralizing antibody response, key cytokine and chemokine mRNA production and changes in mRNA levels of cell surface markers of immunocompetent cells in brain were measured in mice with different susceptibilities to TBE virus infection. RESULTS: An animal model of TBE based on BALB/c-c-STS/A (CcS/Dem) recombinant congenic mouse strains showing different severities of the infection in relation to the host genetic background was developed. After subcutaneous inoculation of TBE virus, BALB/c mice showed medium susceptibility to the infection, STS mice were resistant, and CcS-11 mice were highly susceptible. The resistant STS mice showed lower and delayed viremia, lower virus production in the brain and low cytokine/chemokine mRNA production, but had a strong neutralizing antibody response. The most sensitive strain (CcS-11) failed in production of neutralizing antibodies, but exhibited strong cytokine/chemokine mRNA production in the brain. After intracerebral inoculation, all mouse strains were sensitive to the infection and had similar virus production in the brain, but STS mice survived significantly longer than CcS-11 mice. These two strains also differed in the expression of key cytokines/chemokines, particularly interferon gamma-induced protein 10 (IP-10/CXCL10) and monocyte chemotactic protein-1 (MCP-1/CCL2) in the brain. CONCLUSIONS: Our data indicate that the genetic control is an important factor influencing the clinical course of TBE. High neutralizing antibody response might be crucial for preventing host fatality, but high expression of various cytokines/chemokines during TBE can mediate immunopathology and be associated with more severe course of the infection and increased fatality.

• MeSH
• buněčná imunita imunologie   MeSH
• centrální nervový systém patologie   MeSH
• chemokiny biosyntéza   MeSH
• cytokiny biosyntéza   MeSH
• genotyp MeSH
• klíšťová encefalitida imunologie   patologie   MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• membránové proteiny biosyntéza   MeSH
• messenger RNA biosyntéza   genetika   MeSH
• mozek - chemie fyziologie   MeSH
• myši inbrední BALB C MeSH
• myši MeSH
• náchylnost k nemoci MeSH
• neutralizující protilátky biosyntéza   MeSH
• odolnost vůči nemocem MeSH
• plakové testy MeSH
• protilátky virové biosyntéza   genetika   MeSH
• virová nálož MeSH
• viry klíšťové encefalitidy * MeSH
• zánět patologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chemokiny MeSH
• cytokiny MeSH
• membránové proteiny MeSH
• messenger RNA MeSH
• neutralizující protilátky MeSH
• protilátky virové MeSH