Resumo
Arboviruses are worldwide distributed arthropod-borne viruses representing a constant threat to public health. Among these arboviruses, the Chikungunya (CHIKV) and Zika (ZIKV) viruses have a high prevalence in Brazil being responsible for recent outbreaks resulting mainly in irreparable socioeconomic damages such as the high rate of cases of comorbidities and microcephaly in newborns, respectively. Therefore, it is necessary to understand the biology of these arboviruses and develop effective treatments against them; moreover, appropriate mice animal models are strongly encouraged. Here we reviewed the scientific literature aiming to improve the search for the best murine animal model, specific for the arboviruses, specifically, CHIKV and ZIKV. In this way, we performed a comparison between the various mice models currently available, among them: genetically modified immunosuppressed animals, as the A129 and AG129 which are knockout animals for the α/ß and α/ß/γ receptors, respectively, neonatal immunocompetent models C57BL/6 strains used between 6-8 days old for neuropathogenesis studies or 1 day old for vaccine safety studies and finally immunosuppressed induced by dexamethasone or interferon 1 blocker for pathogenesis studies. Mice models are the first option after in vitro analysis, as they are small animals, which facilitates handling and maintenance, in addition to being more inexpensive and abundantly available in different genetic strains, both wild and modified. If the results of this stage are promising, the studies move forward to the use of models with animals of greater complexity, such as rats, non-human primates and finally humans. For this review, we searched through articles in PubMed, Scopus and ScienceDirect databases using the criteria of date publications, titles, abstracts and complete manuscripts. The correct choice of these models during experimental planning is essential, since increases the confidence and the rational use of animals in experimentation in accordance to current bioethics guidelines.
Assuntos
Animais , Camundongos , Vírus Chikungunya , Modelos Animais , Zika virus , Animais de Laboratório , CamundongosResumo
Purpose: To investigate the effect of hyperbaric oxygen therapy (HBOT) on traumatic brain injury (TBI) outcome. Methods: The modified Marmarou's weight drop device was used to generate non-lethal moderate TBI rat model, and further developed in vitro astrocytes culturing system. Then, we analyzed the expression changes of interested genes and protein by quantitative PCR and western blot. Results: Multiple HBO treatments significantly reduced the expression of apoptosis promoting genes, such as c-fos, c-jun, Bax and weakened the activation of Caspase-3 in model rats. On the contrary, HBOT alleviated the decrease of anti-apoptosis gene Bcl-2 and promoted the expression of neurotrophic factors (NTFs), such as NGF, BDNF, GDNF and NT-3 in vivo. As a consequent, the neuropathogenesis was remarkably relied with HBOT. Astrocytes from TBI brain or those cultured with 21% O2 density expressed higher NTFs than that of corresponding controls, from sham brain and cultured with 7% O2, respectively. The NTFs expression was the highest in astrocytes form TBI brain and cultured with 21% O2, suggesting a synergistic effect existed between TBI and the following HBO treatment in astrocytes. Conclusion: Our findings provided evidence for the clinical usage of HBO treating brain damages.(AU)
Assuntos
Animais , Masculino , Adulto , Ratos , Lesões Encefálicas Traumáticas/induzido quimicamente , Lesões Encefálicas Traumáticas/terapia , Oxigenoterapia Hiperbárica , Fatores de Crescimento Neural , Ratos Sprague-Dawley , Modelos AnimaisResumo
O objetivo deste estudo foi revisar a neuropatogenia causada pelo Morbillivirus em cães, através de revisão de literatura sistemática com ênfase na fisiopatogenia da cinomose. Foram analisadas amostras de encéfalos de cães com diagnóstico de cinomose que vieram a óbito, com base em análise histopatológica das lesões. A cinomose é uma doença multissistêmica grave, altamente contagiosa, com altas taxas de morbidade e mortalidade. É necessário um estudo completo da neuropatogenia da cinomose, abrangendo a etiopagenia e as complicações neurológicas da doença para elucidar detalhadamente as consequências dessa infecção, como as lesões desmielinizantes multifocais crônicas. Nessas patologias, os nucleocapsídeos virais por não utilizarem envelopamento em partículas virais para a sua multiplicação, são transmitidos para as células nervosas através de receptores, principalmente os astrócitos. A fusão célula a célula depende da nectina-4, presente nas cepas desmielinizantes do vírus da cinomose (VCC). Algumas pesquisas mostram que o vírus da cinomose acomete o SNC, mesmo que a infecção não tenha gerado alterações neurológicas. Os mecanismos de invasão e disseminação cerebral do Morbillivirus ainda não são totalmente conhecidos. A análise das alterações neuropatológicas causadas pela presença do VCC pode auxiliar estudos de novos tratamentos que visam à recuperação de sinais neurológicos de cães acometidos pela cinomose, que resultam nas principais causas de óbitos.
The aim of this study was to review the neuropathogenesis caused by Morbillivirus in dogs through systematic literature review with emphasis on the pathophysiology of canine distemper. Encephalic samples from dogs diagnosed with canine distemper that died were analyzed based on histopathological analysis of the lesions. Canine distemper is a severe, highly contagious, multisystemic disease with high morbidity and mortality rates. A complete study of the neuropathogenesis of canine distemper, covering the etiopathogenesis and neurological complications of the disease is necessary to elucidate in detail the consequences of this infection in the Central Nervous System (CNS), such as chronic multifocal demyelinating lesions. In these pathologies, the viral nucleocapsids by they do not use envelopment in viral particles for their multiplication, they are transmitted to the nerve cells through receptors, mainly the astrocytes. Cell-to-cell fusion depends on nectin-4, which is present in demyelinating strains of cynomatosis virus (DCV). Some research shows that the distemper virus affects the CNS, even if the infection has not generated neurological changes. The mechanisms of brain invasion and dissemination of Morbillivirus are not yet fully known. The analysis of neuropathological changes caused by the presence of CCV to support studies of new treatments aimed at the recovery of neurological signs in dogs affected by canine distemper, which results in the main causes of death.
Resumo
A leishmaniose visceral (LV) é uma doença parasitária que apresenta distribuição mundial e que pode afetar homens e animais, sendo que o cão é considerado o principal hospedeiro da doença. Cães infectados pelo parasito Leishmania podem apresentar-se assintomáticos ou com desordens generalizadas, incluindo alterações neurológicas. Existem alguns relatos do acometimento do encéfalo durante a infecção, mas a neuropatogenia da doença não foi completamente elucidada. Há evidências do comprometimento das barreiras encefálicas e da presença do DNA do parasito no encéfalo. Os receptores tipo Toll (TLRs) são sensores do sistema imune inato capazes de detectar padrões moleculares associados aos patógenos (PAMPs), desencadeando uma resposta inflamatórias com produção de diversos mediadores inflamatórios, incluindo citocinas. Desta forma, o objetivo deste estudo foi avaliar o perfil de expressão gênica dos Tolls 1-10, assim como a produção de citocinas pró-inflamatórias TNF-, IFN-, IL-1 e IL-6 no encéfalo e no baço de cães com leishmaniose visceral. No baço houve aumento de expressão gênica de TLR-5 e TLR-9, enquanto no encéfalo houve aumento de TLR-4 em uma pequena população de cães infectados. Em relação às citocinas, todas as citocinas foram detectadas nos dois tecidos avaliados, com excessão de IL-6. Nos cães infectados, TNF- e IL-1 estavam presentes em maiores concentrações no encéfalo e no baço, respectivamente. Este estudo fornece suporte para explicar o envolvimento de TLRs na LV e nossos dados confirmam o envolvimento encefálico durante a doença.
Visceral leishmaniasis (VL) is a parasitic disease that presents world distribution, affecting humans and animals. Dogs are considered the main hosts of the disease. Infected dogs with the Leishmania parasite can be asymptomatic or present generalized disorders, including neurological alterations. There are some reports of brain commitment during infection. Nevertheless, neuropathogenesis of VL is not completely elucidated. There are evidences of brain barriers breakdown and of the presence of Leishmania DNA in the brain. Toll-like receptors (TLRs) are innate immune sensors capable of detecting pathogen-associated molecular patterns (PAMPs), trigger an inflammatory response with production of several inflammatory mediators, including cytokines. Therefore, the aim of this study was to evaluate gene expression profile of TLRs1-10, along with the production of proinflammatory cytokines in both brain and spleen in dogs with VL. In spleen there was an up-regulation of TLR-5 and TLR-9 while in the brain there was up-regulation of TLR-4 in a few number of infected animals. Regarding cytokines, all cytokines were detected in both tissues, except IL-6. In the infected dogs, TNF- and IL-1 were present at higher concentrations in the brain and spleen, respectively. This study provides support to explain the involvement of TLRs in VL and our data confirm the brain as an affected organ in this disease.
Resumo
Bovine herpesvirus type 5 (BoHV-5) is the agent of meningoencephalitis, an important disease of cattle in South America. The neuropathogenesis of BoHV-5 infection is poorly understood and most previous research focused on the role of envelope glicoproteins in neurovirulence. Thymidine kinase (TK) is a viral enzyme necessary for virus replication in neurons and, therefore, represents a potential target for virus attenuation. The selection and characterization of BoHV-5 variants resistant to the nucleoside analog brivudin (BVDU), which selects TK-defective viruses is here described. Several BVDU-resistant clones were obtained after multiple passages in tissue culture in the presence of BVDU and one clone (BoHV-5/R-27) was further characterized. The selected clone replicated to similar titers and produced plaques with similar size and morphology to those of wild-type virus (SV507/99). The genetic stability of the resistant virus was demonstrated after ten passages in cell culture in the absence of the drug. Moreover, the drug-resistant virus showed reduced virulence in a rabbit model: virus inoculation in four rabbits did not result in disease, in contrast with 75% morbidity (3/4) and 50% mortality (2/2) among rabbits inoculated with the parental virus. These results demonstrate that BoHV-5 is sensitive to BVDU and that drug-resistant mutants can be readily selected upon BVDU treatment. BVDU-resistant mutants, likely defective in TK, retained their ability to replicate in tissue culture yet were attenuated for rabbits. This strategy to obtain TK-defective BoHV-5 may be useful to study the role of TK in BoHV-5 neuropathogenesis and for vaccine development.
Resumo
A doença neurológica causada pela replicação do BoHV-5 no cérebro de animais infectados, em sua fase inicial, pode não estar associada à alterações histológicas significativas ou à um número significativo de neurônios antígeno-positivos. Assim, outros mecanismos, como os mecanismos oxidantes e antioxidantes e importantes enzimas do sistema purinérgico, poderiam modular a resposta imune e inflamatória em animais infectados pelo herpesvírus bovino tipo 5 (BoHV-5) e contribuir para o desenvolvimento dos sinais neurológicos observados durante a infecção. Com este trabalho buscou-se compreender mecanismos da neuropatogenia do BoHV-5 durante infecção aguda. Para isso, no artigo I, foram avaliadas alterações na hidrólise de nucleotídeos de adenina em sinaptossomas de córtex e hipocampo, de coelhos experimentalmente infectados com BoHV-5, através da atividade das enzimas NTPDase e 5-nucleotidase; no artigo II, foi avaliada a participação de mecanismos oxidantes e antioxidantes na infecção pelo BoHV-5 em nível sistêmico e de sistema nervoso central de coelhos, através da atividade dos antioxidantes catalase (CAT), glutationa reduzida (GSH) e tióis não-proteicos (TSH) e quantificação de espécies reativas de oxigênio totais (ERO-totais) e as espécies reativas ao ácido tiobarbitúrico (TBARS). Para realização dos experimentos, os animais foram divididos em grupos, grupos controle, não infectados, e grupos teste, inoculados com BoHV-5. Os animais dos grupos teste foram inoculados com a cepa parental SV-507/99 contendo aproximadamente 107,5TCID50 ou 108TCID50, os demais animais receberam apenas meio essencial mínimo (MEM). Todos os coelhos foram inoculados pela via intranasal e monitorados quanto aos aspectos clínicos e virológicos durante todo o experimento. Aos 7 e 12 dias p.i. os coelhos foram anestesiados e submetidos à eutanásia para coleta sanguínea e de estruturas encefálicas. Em relação à atividade da NTPDase e 5-nucleotidase (artigo I), os resultados revelaram uma diminuição na hidrólise do ATP e ADP aos 7 dias p.i., e na hidrólise do ADP e AMP aos 12 dias p.i. em sinaptossomas de córtex cerebral dos animais infectados; e um aumento da atividade ectonucleotidásica em sinaptossomas de hipocampo no grupo infectado em relação ao grupo controle, com exceção da hidrólise do AMP aos 12 dias p.i. A reduzida hidrólise do ATP no córtex cerebral poderia causar acúmulo deste nucleotídeo no meio extracelular, sabe-se que o excesso de ATP pode ser citotóxico. Além disso, com a diminuição da atividade enzimática menos adenosina é produzida, sendo esta uma molécula neuroprotetora e anticonvulsivante. O aumento da atividade enzimática no hipocampo poderia ocorrer para produzir adenosina e conferir neuroproteção, no entanto, aos 12 dias p.i. não ocorreu aumento na hidrólise do AMP à adenosina. No artigo II, a avaliação dos parâmetros oxidativos e antioxidantes revelou que ocorre um aumento nos níveis de TBARS e de ERO-totais no grupo infectado em relação ao grupo controle, principalmente no córtex aos 7 e 12 dias p.i., mas também no cerebelo aos 7 dias p.i. e no hipocampo e estriado aos 12 dias p.i. Ainda, os níveis de GSH estavam diminuídos no estriado e cerebelo dos animais infectados aos 7 dias p.i. O processo de estresse oxidativo pode ocorrer em resposta à infecção viral, porém por ter baixa especificidade acaba resultando em danos oxidativos à célula. Desta forma, foi possível observar que existe uma participação do sistema purinérgico e do processo de estresse oxidativo na patogênese da infecção pelo BoHV-5, sugerindo que estes mecanismos possam estar envolvidos na imunomodulação, disfunção e morte neuronal, contribuindo, assim, para a processo da meningoencefalite herpética.
The neurological disorder caused by BoHV-5 replication in the brain of infected animals, in the acute phase, may not be associated with significant histological changes or a significant number of antigen-positive neurons. Thus, other mechanisms, such as oxidants and antioxidants mechanisms and important enzymes of the purinergic system might modulate the immune and inflammatory response in animals infected with bovine herpesvirus type 5 (BoHV-5) and contribute to the development of neurological signs observed during infection. This study aimed to understand the mechanisms of the neuropathogenesis of BoHV-5 during acute infection. For this, in article I, alterations in the hydrolysis of adenine nucleotides in synaptosomes of the cortex and hippocampus of rabbits experimentally infected with BoHV-5 were assessed through the activity of enzymes NTPDase and 5'-nucleotidase; in article II, we evaluated the involvement of oxidants and antioxidants mechanisms during BoHV-5 infection in systemic level and central nervous system of rabbits experimentally infected through the evaluation of catalase (CAT), reduced glutathione (GSH) and non-protein thiols (TSH), and the quantitation of reactive oxygen species (ROS) and thiobarbituric acid reactive species (TBARS). For the experiments, the animals were divided into groups, control groups, with no infected animals, and test groups, animals inoculated with BoHV-5. The animals of the test groups were inoculated with the parental strain SV-507/99 containing approximately 107,5TCID50 or 108TCID50, control animals received only minimal essential medium (MEM). All rabbits were inoculated intranasally and monitored for clinical and virological aspects during the experiment. At 7 and 12 days p.i. the rabbits were anesthetized and euthanized for blood and brain structures collection. Regarding the activity of 5'-nucleotidase and NTPDase (Article I), the results showed a decrease in hydrolysis of ATP and ADP at 7 days pi, and in hydrolysis of ADP and AMP at 12 days pi in synaptosomes of cerebral cortex in the infected animals; and increased ectonucleotidases activity in synaptosomes of hippocampus in the infected group compared to the control group, except for the hydrolysis of AMP at 12 days pi. The reduced hydrolysis of ATP in cerebral cortex can cause accumulation of the nucleotide into the extracellular milieu; it is known that the excess of ATP can be cytotoxic. Furthermore, with the decrease in enzyme activity less adenosine is produced, this nucleoside is an anticonvulsant and neuroprotective molecule. The increased enzyme activity in the hippocampus can occur to produce adenosine and confer neuroprotection; however at 12 days pi, it is not observed an increase in the hydrolysis of AMP to adenosine. In article II, the evaluation of oxidative parameters and antioxidants revealed that the levels of TBARS and ROS were higher in the infected group compared to controls, mainly in the cortex at 7 and 12 days pi, but also in the cerebellum at 7 days pi and hippocampus and striatum at 12 days pi. Moreover, GSH levels were decreased in the striatum and cerebellum of animals infected at 7 days pi. The oxidative stress process may occur in response to viral infection, but this process has a low specificity and eventually results in oxidative damage to the cell. Thereby, was possible to observe that there is a participation of the purinergic system and the oxidative stress process in the pathogenesis of BoHV-5, suggesting that these mechanisms may be involved in immunomodulation, dysfunction and neuronal death, and may contribute to the process of herpetic meningoencephalitis
Resumo
Bovine herpesvirus type 5 (BoHV-5) is a major cause of viral meningoencephalitis in cattle. The expression of different viral proteins has been associated with BoHV-5 neuropathogenesis. Among these, gI, gE and US9 have been considered essential for the production of neurological disease in infected animals. To evaluate the role of gI, gE and US9 in neurovirulence, a recombinant from which the respective genes were deleted (BoHV-5 gI-/gE-/US9-) was constructed and inoculated in rabbits of two age groups (four and eight weeks-old). When the recombinant virus was inoculated through the paranasal sinuses of four weeks-old rabbits, neurological disease was observed and death was the outcome in 4 out of 13 (30.7 percent) animals, whereas clinical signs and death were observed in 11/13 (84.6 percent) of rabbits infected with the parental virus. In eight weeks-old rabbits, the BoHV-5 gI-/gE-/US9- did not induce clinically apparent disease and could not be reactivated after dexamethasone administration, whereas wild type BoHV-5 caused disease in 55.5 percent of the animals and was reactivated. These findings reveal that the simultaneous deletion of gI, gE and US9 genes did reduce but did not completely abolish the neurovirulence of BoHV-5 in rabbits, indicating that other viral genes may also play a role in the induction of neurological disease.(AU)
O herpesvírus bovino tipo 5 é uma das principais causas de meningoencefalite viral em bovinos. A expressão de diferentes proteínas virais tem sido associada à neuropatogenia do BoHV-5. Entre estas, a gI, gE e US9 têm sido consideradas essenciais para a indução de sinais neurológicos nos animais infectados. Para avaliar o papel das proteínas gI, gE e US9 na neurovirulência, construiu-se um recombinante no qual os genes que codificam estas proteínas foram deletados, denominado BoHV-5 gI-/gE-/US9-. Este vírus foi inoculado em coelhos de idades diferentes (quatro e oito semanas de idade). Quando o vírus recombinante foi inoculado nos seios paranasais de coelhos de quatro semanas de idade, doença neurológica e morte foram observadas em 4 dos 13 (30,7 por cento) animais, enquanto que sinais clínicos e morte foram observados em 11/13 (84,6 por cento) dos coelhos infectados com o vírus parental. Em coelhos de oito semanas de idade, o BoHV-5 gI-/gE-/US9- não induziu sinais clínicos aparentes e, após tentativa de reativação viral por tratamento com dexametasona, o vírus não foi re-excretado. Por outro lado, o vírus selvagem causou doença clínica em 55,5 por cento dos coelhos e foi re-excretado após tratamento com dexametasona. Estes achados revelam que a deleção simultânea dos genes gI, gE e US9 reduziu mas não aboliu completamente a neurovirulência do BoHV-5 em coelhos, indicando que outros genes virais possam ter papel na indução da doença neurológica.(AU)
Assuntos
Animais , Coelhos , Herpesvirus Bovino 5/genética , Infecções por Herpesviridae/veterinária , Meningoencefalite/veterinária , Regulação Viral da Expressão Gênica/genética , Proteínas Recombinantes/genética , Modelos Animais , Herpesvirus Bovino 5/crescimento & desenvolvimento , Etiquetas de Sequências Expressas/químicaResumo
Vários aspectos da biologia do herpesvírus bovino tipo 5 (BoHV-5) têm sido estudados em coelhos, que desenvolvem infecção aguda e doença neurológica após inoculação experimental. A infecção aguda é seguida pelo estabelecimento de infecção latente, que pode ser reativada natural ou artificialmente. Os primeiros experimentos nesta espécie estabeleceram um protocolo de inoculação e monitoramento da infecção, e caracterizaram os principais aspectos virológicos, clínicos e patológicos da infecção aguda. A patogenia da infecção aguda, desde a replicação viral nos sítios de inoculação, vias e cinética de transporte viral até o encéfalo, distribuição e replicação viral no sistema nervoso central (SNC), tropismo celular e tecidual, manifestações clínicas e patologia no SNC foram detalhadamente estudados nestes animais. Posteriormente, vários aspectos biológicos e moleculares da infecção latente também foram elucidados a partir de inoculações de coelhos. Os coelhos também têm sido utilizados para estudar o fenótipo (neuroinvasividade, neurovirulência) de isolados de campo e de cepas vacinais recombinantes, proteção por imunidade passiva, proteção vacinal, eficácia de drogas anti-virais e terapêuticas de suporte da infecção neurológica. Este modelo experimental também foi utilizado para o estudo da origem e distribuição dos estímulos elétricos produzidos durante as convulsões - uma característica da infecção neurológica pelo BoHV-5 -, e para testes de medicamentos anti-convulsivantes. Ressalvadas as diferenças que certamente existem entre bovinos - os hospedeiros naturais - e coelhos, as observações oriundas deste modelo experimental tem contribuído sobremaneira para o conhecimento da biologia do BoHV-5. O presente trabalho apresenta uma coletânea de resultados e observações, publicadas ou não pelo grupo, ao longo de mais de uma década, envolvendo inoculações de coelhos para estudar diversos aspectos da infecção pelo BoHV-5.(AU)
Several aspects of the biology of bovine herpesvirus 5 (BoHV-5) have been studied in rabbits, which develop acute infection and neurological disease upon experimental inoculation. The acute infection is followed by the establishment of latent infection, which can be naturally or artificially reactivated. The first experiments in rabbits established a protocol for virus inoculation and monitoring the infection, and characterized the main virological, clinical and pathological aspects of the acute infection. The pathogenesis of acute infection, from the initial viral replication at site of inoculation, pathways and kinetics of viral transport to the brain, distribution and virus replication in the central nervous system (CNS), cellular and tissue tropism, clinical signs and CNS pathology have been extensively studied using this animal model. Subsequently, several biological and molecular aspects of latent BoHV-5 infection have also been elucidated upon inoculation of rabbits. Rabbits have also been used to investigate the phenotype (neuroinvasiveness, neurogrowth) of field isolates and recombinant vaccine candidates, protection by passive immunity, vaccine protection, the efficacy of anti-viral drugs and support therapies for neurological disease. This animal model was also used to investigate the origin and distribution of electric impulses involved in seizures - a hallmark of BoHV-5 induced neurological infection - and also to test the efficacy of anti-convulsivants. In spite of the possible differences between rabbits and cattle - the natural host of the virus - the observations taken from this experimental model have greatly contributed to the knowledge of the biology of BoHV-5 infection. The present article presents a review of the main published and unpublished results and observations by our group, comprising more than a decade of studies on the pathogenesis of BoHV-5 infection in the rabbit model.(AU)