Japanese encephalitis virus: Associated immune response and recent progress in vaccine development
Introduction
Japanese encephalitis virus (JEV) is a member of genus Flavivirus that belongs to family Flaviviridae. It is an enveloped virus consisting of single stranded positive sense RNA genome surrounded by a nucleocapsid. The virus is transmitted to humans by mosquito vector, principally by Culex tritaeniorhynchus that proliferates in close proximity with other vertebrate hosts like pigs, water birds and chicks. JEV is an etiological agent of acute zoonotic infection, commonly affecting children and is the major cause of epidemic Japanese encephalitis (JE) in Asia [1]. Since 1870s, JE has emerged as the most important form of viral encephalitis throughout South-east Asia, China, and in Asia-pacific belt [2]. It is a serious central nervous system (CNS) disease with high rate of mortality and morbidity, and has a potential to spread to far unaffected areas like Australian continent [3]. JE has been attributed to an estimated worldwide incidence of 50,000 to 70,000 cases with a high case fatality rate of 30–50%. A large proportion of survivors suffer from long-term neurological manifestations in the form of convulsions, tremors, paralysis, ataxia and other such symptoms [2]. Different disease manifestations however occur in the host as a result of JEV infection, ranging from mild subclinical febrile illness where infection is limited to the extraneural tissues to the clinical infections leading to encephalitis.
JEV is transmitted to humans from the bite of carrier mosquito and it replicates in local tissues and regional lymph nodes from where it is carried via lymphatics to the thoracic duct and then into the bloodstream. From blood, the virus penetrates into the CNS and causes encephalitis. A rapid immune response is generated in the organs where JEV replicates with mononuclear and polymorphonuclear cell infiltrations [4]. However, the mechanism involved in producing this inflammatory state is not clearly understood [5] and an elucidation of immune responses to JEV infection has been an active area of research during the past few decades in both humans and experimental animals [[6], [7], [8], [9], [10]]. At present, both humoral [11] and cell-mediated immunity (CMI) have been implicated in providing protection against JEV infection, where early host defense against JEV infection is mediated by phagocytic cells [12], and B and T effector cells [6]. However, the role of immune responses in recovery from JEV infection is poorly understood.
The humoral response is indispensable for the control and dissemination of JEV infection. Much of this control is provided by the neutralizing antibodies, which recognize epitopes located predominately in the viral envelope (E) glycoprotein. These antibodies inhibit viral attachment, internalization, and/or replication within the cells. Apart from anti-E antibodies, anti-non-structural-1 (NS-1) and precursor membrane protein (PrM) antibodies are also important in mediating protective immunity to JEV infection [[13], [14], [15]]. A failure to mount adequate humoral immune response by the host can increase the susceptibility to encephalitis caused by JEV. In such a condition, cellular immunity can play an important protective role against JEV infection; however, it is less well characterized. CD4+ and CD8+ T cells are reported to be important in controlling experimental murine flaviviral infections [6,8,[16], [17], [18]]. Pan et al. (2001) have however reported that CD8+ cytotoxic T cell activity is not required for protection and presence of CD4+ T helper (Th) cells assist in the induction of optimal antibody responses by DNA or live JEV vaccines. CD4+ Th cells exert most of their functions through secreted cytokines [19]. Changes in the pattern of cytokines produced by Th cells can change the type of immune responses that develop among other leukocytes. The Th1 response generates a cytokine profile that supports inflammation and activation of certain T cells and macrophages, whereas Th2 response activates B cells and immune responses that depend on antibodies. The Th1 cells produce proinflammatory cytokines such as IFN-γ, IL-2 and IL-12, while the Th2 cells produce antiinflammatory cytokines that are involved in the regulation of the B cell response such as IL-4, IL-5, IL -6, and IL-10. However, there is dearth of information in JEV infection regarding the type of Th responses (Th1/Th2) generated as well as the role of cytokines involved. In the study where cytokines and IgG subtypes responses have been studied, the Th responses were found to be mice strain and route specific. These studies involved three stains of mice (C57BL/6J, Swiss Albino and BALB/c), which were immunized with either live JEV (Vellore strain P20778) or killed JEV vaccine by intraperitoneal, subcutaneous and peroral routes. The live JEV was found to induce Th1, while killed vaccine induced a predominant Th2 profile [9].
After the viral exposure, T cells activation leads to the expression of large number of cytokines that facilitate in generation of both cell-mediated and antibody responses against foreign antigen. The early innate and cellular immune responses with special reference to cytokines during JEV infection has been less studied, though there is increasing evidence for the critical role of cytokines during JE. Proinflammatory cytokines like IFN-γ, TNF-α, macrophage migration inhibitory factor and chemokine IL-8, have been found to be associated with bad outcome in small studies including mice brain [[20], [21], [22], [23]]. It has also been shown in a large patient based study that elevated levels of proinflammatory cytokines like IL-6 and chemokines like RANTES (regulated upon activation, T cell expressed and secreted) are associated with a poor outcome, but whether these enhanced the levels of proinflammatory cytokines contribute to the immunopathogenesis or are simply correlated to severe illness is still ambiguous [24]. An in vitro study has recently demonstrated that after JEV infection there is microglial activation followed by subsequent release of various proinflammatory mediators, which induces neuronal death. It has also been reported that although initiation of immune responses by microglial cells is an important protective mechanism in the CNS, unrestrained inflammatory responses may result in irreparable brain damage [25].
The mechanisms by which many neurotropic viruses cause the neurological diseases, precise mechanism of inflammation and immune responses are not fully understood. Infiltration of inflammatory cells, perivascular cuffing, gliosis and necrosis are the hallmarks of brain pathology during JE [22,26,27]. In many cases, virus mediates damage to brain tissues indirectly by triggering cell-mediated immune responses like activation of cytotoxic T cells and macrophages, instead of playing a direct role. Activated inflammatory cells secrete various cytokines, such as IL-1, TNF-α, thereby causing toxic effect in the brain [28]. Although, these immune/inflammatory infiltrates are aimed at restricting the viral growth in the brain, their uncontrolled production is detrimental to the host. Most cytokine studies have been focused on the importance of innate immune response in brain against this neurotropic virus in mice model [22,25]. Since the immune responses in the CNS differs from that in the periphery, the importance of immune response with special reference to cytokines cannot be overlooked in the later system. Here in this review, we discuss JEV-associated immune responses and the contribution of cytokines in the immunopathogenesis of JEV during the course of infection. An understanding of immunopathogenesis associated with JEV can aid in improving the strategies for development of future vaccines against JEV.
Section snippets
Japanese encephalitis virus
JEV was first isolated in 1924 from a clinical case during the first reported epidemic of Japan [29]. In 1935, the prototype Nakayama strain was isolated from the brain of a patient suffering from encephalitis. Thereafter the virus was classified with other flavivirus as group B arbovirus in family Togaviridae, but in 1985 it has been designated under a separate family Flaviviridae as a member of genus Flavivirus [30]. The genus Flavivirus has been named after the prototype yellow fever virus
Immune response
In the early phase of the disease, when the virus replicates and spreads by haematogenous route to other parts of the body, the immune responses are likely to determine the outcome of the host-pathogen interactions [56]. A differential clinical presentation of disease symptoms is observed in JEV-infected individuals where only a small proportion of infected individuals develop clinical features, and these may range from a non-specific flu-like illness to a severe fatal meningoencephalitis,
Vaccination strategies for JEV
There are majorly three types of JE vaccines, which are currently in use: mouse-brain derived inactivated, cell-culture derived inactivated and cell-culture derived live attenuated JE vaccine. The formalin-inactivated vaccines are reported to be safe and effective against JEV for at least 30 years [33]. However, the mouse-brain inactivated vaccines were the most widely produced and internationally distributed vaccines, which are no longer in use owing to concerns regarding adverse effects of
Conclusions
The fundamental basis of vaccination is the generation and maintenance of an antigen-specific immune response, which is sufficient to mediate protection from infection and trigger a long-lived humoral response through the sufficient production of antibodies. In order to develop such an effective vaccine against JEV, it is important to understand the mechanisms of the immune response to JEV infection, and we have critically dealt with it in this review.
A variety of approaches have been tested
Conflicts of interest
The authors have no competing interests to declare.
Acknowledgements
The author KKN is the recipient of Ramalingaswami re-entry fellowship and acknowledges the Department of Biotechnology, New Delhi, India. The authors are thankful to the All India Institute of Medical Science, Jodhpur, India and the National Institute of Technology Raipur, India for providing the facility and space.
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