Elsevier

Vaccine

Volume 22, Issues 11–12, 29 March 2004, Pages 1433-1440
Vaccine

Vaccination with inactivated murine gammaherpesvirus 68 strongly limits viral replication and latency and protects type I IFN receptor knockout mice from a lethal infection

https://doi.org/10.1016/j.vaccine.2003.10.015Get rights and content

Abstract

Human gammaherpesviruses such as Epstein-Barr virus (EBV) cause lifelong infections and associated diseases, including malignancies, and the development of an effective vaccine against this class of viral infections is of considerable interest. The murine herpesvirus 68 (MHV-68) model provides a useful experimental setting to investigate the immune response to gammaherpesvirus infections and to evaluate the efficacy of vaccination strategies. In this study, we tested a heat-inactivated MHV-68 vaccine in immunocompetent mice as well as in B cell-deficient or type I IFN receptor knockout mice. Vaccination with heat-inactivated MHV-68 protected immunocompetent mice from the acute MHV-68 infection in the lung and strongly reduced the expansion of latently infected cells in the spleen and the development of splenomegaly. A similar inhibition of the acute viral replication in the lung was also observed in vaccinated B cell-deficient mice. Of note, the inactivated MHV-68 vaccine completely protected type I IFN receptor knockout mice from the infection with a lethal dose of MHV-68.

Introduction

Gammaherpesviruses cause widespread infections in their natural host and are linked to a wide variety of diseases. The two gammaherpesviruses infecting humans, Epstein-Barr virus (EBV) and Human Herpes virus 8 (HHV8), are generally well controlled through life by immune mechanisms. After the primary infection, during which EBV causes infectious mononucleosis (IM), both viruses establish latency in lymphoid compartments. Under conditions of massive immunosuppression, often in association with AIDS or organ transplantation, latently infected cells can undergo uncontrolled proliferation, leading to the development of malignancies. EBV is associated with undifferentiated nasopharyngeal carcinoma (NPC), endemic Burkitt’s lymphoma (BL), post-transplant lymphoproliferative disease (PTLPD) and Hodgkin’s lymphoma [1]. HHV8 is associated with Kaposi’s sarcoma, several forms of primary effusion lymphoma and multicentric Castleman’s disease [2], [3].

The development of an effective vaccine against EBV has been a goal for many years [4], [5], [6]. A vaccine capable of either blocking EBV primary infection or significantly reducing virus load during this stage might prevent significant acute illness typical of IM, and this would be the principal aim of an EBV vaccine in western societies. On the other hand, EBV-associated malignancies arise in patients years after their primary infection, and protection from these long-term consequences of EBV infection would require a vaccine that ideally confers sterile immunity and prevents the establishment of the carrier state. Furthermore, a reduced virus replication during primary infection might result in a smaller reservoir of latent virus in the individual, thus limiting the risk of harmful virus reactivation and development of lymphoproliferative diseases in immunosuppressed subjects.

Progress on testing vaccine strategies against gammaherpesviruses suffered for many years from the lack of a relevant small animal model system. In recent years, murine herpesvirus 68 (MHV-68) mouse model has been established, and provides a practical route for dissecting gammaherpesvirus infection in vivo. MHV-68 was isolated from free-living small rodents and is able to infect inbred strains of mice. It shares genetic and pathogenic features with human gammaherpesviruses [7], [8]. When administered intranasally (i.n.) to mice, MHV-68 establishes a productive infection in lung alveolar epithelial cells that lasts for 7–10 days. Following the primary infection, MHV-68 establishes a latent infection in B lymphocytes and also persists in macrophages and lung epithelial cells [8], [9], [10], [11]. MHV-68 infection also causes a notable splenomegaly and the expansion of CD8+ T lymphocytes [12]. Both of these features are characteristic of IM in EBV infected individuals [1]. A variety of studies have demonstrated that all components of the adaptive immune response are involved in the control of the various phases of MHV-68 infection, similar to what happens during EBV infection. CD8+ T cells play a major role in controlling MHV-68 replication in the lung, during the acute and persistent infection [13], [14], whereas the antibody response appears to play a major role in anti-viral surveillance in the lung at late times post-infection. Although not required for the initial control of the lytic infection, CD4+ T cells may play a direct anti-viral role through IFN-γ production [15], [16], [17], [18], and are also implicated in the maintenance of a virus-specific antibody response [19]. Immune control of latency is also largely dependent on CD8+ T cells, but CD4+ T cells appear to be important for maintaining an effective CD8+ T cell response during this stage of the infection [13].

The ability to readily characterise MHV-68 pathogenic and immunologic features renders this a powerful model to test the efficacy of vaccination strategies against gammaherpesviruses [20], [21], [22], [23], [24], [25]. In the present study, we evaluated the efficacy of a heat-inactivated MHV-68 vaccine in normal and immunocompromised mice. The use of heat-inactivated virus represents an advantageous strategy for at least two reasons. Firstly, vaccines based on whole inactivated virus contain a mixture of virtually all viral proteins, and may prime the immune system against a wider spectrum of immunologically relevant viral antigens with respect to the use of recombinant or purified viral proteins or subunits. Secondly, the use of inactivated viral particles does not imply the risk of infection associated with live-attenuated virus vaccines.

Our results show that vaccination with heat-inactivated MHV-68 is highly effective in protecting immunocompetent mice against lytic MHV-68 infection as well as against the amplification of viral latency and the development of splenomegaly. The inactivated MHV-68 vaccine protected against viral replication in the lung of normal and B cell-deficient mice and, importantly, protected type I IFN receptor knockout mice from the lethal outcome of MHV-68 infection observed in unvaccinated counterparts.

Section snippets

Mice

C57BL/6 mice (H-2b) were purchased from Banton and Kingman Universal Limited (Grimston, Aldborough, Hull, UK). Inbred μMT on a C57BL/6 background [26] have a targeted lesion in the μ immunoglobulin chain resulting in the failure to express IgM; consequently, B cell development is arrested at the pre-B cell stage. A colony was established at the Department of Veterinary Pathology, University of Edinburgh. Inbred wild-type 129Sv and IFNARI−/− 129Sv mice [27] were obtained from B&K Universal

Protective effect of inactivated MHV-68 vaccination in C57BL/ 6 mice

To assess the effect of inactivated virus vaccination on the acute phase of MHV-68 infection, C57BL/6 mice were given one or two subdermal injections of 107 PFU of heat-inactivated virus. Four weeks after the single or second vaccination, the mice were challenged intranasally with 4×105 PFU of MHV-68. Lungs were harvested from these mice at 3 and 6 days post-infection and viral titres in these organs determined by plaque assays. As shown in Fig. 1, vaccination with heat-inactivated MHV-68 was

Discussion

The MHV-68 infection of inbred mice represents an animal model well suited for the study of gammaherpesviruses biology, and has been used over the past few years to investigate the requirements for the effective control of this class of viruses and to evaluate the efficacy of vaccination strategies [20], [21], [22], [23], [24], [25]. The recent knowledge of the complex progression of MHV-68 infection and the corresponding virus-specific host immune response has led to the design of vaccines

Acknowledgements

This work was supported by the European Community (contract B104-CT98-0466).

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