Elsevier

Antiviral Research

Volume 108, August 2014, Pages 1-9
Antiviral Research

Protection against Chikungunya virus induced arthralgia following prophylactic treatment with adenovirus vectored interferon (mDEF201)

https://doi.org/10.1016/j.antiviral.2014.05.004Get rights and content

Highlights

  • The DBA/1J mouse model mimics several human disease signs of Chikungunya virus infection.

  • Infection results in footpad swelling, cytokine elevation, and arthritis.

  • Prophylactic treatment with mDEF201 results in reduced virus titer, footpad swelling and cytokine levels.

Abstract

Recent outbreaks of Chikungunya virus (CHIKV) infection have resulted in millions of cases of disease with significant morbidity. No approved antiviral treatments exist for the prevention or treatment of this viral disease. Infection with CHIKV results in a high rate of symptomatic disease that primarily includes a debilitating arthralgia. To model this cardinal disease manifestation, adult DBA/1J mice were challenged with CHIKV by footpad injection. Viremia and hind limb virus titers increased ∼100-fold while spleen virus increased >1000-fold within 1 day post-virus infection (dpi). Footpad swelling was measured over a 10-day period, with peak swelling observed between 6 and 7 dpi. Histology of the hind leg at the site of virus challenge showed evidence of myositis and synovitis starting on 5 dpi. Cytokine profiling of the hind limb at the site of inoculation revealed a biphasic inflammatory response represented by an increase in IL-6, MCP-1, IFN-γ, MIP-1α, RANTES, and IL-17. To investigate the prophylactic capacity of IFN, mice were treated with mDEF201, an adenovirus-vectored IFN-α. Intranasal administration of a single 107 pfu/ml dose of mDEF201 administered 21 days to 24 h prior to infection, significantly reduced footpad swelling, virus titers in the hind leg and spleen, and several inflammatory cytokines. Efficacy was not observed when treatment was initiated 24 h after virus challenge. This arthralgia model of CHIKV recapitulates relevant disease features commonly observed in human disease making it applicable to preclinical testing of therapies that target both viral replication and the associated joint disease.

Introduction

Chikungunya virus (CHIKV) is associated with significant morbidity and mortality worldwide. Historically, the primary vector for spread of CHIKV was Aedes aegypti, although recent adaptive mutations within the virus have resulted in a more efficient infection of Aedes albopictus (Tsetsarkin et al., 2006). This has resulted in outbreaks outside the historical range, making this emerging virus a major public health concern.

The rapid emergence of CHIKV and the millions of cases of disease underscore the importance of the development of countermeasures to prevent or treat the symptoms of CHIKV. Fever and arthralgia are common symptoms of CHIKV infection, although a small number (5–18%) of infected people, especially those below age 25, are asymptomatic (Dupuis-Maguiraga et al., 2012). Mortality as a result of CHIKV infection is rare and is generally associated with underlying health issues (Thiberville et al., 2013). Rheumatic manifestations typically affect the extremities, primarily including ankles, wrists and phalanges (Kennedy et al., 1980). A study conducted after the La Reunion Island outbreak identified persistent arthritis in over half of the participants 15 months after acute infection, with 43% of those with persistent arthritis significantly impaired in carrying out daily or household activities (Sissoko et al., 2009). As the rheumatic disease is debilitating and very common in persons infected with CHIKV, it would be important to prevent or treat this aspect of the disease.

An animal model that replicates human arthralgia is essential for the development of countermeasures for the prevention and treatment of disease associated with CHIKV infection. Macaques have been used to study CHIKV, and this model mimics many acute symptoms seen in humans such as fever, rash, and high viral titers but displays inconsistent swelling of the joints and lacks the joint and muscle pathology typically observed after infection with CHIKV (Higgs and Ziegler, 2010).

Various mouse models have been developed which model different aspects of the disease. The C57BL/6 neonatal mouse model includes relevant disease manifestations such as joint swelling. There is also consistency between disease seen in children less than one year of age and this model, with virus being found in similar organs and a strong age-dependent correlation with severity of disease (Couderc et al., 2008). Interferon receptor knockout mice were highly susceptible to infection with CHIKV and rapidly succumbed to infection (Couderc et al., 2008), which serves to model severe infection and mortality that is occasionally seen. Infection of these immunocompromised mice results in dissemination of the virus to all tissue types, including the central nervous system (Partidos et al., 2011). This pathology may have relevance in light of recently reported neurologic complications of encephalopathy in newborns and meningitis and encephalitis in older children and adults (Arpino et al., 2009). Inoculation of the footpad of 14-day old C57BL/6 mice resulted in swelling, arthritis, tenosynovitis and myositis (Morrison et al., 2011), but a neonatal model has limitations, especially in regard to testing vaccines. Others have developed a model of footpad swelling and joint involvement in adult C57BL/6 by injecting virus into the footpad (Gardner et al., 2010).

A replication-deficient adenovirus type 5 vector containing a gene for mouse IFN-α, mDEF201, is administered intranasally where it enters nasal epithelial cells and expresses the transgene (Julander et al., 2011). This results in the production of systemic levels of IFN several hours after treatment (Wu et al., 2007). With non-lethal infections such as CHIKV, type I interferon (IFN) plays a role in coordinating early antiviral immune response (Havenar-Daughton et al., 2006), and plays a central role in controlling CHIKV infection, acting indirectly as an antiviral through the induction of IFN-stimulated genes that inhibit viral replication (Schilte et al., 2010). The nonstructural proteins (nsPs) of CHIKV antagonize the IFN response as infection progresses, so the timing of IFN intervention is critical (Fros et al., 2010).

This manuscript details the establishment of an adult mouse model of CHIKV arthritis and swelling. Footpad swelling, virus replication, and cytokine profiles are used as parameters for the characterization of the antiviral effect of mDEF201 against CHIKV. These results demonstrate the utility of this model for use in antiviral studies.

Section snippets

Cells

C6/36 cells were obtained from ATCC (Manassas, VA) and were used for virus propagation. These cells were grown in RPMI at 28 °C with 5% fetal bovine serum (FBS). Vero 76 cells, maintained in minimal essential medium (MEM) supplemented with 10% FBS, were also obtained from ATCC and were used in viral titer assays. Media was obtained from Hyclone Laboratories, Logan, UT.

Virus

Chikungunya virus strain S27 (VR-64) was obtained from ATCC. Stock virus was prepared by harvesting supernatant from C6/36 cells 2

Characterization of CHIKV infection of DBA/1 mice

Several mouse strains, including C57BL/6, AG129, CAST/EiJ and DBA/1J, were inoculated with CHIKV and disease signs, including footpad swelling, differences in behavior, weight change and mortality, were observed. The DBA/1J strain was selected based on the development of robust footpad swelling, availability and use in arthritis research (Chiba et al., 2012, Eros et al., 2009, Nishida et al., 2002). To characterize CHIKV replication and disease in this mouse strain, DBA/1J mice were inoculated

Discussion

Infection of DBA/1J mice with CHIKV mirrors several disease signs observed in humans, including arthralgia, high acute viral titers and increased cytokine levels (Couderc and Lecuit, 2009). This non-lethal model focuses primarily on the arthritic disease manifestations that occur most commonly in infected individuals. The primary clinical and economic burden of human CHIKV disease is arthralgia, which renders infected individuals unable to work and perform other necessary activities, which in a

Acknowledgements

We thank Isaac Wong, Joseph Hagloch, Brett Hurst, Makda Gebre, and Shelby Wilcox for expert technical assistance. This work was supported by funding from the Virology Branch, DMID, NIAID, NIH (HHSN272201000039I/HHSN27200004/A21).

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