Complement Receptor 3 Promotes Severe Ross River Virus-Induced Disease

Arthritis/myositis-associated alphaviruses, such as Ross River virus (RRV), chikungunya virus, o'nyong-nyong virus, mayaro virus, and others, are mosquito-transmitted viruses that cause debilitating inflammatory disease in humans (18, 37, 46). In addition to causing endemic disease, this group of viruses is capable of causing explosive epidemics that can involve millions of infected individuals. Past epidemics include a 1979-to-1980 epidemic of RRV disease in the South Pacific that involved more than 60,000 patients (20) and a 1959-to-1962 epidemic of o'nyong-nyong fever in Africa that involved at least 2 million patients (57). More recently, a reemergence of chikungunya virus has resulted in an unprecedented epidemic in multiple countries, including Indian Ocean islands such as the Comoros, Reunion, Mayotte, Seychelles, and Mauritius as well as India, Sri Lanka, and Indonesia (36, 37). The number of infected individuals involved thus far in this ongoing epidemic is in the millions, with an estimated 1.4 to 6.5 million cases in India alone (29). In addition, from July to September 2007, an outbreak of chikungunya virus occurred in northeastern Italy, resulting in 205 confirmed human cases and detection of the virus within the local mosquito population (4, 40). Clinical signs and symptoms following infection with arthritis/myositis-associated alphaviruses include fever, rash, muscle pain, and severe joint pain, with the peripheral joints, such as the ankles, knees, wrists, and joints of the hands, most commonly affected (19, 20, 37). Treatment is palliative and based on analgesics and anti-inflammatory drugs. The resolution of disease symptoms generally occurs over several weeks; however, many patients complain of chronic pain lasting 6 months or longer, suggesting that these viruses may be responsible for long-term health effects (20, 39). The disease caused by this group of viruses is believed to be initiated by viral replication and the induction of host inflammatory responses within the affected tissues. This is based on the detection of RRV RNA and antigen in joint tissues of affected patients as well as the presence of large number of inflammatory cellular infiltrates, consisting primarily of monocytes and macrophages, within synovial effusions (16, 21, 45). Additionally, chikungunya virus antigen, severe myositis with inflammatory macrophages and T lymphocytes, and myofibril pathology have been detected in quadriceps muscle biopsy samples collected from chikungunya virus-infected patients (34). The complement system, which consists of more than 30 soluble and cell surface proteins, is a major component of innate immunity that functions to recognize and eliminate invading pathogens (9). Activation of the complement system occurs through multiple mechanisms that include three well-described pathways: the classical, lectin, and alternative complement activation pathways. Activation of the complement system results in a cascade of enzymatic proteolysis of various components of the complement system, including complement component 3 (C3), C4, and C5. The proteolytic processing of C3 generates an array of cleavage products that are involved in the amplification of complement activity through the formation of C3 and C5 convertases, the opsonization of pathogens, and the attraction and activation of leukocytes of both the innate and adaptive arms of the immune response. Regulation of trafficking and effector functions of leukocytes via C3-, C4-, and C5-derived cleavage products occurs through interactions with a number of cell surface receptors. Complement receptor 3 (CR3) (CD11b/CD18, Mac-1, αmβ2) is a member of the β2 integrins, which are heterodimeric cell surface proteins composed of a common β chain (CD18) and one of four α chains (14, 41). The β2 integrins regulate the accumulation and activation of neutrophils, macrophages, and natural killer cells at sites of tissue inflammation and injury. CR3 binds a number of different ligands, including iC3b, ICAM-1 and ICAM-2, and fibrinogen, within the I-domain of the CD11b subunit (3, 11, 44, 58, 59). Additionally, CR3 contains a lectin domain within the C-terminal region of CD11b that interacts with polysaccharides (42, 50, 60). Recently, our group developed an experimental mouse model to investigate the immunopathogenesis of arthritis/myositis-associated alphaviruses based on RRV infection of C57BL/6 mice (32). Following the inoculation of 3- to 4-week-old mice, RRV rapidly spreads to joint and skeletal muscle tissue via a high-titer serum viremia (32). Viral titers peak in tissues at 24 to 48 h postinfection and decline thereafter. At later times postinfection, RRV-infected mice develop severe inflammation within joint and skeletal muscle tissues that correlates with observable disease signs such as a loss of gripping ability, altered gait, and hind-limb weakness (28, 32). Utilizing this model, we demonstrated that C3 activation products, such as the CR3 ligand iC3b, were detected at the sites of RRV-induced inflammation and that C3 was critical for the tissue destruction phase of RRV-induced inflammatory disease (31). In addition, work from that study indicated that C3 was not required for recruitment of inflammatory leukocytes to the sites of RRV-induced inflammation, suggesting that complement activation may regulate the activation phenotypes of inflammatory leukocytes at the sites of RRV-induced inflammation. In this study, we assessed the role of CR3 in the pathogenesis of RRV infection. We found that mice deficient in CR3 (CD11b−/−) developed inflammation of joint and skeletal muscle tissue following RRV infection. However, CD11b−/− mice exhibited less-severe disease signs and tissue damage compared to wild-type (WT) mice, suggesting that like C3, CR3 contributes to the tissue destruction phase of the inflammatory disease. These findings are consistent with a model in which CR3-dependent regulation of inflammatory cell function at the sites of inflammation promotes alphavirus-induced disease.