Tissue-Specific Deletion of the Coxsackievirus and Adenovirus Receptor Protects Mice from Virus-Induced Pancreatitis and Myocarditis

Acute coxsackievirus B3 (CVB3) infection is one of the most prevalent causes of acute myocarditis, a disease that frequently is identified only after the sudden death of apparently healthy individuals. CVB3 infects cardiomyocytes, but the infection is highly focal, even in the absence of a strong adaptive immune response, suggesting that virus spread within the heart may be tightly constrained by the innate immune system. Type I interferons (T1IFNs) are an obvious candidate, and T1IFN receptor (T1IFNR) knockout mice are highly susceptible to CVB3 infection, succumbing within a few days of challenge. Here, we investigated the role of T1IFNs in the heart using a mouse model in which the T1IFNR gene can be ablated in vivo, specifically in cardiomyocytes. We found that T1IFN signaling into cardiomyocytes contributed substantially to the suppression of viral replication and infectious virus yield in the heart; in the absence of such signaling, virus titers were markedly elevated by day 3 postinfection (p.i.) and remained high at day 12 p.i., a time point at which virus was absent from genetically intact littermates, suggesting that the T1IFN-unresponsive cardiomyocytes may act as a safe haven for the virus. Nevertheless, in these mice the myocardial infection remained highly focal, despite the cardiomyocytes' inability to respond to T1IFN, indicating that other factors, as yet unidentified, are sufficient to prevent the more widespread dissemination of the infection throughout the heart. The absence of T1IFN signaling into cardiomyocytes also was accompanied by a profound acceleration and exacerbation of myocarditis and by a significant increase in mortality. IMPORTANCEAcute coxsackievirus B3 (CVB3) infection is one of the most common causes of acute myocarditis, a serious and sometimes fatal disease. To optimize treatment, it is vital that we identify the immune factors that limit virus spread in the heart and other organs. Type I interferons play a key role in controlling many virus infections, but it has been suggested that they may not directly impact CVB3 infection within the heart. Here, using a novel line of transgenic mice, we show that these cytokines signal directly into cardiomyocytes, limiting viral replication, myocarditis, and death. Myocarditis is a potentially serious and sometimes fatal disease, with several infectious and noninfectious causes. In developed countries, it most commonly results from a virus infection, often by type B coxsackieviruses (CVB), and in particular by serotype 3 (CVB3) (1-8). It is found at routine necropsy at a prevalence that greatly exceeds its rate of clinical diagnosis (9, 10), a discrepancy that may be explained by the fact that only a subset of individuals with acute myocarditis develop marked symptoms such as chest pains, palpitations, or signs of heart failure. Consistent with this, acute myocarditis was identified as the cause of death in 39 of 2,560 serial autopsies (ϳ1.5%), but it had been clinically suspected in only one of the cases (11); thus, in this s...

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confidence: 99%

In VivoAblation of Type I Interferon Receptor from Cardiomyocytes Delays Coxsackieviral Clearance and Accelerates Myocardial Disease

Althof

Harkins

Kemball

et al. 2014

“…They are one of the most common causes of myocarditis, with up to 33% of patients positive for coxsackievirus group B (CVB) (2). The classification of coxsackieviruses is based on the systemic versus localized pathology in the mouse, which is the experimental model of choice to study the CVB3 serotype (3,4). In addition to mouse and human, other species are also infected by CVB3, which has been reported in diverse primate species with myocarditis (5-7).…”

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confidence: 99%

Interspecies Differences in Virus Uptake versus Cardiac Function of the Coxsackievirus and Adenovirus Receptor

Freiberg

Sauter

Pinkert

et al. 2014

The coxsackievirus and adenovirus receptor (CAR) is a cell contact protein with an important role in virus uptake. Its extracellular immunoglobulin domains mediate the binding to coxsackievirus and adenovirus as well as homophilic and heterophilic interactions between cells. The cytoplasmic tail links CAR to the cytoskeleton and intracellular signaling cascades. In the heart, CAR is crucial for embryonic development, electrophysiology, and coxsackievirus B infection. Noncardiac functions are less well understood, in part due to the lack of suitable animal models. Here, we generated a transgenic mouse that rescued the otherwise embryonic-lethal CAR knockout (KO) phenotype by expressing chicken CAR exclusively in the heart. Using this rescue model, we addressed interspecies differences in coxsackievirus uptake and noncardiac functions of CAR. Survival of the noncardiac CAR KO (ncKO) mouse indicates an essential role for CAR in the developing heart but not in other tissues. In adult animals, cardiac activity was normal, suggesting that chicken CAR can replace the physiological functions of mouse CAR in the cardiomyocyte. However, chicken CAR did not mediate virus entry in vivo, so that hearts expressing chicken instead of mouse CAR were protected from infection and myocarditis. Comparison of sequence homology and modeling of the D1 domain indicate differences between mammalian and chicken CAR that relate to the sites important for virus binding but not those involved in homodimerization. Thus, CAR-directed anticoxsackievirus therapy with only minor adverse effects in noncardiac tissue could be further improved by selectively targeting the virus-host interaction while maintaining cardiac function. IMPORTANCECoxsackievirus B3 (CVB3) is one of the most common human pathogens causing myocarditis. Its receptor, the coxsackievirus and adenovirus receptor (CAR), not only mediates virus uptake but also relates to cytoskeletal organization and intracellular signaling. Animals without CAR die prenatally with major cardiac malformations. In the adult heart, CAR is important for virus entry and electrical conduction, but its nonmuscle functions are largely unknown. Here, we show that chicken CAR expression exclusively in the heart can rescue the otherwise embryonic-lethal CAR knockout phenotype but does not support CVB3 infection of adult cardiomyocytes. Our findings have implications for the evolution of virus-host versus physiological interactions involving CAR and could help to improve future coxsackievirus-directed therapies inhibiting virus replication while maintaining CAR's cellular functions.

“…injection, readily infect mice, causing myocarditis (11) and pancreatitis (12); infection of the heart and pancreas is mediated by murine CAR (13,14). Although infection can be established by the enteral (oral [p.o.])…”

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confidence: 99%

Expression of Human Decay-Accelerating Factor on Intestinal Epithelium of Transgenic Mice Does Not Facilitate Infection by the Enteral Route

Pan

Zhang

Odenwald

et al. 2015

Self Cite

In vitro, infection of polarized human intestinal epithelial cells by coxsackievirus B3 (CVB3) depends on virus interaction with decay-accelerating factor (DAF), a receptor expressed on the apical cell surface. Although mice are highly susceptible to CVB3 infection when virus is delivered by intraperitoneal injection, infection by the enteral route is very inefficient. Murine DAF, unlike human DAF, does not bind virus, and we hypothesized that the absence of an accessible receptor on the intestinal surface is an important barrier to infection by the oral route. We generated transgenic mice that express human DAF specifically on intestinal epithelium and measured their susceptibility to infection by a DAF-binding CVB3 isolate. Human DAF permitted CVB3 to bind to the intestinal surface ex vivo and to infect polarized monolayers of small-intestinal epithelial cells derived from DAF transgenic mice. However, expression of human DAF did not facilitate infection by the enteral route either in immunocompetent animals or in animals deficient in the interferon alpha/beta receptor. These results indicate that the absence of an apical receptor on intestinal epithelium is not the major barrier to infection of mice by the oral route. IMPORTANCE CVB3 infection of human intestinal epithelial cells depends on DAF at the apical cell surface, and expression of human DAF on murine intestinal epithelial cells permits their infection in vitro. However, expression of human DAF on the intestinal surface of transgenic mice did not facilitate infection by the oral route. Although the role of intestinal DAF in human infection has not been directly examined, these results suggest that DAF is not the critical factor in mice.C oxsackieviruses belong to the genus Enterovirus of the family Picornaviridae, a group of small, nonenveloped viruses with a single-strand, positive-sense RNA genome (1). Group B coxsackieviruses (CVBs) are important causes of viral myocarditis and meningitis, and they have been proposed to be a possible triggering agent for juvenile diabetes (1).CVBs, like other enteroviruses, are transmitted by the fecaloral route and are believed to initiate infection by crossing the intestinal mucosa, which is lined by polarized epithelial cells with distinct apical and basolateral surfaces. In polarized epithelial cells, the primary CVB receptor, the coxsackievirus and adenovirus receptor (CAR) (2-4), is not expressed on the apical cell surface but is instead confined to the basolateral surface, where it functions as part of the intercellular tight junction (5); CAR is thus inaccessible to virus approaching the apical cell surface from the intestinal lumen. Some CVB isolates bind to a second receptor, decay-accelerating factor (DAF) (6-8). Unlike CAR, DAF is highly expressed on the apical surface of polarized epithelial cells, and infection of polarized cells depends on virus attachment to DAF (9). In addition to providing a docking site for virus, DAF mediates a series of intracellular signals that permit virus to move ac...