The coxsackie-adenovirus receptor induces an inflammatory cardiomyopathy independent of viral infection.

The coxsackie-adenovirus receptor (CAR) is a viral receptor for Group B coxsackieviruses (CVBs) and adenoviruses. CAR has been linked with the innate immune response to CVB myocarditis, and with activation of inflammatory cells in vitro. We hypothesized that CAR activates signals that promote inflammation in the myocardium independent of viral infection. To test this we conditionally overexpressed murine CAR in cardiomyocytes of adult binary transgenic mice under the control of a tetracycline-responsive (tet-off) α-myosin heavy chain (αMtTA) promoter (mCAR(+)/αMtTA(+) mice). An inflammatory cardiomyopathy developed in both lines generated (6-mCAR(+)/αMtTA(+) and 12-mCAR(+)/αMtTA(+)) following withdrawal of doxycycline. Cardiac CAR was upregulated at 4weeks of age in 6-mCAR(+)/αMtTA(+) mice and induced a mild inflammatory infiltrate (score 1.3 of 4.0±0.3) at 6weeks, with 95% of mice surviving to that time. In the second line, 12-mCAR(+)/αMtTA(+) mice, CAR was upregulated in the majority of mice by 3weeks of age, and by 5weeks of age more severe cardiac inflammation (score 2.8 of 4.0±0.4) developed with only 56% of mice surviving. The cardiac inflammatory infiltrate was primarily natural killer cells and macrophages in both mCAR(+)/αMtTA(+) lines. A proinflammatory cytokine response with increased cardiac interferon-γ, interleukin (IL)-12, IL-1ß, tumor necrosis factor-α and IL-6 was detected by real-time RT-PCR. CAR has been linked to signaling via the inflammatory mitogen-activated protein kinase (MAPK) cascades; therefore, we evaluated the response of these pathways in hearts with upregulated CAR. Both stress-activated JNK and p38MAPK were activated in mCAR(+)/αMtTA(+) hearts prior to onset of inflammation and in isolated mCAR(+)/αMtTA(+) cardiomyocytes. In conclusion, we show for the first time that CAR upregulation in the adult mouse heart induces cardiac inflammation reminiscent of early viral myocarditis. CAR-induced stress-activated MAPK signaling may contribute to the development of cardiac inflammation unrelated to viral infection per se.

Regulatory T cells protect mice against coxsackievirus-induced myocarditis through the transforming growth factor beta-coxsackie-adenovirus receptor pathway.

BACKGROUND: Coxsackievirus B3 infection is an excellent model of human myocarditis and dilated cardiomyopathy. Cardiac injury is caused either by a direct cytopathic effect of the virus or through immune-mediated mechanisms. Regulatory T cells (Tregs) play an important role in the negative modulation of host immune responses and set the threshold of autoimmune activation. This study was designed to test the protective effects of Tregs and to determine the underlying mechanisms. METHODS AND RESULTS: Carboxyfluorescein diacetate succinimidyl ester-labeled Tregs or naïve CD4(+) T cells were injected intravenously once every 2 weeks 3 times into mice. The mice were then challenged with intraperitoneal coxsackievirus B3 immediately after the last cell transfer. Transfer of Tregs showed higher survival rates than transfer of CD4(+) T cells (P=0.0136) but not compared with the PBS injection group (P=0.0589). Interestingly, Tregs also significantly decreased virus titers and inflammatory scores in the heart. Transforming growth factor-beta and phosphorylated AKT were upregulated in Tregs-transferred mice and coxsackie-adenovirus receptor expression was decreased in the heart compared with control groups. Transforming growth factor-beta decreased coxsackie-adenovirus receptor expression and inhibited coxsackievirus B3 infection in HL-1 cells and neonatal cardiac myocytes. Splenocytes collected from Treg-, CD4(+) T-cell-, and PBS-treated mice proliferated equally when stimulated with heat-inactivated virus, whereas in the Treg group, the proliferation rate was reduced significantly when stimulated with noninfected heart tissue homogenate. CONCLUSIONS: Adoptive transfer of Tregs protected mice from coxsackievirus B3-induced myocarditis through the transforming growth factor beta-coxsackie-adenovirus receptor pathway and thus suppresses the immune response to cardiac tissue, maintaining the antiviral immune response.

Cardiomyocyte-targeted overexpression of the coxsackie-adenovirus receptor causes a cardiomyopathy in association with beta-catenin signaling.

The coxsackie-adenovirus receptor (CAR) is an adhesion molecule found at the intercalated disc of cardiomyocytes in association with other adherens and tight junction proteins. CAR expression is increased at cardiomyocyte junctions in patients with heart failure. It is not known what contribution elevated CAR expression makes to cardiac pathology. We generated a binary transgenic mouse enabling cardiac-restricted doxycycline-regulated expression of Flag-tagged murine CAR (mCAR(+)/alpha MtTA(+) mice). Myocardial CAR levels were increased 6-fold in mCAR(+)/alpha MtTA(+) mice, localizing to intercalated discs and sarcolemma. Well at birth, mCAR(+)/alpha MtTA(+) mice developed a severe cardiomyopathy and died by 4 weeks. Cardiomyocyte hypertrophy was evident at 1 week, with increased heart:body weight ratios by 3 weeks. Disorganization and degeneration of cardiomyocytes were evident with disrupted adherens junctions. Doxycycline administration turned off transgene expression and rescued mice from the development of the cardiomyopathic phenotype. In CAR-overexpressing mCAR(+)/alpha MtTA(+) mice, adherens junction proteins were abnormally expressed. N-cadherin protein levels were 83% lower in mCAR(+)/alpha MtTA(+) hearts vs controls at 1 week, with levels subsequently increased above controls at 3 weeks. beta-catenin expression was 90% and 135% above controls at 1 and 3 weeks, respectively. Nuclear translocation of beta-catenin in cardiomyocytes of mCAR(+)/alpha MtTA(+) mice was associated with increased c-myc RNA, a target of active beta-catenin known to be associated with cardiac hypertrophy. Our study is the first to demonstrate that increased CAR expression can induce a cardiomyopathy and supports a model whereby the pathogenesis is determined by CAR stimulated beta-catenin signaling, and/or disruption of the adherens junction.

Eosinophilic myocarditis temporally associated with conjugate meningococcal C and hepatitis B vaccines in children.

We report the first cases of tissue-proven eosinophilic myocarditis after single vaccine administration of conjugate meningococcal C and hepatitis B vaccine, respectively. The nature of histopathologic findings strongly supports hypersensitivity reaction and negates viral etiology, which is typically characterized by a lymphocytic infiltrate. Both episodes resolved with corticosteroid therapy. To enhance discussion of our cases, we performed a systematic review of the literature on postimmunization myocarditis or pericarditis, and identified 37 publications, reporting 269 cases during the search period (1966-2007). Time of onset of cardiac symptoms in all patients ranged from 1 to 30 days postimmunization.

Enhanced ERK-1/2 activation in mice susceptible to coxsackievirus-induced myocarditis.

Group B coxsackieviral (CVB) infection commonly causes viral myocarditis. Mice are protected from CVB3 myocarditis by gene-targeted knockout of p56(Lck)(Lck), the Src family kinase (Src) essential for T cell activation. Extracellular signal-regulated kinase 1 and 2 (ERK-1/2) can influence cell function downstream of Lck. Using T cell lines and neonatal cardiac myocytes we investigated the role of ERK-1/2 in CVB3 infection. In Jurkat T cells ERK-1/2 is rapidly activated by CVB3; but, this response is absent in Lck-negative JCaM T cells. Inhibition of ERK-1/2 with UO126 reduced CVB3 titers in Jurkat cells, but not in JCaM cells. In cardiac myocytes CVB3 activation of ERK-1/2 is blocked by the Src inhibitor PP2. In addition, viral production in myocytes is decreased by Src or ERK-1/2 inhibition. In vitro, in both immune and myocardial cells, ERK-1/2 is activated by CVB3 downstream of Lck and other Src's and is necessary for efficient CVB3 replication. In vivo, following CVB3 infection, ERK-1/2 activation is evident in the myocardium. ERK-1/2 activation is intense in the hearts of myocarditis-susceptible A/J mice. In contrast, significantly less ERK-1/2 activation is found in the hearts of myocarditis-resistant C57BL/6 mice. Therefore, the ERK-1/2 response to CVB3 infection may contribute to differential host susceptibility to viral myocarditis.

A child with raccoon roundworm meningoencephalitis: A pathogen emerging in your own backyard?

Raccoon roundworm (Baylisascaris procyonis) is a cause of devastating neural and ocular disease. The first documented case of raccoon roundworm encephalitis in Canada, in a seven-year-old boy who presented with severe neurological impairment, is presented. His significant recovery illustrates the importance of clinical suspicion and the benefit of early treatment.

Children hospitalized with severe acute respiratory syndrome-related illness in Toronto.

OBJECTIVE: An outbreak of severe acute respiratory syndrome (SARS) occurred in the greater Toronto area between February and June 2003. We describe the clinical, laboratory, and epidemiologic features of children who were admitted to the Hospital for Sick Children, Toronto, with a presumptive diagnosis of suspect or probable SARS. METHODS: A prospective investigational study protocol was established for the management of children with a presumptive diagnosis of suspect or probable SARS. All were ultimately classified as having probable SARS, suspect SARS, or another cause on the basis of their epidemiologic exposure, clinical and radiologic features, and results of microbiologic investigations. RESULTS: Twenty-five children were included; 10 were classified as probable SARS and 5 were classified as suspect SARS, and in 10 another cause was identified. The exposure consisted of direct contact with at least 1 adult probable SARS case in 11 children, travel from a World Health Organization-designated affected area in Asia in 9 children, and presence in a Toronto area hospital in which secondary SARS spread had occurred in 5 children. The predominant clinical manifestations of probable cases were fever, cough, and rhinorrhea. With the exception of 1 teenager, none of the children developed respiratory distress or an oxygen requirement, and all made full recoveries. Mild focal alveolar infiltrates were the predominant chest radiograph abnormality. Lymphopenia; neutropenia; thrombocytopenia; and elevated alanine aminotransferase, aspartate aminotransferase, and creatine kinase were present in some cases. Nasopharyngeal swab specimens were negative for the SARS-associated coronavirus by an in-house reverse transcriptase-polymerase chain reaction in all 25 children. CONCLUSIONS: Our results indicate that SARS is a relatively mild and nonspecific respiratory illness in previously healthy young children. The presence of fever in conjunction with a SARS exposure history should prompt one to consider SARS as a possible diagnosis in children irrespective of the presence or absence of respiratory symptoms. Reverse-transcriptase polymerase chain reaction analysis of nasopharyngeal specimens seems to be of little utility for the diagnosis of SARS during the early symptomatic phase of this illness in young children.