Burton J. Landau

The Early Interaction of Coxsackievirus B3 with HeLa Cells

Summary Purified coxsackievirus B3 labeled with 32P- or 14C-valine interacted with HeLa cells in a manner resembling that found by others for poliovirus. Radioactivity eluted from cells at 37°, pH 7, as noninfectious virus, but contained infectious RNA. Both elution and “eclipse” of virus were inhibited irreversibly at low pH to provide further evidence that these processes may be related properties of the cell surface.

Induction of heterotypic virus resistance in adult inbred mice immunized with a variant of Coxsackievirus B3

Infection of adult male C3H/HeJ mice with a host range variant of Coxsackievirus B3 (CB3W-RD) induced resistance in these mice to an otherwise lethal dose of Coxsackievirus B1 (CB1). The protective effect induced by CB3W-RD was detectable as early as 1 day post-vaccination and was still present 10 weeks later. While untreated mice infected with CB1 died within 5 days because of massive hepatic necrosis, the liver was spared in mice immunized with CB3W-RD and then challenged with CB1. In general, CB1 titers in heart, liver, and pancreas of CB3W-RD-vaccinated animals were lower than that found in unvaccinated animals. Virus neutralizing antibody was not a mediator of this heterotypic, virus-induced protective effect. In addition, the outcome of CB1 infection could be modified if superinfection with CB3W-RD took place within 1-4 days following CB1 infection. In this regard, maximum therapeutic efficacy was observed when CB1 infected mice were superinfected 2 days after CB1 infection. CB1-infected mice that survived as a result of treatment with CB3W-RD exhibited liver regeneration but did develop myocardial necrosis.

Pathogenesis of acute myocardial necrosis in inbred mice infected with coxsackievirus B3

The pathogenesis of myocardial necrosis due to CB3W infection was studied in BALB/c and C3H/HeJ mice. BALB/c mice infected with 5 x 10(4) pfu were found to die of massive hepatic coagulative necrosis before myocardial changes occurred. Reducing the inoculum size to 5 x 10(2) pfu resulted in sublethal hepatic involvement and multifocal myocardial coagulative necrosis by day 7 p.i. In contrast, C3H/HeJ mice survived infection and developed multifocal myocardial coagulative necrosis, but not liver disease following inoculation with as much as 5 x 10(6) pfu of CB3W. As with BALB/c mice infected with 5 x 10(2) pfu, myocardial lesions became apparent in C3H/HeJ mice a few days after peak cardiac virus titer was attained. Minimal inflammatory infiltrate was seen following development of cellular necrosis and was restricted to the areas of virus-induced pathologic change. However, no evidence was found for virus-specific cytotoxic T cell activity or for delayed type hypersensitivity responses. Furthermore, myocardial necrosis in CB3W-infected, T cell-depleted C3H/HeJ mice was as severe as in CB3W-infected, immunocompetent mice. These data have led us to conclude that cardiac lesions were due to virus-induced cytopathology rather than immunopathogenic mechanisms.

Limited Persistence of Viral Antigen in Coxsackievirus B3 Induced Heart Disease in Mice

Summary Following acute Coxsackievirus B3 infection of young adult mice, peak virus titers in the heart were found 7 days postinfection, after which virus was cleared. Though postinfection cardiomyopathy was observed following disappearance of virus, evidence for the continued presence of structural viral antigen in cardiac cells could not be found. Postinfection myocarditis is likely to be due to cytotoxic T-lymphocytes directed against nonstructural viral antigen or, alternatively, to a cellular neoantigen.

A Murine Model for Coxsackievirus B3-Induced Acute Myocardial Necrosis for Study of Cellular Receptors as Determinants of Viral Tropism

A number of studies have contributed to the working hypothesis that specific cellular receptors, if present in sufficient quantity, serve as major determinants of both the host range and tissue tropism in the pathogenesis of human and animal picornavirus infections [14, 21, 28, 36, 42]. Though picornaviruses contain relatively small genomes, they produce a bewildering array of diseases. It is likely that one of the several factors contributing to disease diversity is due to the specific requirement of individual viruses for host cell receptors, For example, we have postulated [12a] that genetic changes in the virion attachment site could occur during virus replication in infected individuals, These changes could influence the course of the disease [9, 23, 51] by altering the virus receptor specificity, This would be of special importance if different receptors occurred on different tissues.

The permissiveness of differentiating mouse muscle-cell cultures to infection by group A coxsackievirus types 1 and 5.

Summary Differentiating primary fetal-mouse muscle-cell cultures have been shown to support the replication of prototype strains of coxsackieviruses A1 and A5. Heretofore, mouse cell cultures had been found to be restrictive to the growth of these agents. Myoblastic elements rather than fibroblasts appeared to be the primary cell type susceptible to infection by those coxsackieviruses of Group A tested. This investigation was supported in part by U.S. Public Health Service Research Grant AI-03771 from the National Institute of Allergy and Infectious Diseases. We thank Barbara Goldberg and Christine Sturm for competent technical assistance.

[35] Methods for assay of cellular receptors for picornaviruses

Publisher Summary This chapter provides an overview of various methods for assay of cellular receptors for picornaviruses. The picornaviruses are small, single-stranded RNA-containing, nonenveloped viruses comprising over 170 immunologically distinct human viruses and a multitude of animal viruses. The several genera include the human enteroviruses and rhinoviruses, the murine cardioviruses, and the viruses of foot-and-mouth disease (FMDV). Factors influencing the attachment rate of virions to receptors include cell and virion concentration, pH, temperature, ion species and concentration, viscosity, presence of sulfhydryl groups, proteolytic enzymes, and assorted inhibitors. The chapter discusses assays of receptors for virus attachment; assays for virus attachment to subcellular fractions, and; to solubilized membranes, assay of virus penetration, and assay of virus eclipse and virus uncoating. It is believed that the methods and comments presented in the chapter will stimulate others to explore further the role of receptors in the molecular events in viral infections and in the cellular function(s) that receptors serve.

Determination of the Success of the Integration of a Business of Healthcare Module into the Medical School Curriculum

Purpose: Assessment of the knowledge of health care economics of four consecutive classes of second year medical students before and after an 8 hour module on the business of health care was performed. Methods: Through pre and post-tests, four consecutive classes of second year medical students were assessed as to their knowledge of basic principles of the business of health care before and after an 8 hour lecture module given over a 2 week period in the middle of their academic year. Student evaluations of the module were collected at the end of the course in each of the four years of the study. Results: Although the students came from diverse educational backgrounds and had many different life experiences before entering medical school, few students in any of the four consecutive classes could pass a basic examination of these principles. At the completion of the 2 week module, nearly all of the students passed this test, suggesting that the concentrated lecture format successfully addressed the material. Student evaluations of the course were positive and indicated that delivery of the material in a concentrated lecture format was effective. Conclusions: Second year medical students of diverse backgrounds have little working knowledge of the US health care system. An 8 hour concentrated lecture course can be effective in teaching basic principles. Continued review and application of these principles throughout the clinical years of training in an on-line and interactive session format is discussed by the authors. Further studies assessing optimal teaching methods need to be performed.

Perspectives on Cellular Receptors as Determinants of Viral Tropism

Coxsackievirus (CV) infections of humans were recently reviewed1 and are also discussed elsewhere in this volume. These agents cause a wide variety of syndromes, including meningitis, pleurodynia, myositis, herpangina, myocarditis, pericarditis, pancreatitis, hepatitis, nephritis, orchitis, gastroenteritis, exanthems, acute respiratory disease, and congenitally acquired disease. In addition, inapparent or mild infections may be common and provide a source of virus to help explain the ubiquitous spread of infections. No conclusive evidence is available to explain why some individuals develop acute self-limiting infections, while others develop a severe or chronic disease. Nevertheless, it is predicted that cellular receptors play an important role in determining the tropism of CV in the pathogenesis of infection.

Picornarvirus Receptors in Pathogenesis

The picornaviruses comprise a large number of viruses with a restricted host range. The diverse diseases produced in man and animals (Table 9.1) suggest that the different viruses in this family are more heterogeneous in disease potential than suggested from the similarity in their biochemical and biophysical properties (Newman et al., 1973; Rueckert, 1976). Although polioviruses are considered the prototypes of this group, there appear to be differences between the cellular receptors for polioviruses and members of the other virus subgroups. These differences may reflect the variation in tissue tropisms and host range observed for these viruses. The major tissue tropisms of picornaviruses in humans are tabulated in Table 9.2 based on the embryological origin of the tissues. The purpose of this chapter is to explore the concept that specific receptors for picornaviruses determine cellular tropism and thus influence the course of events leading to disease. Since many studies have been performed with cells grown in culture, we will consider their applicability to the in vivo situation.