Viral Protein VP4 Is a Target of Human Antibodies Enhancing Coxsackievirus B4- and B3-Induced Synthesis of Alpha Interferon

Abstract

Coxsackievirus B4 (CVB4)-induced production of alpha interferon (IFN-␣) by peripheral blood mononuclear cells (PBMC) is enhanced in vitro by nonneutralizing anti-CVB4antibodies from healthy subjects and, to a higher extent, from patients with insulin-dependent diabetes mellitus. In this study, we focused on identification of the viral target of these antibodies in CVB systems. High levels of IFN-␣ were obtained in supernatants of PBMC incubated with CVB4E2 or CVB3 and plasma from healthy subjects and, to a higher extent, from patients. The VP4 capsid proteins dissociated by heating at 56°C from CVB4E2 (VP4 CVB4 ) and CVB3 (VP4 CVB3 ) but not H antigen preincubated with plasma from healthy subjects or patients inhibited the plasma-dependent enhancement of CVB4E2-and CVB3-induced IFN-␣ synthesis. There was no cross-reaction between VP4 CVB4 and VP4 CVB3 in the inhibiting effect. IFN-␣ levels in culture supernatants showed dosedependent correlation with anti-VP4 antibodies eluted from plasma specimens using VP4-coated plates. There were higher index values for anti-VP4 antibodies detected by enzyme-linked immunosorbent assay (ELISA) and higher proportions of positive detection in 40 patients than in 40 healthy subjects (80% versus 15% for anti-VP4 CVB4 ). There was no relationship between the levels of anti-CVB neutralizing antibodies and the detection of anti-VP4 antibodies by ELISA. The CVB plasma-induced IFN-␣ levels obtained in PBMC cultures in the anti-VP4 antibody-positive groups were significantly higher than those obtained in the anti-VP4 antibody-negative groups regardless of the titers of anti-CVB neutralizing antibodies. These results show that VP4 is the target of antibodies involved in the plasma-dependent enhancement of CVB4E2-and CVB3-induced IFN-␣ synthesis by PBMC.

Anti-coxsackievirus B4 (CV-B4) enhancing activity of serum associated with increased viral load and pathology in mice reinfected with CV-B4

“…Interestingly, in patients with type 1 diabetes, an anti-CV-B4 enhancing activity of serum was observed. 11,26,27 Whether enhancing antibodies play a role in the pathogenesis of CV-B -induced diseases in vivo, especially type 1diabetes deserves further studies. Future studies will be directed along this line in our laboratory.…”

Immunoglobulin G-dependent enhancement of the infection with Coxsackievirus B4 in a murine system

“…This is reminiscent of previous studies describing that human serum harbouring an anti-CV-B4 E2 neutralizing activity had an enhancing anti-CV-B4 E2 activity, due to anti-CV-B4 E2 IgG, in human PBMC or cell line cultures, provided the serum was diluted. 6,12,13 Intracellular viral RNA was detected by RT-PCR when spleen cells cultures were inoculated with CV-B4 E2 mixed with diluted serum or serum-derived IgG from CV-B4 E2-infected mice, whereas it was only detected by sn-RT-PCR, a more sensitive technique, when cells where inoculated with CV-B4 E2 or with CV-B4 E2 mixed with serum or serum-derived IgG from mockinfected mice. These observations suggest that the level of intracellular viral RNA was higher in the first case.…”

Production of Cross‐Reactive Peptide Antibodies against Viral Capsid Proteins of Human Enterovirus B to Apply Diagnostic Reagent

“…All anti-peptide rabbit sera were tested by ELISA for the induction of specific antibody against the corresponding peptide, as modified from a previously reported method (4). Briefly, the wells of 96-well ELISA plates (Dynatech Lab., Chantilly, Va., U.S.A.) were coated with peptide antigen at 250 ng/well overnight at 4 C. The sera of rabbits immunized with peptide were diluted between 1:1,000 and 1:100,000, added to the wells, and incubated for 1 hr at 37 C. Rabbit IgG-horseradish peroxidase was added and incubated for 1 hr at 37 C, and was then visualized with the substrate 3,3',5,5'-tetramethylbenzidine.…”