Free Communication: Immunology

Abstract

*††YukitoshiTakahashi, *NaomiSakaguchi, ‡MasahikoWatanabe, ‡HisashiMori, ‡MasayoshiMishina, §HideoAiba, ∥TomokoMiyajima, ¶YoshiakiSaitou, **YasushiUchida, †NaokiMatsuo, †SatomiHattori, and *NaomiKondo *Department of Pediatrics, Gifu University School of Medicine, Gifu ; †Department of Anatomy and Embryology, Hokkaido University Graduate School of Medicine, Hokkaido ; ‡Department of Molecular Neurobiology and Pharmacology, School of Medicine, University of Tokyo, Tokyo ; §Shizuoka Children's Hospital, Shizuoka ; ∥Shiga Medical Center for Children, Shiga ; ¶Department of Pediatrics, Tokyo Women's Medical University, Tokyo ; **Department of Pediatrics, Nagahama Municipal Hospital, Nagahama ; and ††Gifu Prefectural Gifu Hospital, Gifu, Japan Purpose: Glutamate receptors (GluRs) in the central nervous system play an important role in the excitatory synapse and have been implicated in neurodegenerative diseases and symptomatic epilepsy. After Rogers et al. reported that autoantibodies against GluR3 were involved in some cases of Rasmussen encephalitis (Science 1994; 265:648–51), studies have focused on other neurologic diseases with a possible causal relation to autoantibodies against GluRs. To reveal the pathophysiologic mechanisms of severe epilepsia partialis continua (Y. Takahashi et al. ActaNeurolScand 1997; 96:345–52), we developed a detection system highly sensitive to autoantibodies against the N‐methyl‐d‐aspartate (NMDA) GluRα2 subunit and found autoantibodies against GluRα2 in patients with chronic progressive epilepsia partialis continua of childhood (Y. Takahashi et al. AnnRepJpnEpilepsyResFound 1999; 11:45–52). Serum autoantibodies against GluRα2 in patients with chronic progressive epilepsia partialis continua of childhood have epitopes on the C‐terminal of GluR2 molecules. In this study, we tried to reveal the site of immunogenesis of autoantibodies by comparing the epitopes in serum and cerebrospinal fluid (CSF). Methods: Six patients with immunoglobulin G (IgG) autoantibodies against the NMDA GluRα2 subunit in serum and CSF were examined. The age at onset of their disease ranged from 2 months to 12.5 years for epilepsy, and 7 months to 12.9 years for epilepsia partialis continua. Probable causative factors were aseptic meningitis, encephalopathy, IgA deficiency, influenza infection, and immunization. The patients had intractable epilepsia partialis continua and complex partial seizures. Neuropsychological outcomes were poor in three patients; quadriplegia developed in four patients, and hemiplegia, in one. Four sequences of GluRα2 cDNA (an N‐terminal fragment, and three C‐terminal fragments) were inserted into bacterial fusion protein vectors (pGEX and pMAL), and the production of those bacterial fusion proteins was induced by isopropyl thiogalactose (IPTG). Three C‐terminal peptides existed after the fourth transmembrane region in the order of C1, C2, and C3. Western blots transferred bacterial fusion proteins composed of GluRα2 fragments to nitrocellulose membranes, and the membranes were incubated with the patients' serum or CSF and stained with IgG‐type second antibodies coupled to alkaline phosphatase. Results: In five of the six patients, IgG autoantibodies against GluRα2 in CSF had all four epitopes, and in a patient with mild disease, without mental retardation or motor dysfunction, IgG autoantibodies against GluR2 in CSF had all epitopes except C1. In the five patients with all epitopes in CSF, one patient had one serum epitope, two patients had two serum epitopes, and two patients had three serum epitopes. Their sera showed epitope N in two patients, epitope C1 in two patients, epitope C2 in five patients, and epitope C3 in two patients. The patient with mild disease and three epitopes in CSF had only one epitope, C2, in serum. Conclusions: The diversity of epitopes of IgG autoantibodies against GluRα2 was broader in the CSF than in the sera of all six patients. It is, therefore, suggested that the immunogenesis of IgG autoantibodies against GluRα2 occurs mainly in the central nervous system of patients with chronic progressive epilepsia partialis continua of childhood.