Autoantibodies to neurofibrillary tangles and brain tissue in Alzheimer's disease. Establishment of Epstein-Barr virus-transformed antibody-producing cell lines.

Abstract

Alzheimer's disease (AD) is a major form of dementia . The pathological hallmark is a marked increase of neuritic plaques and neurofibrillary tangles (NFT) in certain regions of the AD brain. Despite considerable advances, the etiology and pathogenesis of AD remain unknown (reviewed in reference 1). Although autoimmunity and other immunological factors have been proposed to play certain roles, previous studies to demonstrate serum antibrain and other relevant antibodies have been complicated by the presence of high titers of other autoantibodies such as antinuclear antibodies (2) . Our approach to circumvent this difficulty is to derive multiple B cell lines by Epstein-Barr viral (EBV) transformation of a limited number of B cells from the blood of patients with clinically diagnosed AD and age-matched controls . The present studies describe our findings, by immunocytochemical techniques, that certain EBV-transformed cell lines secrete autoantibodies reactive with NFT and neuronal tissue.Volume 165 January 1987 245-250 Materials and Methods BriefDefinitive ReportDerivation of EBV-transformed B Cell Lines. Non-T cells were isolated according to Fu et al. (3) from the peripheral blood of five patients with clinically diagnosed AD, ages 59, 61, 66, 68, and 75, respectively and six normal age-matched controls ages, 60, 61, 66, 66, 71, and 72, respectively . They were seeded at 1 .5-3.0 X 10s cells/well in RPMI 1640, 10% FCS, with 10 allogeneic mononuclear cells irradiated with 3,000 rad as feeder cells, in 96-well plates . EBV-containing B95-8 cell line supernatant was added and culture medium was replaced weekly. Cell lines were considered to be established when the cell cultures were expanded to 7 ml at 5 X 105 cells/ml. They were cryopreserved and the supernatants were analyzed for antibodies (Ab) of desired reactivity. Passive hemagglutination inhibition (3) showed that most of the supernatants secreted IgM at 0.5-5 pg/ml. The cell lines were either monoclonal or oligoclonal as indicated by the presence of a single class of light chains.Immunocytochemistry. SDS-treated isolated NFT were prepared by the long procedure of Igbal et al. (4) using frontal and temporal cortex from two autopsied brains with a neuropathological confirmation of AD . 2 Al of isolated NFT containing 50-100 NFT

Rheumatoid factors isolated from patients with autoimmune disorders are derived from germline genes distinct from those encoding the Wa, Po, and Bla cross-reactive idiotypes.

“…The screening oligomers used for each antibody are as follows: (a) RF-KES1 studies used a CDR1 probe corresponding to amino acids [27][28][29][30][31][32][33][34] The analysis of the JKV locus of patient TS was conducted in a similar manner. The genomic JKV gene segment was amplified using oligomers corresponding to the 5' and 3' flanking regions of this locus.…”

“…The two monospecific rheumatoid factors RFKESl and RF-TMCl were isolated from EBV-transformed peripheral blood B-lymphocytes obtained from a patient with systemic lupus erythematosus (KES) and a normal individual (TMC). These cell lines were derived by EBV transformation of a limited number of non-T lymphocytes as described (27) and subsequently single cell cloned by limiting dilution. The remaining seven rheumatoid factors were isolated from EBV-transformed B-lymphocytes obtained from the synovial tissue of two patients with rheumatoid arthritis (TS and SJ) and one patient with polyarticular juvenile rheumatoid arthritis (KL) (28).…”

Evidence for Multiple Shared Antigenic Determinants within Ro60 and Other Lupus-Related Ribonucleoprotein Autoantigens in Human Autoimmune Responses

“…Immunofluorescence mAbs were assayed for reactivity on HeLa cells as previously described using methanol fixation (20). The bound Ab was detected with our goat anti-human F(abЈ) 2 rhodamine conjugate.…”

Microglial in Neurodegenerative Disorders: Emphasis on Alzheimer’s Disease

“…Reports indicate that antibrain antibodies in AD recognize cholinergic elements [73][74][75][76], neurofibrillary tangles [77], thyroglobulin [78] and vascular proteoglycans [79], All of these antigens have a relationship to AD. Au toantibodies may even be harmful.…”