ab21503

Use of human chondrocyte cell cultures to identify and characterize reactive antibodies in rheumatoid arthritis sera

A. Sabbatini, C. Tacchetti¹, S. Tommasi, F. Bongiorni, P. Migliorini

Clinical Immunology Unit, Department of Internal Medicine, University of Pisa and the ¹Department of Experimental Medicine, Section of Anatomy, University of Genova, Italy.

ABSTRACT
Objective
To study the reactivity of rheumatoid arthritis (RA) sera with human chondrocyte populations isolated from normal cartilage and expanded in vitro.

Methods
Human articular chondrocytes were cultured as adherent (non-differentiated) cells on plastic dishes or in suspension (differentiated) on dishes previously coated with a thin layer of 1% agarose. Sera from 28 RA patients and 5 paired synovial fluids were tested on lysates from chondrocytes and fibroblasts as control by immunoblot. Antigen expression on the cell membrane was evaluated by flow cytometry in a few sera.

Results
In 9/28 RA sera IgG antibodies specific for chondrocyte antigens (97kDa, 74kDa, 67kDa, 60kDa, 54kDa, 48kDa and 37kDa) were detected. Twelve sera reacted with proteins expressed both on chondrocytes and fibroblasts and 7 with fibroblasts only; two sera had no reactivity. When lysates from adherent or suspension chondrocytes were compared, RA sera reacted with higher intensity and detected more antigens on chondrocytes cultured in suspension. Flow cytometry assay demostrated that RA sera are able to recognize antigens expressed on the cell membrane of the human chondrocytes.

Conclusion
Our data indicate that: a) 32% of the RA sera contain antibodies reactive with antigens expressed exclusively by chondrocytes, but this value rises to 75% if antigens expressed both by chondrocytes and fibro-blasts are considered; b) the reactivity of fully differentiated chondrocytes in suspension culture is higher than the reactivity of chondrocytes cultured in monolayer; and c) some of the chondrocyte-specific antigens identified are associated with the chondrocyte membrane. Thus, in vitro cultured chondrocytes may be used to study both the specificity and the biological activity of autoantibodies in RA.

Key words
Chondrocytes, rheumatoid arthritis, autoantibodies.

Supported by a grant from the Italian Ministry of University and Research and the National Research Council (Target project Biotechnology).
Please address correspondence and reprint requests to: Paola Migliorini, MD, Clinical Immunology Unit, Department of Internal Medicine, Via Roma 67, 56126 Pisa, Italy.
E-mail: p.migliorini@int.med.unipi.it