Detection of nucleosome–IgG immune complexes in ascites from mice transplanted with anti-DNA antibody-secreting hybridomas and in plasmas from MRL-lpr/lpr mice

Abstract

SUMMARYAutoantibodies directed against chromatin components characterize lupus diseases. Immune complexes made of these autoantibodies bound to nucleosomes released from dead cells could play some pathogenic role. The aims of this study were to investigate if nucleosome-IgG complexes could contaminate IgG anti-DNA MoAb preparations, and if such complexes circulate in lupus diseases. A new method was set up using preformed nucleosome-IgG complexes. Complexes were adsorbed onto microplate through Fc binding and nucleosomal DNA was detected by internal incorporation of labelled nucleotide. Using this method, high amounts of complexes were found in ascites from mice transplanted with anti-DNA antibody-secreting hybridomas. In some ascites, nucleosome was found to be strongly associated with the MoAb, confirming that nucleosome-IgG complexes could contaminate monoclonal autoantibody preparations. In MRL-lpr/lpr mice, nucleosome-IgG complexes were detected at 16-24 weeks of age at a time when kidney lesions are rapidly worsening, raising the question of their pathogenic significance.

The key role of nucleosomes in lupus

“…Likewise, mouse inoculation with anti-dsDNA and antihistone B cell hybridomas leads to the formation of nucleosome-containing immune complexes in both ascitic fluid and plasma (15,31). These findings illustrate the ability of anti-dsDNA and antihistone lupus antibodies to react, via their respective nucleosomal targets, with nucleosomes.…”

Circulating plasma levels of nucleosomes in patients with systemic lupus erythematosus. Correlation with serum antinucleosome antibody titers and absence of clear association with disease activity

“…Nonetheless, the presence of circulating DNA(nucleosome? )-anti-DNA immune complexes has been difficult to demonstrate in the past (37,38), and in recent studies with mouse and human sera, direct assessment of such complexes in the circulation (16) or through glomerular basement membrane binding in vitro have revealed low levels of circulating immune complexes (39,40).…”

“…In this regard, Kramers et a1 (15) showed that nucleosome-antinucleosome immune complexes bound to the rat glomerular basement membrane when perfused in vivo, while antinucleosome antibodies alone did not. Finally, recent data have demonstrated the presence of nucleosome-IgG complexes in the circulation of 16-28-week-old MRL-lpr/lpr mice (16), a time when kidney lesions are rapidly worsening.…”

Calreticulin, a Potential Cell Surface Receptor Involved in Cell Penetration of Anti-DNA Antibodies

“…One of the major problems with the anti-DNA mAbs is that these IgG may be complexed with nucleosomes (DNA complexed with histones) released from dead cells in the culture supernatant of hybridomas and may give false positive results in assays of cross-reactive binding to several Ags (7,30,31). To assess the purity of the mouse mAbs used, we estimated the possible extent of nucleosome contamination.…”