Pathogenic Profiles and Molecular Signatures of Antinuclear Autoantibodies Rescued from NZM2410 Lupus Mice

Abstract

Two outstanding questions concerning antinuclear antibodies (ANAs) in lupus involve their pathogenic potential and their molecular signatures. To address these questions, a panel of 56 antinuclear and 47 nonnuclear binding monoclonal antibodies was rescued from four seropositive NZM2410 lupus mice. The monoclonals varied in their reactivity to nucleosomes, ssDNA, dsDNA, and glomerular substrate. A large fraction of the antibodies demonstrated apparent polyreactivity (to DNA, histones, and glomerular antigens) due to bound, DNase-1 sensitive nuclear antigenic bridges. Although nephrophilic immunoglobulin (Ig) M and IgG antibodies were the most pathogenic, the dsDNA-binding antibodies were modestly so; in contrast, antinucleosome antibodies were clearly not pathogenic. Compared with the nonnuclear antigen-binding monoclonal antibodies rescued from the same mice, ANAs exhibited increased utilization of VH5/7183 genes and highly cationic heavy chain (HC) CDR3 regions. Most intriguingly, the CDR3 regions of the ANAs exhibited alternating arginine/lysine peaks at H96, H98, and H100, with neutral troughs at H95, H97, and H99. To summarize, glomerular-binding anti-dsDNA antibodies appear to be the most pathogenic variety of lupus autoantibodies. The presence of an alternating charge pattern in their HC CDR3 regions appears to be a prominent hallmark of ANAs.

The Lupus Susceptibility Locus Sle1 Facilitates the Peripheral Development and Selection of Anti-DNA B Cells through Impaired Receptor Editing

“…Thus, the primary repertoire of B cells in B6.Sle1 mice might have higher numbers of autoreactive B cells compared with those of normal mice. Such autoreactive B cell-enriched primary repertoires were confirmed by the observation of mature naive B cell repertoires in patients with SLE or rheumatoid arthritis (6)(7)(8), as well as by a lupus-prone murine model (9,10).…”

“…These primers allow the amplification of transgenes, as well as endogenous sequences, including those with nonproductive rearrangements and edited sequences with VH replacements or D invasions. The Ig-k LC primers used have the potential to amplify most Ig-k gene families, including Vk1, Vk2, Vk4, Vk8, Vk9, Vk2, Vk19, Vk20, Vk21-3, Vk21-4, Vk23, Vk31, Vk33, Vk38, L1, L2, Lx, and RF, based on our previous work (9,13,22).…”

“…Splenocytes from 9-12-mo-old seropositive KI mice were fused to the SP2/0 fusion partner, as described previously (9). All wells with single colonies were screened for Ab production 7-10 d after fusion.…”

“…The primers and conditions for HC and LC RT-PCR were as described previously (9,13,22). The HC was amplified using HC primers as follows: 59 primer, 59-AGGT(G/C)(A/C)A(A/G)CTGCAG(G/ C)AGTC(A/T)GG-39, 39 primers to Cm, 59-CAGGGGGCTCCGCAGGA-GACGAGG-39, Cg, 59-GGACAGGGATCCAGAGTTCC-39.…”

Peripheral B cell receptor editing may promote the production of high‐affinity autoantibodies in CD22‐deficient mice

“…We therefore compared the frequency of somatic mutations in framework regions (FR) and CDR of D42H/VjRF-Jj5 anti-DNA mAb derived from CD22 -/-D42H +/-mice to those of diseased D42H +/-NZB/NZW female mice. For comparison, the frequency of somatic mutations in anti-nuclear mAb from non-Tg, genetically related, lupus-prone NZM2410 mice [31] is also shown (Fig. 2D).…”