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. 2024 Oct 21;83(11):1549-1560.
doi: 10.1136/ard-2024-225773.

Pathological autoantibody internalisation in myositis

Iago Pinal-Fernandez  1   2 , Sandra Muñoz-Braceras  3 , Maria Casal-Dominguez #  3   2 , Katherine Pak #  3 , Jiram Torres-Ruiz #  3   4 , Jon Musai  3 , Stefania Dell'Orso  3 , Faiza Naz  3 , Shamima Islam  3 , Gustavo Gutierrez-Cruz  3 , Maria Dolores Cano  5 , Ana Matas-Garcia  5   6   7 , Joan Padrosa  7 , Ester Tobias-Baraja  5   6   7 , Gloria Garrabou  5   6   7 , Iban Aldecoa  8 , Gerard Espinosa  6   9 , Carmen Pilar Simeon-Aznar  10   11 , Alfredo Guillen-Del-Castillo  10   11 , Albert Gil-Vila  10   11 , Ernesto Trallero-Araguás  10   11   12 , Lisa Christopher-Stine  2   13 , Thomas E Lloyd  2 , Teerin Liewluck  14 , Elie Naddaf  14 , Werner Stenzel  15 , Steven A Greenberg  16 , Josep Maria Grau  5   6   7 , Albert Selva-O'Callaghan  10   11 , Jose Cesar Milisenda  5   6   7 , Andrew Lee Mammen  1   2   13
Affiliations

Pathological autoantibody internalisation in myositis

Iago Pinal-Fernandez et al. Ann Rheum Dis. .

Abstract

Objectives: Autoantibodies targeting intracellular proteins are common in various autoimmune diseases. In the context of myositis, the pathologic significance of these autoantibodies has been questioned due to the assumption that autoantibodies cannot enter living muscle cells. This study aims to investigate the validity of this assumption.

Methods: Confocal immunofluorescence microscopy was employed to localise antibodies and other proteins of interest in myositis muscle biopsies. Bulk RNA sequencing was used to examine the transcriptomic profiles of 669 samples, including those from patients with myositis, disease controls and healthy controls. Additionally, antibodies from myositis patients were introduced into cultured myoblasts through electroporation, and their transcriptomic profiles were analysed using RNA sequencing.

Results: In patients with myositis autoantibodies, antibodies accumulated inside myofibres in the same subcellular compartment as the autoantigen. Bulk RNA sequencing revealed that muscle biopsies from patients with autoantibodies targeting transcriptional regulators exhibited transcriptomic patterns consistent with dysfunction of the autoantigen. For instance, in muscle biopsies from patients with anti-PM/Scl autoantibodies recognising components of the nuclear RNA exosome complex, an accumulation of divergent transcripts and long non-coding RNAs was observed; these RNA forms are typically degraded by the nuclear RNA exosome complex. Introducing patient antibodies into cultured muscle cells recapitulated the transcriptomic effects observed in human disease. Further supporting evidence suggested that myositis autoantibodies recognising other autoantigens may also disrupt the function of their targets.

Conclusions: This study demonstrates that, in myositis, autoantibodies are internalised into living cells, causing biological effects consistent with the disrupted function of their autoantigen.

Keywords: Antibodies; Autoantibodies; Dermatomyositis; Polymyositis.

PubMed Disclaimer

Conflict of interest statement

Competing interests: None declared.

Figures

Figure 1.
Figure 1.. Immunoglobulin localization in myositis muscle.
Confocal immunofluorescence of human immunoglobulin (IgG) in different autoantibody-defined types of myositis shows antibody deposition in the nuclei of muscle fibers from anti-Mi2-positive patients (A-C); the nucleoli of anti-PM/Scl-positive patients (D-F) the cytoplasm of anti-MDA5-positive patients (G-I); and the cytoplasm of anti-HMGCR-positive patients (J-L). The square boxes contain higher magnification images of one nucleus.
Figure 2.
Figure 2.. Expression of the anti-Mi2-specific gene set in different types of myositis.
Top 12 specifically overexpressed genes in muscle biopsies from anti-Mi2-positive patients compared to all the other groups included in the study. Each dot represents the gene expression value from a single patient. NT: histologically normal muscle biopsies; DM: dermatomyositis; AS: antisynthetase syndrome; IBM: inclusion body myositis; IM: inflammatory myopathies.
Figure 3.
Figure 3.. Correlation of autoantibody-specific gene sets with markers of disease activity.
The heat maps depict correlations of the anti-Mi2-specific gene set (A), the anti-PM/Scl-specific gene set (B), and the antisynthetase-specific gene set (C) with transcriptomic markers of type 1 interferon-inducible genes (ISG15, MX1), type 2 interferon-inducible genes (GBP2, IFI30), T-cell markers (CD3E, CD4, CD8), macrophage markers (CD14, CD68), markers of muscle differentiation (NCAM1, MYOG, MYH3, MYH8), and structural mature muscle proteins (ACTA1, MYH1, MYH2).
Figure 4.
Figure 4.. The expression of anti-Mi2-specific genes in cultured muscle cells electroporated with antibodies.
Standardized expression levels (Z-scores) of the more than 100 anti-Mi2-specific genes in human skeletal muscle cells electroporated with purified antibodies from different myositis patients (each column corresponds to a different serum from a different patient). Only cells electroporated with antibodies from anti-Mi2-positive patients have strong expression of anti-Mi2-specific genes.
Figure 5.
Figure 5.. Expression of the anti-PM/Scl-specific gene set in different types of myositis.
Top 12 specifically overexpressed genes in muscle biopsies from in muscle biopsies from anti-PM/Scl-positive patients compared to all the other groups included in the study. NT: histologically normal muscle biopsies; DM: dermatomyositis; AS: antisynthetase syndrome; IBM: inclusion body myositis; IM: inflammatory myopathies.
Figure 6.
Figure 6.. The expression of anti-PM/Scl-specific genes in cultured muscle cells electroporated with antibodies.
Standardized expression levels (Z-scores) of the more than 200 anti-PM/Scl-specific genes in human skeletal muscle cells electroporated with purified antibodies from different myositis patients (each column corresponds to a different serum from a different patient). Only cells electroporated with antibodies from anti-PM/Scl-positive patients have strong expression of anti-PM/Scl-specific genes.
Figure 7.
Figure 7.. A model of pathogenic myositis autoantibody internalization.
In myositis, autoantibodies are internalized into the muscle fibers, disrupting the normal biological function of their autoantigen, which mediates the pathogenesis of the disease (A). For instance, anti-Mi2 autoantibodies (B) interfere with the Mi2/NuRD complex inducing the derepression of more than 100 genes. Similarly, anti-PM/Scl autoantibodies (C) cause a dysfunction of the nuclear RNA exosome complex, impairing the normal degradation of various types of RNA.
See this image and copyright information in PMC

Update of

  • Pathogenic autoantibody internalization in myositis.
    Pinal-Fernandez I, Muñoz-Braceras S, Casal-Dominguez M, Pak K, Torres-Ruiz J, Musai J, Dell'Orso S, Naz F, Islam S, Gutierrez-Cruz G, Cano MD, Matas-Garcia A, Padrosa J, Tobías-Baraja E, Garrabou G, Aldecoa I, Espinosa G, Simeon-Aznar CP, Guillen-Del-Castillo A, Gil-Vila A, Trallero-Araguas E, Christopher-Stine L, Lloyd TE, Liewluck T, Naddaf E, Stenzel W, Greenberg SA, Grau JM, Selva-O'Callaghan A, Milisenda JC, Mammen AL. Pinal-Fernandez I, et al. medRxiv [Preprint]. 2024 Jan 17:2024.01.15.24301339. doi: 10.1101/2024.01.15.24301339. medRxiv. 2024. Update in: Ann Rheum Dis. 2024 Oct 21;83(11):1549-1560. doi: 10.1136/ard-2024-225773. PMID: 38313303 Free PMC article. Updated. Preprint.

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