Background/Purpose:

Juvenile Dermatomyositis (JDM) is a systemic and inflammatory vasculopathy that affects mainly proximal muscles and skin in children.  It is a rare but severe autoimmune disease featuring the presence of autoantibodies against the translational machinery for protein synthesis. Proteolytic products of antibody-mediated complement activation are often observed in affected tissues of JDM.  The causal factors for JDM are unknown, but genetic variants of the HLA are strongly implicated, particularly in haplotypes with HLA-DR3.  Recently, differential copy number variations (CNVs) for the HLA-associated complement C4A and C4B genes were demonstrated in healthy subjects and autoimmune disease patients. A deficiency of C4A is strongly associated with HLA-DR3 and systemic autoimmunity.  Our objective is to investigate CNVs of total C4, C4A, and C4B on the pathogenesis of JDM.

Methods:

The study population included 56 JDM patients (53 Whites, 3 Blacks) and 577 geographically matched healthy subjects, recruited upon informed consents according to IRB-approved protocols. The mean (±SD) age for JDM disease diagnosis was 7.91±4.27 years old.  Calcinosis, lipodystrophy, and skin ulceration were observed in 19.6%, 9.1% and 16.1% of patients, respectively.  Genomic DNA, PAXGENE RNA, and EDTA-plasma were isolated from peripheral blood samples. Gene copy numbers (GCN) for total C4, C4A, and C4B were determined by genomic Southern blot and TaqMan-based, realtime qPCR.  Global gene expression array were determined using PAXGENE RNA and Agilent SurePrint G3 8x60K microarrays.  The expression of IFI44 was determined by SYBR-Green based qPCR using PAXGENE RNA and GADPH as standard.

Results:

Among White subjects, the GCNs vary from 2 to 6 for total C4, 0 to 5 for C4A, and 0 to 4 for C4B.  There were significant decreases in JDM (vs. controls) for GCNs of total C4 (cases: 3.42±0.72, controls: 3.85±0.69; p=0.00007, t-test) and C4A (cases: 1.74±0.86, controls: 2.10±0.76; p=0.0009, t-test).  Remarkably, 43.4% of the JDM patients had a heterozygous or homozygous deficiency of C4A, compared to 18.5% in controls (p=0.00008, Χ²analysis). The odds ratio (95% CI) for C4A deficiency in JDM was 3.37 (1.88- 6.04).  Intra-group comparison of clinical presentations revealed that patients with calcinosis and lipodystrophy both had significantly higher GCN of C4B (p=0.001, Fisher’s exact), reflecting a role of C4B in complement-mediated tissue injuries. Global gene expression studies using peripheral blood RNA (19 JDM, 8 controls) and Agilent microarrays revealed up-regulation of genes typical for interferon-α response signature and down-regulation for chemokine genes. SYBR-green labeled qPCR on transcripts for IFN-α response gene IFI44 showed a 4.7 and 2.6 -fold increase in expression among patients with and without a C4A deficiency, respectively, vs. controls (p=0.004, t-test).

Conclusion:

A deficiency of C4A is a common genetic risk factor for JDM with medium to high effect size. Peripheral blood RNA from JDM exhibited upregulation of IFN-α response gene IFI44, particularly among C4A deficient patients. Among established JDM patients, normal/high GCN of C4B was associated with disease complications, including calcinosis and lipodystrophy.

ACR Meeting Abstracts - https://acrabstracts.org/abstract/copy-number-variations-of-complement-c4a-and-c4b-genes-are-genetic-risk-factor-and-disease-modification-factor-respectively-for-juvenile-dermatomyositis/