Detection of Catalase as a major protein target of the lipid peroxidation product 4-HNE and the lack of its genetic association as a risk factor in SLE

Abstract

BackgroundSystemic lupus erythematosus (SLE) is a multifactorial disorder characterized by the presence of autoantibodies. We and others have implicated free radical mediated peroxidative damage in the pathogenesis of SLE. Since harmful free radical products are formed during this oxidative process, including 4-hydroxy 2-nonenol (4-HNE) and malondialdehyde (MDA), we hypothesized that specific HNE-protein adducts would be present in SLE red blood cell (RBC) membranes. Catalase is located on chromosome 11p13 where linkage analysis has revealed a marker in the same region of the genome among families with thrombocytopenia, a clinical manifestation associated with severe lupus in SLE affected pedigrees. Moreover, SLE afflicts African-Americans three times more frequently than their European-American counterparts. Hence we investigated the effects of a genetic polymorphism of catalase on risk and severity of SLE in 48 pedigrees with African American ancestry.MethodsTryptic digestion followed by matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) analysis was used to identify the protein modified by HNE, following Coomassie staining to visualize the bands on the acrylamide gels. Genotyping analysis for the C → T, -262 bp polymorphism in the promoter region of catalase was performed by PCR-RFLP and direct PCR-sequencing. We used a "pedigree disequilibrium test" for the family based association analysis, implemented in the PDT program to analyze the genotyping results.ResultsWe found two proteins to be HNE-modified, migrating around 80 and 50 kD respectively. Tryptic digestion followed by matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) analysis of the Coomassie stained 80 kD band revealed that the target of HNE modification was catalase, a protein shown to associate with RBC membrane proteins. All the test statistics carried out on the genotyping analysis for the C → T, -262 bp polymorphism in the promoter region of catalase were non-significant (p > 0.05) in our data, which suggested that this SNP is not associated with SLE.ConclusionOur results indicate that catalase is one of the proteins modified due to oxidative stress. However, catalase may not be a susceptibility gene for SLE. Nonetheless, catalase is oxidatively modified among SLE patients. This suggests a possible role between oxidative modification of catalase and its affects on enzymatic activity in SLE. An oxidatively modified catalase could be one of the reasons for lower enzymatic activity among SLE subjects, which in turn could favor the accumulation of deleterious hydrogen peroxide. Furthermore, HNE-products are potential neoantigens and could be involved in the pathogenesis of SLE. Decrease in catalase activity could affect the oxidant-antioxidant balance. Chronic disturbance of this balance in patients with SLE may work favorably for the premature onset of atherogenesis with severe vascular effect.

Relationships between single nucleotide polymorphisms of antioxidant enzymes and disease

“…This silent mutation, rs769217, found in exon 9, changes T > C and leaves the amino acid at position 389 as aspartic acid (Asp). It has been shown to have no association with the prevalence of T1DM (Pask et al, 2006), susceptibility to systemic lupus erythematosus (SLE) (D'Souza et al, 2008) or non-pathological cognitive ageing (Harris et al, 2007). However, the Asp allele is associated with vitiligo in North Americans (Casp et al, 2002) and UK English residents (Gavalas et al, 2006) but not Chinese subjects (Liu et al, 2010) and there is no association between breast cancer and the rs769217 SNP and three other catalase SNPs, rs511895, rs7104301 and rs1049982 (Oestergaard et al, 2006).…”

Relationship Between Redox Status and Cell Fate in Immunity and Autoimmunity

“…Interestingly, ROS-modified IgG have been associated with the induction of circulating SLE autoantibodies where neoepitopes elicit an immune response in SLE patients (3). Moreover, the formation of MDA-and 4-HNE-modified proteins are associated with SLE (33,184). Several studies, both in animal models and in humans, suggest that the overexpression of inducible NO synthase (iNOS) may contribute to pathologic processes of SLE by altering multiple signaling pathways in T cells and leading to glomerular and vascular manifestations (120,125,186).…”

Significantly reduced lymphadenopathy, salivary gland infiltrates and proteinuria in MRL-lpr/lprmice treated with ultrasoluble curcumin/turmeric: increased survival with curcumin treatment

“…Our studies showing significantly increased oxidative damage in lupus patients [19][20][21] led us to investigate an antioxidant/anti-inflammatory agent known as curcumin, [22][23][24][25][26][27][28][29][30][31] obtained from the rhizome of Curcuma longa (turmeric). Curcumin (1,7-bis[4-hydroxy-3-methoxyphenyl]-1,6-heptadiene-3,5-dione), a naturally occurring 'nutraceutical', comprises 4-6% of turmeric.…”