Contribution of toll-like receptor 9 gene single-nucleotide polymorphism to systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is a progressive and chronic autoimmune disorder in which the immune system cannot distinguish between the body’s own tissues and foreign antigens, producing antibodies (Ab) to self-antigens [1]. Flare-ups of SLE can be initiated by disparate environmental factors, such as exposure to ultraviolet light, drugs, chemicals, as well as viral and bacterial infections [2, 3]. The underlying cause of SLE is elusive; however, it is well established that both environmental and genetic factors are involved in this disease [3‐7]. Genome-wide association studies have revealed many SLE prone genes and the contribution of some of these genes to the risk of SLE have been studied in different ethnicities [7]. One of these genes is the toll-like receptor 9 (TLR9) gene located in susceptibility regions for SLE [7].

TLR9 plays an elementary role in pathogen recognition and activation of innate immunity [8‐10]. This receptor recognizes unmethylated cytosine–phosphate–guanine (CpG) dinucleotide motifs located in bacterial, viral and fungal DNA [8‐12]. The TLR9 gene is expressed in macrophages and dendritic cells, and TLR9 has been shown to be present almost exclusively in endosomes [8, 13‐15].

It has been demonstrated that TLR9 is involved in the development of autoimmunity in SLE patients [16‐18]. Stimulation of TLR9 in the endosomes by host DNA leads to plasmacytoid dendritic cell activation and type I interferon biosynthesis, which is implicated in lupus pathophysiology [19]. TLR9 activation also promotes the production of IgG2a and IgG2b autoantibodies recognizing host DNA, which further develop autoimmunity pathology [20]. The role of TLR9 in autoimmunity was also demonstrated by Leadbetter et al. [21], who indicated that DNA-containing complexes interacting with TLR9 may activate both autoreactive B cells and other antigen presenting cells.

There are several studies on the contribution of TLR9 polymorphisms to the risk of SLE in different ethnicities; however, the results are inconsistent [22‐27]. The four TLR9 single-nucleotide polymorphisms rs187084, rs5743836, rs352139 and rs352140 in Caucasians are located in the same block of linkage disequilibrium (LD) HapMap CEU data (http://​hapmap.​ncbi.​nlm.​nih.​gov/​). Therefore, we aimed to study whether TLR9 C > T (rs352140) can be a genetic risk factor of SLE in the Polish population. Because SLE is a heterogeneous disorder, we also evaluated the contribution of this polymorphism to different clinical symptoms of SLE.

Some studies have demonstrated increased TLR9 expression in B cells from patients with SLE [30, 31]. Papadimitraki et al. [30] reported an increased proportion of peripheral blood memory B cells and plasma cells expressing TLR9, which correlated with the presence of anti-dsDNA Ab in patients with active SLE. Another study revealed that the level of TLR9 mRNA in B cells was increased in SLE patients, and TLR9 expression on CD20⁺ B cells correlated with SLE activity and CH50 [31]. This may suggest that genetic variations of TLR9 that affect their expression may have an effect on SLE development and clinical manifestation of this disease.

We did not observe a contribution of the TLR9 C > T (rs352140) polymorphism to the risk of SLE in a Polish population. Recent studies conducted by Huang et al. (2011) suggested that the TLR9 −1486 T/C (rs187084) polymorphism, located in the LD block with rs352140, is related to SLE in Taiwanese patients. [22]. In addition to this finding, Xu et al. [23] demonstrated a significant association of rs352140 gene variants with the susceptibility to SLE in a Chinese population. Additionally, the TLR9 G allele at position +1174 of TLR9 (rs352139) conferred an increased risk for SLE in a Japanese population [24]. The contributions of rs5743836 to SLE have been observed in individuals of European descent from Southern Brazil; however, this was not confirmed in Caucasian American individuals [25, 32]. TLR9 polymorphisms were not significantly associated with the susceptibility to SLE and related phenotypes in Korean patients with SLE [26]. Furthermore, Zhou et al. [27] did not find a significant contribution of the rs352140 polymorphism to SLE development in a Chinese Han population.

In our study, the TRL9 T/T and T/C genotypes exhibited a significantly increased risk of developing renal disease in patients with SLE. The significant association between the rs352140 gene variant and lupus nephritis was also observed in a Chinese Han population [27]. We also observed that the TRL9 T/T and T/C genotypes were significantly associated with immunologic disorders and the presence of anti-dsDNA Ab in patients with SLE. To date, an increased frequency of the TRL9 rs5743836 C allele has been observed in patients of European descent from Southern Brazil bearing the Anti-SSa/Ro Ab [25].

The function of +1174 G/A of TLR9 rs187084 located in the LD block with rs352140 has been studied by Tao et al. [24], who demonstrated that the +1174 G variant may down-regulate TLR9 expression. Moreover, they indicated that TLR9 rs187084 may contribute to differences in the B cell response to autoantigens and the production of autoantibodies [24].

Recently, the role of TLR9 expression in the production of anti-dsDNA Ab was confirmed by Chen et al. [33], who performed knockdown of TLR9 by siRNA in B cells, resulting in a reduction of anti-dsDNA Ab levels and amelioration of the disease in SLE murine model. Recent studies also suggest that the changes in TLR9 expression may have an effect on renal disease development in SLE [34, 35]. Machida et al. [34] demonstrated that injured podocytes express TLR9 in active lupus nephrites accompanied by proteinuria and elevated anti-dsDNA Ab. In addition to this finding, Anders et al. [35] revealed that activation of TLR9 by CpG oligonucleotides in MRL-Fas (lpr) mice induces anti-dsDNA Ab production and renal disease.