Introduction

Kawasaki disease (KD)1 is an acute, febrile systemic vasculitis that was first described by Kawasaki et al2. In developed countries, it is the leading cause of acquired heart diseases in children, though its etiology remains unknown3, 4, 5. KD occurs worldwide, most commonly in Asian countries, and mainly affects children less than 5 years of age. Japan, Korea, and Taiwan region have the highest incidence of KD ranging from 69 to 213 cases per 100 000 children under 5 years of age6, 7, 8. The incidence of KD has been increasing globally in recent years. The most serious complications of KD are coronary artery lesions (CAL), including myocardial infarction, coronary artery fistula formation9, coronary artery dilatation/ectasia and coronary artery aneurysm10.

The clinical characteristics of KD patients include prolonged fever longer than five days, diffuse mucosal inflammation, bilateral non-purulent conjunctivitis, dysmorphous skin rashes, indurative angioedema over the hands and feet, and cervical lymphadenopathy. In addition to the diagnostic criteria, there is a broad range of non-specific clinical features, including irritability, uveitis, aseptic meningitis, cough, vomiting, diarrhea, abdominal pain, gallbladder hydrops, urethritis, arthralgia, arthritis, hypoalbuminemia5, liver function impairment and heart failure4, 11.

Although the clinical features of KD are recognizable, the immuno-pathogenetic mechanism of this disease is still unclear, particularly the causative agent for CAL formation. Transforming growth factor-beta (TGF-β) is a candidate gene for KD pathogenesis because TGF-β-mediated T-cell activation and cardiovascular remodeling are regarded as important features of KD. Indeed, genetic polymorphisms of the TGF-β pathway, including TGFB2, TGFBR2, and SMAD3, are associated with susceptibility to KD and development of CAL in the European and US populations12. In the Asian population, we reported that monocytosis, eosinophilia, and eosinophil-related Th2 immune response (especially, plasma level of IL-5) are associated with CAL formation and/or initial intravenous immunoglobulin (IVIG) treatment response13, 14, 15, 16. Immune-related genes, such as CTLA-4, CASP3 and ITPKC, have also been suggested to influence the susceptibility to and the clinical status of KD14, 15, 17, 18, 19, 20, 21.

The efficacy of IVIG administered during the acute phase of KD to reduce the prevalence of coronary artery abnormalities has been well established22. However, the mechanism of action of IVIG is still elusive. IVIG appears to have a generalized anti-inflammatory effect. Possible mechanisms of IVIG include the modulation of cytokine production, neutralization of bacterial super-antigens, suppression of antibody synthesis and inflammatory markers (CD40L, nitric oxide and iNOS expression)23, 24, and provision of anti-idiotypic antibodies9, 13, 16.

The role of genetic polymorphisms in immune-related genes in the susceptibility to Kawasaki disease

The higher incidence of KD in Asia, in conjunction with a higher incidence of the disease in Asian descendants compared with other ethnic populations in the United States and Europe, suggests that a genetic predisposition might play an important role in the susceptibility to this disease3, 4, 7, 10, 25, 26, 27. There is also evidence that the incidence of KD is higher among siblings than in the general population28. Additionally, KD has a higher incidence rate among males than in females5, 19. Further evidence supports the hypothesis that genetic factors contribute to the susceptibility to KD17. For example, a number of genes have been reported to have significant associations with the susceptibility to KD in different populations. For instance, single nucleotide polymorphisms (SNPs) in the monocyte chemoattractant protein 1 (MCP-1)29, IL-1030, 31, 32, CD40L33, IL-426, CASP320, 34, IL-1835, IL-1B36, HLA-E37, C-C chemokine receptor 5 (CCR5)38, and inositol 1, 4, 5-trisphosphate 3-kinase C (ITPKC)21, 39 have been reported to be associated with the development of KD. In early 2011, Shimizu et al12 first reported that genetic polymorphisms of TGFB2, TGFBR2, and SMAD3 are associated with susceptibility to Kawasaki disease and the development of coronary artery lesions. Taken together, these findings suggest that multiple polymorphic alleles influence KD susceptibility and that different ethnic populations, which have distinct allelic expression patterns, and different sexes may have different susceptibilities to KD3. Interestingly, there are some genes associated with susceptibility to KD, but not CAL formation. We hypothesize that the genes responsible for susceptibility and CAL formation may be distinct20, 39, 40.

Association between the genetic polymorphisms and CAL formation in KD

All KD patients were treated with IVIG, 2 g/kg in a single infusion for 12 h, together with aspirin22. This therapy was within 10 d of illness and, if possible, within 7 d of illness. From a serial analysis of coronary artery lesions (n=341) in Chang Gung Memorial Hospital-Kaohsiung20, 35% of KD patients had dilatation during the acute phase, 17.2% had dilatation one month after disease onset, 10.2% still had dilatation at two months of follow-up, and 4% had persistent CAL for more than one year39.

The most commonly used definition of CAL (also known as coronary artery abnormality, CAA or CALs) is based on the Japanese Ministry of Health criteria: maximum absolute internal diameter >3 mm in children younger than 5 years of age or >4 mm in children 5 years and older, or a segment 1.5 times larger than an adjacent segment, or the presence of luminal irregularity41, 42, 43. If the body surface area is known, then coronary arteries are normalized to this surface area and expressed as standard deviation units from the mean (Z scores)44. Several studies have analyzed CAL using other methods, including the aorta route dimension12 and transient CAL (although the definition of “transient” varies among studies, from 30 d to 6–8 weeks after disease). Interestingly, some results have indicated that the genetic association was observed only with susceptibility, not with CAL formation. However, other studies have revealed inconsistent results (Table 1)45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57. These results indicate that the genes responsible for susceptibility and CAL formation may be different between populations7, 32, 37, 38, 58, 59, 60. Recently, several candidate genes have been proposed for the susceptibility to KD or the formation of CAL in different populations. Although there is evidence to support a role for each candidate gene in the susceptibility to KD and/or development of CAL, there is also evidence that cannot be easily fitted into any (Table 2). Most studies addressing this question are plagued with inconsistencies. First, the sample size varies dramatically across studies. Hence, a small sample size may not have sufficient power to detect minor genetic effects. Second, it is becoming clear that there are different genetic backgrounds within populations that due to variations in allele frequencies or heterogeneity of the phenotype, may also influence the results. Third, the incidence of KD in Asia is much higher than in other places. Thus, the role of environmental factors or infectious agents in the development of KD should also be considered.

Table 1 Genes associated with susceptibility or coronary artery lesion (CAL) formation in KD.
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Table 2 Controversial genetic effects on the susceptibility to KD or CAL formation between populations.
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Genetic polymorphisms of the ITPKC signaling pathway in patients with Kawasaki disease

A major advancement in the genetic study of KD was provided by the discovery of ITPKC. ITPKC is an important molecule in the regulation of T cell activation, and it may function as a calcium channel modulator21. In 2008, Onouchi and colleagues first identified the functional polymorphism of ITPKC (rs28493229) that is significantly associated with the susceptibility to KD and coronary artery lesions in both Japanese and US children21, 61. Using cell-based functional studies, Onouchi et al further indicated that the risk allele (C allele) of ITPKC reduces the splicing efficiency of the ITPKC mRNA that, in turn, may contribute to the hyperactivation of Ca2+-dependent NFAT pathways in T cells21. The identification of ITPKC has had an enormous effect on the field of genetic association studies of KD. These novel insights into genetic mechanism clearly provide a new understanding of the pathogenesis of KD. In contrast, the results obtained from the replication studies in the Taiwanese populations are strikingly controversial62, 63. A similar approach was taken by Chi et al62. These authors genotyped 385 KD patients and 1158 normal subjects. However, there were no significant differences in the genotype of rs28493229 between the controls and children with KD. The results from a study by Lin et al63 in another independent medical center in Taipei indicated that the C allele of rs28493229 is associated with KD susceptibility. Recently, new results using meta-analysis showed that the rs28493229 SNP of ITPKC is associated with the susceptibility to KD in the Taiwanese population39. Regarding to the controversial results obtained for ITPKC genetic association studies in the Taiwanese population, we attribute them to population migration, due to the increase in genetic diversity between cities in the south or north of Taiwan39.

ITPKC is involved in the Ca2+-dependent NFAT signaling pathways in T cells21. In the non-excitable cells, such as T cells and mast cells, one of the main pathways to increase the intracellular Ca2+ concentration is through store-operated calcium channels (SOC)64. The activation of store-operated calcium channels can be controlled by the expression level of IP3. IP3 binds to its receptor (IP3R) on the endoplasmic reticulum (also called the calcium store) causing the release of calcium from stores64. An empty store results in the activation of both store-operated calcium channels and Ca2+-dependent signaling pathways, including inflammatory reactions65, 66 and apoptosis64. Onouchi et al reported that a G to A substitution in the 5′-untranslated region of CASP3 (rs72689236) is associated with susceptibility to Kawasaki disease in Japanese and in Americans of European descent34. The CASP3 (rs72689236) is replicated in the KD patients in the Taiwanese population. Kuo et al provided further evidence that the A allele of rs72689236 is very likely to be a risk allele in the development of aneurysms in patients with KD20.

Another important molecule in the ITPKC signaling pathway is ORAI1 (also known as CRACM1). ORAI1 was identified by Feske et al in 200667. Modified linkage analysis completed on data generated by single-nucleotide polymorphism arrays and an RNA interference screen led to an important finding. A single missense mutation in ORAI1 was found in patients with severe combined immune deficiency (SCID) syndrome67. Furthermore, this mutation in ORAI1 has been shown to cause dysfunctions in Ca2+ release-activated Ca2+ (CRAC) channels and impairs the immune system67. In 2011, a genetic polymorphism of ORAI1 was reported to be associated with the risk and recurrence of calcium nephrolithiasis68. In the study of Kawasaki disease, no significant association between genotype and allele frequency of the five ORAI1 tSNPs was found. Additionally, there is no association between ORAI1 polymorphisms and CAL formation or IVIG treatment responses69. However, this lack of association does not rule out the possibility that other genes in the ITPKC signaling pathways might contribute to the susceptibility or clinical status (CAL or IVIG treatment responses) of KD (Figure 1). For example, a mutation in STIM1, a key molecule in the regulation of SOC, has also been reported to be associated with an immunodeficiency syndrome70. Therefore, identification of genetic polymorphisms for STIM1 and other genes related to the ITPKC pathway by direct DNA sequencing in a larger population may be helpful to better understand the pathogenesis of KD.

Figure 1
figure 1

Model depicting the cellular pathways of ITPKC in T cells.

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Genome-wide association study (GWAS) in Kawasaki disease

In 2009, Burgner et al firstly performed a genome-wide association study (GWAS) on 119 Caucasian KD cases and 135 matched controls. Forty SNPs and six haplotypes were confirmed in an independent cohort of KD families71. This insightful work led to the identification of a SNP within the N-acetylated alpha-linked acidic dipeptidase-like 2 gene (NAALADL2; rs17531088), which was significantly associated with the susceptibility to KD. Although the function of NAALADL2 remains unclear, mutations in the gene may be involved in the development of Cornelia de Lange syndrome72. In 2010, another GWAS was conducted by Kim et al in a Korean population. A total of 786 subjects (186 KD patients and 600 controls) were recruited. A locus in the 1p31 region was identified as a susceptibility locus for KD. Furthermore, the PELI1 gene locus in the 2p13.3 region was confirmed to associate with the development of CAL in KD patients73. GWAS results from a Taiwanese population suggest another three novel susceptibility loci for KD74. However, the susceptibility loci reported by Kim et al in the Korean population cannot be replicated in a Taiwanese or Caucasian population. Hence, the results of the GWAS from independent groups support the hypothesis that susceptibility loci for KD and CAL formation can be distinct between different ethnic populations.

Conclusion

Several major advances have been made in understanding the genetic effects of the susceptibility and clinical status of KD over the past decade. Very recently, genome-wide association has led two groups to identify novel susceptibility loci as being important for KD in the Asian population. Although the exact genes in the loci are still unclear at present, these loci could provide a new direction for future studies. Now that the sequencing of the human genome is complete and advanced genotyping tools are readily available to help identify candidate genes, we can expect to see more insightful researches begin to elucidate the genes responsible for KD susceptibility.