Association of the programmed cell death 1 (PDCD1) gene polymorphism with ankylosing spondylitis in the Korean population
© Lee et al.; licensee BioMed Central Ltd. 2006
Received: 15 March 2006
Accepted: 25 October 2006
Published: 25 October 2006
The PD-1 (programmed death 1) molecule is a negative regulator of T cells. PDCD1 (programmed cell death 1) has been reported to have a genetic association in systemic lupus erythematosus and rheumatoid arthritis in Caucasians. However, there are no reports on the association between this gene and ankylosing spondylitis (AS). The present study investigated the association of the PD-1 polymorphisms and the haplotypes with AS in a Korean population sample. In a case-control association study, two single-nucleotide polymorphisms, PD-1.5 C/T and PD-1.9 T/C, were genotyped in 95 AS patients and 130 healthy controls. The T allele of the PD-1.9 polymorphism was more frequent in the Korean male population with AS than in the Korean male controls (21.0% versus 6.9%, odds ratio 1.89, 95% confidence interval 1.483 to 2.408). The frequency of the CT haplotype (PD-1.5 C/T and PD-1.9 T/C) was higher in the AS patients (19%) than the controls (5.4%) (odds ratio 1.83, 95% confidence interval 1.559 to 2.521). The PD-1 polymorphism was demonstrated in Korean AS patients. The results suggest a genetic association between the PD-1 polymorphism and susceptibility to AS.
Ankylosing spondylitis (AS) is an important chronic inflammatory disease with an incidence range of between 0.5% and 1.0% [1, 2]. Many genetic and environmental factors have been suggested to have some role in the development of AS . However, the pathogenesis of AS is still unclear. Human leukocyte antigen (HLA) B27 makes up only a small proportion of the overall risk for spondyloarthritis. Fewer than 5% of HLA-B27-positive people in the general population develop these diseases . In contrast, 20% of the HLA-B27-positive relatives of AS patients are affected. Family studies have suggested that HLA-B27 contributes to about 37% of the overall genetic risk for spondyloarthritis [5–7], which suggests the involvement of other genes in the development of AS. Another new susceptible gene therefore needs to be identified.
Recently, the PDCD1 (programmed cell death 1) gene polymorphism was reported to be associated with systemic lupus erythematosus (SLE) [8, 9], lupus nephritis [10, 11] and seronegative rheumatoid arthritis (RA) . It was also suggested that lupus nephritis and the seronegative RA susceptibility was associated with the PD-1.3 (position 7,146) A allele in a northern Sweden population.
The PD-1 molecule is a negative regulator of T cells  that belongs to the immunoglobulin receptor superfamily. It encodes a 55 kDa type 1 transmembrane inhibitory immunoreceptor and is responsible for the negative regulation of T-cell activation and peripheral tolerance . Nishimura and Honjo  reported that PD-1 expression was observed only in activated T and B cells as well as in the early lymphoid precursors. PD-1 is actively expressed on the cell surface during the activation of T and B cells. The cytoplasmic immunoreceptor tyrosine-based inhibitory motif of PD-1 was activated by an interaction between PD-1 and its corresponding ligands, PDL-1 (B7-H1) and PDL-2 (B7-DC) [15, 16], which induces the inhibitory signal to inhibit the proliferation of T and B cells to maintain peripheral tolerance [14–16].
The human gene encoding PD-1, PDCD1, is localized on 2q37.3, which is a susceptibility locus for SLE . Allele T of a single-nucleotide polymorphism (SNP) corresponding to PD-1.5 C/T (dbSNP rs#cluster id rs2227981) was found to be associated with the development of RA (odds ratio (OR) 1.94, 95% confidence interval (CI) 1.25 to 3.01, p < 0.0025) but not SLE in Chinese patients living in Taiwan .
So far more than 30 SNPs have been identified. Among the 30 SNPs, 7 (namely PD-1.1, PD-1.2, PD-1.3, PD-1.4, PD-1.5, PD-1.6 and PD-1.9) were examined in SLE . Two SNPs (PD-1.5 C/T (rs2227981) and PD-1.9 T/C/(rs2227982)) were selected because they occur in an exon, which affects protein synthesis. The change in PD-1.9 T/C causes a change in their synthesized amino acid from valine to alanine. Our study was therefore focused on determining whether PD-1.9 T/C SNP affects the development of AS.
Overall, PDCD1 is a strong candidate for susceptibility to autoimmune disease. This study therefore investigated whether or not PDCD1 is associated with AS in the Korean population.
Materials and methods
Patients and control subjects
Case-control genetic association studies were performed to test the association between PDCD1 and the development of AS. The patient group consisted of 95 Korean patients with AS who were recruited from the Kyunghee University Medical Center, Seoul, Korea. A diagnosis of AS was established by using the classification criteria reported by the American College of Rheumatology (Modified New York Criteria). At least two rheumatologists confirmed the diagnosis. The patients were diagnosed before March 2005 and samples of whole blood was extracted between March 2003 and February 2005. The other control group was made up of 130 normal individuals who underwent a health examination in Kyunghee University Medical Center. They had no previous medical history and no abnormal laboratory results. All subjects provided written informed consent. All medical records in both groups were reviewed and analysed according to age, gender, combined disease, complications and disease duration.
This study was approved by the ethics review committee of the Medical Research Institute at Kyung Hee University Medical Center, Seoul, Korea in accordance with the World Medical Association Declaration of Helsinki: Ethical Principles for Medical Research Involving Human Subjects.
Genotyping of PD-1.5 and 1.9
PCR-RFLP methods, PCR primers and restriction enzymes (RE).
The SPSS statistical package (version 10.0) was used for statistical analysis. The data are expressed as means ± SD. The baseline differences between the groups were tested with Student's t test and a χ2 test. All the SNP analyses, such as Hardy–Weinberg equilibrium test, linkage disequlibrium and haplotype inference, were performed with SNP Alyze (version 5.1; DYNACOM). Hardy–Weinberg equilibrium, which indicates the absence of any discrepancies between the genotype and allele frequencies, was examined and no discrepancies were noted. SNP Alyze was used to examine the allele frequency and haplotype. There was no linkage disequilibrium or haplotype inference. The ORs and 95% CIs of the allele frequencies and haplotypes were calculated in the control and disease groups. The allele frequencies were compared by using a χ2 test with a 2 × 2 contingency table. A χ2 test was also used to compare the haplotype frequencies. p ≤ 0.05 was considered significant.
Basal characteristics of patients with ankylosing spondylitis and of normal controls
Basal characteristics and clinical features of patients with AS and of normal controls.
Sex, M:F (%)
Age (mean ± SD)
33.4 ± 9.4
66.3 ± 5.8
Disease subgroup, n
Age on initial symptom, years (mean ± SD)
23.1 ± 7.6
Disease duration, months (mean ± SD)
25.9 ± 8.5
Delayed diagnosis, months (mean ± SD)
32.1 ± 38.2
HLA B27+ (n = 84) (%)
Complications, n (%)
Peripheral joint involvement
Total hip replacement
Heart conduction abnormality
Cauda equina syndrome
PD-1.5 SNP in patients with ankylosing spondylitis and in normal controls
Genotyping and allele frequency and haplotype of PD-1.5 and PD-1.9 SNP in patients and controls.
OR (95% CI)
PD-1.9 SNP in patients with ankylosing spondylitis and normal controls
A total of 225 subjects (95 patients and 130 controls) were genotyped for the PD-1.9 T/T SNPs. Among the 95 AS patients, 2.5%, 37.5% and 60.5% had the T/T, C/T and C/C genotypes, respectively; among the 130 normal controls the proportions were 2.3%, 9.2% and 88.5%, respectively. There was a significant difference between the groups in C/T versus C/C but no significant difference in C/C versus T/T. Among the 95 AS patients, the allele frequencies of T and C were 21.0% and 79.0%, respectively, and 6.9% and 93.1% among the controls. There was a statistically significant difference between the groups, and the 95% CI 1.483 to 2.408, OR was 1.89 according to the χ2 test (Table 3).
Among the 95 AS patients, 19.1%, 47.5%, 1.9% and 31.5% had the CT (PD-1.5 C/T and PD-1.9 T/C), CC, TT and TC haplotype, respectively; among the 130 normal controls the proportions were 5.4%, 55.4%, 1.5% and 37.7%, respectively. The frequency of the CT haplotype was higher in the AS patients than the controls (OR 1.83; 95% CI 1.559 to 2.521; Table 3).
The results show that a functionally important polymorphism in PDCD1 is associated with a subset of AS. A polymorphism in this gene has previously been shown to be associated with SLE [8, 9] and RA . These results suggest that this gene may commonly have some role in the development of autoimmune disease. PDCD1 has a key role in the immune response as follows.
The ligands for PD-1 have been identified as PD-L1 (B7-H1), which is expressed in all haemopoietic cells as well as in many non-haemopoietic tissues, and PD-L2 (B7-DC), which is expressed primarily on dendritic cells and macrophages [14, 16, 18, 19]. This molecule has a role in the negative co-stimulated pathway involving the CD28 homologue PD-1 receptor . PD-1 is expressed by activated, but not unstimulated, T cells, B cells and myeloid cells, which is in contrast with the predominantly T-cell-restricted expression of CD28 and CTLA-4 [21–23]. PD-1-/- mice display a variety of autoimmune pathologies, including dilated cardiomyopathy (in BALB/c mice) and a lupus-like syndrome (in B6 mice). This indicates that PD-1 has a key role in the maintenance of peripheral tolerance to self-antigens, which is analogous to that of CTLA-4 [13, 24]. The development of AS can therefore be affected by PD-1 expression or the function of the PD-1 molecule, and is associated with the continuous activation of immune cells such as CD4+ T cells or dendritic cells. HLA B27, which is the most important genetic factor for AS, is a class I antigen and reacts with CD 8+ T cells. However, most autoimmune diseases are associated with CD4+ T cells. Recently, misfolding HLA B27 was reported to stimulate CD4+ T cells or natural killer cells in HLA B27 transgenic rats . In contrast, it was suggested that HLA B27 itself also could act as an autoantigen, because it is recognized by the T-cell receptors on CD4+ T cells  or by its presentation by HLA class II molecules. This suggests the importance of the CD4+ T-cell activation pathway and termination of its activation in AS. It is therefore possible that AS is associated with the function of the PD-1 molecule.
Differences in PD-1.5C allele frequencies between Korean female control population and other female populations.
Differences in PD-1.5 genotype and alleles between Korean and Chinese populations.
Korean controls (%)
Chinese controls (%)
Korean controls (%)
Chinese controls (%)
The limitation of our study was that the control group was mismatched to the disease group in sex and age. With regard to sex, the polymorphism of this gene is not affected by the sex difference in our control group. With regard to age, with increasing age in the control group the possibility of being in the disease-free group increased because AS develops at a relatively young age.
The PD-1.9 T allele was found to be a susceptibility factor for AS; however, 1.5 SNP was not a factor. The PD-1.9 T allele substitutes valine for alanine 215 during PD-1 protein synthesis. However, other subgroup analyses according to gender, combined uveitis, the involvement of a peripheral joint and the existence of HLA B27 showed no significant differences. Therefore, further study will be needed to determine whether this amino acid change alters the protein structure and affects its function.
PDCD1 was known as another genetic risk factor in RA or SLE patients. However, there was no known study on this gene with regard to AS. We have examined the association between this autoimmune-associated gene and AS and concluded that the gene is associated with susceptibility to AS, although only a small sample was used. We believe that these results are sufficient to suggest another genetic susceptibility (PDCD1) for AS.
= ankylosing spondylitis
= human leukocyte antigen
= odds ratio
= rheumatoid arthritis
= restriction-fragment-length polymorphism
= systemic lupus erythematosus
= single-nucleotide polymorphism.
This study was supported by an Oriental Medicine Research Center for Bone and Joint Disease grant from the Ministry of Health and Welfare of the Republic of Korea (03-PJ9-PG6-SO01-0002).
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