DNA methylation of ERAP1 gene
In total, two CpG islands containing 31 loci were tested the proportion
of methylated cytokine accounting for the total cytokine. Among the 15
loci of ERAP1_1 island and 16 loci of ERAP2 island, 13 CpG sites,
ERAP1_1_46, ERAP1_1_91, ERAP1_1_107, ERAP1_1_118, ERAP1_1_222,
ERAP1_2_28,
ERAP1_2_31, ERAP1_2_46, ERAP1_2_49, ERAP1_2_69, ERAP1_2_95,
ERAP1_2_121 and ERAP1_2_136, were significant hypermethylated in AS
patients compared to HCs used Mann-Whitney U test detailed in Table 2.
Logistic regression also indicated that methylation of six loci in
ERAP1_2 island, ERAP1_2_28, ERAP1_2_31, ERAP1_2_49, ERAP1_2_69,
ERAP1_2_121 and ERAP1_2_136, were significantly associated with
increased risk of AS visualized in Figure 1A. Among them, ERAP1_2_28
located at chr5: 96143595 was reported to have highest effect size with
OR = 5.390 (95% CI: 2.048 to 14.184) of as a percent of total cytokine
methylated. The average methylation levels of ERAP1_1
(Z= -4.831, P < 0.001) and ERAP1_2 (Z = -6.140,P < 0.001) islands were also calculated to associated
with AS (Figure 1B & 1C). ROC curve analysis on the average methylation
levels of the two CpG islands also indicated their potential as
biomarkers of AS. The AUC of ROC curve for ERAP1_1 island was 0.669
(95% CI: 0.626 to 0.707, P < 0.001), with a
sensitivity of 0.818 and a specificity of 0.531 for the best cutoff
point 0.671 (Figure 1D, Table 3). Correspondingly, the AUC of ERAP1_2
was 0.750 (95% CI: 0.683 to 0.818,P < 0.001), and
the
best cutoff point was 0.681, with a sensitivity of 0.707 and specificity
of 0.684 (Figure 1D, Table 3). We
also established a regression model included the above mentioned 13
significant CpG loci detailed in Table S1, with each sample obtained a
probability value. ROC curve analysis on the probability value also
reported that the AUC was 0.779 (95% CI: 0.714 to 0.844, P< 0.001), and best cutoff point was 0.500, with a sensitivity
of 0.717 and specificity of 0.737 (Figure 1D, Table 3).