Polymorphism Of Human Leukocyte Antigen-DQB1 Genes In Patients With Vogt-Koyanagi-Harada Syndrome

Vogt—Koyanagi—Harada syndrome(VKH) is featured as bilateral, diffuse, granulomatous panuveitis often associated with alopecia, vitiligo, poliosis, tinnitus dysacusis and meningeal symptoms. Pigmented populations, such as Mongolian ancestry, Aficans and some Mestizo were often affected for the auto- immunological mechanisms against the pigment cells in the skin, meninges, inner ear and ocular tissues. The Major Histocompatibility Complex (MHC) or Human Leukocyte Antigen (HLA) plays an important role on its course. Polymorphisms of HLA in patients with VKH have been reported especially in the past 10 years. HLA—DR4 by serology, HLA—DRBI~*0405, DQBI~*0401 by PCR—SSO or RFLP were reported to be associated with VKH in various degree in Japanese, Chinese, Mexico and American Mestizo, Korea, ect. However no one focuses on the upper regulatory regions(URRs) or promoter regions(QBP) of HLA- DQB molecules and combinatory analysis between coding region and QBPs for VKH syndrome. In our study 88 North Chinese Han patients with VKH matched with 88 healthy controls were admitted. Polymorphism of coding region and upper regulatory region for HLA—DQB1 were investigated by PCR- SSP and PCR- SSCP respectively. Relationships among HLA--DQB1 alleles and clinical manifestations, association among coding regions and promoter of HLA--DQB1 were analyzed.12 of 13 already known HLA—DQB1 Alleles were detected in patients with VKH by PCR—SSE The most frequent allele was HLA—DQBI~*0401(31.82%). It was in strong association with VKH syndrome as compared to the healthy controls (31.82% vs. 4.55%,χ~2=44.00, OR=9.8). So was for HLA—DQBl~*0303 (6.82%vs. 0.57%,χ~22=9.67, OR=12.81). On the contrary, the frequency of HLA—DQB1~*0601 (1.70% vs. 9.66%,χ~2=10.39, P=0.001, OR=0.16)and HLA—DQB1~*0302 (6.25% vs. 19.32%,χ~2= 13.48, P=0.000, OR=0.28) were lower in patients with VKH than that of controls, the differences were statistically significant. We found no relationships among HLA—DQBI~*0401 and clinical manifestations such as age of onsets, visual acuity, cataract, complicated glaucoma, exudative retinal detachment.By PCR—SSCP, 16 representative gels on denatured polyacrylamide gel electrophoresis for all subjects were found. The types of gels among patients and controls were significantly different in 2—, 3—, 4—band gels. After ligation with plasmids, transfection into recipient cells and sequencing of the culturing bacteria, the sequences of the corresponding fragments were confirmed. Matched by already known HLA--DQB1 promoters sequences(QBP1), five kinds of QBPls, ~*0201, ~*0301, ~*0302, ~*0401 ~*06011, were summed up. The frequency of HLA--DQB1 promoter(QBP1) ~*0601 was significantly lower than that of controls (χ~2=11.68, P=0.001). 12 SNPs, including—21 ACT/———,—40 G/A,—94 G/A,—103 G/A,—151 G/A,—171 G/A,—173 T/C,—182—/A,—182———/AAA,—189 C/A,—214 G/A and—227 G/A, were discovered. Among them, the frequency of—189 C/A in patients with VKH was significantly higher than that in controls(χ~2= 45.92, P=0.000). On the contrary the frequency of—227 G/A in patients with VKH was significantly lower than that in controls(χ~2= 15.63, P=0.000).Combinational analysis of the relationships among the coding regions and promoters was established. The cumulative effects among HLA—DQBI~*0401,—DQBI~*0601 and QBP1~*0601 were found. The same phenomenons were found among—189 C/A,—227 G/A, HLA--DQB1~*0401 and HLA—DQB~*0601. These findings matched with the law of cumulative effects of multiple genotypes.It can be concluded: 1) Alleles of HLA—DQB1~*0401 and HLA--DQB1~*0303 were susceptible to VKH syndrome, 2) HLA—DQB1~*0601 and HLA—DQB1~*0302 were resistant to VKH syndrome. 3) PCR--SSP was a rapid method to identify the HLA—DQB1 alleles and can be used as routine. 4) QBPI~*0601 was the resistant gene in patients with VKH. 5)—189 C/A and—227 G/A were the susceptible and resistant SNPs of QBP1 in patients with VKH respectively. 6) With respective corresponding coding alleles (HLA—DQB1~*0401 and ~*0601), QBP1~*0601,—189 C/A and—227 G/A play accumulative effects on patients with VKH.