Masami Yanagisawa

Lack of primary association between transporter associated with antigen processing genes and atopic dermatitis

We examined polymorphisms of transporter associated with antigen processing (TAP) genes in 37 Japanese patients with atopic dermatitis and 52 control subjects. We have evaluated, again, the specificity polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method since our previous report and have also developed a mismatch PCR-RFLP method for discriminating two dimorphic sites of TAP1 gene and four dimorphic sites of TAP2 gene. Amplified products from genomic DNA were digested with restriction endonucleases: Sau3A1 for TAP1 codon 333 (Ile-Val), AccI for TAP1 codon 637 (Asp-Gly), and BfaI for TAP2 codon 687 (Stop-Gln). The other sites of TAP2 gene were analyzed by mismatch PCR-RFLP: AccII for TAP2 codon 379 (Val-Ile), RsaI for codon 565 (Ala-Thr), and MspI for codon 665 (Thr-Ala). We observed three TAP1 alleles and six TAP2 alleles. Infrequent allele TAP1 C was observed in one patient and one control subject. We did not identify any differences in TAP allele frequencies between those patients with atopic dermatitis and Japanese control subjects. Analysis of TAP gene polymorphisms will provide better understanding of susceptibility loci in HLA class II-associated disease because TAP genes are located between HLA-DQB1 and HLA-DPB1 loci.

Polymorphisms of transporter associated with antigen processing genes in atopic dermatitis

We investigated polymorphisms of transporter associated with antigen process (TAP) genes in atopic dermatitis. We developed a polymerase chain reaction-restriction fragment length polymorphism method for discriminating TAP alleles. Genomic DNA was obtained from 29 Japanese patients with atopic dermatitis and 35 control subjects. Dimorphic regions of TAP1 and TAP2 genes were amplified by polymerase chain reaction. Amplified products were digested with restriction endonucleases to determine TAP alleles: Sau3A1 for TAP1 codon 333 (Ile-Val), AccI for TAP1 codon 637 (Asp-Gly), and EcoRII for TAP2 codon 687 (Gln-Stop). We observed four alleles for TAP1 and dimorphism for TAP2 codon 687. Six of 35 controls had the TAP1 D allele, which has been reported to be a rare allele in Caucasian populations. Gene frequency of TAP1 637Asp exhibited a tendency to increase in the patients with atopic dermatitis. TAP1 637Asp and TAP1 A alleles were estimated to constitute a haplotype with DRB1* 1302-DQB1*0604 and DRB1*0803-DQB1*0601 in the Japanese population. Because TAP1 and TAP2 genes are located between HLA-DQB1 and -DPB1 loci, analysis of TAP gene polymorphisms will be useful for a better understanding of susceptibility loci in HLA class II-associated disease.

Analysis of disease-associated amino acid epitopes on HLA class II molecules in atopic dermatitis.

We investigated the association between HLA class II alleles and severe atopic dermatitis with high serum IgE levels (greater than 8000 U/ml). The frequencies of HLA-DRB1*1302 and DQB1*0604 were increased, whereas the frequency of HLA-DQB1*0302 was decreased. A strong haplotype, HLA-DRB1*1302-DQB1*0604, has been reported in the Japanese population, and this haplotype is conserved in patients with AD. Further analysis of the amino acid epitopes on the HLA-DR beta 1 and DQ beta 1 domains revealed that DR beta 1 71Glu (RR = 5.71, p < 0.05) and DQ beta 1 30His (RR = 3.25 p < 0.01), and 57Val (RR = 3.13, p < 0.05) were increased in frequency. DR beta 1 71Glu was exclusively unique to DRB1*1302. DQ beta 1 30His was shared by the following alleles: DQ beta 1*0501, *0502, *0503, and *0604, which were all increased in patients with AD. DQ beta 1 57Val was shared by DQB1*0501 and *0604. A well-known haplotype, HLA-DRB1*1302-DQB1*0604, contains DR beta 1 71Glu and DQ beta 1 30His and 57Val. Therefore HLA-DR beta 1 71Glu and/or DQ beta 1 30His/57Val are considered to play the most important role in the development of atopic dermatitis.

HLA-DM gene polymorphisms in atopic dermatitis

HLA-DM molecules are involved in the antigen-processing pathway of HLA class II-restricted antigen presentation. We investigated polymorphisms of HLA-DM genes in atopic dermatitis by using the polymerase chain reaction-restriction-fragment length polymorphism method to examine a possible contribution of these genes to the pathogenesis of atopic dermatitis. Genomic DNA was extracted from 37 Japanese patients with atopic dermatitis and 52 control subjects. After polymerase chain reaction amplification of the polymorphic third exon of DMA and DMB genes, amplified products were digested with restriction endonucleases to determine HLA-DM alleles. FokI, HinfI, AciI and SfaNI were used for DMA; HhaI, BsrI, ApaLI, and Bsp1286I for the DMB gene. We identified three DMA alleles and also three DMB alleles. One of 37 patients possessed the DMA*0103 allele, which has been reported as a rare allele in Caucasian populations. Any DMA and DMB alleles were not increased in the patients. The DMA*0102 allele was estimated to constitute a haplotype with DRB1*1201/DQB1*0301 and DRB1*0901/DQB1*0301 in a Japanese population. HLA-DM genes are not considered to contribute primarily to the susceptibility of atopic dermatitis. Further investigation of the functional roles of HLA-DM gene polymorphisms will be useful for a better understanding of susceptibility loci in HLA class II-associated disease.

HLA-DR types in Japanese schizophrenics: analysis by group-specific PCR amplification

An association of HLA-DR8 and DR1 with DSM-III schizophrenia has been reported in Japan (Miyanaga et al. (1984) Biol. Psychiatr. 19, 121-129). To further investigate this preliminary finding, we compared HLA-DR types in 44 unrelated Japanese schizophrenics (DSM-III-R) with those in 51 unrelated, healthy Japanese volunteers. Group-specific PCR amplification was used in the determination of HLA-DR in the patients. No significant difference was observed in the frequency of any DR types between patients and controls, after statistical correction for multiple testing. However, the frequency of DR1 in our patients (23%) and controls (10%) was almost the same as those in the previous report (22% vs. 10%), which means that there is a suggestive trend which could become significant if numbers were larger. It is argued that an exact determination of HLA-DR by DNA typing is important in current HLA studies of schizophrenia.

HLA and atopic dermatitis with high serum IgE Levels

Patients with atopic dermatitis usually exhibit allergen-specific IgE antibodies against several environmental antigens. HLA restriction is presumed to be involved in the recognition of such antigens, but several previous reports have so far failed to find a significant association between atopic dermatitis and HLA antigens. In this study we examined 38 unrelated Japanese patients with severe atopic dermatitis and high serum IgE levels (greater than 800 U/ml). We investigated the serological HLA types and HLA class II alleles in this group of patients with atopic dermatitis. Frequencies of HLA-A24, A33, Cwblank, B44, DR13 and HLA-DRB1*1302, DQB1*0604, DPB1*0301 alleles were increased in the patients. In contrast, frequencies of HLA-Cw1, Bw6, DR4, DR53, and HLA-DQB1*0302 allele were decreased. However, none of these remained significant after p values were corrected. Further study on HLA association with atopic dermatitis through characterization of specific antigens or antigen epitopes is needed.

Ethnic Differences in Allele Frequencies of Two Microsatellite Markers Closely Linked to the Locus for Polycystic Kidney Disease 1 (PKD1)

Two microsatellite markers, D16S283 and D16S284, closely linked to the locus of autosomal polycystic kidney disease (PKD1), were amplified in a Japanese population sample by the polymerase chain reaction to investigate differences in allele frequencies between Japanese and Caucasian populations. Ten D16S283 alleles and four D16S284 alleles were discerned among 53 Japanese. The observed heterozygosities of D16S283 and D16S284 in this study were 76.8 and 9.1%, and the polymorphism-informative contents were 0.75 and 0.09, respectively. Several allele frequencies of D16S283 exhibited significant differences. The Y9 allele (19 CA repeats, 91 bp) was the most frequent in both populations but its frequency in Caucasians was higher. The second most frequent allele was different: Y6 (22 CA repeats, 97 bp) in Japanese and Y10 (18 CA repeats, 89 bp) in Caucasians. Y8 (20 CA repeats, 93 bp) and Y11 (17 CA repeats, 87 bp) alleles also showed different frequencies. For D16S284, no differences in allele frequencies were observed between the two populations. As we observed differences in allele frequencies for D16S283, an association study with this polymorphism should carefully control for the ethnic origin of subjects.

Genetic polymorphisms of complement components C6 and C7 in Korean

SummaryGenetic polymorphisms of the complement components C6 and C7 were investigated in Korean living in Seoul using isoelectric focusing and immunoblotting. Three common and four rare C6 allotypes were observed. The allele frequencies estimated were as follows: C6* A 0.433, C6* B 0.523, C6* B2 0.039, and rare alleles (M91, M92, M11 and M2) 0.005. Three C7 variants, besides a common allele, were observed with polymorphic frequencies. Two rare C7 variants, considered to be new, were also observed. The allele frequencies estimated for C7*1, C7*2, C7*3, C7*4, and rare variants (tentatively named K1 and K2) were 0.843, 0.073, 0.034, 0.048 and 0.002, respectively. The C6 and C7 allele frequencies are similar to those in Japanese and Chinese. The association analysis between C6 and C7 showed a significant negative association between C6 B and C7 4 allotypes (p<0.02).