: 58 1 2000 ( ) ( ) ( ) HLA- DR ( ) polymorphis m, *, (SCL), * * = A bs tra c t= A s tudy of HLA- DR polymorphis m in f our physical cons titution g roups clas s if ied by Korean traditional medicine T aek- Boo kwon, M.D., Chee- Won Whang*, M.D., In- Chan Sul*, M.D,.Kyung- Ok Lee, M.D. Kyu- Yoon Hwang, M.D., Dong- Ho Yang, M.D. and Sae- Yong Hong, M.D. Department of Internal M edicine, Soonchunhyang University. Department Preventive M edicine, Soonchunhyang University. Oriental M edical School, T ae J un University.* Seoul M edical Science Institute, Seoul Clinical Laboratory(SCL). Backg round : In Korean traditional medicine, human beings are classified into 4 groups accordance with the characteristics of physical constitution, named ; Taeum, Taeyang, Soum, and Soyang. Most of Korean traditional doctors believe that the back ground of pathophysiology is differ among each group in many diseases. HLA is the most polymorphic gene and related with human immune response. It has been used for determination of individual identification, genetic diversity and a study of disease association in different ethnic group. It has well known that the most human inflammatory disease which thought to have some autoimmune basis are in some way promoted by genes of the HLA region. The purpose of this study is to observe the distribution of HLA- DR genotypes in 4 types of physical constitution in Korean traditional medicine. Methods : T hree hundred and ten Korean subjects(soyang : 58cases, Soum :142cases, Taeum : 92cases, Taeyang : 18cases) were enrolled for this study. Genomic DNAs extracted by phenol/chroloform were amplified with allele specific primers and determined by 2% agarose gel electrophoresis with ethidium bromide staining for analysis of HLA- DR polymorphism.. Results : The genotype frequency of HLA- DR in a total or 310 Koreans was DR1 ; 7.2%, DR 2; 10.5%, DR 3;1.7%, DR 4; 19.9%, DR 5; 11.9%, DR 6;19.9%, DR 7; 7.0%, DR 8; 10.6%, DR 9; 8.7%, DR10;1.9%. There was no significant diffrence of HLA- DR genotype distribution among four physical constitution groups classified by Korean traditional medicine. Conclusion : Our results suggest that the classification of physical constitution in Korean traditional medicine is not related to HLA-DR genotypes.(korean J Med 58:91-101, 2000) Key Words : HLA-DR, Gene, Physical constitution, Korean traditional medicine. : 1999 6 15 : 1999 8 13 :, 23-20 (330-100) E-mail : syhong@sparc.schch.co.kr - 91 -
Korean Journal of Medicine : Vol. 58, No. 1, 2000 ( ) ( ) ( ) ( ) ( ) ( ) 100 ( ;1837-1900) 1)., 2)., ( ) 1, 2). ( ) ( ), ( ), ( ), ( ) ( ) ( ), ( ), ( ), ( ) 3, 4). ( ) ( ), ( ) ( ), ( ), ( ) (gene) (polymorphism) ( ).,, ABO DNA (phenotype) 1-1 (one gene- one protein) (genome) 30 DNA (base pairs) 5-10 (genes) 5). Major Histocompatibility Complex (MHC) 6). MHC (histocompatible) (incompatible) (antigen) 7, 8). HLA class 1 class II 6 coading Class l glycoprotein( ) HLA- A, HLA- B HLA- C 9). Class ll B,, dendritic cell antigen presenting cell T HLA- DR, HLA- DQ, HLA- DP 34 KD a 28KD b heterodimer a b domain a A, b B 10). HLA,, HLA, 5). HLA 11). HLA HLA - 92 -
T aek- Boo kwon, et al : A study of HLA- DR polymorphism in four physical constitution groups classified by Korean traditional medicine HLA HLA HLA class ll antigens(dr, DQ, DP) DNA HLA 12). HLA- DR locus A B 13-21). (genome) (gene) HLA (association) HLA- DR 1. 1997 3 1 ( ) 310 HLA- DR 45 (45 11 ) 2. 2 ( ) 22,23) ( ) ( ) 1, 2, 3, 4 (5 ) 22, 23), ( ) 3. HLA-DR cubital vein EDT A DNA SSC(15mM sodium citrate, 150 mm NaCl) 12,000RPM 3 0.2M sodium acetate(ph 7.0) 400 µl, proteinase K(10mg/ml) 10 µl, 10% SDS 40 µl 56oC 1 phenol/chloroform 120 µl 200 µl 100% ethanol Ethanol PCR- SSP (polymerase chain reaction- sequence specific primer) Sequence specific primers Marsh24) HLA- DRB1 gene exon 2 nucleic acid sequence 393 DNA/RNA synthesizer oligonucleotide purification cartridge (Applied Biosystem, USA) HLA- DR PCR HLA- DR PCR 1 group specific - 93 -
: 58 1 473 2000 Table 1. Sequences for the HLA- DR allele specific primers DR Primer Sequence Orientation DR 1 5' - TT TCTT GT GGCAGCT TAAGT TT GAA- 3' Sense DR 2 5' - ACGT TT CCT GT GGCAGCCTAAGAGG- 3' Sense DR 4 5' - CGTT TCT TGGAGCAGGTT AAACA- 3' Sense DR 7 5' - CACGTT TCCT GT GGCAGGG- 3' Sense DR 9 5' - CACGTT TCT TGAAGCAGGA- 3' Sense DR 10 5' - CACGTT TCT TGGAGG- 3' Sense DR(3,5,6,8) 5' - CACGTT TCT TGGAGTACT CTAC- 3' Sense DR(3,5,6,8) 5' - CACGTT TCT TGGAGTACT CTACGTC- 3' Sense DR 3 5' - TGTCCACCCGGCCCCGCT - 3' Antisence DR 5(11) 5' - GGCT GT TCCAGTACT CCT - 3' Antisence DR 5(12) 5' - TGCGGT TACT GGAGAGAC- 3' Sense DR 8 5' - TGCAGT AGGTGTCCACCA- 3' Antisence DR ALL 5' - CCCCACAGCACGTT TCT TG- 3' Sense DR ALL 5' - CCGCTGCACT GT GAAGCT - 3' Antisence primer 7 - DR1, DR2, DR4, DR6, DR7, DR9, DR10, DR52 DR52 DR3, 5, 8 2 PCR PCR 20ul 200ng DNA primer(t able 1) 20pmol 200mM dnt Ps 1 unit T aq polymerase, 56 mm KCL, 1.7mM MgCl2, 11mM T ris Hcl(pH 8.3), 0.0011%(W/V) gelatin 94oC 5 94oC 30, 65oC 45, 72oC 40 1 25 72oC 10 2% agarose ethidium bromide DNA band DR1, DR2, DR4, DR52, DR7, DR9, DR10 DR52 2 PCR PCR 1 l 10 x buffer(50mm KCl, 10 mm T ris- HCl ph 8.8), 1.5 mm MgCl2, 0.1% T riton X- 100), 0.5 unit T aq DNA polymerase, 0.4 mm dnt Ps, primer(10 pmol) PCR 2% agarose gel ethidium bromide DNA band DR52 3, 8, 11, 12 4. HLA- DR Chi square test HLA- DR1, 3, 7, 10 DR 5 chi- square test,, 1. 310 58 (18.7%), 142 (45.8%), 92 (29.7%), 18 (5.8%) 2. HLA DR 4 HLA- DR - 94 -
6 : ( ) ( ) ( ) HLA- DR ( ) polymorphism Table 2. Distribution of HLA- DR g enotype on the classes based on physical constitution in korean traditional medicine HLA DR1 DR2 DR3 DR4 DR5 DR6 DR7 DR8 DR9 DR10 Soyang (%) 5.2 10.3 3.4 24.1 15.5 15.5 3.4 6.9 13.8 1.7 Soum (%) 5.6 9.9 3.5 23.3 12.7 14.1 7.7 4.2 16.2 2.8 T aeum (%) 6.5 7.5 1.1 23.9 13.0 20.7 10.0 4.3 8.7 4.3 T aeyang (%) 0 16.7 0 22.2 27.8 16.7 0 11.1 0.6 0 T otal (%) 5.4 9.7 2.6 23.5 14.2 16.5 7.1 5.2 12.9 2.90 T otal n (58) (142) (92) (18) 100(310) p value *Taeyang group was not included for Chi square test. Chi square test of each HLA- DR genotype among different types of physical constitution in Korean Traditional Medicine. 0.74 0.68 0.58 0.99 0.37 0.62 0.30 0.57 0.29 0.86 T able 2 HLA- DR (p> 0.05).,, 10, DR1, DR3, DR7, DR10 DR4 20% (22.2%- 24.1%) DR5 DR6 10% HLA- DR (58 ) DR4 24.1% DR5 DR6 15.5% DR9(13.8%), DR2(10.2%) DR3 DR7 5% DR10(1.7%) (142 ) DR4 23.3% DR9(16.2%), DR6(14.1%), DR5(12.7%) (92 ) DR4(23.9%) DR6(20.7%), DR5(13.0%) DR6(20.7%) DR7(10.0%) 4 DR3(1.1%) (18 ) DR5(27.8%) DR4(22.2%), DR6(16.7%), DR2(16.7%) DR5 DR1, DR3, DR7, DR10,, HLA DR (p> 0.05). HLA- DR HLA- DR 25, 26). HLA HLA 12). 310 4 142 (45.8%) 18 (5.8%) 3) 310 HLA- DR 13) DR4 (23.5% vs 19.9%), - 95 -
Korean Journal of Medicine : Vol. 58, No. 1, 2000 DR6 (16.5% vs 19.9%) DR8 (5.2% vs 10.6%) HLA (relative risk) HLA HLA HLA (linkage disequilibrium) HLA HLA idiopathic hemochromatosis HLA- A3, HLA- B27, HLA- DR4, Sjoren DRw52, DQw8, celiac disease DP 6). class ll class ll CD4 T 27). HLA- DR3 28). HLA (association) HLA HLA- DR ( ) ( ) ( ) 29-31). ( ) ( ) ( ) ( ) 5.8%(310 18 ) DR1, 3, 6, 10 " " HLA- DR DR3(relative risk 6.8)6) 18 ( ) ( ) ( ) ( ) ( ) ( ),, 92 29.7% DR4 (23.9%) DR6(20.7%) ( ) ( ).. - 96 -
T aek- Boo kwon, et al : A study of HLA- DR polymorphism in four physical constitution groups classified by Korean traditional medicine 142 45.8% DR4 (23.3%) DR9(16.2%), DR6(14.1%) HLA- DR DR2(gluten- sensitive enteropathy, relative risk 11.6) 9.9% 6). ( ) ( ) ( ) ( ) ( ) ( ) 58 18.7% HLA DR2(Goodpasture' s disease, relative risk 15.9), DR3(membranous GN, relative risk 5.7), DR4(IgA GN relative risk 3.1)6) 10.3%, 3.4%, 24.1% ( ) ( ) ( ), ( ) ( ),,,, HLA- DR : ( ), ( ), ( ), ( ) ( ) HLA,, HLA- DR : 1997 3 1 310 HLA- DR : 310 58 (18.7%), 142 (45.8%), 92 (29.7%), 18 (5.8%), HLA- DR (p> 0.05). : HLA- DR - 97 -
: 58 1 473 2000-1-. ( ), " " " ". A. ( ) 1.? 1) 2) 3) 4) 5) ( ) ( ) 2.? 1) 2) 3) ( ). 4) 5) ( ) ( ) 3.? 1) 2) 3) 4) 5) ( ) ( ) 4.? 1) 2) 3) 4) 5) ( ) ( ) 5.? 1) 2) 3) 4) 5) ( ) ( ) 6.? 1) 2) 3) 4) 5) ( ) ( ) 7.? 1) 2) 3) 4) 5) ( ) ( ) 8.? 1) 2) 3) 4) 5) ( ) ( ) - 98 -
6 : ( ) ( ) ( ) HLA- DR ( ) polymorphism 9.? 1) 2) 3) 4) 5) ( ) ( ) 10.? 1) ( ) 2) ( ) 3) ( ) 4) ( ) 5) ( ) ( ) 11.? 1) 2) 3) 4) 5) ( ) ( ) 12.? 1) ( ) 2) 3) 4) 5) ( ) ( ) 13. ( )? 1) 2) 3) ( ) 4) 5) ( ) ( ) 14. ( )? 1) 2) 3) (, ),. 4),, 5) ( ) ( ) 15.? 1) 2) 3) 4) 5) ( ) ( ) 16. ( )? 1)., 2)., 3). (,,,, ) 4). (,,,,, ) 5). ( ) 17. ( )? 1),,,, 2),,, 3),,, 4),,,, 5) ( ) 18.? 1) ( ). 2) 3) 19.? 1) 2) 3) 4) 5) - 99 -
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