HLA-A, B, DR, and DQ Antigens in Indian Patients with Severe Chronic Rheumatic Heart Disease

HLA-A, B, DR, and DQ Antigens in Indian Patients with Severe Chronic Rheumatic Heart Disease Breminand MAHARAJ, MD, Shaun M. KHEDUN, M Med Sci, Michael G. HAMMOND, DSc, and Kenneth van der Byl, BSc R SUMMARY To determine whether genetic factors could be involved in the pathogenesis of rheumatic heart disease, we performed HLA-A and HLA-B typing in 59 Indian patients with severe chronic rheumatic heart disease requiring cardiac surgery, and HLA-DR and HLA-DQ typing in 58 of these patients. The HLA typing was done by a standard microlymphocytotoxicity method. Patients were 12 to 59 years old (mean 32.9 years). No significant differences in HLA- A, HLA-B, HLA-DR and HLA-DQ frequencies between patients and controls were noted. The role of genetically determined immune-response factors in the pathogenesis of chronic rheumatic heart disease was not evident in this study. (Jpn Heart J 1997; 38: 663-668) Key words: heart disease HLA-A, HLA-B, HLA-DR, HLA-DQ Antigens, Rheumatic HEUMATIC fever is a major problem in developing countries. Many patients develop chronic rheumatic heart disease which has devastating sequelae. It has been suggested that genetic factors contribute towards a predisposition to rheumatic fever and rheumatic heart disease.1) The HLA antigens, which are encoded by closely arranged genes on the short arm of the sixth chromosome, influence the predisposition to several diseases.2) Some with initially weak associations with HLA-A and HLA-B antigens are found to have stronger associations with HLA-DR antigens.3) Against this background, we wished to determine if genes within or closely linked to the major histocompatibility complex (MHC) were etiologically linked to rheumatic heart disease. We chose to study Indian patients with this disease because our earlier work in black patients had supported the hypothesis.4) Another reason for selecting patients with rheumatic heart disease was that rheumatic fever normally occurs in the pediatric age group and we therefore had reservations about taking the relatively large volume From the Department of Experimental and Clinical Pharmacology, University of Natal Medical School, and Natal Institute of Immunology, Durban, South Africa. Address for correspondence: Breminand Maharaj, MD, Department of Experimental and Clinical Pharmacology, University of Natal Medical School, P Bag 7, Congella 4013, Durban, South Africa. Received for publication November 8, 1996. Accepted March 17, 1997. 663

664 MAHARAJ ET AL Jpn Heart J September 1997 of blood required for HLA studies from children. Accordingly we performed HLA typing in a group of Indian patients with chronic rheumatic heart disease. MATERIALS AND METHODS The control group consisted of 1444 black adults. HLA-A and HLA-B typing was carried out in 59 Indian patients with severe chronic rheumatic heart disease, as defined by the World Health Organization,5) who required cardiac surgery at the Cardiothoracic Surgical Unit, Wentworth Hospital, Durban; HLA-DR and HLA-DQ typing was performed in 58 of these patients. The distribution of valvular lesions was as follows: isolated mitral stenosis, 22 patients mitral stenosis plus aortic incompetence, 1 patient; mitral incompetence and aortic incompetence with aortic stenosis (mixed aortic valve disease), 1 patient; mitral plus aortic incompetence, 5 patients; mitral stenosis with mitral incompetence (mixed mitral valve disease), 16 patients; mixed mitral valve disease plus aortic incompetence, 8 patients; mixed mitral valve disease plus mixed aortic valve disease, 1 patient; isolated aortic incompetence, 5 patients. In each case the rheumatic etiology of the valve lesions was confirmed by inspection of the valve at surgery or on histological examination of the valve. There were 33 female and 26 male patients between 12 and 59 (mean: 32.9) years. The control group consisted of 1444 normal black adults for HLA-A and HLA-B typing, 538 for HLA-DR typing and 370 for HLA-DQ typing. Although over 2000 individual have been tested for the HLA-DR and the HLA-DQ locus in our laboratory, the majority were white or patients with selected diseases.6) The HLA-A and HLA-B antigens were identified with a 2-stage lymphocytotoxicity test.7) These antigens were identified with 180 antisera, which consisted of local sera which had been requested for use in International Histocompatibility Workshops, local sera that had been verified by use in parallel with International Workshop sera and sera that had been exchanged with other laboratories world wide,8-12) Similarly, 120 sera were used to define the HLA-DR and HLA-DQ antigens in B-cell enriched lymphocyte suspensions prepared with the use of straw packed with nylon wool.13) The difference in the frequency of various antigens between patients and control subjects was tested for significance by means of the chi-square test (without Yates' correction). The resulting p value was multiplied by the number of HLA antigens tested to determine the corrected p value. Relative risk was calculated according to the method of Svejgaard et al.1 RESULTS The percentages of HLA-A, HLA-B, HLA-DR and HLA-DQ antigens in

Vol38 No5 HLA ANTIGENS IN RHEUMATIC HEART DISEASE 665 Table I. Frequencies of HLA-A Antigens (%) for all comparisons. Table II. Frequencies of HLA-B Antigens (%) for all comparisons.

666 MAHARAJ ET AL Jpn Heart J September 199 Table III. Frequency of HLA-DR Antigens (%) for all comparisons Table IV. Frequencies of HLA-DQ Antigens (%) for all comparisons patients with chronic rheumatic heart disease and control subjects are shown in Tables I to IV. There was no significant difference in the frequency of the antigens at the A, B, DR and DQ loci between patients and control subjects. DISCUSSION Associations between disease and the HLA system may involve class I (HLA-A, B or C) or class II (HLA-DR or HLA-DQ) antigens. In this study, no differences in frequency of any of the HLA-A, B, DR or DQ antigens between Indian patients with chronic rheumatic heart disease and control subjects were found. Our observations confirm our impression obtained from analysis of previous studies1,4,15-26) that no association exists between rheumatic heart disease and any of the antigens at the A or B loci. However, our results with respect to class II antigens are at variance with those we obtained in an earlier study in black patients.4) We had identified an allelic association with rheumatic heart disease in black patients; there was a significantly increased expression of HLA DR1 and HLA DR w6 antigens.4) We were also unable to confirm the findings of workers in India.22,24) One group of workers reported an increased frequency of HLA-DR3 and a decreased frequency of HLA-DR2 in North Indian patients with rheumatic heart disease.

Vol38 No5 HLA ANTIGENS IN RHEUMATIC HEART DISEASE 667 Another team of investigators found a decreased frequency of HLA-DR2 and an increased frequency of HLA-DQw2 in North Indian patients with rheumatic heart disease.24) An increased frequency of HLA-DR2,25) HLA-DR3,26) HLA- DR4,1,23) HLA-DR623) and HLA-DR3 and HLA DR726) antigens has been observed in patients with this disease. The hypothesis that predisposing genetic factors are important in the development of rheumatic heart disease is not new. From the diversity of findings in published reports on the HLA system in patients with rheumatic heart disease and our present findings it is evident that this hypothesis remains unproven. ACKNOWLEDGEMENTS We thank the late Professor B.T. Le Roux, Department of Cardiothoracic Surgery, Wentworth Hospital, Durban, for allowing us to study patients under his care. REFERENCES 1. Rajapakse CNA, Halim K, Al-Orainey I, Al-Nozha M, Al-Aska AK. A genetic marker for rheumatic heart disease. Br Heart J 1987; 58: 659-62. 2. Stastny P, Ball EJ, Dry PJ, Nunez G. The human immune response region (HLA-D) and disease susceptibility. Immunol Rev 1980; 70: 113-53. 3. Bodmer WF. The HLA systems. Br Med Bull 1978; 34: 213-6. 4. Maharaj B, Hammond MG, Appadoo B, Leary WP, Pudifin DJ. HLA-A, B, DR and DQ antigens in black patients with severe chronic rheumatic heart disease. Circulation 1987; 76: 259-61. 5. World Health Organisation: Prevention of rheumatic fever. WHO Technical Report Series, No 342. 6. Hammond MG, Appadoo B. HLA antigens in African blacks. In: Aizawa M, Natori T, Wakisaka A, Konoeda Y, editors. HLA in Asia-Oceania, 1986. Sapporo: Hokkaido University Press, 1986: 316-9. 7. Mittal KK, Mickey MR, Singal DP, Terasaki PI. Serotyping for homotransplantation. XV111: refinement of microdroplet lymphocyte cytotoxicity test. Transplantation 1968; 6: 913-27. 8. Hammond MG, Appadoo B, Brain P. HLA antigens in Bantu and Indians. In: Kissmeyer-Nielsen F, editor. Histocompatibility Testing. Copenhagen: Munksgaard, 1975: 173-8. 9. Hammond MG, Appadoo B, Brain P. HLA in non-caucasian populations. In: Bodmer WF, editor. Histocompatibility Testing. Copenhagen: Munksgaard, 1977: 407. 10. Hammond MG. HLA Bw53. In: Terasaki PI, editor. Histocompatibilty testing 1980. Los Angeles: UCLA Tissue Typing Laboratory, 429-32, 1980. 11. Hammond MG. HLA Bw53. In: Simons MJ, Tait BD, editors. Proceedings of the Second Asia- Oceania Histocompatibilty Workshop. Toorak: Immunopublishing, 1983: 112-4. 12. Hammond MG, Betuel H, Gebuhrer L. HLA A29. In: Albert ED, Baur MP, Mayr WR, editors. Histocompatibilty Testing. Berlin: Springer Verlag, 1984: 126. 13. Danilov JA, Ayoub G, Terasaki PI. Joint report: B lymphocyte isolation by thrombin-nylon wool. In: Terasaki PI, editor. Histocompatibilty testing 1980. Los Angeles: UCLA Tissue Typing Laboratory, 1980: 287-8. 14. Svejgaard A, Platz P, Ryder LP, Staub-Nielsen L, Thomsen M. HLA and disease associations. Transplant Rev 1975; 22: 3-43. 15. Falk JA, Fleischman JL, Zabriskie JB, Falk RE. A study of HLA antigen phenotype in rheumatic fever and rheumatic heart disease patients. Tissue Antigens 1973; 3: 173-8.

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