DNA sequencing validation of PCR-RFLP technique for evaluating butyrophilin-like 2 rs2076530 polymorphism in Iranian patients with sarcoidosis

Main Article Content

Alireza Javadi
Masoud Zarei
Mihan Pourabdollah
Makan Sadr
Arda Kiani
Atefeh Abedini
Masoud Shamaei


BTNL2, Sarcoidosis, RFLP, Polymorphism, Single Nucleotide


Background: Sarcoidosis is a multifactorial immune disorder with uncertain origin. A single nucleotide polymorphism (G→A, rs2076530) in the butyrophilin-like 2 (BTNL2) gene results in the formation of truncating protein. This study aimed to genotype the predisposition of the BTNL2 rs2076530 polymorphism in Iranian patients with sarcoidosis using the newly designed RFLP technique.

Materials and Methods: In this study, 80 patients with sarcoidosis and 80 healthy individuals were included. The rs2076530 polymorphism of the BTNL2 gene was genotyped using the PCR-RFLP method by AvrII restriction enzyme and confirmed by DNA sequencing (Capillary electrophoresis 3130, ABI).

Results: A statistical significant was observed in proportion of patient's AA (67.9%) in comparison to AA (40.7%) (OR=3.07,95%CI:1.40-6.7, P=0.004) in controls. Also, a significant difference was statistically found between the A (62.5%) in subjects and A (43.8%) allele in controls (OR=2.14,95%CI:1.37-3.35, P=0.001). However, no statistical difference was seen between cases and controls based on AA and AG proportion.

Conclusion: Our data emphasized the association of the BTNL2 rs2076530 A allele with sarcoidosis susceptibility in Iranian patients. The newly designed PCR-RFLP method for the rs2076530 A allele could also be applicable in resource-limited countries.

Abstract 447 | PDF Downloads 305


1. Moller DR, Rybicki BA, Hamzeh NY, et al. Genetic, immunologic, and environmental basis of sarcoidosis. Annals of the American Thoracic Society. 2017;14(Supplement 6):S429-S36.
2. Baughman R, Culver D, Judson M, Gerke A, Maier L, Spitzer G. Self Reported Organ Involvement in Sarcoidosis: Results of a Multi-National Registry. D103 TRANSPLANT AND ILD POTPOURRI: American Thoracic Society; 2018. p. A7482-A.
3. Fontenot A, King Jr TE, Nicholson A. Pathology and pathogenesis of sarcoidosis. Uptodate/Internet. 2018.
4. Newman LS, Rose CS, Bresnitz EA, et al. A case control etiologic study of sarcoidosis: environmental and occupational risk factors. American journal of respiratory and critical care medicine. 2004;170(12):1324-30.
5. Terwiel M, van Moorsel CH. Clinical epidemiology of familial sarcoidosis: a systematic literature review. Respiratory medicine. 2018.
6. Shamaei M, Pourabdollah M, Mousaei H, Mirsaeidi M, Masjedi MR. The presence of mycobacterial antigens in sarcoidosis associated granulomas. Sarcoidosis vasculitis and diffuse lung disease. 2017;34(3):236-41.
7. Wang SH, Chung CH, Huang TW, et al. Bidirectional association between tuberculosis and sarcoidosis. Respirology. 2019;24(5):467-74.
8. Shamaei M, Javadi A, Sadr M, Pourabdollah M. Sarcoidosis or Tuberculosis? Detecting Mycobacterium tuberculosis Complex DNA in Sarcoidosis Granulomas. Current Respiratory Medicine Reviews. 2018;14(2):100-4.
9. Ishihara M, Ishida T, Inoko H, Ando , et al. HLA serological and class II genotyping in sarcoidosis patients in Japan. Japanese journal of ophthalmology. 1996;40(1):86-94.
10. Foley PJ, McGrath DS, Puscinska E, et al. Human leukocyte antigen-DRB1 position 11 residues are a common protective marker for sarcoidosis. American journal of respiratory cell and molecular biology. 2001;25(3):272-7.
11. Rossman MD, Thompson B, Frederick M, et al. HLA-DRB1* 1101: a significant risk factor for sarcoidosis in blacks and whites. The American Journal of Human Genetics. 2003;73(4):720-35.
12. Rybicki BA, Maliarik MJ, Poisson LM, Sheffer, et al. The major histocompatibility complex gene region and sarcoidosis susceptibility in African Americans. American journal of respiratory and critical care medicine. 2003;167(3):444-9.
13. Sato H, Woodhead FA, Ahmad T, et al. Sarcoidosis HLA class II genotyping distinguishes differences of clinical phenotype across ethnic groups. Human molecular genetics. 2010;19(20):4100-11.
14. Stammers M, Rowen L, Rhodes D, Trowsdale J, Beck S. BTL-II: a polymorphic locus with homology to the butyrophilin gene family, located at the border of the major histocompatibility complex class II and class III regions in human and mouse. Immunogenetics. 2000;51(4-5):373-82.
15. Rhodes D, Stammers M, Malcherek G, Beck S, Trowsdale J. The cluster of BTN genes in the extended major histocompatibility complex. Genomics. 2001;71(3):351-62.
16. Rybicki BA, Walewski JL, Maliarik MJ, Kian H, Iannuzzi MC, Group AR. The BTNL2 gene and sarcoidosis susceptibility in African Americans and Whites. The American Journal of Human Genetics. 2005;77(3):491-9.
17. Valentonyte R, Hampe J, Huse K, et al. Sarcoidosis is associated with a truncating splice site mutation in BTNL2. Nature genetics. 2005;37(4):357.
18. Shahinian A, Pfeffer K, Lee KP, et al. Differential T cell costimulatory requirements in CD28-deficient mice. Science. 1993;261(5121):609-12.
19. Krinzman SJ, De Sanctis GT, Cernadas M, et al. Inhibition of T cell costimulation abrogates airway hyperresponsiveness in a murine model. The Journal of clinical investigation. 1996;98(12):2693-9.
20. Harding FA, McArthur JG, Gross JA, Raulet DH, Allison JP. CD28-mediated signalling co-stimulates murine T cells and prevents induction of anergy in T-cell clones. Nature. 1992;356(6370):607.
21. Society AT. Statement on sarcoidosis. Joint Statement of the American Thoracic Society (ATS), the European Respiratory Society (ERS) and the World Association of Sarcoidosis and Other Granulomatous Disorders (WASOG) adopted by the ATS Board of Directors and by the ERS Executive Committee. Am J Respir Crit Care Med. 1999;160(2):736-55.
22. Shamaei M, Mortaz E, Pourabdollah M, et al. Evidence for M2 macrophages in granulomas from pulmonary sarcoidosis: a new aspect of macrophage heterogeneity. Human immunology. 2018;79(1):63-9.
23. Nguyen T, Liu XK, Zhang Y, Dong C. BTNL2, a butyrophilin-like molecule that functions to inhibit T cell activation. The Journal of Immunology. 2006;176(12):7354-60.
24. Li Y, Wollnik B, Pabst S, Lennarz M, Rohmann E, Gillissen A, et al. BTNL2 gene variant and sarcoidosis. Thorax. 2006;61(3):273-4.
25. Milman N, Svendsen CB, Nielsen FC, van Overeem Hansen T. The BTNL2 A allele variant is frequent in Danish patients with sarcoidosis. The clinical respiratory journal. 2011;5(2):105-11.
26. Morais A, Lima B, Peixoto MJ, Alves H, Marques A, Delgado L. BTNL2 gene polymorphism associations with susceptibility and phenotype expression in sarcoidosis. Respiratory medicine. 2012;106(12):1771-7.
27. Suzuki H, Ota M, Meguro A, Katsuyama Y, Kawagoe T, Ishihara M, et al. Genetic characterization and susceptibility for sarcoidosis in Japanese patients: risk factors of BTNL2 gene polymorphisms and HLA class II alleles. Investigative ophthalmology & visual science. 2012;53(11):7109-15.
28. Lin Y, Wei J, Fan L, Cheng D. BTNL2 gene polymorphism and sarcoidosis susceptibility: a meta-analysis. PloS one. 2015;10(4):e0122639.

Most read articles by the same author(s)