- Email: [email protected]
Interferon gamma gene in rheumatoid arthritis
CORRESPONDENCE
population. Conversely, the carriage rate of the 126 bp IFNG allele (13 CA repeat units) is 12% in the Canadian population, but amounts to 62–68% in our European populations. Since the white Canadian population is generally of recent European descent, a similar allele distribution to European populations would be anticipated. In view of this discrepancy, that the Sardinian IFNG carriage rates are similar to the continental European ones is remarkable. While the phylogeny of the Sardinian population is related to white ethnic origin, it is, in contrast to that of white Canadians, genetically distinct from that of other Europeans.4 If the Canadian IFNG alleles are reassigned by shifting them one position upward (eg, 122 bp→124 bp), a striking resemblance to the European carriage rates is generated. Our data substantiate the concern of Ollier that case-control mismatching of alleles might have led to the differences seen. If allele slippage occurred as we show in the table, it follows that IFNG alleles have been wrongly assigned in the control and mild rheumatoid arthritis groups, but were correctly assigned in the severe rheumatoid arthritis group in Khani-Hanjani and colleagues’ study. This finding would explain why significant differences were seen only on comparison of the latter with each of the former groups. Various errors in methods might account for allele slippage. Apart from those already mentioned by Ollier, the use of size standards between rather than within lanes during electrophoresis could have hampered accurate assignment of alleles as well. Our observations might jeopardise the validity of the findings of KhaniHanjani and colleagues, but IFNG remains an interesting candidate for conferring susceptibility to various autoimmune diseases. By use of a family-based association method (transmission disequilibrium test), we have shown significant distortion of parental transmission of this IFNG microsatellite polymorphism to male offspring with multiple sclerosis in Sardinia.3,5 Ollier reports an odds ratio of 1·8 (95% CI 1·0–3·1) for an IFNG
on Perinatal and Women’s Health, INSERM, Villejuif, France; Unit of Research on Reproduction, INSERM, Toulouse, France; and Division of Epidemiology, IFC, National Research Council, Pisa, Italy 1
2
Pochard F, Azoulay E, Grassin M. End-oflife decisions for newborn infants. Lancet 2000; 356: 946. Cuttini M, Nadai M, Kaminski M, et al. End-of-life decisions in neonatal intensive care: physicians’ self-reported practices in seven European countries. Lancet 2000; 355: 2112–18.
Interferon gamma gene in rheumatoid arthritis Sir—A Khani-Hanjani and colleagues (Sept 2, p 820)1 report a striking association between a dinucleotide polymorphism in the interferon gamma gene (IFNG) and rheumatoid arthritis in Canadians. In a commentary, W E R Ollier (Sept 2, p 783)2 raised the question of whether their allele typing could have slipped by one position, which would lead to the highly significant differences between cases and controls. We did an extensive analysis of this polymorphism in 1730 white European individuals.3 Since the primers used for amplification of the IFNG polymorphism in our and Khani-Hanjani and colleagues’ studies were identical, the numbers of CA repeat units per allele have been determined unambiguously. Direct comparison of IFNG allele carriage rates (ie, the proportion of individuals carrying a certain allele) between the two studies is possible and scientifically correct. We show in the table the IFNG allele carriage rates in German, Swedish, and Sardinian healthy individuals in our study, compared with those for the Canadian white healthy controls of Khani-Hanjani and colleagues. In Europeans, the 122 bp IFNG allele, which encompasses 11 CA repeat units, is rare. The carriage rate of the 122 bp allele was not higher than 1% in any population in our study. According to Khani-Hanjani and colleagues, the carriage rate of this allele is 80% in the white Canadian Population sample
Canadian (n=65)* Canadian (n=65)† Sardinian (n=413) German (n=367) Swedish (n=54)
Interferon gamma 122 bp (11 CA) 124 bp (12 CA) 126 bp (13 CA)
128 bp (14 CA) 130 bp (15 CA)
Other
52 (80%) 0 0 3 0
8 (12%) 8 (12%) 71 (17%) 46 (12·5%) 9 (17%)
0 0 0 14 (14%) 0
44 (68%) 52 (80%) 326 (79%) 248 (68%) 32 (59%)
8 (12%) 44 (68%) 257 (62%) 251 (68%) 35 (65%)
0 8 (12%) 26 (6%) 21 (6%) 6 (11%)
*Original assignment of alleles from Khani-Hanjani and colleagues. †Reassignment of alleles from Khani-Hanjani and colleagues.
Comparison of IFNG allele carriage rates in Canadian and European healthy population samples
THE LANCET • Vol 356 • December 23/30, 2000
single nucleotide polymorphism in male rheumatoid arthritis patients.2 Taken together, these data suggest that the IFNG, or a nearby locus, is functionally associated with sex bias in susceptibility to autoimmune diseases. *Koen Vandenbroeck, An Goris, Alfons Billiau, Ghislain Opdenakker, Cornelia Hardt *School of Pharmacy, Queen’s University of Belfast, Belfast BT9 7BL, UK; Rega Institute for Medical Research, University of Leuven, Leuven, Belgium; Institut für Humangenetik, Universitätsklinikum Essen, Essen, Germany (e-mail: [email protected]) 1
2
3
4
5
Khani-Hanjani A, Lacaille D, Hoar D, et al. Association between dinucleotide repeat in non-coding region of interferon-gamma gene and susceptibility to, and severity of, rheumatoid arthritis. Lancet 2000; 356: 820–25. Ollier WER. Role of interferon-␥ gene in rheumatoid arthritis? Lancet 2000; 356: 783–84. Goris A, Epplen C, Fiten P, et al. Analysis of an IFN-␥ gene (IFNG) polymorphism in multiple sclerosis in Europe: effect of population structure on association with disease. J Interferon Cytokine Res 1999; 19: 1037–46. Cavalli-Sforza LL, Piazza A, Menozzi P, Mountain J. Reconstruction of human evolution: bringing together genetic, archeological and linguistic data. Proc Natl Acad Sci USA 1988; 85: 6002–06. Vandenbroeck K, Opdenakker G, Goris A, Murru R, Billiau A, Marrosu MG. Interferon-␥ gene polymorphism-associated risk for multiple sclerosis in Sardinia. Ann Neurol 1998; 44: 841–42.
Authors’ reply Sir—Koen Vandenbroek and colleagues cite their analysis of data about allele carriage rates in IFNG and the severity of rheumatoid arthritis. Although the primers encompass a similar number of nucleotides, those used in our study were not identical to the ones reported by Vandenbroek and colleagues.1 The typing of all patients and controls was not segregated by run number (about 62 samples per gel). The Genescan-500 ROX standards used were within lanes (ie, included in each sample), and allele sizing was derived with the ABI software (version 1.0.2) as described. With the exception of the longest allele, all allele sizes were identified in each of the three groups of study participants, and the difference in allele carriage rates between controls and patients with rheumatoid arthritis was seen in a second set of participants tested to confirm this finding. Although we have no immediate explanation for the discrepancy in allele assignment, this difference can be quickly resolved through an exchange of samples. However, a 2 bp change in
2191
For personal use only. Not to be reproduced without permission of The Lancet.
population. Conversely, the carriage rate of the 126 bp IFNG allele (13 CA repeat units) is 12% in the Canadian population, but amounts to 62–68% in our European populations. Since the white Canadian population is generally of recent European descent, a similar allele distribution to European populations would be anticipated. In view of this discrepancy, that the Sardinian IFNG carriage rates are similar to the continental European ones is remarkable. While the phylogeny of the Sardinian population is related to white ethnic origin, it is, in contrast to that of white Canadians, genetically distinct from that of other Europeans.4 If the Canadian IFNG alleles are reassigned by shifting them one position upward (eg, 122 bp→124 bp), a striking resemblance to the European carriage rates is generated. Our data substantiate the concern of Ollier that case-control mismatching of alleles might have led to the differences seen. If allele slippage occurred as we show in the table, it follows that IFNG alleles have been wrongly assigned in the control and mild rheumatoid arthritis groups, but were correctly assigned in the severe rheumatoid arthritis group in Khani-Hanjani and colleagues’ study. This finding would explain why significant differences were seen only on comparison of the latter with each of the former groups. Various errors in methods might account for allele slippage. Apart from those already mentioned by Ollier, the use of size standards between rather than within lanes during electrophoresis could have hampered accurate assignment of alleles as well. Our observations might jeopardise the validity of the findings of KhaniHanjani and colleagues, but IFNG remains an interesting candidate for conferring susceptibility to various autoimmune diseases. By use of a family-based association method (transmission disequilibrium test), we have shown significant distortion of parental transmission of this IFNG microsatellite polymorphism to male offspring with multiple sclerosis in Sardinia.3,5 Ollier reports an odds ratio of 1·8 (95% CI 1·0–3·1) for an IFNG
on Perinatal and Women’s Health, INSERM, Villejuif, France; Unit of Research on Reproduction, INSERM, Toulouse, France; and Division of Epidemiology, IFC, National Research Council, Pisa, Italy 1
2
Pochard F, Azoulay E, Grassin M. End-oflife decisions for newborn infants. Lancet 2000; 356: 946. Cuttini M, Nadai M, Kaminski M, et al. End-of-life decisions in neonatal intensive care: physicians’ self-reported practices in seven European countries. Lancet 2000; 355: 2112–18.
Interferon gamma gene in rheumatoid arthritis Sir—A Khani-Hanjani and colleagues (Sept 2, p 820)1 report a striking association between a dinucleotide polymorphism in the interferon gamma gene (IFNG) and rheumatoid arthritis in Canadians. In a commentary, W E R Ollier (Sept 2, p 783)2 raised the question of whether their allele typing could have slipped by one position, which would lead to the highly significant differences between cases and controls. We did an extensive analysis of this polymorphism in 1730 white European individuals.3 Since the primers used for amplification of the IFNG polymorphism in our and Khani-Hanjani and colleagues’ studies were identical, the numbers of CA repeat units per allele have been determined unambiguously. Direct comparison of IFNG allele carriage rates (ie, the proportion of individuals carrying a certain allele) between the two studies is possible and scientifically correct. We show in the table the IFNG allele carriage rates in German, Swedish, and Sardinian healthy individuals in our study, compared with those for the Canadian white healthy controls of Khani-Hanjani and colleagues. In Europeans, the 122 bp IFNG allele, which encompasses 11 CA repeat units, is rare. The carriage rate of the 122 bp allele was not higher than 1% in any population in our study. According to Khani-Hanjani and colleagues, the carriage rate of this allele is 80% in the white Canadian Population sample
Canadian (n=65)* Canadian (n=65)† Sardinian (n=413) German (n=367) Swedish (n=54)
Interferon gamma 122 bp (11 CA) 124 bp (12 CA) 126 bp (13 CA)
128 bp (14 CA) 130 bp (15 CA)
Other
52 (80%) 0 0 3 0
8 (12%) 8 (12%) 71 (17%) 46 (12·5%) 9 (17%)
0 0 0 14 (14%) 0
44 (68%) 52 (80%) 326 (79%) 248 (68%) 32 (59%)
8 (12%) 44 (68%) 257 (62%) 251 (68%) 35 (65%)
0 8 (12%) 26 (6%) 21 (6%) 6 (11%)
*Original assignment of alleles from Khani-Hanjani and colleagues. †Reassignment of alleles from Khani-Hanjani and colleagues.
Comparison of IFNG allele carriage rates in Canadian and European healthy population samples
THE LANCET • Vol 356 • December 23/30, 2000
single nucleotide polymorphism in male rheumatoid arthritis patients.2 Taken together, these data suggest that the IFNG, or a nearby locus, is functionally associated with sex bias in susceptibility to autoimmune diseases. *Koen Vandenbroeck, An Goris, Alfons Billiau, Ghislain Opdenakker, Cornelia Hardt *School of Pharmacy, Queen’s University of Belfast, Belfast BT9 7BL, UK; Rega Institute for Medical Research, University of Leuven, Leuven, Belgium; Institut für Humangenetik, Universitätsklinikum Essen, Essen, Germany (e-mail: [email protected]) 1
2
3
4
5
Khani-Hanjani A, Lacaille D, Hoar D, et al. Association between dinucleotide repeat in non-coding region of interferon-gamma gene and susceptibility to, and severity of, rheumatoid arthritis. Lancet 2000; 356: 820–25. Ollier WER. Role of interferon-␥ gene in rheumatoid arthritis? Lancet 2000; 356: 783–84. Goris A, Epplen C, Fiten P, et al. Analysis of an IFN-␥ gene (IFNG) polymorphism in multiple sclerosis in Europe: effect of population structure on association with disease. J Interferon Cytokine Res 1999; 19: 1037–46. Cavalli-Sforza LL, Piazza A, Menozzi P, Mountain J. Reconstruction of human evolution: bringing together genetic, archeological and linguistic data. Proc Natl Acad Sci USA 1988; 85: 6002–06. Vandenbroeck K, Opdenakker G, Goris A, Murru R, Billiau A, Marrosu MG. Interferon-␥ gene polymorphism-associated risk for multiple sclerosis in Sardinia. Ann Neurol 1998; 44: 841–42.
Authors’ reply Sir—Koen Vandenbroek and colleagues cite their analysis of data about allele carriage rates in IFNG and the severity of rheumatoid arthritis. Although the primers encompass a similar number of nucleotides, those used in our study were not identical to the ones reported by Vandenbroek and colleagues.1 The typing of all patients and controls was not segregated by run number (about 62 samples per gel). The Genescan-500 ROX standards used were within lanes (ie, included in each sample), and allele sizing was derived with the ABI software (version 1.0.2) as described. With the exception of the longest allele, all allele sizes were identified in each of the three groups of study participants, and the difference in allele carriage rates between controls and patients with rheumatoid arthritis was seen in a second set of participants tested to confirm this finding. Although we have no immediate explanation for the discrepancy in allele assignment, this difference can be quickly resolved through an exchange of samples. However, a 2 bp change in
2191
For personal use only. Not to be reproduced without permission of The Lancet.