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Molecular analysis of HLA-D region genes in seropositive rheumatoid arthritis
D O McDaniel1, B O Barger, R T Acton
1Department of Medicine, School of Medicine, University of Albama, Birmingham.
Insights
Specific human leukocyte antigen (HLA) gene fragments are more common in Caucasians with rheumatoid arthritis (RA). These findings highlight potential genetic markers for RA susceptibility in this population.
Area of Science:
- Immunogenetics
- Rheumatology
- Molecular Biology
Background:
- Rheumatoid arthritis (RA) is a chronic autoimmune disease with a significant genetic component.
- Human Leukocyte Antigen (HLA) genes, particularly DRB1 and DQB1, are strongly associated with RA susceptibility.
- Restriction Fragment Length Polymorphisms (RFLPs) can identify variations within these genes.
Purpose of the Study:
- To investigate the association of specific HLA DRB1 and DQB1 BamHI RFLPs with seropositive rheumatoid arthritis in Caucasians.
- To determine the relative risk and frequency of these RFLPs in RA patients compared to healthy controls.
- To correlate identified RFLPs with specific HLA serotypes and map their location within the DRB1 gene.
Main Methods:
- Analysis of BamHI RFLPs in HLA DRB1 and DQB1 genes using DNA from Caucasian RA patients and healthy controls.
- Statistical analysis including calculation of p-values and relative risk (RR) for fragment frequencies.
- Correlation analysis between RFLPs and HLA serotypes (DR4, -7, -9, -w53, DR1, DQw1).
- Gene mapping of RFLPs using 5' and 3' specific probes for the DRB1 gene and a probe for the DRB4 (DRw53) gene.
- Segregation analysis of RFLPs with HLA phenotypes in family studies.
Main Results:
- Four DRB1 fragments (4.8, 5.2, 6.0, 7.0 kb) and one DQB1 fragment (3.2 kb) were significantly increased in Caucasian RA patients.
- The 6.0 kb DRB1 RFLP showed the highest association (p = 2 x 10(-5), RR = 8.0), followed by the 4.8, 5.2, 7.0 kb DRB1 fragments (p = 10(-3), RR = 5.0).
- The 3.2 kb DQB1 fragment was also significantly associated with RA (p = 10(-2), RR = 2.8).
- Specific RFLPs were correlated with distinct HLA serotypes, and mapping localized DRB RFLPs to the 5' and 3' ends of the DRB1 gene.
Conclusions:
- Specific HLA DRB1 and DQB1 BamHI RFLPs are significantly associated with seropositive rheumatoid arthritis in Caucasians.
- These RFLPs may serve as genetic markers for RA susceptibility.
- The findings provide further insight into the genetic basis of RA and the role of HLA complex variations.
Abstract:
Analysis of HLA DRB1 and DQB1 Bam HI RFLPs revealed four DRB1 (4.8, 5.2, 6.0 and 7.0 kb) fragments and a 3.2 kb DQB1 fragment to be significantly increased in Caucasians with seropositive RA compared to healthy individuals. The 4.8, 5.2 and 7.0 kb DRB1 fragments were found in 86.5% of RA patients and in 56% of the controls (p = 10(-3), relative risk (RR) = 5.0), while the 6.0 kb fragment was found in 79% of RA patients compared to 32% of controls (p = 2 x 10(-5), RR = 8.0). The 3.2 kb DQB1 fragment was observed in 63.5% of RA patients versus 38.0% of controls (p = 10(-2), RR = 2.8). Analysis of these fragments relative to HLA phenotypes revealed that the 4.8, 5.2 and 7.0 kb DRB1 fragments were strongly correlated with DR4, -7, -9 and -w53 serotypes, the 6.0 kb RFLP with DR4 and the 3.2 kb DQB1 fragment with DR1 and DQw1. Using probes specific for the 5' or 3' regions of the DRB1 gene, the 5.2 and 6.0 kb DRB RFLPs were mapped to the 5' end and the 4.8 and 7.0 kb RFLPs to the 3' end of the DRB1 gene. A probe generated from the second exon of the DRB4 (DRw53) gene recognized only the 5.2 and the 6.0 kb RFLPs corroborating the 5' location of these RFLPs. Family studies further confirmed that these RFLP's segregated with HLA phenotypes.