A functional haplotype of UBE2L3 confers risk for systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is an autoimmune disease with diverse clinical manifestations characterized by the development of pathogenic autoantibodies manifesting in inflammation of target organs such as the kidneys, skin and joints. Genome-wide association studies have identified genetic variants in the UBE2L3 region that are associated with SLE in subjects of European and Asian ancestry. UBE2L3 encodes an ubiquitin-conjugating enzyme, UBCH7, involved in cell proliferation and immune function. In this study, we sought to further characterize the genetic association in the region of UBE2L3 and use molecular methods to determine the functional effect of the risk haplotype. We identified significant associations between variants in the region of UBE2L3 and SLE in individuals of European and Asian ancestry that exceeded a Bonferroni-corrected threshold (P<1 × 10(-4)). A single risk haplotype was observed in all associated populations. Individuals harboring the risk haplotype display a significant increase in both UBE2L3 mRNA expression (P=0.0004) and UBCH7 protein expression (P=0.0068). The results suggest that variants carried on the SLE-associated UBE2L3 risk haplotype influence autoimmunity by modulating UBCH7 expression.

Role of MYH9 and APOL1 in African and non-African populations with lupus nephritis.

Systemic lupus erythematosus (SLE) is a complex autoimmune disease characterized by autoantibody production and organ damage. Lupus nephritis (LN) is one of the most severe manifestations of SLE. Multiple studies reported associations between renal diseases and variants in the non-muscle myosin heavy chain 9 (MYH9) and the neighboring apolipoprotein L 1 (APOL1) genes. We evaluated 167 variants spanning MYH9 for association with LN in a multiethnic sample. The two previously identified risk variants in APOL1 were also tested for association with LN in European-Americans (EAs) (N = 579) and African-Americans (AAs) (N = 407). Multiple peaks of association exceeding a Bonferroni corrected P-value of P < 2.03 × 10(-3) were observed between LN and MYH9 in EAs (N = 4620), with the most pronounced association at rs2157257 (P = 4.7 × 10(-4), odds ratio (OR) = 1.205). A modest effect with MYH9 was also detected in Gullah (rs8136069, P = 0.0019, OR = 2.304). No association between LN and MYH9 was found in AAs, Asians, Amerindians or Hispanics. This study provides the first investigation of MYH9 in LN in non-Africans and of APOL1 in LN in any population, and presents novel insight into the potential role of MYH9 in LN in EAs.

The role of HLA-DR-DQ haplotypes in variable antibody responses to anthrax vaccine adsorbed.

Host genetic variation, particularly within the human leukocyte antigen (HLA) loci, reportedly mediates heterogeneity in immune response to certain vaccines; however, no large study of genetic determinants of anthrax vaccine response has been described. We searched for associations between the immunoglobulin G antibody to protective antigen (AbPA) response to Anthrax Vaccine Adsorbed (AVA) in humans, and polymorphisms at HLA class I (HLA-A, -B, and -C) and class II (HLA-DRB1, -DQA1, -DQB1, -DPB1) loci. The study included 794 European-Americans and 200 African-Americans participating in a 43-month, double-blind and placebo-controlled clinical trial of AVA (clinicaltrials.gov identifier NCT00119067). Among European-Americans, genes from tightly linked HLA-DRB1, -DQA1, -DQB1 haplotypes displayed significant overall associations with longitudinal variation in AbPA levels at 4, 8, 26 and 30 weeks from baseline in response to vaccination with three or four doses of AVA (global P=6.53 × 10(-4)). In particular, carriage of the DRB1-DQA1-DQB1 haplotypes (*)1501-(*)0102-(*)0602 (P=1.17 × 10(-5)), (*)0101-(*)0101-(*)0501 (P=0.009) and (*)0102-(*)0101-(*)0501 (P=0.006) was associated with significantly lower AbPA levels. In carriers of two copies of these haplotypes, lower AbPA levels persisted following subsequent vaccinations. No significant associations were observed amongst African-Americans or for any HLA class I allele/haplotype. Further studies will be required to replicate these findings and to explore the role of host genetic variation outside of the HLA region.

Factors associated with arterial vascular events in PROFILE: a Multiethnic Lupus Cohort.

The objective of this study was to determine the factors associated with the occurrence of arterial vascular events in a multiethnic systemic lupus erythematosus (SLE) cohort. The PROFILE cohort, comprised SLE patients (n = 1333) of defined ethnicity from five different US institutions, was studied to determine demographic, clinical and biological variables associated with vascular events. An arterial vascular event (first episode) was either a myocardial infarction, angina pectoris and/or a vascular procedure for myocardial infarction, stroke, claudication and/or evidence of gangrene. Patient characteristics were analyzed by univariable and multivariable Cox proportional hazards regression analyses. One-hundred twenty-three (9.8%) patients had at least one incident arterial event. Age at cohort enrollment (HR = 1.04, 95% CI 1.03-1.06), smoking (HR = 2.20, 95% CI 1.40-3.46) and the CRP2* C alleles (HR = 1.91, 95% CI 1.04-3.49) were associated with a shorter time-to-the occurrence of arterial vascular events. Some clinical manifestations of disease activity were associated with a shorter time-to-occurrence [psychosis (HR = 2.21, 95% CI 1.10-4.44), seizures (HR = 1.85, 95% CI 1.00-3.24) and anaemia (HR = 1.83, 95% CI 1.02-3.31)], but others were not [arthritis (HR = 0.32, 95% CI 0.18-0.58)]. In conclusion, older patients, especially in the context of a predisposing environmental factor (smoking) and severe clinical manifestations, are at higher risk of having arterial vascular events. The genetic contribution of the variation at the CRP locus was not obscured by demographic or clinical variables. Awareness of these factors should lead to more effective management strategies of patients at risk for arterial vascular events.

Fcgamma receptors: structure, function and role as genetic risk factors in SLE.

Over 30 years ago, receptors for the Fc region of IgG (FcgammaR) were implicated in the pathogenesis of systemic lupus erythematosus (SLE). Since those pioneering studies, our knowledge of the structure and function of these FcgammaRs has increased dramatically. We now know that FcgammaR contributes to the regulation of acquired immunity and to the regulation of innate immune responses where FcgammaRs act as specific receptors for innate opsonins (CRP and SAP). Our understanding of the genomic architecture of the genes encoding the FcgammaR has also witnessed remarkable advances. Numerous functionally relevant single-nucleotide polymorphism (SNP) variants and copy number (CN) variants have been characterized in the FcgammaR genes. Many of these variants have also been shown to associate with risk to development of SLE and some have been associated with disease progression. This review will provide an overview of the FcgammaR in relation to SLE, including consideration of the role of genetic variants in FcgammaR in SLE pathogenesis. The difficulties in assessing genetic variation in these genes will be discussed. To enhance our understanding of the functional roles of these receptors in SLE, future research will need to integrate our knowledge of SNP variants, CN variants and the functional diversity of these receptors.

Identification of new SLE-associated genes with a two-step Bayesian study design.

In our earlier study, we utilized a Bayesian design to probe the association of approximately 1000 genes (approximately 10,000 single-nucleotide polymorphisms (SNPs)) with systemic lupus erythematosus (SLE) on a moderate number of trios of parents and children with SLE. Two genes associated with SLE, with a multitest-corrected false discovery rate (FDR) of <0.05, were identified, and a number of noteworthy genes with FDR of <0.8 were also found, pointing out a future direction for the study. In this report, using a large population of controls and adult- or childhood-onset SLE cases, we have extended the earlier investigation to explore the SLE association of 10 of these noteworthy genes (109 SNPs). We have found that seven of these genes exhibit a significant (FDR<0.05) association with SLE, both confirming some genes that have earlier been found to be associated with SLE (PTPN22 and IRF5) and presenting novel findings of genes (KLRG1, interleukin-16, protein tyrosine phosphatase receptor type T, toll-like receptor (TLR)8 and CASP10), which have not been reported earlier. The results signify that the two-step candidate pathway design is an efficient way to study the genetic foundations of complex diseases. Furthermore, the novel genes identified in this study point to new directions in both the diagnosis and the eventual treatment of this debilitating disease.

Replication of the BANK1 genetic association with systemic lupus erythematosus in a European-derived population.

Systemic lupus erythematosus (SLE) is an autoimmune disease with highly variable clinical presentation. Patients suffer from immunological abnormalities that target T-cell, B-cell and accessory cell functions. B cells are hyperactive in SLE patients. An adapter protein expressed in B cells called BANK1 (B-cell scaffold protein with ankyrin repeats) was reported in a previous study to be associated with SLE in a European population. The objective of this study was to assess the BANK1 genotype-phenotype association in an independent replication sample. We genotyped 38 single nucleotide polymorphisms (SNPs) in BANK1 on 1892 European-derived SLE patients and 2652 European-derived controls. The strongest associations with SLE and BANK1 were at rs17266594 (corrected P-value=1.97 x 10(-5), odds ratio (OR)=1.22, 95% CI 1.12-1.34) and rs10516487 (corrected P-value=2.59 x 10(-5), OR=1.22, 95% CI 1.11-1.34). Our findings suggest that the association is explained by these two SNPs, confirming previous reports that these polymorphisms contribute to the risk of developing lupus. Analysis of patient subsets enriched for hematological, immunological and renal ACR criteria or the levels of autoantibodies, such as anti-RNP A and anti-SmRNP, uncovers additional BANK1 associations. Our results suggest that BANK1 polymorphisms alter immune system development and function to increase the risk for developing lupus.

Complement receptor 2 polymorphisms associated with systemic lupus erythematosus modulate alternative splicing.

Genetic factors influence susceptibility to systemic lupus erythematosus (SLE). A recent family-based analysis in Caucasian and Chinese populations provided evidence for association of single-nucleotide polymorphisms (SNPs) in the complement receptor 2 (CR2/CD21) gene with SLE. Here we confirmed this result in a case-control analysis of an independent European-derived population including 2084 patients with SLE and 2853 healthy controls. A haplotype formed by the minor alleles of three CR2 SNPs (rs1048971, rs17615, rs4308977) showed significant association with decreased risk of SLE (30.4% in cases vs 32.6% in controls, P=0.016, OR=0.90 (0.82-0.98)). Two of these SNPs are in exon 10, directly 5' of an alternatively spliced exon preferentially expressed in follicular dendritic cells (FDC), and the third is in the alternatively spliced exon. Effects of these SNPs and a fourth SNP in exon 11 (rs17616) on alternative splicing were evaluated. We found that the minor alleles of these SNPs decreased splicing efficiency of exon 11 both in vitro and ex vivo. These findings further implicate CR2 in the pathogenesis of SLE and suggest that CR2 variants alter the maintenance of tolerance and autoantibody production in the secondary lymphoid tissues where B cells and FDCs interact.

Genetic associations of LYN with systemic lupus erythematosus.

We targeted LYN, a src-tyosine kinase involved in B-cell activation, in case-control association studies using populations of European-American, African-American and Korean subjects. Our combined European-derived population, consisting of 2463 independent cases and 3131 unrelated controls, shows significant association with rs6983130 in a female-only analysis with 2254 cases and 2228 controls (P=1.1 x 10(-4), odds ratio (OR)=0.81 (95% confidence interval: 0.73-0.90)). This single nucleotide polymorphism (SNP) is located in the 5' untranslated region within the first intron near the transcription initiation site of LYN. In addition, SNPs upstream of the first exon also show weak and sporadic association in subsets of the total European-American population. Multivariate logistic regression analysis implicates rs6983130 as a protective factor for systemic lupus erythematosus (SLE) susceptibility when anti-dsDNA, anti-chromatin, anti-52 kDa Ro or anti-Sm autoantibody status were used as covariates. Subset analysis of the European-American female cases by American College of Rheumatology classification criteria shows a reduction in the risk of hematological disorder with rs6983130 compared with cases without hematological disorders (P=1.5 x 10(-3), OR=0.75 (95% CI: 0.62-0.89)). None of the 90 SNPs tested show significant association with SLE in the African American or Korean populations. These results support an association of LYN with European-derived individuals with SLE, especially within autoantibody or clinical subsets.

Time to seizure occurrence and damage in PROFILE, a multi-ethnic systemic lupus erythematosus cohort.

The objective of this study was to determine risk factors predicting seizures and damage caused by seizures in a multi-ethnic systemic lupus erythematosus cohort (PROFILE) that includes systemic lupus erythematosus patients (n = 1295) from five different US institutions. Only patients with seizures after systemic lupus erythematosus diagnosis (incident) were included in the analyses of clinical seizures (80/1295, 6.2%), but all patients (prevalent and incident) were included in the analyses of damage caused by seizures (51/1295, 3.9%). We examined socioeconomic-demographic, clinical, and genetic variables predictive of clinical seizures and damage from seizures by Cox proportional hazard ratios (HR) and 95% confidence intervals (CI). Independent predictors of a shorter time to the occurrence of clinical seizures were younger age (HR = 1.0; 95% CI 0.9-1.0), having Hispanic-Texan ethnicity (HR = 2.7; 95% CI 1.3-5.7) or African-American ethnicity (HR = 1.8; 95% CI 1.0-3.1), and the previous occurrence of a cerebrovascular accident (HR = 3.3; 95% CI 1.6-7.1) or an episode of psychosis (HR = 2.4; 95% CI 1.1-5.0), whereas the previous occurrence of photosensitivity (HR = 0.5; 95% CI 0.3-0.9) was the only independent predictor of a longer time to the occurrence of clinical seizures. Independent predictors of a shorter time to the occurrence of damage caused by seizures were younger age (HR = 1.0; 95% CI 0.9-1.0), male gender (HR = 2.4; 95% CI 1.1-5.4), and the occurrence of a previous cerebrovascular accident (HR = 2.7; 95% CI 1.0-7.0) or an episode of psychosis (HR = 4.7; 95% CI 2.3-9.9). No allele from the candidate genes examined (HLA-DRB1, HLA-DQB1, FCGR2A, FCGR3A, or FCG3B) predicted clinical seizures or damage caused by seizures.

Interferon regulatory factor-5 is genetically associated with systemic lupus erythematosus in African Americans.

Increased expression of interferon (IFN)-inducible genes is implicated in the pathogenesis of systemic lupus erythematosus (SLE). One transcription factor responsible for regulating IFN, interferon regulatory factor-5 (IRF5), has been associated with SLE in genetic studies of Asian, Caucasian and Hispanic populations. We genotyped up to seven polymorphic loci in or near IRF5 in a total of 4870 African-American and Caucasian subjects (1829 SLE sporadic cases and 3041 controls) from two independent studies. Population-based case-control comparisons were performed using the Pearson's chi(2)-test statistics and haplotypes were inferred using HaploView. We observed significant novel associations with the IRF5 variants rs2004640 and rs3807306 in African Americans and replicated previously reported associations in Caucasians. While we identified risk haplotypes, the majority of haplotypic effects were accounted for by one SNP (rs3807306) in conditional analyses. We conclude that genetic variants of IRF5 associate with SLE in multiple populations, providing evidence that IRF5 is likely to be a crucial component in SLE pathogenesis among multiple ethnic groups.

Copy number variants in genetic susceptibility and severity of systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a systemic autoimmune disorder characterized by the presence of auto-antibodies to nuclear antigens, immune complex deposition, and subsequent tissue destruction. Early studies in twins suggested that SLE has, at least in part, a genetic basis, and a role for class II alleles in the major histocompatibility complex has been known for over 30 years. Through both linkage studies and candidate gene studies, numerous additional genetic risk factors have been identified. The recent publication of two SNP-based genome-wide association studies (GWAS) has resulted in the confirmation of a number of previously identified genetic risk loci and has identified new previously unappreciated loci conferring risk for development of SLE. A role for gene copy number variation (CNV) in SLE has also been appreciated through studies of the complement component 4 (C4) loci and more recent work in the IgG Fc receptor loci. The availability of large SNP-based GWAS datasets will undoubtedly lead to the genome-wide analysis and identification of copy number variants related to genetic susceptibility for development of SLE. We review current studies of CNV in SLE susceptibility that include reports of association between SLE and CNV in C4, IgG Fc receptors, TLR7, and CCL3L1.

Single-nucleotide polymorphisms in the C-reactive protein (CRP) gene promoter that affect transcription factor binding, alter transcriptional activity, and associate with differences in baseline serum CRP level.

To investigate whether functional polymorphisms exist in the C-reactive protein (CRP) gene, i.e., ones that contribute directly to differences in baseline CRP among individuals, we sequenced a 1,156-nucleotide-long stretch of the CRP gene promoter in 287 ostensibly healthy people. We identified two single-nucleotide polymorphisms (SNPs), a bi-allelic one at nucleotide -409 (G-->A), and a tri-allelic one at -390 (C-->T-->A), both resident within the hexameric core of transcription factor binding E-box elements. Electrophoretic mobility shift assays confirmed that the SNP within the sequence (-412)CACGTG(-407) (E-box 1) modulates transcription factor binding, and that the one within (-394)CACTTG(-389) (E-box 2) supports transcription factor binding only when the -390 T allele is present. The commonest of four E-box 1/E-box 2 haplotypes (-409G/-390T) identified in the population supported highest promoter activity in luciferase reporter assays, and the rarest one (-409A/-390T) supported the least. Importantly, serum CRP in people with these haplotypes reproduced this rank order, i.e., people with the -409G/-390T haplotype had the highest baseline serum CRP (mean +/- SEM 10.9 +/- 2.25 microg/ml) and people with the -409A/-390T haplotype had the lowest (5.01 +/- 1.56 microg/ml). Furthermore, haplotype-associated differences in baseline CRP were not due to differences in age, sex, or race, and were still apparent in people with no history of smoking. At least two other SNPs in the CRP promoter lie within E-box elements (-198 C-->T, E-box 4, and -861 T-->C, E-box 3), indicating that not only is the quality of E-box sites in CRP a major determinant of baseline CRP level, but also that the number of E-boxes may be important. These data confirm that the CRP promoter does encode functional polymorphisms, which should be considered when baseline CRP is being used as an indicator of clinical outcome. Ultimately, development of genetic tests to screen for CRP expression variants could allow categorization of healthy people into groups at high versus low future risk of inflammatory disease.

Systemic lupus erythematosus in a multiethnic US Cohort (LUMINA). XXX: association between C-reactive protein (CRP) gene polymorphisms and vascular events.

OBJECTIVES: To determine if a polymorphic GTn repeat in the intron of the C-reactive protein (CRP) gene associates with occurrence of vascular arterial events in systemic lupus erythematosus (SLE). METHODS: We performed a nested case-control study on the LUMINA cohort of 546 Hispanic, African-American and Caucasian SLE patients. Twenty-five patients who developed vascular arterial events (i.e. myocardial infarction, angina, coronary artery bypass graft surgery, stroke, claudication, gangrene or significant tissue loss and/or arterial peripheral thrombosis) after enrolment were selected as cases and 32 ethnically matched patients with no previous vascular arterial events served as controls. Their CRP gene GTn polymorphism and plasma CRP was determined. RESULTS: Patients with vascular events had more severe SLE and were more likely to have plasma CRP in the highest quintile of measured values. The overall distribution of GTn alleles for patients with vascular events had a greater number of the GT20 variant compared with controls [26.0% of alleles (13/50) vs 15.6% (10/64)]. This greater number of GT20 in patients with vascular events was observed for African-Americans [29.2% (7/24) vs 21.0% (8/38)] and Hispanics [33.0% (4/12) vs 0% (0/16)] but not for Caucasians [14.3% (2/14) vs 20.0% (2/10)]. For African-Americans and Hispanics combined (45 patients), the frequency of GT20 in those with vascular events (30.6%, 11/36) was significantly higher than in those without them (14.8%, 8/54) (P<0.05, one-tailed test for difference in proportions). When patients were categorized according to the number of GT20 alleles they carried (thus GT20/GT20, GT20/GTx or GTx/GTx, where x is any allele other than GT20), for both African-Americans and Hispanics the likelihood of vascular arterial events increased in proportion with the GT20 dose, and all GT20-homozygous patients developed vascular arterial events. CONCLUSIONS: The CRP GT20 variant is more likely to occur in African-American and Hispanic SLE patients than in Caucasian ones, and SLE patients carrying the GT20 allele are more likely to develop vascular arterial events.

Genetic risk factors for infection in patients with early rheumatoid arthritis.

We analyzed clinical and genetic factors contributing to infections in 457 subjects with early rheumatoid arthritis (RA) enrolled in a prospective, 1-year clinical trial of methotrexate and the TNF inhibitor etanercept. Subjects were genotyped for the following single nucleotide polymorphisms (SNPs): (TNF -308, -238, and + 488); lymphotoxin-alpha (LTA) (LTA + 249, + 365, and + 720); and Fc gamma receptors FCGR2A 131 H/R; FCGR3A 176 F/V; and FCGR3B NA 1/2 and genotypes were correlated with infections. At least one URI was noted in 52% of subjects (99/191) with the NA2/NA2 genotype of the neutrophil-specific FCGR3B gene, compared to 42% (77/181) of those with the NA1/NA2 genotype and 39% (23/59) of those with the NA1/NA1 genotype (P = 0.038). Urinary tract infection (UTI) was associated with the TNF -238 A (odds ratio(OR) 2.56, 95% confidence interval (CI) 1.05-6.25) and LTA +365 C (OR 1.73, 95% CI 1.07-2.79) alleles, and marginally with the FCGR3A F allele (OR 1.72, 95% CI 0.99-3.00). There was a striking linear correlation between UTI and the number of risk alleles defined by these three SNPs (P < 0.001), suggesting an additive effect on susceptibility. These findings have important implications for the role of genetics in susceptibility to bacterial and viral infections.

Genomic organization of classical human low-affinity Fcgamma receptor genes.

The classical low-affinity Fcgamma receptor genes (FcgammaRIIA, B, C and FcgammaRIIIA, B) are located on chromosome 1q23, a region that shows strong linkage with human systemic lupus erythematosus (SLE) in several genome-wide scans, and family-based association between FcgammaRIIIA and SLE is now established. High homology among the Fcgamma receptor genes, however, has hampered further study of this region. We have used a human bacterial artificial chromosome (BAC) library to determine the order and orientation of these Fcgamma receptor genes and have sequenced the very highly homologous 5' region (including 3.4 kb of the promoter and the 8 kb from exon 1 to exon 3) of the FcgammaRIIB and FcgammaRIIC genes to enable study of their unique single nucleotide polymorphisms (SNP). We have utilized these data to characterize a linked set of three coding region SNPs in the FcgammaRIIC exon 3 (EC1) that includes the stop codon SNP, which provides an important insight into natural killer cell function. Together, these data provide the basis for the study of additional SNPs in FcgammaR genes in SLE disease susceptibility.

Conservation of FcepsilonRI gamma chain coding region in normals and in SLE patients.

High-frequency single nucleotide polymorphism (SNP) alleles are useful in mapping genes responsible for disease susceptibility. Functionally, Fcgamma receptors (FcgammaR) have been implicated in autoimmune disease, and the gene encoding the signaling element for several FcgammaR, Fc-epsilon-receptor gamma-chain (FcepsilonRIgamma), has several SNPs in the immunoreceptor tyrosine activation motif (ITAM) recorded in GenBank. Direct sequencing of the FcepsilonRIgamma coding region found potentially polymorphic sites in the 5'-->3' direction in control donors, which were not confirmed in the reverse direction (n = 66), and further exploration of 80 SLE patients revealed no non-synonymous SNPs. One normal donor was heterozygous for a non-synonymous SNP at nt 38 which changed the fifth codon from valine (GTG) to methionine (ATG). Although the EST databases suggest candidate SNPs, insertions and deletions, these appear to be artifacts, most probably due to secondary structure. The coding region of FcepsilonRIgamma shows a remarkable absence of nucleotide diversity. Either as yet unidentified regulatory elements of FcepsilonRIgamma or other genes in the region of human chromosome 1q23 are likely to be systemic lupus erythematosus disease susceptibility and severity genes.

Novel single nucleotide polymorphisms in the distal IL-10 promoter affect IL-10 production and enhance the risk of systemic lupus erythematosus.

Family studies of first-degree relatives and analysis of twins indicate that as much as 75% of the differences in quantitative IL-10 production in man derive from heritable genetic factors. Studies of single nucleotide polymorphisms (SNP) in the proximal 1.0 kb of the IL-10 promoter have yielded inconsistent association with IL-10 production and variable results in promoter-reporter studies. However, in normal donors, an association of quantitative production with certain alleles of the IL-10.R short tandem repeat polymorphism at -4.0 kb suggested that SNPs in the more distal promoter might be informative. We have identified seven novel SNP sites in the genomic sequence of the first 4 kb of the IL-10 promoter region 5' to the ATG start site from Caucasian individuals with either a high or a low IL-10 production phenotype. We have also identified eight SNP haplotypes in the distal promoter that segregate with significant differences in quantitative IL-10 production in normal donors. These SNPs are significantly associated with systemic lupus erythematosus in African-Americans and may define one component of the genetic susceptibility to systemic lupus erythematosus in this group.

C-reactive protein binding to FcgammaRIIa on human monocytes and neutrophils is allele-specific.

C-reactive protein (CRP) is involved in host defense, regulation of inflammation, and modulation of autoimmune disease. Although the presence of receptors for CRP on phagocytes has been inferred for years, their identity was determined only recently. FcgammaRIa, the high-affinity IgG receptor, binds CRP with low affinity, whereas FcgammaRIIa, the low-affinity IgG receptor, binds CRP with high affinity. Because the single nucleotide polymorphism in FcgammaRIIA - which encodes histidine or arginine at position 131 - strongly influences IgG2 binding, we determined this polymorphism's effect on CRP binding. CRP bound with high avidity to monocytes and neutrophils from FcgammaRIIA R-131 homozygotes, and binding was inhibited by the R-specific mAb 41H16. CRP showed decreased binding to cells from FcgammaRIIA H-131 homozygotes (which bind IgG2 with high affinity). However, IFN-gamma enhanced FcgammaRI expression by H-131 monocytes and increased CRP binding. FcgammaRIIa heterozygotes showed intermediate binding. CRP initiated increases in [Ca(2+)](i) in PMN from R-131, but not from H-131 homozygotes. These data provide direct genetic evidence for FcgammaRIIa as the functional, high-affinity CRP receptor on leukocytes while emphasizing the reciprocal relationship between IgG and CRP binding avidities. This counterbalance may affect the contribution of FcgammaRIIA alleles to host defense and autoimmunity.

Differential expression of integrin subunits in canine knee ligament fibroblasts.

A method for measuring the expression of integrin subunits on the cell surface of knee ligament fibroblasts was developed with use of flow cytometry and immunofluorescence. The ligament cells exhibited uniform size and density, as shown by forward and side-scatter properties, and showed minimal nonspecific binding of isotype control antibodies compared with unstained cells. All cells expressed the alpha5 integrin subunit; lateral collateral ligament cells stained with antibody to alpha5 showed a mean fluorescence intensity 2-fold higher than that of medial collateral ligament cells, 1.5-fold higher than that of posterior cruciate ligament cells, and 3-fold higher than that of anterior cruciate ligament cells, indicating a greater expression of the alpha5 subunit by lateral collateral ligament cells than by medial collateral, posterior cruciate, and anterior cruciate ligament cells. All cells expressed the beta1 integrin subunit; the expression by posterior cruciate ligament cells was 3-fold higher than that by medial collateral ligament or lateral collateral ligament cells and 5-fold higher than that by anterior cruciate ligament cells. All cells expressed the beta3 integrin subunit; the expression by posterior cruciate ligament cells was 1.5, 3, and 4.5-fold greater than that by lateral collateral, anterior cruciate, and medial collateral ligament cells, respectively. Our data suggest there is a differential expression of integrin subunits in knee ligament fibroblasts, and this in part may explain differences in their attachment and adherence to extracellular matrix molecules.