Interferon‐regulated chemokines as biomarkers of systemic lupus erythematosus disease activity: A validation study

Objective

Systemic lupus erythematosus (SLE) is a complex autoimmune disease characterized by unpredictable flares of disease activity and irreversible damage to multiple organ systems. An earlier study showed that SLE patients carrying an interferon (IFN) gene expression signature in blood have elevated serum levels of IFN‐regulated chemokines. These chemokines were associated with more‐severe and active disease and showed promise as SLE disease activity biomarkers. This study was designed to validate IFN‐regulated chemokines as biomarkers of SLE disease activity in 267 SLE patients followed up longitudinally.

Methods

To validate the potential utility of serum chemokine levels as biomarkers of disease activity, we measured serum levels of CXCL10 (IFNγ‐inducible 10‐kd protein), CCL2 (monocyte chemotactic protein 1), and CCL19 (macrophage inflammatory protein 3β) in an independent cohort of 267 SLE patients followed up longitudinally over 1 year (1,166 total clinic visits).

Results

Serum chemokine levels correlated with lupus activity at the current visit (P = 2 × 10⁻¹⁰), rising at the time of SLE flare (P = 2 × 10⁻³) and decreasing as disease remitted (P = 1 × 10⁻³); they also performed better than the currently available laboratory tests. Chemokine levels measured at a single baseline visit in patients with a Systemic Lupus Erythematosus Disease Activity Index of ≤4 were predictive of lupus flare over the ensuing year (P = 1 × 10⁻⁴).

Conclusion

Monitoring serum chemokine levels in SLE may improve the assessment of current disease activity, the prediction of future disease flares, and the overall clinical decision‐making.

     

Linkage analysis of human systemic lupus erythematosus–related traits: A principal component approach

Objective

To identify chromosomal regions containing genes involved in the susceptibility to human systemic lupus erythematosus (SLE)–related traits.

Methods

In the context of a genome scan, we analyzed 101 SLE‐affected sibpairs with respect to dermatologic, renal, immunologic, hematologic, neurologic, cardiopulmonary, and arthritic characteristics. Phenotypes were redefined in terms of principal components, which are synthetic variables composed of linear combinations of the original traits. Using 9 principal components obtained from these 7 traits plus age at SLE onset and race, we analyzed genome scan data with the multivariate version of the new Haseman‐Elston regression model.

Results

The largest linkage for an individual trait was on chromosome 2 at 228 cM (immunologic; P = 0.00048). The most significant linkage to an individual principal component was on chromosome 4 at 208 cM (P = 0.00007). The largest multivariate linkage was on chromosome 7 at 69 cM (P = 0.0001). Of the individual organ systems, dermatologic involvement had the largest effect (P = 0.0083) at this peak at 7p13 on chromosome 7. Further analyses revealed that malar rash, a subtype of dermatologic involvement, was linked significantly (P = 0.00458) to this location.

Conclusion

These results provide evidence of the presence and locations of genes that are involved in the genetic susceptibility to SLE‐related traits in humans.

     

Identification of novel susceptibility genes in childhood‐onset systemic lupus erythematosus using a uniquely designed candidate gene pathway platform

Objective

Childhood‐onset systemic lupus erythematosus (SLE) presents a unique subgroup of patients for genetic study. The present study was undertaken to identify susceptibility genes contributing to SLE, using a novel candidate gene pathway microarray platform to investigate gene expression in patients with childhood‐onset SLE and both of their parents.

Methods

Utilizing bioinformatic tools, a platform of 9,412 single‐nucleotide polymorphisms (SNPs) from 1,204 genes was designed and validated. Molecular inversion probes and high‐throughput SNP technologies were used for assay development. Seven hundred fifty three subjects, corresponding to 251 full trios of childhood‐onset SLE families, were genotyped and analyzed using transmission disequilibrium testing (TDT) and multitest corrections.

Results

Family‐based TDT showed a significant association of SLE with a N673S polymorphism in the P‐selectin gene (SELP) (P = 5.74 × 10⁻⁶) and a C203S polymorphism in the interleukin‐1 receptor–associated kinase 1 gene (IRAK1) (P = 9.58 × 10⁻⁶). These 2 SNPs had a false discovery rate for multitest correction of <0.05, and therefore a >95% probability of being considered as proven. Furthermore, 7 additional SNPs showed q values of <0.5, suggesting association with SLE and providing a direction for followup studies. These additional genes notably included TNFRSF6 (Fas) and IRF5, supporting previous findings of their association with SLE pathogenesis.

Conclusion

SELP and IRAK1 were identified as novel SLE‐associated genes with a high degree of significance, suggesting new directions in understanding the pathogenesis of SLE. The overall design and results of this study demonstrate that the candidate gene pathway microarray platform used provides a novel and powerful approach that is generally applicable in identifying genetic foundations of complex diseases.

     

Association of a transmembrane polymorphism of Fcγ receptor IIb (FCGR2B) with systemic lupus erythematosus in Taiwanese patients

Objective

To investigate the possible association of the Fcγ receptor IIb (FcγRIIb) Ile/Thr187 transmembrane domain polymorphism, which significantly affects receptor signaling, with susceptibility to systemic lupus erythematosus (SLE) in Taiwanese patients.

Methods

We used matrix‐assisted laser desorption ionization−time‐of‐flight mass spectrometry to genotype 351 Taiwanese SLE patients and 372 age‐ and sex‐matched healthy individuals from the same geographic area. Allele frequencies and genotype distributions were compared between the patients and controls, both as an aggregate and as stratified by sex, autoantibody profile, and clinical parameters. A combined analysis was conducted to assess the FCGR2B Thr187 allele as a common risk factor in different ethnic populations.

Results

The minor Thr187 allele was significantly associated with SLE in Taiwanese subjects (P = 0.017, odds ratio [OR] 1.989 [95% confidence interval (95% CI) 1.119–3.553]). Interestingly, male SLE patients showed enrichment of the Thr/Thr187 genotype (24%; 7 of 29) as compared with female SLE patients (10%; 32 of 322) (P = 0.043, OR 2.884 [95% CI 1.028–7.839]). Additionally, SLE patients with Thr/Thr187 and Ile/Thr187 genotypes were more likely to have pleural effusions (P = 0.038, OR 1.874 [95% CI 1.033–3.411]) and anti‐SSA/Ro antibody production (P = 0.046, OR 2.221 [95% CI 1.013–4.897]). Combined analysis of 4 groups of Asian patients strongly supported the association of the FCGR2B Thr187 allele with the lupus phenotype (P = 0.000159).

Conclusion

The FcγRIIb transmembrane polymorphism is a strong disease susceptibility candidate in epistasis with other genetic effects in Taiwanese and other Asian populations. It may also play a more prominent role in male patients with SLE.

     

Assessment of atherosclerotic risk factors and endothelial function in children and young adults with pediatric‐onset systemic lupus erythematosus

Objective

To characterize atherosclerotic risk factors and endothelial function in pediatric‐onset systemic lupus erythematosus (SLE).

Methods

Lipoproteins, oxidized state, and autoantibodies to oxidized low‐density lipoprotein (Ox‐LDL) were assessed. Endothelial function was evaluated using brachial artery reactivity.

Results

Thirty‐three SLE patients and 30 controls were studied. SLE subjects had significantly decreased mean high‐density lipoprotein (HDL) cholesterol (41 mg/dl versus 51 mg/dl; P = 0.002) and apolipoprotein A‐I (97 mg/dl versus 199 mg/dl; P = 0.0004). There was no difference between groups in markers of oxidized state (including nitric oxide metabolites, isoprostanes, and Ox‐LDL) or in endothelial function. However, SLE subjects had increased median anti‐Ox‐LDL IgG (2,480 relative light units [RLU] versus 1,567 RLU; P = 0.0007) and IgG immune complexes with LDL (4,222 RLU versus 2,868 RLU; P = 0.002).

Conclusion

Pediatric SLE patients had significantly decreased levels of HDL cholesterol and apolipoprotein A‐I and elevated titers of autoantibodies to Ox‐LDL. Despite these atherosclerotic risk factors, SLE patients had normal measures of oxidized state and endothelial function.

     

A polymorphism within IL21R confers risk for systemic lupus erythematosus

Objective

Interleukin‐21 (IL‐21) is a member of the type I cytokine superfamily that has a variety of effects on the immune system, including B cell activation, plasma cell differentiation, and immunoglobulin production. The expression of IL‐21 receptor (IL‐21R) is reduced in the B cells of patients with systemic lupus erythematosus (SLE), while serum IL‐21 levels are increased both in lupus patients and in some murine lupus models. We recently reported that polymorphisms within the IL21 gene are associated with increased susceptibility to SLE. The aim of this study was to examine the genetic association between single‐nucleotide polymorphisms (SNPs) within IL21R and SLE.

Methods

We genotyped 17 SNPs in the IL21R gene in 2 large cohorts of lupus patients (a European‐derived cohort and a Hispanic cohort) and in ethnically matched healthy controls.

Results

We identified and confirmed the association between rs3093301 within the IL21R gene and SLE in the 2 cohorts (meta‐analysis odds ratio 1.16 [95% confidence interval 1.08–1.25], P = 1.0 × 10⁻⁴).

Conclusion

Our findings indicate that IL21R is a novel susceptibility gene for SLE.

     

Prevalence of conventional and lupus‐specific risk factors for cardiovascular disease in patients with systemic lupus erythematosus: A case–control study

Objective

Patients with systemic lupus erythematosus (SLE) are significantly more likely to experience a myocardial infarction or a stroke than age‐matched controls. We compared the prevalence of conventional and lupus‐specific risk factors in patients with SLE just before a cardiovascular event and in matched controls with SLE but no cardiovascular disease (CVD).

Methods

Twenty‐nine patients with SLE and CVD were enrolled. For each patient, 2 ethnically‐ and sex‐matched controls were obtained, 1 matched for age and 1 for SLE duration. Data regarding risk factors were collected for the time immediately preceding the relevant cardiovascular event, or at an equivalent time for controls.

Results

Patients' median age at event was 49 years (interquartile range 43–54 years) and mean disease duration was 12.0 ± 7.1 years. Patients with SLE and CVD were more likely than both age and duration controls to be treated for hypertension (P = 0.01 and P = 0.001, respectively) and to have elevated triglyceride levels (P = 0.05 and P = 0.01, respectively). Compared with duration controls, CVD patients were more likely to have lupus anticoagulant (P = 0.03), but less likely to be receiving treatment with hydroxychloroquine (P = 0.003). Compared with age controls, patients were more likely to be current smokers (P = 0.03), to have taken a mean dosage >7.5 mg/day of prednisolone (P = 0.04), and to have been treated with pulsed methylprednisolone (P = 0.03). In multivariable analysis, only hypertension treatment was an independent risk factor for CVD.

Conclusion

We identified significantly increased prevalence of some conventional and lupus‐specific risk factors in patients with SLE immediately before a CVD event compared with controls matched for age or disease duration.

     

Systemic lupus erythematosus in adults is associated with previous Epstein‐Barr virus exposure

Objective

The possible molecular mimicry of the Epstein‐Barr virus (EBV) peptide PPPGRRP by the peptide PPPGMRPP from Sm B′/B of the human spliceosome is consistent with the possibility that EBV infection is related to the origin of systemic lupus erythematosus (SLE) in some patients. Association of EBV exposure with SLE was therefore tested for and subsequently found in children and adolescents (odds ratio [OR] 49.9, 95% confidence interval [95% CI] 9.3–1,025, P < 10⁻¹¹). These results were confirmed at the level of EBV DNA (OR > 10, 95% CI 2.53–∞, P < 0.002). Much smaller seroconversion rate differences were found against 4 other herpes viruses. Herein, we extend these studies to adults and test the hypothesis that EBV infection is associated with adult SLE.

Methods

We selected 196 antinuclear antibody–positive adult SLE patients (age ≥20 years) and 2 age‐, race‐, and sex‐matched controls per patient. SLE patients and matched controls were tested for evidence of previous infection with EBV, cytomegalovirus (CMV), herpes simplex virus types 1 and 2 (HSV‐1 and HSV‐2), or varicella‐zoster virus (VZV) by standardized enzyme‐linked immunosorbent assays.

Results

Of the 196 lupus patients tested, all but 1 had been exposed to EBV, while 22 of the 392 controls did not have antibodies consistent with previous EBV exposure (OR 9.35, 95% CI 1.45–∞, P = 0.014). No differences were observed between SLE patients and controls in the seroconversion rate against CMV, HSV‐2, or VZV.

Conclusion

These new data from adults, along with the many suggestive features of EBV infection, are consistent with the contribution of this infection to the etiology of SLE.

     

Effects of a stress‐reduction program on psychological function,pain, and physical function of systemic lupus erythematosus patients: A randomized controlled trial

Objective

To assess the effects of a stress‐reduction program on pain, psychological function, and physical function in persons with systemic lupus erythematosus (SLE) who experience pain.

Methods

Ninety‐two SLE patients were assigned randomly to receive either biofeedback‐assisted cognitive‐behavioral treatment (BF/CBT), a symptom‐monitoring support (SMS) intervention, or usual medical care (UC) alone.

Results

BF/CBT participants had significantly greater reductions in pain and psychological dysfunction compared with the SMS group (pain, P = 0.044; psychological functioning, P < 0.001) and the UC group (pain, P = 0.028; psychological functioning, P < 0.001). BF/CBT had significantly greater improvement in perceived physical function compared with UC (P = 0.035), and improvement relative to SMS was marginally significant (P = 0.097). At a 9‐month followup evaluation, BF/CBT continued to exhibit relative benefit compared with UC in psychological functioning (P = 0.023).

Conclusion

This study supports the utility of a brief stress management program for short‐term improvement in pain, psychological function, and perceived physical function among persons with SLE who experience pain.

     

Linkage at 5q14.3–15 in multiplex systemic lupus erythematosus pedigrees stratified by autoimmune thyroid disease

Objective

To identify genetic effects potentially shared between systemic lupus erythematosus (SLE) and autoimmune thyroiditis (AITD).

Methods

Families from the Lupus Multiplex Registry and Repository were studied in which there was at least 1 member who had both SLE and AITD (Graves' disease or Hashimoto thyroiditis). Genome scan genotyping findings in these pedigrees were evaluated for evidence of genetic linkage, by the maximum‐likelihood parametric method. Nineteen pedigrees were used in the initial genome scan. Subsequently, an independent sample of 16 pedigrees was used to replicate findings.

Results

Studies of the first set of 19 pedigrees yielded a 2‐point parametric logarithm of odds (LOD) of 4.97, which was independently confirmed in the replication sample of 16 pedigrees (LOD 2.89). For all 35 pedigrees together, the 2‐point LOD was 7.86, under a dominant model used for screening with 90% penetrance and a disease allele frequency of 10%. The multipoint locus homogeneity LOD in the 35 pedigrees was 6.90 (α = 1.0) at 5q14.3–15 between D5S1725 and D5S1453, a 12‐cM interval, with the peak at D5S1462 at 96.64 cM (nonparametric linkage P = 0.00002). Fine mapping further confirmed the genetic linkage effect and narrowed the region likely to contain the gene to ∼5 Mb.

Conclusion

These results suggest that stratifying SLE pedigrees by the presence of other autoimmune disorders may facilitate the discovery of genes related to SLE and that 5q14.3–15 harbors a susceptibility gene shared by SLE and AITD.

     

Rheumatologists' quality of care for lupus: Comparison study between a university and county hospital

Objective

Quality of care received from physicians may impact health outcomes in systemic lupus erythematosus (SLE). We compared physician quality of care (P‐QOC), health‐related quality of life (HRQOL), and disease activity and damage between SLE patients receiving outpatient care at a university and a county rheumatology clinic.

Methods

Forty‐two university and 41 county clinic SLE patients provided data on 5 P‐QOC parameters and HRQOL health outcomes (Short Form 36 [SF‐36] health survey and EuroQol 5‐domain instrument [EQ‐5D]). Disease activity and damage were measured. Chi‐square analysis and Student's t‐tests were used for comparisons.

Results

Overall satisfaction with medical care was similar; however, university patients had higher P‐QOC scores than county patients in “perception of doctor's understanding of impact of SLE on patient's life” (P = 0.02) and “providing education/educational information to understand their disease” (P = 0.05). HRQOL, disease activity, and damage were similar in the 2 groups. Overall satisfaction with medical care was directly related SF‐36 general health (r = 0.34, P = 0.03) and EQ‐5D visual analog scale on state of health (r = 0.39, P = 0.01), and inversely related EQ‐5D pain (r = ?0.37, P = 0.02).

Conclusion

Patient perceptions of P‐QOC differed across the 2 centers despite similar demographics, clinical and HRQOL outcomes, and significant overlap in the physicians serving each clinic. Patients' overall satisfaction with medical care is associated with better HRQOL.

     

Anti‐C1q antibodies are associated with systemic lupus erythematosus global activity but not specifically with nephritis: A controlled study of 126 consecutive patients

Objective

Several studies have shown that anti‐C1q antibodies correlate with the occurrence and activity of nephritis in systemic lupus erythematosus (SLE). However, the significance of anti‐C1q antibodies in SLE has not been fully characterized. The aim of this study was to investigate associations between anti‐C1q antibodies and clinical and serologic parameters of SLE.

Methods

An enzyme‐linked immunosorbent assay kit was used to measure anti‐C1q antibodies in the sera of 126 consecutive patients with active SLE who were admitted to our university hospital from 2007 through 2009. Sera obtained from patients with high titers of anti‐C1q antibodies at the initial evaluation (n = 20) were reevaluated following treatment. Control sera were obtained from patients with other autoimmune diseases and from normal healthy control subjects (n = 20 in each group). Associations between anti‐C1q antibodies and clinical and serologic parameters of SLE were statistically analyzed.

Results

Anti‐C1q antibodies were detected in the sera of 79 of 126 patients with SLE. The prevalence and titers of anti‐C1q antibodies were significantly (P < 0.0001) higher in SLE patients than in patients with rheumatoid arthritis, patients with systemic sclerosis, and normal healthy control subjects. The prevalence and titers of anti‐C1q antibodies were not significantly associated with active lupus nephritis (P = 0.462 and P = 0.366, respectively). Anti‐C1q antibody titers were significantly correlated with SLE Disease Activity Index 2000 scores and the levels of anti–double‐stranded DNA antibodies, C3, C4, CH50, and C1q (P < 0.0001 for all comparisons). Moreover, anti‐C1q antibody titers significantly decreased as clinical disease was ameliorated following treatment (P = 0.00097).

Conclusion

These findings indicate that anti‐C1q antibodies are associated with SLE global activity but not specifically with active lupus nephritis.

     

Lymphopenia is a risk factor in the progression of carotid intima‐media thickness in juvenile‐onset systemic lupus erythematosus

Objective

To characterize the atherosclerotic risk factors in the progression of subclinical atherosclerosis in patients with juvenile‐onset systemic lupus erythematosus (SLE).

Methods

This was a longitudinal study of 76 patients with juvenile‐onset SLE. Carotid arteries were evaluated using ultrasonography at baseline and at followup visits at 6‐month intervals over the 6‐year study period. Clinical and laboratory parameters, disease activity, treatment, and traditional risk factors for atherosclerosis were evaluated. Data were analyzed using generalized estimating equations.

Results

The mean ± SD age of the patients at baseline was 15.01 ± 3.48 years and the mean ± SD disease duration was 2.65 ± 2.5 years. The mean ± SD duration of followup was 3.74 ± 1.24 years. The mean ± SD intima‐media thickness (IMT) of the common carotid arteries differed significantly between the patient and control (n = 38) groups (0.63 ± 0.08 mm versus 0.54 ± 0.06 mm; P < 0.001). The presence of lymphopenia at diagnosis and at baseline and higher levels of serum creatinine and C‐reactive protein at baseline were positively associated with progression of carotid IMT (P = 0.006, P = 0.043, P = 0.037, and P = 0.049, respectively). In multivariate analysis, only lymphopenia at baseline and at diagnosis were consistently associated with progression of IMT (P = 0.012 and P = 0.045, respectively).

Conclusion

In patients with juvenile‐onset SLE, some nontraditional risk factors for the progression of subclinical atherosclerosis were identified. Lymphopenia was the only independent risk factor for the progression of IMT. The pathogenic mechanisms warrant further investigation.

     

Effect of glutathione S‐TRANSFERASE polymorphisms and proximity to hazardous waste sites on time to systemic lupus erythematosus diagnosis: Results from the roxbury lupus project

Objective

The high prevalence of systemic lupus erythematosus (SLE) among African American women may be due to environmental exposures, genetic factors, or a combination of factors. Our goal was to assess association of residential proximity to hazardous waste sites and genetic variation in 3 glutathione Stransferase (GST) genes (GSTM1, GSTT1, and GSTP1) with age at diagnosis of SLE.

Methods

Residential histories were obtained by interviewing 93 SLE patients from 3 predominantly African American neighborhoods in Boston. Residential addresses and locations of 416 hazardous waste sites in the study area were geocoded using ArcView software. Time‐varying Cox models were used to study the effect of residential proximity to hazardous sites, GST genotype, and interaction between genotype and exposure in determining age at diagnosis.

Results

The prevalence of SLE among African American women in these neighborhoods was 3.56 SLE cases per 1,000. Homozygosity for GSTM1‐null and GSTP1 Ile105Val in combination was associated with earlier SLE diagnosis (P = 0.03), but there was no association with proximity to 416 hazardous sites. Available data on specific site contaminants suggested that, at a subset of 67 sites, there was higher potential risk for exposure to volatile organic compounds (P < 0.05 with Bonferroni correction). GST genotypes had a significant interaction with proximity (P = 0.03) in analyses limited to these sites.

Conclusion

There was no independent association between residential proximity to hazardous waste sites and the risk of earlier SLE diagnosis in this urban population. However, analysis of a limited number of sites indicated that the risk of earlier SLE associated with proximity to hazardous sites might be modulated by GST polymorphisms.

     

Association of the PD‐1.3A allele of the PDCD1 gene in patients with rheumatoid arthritis negative for rheumatoid factor and the shared epitope

Objective

To study the frequency of allele A of polymorphism PD‐1.3 of the PDCD1 gene in patients with rheumatoid arthritis (RA) and its subsets, based on the presence of rheumatoid factor (RF) and the shared epitope (SE) alleles.

Methods

A total of 1,175 patients with RA and 3,404 controls were genotyped for the PD‐1.3 A/G polymorphism, which previously was identified as being involved in susceptibility to systemic lupus erythematosus (SLE) in patients of European descent.

Results

We first detected a trend for association of allele A of the single‐nucleotide polymorphism PD‐1.3 with RA (P = 0.053, odds ratio [OR] 1.18, 95% confidence interval [95% CI] 0.99–1.41). To further clarify the nature of this association, patients with RA were divided into 4 groups according to the presence of RF and the SE alleles. Association was found only in the group of patients negative for both RF and the SE alleles (P = 0.0054 [corrected P = 0.015], OR 1.75, 95% CI 1.15–2.65).

Conclusion

Patients negative for both RF and the SE alleles showed association with the same allele that we previously identified as being involved in susceptibility to SLE. These results provide the first evidence of the involvement of the human PDCD1 gene in arthritis.

     

Association between physician volume and in‐hospital mortality in patients with systemic lupus erythematosus

Objective

Systemic lupus erythematosus (SLE) is an uncommon, clinically complex disease for which prior experience treating similar patients may be particularly important. This study was undertaken to determine if physician volume is associated with the outcome of hospitalization of patients with SLE.

Methods

Data on in‐hospital mortality in a population‐based sample of 15,509 patients with SLE ages 18 years or older who were hospitalized in 2000, 2001, or 2002 in New York or Pennsylvania were obtained from state health planning agencies. Risks of in‐hospital mortality were examined in relation to the average annual number of patients with SLE hospitalized by the admitting physician.

Results

Physician volume was inversely associated with mortality. Mortality was 4.1% among patients of physicians who treated <1 hospitalized patient with SLE per year, 3.5% among patients of physicians who treated 1–3 patients per year, and 2.5% among patients of physicians who treated >3 patients per year. After adjustment for demographic characteristics, severity of illness, and hospital characteristics, the mortality risk was 20% lower among patients in the middle category of physician volume (odds ratio 0.80, 95% confidence interval 0.66–0.96, P =0.02), and 42% lower among patients in the highest category of physician volume (odds ratio 0.58, 95% confidence interval 0.42–0.82, P = 0.002), compared with patients in the lowest category. The association was stronger among patients with nephritis (n = 2,673), for whom the adjusted odds of mortality were ∼60% lower among those in the highest category of physician volume.

Conclusion

Our findings indicate that higher disease‐specific physician volume is associated with lower risks of in‐hospital mortality in patients with SLE.

     

Association of increased interferon‐inducible gene expression with disease activity and lupus nephritis in patients with systemic lupus erythematosus

Objective

To study 5 type I interferon (IFN)–inducible genes (LY6E, OAS1, OASL, MX1, and ISG15) in patients with systemic lupus erythematosus (SLE) and to correlate expression levels with disease activity and/or clinical manifestations.

Methods

Peripheral blood cells were obtained from 48 SLE patients, 48 normal controls, and 22 rheumatic disease controls, and total RNA was extracted and reverse transcribed into complementary DNA. Gene expression levels were measured by real‐time polymerase chain reaction, standardized to a housekeeping gene, and summed to an IFN score. Disease activity was determined by the Safety of Estrogens in Lupus Erythematosus: National Assessment–Systemic Lupus Erythematosus Disease Activity Index (SELENA‐SLEDAI) composite.

Results

Each gene was highly expressed in SLE patients compared with normal controls (P ≤ 0.0003) or disease controls (P ≤ 0.0008 except for MX1). IFN scores were positively associated with the SELENA‐SLEDAI instrument score (P = 0.001), the SELENA‐SLEDAI flare score (P = 0.03), and the physician's global assessment score (P = 0.005). Compared with patients without nephritis, lupus nephritis patients had higher IFN scores (overall P < 0.0001), especially during active renal disease. IFN scores were weakly associated with neurologic manifestations. Elevated IFN scores were positively associated with the current presence of anti–double‐stranded DNA (anti‐dsDNA) antibodies (P = 0.007) or hypocomplementemia (P = 0.007). LY6E expression levels distinguished active from inactive lupus nephritis (P = 0.02) and were positively associated with proteinuria (P = 0.009).

Conclusion

The 5 IFN‐inducible genes were highly expressed in SLE patients, and increased levels were correlated with disease activity defined by several methods. IFN scores, or LY6E levels, were elevated in lupus nephritis patients, especially during active renal disease, and in patients with anti‐dsDNA antibody positivity and hypocomplementemia. IFN scores, or LY6E levels, may be useful as a biomarker for lupus nephritis therapy.

     

Defective DNA double‐strand break repair in pediatric systemic lupus erythematosus

Objective

Previous reports of cells from patients with systemic lupus erythematosus (SLE) note that repair of single‐strand breaks is delayed, and these lesions may be converted to double‐strand breaks (DSBs) at DNA replication forks. We undertook this study to assess the integrity of DSB recognition, signaling, and repair mechanisms in B lymphoblastoid cell lines derived from patients with pediatric SLE.

Methods

Nine assays were used to interrogate DSB repair and recognition in lymphoblastoid cell lines from patients with pediatric SLE, including the neutral comet assay (NCA), colony survival assay (CSA), irradiation‐induced foci formation for γ‐H2AX and 53BP1 proteins, kinetics of phosphorylation of structural maintenance of chromosomes protein 1 (SMC1), postirradiation bromodeoxyuridine incorporation to evaluate S phase checkpoint integrity, monoubiquitination of Fanconi protein D2, ATM protein expression, and non‐homologous DNA end joining protein expression and function.

Results

Three of the 9 assays revealed abnormal patterns of response to irradiation‐induced DNA damage. The NCA and CSA yielded aberrant results in the majority of SLE lymphoblastoid cell lines. Abnormal prolongation of SMC1 phosphorylation was also noted in 2 of 16 SLE lymphoblastoid cell lines.

Conclusion

Our data suggest that DSB repair is defective in some lymphoblastoid cell lines from pediatric patients with SLE, especially when assessed by both NCA and CSA. Since these studies are nonspecific, further studies of DNA repair and kinetics are indicated to further delineate the underlying pathogenesis of SLE and possibly identify therapeutic targets.

     

Dysregulated expression of CXCR4/CXCL12 in subsets of patients with systemic lupus erythematosus

Objective

CXCR4 is a chemokine with multiple effects on the immune system. In murine lupus models, we demonstrated that monocytes, neutrophils, and B cells overexpressed CXCR4 and that its ligand, CXCL12, was up‐regulated in diseased kidneys. We undertook this study to determine whether CXCR4 expression was increased in peripheral blood leukocytes from patients with systemic lupus erythematosus (SLE) and whether CXCL12 expression was increased in kidneys from patients with SLE.

Methods

Peripheral blood leukocytes from 31 SLE patients, 8 normal controls, and 9 patients with rheumatoid arthritis were prospectively analyzed by flow cytometry for CXCR4 expression. Biopsy samples (n = 14) from patients with lupus nephritis (LN) were immunostained with anti‐CXCL12 antibody.

Results

CD19+ B cells and CD4+ T cells from SLE patients displayed a >2‐fold increase (P = 0.0001) and >3‐fold increase (P < 0.0001), respectively, in median CXCR4 expression compared with that in controls (n = 7–8). Moreover, CXCR4 expression on B cells was 1.61‐fold higher in patients with SLE Disease Activity Index (SLEDAI) scores >10 (n = 8) than in patients with SLEDAI scores ≤10 (n = 16) (P = 0.0008), 1.71‐fold higher in patients with class IV LN (n = 5) than in patients with other classes of LN (n = 7) (P = 0.02), and 1.40‐fold higher in patients with active neuropsychiatric SLE (NPSLE) (n = 6) than in patients with inactive NPSLE (n = 18) (P = 0.01). CXCL12 was significantly up‐regulated in the tubules and glomeruli of kidneys in patients with LN (n = 14), with the percentage of positive cells correlating positively with the severity of LN.

Conclusion

CXCR4 appears to be up‐regulated in multiple leukocyte subsets in SLE patients. The heightened expression of CXCR4 on B cells in active NPSLE and of CXCL12 in nephritic kidneys suggests that the CXCR4/CXCL12 axis might be a potential therapeutic target for SLE patients with kidney and/or central nervous system involvement.

     

IgG glycan hydrolysis by endoglycosidase S diminishes the proinflammatory properties of immune complexes from patients with systemic lupus erythematosus: A possible new treatment?

Objective

Systemic lupus erythematosus (SLE) is an autoimmune disease with chronic or episodic inflammation in several organ systems, related to the presence of circulating and tissue‐deposited immune complexes (ICs) that stimulate leukocytes through Fcγ receptors (FcγR) with subsequent inflammation. Treatment with endoglycosidase S (EndoS), an IgG glycan–hydrolyzing bacterial enzyme from Streptococcus pyogenes, has shown beneficial effects in several experimental animal models of chronic inflammatory disease. This study was undertaken to investigate whether EndoS affects the proinflammatory properties of ICs and has the potential to be developed as a therapy for SLE.

Methods

ICs purified from SLE patients or RNA‐containing ICs formed in vitro were treated with EndoS and used in several assays reflecting different important features of SLE pathogenesis, such as phagocytosis by polymorphonuclear cells (PMNs) and plasmacytoid dendritic cells (PDCs), complement activation, and interferon‐α (IFNα) production by PDCs.

Results

EndoS treatment abolished all proinflammatory properties of the ICs investigated. This included FcγR‐mediated phagocytosis by PDCs (P = 0.001) and subsequent production of IFNα (P = 0.002), IC‐induced classical pathway of complement activation (P = 0.008), chemotaxis, and oxidative burst activity of PMNs (P = 0.002). EndoS treatment also had a direct effect on the molecular structure of ICs, causing decreased IC size and glycosylation.

Conclusion

Our findings indicate that EndoS treatment has prominent effects on several pathogenetically important IC‐mediated events, and suggest that EndoS has the potential to be developed as a novel therapy for SLE.