Reticulocytes bearing C4d as biomarkers of disease activity for systemic lupus erythematosus

OBJECTIVE: There is an urgent need for biomarkers with which to monitor disease activity in patients with systemic lupus erythematosus (SLE). We recently showed that abnormal levels of C4d, an activation-derived fragment of complement component C4, are deposited on the surface of erythrocytes from patients with SLE. This study focused on reticulocytes, the youngest and shortest-lived erythrocytes (lifespan 24-48 hours), with the objective of testing our hypothesis that when reticulocytes emerge from the bone marrow, they are immediately exposed to and acquire C4d at levels proportionate to the extent of complement activation at that time, thereby reflecting disease activity in SLE. METHODS: We conducted a cross-sectional study of 156 patients with SLE, 140 patients with other diseases, and 159 healthy controls. Levels of C4d on the surface of reticulocytes were examined using a 2-color flow cytometric assay. The results were analyzed for correlations with SLE disease activity. RESULTS: A wide range of increased levels of reticulocyte C4d was specifically detected in SLE patients. These levels fluctuated in SLE patients and correlated with clinical disease activity, as determined by the Safety of Estrogens in Lupus Erythematosus: National Assessment (SELENA) version of the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) and the Systemic Lupus Activity Measure (SLAM). Specifically, in cross-sectional analyses, patients with reticulocyte C4d levels in the highest quartile compared with those in the lowest quartile had significantly higher SELENA-SLEDAI (P = 0.00002) and SLAM (P = 0.02) scores. Longitudinal observation demonstrated that the reticulocyte C4d levels changed in relation to the clinical course in individual patients. CONCLUSION: These findings support our hypothesis that C4d-bearing reticulocytes may serve as biomarkers of disease activity in patients with SLE.     

Complement split product C3d as an indicator of disease activity in systemic lupus erythematosus

Summary In order to investigate, if complement levels can be used as an indicator of clinical activity in systemic lupus erythematosus (SLE), levels of C3, C4, CH50, and C3d were measured in 79 patients, 41 with inactive, 31 with moderately active and 7 with severely active disease. Our study shows that C3d, and particularly the C3d/C3 ratio, provide sensitive markers for disease activity in SLE. Since C3d is a direct measurement of complement turnover, it reflects complement activation better than C3, C4 and CH50.     

Platelet C4d is highly specific for systemic lupus erythematosus

OBJECTIVE: Complement-activation product C4d is deposited on normal erythrocytes, while abnormal levels have been observed on the surface of erythrocytes of patients with systemic lupus erythematosus (SLE). This study examines whether C4d also deposits on human platelet surfaces, and whether platelet-bound C4d may provide a biomarker for SLE. METHODS: We conducted a cross-sectional study of 105 patients with SLE, 115 patients with other diseases, and 100 healthy controls. Levels of C4d on the surface of platelets were examined by flow cytometry and scanning confocal microscopy. Statistical analyses were performed to determine the clinical variables associated with platelet C4d. RESULTS: Abnormal levels of platelet C4d were found to be highly specific for SLE. Platelet C4d was detected in 18% of patients with SLE, being 100% specific for a diagnosis of SLE compared with healthy controls and 98% specific for SLE compared with patients with other diseases (P < 0.0001). In addition, platelet C4d was significantly associated with positivity for lupus anticoagulant (P < 0.0001) and anticardiolipin antibodies of the IgG (P = 0.035) or the IgM (P = 0.016) isotype. Platelet C4d was also significantly associated with SLE disease activity according to the SLE Disease Activity Index (P = 0.039), low serum C4 (P = 0.046), an elevated erythrocyte sedimentation rate (P = 0.006), and abnormal levels of C4d on erythrocytes (P < 0.0001). CONCLUSION: This observation suggests that platelet-bound C4d may be a useful biomarker for SLE and may be a clue to the pathogenic mechanisms responsible for the myriad thrombotic and vascular complications of lupus associated with antiphospholipid antibodies.     

The use of C3d as a means of monitoring clinical activity in systemic lupus erythematosus and rheumatoid arthritis.

Plasma samples from 44 patients with systemic lupus erythematosus (SLE) and 43 with rheumatoid arthritis (RA) were assayed for C3d, a breakdown product of the third component of complement (C3), which was also measured in parallel. Levels of C3d varied in direct proportion with disease activity in RA, whereas C3 showed little change. Although C3d values also increased with worsening clinical condition in SLE, this trend was not considered to be sufficiently clear to be useful and did not provide any advantage over the routinely performed C3 assay.     

Serum C4 concentration in the monitoring of systemic lupus erythematosus: requirement for C4 allotyping

Summary We have examined the influence of genetic and other factors on the serum concentration of the fourth component of complement (C4) in four patients with systemic lupus erythematosus (SLE) studied over 3 to 8 years. Complement allotyping was performed to determine the number of C4 null genes in each patient. Two patients with C4 null genes had relatively low serum C4 concentrations with normal serum anti-DNA binding and no evidence of active disease. By contrast two patients without null alleles appeared to be consuming C4 when the serum C4 concentrations were within the conventional reference range. We therefore propose the use of appropriate reference ranges adjusted for the number of null alleles. Such adjusted reference ranges may improve the utility of serum C4 concentration in monitoring disease activity.     

Patterns of disease activity in systemic lupus erythematosus

OBJECTIVE: To describe patterns of systemic lupus (SLE) disease activity over time. METHODS: Disease activity was measured in a prospective cohort of 204 consecutive SLE patients followed up quarterly for 2.0-7.5 years (911 person-years of followup). Physician's global assessment (PGA) and modified SLE Disease Activity Index (M-SLEDAI; omitting serology) scores were plotted against time for each patient. Definitions for disease activity patterns were developed by consensus using these plots, and the proportion of total follow-up time represented by each pattern was calculated. RESULTS: Three patterns of SLE activity were apparent: relapsing-remitting (RR), chronic active (CA), and long quiescent (LQ). The CA pattern was the most frequent for both the PGA and M-SLEDAI, representing 58% and 40% of total person-years, respectively. The least common pattern was LQ (PGA 16%, M-SLEDAI 25%), while the RR pattern was intermediate in frequency (PGA 26%, M-SLEDAI 35%). Average disease activity during RR periods tended to be mild, while that during CA periods was more likely to be moderately severe. The most common discrepancy between instruments was that the PGA depicted CA when the M-SLEDAI showed an RR pattern. The M-SLEDAI did not appear to capture mild degrees of activity. CONCLUSION: SLE activity was readily classified into 1 of 3 patterns: RR, CA, or LQ. The CA pattern was most common, suggesting that significant morbidity may arise from persistent disease activity. These findings may have important implications regarding the choice of outcome measures in SLE clinical trials, since comparison of flare rates may not account for chronic disease activity.     

Measurement of erythrocyte C4d and complement receptor 1 in systemic lupus erythematosus

OBJECTIVE: C4-derived activation fragments are the only complement ligands present on the surfaces of normal erythrocytes. The significance of this observation is unknown, and the role of erythrocyte-bound C4 (E-C4) in human disease has not been explored. More than any other human disease, the pathogenesis of systemic lupus erythematosus (SLE) has been characterized by defects in clearance of complement-bearing immune complexes via erythrocytes expressing complement receptor 1 (CR1). This study was undertaken to determine whether these functional defects might be reflected by abnormal patterns of E-C4 and E-CR1 expression on erythrocytes of patients with SLE. METHODS: We conducted a cross-sectional study of 100 patients with SLE, 133 patients with other diseases, and 84 healthy controls. Erythrocytes were characterized by indirect immunofluorescence and by flow cytometry for determination of levels of C4d and CR1. RESULTS: Patients with SLE had higher levels of E-C4d and lower levels of E-CR1 than did patients with other diseases (P < or = 0.001) or healthy controls (P < or = 0.001). The test was 81% sensitive and 91% specific for SLE versus healthy controls and 72% sensitive and 79% specific for SLE versus other diseases, and it had an overall negative predictive value of 92%. CONCLUSION: This is the first report of abnormal levels of E-C4d in human disease. We found that abnormally high levels of E-C4d and low levels of E-CR1 are characteristic of SLE, and combined measurement of the 2 molecules has high diagnostic sensitivity and specificity for lupus. Determination of E-C4d/E-CR1 levels may be a useful addition to current tests and criteria for SLE diagnosis.     

Erythrocyte aggregation in patients with systemic lupus erythematosus

Systemic Lupus Erythematosus (SLE) is an autoimmune, chronic inflammatory, non-organ specific disease. SLE patients present a high prevalence of thrombotic and arteriosclerotic disease. The aim of the present work was to study the erythrocyte aggregation kinetics, and the effect of plasma factors, namely, immunoglobulin and fibrinogen concentration, as well as cell factors such as deformability and erythrocyte membrane lipid fluidity on the erythrocyte aggregation, in SLE patients and healthy controls. The results show that SLE patients red blood cells aggregate at higher rate and the aggregates size are also greater than controls due to an increase of immunoglobulin and plasma fibrinogen. The negative correlation between aggregation parameters and rigidity index could point out that the altered deformability diminishes the erythrocyte aggregation. Correlation between rigidity index and anisotropy suggests that the decrease of membrane lipid fluidity might be a cause of deformability decrease. The erythrocyte aggregation increase in these patients could induce a decreased flow that might contribute to the thromboembolic process present in SLE patients.     

Cigarette smoking and disease activity in systemic lupus erythematosus

OBJECTIVE: To investigate the effect of cigarette smoking on disease activity and cumulative organ damage in systemic lupus erythematosus (SLE). Methods. Extensive clinical and demographic variables, including current and previous cigarette smoking, were collected from 111 SLE patients using a detailed interview-administered questionnaire. Disease activity was estimated with the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI). Cumulative organ damage was measured by the Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index (SLICC/ACR). Smoking status was correlated with disease activity and cumulative organ damage, while statistically adjusting for the individual effects of potentially confounding demographic and clinical variables using analysis of variance followed by Fisher's least significant difference method. Results. Current smokers demonstrated significantly higher (p < 0.001) SLEDAI scores (15.6 +/- 7.8) than ex-smokers (9.63 +/- 6.00), and never smokers (9.03 +/- 5.75). This association remained significant (p = 0.001) after adjusting for all covariates, including ethnicity, education level, income level, alcohol use, age of onset of SLE, current age, mean duration of SLE, marital status, and hydroxychloroquine therapy. Current smokers also demonstrated significantly (p = 0.003) higher scores for both the neurological and non-neurological components of SLEDAI. There was no significant difference in the SLICC/ACR scores across the various smoking groups, although there was a trend for more severe disease in current smokers. Conclusion. Cigarette smoking is associated with increased disease activity in SLE. These data further establish the association of SLE with cigarette smoking, and suggest that individuals with SLE should avoid all exposure to tobacco products.     

Health status and disease activity in systemic lupus erythematosus

The physical manifestations of disease activity and health status of patients with systematic lupus erythematosus (SLE) were measured in this study. Forty-nine patients completed the Arthritis Impact Measurement Scale (AIMS) and consented to examination for physical features of SLE documented by a Clinical Activity Index (CAI). Results showed a mean score of 22 on the AIMS and 6.6 on the CAI. The total scores for each measure were significantly correlated (r = 0.55, p < 0.001), indicating a relationship between health status and clinical disease activity. The total score for CAI was significantly correlated with the physical activity, pain, and depression subscales of health status. The total score for health status was significantly correlated with mucocutaneous, musculoskeletal, and general features of CAI. Within scale correlations were also found. Mucocutaneous aspects of disease activity were significantly correlated with pain and depression. Musculoskeletal features were significantly correlated with physical activity and pain. General aspects of SLE, including fatigue, were significantly correlated with physical activity. The study concludes that there is a relationship between certain physical features of SLE and key components of health status.     

Value of anticardiolipin antibodies for monitoring disease activity in systemic lupus erythematosus and other rheumatic diseases

Summary The prevalence of anticardiolipin antibodies in active systemic lupus erythematosus (SLE) was compared with that in inactive SLE and other rheumatic and non-rheumatic diseases to determine the value of these autoantibodies in monitoring rheumatic diseases. Pairs of IgG- and IgM-aCL were measured by ELISA in 173 consecutive hospitalised patients, including 141 with rheumatic diseases (18 active SLE, 21 inactive SLE, 19 rheumatoid arthritis, 13 reactive arthritis, 7 other spondyloarthropathies, 16 vasculitis, 47 other autoimmune diseases) and 32 non-rheumatic controls. A further 101 aCL pairs were determined during follow-up in 19 patients with SLE. Serum concentrations were analysed with respect to SLE activity and compared between the different patient groups. IgG- and IgM-aCL levels in excess of 10 GPL and 9 MPL respectively were considered positive. 30.6% of all patients (53/173) were found to be positive for IgG-aCL, as against only 9.8% (17/173) for IgM-aCL. IgG-aCL serum levels in active SLE differed significantly from all other groups, including inactive SLE (all p<0.005). Median IgM-aCL levels were below the cut off point in all groups, although measurable values were obtained almost exclusively in active SLE and RA. In this study IgM-aCL measurement was of less value in monitoring rheumatic diseases. IgG-aCL positivity in SLE was associated with a significantly higher odds ratio (OR) for active disease (OR 16.0, 95% confidence interval: 2.8–90.0). The results show that disease activity in SLE was accompanied by significantly increased IgG-aCL, whereas no elevation was found in other diseases. This parameter may therefore be useful in monitoring SLE activity.     

Hypercatabolism of C3 and C4 in active and inactive systemic lupus erythematosus.

The metabolism of the complement proteins C3 and C4 was studied in patients with active and inactive systemic lupus erythematosus (SLE) using highly purified, functionally active preparations. Nine patients with active and eight with inactive SLE were examined and 11 control subjects. There was a significant difference in the level of double stranded DNA antibodies, immune complexes, and serum C4 between the patients with active and inactive disease. Seven of 16 patients had detectable C4 null alleles and four had low serum concentrations of complement inhibitors. Each subject received approximately 370 kBq [125I]C4 and 93 kBq [131I]C3. Both patient groups showed significant C4 hypercatabolism compared with control subjects, but there was no difference between patients with active and inactive disease. The fractional catabolic rate (FCR) of C4 was comparable in subjects with and without detectable C4 null alleles. C4 production rate was significantly lower in patients with active SLE than in control subjects. There was significant C3 hypercatabolism for both patient groups, but C3 production was normal. An inverse correlation was observed between serum concentration and FCR. There was a highly significant correlation between C4 FCR and C3 FCR for control subjects + patients with inactive disease but not for those with active SLE combined with either controls or the inactive group. We conclude that complement hypercatabolism occurs in SLE irrespective of disease activity and that accelerated turnover does not account completely for the low C4 concentration observed in patients with active disease. This low concentration also results from impaired plasma production, which could reflect a high incidence of C4 null alleles or (inhibitory) factors associated with pathological complement activation, or both. Low C4 production could affect generation of the C3 converting enzyme C4b, 2b and thus influence proceeding complement activation.     

C4 concentrations and C4 deficiency alleles in systemic lupus erythematosus.

In a study of 66 patients with systemic lupus erythematosus (SLE) and 80 controls it was found that the presence of two deficiency (null) alleles of C4 had a significant effect on mean C4 concentrations in serum. In six controls who each had two C4 null alleles the mean C4 concentration in serum was 56% lower than in 43 controls without C4 null alleles; the nadir of the C4 concentration in four patients with SLE with two null alleles was also lower by a mean of 55% than in 32 patients who did not have null alleles. Reduced production of C4 allotypes in subjects with two null alleles may be an important determinant of total C4 concentration in patients with SLE. For optimal interpretation of C4 concentrations in SLE, C4 allotyping appears to be indicated, particularly to identify patients who have two null alleles of C4.     

Selective C4 deficiency, systemic lupus erythematosus, and Whipple''s disease.

A 45-year-old female with selective deficiency of C4 and systemic lupus erythematosus developed puzzling gastrointestinal and systemic symptoms in the last 6 months of her life. Extensive investigation of the gastrointestinal tract did not yield any diagnosis, and the patient died shortly afterwards. Autopsy revealed evidence of a typical Whipple's disease of the jejunum and lymph nodes. This association has not been previously described. The disease is reviewed with emphasis on its being an opportunistic infection in an immunosuppressed host with a complement deficiency and SLE.     

Depression predicts self-reported disease activity in systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is an autoimmune disease that can significantly impact both physiological and psychological functioning. In order to examine the relationship between psychological functioning and disease activity in SLE, we administered instruments that collected sociodemographic information and measured indices of disease activity and psychosocial functioning from 125 adult Hispanic and White patients with SLE. Patients were recruited from four healthcare settings in the greater Southern California area. Both cross-sectional and longitudinal relationships between depression and disease activity were evaluated. Cross-sectional findings revealed that depression and ethnicity were independently correlated with self-reported disease activity. Longitudinally, depression alone predicted self-reported disease activity. These data suggest that depression may play a significant role in the health status of SLE patients and serve as an important target for clinical intervention.     

Urinary soluble VCAM-1 in systemic lupus erythematosus: a clinical marker for monitoring disease activity and damage

OBJECTIVE: To determine the urinary levels of soluble vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) in patients with systemic lupus erythematosus (SLE) and to assess their relationship with clinical and laboratory features and the degree of activity and damage associated with the disease. METHODS: The study sample included 24 consecutive patients with SLE. 24-hour urine samples were collected for the determination of soluble VCAM-1 and ICAM-1 levels by ELISA. Disease activity was defined by the SLE Disease Active Index (SLEDAI) and disease outcome by the Systemic Lupus International Collaborating Clinics/American College of Rheumatology (SLICC/ ACR) damage index. RESULTS: The urinary soluble VCAM-1 level was significantly higher in patients with SLE compared to normal controls (32.35+/-34.27 vs. 4.66+/-3.8 ng/mg creatinine, p = 0.0005) and statistically significantly correlated with disease activity (SLEDAI), a low serum C3 level, decreased creatinine clearance and albuminuria, as well as with disease damage (SLICC/ACR damage index). In contrast, the urinary soluble ICAM-1 level was not significantly higher in the patients' group compared with the controls (4.5+/-5.19 vs. 2.72+/-2.31 ng/mg creatinine, p=0.2), but was statistically significantly correlated with hematuria and albuminuria. CONCLUSION: Our data suggest that the urinary level of soluble VCAM-1 significantly correlates with overall disease activity and damage scores, but not with nephritis in SLE.     

Retinopathy in systemic lupus erythematosus: relationship to disease activity

Forty-three patients taking chloroquine for systemic lupus erythematosus were followed by one ophthalmologist over a 5-year period. Visual field testing, color vision testing, and fluorescein angiography were performed. Retinopathy was detected in 7 patients (16%), none of whom had hypertension or diabetes mellitus. Retinal abnormalities included cotton-wool spots in 4 patients, microaneurysms in 3, and vascular tortuosity in 4. In 4 patients, these abnormalities were associated with retinal dysfunction, measured in terms of abnormal hue discrimination. In 6 of the 7 patients, the finding of retinopathy coincided with a flare of lupus activity. In 5 patients, retinopathy improved when the disease was controlled.