Patients with systemic lupus erythematosus are deficient in complement-dependent prevention of immune precipitation
G. J. Arason,
K. Steinsson1,
R. Kolka,
Th. Víkingsdóttir,
M. S. D'Ambrogio2 and
H. Valdimarsson
Department of Immunology, Institute of Laboratory Medicine and 1 Division of Rheumatology and Center for Rheumatology Research, Landspitalinn University Hospital, 101 Reykjavík, Iceland and 2 Calderdale Royal Hospital, Halifax, UK.
Correspondence to: G. J. Arason. E-mail: garason{at}landspitali.is
 |
Abstract
|
---|
Objective. A functional deficiency of complement has been implicated but not conclusively demonstrated in the pathogenesis of systemic lupus erythematosus (SLE). To test this, we studied several aspects of complement in 44 patients with SLE, 46 patients with rheumatoid arthritis and 102 blood donors.
Methods. Prevention of immune precipitation (PIP) was measured by an enzyme immunoassay, levels of C1q, C4 and C3 by rocket immunoelectrophoresis, C4A, C4B and C3d by enzyme-linked immunosorbent assay (ELISA), complement haemolysis (CH50) by standard methods and C4 allotypes by high-voltage agarose electrophoresis and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE).
Results. PIP was significantly reduced in SLE (P<0.001); the defect was revealed by a sensitive assay measuring this function of complement but not by the other tests employed. The patients were clinically well at the time of study, and levels of C3d, which have been shown to correlate with disease activity, were normal. The defect was more common in patients with early disease (P = 0.009), supporting a role in aetiology or early pathophysiology. PIP was positively correlated with levels of C4 (P = 3 x 105) and in particular the C4A isotype (P = 9 x 1010) whereas C4B was redundant.
Conclusions. Our results reveal a defect in prevention of immune precipitation in SLE that is apparent at an early stage in the disease and correlates with low levels of C4A. These results indicate that subtle deficiencies of complement may predispose to SLE.
KEY WORDS: Complement, SLE, Antigenantibody complex, Autoimmune disease, Autoimmunity.
 |
Introduction
|
---|
Systemic lupus erythematosus (SLE) is a genetically complex autoimmune disease characterized by organ-non-specific autoantibodies and endothelial deposition of immune complexes in multiple organ systems [1]. The aetiology is unknown, but it is widely believed that SLE may be caused by subtle defects of classical pathway activity, resulting in inefficient handling and chronic tissue deposition of immune complexes and the consequent induction of an autoimmune response [24]. It is clear that this chain of events can lead to SLE, as shown by the high frequency of SLE in subjects with inherited absolute deficiencies in classical pathway components [5, 6], especially C1 and C4. However, complete deficiency of classical pathway activity can only account for a minute fraction of SLE cases, and the notion that more subtle complement abnormalities may also predispose to SLE rests on indirect evidence, i.e. the association of SLE with C4A null alleles [2, 7] combined with the results of in vitro studies indicating that C4A is more efficient than C4B in binding to immune complexes [8, 9].
An association of SLE with C4A null alleles (C4A*Q0) has been observed in diverse populations and against a variable major histocompatibility complex (MHC) background [2, 4, 7, 1015]. Nevertheless, several studies emphasize the possibility of a confounding influence from genes in linkage disequilibrium with C4 genes [2, 16, 17], and studies on the effect of null alleles on serum levels of C4 have yielded conflicting results [18, 19]. Moreover, the advantage of C4A over C4B in binding to immune complexes has been questioned in recent studies [20]. Thus, the effect of C4 null alleles on serum protein levels and their relative contribution to classical pathway function remains controversial.
The relationship between SLE and complement is unlikely to be revealed through genetic studies alone; a central issue in this context is to resolve whether a defect in immune complex handling is common in SLE. Direct studies on this issue have not been reported, and we reasoned that this might be due to the lack of sensitive assays for measuring immune complex handling, as previous studies have been based on assays in which bond formation within and between immune complexes may have been weakened by chemical modification of the antigen, with loss of sensitivity around the lower limit of normal [21]. To avoid this we developed an immunoassay for measuring prevention of immune precipitation (PIP) in which alkaline phosphatase serves the dual role of a label and antigen; this assay is optimally sensitive at all serum dilutions within and below the normal range [22]. Employing this assay we observed a defect in complement function in SLE which appears to be intrinsic and operative at an early stage of the disease; analysis of clinical and laboratory data indicates that it is specifically associated with low levels of C4A.
 |
Patients and methods
|
---|
The study group consisted of five males and 39 females, aged 2384 yr, with a disease history ranging from 4 months to 48 yr (Table 1). The patients were studied during periods of low clinical activity (<10, average 5) according to the SLE disease activity index (SLEDAI) [23]. Rheumatoid arthritis (RA) patients (12 males and 34 females, aged 21 to 81 yr) and blood donors (n = 102) served as controls. Informed consent was obtained from all patients. All patients satisfied the American College of Rheumatology classification criteria for SLE or RA [24, 25].
The study has been approved by all relevant ethical committees.
Materials
Mouse monoclonal anti-C4A (RgD1) and anti-C4B (1228) were a kind gift from Professor Peter Schneider. Alkaline phosphatase (AP) was purchased from Sigma (St Louis, MO), anti-AP from ICN/Cappel (Aurora, OH), anti-C4 from DiaSorin (Stillwater, MN), other antibodies from Dako (Copenhagen, Denmark) and microtitre plates from Nunc (Roskilde, Denmark). In enzyme-linked immunosorbent assay (ELISA) measurements, phosphate-buffered saline (PBS) containing 0.05% Tween was used for diluting sera, and PBS with 0.005% Tween for washing.
The PIP assay
All measurements were performed on serum aliquots kept at 70°C throughout. PIP was measured as previously described [22]. Briefly, AP (1/50, 5 µl) and goat anti-AP (1/5, 5 µl) were added to serum (40 µl); after incubation (37°C, 1 h) and centrifugation (5500 g, 10 min), supernatants were diluted 1:10 in PBS and reacted with substrate (p-nitrophenylphosphate; Sigma). Absorbance was converted to arbitrary units (AU) by comparison with a serially diluted reference serum pool, defined as 100 AU. This assay is sensitive to minor variations within and below the normal range (Fig. 1).

View larger version (10K):
[in this window]
[in a new window]
|
FIG. 1. Sensitivity of the PIP assay. At standard conditions there is a linear relationship between PIP and serum dilutions.
|
|
C4 allotypes and their levels
C4 allotypes were determined by high-voltage agarose electrophoresis on carboxypeptidase- and neuraminidase-treated samples with subsequent immunofixation and staining [26]. Null alleles were determined by visual scoring of the relative intensities of C4A and C4B bands and confirmed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) of C4
-chains [27, 28]. Levels of C4A and C4B were measured by a modified ELISA [29] using goat anti-C4 for coating (1:200) and mouse monoclonal anti-human C4A (RgD1; 1/1000) or C4B (1228; 1/500) followed by AP-conjugated rabbit anti-mouse Ig (1/1000) for catching; sera were diluted 1:800. Results were expressed in g/l by comparison with serially diluted (1/200 to 1/12 800) reference plasma samples from phenotypically homozygous C4A- or C4B-deficient probands; their C4 concentrations had previously been determined twice by rocket immunoelectrophoresis.
CH50, complement components and autoantibodies
C1q, C4 and C3 were measured by rocket immunoelectrophoresis and total haemolytic complement (CH50) by standard titration methods. Results were expressed in g/l by comparison with a known standard, or as AU by comparison with a standard serum pool. Complement activation was assessed by monitoring the level of the C3d cleavage product by ELISA [30, 31], using rabbit anti-C3d (1:1000) for coating and AP-conjugated anti-C3d (1:2000) for catching. Sera were diluted 1:100 and results expressed in AU by comparison with a serially diluted zymosan-activated reference serum, defined as 100 AU. The ELISA was made specific for C3d by removing larger C3 fragments (C3, C3b, iC3b) from sera before adding them to the wells of the microtitre plate [30]. This was done by adding ethylenediaminetetra-acetic acid (EDTA) (10 mM) and mixing the sera with equal volumes of 22% polyethylene glycol, followed by incubation on ice (1 h) and centrifugation (1500 g, 30 min) at 4°C. Normal levels for C3d were determined by measuring sera from 100 blood donors.
Antibodies to double-stranded deoxyribonucleic acid (DNA) were measured by the Farr assay using 14C-labelled DNA from Amersham/Pharmacia (Uppsala, Sweden). Anti-cardiolipin antibodies were measured by the Varelisa test with ß2-glycoprotein I as cofactor (Amersham/Pharmacia).
Statistical analysis
The means of test values between patients and controls were compared using the MannWhitney U-test. PIP was compared with other laboratory and clinical parameters using analysis of variance (ANOVA),
2 and Pearson's correlation statistics. Significance was set at P<0.05.
 |
Results
|
---|
PIP
Prevention of immune precipitation was markedly reduced in 74 SLE patients sampled consecutively at study initiation (Fig. 2). Several of these patients had quiescent disease and normal C4, C3 and CH50. However, the group also included patients who were sampled during active disease and thus had other detectable complement abnormalities. To minimize the effect of such confounding factors, our main study was focused on patients with minimal disease activity as determined by clinical criteria. Patients (n = 57) were therefore evaluated retrospectively for 4 yr and followed prospectively for additional 7 yr, and clinical and laboratory data recorded during both active and inactive disease. The 44 patients who were available for analysis at least once during a quiescent period (SLEDAI<10) formed the study group. As shown in Fig. 2, PIP was markedly reduced in these patients (P<0.001).

View larger version (15K):
[in this window]
[in a new window]
|
FIG. 2. PIP is deficient in SLE regardless of disease activity. PIP was impaired in 74 SLE patients sampled consecutively at study initiation, compared with 46 RA patients and 102 blood donors. This defect was apparent also when 44 SLE patients were sampled during low disease activity (SLEDAI score <10). The lower limit of normal (2 S.D.) is denoted by a dashed line.
|
|
CH50, C1q, C4, C3 and C3d
Measurements of CH50, C1q, C4 and C3 were normal at the time of sampling (Fig. 3) and the defect observed in the PIP assay was thus not associated with any other detectable complement abnormality. Although levels of the complement split product C3d were slightly raised in four patients (Fig. 3) this parameter was not correlated with low PIP. Thus, of the 23 patients with deficient PIP, 20 had no other complement abnormalities and the remaining three patients did not have reduced levels of complement components nor CH50, although some activation of complement may have taken place.

View larger version (23K):
[in this window]
[in a new window]
|
FIG. 3. The functional complement defect in SLE is not revealed by conventional methods. Levels of CH50, C1q, C4, C3 and C3d in 44 patients with quiescent SLE are normal (y-axis). Broken horizontal lines denote normal distribution (2 S.D.).
|
|
Correlation statistics
Studies on the interrelationship of clinical and laboratory parameters revealed a strong positive correlation of PIP to levels of C4 (r = 0.60, P = 0.00003) and this was entirely due to the C4A isotype (r = 0.78; P = 0.00000000086) as no association was seen between PIP and C4B (Fig. 4, Table 2). PIP was negatively correlated to anti-DNA and anti-cardiolipin antibodies (Table 2); no association was found between PIP and organ involvement (data not shown). PIP was also positively correlated with levels of C3 but ANOVA suggested that unlike the other associations found, this was secondary to the correlation of PIP and C3 to C4A. Compared with population values [14], the frequency of C4A*Q0 was increased in the patients (0.23 vs 0.13; P = 0.02), with two patients being homozygous and 17 heterozygous. PIP values tended to be lower in patients with C4A*Q0 (median 64) than in the remaining patients (median 72) but the difference was not significant (P = 0.09).

View larger version (17K):
[in this window]
[in a new window]
|
FIG. 4. The correlation of complement-dependent prevention of immune precipitation (PIP) to levels of (a) total C4, (b) C4A and (c) C4B. The correlation coefficients and P-values according to Pearson's statistics are shown
|
|
PIP and disease progression
The PIP defect is especially pronounced in early disease, as shown in Table 2. Analysis of data from individual patients (Table 1) is consistent with this, as quite low values were observed in seven patients within 18 months of diagnosis, including four who were diagnosed within months after the onset of symptoms. PIP normalized in two patients after treatment with immunosuppressive drugs (recorded as normal in Fig. 2).
 |
Discussion
|
---|
Our results show that prevention of immune precipitation is markedly defective in the majority of patients with SLE, even when the disease is inactive and no deficiencies are detected in CH50 or individual classical pathway components. C3d measurements which have been shown to correlate with disease activity in SLE [32, 33] confirmed that the patients had been sampled during remission, and values of CH50, C1q, C4 and C3 were normal. Although definite conclusions on causal relationships cannot be made at this stage, the fact that the defect was especially prominent in early disease supports a role in the aetiology or early pathophysiology and the results thus provide, for the first time, direct experimental support for the hypothesis that subtle complement deficiencies may be a common inductive factor in SLE.
Our results indicate that the PIP defect mainly reflects low levels of the C4A protein, and this is consistent with the known functional difference between the two C4 isotypes [8, 9, 21], C4A binding more readily to immune complexes while C4B binds preferentially to hydroxyl groups on cell membranes, including erythrocytes. This may also form the basis for the association of SLE with C4A*Q0 in different ethnic groups [2, 7, 1015, 34]. However, it should be noted that in our study the defect was associated with low concentration of the C4A protein rather than the C4A*Q0 genotype; in line with this, low levels of C4A were observed in several patients who did not carry the C4A*Q0 allotype, while many patients with this allotype had normal C4A concentration. Discrepancies between serum C4 concentrations and the number of null alleles have been noted in previous studies [18, 19] and may be explained by a combination of non-MHC linked genetic factors [35, 36], MHC haplotype-specific differences in the relative expression of C4A and C4B [37] and compensation for the lack of one gene product by increased expression of the remaining functional allele. The lack of direct association of PIP with C4A*Q0 in our study is probably due to compensation.
Of the components studied, only C4A proved to be independently correlated with PIP. Neither C1q nor C3 has been demonstrated to show functional heterogeneity, and both proteins are coded for by one locus only. Thus it appears likely that levels of these components are under more rigid control than levels of the subcomponents C4A and C4B. The lack of positive results with the other functional test of complement (CH50) illustrates the insensitivity of this method for measuring complement functions which do not rely upon lytic activity as the endpoint. Taken together, our results underscore the benefit of functional complement assays over those measuring protein concentrations or the frequency of gene allotypes for estimating complement abnormalities, and invite a reappraisal of the notion that a combined qualitative and quantitative defect in classical pathway activity may be a relatively common predisposing factor for SLE [38]. Although studies on lupus-prone mice clearly indicate that SLE may be genetically complex and result in part from immunological defects not associated with complement, it appears quite likely that a hitherto underestimated proportion of SLE cases may result from subtle defects in classical pathway activity. Our working hypothesis is that individuals with impaired PIP due to insufficient levels or activity of C4A or other classical pathway proteins are at increased risk of developing SLE; although complete genetic deficiencies probably confer the highest risk it appears likely that transient insufficiencies due to temporary depletion or reduced production also confer a risk of developing immune complex disease, as indicated by the increased incidence of SLE after infections and in genetically determined secondary deficiencies of classical pathway proteins including C1 esterase inhibitor [2, 3, 39, 40]. In addition to the evidence quoted above, this hypothesis is supported by studies indicating that the B-cell hyperactivity in SLE is driven by antigen [41, 42], as well as by studies showing that drug-induced lupus erythematosus is associated with null alleles of C4A [43] and elevated levels of circulating immune complexes [44] and the compounds implicated are all strong inhibitors of classical pathway function, acting on C4 either directly (isoniazid, hydralazine, penicillamine) [4547] or through a metabolic product (procainamide) [48]. Although the overall impact of these arguments is quite strong, it should be noted that our study provides the first direct experimental evidence that individuals with impaired handling of immune complexes due to subtle complement defects are at increased risk of developing SLE.
In conclusion, prevention of immune precipitation is defective in the majority of SLE patients even during remission when other complement parameters are normal. The defect was prominent at an early stage of the disease and correlated strongly with low levels of C4A. These results indicate that complement dysfunction may be an important factor in the pathophysiology of SLE.
 |
Acknowledgments
|
---|
This work was supported by the Icelandic Research Council (grant no 971310097) and the Science Fund of Landspitali University Hospital, and was approved by the Icelandic Data Protection Commission and the National Bioethics Committee. Monoclonal antibodies RgD1 and 1228 were a kind gift from Professor Peter Schneider (Johann Gutenberg Universität Mainz). We thank the National Blood Bank of Iceland for supplying sera from blood donors, and Alfreð Árnason, Jón þorsteinsson, Kristín Traustadóttir, Kristján Erlendsson, Sigríður Rut Franzdóttir and Örn Ólafsson for their contribution.
The authors have declared no conflicts of interest.
 |
References
|
---|
- Wallace DH, Hahn BH (eds). Dubois Lupus erythematosus, 5th edn. Baltimore: Lippincott, Williams & Wilkins, 1997.
- Lachmann PJ, Walport, MJ. Deficiency of the effector mechanisms of the immune response and autoimmunity. Ciba Found Symp 1987;129:14971.[ISI][Medline]
- Walport MJ, Davies KA, Morley BJ, Botto M. Complement deficiency and autoimmunity. Ann NY Acad Sci 1997;815:26781.[ISI][Medline]
- Sullivan KE. Complement deficiency and autoimmunity. Curr Opin Pediatr 1998;10:6006.[Medline]
- Schifferli JA, Ng YC, Peters DK. The role of complement and its receptor in the elimination of immune complexes. N Engl J Med 1986;315:48895.[ISI][Medline]
- Agnello V. Lupus diseases associated with hereditary and acquired deficiencies of complement. Springer Semin Immunopathol 1986;9:16178.[ISI][Medline]
- Christiansen FT, Zhang WJ, Griffiths M, Mallal SA, Dawkins RL. Major histocompatibility complex (MHC) complement deficiency, ancestral haplotypes and systemic lupus erythematosus (SLE): C4 deficiency explains some but not all of the influence of the MHC. J Rheumatol 1991;18:13508.[ISI][Medline]
- Law SK, Dodds AW, Porter RR. A comparison of the properties of two classes, C4A and C4B, of the human complement component C4. EMBO J 1984;3:181923.[Abstract]
- Kishore N, Shah D, Skanes VM, Levine RP. The fluid-phase binding of human C4 and its genetic variants, C4A3 and C4B1, to immunoglobulins. Mol Immunol 1988;25:81119.[CrossRef][ISI][Medline]
- Yamada H, Watanabe A, Mimori A. et al. Lack of gene deletion for complement C4A deficiency in Japanese patients with systemic lupus erythematosus. J Rheumatol 1990;17:10547.[ISI][Medline]
- Hong GH, Kim HY, Takeuchi F. et al. Association of complement C4 and HLA-DR alleles with systemic lupus erythematosus in Koreans. J Rheumatol 1994;21:4427.[ISI][Medline]
- Ratnoff WD. Inherited deficiencies of complement in rheumatic diseases. Rheum Dis Clin N Am 1996;22:7594.[ISI][Medline]
- Schur PH. Complement and systemic lupus erythematosus. In: Wallace DH, Hahn BH (eds). Dubois Lupus erythematosus. Baltimore: Lippincott, Williams & Wilkins, 1997:24561.
- Steinsson K, Jonsdottir S, Arason GJ, Fossdal R, Skaftadottir I, Arnason A. A study of the association of HLA DR, DQ, and complement C4 alleles with systemic lupus erythematosus in Iceland. Ann Rheum Dis 1998;57:5035.[Abstract/Free Full Text]
- Sullivan KE, Kim NA, Goldman D, Petri MA. C4 deficiency due to a 2 bp insertion is increased in patients with systemic lupus erythematosus. J Rheumatol 1999;26:21447.[ISI][Medline]
- Schur PH, Marcus-Bagley D, Awdeh Z, Yunis EJ, Alper CA. The effect of ethnicity on major histocompatibility complex complement allotypes and extended haplotypes in patients with systemic lupus erythematosus. Arthritis Rheum 1990;33:98592.[ISI][Medline]
- Hartung K, Baur MP, Codewey R. et al. Major histocompatibility complex haplotypes and complement C4 alleles in systemic lupus erythematosus. Results of a multicenter study. J Clin Invest 1992; 90:134651.[ISI][Medline]
- Hammond A, Ollier W, Walport MJ. Effects of C4 null alleles and homoduplications on quantitative expression of C4A and C4B. Clin Exp Immunol 1992;88:1638.[ISI][Medline]
- Moulds JM, Warner NB, Arnett FC. Complement component C4A and C4B levels in systemic lupus erythematosus: quantitation in relation to C4 null status and disease activity. J Rheumatol 1993;20:4437.[ISI][Medline]
- Reilly BD. Analysis of human C4A and C4B binding to an immune complex in serum. Clin Exp Immunol 1999;117:1218.[CrossRef][ISI][Medline]
- Schifferli JA, Steiger G, Paccaud JP, Sjoholm AG, Hauptmann G. Difference in the biological properties of the two forms of the fourth component of human complement (C4). Clin Exp Immunol 1986;63:4737.[ISI][Medline]
- Arason GJ, DAmbrogio MS, Vikingsdottir T, Sigfusson A, Valdimarsson H. Enzyme immunoassays for measuring complement-dependent prevention of immune precipitation (PIP) and solubilization of preformed antigen-antibody complexes (SOL). J Immunol Methods 1999;223:3746.[CrossRef][ISI][Medline]
- Bombardier C, Gladman DD, Urowitz MB, Caron D, Chang CH. Derivation of the SLEDAI. A disease activity index for lupus patients. Arthritis Rheum 1992;35:63040.[ISI][Medline]
- Arnett FC, Edworthy SM, Bloch DA et al. The American Rheumatism Association 1987 revised criteria for the classification of rheumatoid arthritis. Arthritis Rheum 1988;31:31524.[ISI][Medline]
- Tan EM, Cohen AS, Fries JF et al. The 1982 revised criteria for the classification of systemic lupus erythematosus. Arthritis Rheum 1982;25:12717.[ISI][Medline]
- Schneider PM, Rittner C. Complement genetics. In: Dodds AW, Sim RB (eds). Complement, a practical approach. Oxford: Oxford University Press, 1997:16599.
- Roos MH, Mollenhauer E, Demant P, Rittner CA. A molecular basis for the two locus model of human complement component C4. Nature 1982;298:8546.[ISI][Medline]
- Mauff G, Steuer M, Weck M, Bender K. The C4 beta-chain: evidence for a genetically determined polymorphism. Hum Genet 1983;64:1868.[ISI][Medline]
- Rebmann V, Doxiadis I, Kubens BS, Grosse-Wilde H. Quantitation of the human component C4: definition of C4 Q0 alleles and C4A duplications. Vox Sang 1992;62:11723.[ISI][Medline]
- Mollnes TE. Quantification of the C3d split products of human complement by a sensitive enzyme-linked immunosorbent assay. Scand J Immunol 1985;21:60713.[ISI][Medline]
- Traustadottir KH, Rafnar BO, Steinsson K, Valdimarsson H, Erlendsson K. Participation of factor B in residual immune complex red cell binding activity observed in serum from a C2-deficient systemic lupus erythematosus patient may delay the appearance of clinical symptoms. Arthritis Rheum 1998;41:42734.[CrossRef][ISI][Medline]
- Senaldi G, Makinde VA, Vergani D, Isenberg DA. Correlation of the activation of the fourth component of complement (C4) with disease activity in systemic lupus erythematosus. Ann Rheum Dis 1988;47:91317.[Abstract]
- Röther E, Lang B, Coldewey R, Hartung K, Peter HH. Complement split product C3d as an indicator of disease activity in systemic lupus erythematosus. Clin Rheumatol 1993;12:315.[ISI][Medline]
- Traustadottir KH, Steinsson K, Erlendsson K. C4AQ0 superimposed on a primary defect increases the susceptibility to systemic lupus erythematosus (SLE) in a family with association between C4AQ0 and SLE. J Rheumatol 1998;25:211825.[ISI][Medline]
- Wisnieski JJ, Nathanson MH, Anderson JE, Davis AE, 3rd, Alper CA, Naff GB. Metabolism of C4 and linkage analysis in a kindred with hereditary incomplete C4 deficiency. Arthritis Rheum 1987;30:91927.[ISI][Medline]
- Mayumi M, Heike T, Mikawa H. Transient selective C4 deficiency of infancy. Lancet 1992;339:752.
- Truedsson L, Awdeh Z, Yunis EJ, Mrose S, Moore B, Alper CA. Quantitative variation of C4 variant proteins associated with many MHC haplotypes. Immunogenetics 1989;30:41421.[ISI][Medline]
- Porter RR. Complement polymorphism, the major histocompatibility complex and associated diseases: a speculation. Mol Biol Med 1983;1:1618.[Medline]
- Hory B, Haultier JJ. Glomerulonephritis and hereditary angioedema: report of 2 cases. Clin Nephrol 1989;31:25963.[ISI][Medline]
- Walport MJ, Davies KA, Botto M. et al. C3 nephritic factor and SLE: report of four cases and review of the literature. Q J Med 1994;87:60915.[ISI]
- Tan EM. Pathophysiology of antinuclear antibodies in systemic lupus erythematosus and related diseases. Adv Dent Res 1996;10:446.[Abstract]
- Casciola-Rosen L, Rosen A. Ultraviolet light-induced keratinocyte apoptosis: a potential mechanism for the induction of skin lesions and autoantibody production in LE. Lupus 1997;6:17580.[ISI][Medline]
- Speirs C, Fielder AH, Chapel H, Davey NJ, Batchelor JR. Complement system protein C4 and susceptibility to hydralazine-induced systemic lupus erythematosus. Lancet 1989;1:9224.[ISI][Medline]
- Mitchell JA, Batchelor JR, Chapel H, Spiers CN, Sim E. Erythrocyte complement receptor type 1 (CR1) expression and circulating immune complex (CIC) levels in hydralazine-induced SLE. Clin Exp Immunol 1987;68:44656.[ISI][Medline]
- Sim E, Gill EW, Sim RB. Drugs that induce systemic lupus erythematosus inhibit complement component C4. Lancet 1984;2:4224.[CrossRef][Medline]
- Sim E, Law SK. Hydralazine binds covalently to complement component C4. Different reactivity of C4A and C4B gene products. FEBS Lett 1985;184:3237.[CrossRef][ISI][Medline]
- Sim E, Dodds AW, Goldin A. Inhibition of the covalent binding reaction of complement component C4 by penicillamine, an anti-rheumatic agent. Biochem J 1989;259:41519.[ISI][Medline]
- Sim E, Stanley L, Gill EW, Jones A. Metabolites of procainamide and practolol inhibit complement components C3 and C4. Biochem J 1988;251:32.
Submitted 29 August 2002;
revised version accepted 26 February 2004.