Systematic lupus erythematosus (SLE) is an autoimmune disease characterized by the presence of anti-DNA autoantibodies. The fact that mammalian DNA is not immunogenic makes SLE an immunological mystery (Zouali 1994). Victims of SLE may have skin rashes, but the real danger of the disease is its direct attack of specific tissues and organs. Ultimately the pathogenic autoantibodies bind to self proteins and form immune complexes that cause severe organ damage (Blatt et al., 1999).
Autoantibodies found in individuals with SLE are primarily IgG1 and IgG3 immunoglobulin (Maddison 1999). The autoantibodies are found in high concentrations and bind antigen with a high affinity. A high frequency of positively charged and polar uncharged amino acids are found in the CDRs of autoantibodies of SLE patients. Positively charged amino acids like lysine, arginine, and histidine are able to interact with phosphate residues of the DNA backbone. Polar, uncharged amino acids like glutamine, glycine, and asparagine can bind to nucleic acids (Zouali 1997). Since SLE autoantibodies favor DNA binding, they often attack intracellular nucleoprotein particles such as nucleosomes and spiceosomes, SLE autoantibodies also bind ribonucleoproteins such as Ro and La. Ro is a 60 kD protein that forms complexes with small cytoplasmic RNA species. La complexes with a variety of RNAs including the precursor forms of tRNA (Maddison 1999). Crystallographic and binding analyses of single stranded DNA:DNA complexes show that the single stranded DNA binding site is a long cleft with a 1.2 nm gap between the heavy and light chain variable regions. The single stranded DNA ligand is stabilized by thymine bases of binding cleft aromatic side chains. High resolution structures of anti-double stranded DNA:DNA complexes are currently unavailable (Blatt et al., 1999).
ETIOLOGY OF SLE
Evidence suggests that SLE has a multi-factorial etiology. Sex hormones, environmental factors, immunological dysfunction, and genetics have all been hypothesized to be potential causes of SLE (Balow et al., 2000). 90% of SLE affected individuals are women. And the onset of SLE in women occurs most often during child-bearing years. This data has led scientists to hypothesize that estrogen levels during pregnancy may be a factor in the development of SLE. Researchers have also noted that flares of SLE have been correlated with ultraviolet light photosensitivity, which may mean that environmental agents have a role in the onset of SLE (Blatt et al., 1999). However, the most interesting theories about the etiology of SLE deal with defects of specific elements of the immune system.
B cells originate from pluripotent hematopoetic stem cells in the bone marrow and undergo several developmental stages before expressing immunoglobulin specific for a single antigen. The specificity of the immunoglobulin is determined by rearrangement of light chain and heavy chain genes. In a healthy individual, immature B cells undergo a "negative selection" process during which B cells that react to self proteins are deleted, inactivated, or undergo further gene arrangement. Immature B cells that recognize multivalent self antigens are eliminated through clonal deletion. Immature B cells that bind to soluble self molecules become anergic. In some cases receptor editing replaces a self-reactive immunoglobulin rearrangement with a successive rearrangement. Theoretically, if the immune system is working properly it should not produce B cells with immunoglobulin that react with self proteins such as DNA. However, individuals with SLE produce antibodies that react with self proteins (Janeway et al., 1999). Researches have hypothesized that individuals with SLE may have a defect in the negative selection process during B cell development (Bensimon et al., 1994).
A POSSIBLE RECEPTOR-EDITING DEFECT
One model of SLE focuses on a flaw in the receptor-editing mechanism of B cells. Scientists examined anti-DNA antibodies of individuals with SLE and noticed that SLE autoantibodies express only downstream Vk genes. The kappa light chain variable region of an antibody molecule is constructed from a variable (V) gene segment and a joining (J) gene segment. There are about 30 Vk gene segments and 5 Vk genes that can be combined during immunoglobulin rearrangement (Janeway et al., 1999). However, there is currently no mechanism explaining which genes will be selected during rearrangement. Usually, downstream Vk genes are selected during primary rearrangement. If the immunoglobulin reacts with self protein it can be saved from deletion by secondary rearrangements called receptor-editing. Research has shown that during successive rearrangements upstream Vk genes are utilized. The fact that SLE autoantibodies express only downstream Vk genes indicates that these autoantibodies do not undergo secondary rearrangements. Furthermore, secondary light chain rearrangements seem to favor downstream Jk exons. However, SLE autoantibodies show no preference in expressing downstream Jk exons. This information suggests that anti-DNA antibodies do not undergo receptor-editing during the negative selection process. Studies done on mice indicate that receptor-editing is critical in deleting potentially pathogenic SLE anti-DNA antibodies. Therefore, individuals with SLE may have defects in the receptor-editing process, resulting in the failure to delete self-reactive antibodies (Bensimon et al., 1994).
POLYCLONAL B CELL ACTIVATION
The fact that self-reactive B cells were not clonally deleted during their development leads to complications down the road. SLE patients have naïve B cells with anti-DNA antibodies circulating throughout the body. There are two theories concerning how these B cells are activated. The first theory suggests that polyclonally activation of these B cells is responsible for the autoimmune response of SLE. DNA present at high concentrations stimulate the SLE B cells to proliferate and secrete antibody. However, immunologists do not understand why polyclonal activation in SLE patients targets a specific subset of autoantibodies (Zouali 1997).
ANTIGEN DRIVEN B CELL ACTIVATION
While it is possible that SLE B cells are polyclonally activated, SLE B cells have a pattern of mutations that suggest that an antigen-driven response also plays a role in their activation. Analysis of pathogenic SLE autoantibodies have revealed that these self-reactive antibodies have a high rate of mutations clustered in hypervariable regions (Zouali 1997). Hypervariable regions are segments of the heavy and light chain V regions that contain a high degree of sequence variability. The antigen-binding site of antibodies is composed of these hypervariable regions, or complementarity-determining regions (CDRs). The high variation of amino acids found in CDRs accounts for the antigen specificity of the immune system. When a B cell binds antigen it moves to the thymus where it is stimulated to differentiate and proliferate by the appropriate helper T cell. Proliferating B cells move to a primary focus and form a germinal center. In the germinal center the B cells undergo somatic mutation. Ultimately, only somatically mutated B cells that have the ability to bind antigen better than the original antibody bound to the follicular dendritic cells survive. Therefore, somatically mutated B cells accumulate mutations in CDRs (Janeway et al., 1999). Some current theories hypothesize that polyclonally activated B cells are responsible for producing SLE autoantibodies. However, SLE autoantibodies have mutations clustered specifically in hypervariable regions, and SLE antibodies have a high affinity for antigen. High affinity for antigen and clustered mutations are characteristic of an antigen-driven response, whereas limited random variations are typical of polyclonal activation (Zouali 1997).
A GENERALIZED T CELL TOLERANCE DEFECT
The human immune system is designed to delete or inactivate self-reactive B cells before they leave the bone marrow. The immune system of an individual with SLE malfunctions, and B cells that bind self proteins evade detection and escape into circulation. However, B cells do not act alone. Naïve B cells that bind antigen must be stimulated by the appropriate T cell before they can differentiate and proliferate. A healthy human immune system also has self-reactive "checks" for T cells. T cells that are specific for self antigens are deleted in the thymus (Janeway et al., 1999). One theory for the existence of autoreactive T cells is that individuals with SLE may have an additional flaw in the negative selection mechanism for T cells. However, a study conducted with (New Zealand black (NZB) x New Zealand white (NZW))F1 (NZB/W) mice indicates that the loss of T cell tolerance characteristic in SLE does not result from a generalized defect in T cell tolerance. Instead, T cell tolerance is caused by a more subtle defect in the immune system, such as abnormal activation of T cells that are specific for a for a subset of autoantigens (Wither et al., 2000).
SPONTANEOUS ACTIVATION OF AUTOREACTIVE T CELLS
Current studies suggest that autoantibody Ig may spontaneously activate T cells. Evidence shows that the heavy chain variable region of autoantibodies stimulate spontaneous T cell activation in young lupus-prone mice, whereas nonautoantibodies with the same heavy chain variable region failed to stimulate T cell activation. The activated T cells can then stimulate naive B cells expressing the self-reactive proteins. The stimulated B cells proliferate and begin secreting SLE autoantibodies. Evidence shows that these self-reactive T cells also recognize several different self-Ig peptides. Autoimmune T cell recognition may diminish over time resulting in a T cell repertoire that binds many self-Ig peptides (Singh et al., 1998).
SUPERANTIGEN BINDING SITE
SLE B cells produce a high number of autoantibodies even though DNA is a weak immunogen. This observation has led researchers to hypothesize that SLE autoantibodies may contain novel immunoglobulin structural domains such as superantigen binding sites. Murine research has revealed that SLE autoantibodies contain specific conserved nucleic acid sequences at the D-J junction of the heavy chain variable region. The location of these sequences suggests that this region may encode a three-dimensional solvent-exposed determinant. The three-dimensional solvent-exposed determinant, which is distinct from the classical antigen binding site, may function as a superantigen or autoantigen binding site. Further research is needed to determine what antigens bind to this alternative binding site and the mechanism of B cell activation (Zack et al., 1994).
EFFECTS OF SLE ANTIBODIES
DNA is almost exclusively intracellular. Only a small amount of DNA from rupturing cells is found circulating in the blood. However in individuals with SLE high quantities of DNA are present in the blood serum. SLE autoantibodies bind circulating DNA and form antibody:antigen immune complexes. Large amounts of immune complexes are continuously produced. Many times these complexes become trapped in tissues of blood vessels, joints, kidneys and brain. Immune complex deposits activate phagocytic cells that cause tissue damage. More DNA and nucleoproteins are released as tissue damage progresses, and more immune complexes are formed (Janeway et al., 1999). Complement activity amplifies the inflammatory reaction (Balow et al., 2000). Immune complexes commonly deposit in the renal glomerulus and glomerular basement membrane (Fig 1) causing proliferative and membranous forms of lupus nephritis (Fig 2) (Janeway et al., 1999).
Figure 1. The granular pattern of immunofluorescence indicates the deposition of immune complexes in the
basement membranes of the glomerulus. (Internet Pathology Laboratory for Medical Education 2000)
Link to The Internet Pathology Laboratory for Medical Education
Figure 2. This image shows thickened capillary loops of a glomerulus of an individual with lupus nephritis.
(Internet Pathology Laboratory for Medical Education 2000)
Link to The Internet Pathology Laboratory for Medical Education
There are currently several treatments available
for SLE. Research has shown that testosterone suppresses B cell hyperactivity
in individuals with SLE. However, androgen receptors have yet to be found
on peripheral blood B cells indicating that testosterone may not act directly
on B cells. Studies show that testosterone therapy results in suppression
of anti-DNA antibodies (Kanda et al., 1997).
Since complement activity can lead to dangerous consequences including inflammation, one experimental treatment has focused on blocking terminal complement activity. The monoclonal antibody to C5b blocks formation of membrane attack complexes. Murine studies utilizing anti-C5b have resulted in less severe cases of lupus nephritis (Balow et al., 2000).
Cytotoxic immunosuppressives such as cyclophosphamide and azathioprine have proved to be effective in the treatment of lupus nephritis. Both cyclophosphamide and azathioprine act by inhibiting the proliferation of rapidly dividing B cells. However, both these drugs are also toxic to cells of the bone marrow and gastrointestinal tract, which result in unwanted side effects (Blatt et al., 1999).
Balow JE. Boumpas DT. Austin HA. 2000. New prospects for treatment of lupus nephritis. Seminars in Nephrology 20: 32-39.
Bensimon C, Chastagner P, Zouali M. 1994. Human lupus anti-DNA autoantibodies undergo essentially primary Vk gene arrangements. European Molecular Biology Organization Journal 13: 2951-2962.
Blatt NB, Glick GD. 1999. Anti-DNA autoantibodies and systematic lupus erythematosus. Pharmacology and Therapeutics 83: 125-139.
The Internet Pathology Laboratory for Medical Education. 2000 Apr 14. The Internet Pathology Laboratory for Medical Eduation Homepage. <http://www-medlib.med.utah.edu/WebPath/webpath.html#MENU>. Accessed 2000 Apr 21.
Janeway CA, Travers P, Walport M, Capra JD. 1999. Immunobiology: the immune system in health and disease. New York, NY: Elsevier Science Ltd/Garland Publishing. p. 209-216, 398, 499-500.
Kanda N, Tsuchida T, Tamaki K. 1997. Testosterone suppresses anti-DNA antibody production in peripheral blood mononuclear cells from patients with systematic lupus erythematosus. Arthritis and Rheumatism 40: 1703-1711.
Maddison PJ. 1999. Autoantibodies in SLE: Disease Associations. Advances in Experimental Medicine and Biology 445: 141-145.
Singh RR, Hahn BH, Tsao BP, Ebling FM. 1998. Evidence for multiple mechanisms of polyclonal T cell activation in murine lupus. The Journal of Clinical Investigation 102: 1841-1849.
Wither J, Vukusic B. 2000. T cell tolerance induction is normal in the (NZBxNZW)F1 murine model of systematic lupus erythematosus. Immunology 99: 345-351.
Zack DJ. Wong AL, Weisbart RH. 1994. Novel structural features of autoantibodies
in murine lupus:
a possible superantigen binding site. Immunology and Cell Biology 72: 513-520.
Zouali M. 1994. Human autoantibodies and their genes. Applied Biochemistry and Biotechnology 47: 135-141.
Zouali M. 1997. The structure of human lupus anti-DNA antibodies. Methods:
A companion to Methods in Enzymology 11: 27-35.
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