MΦ from prediseased SLE-prone MRL mice differentially express multiple adhesion-related genes in the presence of SL-Apo
We used gene arrays to identify gene products that are abnormally expressed by SLE-derived MΦ in the presence of apo target cells and normally expressed in their absence. Given the apo cell-dependent abnormality in activity of the cytoplasmic G-protein Rho, as expressed by MΦ from SLE-prone mice [2
], we focused our analysis on genes related to cellular adhesion and/or migration (). We identified 42 genes whose differential expression by SLE-prone MRL vs. control BALB/c MΦ in the presence of SL-Apo exceeded a threshold of 1.75-fold in four independent comparisons at either 8 or 24 h following LPS stimulation. The mean fold-difference in expression (± standard error) for these 42 genes was 3.04 ± 0.27.
Adhesion-related genes differentially expressed by MΦ of MRL vs. BALB/c mice.
We also determined the expression of these same 42 genes by MRL vs. BALB/c MΦ in the absence of SL-Apo. Comparison of their expression patterns under these two conditions (presence or absence of SL-Apo) allowed us to sort the 42 genes into two subsets with different pathophysiologic significance.
Genes in the first and larger subset (n = 32) were differentially expressed by MRL MΦ only when they were allowed to interact with SL-Apo. In the absence of SL-Apo, MRL MΦ expressed these 32 genes at a level comparable to that of BALB/c MΦ (<1.75-fold). Differential expression for this subset of genes cannot be attributed to mutation or allelic variation between MRL and BALB/c mice, since their differential expression is conditional and correctable. Thus, in the absence of SL-Apo, expression of these 32 genes by MRL MΦ is normalized. Importantly, since genes in this subset are differentially expressed by MRL MΦ solely in the presence of SL-Apo, the consequences of their aberrant expression should be manifest only during those times when MRL MΦ are interacting with apo cells (or their surrogate, SL-Apo), an abundant source of self-Ag.
Genes in the second smaller subset (n = 10) were differentially expressed by MRL MΦ in both the presence and the absence of SL-Apo. Their differential expression is more likely to result from fixed strain-specific differences between MRL and BALB/c mice. Such differences may be functionally inconsequential. Alternatively, such differences may be indicative of a true SLE-associated allele or mutation, much like the lpr mutation of the Fas gene.
Abnormalities of adhesion can predispose to the development of systemic autoimmunity
The gene products in may be sorted into groups according to function. The first and most provocative group is involved in cell–cell interactions. Examples include several cell surface adhesion receptors (e.g. VCAM-1, DM-GRASP, CD72, and the CD5-like CT-2) or bridging molecules like thrombospondin-1 that facilitate the interaction between MΦ and other cells.
Adhesive interactions play a major role in formation of the immunological synapse between T cell and antigen-presenting cell (APC; [6
]). Alterations in the formation and strength of the immunological synapse modulate signalling events triggered by T-cell receptor engagement and help to determine the balance between tolerance and activation [7
]. In this regard, it is noteworthy that MRL MΦ under-expressed CD72 by >10-fold. CD72 is a cell surface glyco-protein primarily expressed by B cells that appears to modulate signalling through the B-cell receptor. One of its binding partners is CD5. Mice deficient in CD72 develop AAb to dsDNA and demonstrate abnormalities in the maintenance of peripheral tolerance by B cells [8
]. Of note, CT-2, which possesses homology to CD5, was also differentially expressed by MRL MΦ.
The second group is involved in modulating cell–matrix adhesion and/or migration. Examples include constituents of the extracellular matrix (ECM), (e.g. decorin, α1 chain of type I procollagen, and chondroitin sulfate proteoglycan 2) or secreted molecules that modify the ECM (e.g. urokinase, matrix metalloproteinases-9 and -13). Other examples include gene products that modulate the adhesiveness or directed migration of cells (e.g. MAGE-D1, metadherin, and TGF-β-induced 68 kDa protein). Abnormalities of genes in this group may affect the ability of MΦ to be recruited to sites of inflammation [9
]. In addition, inhibition of adhesion through administration of a moAb to the adhesion receptor CD11b/CD18 abrogated autoimmunity in a model of type I diabetes [10
The final group is involved in regulation of the actin cytoskeleton and microtubular network. Genes in this group have the capacity to modulate both cell–matrix and cell–cell interactions. Examples include several small cytoplasmic G-proteins (e.g. Rab5A, Rab7, and Rap2B) and their upstream GTPase activating proteins (Ras p21 protein activators 3 and 4, TBC1D15), as well as other upstream regulatory proteins (DOCK2, DOCK7, Kif23, centaurin-γ2, and gelsolin-like actin filament capping protein). Other examples include several downstream targets of Rho (e.g. citron kinase) or Rho-family G-proteins (e.g. iNOS2A, Rsk3, subunit 5 of Arp2/3 complex, and stathmin 1).
Cytoskeletal rearrangements triggered by Rho-family G-proteins contribute to the co-stimulation and activation of lymphocytes [11
]. Importantly, abnormalities of adhesion and Rho-family G-proteins have been described in MΦ and lymphocytes derived from patients and mice with SLE [3
]. In strong support of a role for abnormalities of adhesion in the development of autoimmunity, two recent genetic studies on murine and human SLE have identified disease-modifying loci with strong links to adhesion. An inactivating mutation of coronin-1A, a multi-functional regulator of the cytoskeleton that participates in formation of the immunological synapse, conferred resistance to autoimmunity in mice [13
]. In addition, polymorphisms within the locus for ITGAM (integrin alpha-M, or CD11b) conferred susceptibility to SLE in human SLE [14
]. The ITGAM-containing CD11b/CD18 adhesion receptor is one of several MΦ receptors whose engagement in the presence of apo cells leads to impaired Rho activity and abnormalities of adhesion and cytoskeletal arrangement [3