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Target Ag Display in Lupus
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-actinin is a major target of anti-dsDNA Abs in systemic lupus erythematosus in mice and humans and that anti--actinin Ab binds better to mesangial cells (MCs) from lupus-prone MRL-lpr/lpr (MRL/lpr) mice than MCs from nonautoimmune BALB/c mice. On p. 7704
, the same group (Zhao et al.) examined the basis for the differential Ab binding. Although the MRL/lpr and BALB/c ACTN4 gene sequences differed by 8 aas, no differences in expression or localization were noted in human cells transfected with vectors expressing either ACTN4 protein. Both purified recombinant -actinin proteins bound anti--actinin 4 Ab with equal affinity. However, levels of ACTN1 and ACTN4 mRNAs and their proteins were higher in primary MRL/lpr MCs compared with BALB/c MCs as determined by real-time PCR and immunoblotting. More transcripts for ACTN1A, a splice variant of ACTN1 cDNA, than ACTN1 were found in MRL/lpr MCs. Anti--actinin Abs bound equally well to ACTN1 and ACTN1A proteins. Immunofluorescence showed higher -actinin expression on fixed primary MCs from the lupus-prone mice; ACTN1 staining was cytoskeletal, whereas ACTN4 was cytoplasmic and nuclear. In vivo, 5-mo-old MRL/lpr mice had higher serum anti-ACTN1, anti-ACTN1A, and anti-ACTN4 Abs, and their glomeruli stained more intensely with anti--actinin Abs than age-matched BALB/c mice. The authors propose that overexpression of -actinin 1 and -actinin 4, target Ags of anti-DNA Abs, in the cytoplasm and on the surface of MRL/lpr MCs contributes to lupus development. Cardiomyopathy Immune Pathology
The mouse model for spontaneous idiopathic dilated cardiomyopathy described by Elliott and coworkers is an I Ab knockout NOD mouse in which the human HLA-DQ8 replaces the MHC class II molecule (NOD.DQ8/Ab0). In a continuation of their work, Hayward et al. (p. 7715
) showed that NOD/Ab0 mice carrying the human DQ8, but not those carrying human DQ6, develop heart block by 24 wk of age as assessed by electrocardiogram and anti-cardiac myosin Abs. All young Rag1 knockout NOD.DQ8/Ab0 mice (NOD.DQ8/Ab0 Rag10) developed complete heart block and other symptoms of cardiomyopathy within 12 wk of receiving splenic lymphocytes from older NOD.DQ8/Ab0 mice with heart block. Adoptively transferred CD4+ T cells, enriched from the splenic lymphocytes by negative or positive selection, accelerated disease development and increased macrophage numbers in hearts of NOD.DQ8/Ab0 Rag10 recipients. In contrast, adoptive transfer of sorted CD8+ T or B cells from NOD.DQ8/Ab0 spleens did not induce cardiac pathology. NOD.DQ8/Ab0 mice lacking IgH developed heart block with a slight delay compared with NOD.DQ8/Ab0 animals, whereas mice deficient in CD8+ T cells (lacking 
2-microglobulin) did not develop disease. The authors show that CD8+ T cells are required to initiate spontaneous anticardiomyocyte autoimmunity, whereas CD4+ T cells perpetuate the autoimmune response and recruit macrophages to cardiac tissue in a model of idiopathic dilated cardiomyopathy.
Estrogen and Thymic Atrophy
The thymus decreases in size during pregnancy, whereas an increase is seen after ovariectomy. The mechanism by which estrogen influences thymus size is unknown. Zoller and Kersh (p. 7371
) injected synthetic 17--estradiol i.p. daily into male mice to induce serum levels equivalent to those present in late-stage pregnancy. Thymic atrophy began 2 days after initial injection and continued through 6 days. Double positive T cells were preferentially reduced, as were c-Kit+ early T lineage progenitor cells and double negative 2 (DN2) cells in the thymus and Flt3+ cells in the bone marrow. Significantly reduced proliferation of DN3 and DN4 thymocytes was demonstrated by decreased BrdU incorporation in vivo after exposure of mice to estradiol for 2 days. No increase in annexin V+ staining was seen in any thymocyte subset, and no up-regulation of Notch target genes was detected by RT-PCR. Estradiol treatment led to a preferential reduction of thymic DN2 cells and a relative expansion of thymic DN1 cells in Rag1–/– mice. IL-7R–/– mice exhibited thymic atrophy following estradiol treatment. The authors conclude that estradiol induces thymic atrophy by eliminating double positive thymocyte precursors in the bone marrow and early T lineage progenitor cells in the thymus through inhibition of thymopoiesis and not through apoptosis.
PKC and TCR Down-Regulation
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or PKC
induced TCR down-regulation; stimulated cells transfected with a dominant negative form of either PKC had partial down-regulation. Stimulated T cells from mice lacking PKC or PKC had impaired TCR down-regulation. Experiments using transfected mutated forms of the CD3
chain showed a requirement for Ser126 and the di-leucine-based receptor-sorting motif. Down-regulated TCRs entered the endocytotic recycling compartment in stimulated T cells transfected with constitutively expressed PKC or PKC. The V3-TCR complex was down-regulated in human V8+ T cells transfected with a V3-expressing plasmid after stimulation with anti-V3/-CD28 or anti-V8/-CD28 mAbs, whereas only anti-V3/-CD28 mAbs stimulated down-regulation of V3-TCR in cells cotransfected with a small interfering RNA against PKC. In similar experiments, anti-V8/-CD28, but not anti-V3/-CD28, mAb stimulated down-regulation of V3-TCR in the presence of small interfering RNA against PKC. This pattern of PKC and PKC activity was confirmed in a stably transfected mouse T cell line coexpressing V2 and V8 after stimulation with anti-V chain-specific mAbs with or without the appropriate dominant negative PKC. The data show that PKC isotypes and are involved in down-regulation of engaged and nonengaged TCRs, respectively, in mouse and human T cells. Statins Increase Efferocytosis
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TLR2, Porphyromonas gingivalis, and Atherosclerosis
Harokopakis and Hajishengallis showed that binding of fimbriae from Porphyromonas gingivalis, a bacterium associated with increased risk for cardiovascular disease in humans, to CD14 stimulates TLR2/PI3K-mediated inside-out signaling for CD11b activation on monocytes. On p. 7645 , Harokopakis et al. detected increased binding of fimbria-stimulated human monocytes to ICAM-1- or fibrinogen-coated wells; pretreatment of the cells with anti-CD11b mAb, with an allosteric antagonist of CD11b, with mAbs to CD14 or TLR2 or with either of two PI3K inhibitors reduced adhesion. Similarly, fimbria-stimulated CD14–/– or TLR2–/– macrophages did not exhibit enhanced binding. Among several bacterial toxins tested, only pretreatment with Clostridium difficile toxin B inhibited the fimbria-stimulated binding of monocytes. Human myeloid cells transfected with plasmids expressing human CD14 plus a dominant negative Rac1 had reduced adhesion to ICAM-1 after fimbriae stimulation. Plasmids expressing dominant negative Rac1 or TLR2 inhibited fimbriae activation of PI3K. Increased transmigration of fimbriae-exposed monocytes through HUVEC monolayers was abrogated by preincubation of monolayers or monocytes with anti-ICAM-1 mAb or anti-CD11b mAb, respectively. Fimbriae stimulation also increased transmigration of the CD14-transfected cells, whereas transient transfection of the same cells with plasmids expressing a dominant negative Rac1 or TLR2 reduced their CD11b activation, HUVEC adhesion, and transendothelial migration. Wild-type P. gingivalis, but not nonfimbriated mutants, stimulated monocyte adhesion and transmigration. The data show that a novel CD14/TLR2 pathway stimulated by P. ginigivalis fimbriae activates CD11b via Rac1 and PI3K to enhance monocyte binding to ICAM-1 and increase transendothelial migration.
Tyk2 Regulates IL-10 Production
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production by NK and T cells and are highly susceptible to pathogens, presumably due to defective IL-10 signaling. Although current evidence suggests that the same Th1 effector cell produces IFN- and IL-10, factors controlling their production are not known. Shaw et al. (p. 7263
) found that wild-type and Tyk2–/– mice vaccinated with a high dose of Toxoplasma gondii were equally resistant to parasite challenge and had similar frequencies of IFN--producing CD4+ T cells at the site of challenge. Surprisingly, vaccination with a low dose of the parasite protected only the mutant mice from a challenge, even though they had fewer IFN--producing CD4+ T cells than vaccinated and challenged wild-type mice. Phosphorylation of STAT1 and STAT3 proteins in response to IL-10 was less in Tyk2–/– than in wild-type peritoneal macrophages; IL-10 suppressed NO production in the Tyk2–/–cells induced by low-dose, but not high-dose, IFN-. IFN- and IL-10 were produced by peritoneal exudate cells of vaccinated wild-type mice restimulated in vivo; only IFN- was produced after in vitro stimulation. No IL-10 response occurred after challenge of vaccinated Tyk2–/– animals. CD4+ T cell depletion and mAb-blocking experiments showed that CD4+ T cells were the source of IL-10. Peritoneal IL-10 secretion was lower in parasite-challenged vaccinated mice deficient in IL-12- or IFN--signaling components and in wild-type mice treated with anti-IFN- or anti-ICOSL mAb during challenge. Vaccinated and challenged TCR–/– mice reconstituted with CD4+ T cells from Tyk2–/– animals also produced less IL-10. Other adoptive transfer experiments showed that IFN-R–/– CD4+ T cell-reconstituted TCR–/– hosts produced IL-10 upon rechallenge. The authors demonstrate that IL-10 production is controlled by Tyk2 regulation of IL-12-induced IFN- production in effector site CD4+ T cells in rechallenged mice vaccinated with T. gondii. Summaries written by Dorothy L. Buchhagen, Ph.D.
Related articles in The JI:
-Dependent IL-10 Reactivation
-Selected Thymocytes
and PKC Are the Major PKC Isotypes Involved in TCR Down-Regulation
-Actinin
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