Susceptibility to myasthenia gravis (MG) is positively linked to expression of HLA-DQ8 and DR3 molecules and negatively linked to expression of the DQ6 molecule. To elucidate the molecular basis of this association, we have induced experimental autoimmune MG (EAMG) in mice transgenic for HLA-DQ8, DQ6, and DR3, and in DQ8×DQ6 and DQ8×DR3 F1 transgenic mice, by immunization with human acetylcholine receptor (H-AChR) in CFA. Mice expressing transgenes for one or both of the HLA class II molecules positively associated with MG (DQ8 and DR3) developed EAMG. T cells from DQ8 transgenic mice responded well to three cytoplasmic peptide sequences of H-AChR (α320-337, α304-322, and α419-437), of which the response to α320-337 was the most intense. DR3 transgenic mice also responded to this sequence very strongly. H-AChR– and α320-337 peptide–specific lymphocyte responses were restricted by HLA class II molecules. Disease resistance in DQ6 transgenic mice was associated with reduced synthesis of anti-AChR IgG, IgG2b, and IgG2c Ab’s and reduced IL-2 and IFN-γ secretion by H-AChR– and peptide α320-337–specific lymphocytes. Finally, we show that DQ8 imparts susceptibility to EAMG and responsiveness to an epitope within the sequence α320-337 as a dominant trait.
Huan Yang, Elzbieta Goluszko, Chella David, David K. Okita, Bianca Conti-Fine, Teh-sheng Chan, Mathilde A. Poussin, Premkumar Christadoss
Insulin-dependent type 1 diabetes is an autoimmune disease mediated by T lymphocytes recognizing pancreatic islet cell antigens. Glutamic acid decarboxylase 65 (GAD65) appears to be an important autoantigen in the disease. However, T cells from both patients with type 1 diabetes and healthy subjects vigorously proliferate in response to GAD65 stimulation ex vivo, leading us to postulate that the critical event in the onset of human diabetes is the activation of autoreactive T cells. Thus, we investigated whether GAD65-reactive T cells in patients with diabetes functioned as previously activated memory T cells, no longer requiring a second, costimulatory signal for clonal expansion. We found that in patients with new-onset type 1 diabetes, GAD65-reactive T cells were strikingly less dependent on CD28 and B7-1 costimulation to enter into cell cycle and proliferate than were equivalent cells derived from healthy controls. We hypothesize that these autoreactive T cells have been activated in vivo and have differentiated into memory cells, suggesting a pathogenic role in type 1 diabetes. In addition, we observed different effects with selective blockade of either B7-1 or B7-2 molecules; B7-1 appears to deliver a negative signal by engaging CTLA-4, while B7-2 engagement of CD28 upregulates T cell proliferation and cytokine secretion.
Vissia Viglietta, Sally C. Kent, Tihamer Orban, David A. Hafler
P-selectin glycoprotein ligand-1 (PSGL-1) mediates rolling of leukocytes on P-selectin under flow. The glycoproteins that enable leukocyte tethering to or rolling on E-selectin are not known. We used gene targeting to prepare PSGL-1–deficient (PSGL-1–/–) mice, which were healthy but had moderately elevated total blood leukocytes. Fluid-phase E-selectin bound to approximately 70% fewer sites on PSGL-1–/– than PSGL-1+/+ neutrophils. Compared with PSGL-1+/+ leukocytes, significantly fewer PSGL-1–/– leukocytes rolled on E-selectin in vitro, because their initial tethering to E-selectin was impaired. The residual cells that tethered rolled with the same shear resistance and velocities as PSGL-1+/+ leukocytes. Compared with PSGL-1+/+ mice, significantly fewer PSGL-1–/– leukocytes rolled on E-selectin in TNF-α–treated venules of cremaster muscle in which P-selectin function was blocked by an mAb. The residual PSGL-1–/– leukocytes that tethered rolled with slow velocities equivalent to those of PSGL-1+/+ leukocytes. These results reveal a novel function for PSGL-1 in tethering leukocytes to E-selectin under flow.
Lijun Xia, Markus Sperandio, Tadayuki Yago, J. Michael McDaniel, Richard D. Cummings, Sonia Pearson-White, Klaus Ley, Rodger P. McEver
Chemokines are involved in recruitment and activation of hematopoietic cells in sites of infection and inflammation. The M3 gene of the γ-herpesvirus γHV68 encodes an abundant secreted protein that binds CC chemokines with high affinity. We report here that this gene is essential for efficient induction of lethal meningitis by γHV68. An M3 mutant γHV68 (γHV68-M3.stop) was 100-fold less virulent than wild-type or marker rescue control (γHV68-M3.MR) viruses after intracerebral inoculation. After intracerebral inoculation, γHV68-M3.stop grew to lower titers than γHV68 or γHV68-M3.MR in the brain but spread to and grew normally in the spleen and lung. Expression of several CC chemokines was significantly induced in the CNS by γHV68 infection. Consistent with M3 acting by blockade of CC chemokine action, γHV68 induced a neutrophilic meningeal inflammatory infiltrate, while γHV68-M3.stop induced an infiltrate in which lymphocytes and macrophages predominated. In contrast to the important role of M3 in lethal meningitis, M3 was not required for establishment or reactivation from latent infection or induction of chronic arteritis. These data suggest a role for chemokines in the protection of the nervous system from viral infection and that the M3 protein acts in a tissue-specific fashion during acute but not chronic γHV68 infection to limit CC chemokine–induced inflammatory responses.
Victor van Berkel, Beth Levine, Sharookh B. Kapadia, James E. Goldman, Samuel H. Speck, Herbert W. Virgin IV
IL-15, a T cell growth factor, has been linked to exacerbating autoimmune diseases and allograft rejection. To test the hypothesis that IL-15–deficient (IL-15–/–) mice would be protected from T cell–dependent nephritis, we induced nephrotoxic serum nephritis (NSN) in IL-15–/– and wild-type (IL-15+/+) C57BL/6 mice. Contrary to our expectations, IL-15 protects the kidney during this T cell–dependent immunologic insult. Tubular, interstitial, and glomerular pathology and renal function are worse in IL-15–/– mice during NSN. We detected a substantial increase in tubular apoptosis in IL-15–/– kidneys. Moreover, macrophages and CD4 T cells are more abundant in the interstitia and glomeruli in IL-15–/– mice. This led us to identify several mechanisms responsible for heightened renal injury in the absence of IL-15. We now report that IL-15 and the IL-15 receptor (α, β, γ chains) are constitutively expressed in normal tubular epithelial cells (TECs). IL-15 is an autocrine survival factor for TECs. TEC apoptosis induced with anti-Fas or actinomycin D is substantially greater in IL-15–/– than in wild-type TECs. Moreover, IL-15 decreases the induction of a nephritogenic chemokine, MCP-1, that attracts leukocytes into the kidney during NSN. Taken together, we suggest that IL-15 is a therapeutic for tubulointerstitial and glomerular kidney diseases.
Michiya Shinozaki, Junichi Hirahashi, Tatiana Lebedeva, Foo Y. Liew, David J. Salant, Ruth Maron, Vicki Rubin Kelley
Vitiligo is a common depigmenting disorder resulting from the loss of melanocytes in the skin. The pathogenesis of the disease remains obscure, although autoimmune mechanisms are thought to be involved. Indeed, autoantibodies and autoreactive T lymphocytes that target melanocytes have been reported in some vitiligo patients. The objective of this study was to identify pigment cell antigens that are recognized by autoantibodies in vitiligo. Using IgG from vitiligo patients to screen a melanocyte cDNA phage-display library, we identified the melanin-concentrating hormone receptor 1 (MCHR1) as a novel autoantigen related to this disorder. Immunoreactivity against the receptor was demonstrated in vitiligo patient sera by using radiobinding assays. Among sera from healthy controls and from patients with autoimmune disease, none exhibited immunoreactivity to MCHR1, indicating a high disease specificity for Ab’s against the receptor. Inhibition of MCH binding to its receptor by IgG from vitiligo patients was also shown.
E. Helen Kemp, Elizabeth A. Waterman, Brian E. Hawes, Kim O’Neill, Raju V.S.R.K. Gottumukkala, David J. Gawkrodger, Anthony P. Weetman, Philip F. Watson
The antiphospholipid syndrome (APS) is characterized by the presence of pathogenic autoantibodies against β2-glycoprotein-I (β2GPI). The factors causing production of anti-β2GPI remain unidentified, but an association with infectious agents has been reported. Recently, we identified a hexapeptide (TLRVYK) that is recognized specifically by a pathogenic anti-β2GPI mAb. In the present study we evaluated the APS-related pathogenic potential of microbial pathogens carrying sequences related to this hexapeptide. Mice immunized with a panel of microbial preparations were studied for the development of anti-β2GPI autoantibodies. IgG specific to the TLRVYK peptide were affinity purified from the immunized mice and passively infused intravenously into naive mice at day 0 of pregnancy. APS parameters were evaluated in the infused mice on day 15 of pregnancy. Following immunization, high titers of antipeptide [TLRVYK] anti-β2GPI Ab’s were observed in mice immunized with Haemophilus influenzae, Neisseria gonorrhoeae, or tetanus toxoid. The specificity of binding to the corresponding target molecules was confirmed by competition and immunoblot assays. Naive mice infused with the affinity-purified antipeptide Ab’s had significant thrombocytopenia, prolonged activated partial thromboplastin time and elevated percentage of fetal loss, similar to a control group of mice immunized with a pathogenic anti-β2GPI mAb. Our study establishes a mechanism of molecular mimicry in experimental APS, demonstrating that bacterial peptides homologous with β2GPI induce pathogenic anti-β2GPI Ab’s along with APS manifestations.
Miri Blank, Ilan Krause, Mati Fridkin, Nathan Keller, Juri Kopolovic, Iris Goldberg, Ana Tobar, Yehuda Shoenfeld
To date, most studies have focused on the characterization of HIV-1–specific cellular immune responses in the peripheral blood (PB) of infected individuals. Much less is known about the comparative magnitude and breadth of responses in the lymphoid tissue. This study analyzed HIV-1–specific CD8+ T cell responses simultaneously in PB and lymph nodes (LNs) of persons with chronic HIV-1 infection and assessed the dynamics of these responses during antiretroviral treatment and supervised treatment interruption (STI). In untreated chronic infection, the magnitude of epitope-specific CD8+ T cell activity was significantly higher in LNs than in PB. Responses decreased in both compartments during highly active antiretroviral therapy, but this decline was more pronounced in PB. During STI, HIV-1–specific CD8+ T cell responses in PB increased significantly. Enhancement in breadth and magnitude was largely due to the expansion of pre-existing responses in the LNs, with new epitopes infrequently targeted. Taken together, these data demonstrate that HIV-1–specific CD8+ T cells are preferentially located in the LNs, with a subset of responses exclusively detectable in this compartment. Furthermore, the enhanced CD8+ T cell responses observed during STI in chronically infected individuals is largely due to expansion of pre-existing virus-specific CD8+ T cells, rather than the induction of novel responses.
Marcus Altfeld, Jan van Lunzen, Nicole Frahm, Xu G. Yu, Claus Schneider, Robert L. Eldridge, Margaret E. Feeney, Dirk Meyer-Olson, Hans-Juergen Stellbrink, Bruce D. Walker
Agonistic αCD40 Ab’s have been shown to be potent immune adjuvants for both cell- and humoral-mediated immunity. While enhancing short-lived humoral immunity, the administration of a CD40 agonist during thymus-dependent immune responses ablates germinal center formation, prematurely terminates the humoral immune response, blocks the generation of B cell memory, and prevents the generation of long-lived bone marrow plasma cells. Interestingly, some of these effects of heightened CD40 engagement could be mimicked by enhancing the magnitude of antigen-specific T cell help. Taken together, these studies demonstrate that as the magnitude of CD40 signaling intensifies, the fate of antigen-reactive B cells can be dramatically altered. These are the first studies to describe the multifaceted function of CD40 in determining the fate of antigen-reactive B cells and provide novel insights into how CD40 agonists can short-circuit humoral immunity.
Loren D. Erickson, Brigit G. Durell, Laura A. Vogel, Brian P. O’Connor, Marilia Cascalho, Teruhito Yasui, Hitoshi Kikutani, Randolph J. Noelle
Treatment of advanced, poorly immunogenic tumors in animal models, considered the closest simulation available thus far for conditions observed in cancer patients, remains a major challenge for cancer immunotherapy. We reported previously that established tumors in mice receiving an agonistic mAb to the T cell costimulatory molecule 4-1BB (CD137) regress due to enhanced tumor antigen–specific cytotoxic T lymphocyte responses. In this study, we demonstrate that several poorly immunogenic tumors, including C3 tumor, TC-1 lung carcinoma, and B16-F10 melanoma, once established as solid tumors or metastases, are refractory to treatment by anti–4-1BB mAb. We provide evidence that immunological ignorance, rather than anergy or deletion, of tumor antigen–specific CTLs during the progressive growth of tumors prevents costimulation by anti–4-1BB mAb. Breaking CTL ignorance by immunization with a tumor antigen–derived peptide, although insufficient to stimulate a curative CTL response, is necessary for anti–4-1BB mAb to induce a CTL response leading to the regression of established tumors. Our results suggest a new approach for immunotherapy of human cancers.
Ryan A. Wilcox, Dallas B. Flies, Gefeng Zhu, Aaron J. Johnson, Koji Tamada, Andrei I. Chapoval, Scott E. Strome, Larry R. Pease, Lieping Chen