bluuebunny/crossref_metadata_2025 huggingface dataset:
Regulation of Class II Expression in Monocytic Cells after HIV-1 Infection
Authors: Rakoff-Nahoum Seth , Chen Houchu , Kraus Thomas , George Italas , Oei Erwin , Tyorkin Max , Salik Erez , Beuria Prarthana , Sperber Kirk
Published: April 2014

Human macrophage hybridoma cells were used to study HLA-DR expression after HIV-1 infection. HLA-DR surface expression was lost 2 wk after infection that was associated with decreased mRNA transcription. Transfecting HLA-DR-α and HLA-DR-β cDNA driven by a nonphysiological CMV promoter restored expression, suggesting that regulatory DNA-binding proteins may be affected by HIV-1 infection. There was no protein binding to conserved class II DNA elements (W/Z/S box, X-1 and X-2 boxes, and Y box) in a HIV-1-infected human macrophage hybridoma cell line, 43HIV, and in primary monocytes that lost HLA-DR expression after HIV-1BaL infection. PCR analysis of the HIV-1-infected cells that lost HLA-DR expression revealed mRNA for W/Z/S (RFX-5), X-1 (RFX-5), X-2 (hX-2BP), and one Y box DNA-binding protein (NF-YB), and CIITA, a non-DNA-binding protein necessary for class II transcription. There was no mRNA for the Y box-binding protein, NF-YA. However, HLA-DR expression could be restored by transfection with NF-YA driven by a CMV promoter, although HLA-DR failed to localize in either the late endosomes, lysosomes, or acidic compartments. This was associated with a loss of class II-associated invariant chain peptide and leupeptin-induced protein in the 43HIV cells. To address this further, non-HIV-1-infected 43 cells were infected with vaccinia virus containing HIV-1 gag, nef, pol, and env proteins. HLA-DR failed to localize in neither the late endosomes, lysosomes, or acidic compartments in the vaccinia-infected cells containing HIV-1 env protein. HIV-1 appears to have multiple effects on class II expression in monocytic cells that may contribute to the immune defects seen in HIV-1-infected patients.
The Role of αβ+ T Cells and Homeostatic T Cell Proliferation in Y-Chromosome-Associated Murine Lupus
Authors: Lawson Brian R. , Koundouris Stefanos I. , Barnhouse Marlene , Dummer Wolfgang , Baccala Roberto , Kono Dwight H. , Theofilopoulos Argyrios N.
Published: April 2014

Male BXSB mice develop an early life, severe lupus-like disease largely attributed to an undefined Y-chromosome-associated autoimmunity accelerator, termed Yaa. Although the exact disease pathogenesis is uncertain, indirect evidence suggests that T cells play an important role in the male BXSB disease. We have developed TCR α-chain gene-deleted BXSB mice to directly examine the role of αβ+ T cells and the mode by which Yaa promotes disease in this strain. All disease parameters, including hypergammaglobulinemia, autoantibody production, glomerulonephritis, and the unique monocytosis of BXSB males, were severely reduced or absent in the αβ+ T cell-deficient mice. Adoptively transferred CD4+ T cells of either male or female BXSB origin showed equal homeostatic proliferation in αβ+ T cell-deficient male recipients. Moreover, deficient male mice eventually developed equally severe lupus-like disease after adoptive transfer and homeostatic expansion of T cells from wild-type BXSB males or females. The results directly demonstrate that the Yaa-mediated disease requires αβ+ T cells that are not, in themselves, abnormal in either composition or properties, but are engaged by a Yaa-encoded abnormality in a non-T cell component. In addition, homeostatic anti-self proliferation of mature T cells derived from a small number of precursors can induce systemic autoimmunity in an appropriate background.
Fas/Fas Ligand Deficiency Results in Altered Localization of Anti-Double-Stranded DNA B Cells and Dendritic Cells
Authors: Fields Michele L. , Sokol Caroline L. , Eaton-Bassiri Ashlyn , Seo Su-jean , Madaio Michael P. , Erikson Jan
Published: April 2014

Autoantibodies directed against dsDNA are found in patients with systemic lupus erythematosus as well as in mice functionally deficient in either Fas or Fas ligand (FasL) (lpr/lpr or gld/gld mice). Previously, an IgH chain transgene has been used to track anti-dsDNA B cells in both nonautoimmune BALB/c mice, in which autoreactive B cells are held in check, and MRL-lpr/lpr mice, in which autoantibodies are produced. In this study, we have isolated the Fas/FasL mutations away from the autoimmune-prone MRL background, and we show that anti-dsDNA B cells in Fas/FasL-deficient BALB/c mice are no longer follicularly excluded, and they produce autoantibodies. Strikingly, this is accompanied by alterations in the frequency and localization of dendritic cells as well as a global increase in CD4 T cell activation. Notably, as opposed to MRL-lpr/lpr mice, BALB-lpr/lpr mice show no appreciable kidney pathology. Thus, while some aspects of autoimmune pathology (e.g., nephritis) rely on the interaction of the MRL background with the lpr mutation, mutations in Fas/FasL alone are sufficient to alter the fate of anti-dsDNA B cells, dendritic cells, and T cells.
Resistance to Xenobiotic-Induced Autoimmunity Maps to Chromosome 1
Authors: Kono Dwight H. , Park Miyo S. , Szydlik Agnieszka , Haraldsson Katarina M. , Kuan Jason D. , Pearson Deborah L. , Hultman Per , Pollard K. Michael
Published: April 2014

Although evidence indicates that environmental factors play a major role in precipitating systemic autoimmunity in genetically susceptible individuals, little is known about the mechanisms involved. Certain heavy metals, such as mercury, are potent environmental immunostimulants that produce a number of immunopathologic sequelae, including lymphoproliferation, hypergammaglobulinemia, and overt systemic autoimmunity. Predisposition to such metal-induced immunopathology has been shown to be influenced by both MHC and non-MHC genes, as well as susceptibility to spontaneous lupus, in mice and other experimental animals. Among the various mouse strains examined to date, the DBA/2 appears to uniquely lack susceptibility to mercury-induced autoimmunity (HgIA), despite expressing a susceptible H-2 haplotype (H-2d). To define the genetic basis for this trait, two genome-wide scans were conducted using F2 intercrosses of the DBA/2 strain with either the SJL or NZB strains, both of which are highly susceptible to HgIA. A single major quantitative trait locus on chromosome 1, designated Hmr1, was shown to be common to both crosses and encompassed a region containing several lupus susceptibility loci. Hmr1 was linked to glomerular immune complex deposits and not autoantibody production, suggesting that DBA/2 resistance to HgIA may primarily involve the later stages of disease pathogenesis. Identification and characterization of susceptibility/resistance genes and mechanisms relevant to the immunopathogenesis of mercury-induced autoimmunity should provide important insights into the pathogenesis of autoimmunity and may reveal novel targets for intervention.
Inhibition of Autoimmune Diabetes in Nonobese Diabetic Mice by Transgenic Restoration of H2-E MHC Class II Expression: Additive, But Unequal, Involvement of Multiple APC Subtypes
Authors: Johnson Ellis A. , Silveira Pablo , Chapman Harold D. , Leiter Edward H. , Serreze David V.
Published: April 2014

Transgenic restoration of normally absent H2-E MHC class II molecules on APC dominantly inhibits T cell-mediated autoimmune diabetes (IDDM) in nonobese diabetic (NOD) mice. We analyzed the minimal requirements for transgenic H2-E expression on APC subtypes (B lymphocytes vs macrophages/dendritic cells (DC)) to inhibit IDDM. This issue was addressed through the use of NOD stocks transgenically expressing high levels of H2-E and/or made genetically deficient in B lymphocytes in a series of genetic intercross and bone marrow/lymphocyte chimera experiments. Standard (H2-Enull) NOD B lymphocytes exert a pathogenic function(s) necessary for IDDM. However, IDDM was inhibited in mixed chimeras where H2-E was solely expressed on all B lymphocytes. Interestingly, this resistance was abrogated when even a minority of standard NOD H2-Enull B lymphocytes were also present. In contrast, in NOD chimeras where H2-E expression was solely limited to approximately half the macrophages/DC, an active immunoregulatory process was induced that inhibited IDDM. Introduction of a disrupted IL-4 gene into the NOD-H2-E transgenic stock demonstrated that induction of this Th2 cytokine does not represent the IDDM protective immunoregulatory process mediated by H2-E expression. In conclusion, high numbers of multiple subtypes of APC must express H2-E MHC class II molecules to additively inhibit IDDM in NOD mice. This raises a high threshold for success in future intervention protocols designed to inhibit IDDM by introduction of putatively protective MHC molecules into hemopoietic precursors of APC.
Down-Regulation of IgE and IgG4 Antibodies to Tetanus Toxoid and Diphtheria Toxoid by Covaccination with Cellular Bordetella pertussis Vaccine
Authors: Grüber Christoph , Lau Susanne , Dannemann Almut , Sommerfeld Christine , Wahn Ulrich , Aalberse Rob C.
Published: April 2014

Pertussis (P) toxin acts as adjuvant for IgE formation against simultaneously administered Ags in animal models. P vaccination may also have an adjuvant impact on IgE formation against coadministered diphtheria (D) and tetanus (T) Ags in humans. Sera of 103 D-T-P-immunized and 319 D-T-immunized children aged 2 years were analyzed for IgE, IgG4, and IgG to D and T (radioallergosorbent test), total IgE and IgE against common inhalant allergens (CAP radioallergosorbent test fluoroenzyme immunoassay). Fewer D-T-P- than D-T-immunized children had sera positive for T-IgE (12.6 vs 53.6%, p < 0.001), T-IgG4 (71.6 vs 89.2%, p < 0.001), D-IgE (31.0 vs 70.5%, p < 0.001), and D-IgG4 (85.2 vs 93.4%, p = 0.039). Suppression of T-IgE was not dependent on the cutoff chosen for a positive test result, but was dependent on the proportion of D-T immunizations given with P. The risk for sensitization to common environmental allergens did not differ (odds ratio 0.953, 95% confidence interval 0.815–1.114). No significant differences between D-T- and D-T-P-immunized children were found with regard to T-IgG or D-IgG. In summary, IgE and IgG4 (but not IgG) serum levels to coadministered D- and T-Ags are suppressed among P-immunized children as compared with nonimmunized children. These results suggest that the presence of a microbial product during Ag exposure can down-regulate an IgE/IgG4 response in humans.
Cutting Edge: STAT Activation By IL-19, IL-20 and mda-7 Through IL-20 Receptor Complexes of Two Types
Authors: Dumoutier Laure , Leemans Caroline , Lejeune Diane , Kotenko Sergei V. , Renauld Jean-Christophe
Published: April 2014

IL-10-related cytokines include IL-20 and IL-22, which induce, respectively, keratinocyte proliferation and acute phase production by hepatocytes, as well as IL-19, melanoma differentiation-associated gene 7, and AK155, three cytokines for which no activity nor receptor complex has been described thus far. Here, we show that mda-7 and IL-19 bind to the previously described IL-20R complex, composed by cytokine receptor family 2–8/IL-20Rα and DIRS1/IL-20Rβ (type I IL-20R). In addition, mda-7 and IL-20, but not IL-19, bind to another receptor complex, composed by IL-22R and DIRS1/IL20Rβ (type II IL-20R). In both cases, binding of the ligands results in STAT3 phosphorylation and activation of a minimal promoter including STAT-binding sites. Taken together, these results demonstrate that: 1) IL-20 induces STAT activation through IL-20R complexes of two types; 2) mda-7 and IL-20 redundantly signal through both complexes; and 3) IL-19 signals only through the type I IL-20R complex.
Cutting Edge: Differential Regulation of Chemoattractant Receptor-Induced Degranulation and Chemokine Production by Receptor Phosphorylation
Authors: Ahamed Jasimuddin , Haribabu Bodduluri , Ali Hydar
Published: April 2014

Phosphorylation of G protein-coupled receptors and the subsequent recruitment of β-arrestin play an important role in desensitization of receptor-mediated responses, including degranulation in leukocytes. In this study, we report that receptor phosphorylation also provides a stimulatory signal for CCR ligand 2 (CCL2) production. C3a stimulated degranulation in a basophilic leukemia RBL-2H3 cell expressing wild-type C3aR or a phosphorylation-deficient mutant (ΔST-C3aR). In contrast, C3a caused CCL2 production only in C3aR but not ΔST-C3aR cells. Furthermore, overexpression of G protein-coupled receptor kinase 2 resulted in enhancement of both ligand-induced receptor phosphorylation and CCL2 production but inhibition of degranulation. Agonist activation of C3aR, but not ΔST-C3aR, led to the translocation of green fluorescent protein tagged β-arrestin 2 from the cytoplasm to the plasma membrane. These data demonstrate that receptor phosphorylation, which provides a turn off signal for degranulation, is essential for CCL2 production.
Molecular Cloning of F4/80-Like-Receptor, a Seven-Span Membrane Protein Expressed Differentially by Dendritic Cell and Monocyte-Macrophage Subpopulations
Authors: Caminschi Irina , Lucas Karen M. , O’Keeffe Meredith A. , Hochrein Hubertus , Laâbi Yacine , Köntgen Frank , Lew Andrew M. , Shortman Ken , Wright Mark D.
Published: April 2014

A novel dendritic cell (DC) surface molecule termed F4/80-like-receptor (FIRE) has been selected based on its differential expression between DC subsets. The gene encoding FIRE has been cloned and sequenced, and mAbs specific for FIRE have been produced. FIRE is a seven-transmembrane-spanning molecule with two epidermal growth factor-like domains in the extracellular region. It is a novel member of the epidermal growth factor/transmembrane-7 protein subfamily and shows similarity to the macrophage marker F4/80. FIRE is expressed by CD8− DC, but not by CD8+ DC, and it is down-regulated on DC activation. It is expressed by blood monocytes and by some tissue macrophages, but not by most macrophage cell lines or by lymphoid cells. FIRE is a useful marker of myeloid cells with a DC developmental potential.
Plasmid DNA Encoding CCR7 Ligands Compensate for Dysfunctional CD8+ T Cell Responses by Effects on Dendritic Cells
Authors: Eo Seong Kug , Kumaraguru Udayasankar , Rouse Barry T.
Published: April 2014

Lymphotoxin α-deficient (LTα−/−) mice, which lack lymph nodes and possess a disorganized spleen, develop dysfunctional CD8+ T cells upon HSV infection and readily succumb to herpes encephalitis. Such mice do develop apparently normal peptide-specific CD8+ T cell responses, as measured by MHC class I tetramer staining, but the majority of cells fail to become cytotoxic or express peptide-induced IFN-γ production. In the present study, we demonstrate that functional defects of CD8+ T cells in LTα−/− mice can be largely rectified by the administration of plasmid DNA encoding CCR7 ligands before HSV infection. Treated mutant mice developed increased peptide-specific cytotoxic responses, enhanced numbers of CD8+ T cells capable of producing IFN-γ, as well as improved resistance to HSV challenge. The corrective effect of chemokine treatment appeared to result from improved dendritic cell-mediated Ag presentation. Thus, a major consequence of the treatment was an increase in splenic dendritic cell number in CCR7 ligand-treated LTα−/− mice with such splenocyte populations showing improved APC activity in vitro. Our results document that functional defects of CD8+ T cells can be corrected, and indicate the value of plasmid vector encoding appropriate chemokines to achieve such immunotherapy.
TGF-β-Producing CD4+ Mediastinal Lymph Node Cells Obtained from Mice Tracheally Tolerized to Ovalbumin (OVA) Suppress Both Th1- and Th2-Induced Cutaneous Inflammatory Responses to OVA by Different Mechanisms
Authors: Terui Tadashi , Sano Kunio , Shirota Hidekazu , Kunikata Nagisa , Ozawa Maki , Okada Mikiko , Honda Motoko , Tamura Gen , Tagami Hachiro
Published: April 2014

Advances in the treatment of allergic disorders require elucidation of the autoregulatory immune systems induced in averting detrimental inflammatory responses against invading foreign Ags. We previously reported that excessive Ags intruding through the airway mucosa induce a subset of regulatory CD4+ T cells secreting TGF-β in the regional mediastinal lymph nodes (MLNs), which inhibits Th2 cells and subsequent eosinophilic inflammation in the trachea. In the present experiments we examined whether and in what mechanisms TGF-β-secreting CD4+ T cells in the MLNs regulate Th cell-mediated skin inflammation using a previously established murine model. Th1 or Th2 cells injected s.c. into ear lobes of naive mice induced swelling, whereas the concomitant local injection of MLN cells suppressed the inflammation. The suppressor activities of MLN cells were markedly neutralized by anti-TGF-β mAb and were mimicked by rTGF-β. The MLN cell- and rTGF-β-induced inhibition was reversed by anti-IL-10 mAb significantly in Th1-induced inflammation and only partially in Th2-induced inflammation. rIL-10 reduced Th-induced ear swelling, although higher doses of rIL-10 were required in Th2-induced one. Thus, allergen-specific TGF-β-producing CD4+ T cells induced in the respiratory tract controlled cutaneous inflammatory responses by Th1 or Th2 cells either directly by TGF-β or indirectly through IL-10 induction. From a clinical standpoint, these observations might explain the mechanism of spontaneous regression in some patients with atopic dermatitis, which exhibits both Th1- and Th2-mediated skin inflammation in response to airborne protein Ags.
Morphine Inhibits Mucosal Antibody Responses and TGF-β mRNA in Gut-Associated Lymphoid Tissue Following Oral Cholera Toxin in Mice
Authors: Peng Xiaohui , Cebra John J. , Adler Martin W. , Meissler Joseph J. , Cowan Alan , Feng Pu , Eisenstein Toby K.
Published: April 2014

In this study, we investigated the effect of morphine on the mucosal immune system using fragment cultures of ileal segments, Peyer’s patches (PPs), and mesenteric lymph nodes. Mice were implanted s.c. with a morphine slow release pellet. Control groups received a naltrexone slow release pellet, a placebo pellet, or both a morphine and a naltrexone pellet. After 48 h, mice were orally immunized with cholera toxin (CT) and were boosted orally 1 wk later. Animals were sacrificed 1 wk after the booster immunization, and PPs, mesenteric lymph nodes, and ileal segments were cultured in 24-well plates for 12 days. Morphine resulted in a highly significant inhibition of CT-specific IgA and IgG production in fragment culture supernatants of all three tissues compared with placebo. Naltrexone blocked the reduction in Ab levels induced by morphine, indicating that the effect is opioid receptor mediated. Morphine did not significantly alter total IgA levels in any of the tissue culture supernatants. Morphine also inhibited CT-specific IgA and IgG levels in serum. By flow cytometry, morphine did not alter the lymphoid cell composition in PPs compared with placebo. The effect of morphine on TGF-β, IL-5, and IL-6 mRNA expression in PPs and ileal segments was determined following oral immunization with CT. Morphine significantly decreased TGF-β mRNA compared with that in the placebo group, and naltrexone blocked this effect. These results indicate that morphine inhibits Ag-specific IgA responses in gut-associated lymphoid tissue at least partially through the inhibition of TGF-β, a putative IgA switch factor, in the gastrointestinal tract.
A Closer Look at Homeostatic Proliferation of CD4+ T Cells: Costimulatory Requirements and Role in Memory Formation
Authors: Gudmundsdottir Hrefna , Turka Laurence A.
Published: April 2014

Ag-specific proliferation of CD4+ T cells is regulated, in part, by costimulatory signals through CD28. The proliferative response during primary activation is an important determinant of the ability of the T cell to respond to Ag re-encounter. Proliferation of mature CD4+ T cells during lymphopenia (homeostatic proliferation) requires interaction with endogenous peptide MHC. However, the role of costimulation during homeostatic proliferation is unclear, as is the ability of homeostatic proliferation to regulate secondary T cell responses. Using a TCR transgenic system and serial adoptive transfers we find that homeostatic proliferation of CD4+ T cells occurs for at least 5 wk after adoptive transfer into recombination-activating gene (RAG)−/− recipients. Two discrete populations of proliferating T cells can be resolved, one that is highly proliferative and dependent on CD28 signaling, and the other that contains cells undergoing low levels of CD28-independent proliferation. Importantly, naive CD4+ T cells that have undergone homeostatic proliferation acquire both phenotypic and functional characteristics of true memory cells. These studies indicate that functional memory T cells can be generated by encounters with endogenous Ags only. This mechanism of T cell regeneration is possibly active during lymphopenia due to viral infections, such as HIV, transplantation, or cancer therapy, and may explain selected autoimmune diseases.
Antigen-Specific Modulation of an Immune Response by In Vivo Administration of Soluble MHC Class I Tetramers
Authors: Maile Robert , Wang Bo , Schooler Wesley , Meyer Anthony , Collins Edward J. , Frelinger Jeffrey A.
Published: April 2014

Soluble MHC/peptide tetramers that can directly bind the TCR allow the direct visualization and quantitation of Ag-specific T cells in vitro and in vivo. We used HY-Db tetramers to assess the numbers of HY-reactive CD8+ T cells in HYTCR-transgenic mice and in naive, wild-type C57BL/6 (B6) mice. As expected, tetramer staining showed the majority of T cells were male-specific CD8+ T cells in female HY-TCR mice. Staining of B6 mice showed a small population of male-specific CD8+ T cells in female mice. The effect of administration of soluble MHC class I tetramers on CD8+ T cell activation in vivo was unknown. Injection of HY-Db tetramer in vivo effectively primed female mice for a more rapid proliferative response to both HY peptide and male splenocytes. Furthermore, wild-type B6 female mice injected with a single dose of HY-Db tetramer rejected B6 male skin grafts more rapidly than female littermates treated with irrelevant tetramer. In contrast, multiple doses of HY-Db tetramer did not further decrease graft survival. Rather, female B6 mice injected with multiple doses of HY-Db tetramer rejected male skin grafts more slowly than mice primed with a single injection of tetramer or irradiated male spleen cells, suggesting clonal exhaustion or anergy. Our data highlight the ability of soluble MHC tetramers to identify scarce alloreactive T cell populations and the use of such tetramers to directly modulate an Ag-specific T cell response in vivo.
Integrin Functions Play a Key Role in the Differentiation of Thymocytes In Vivo
Authors: Schmeissner Peter J. , Xie Haichun , Smilenov Lubomir B. , Shu Fengyu , Marcantonio Eugene E.
Published: April 2014

T cells express a variety of surface proteins as they develop to maturity in the thymus. In addition to the TCR-CD3 complex and the two major coreceptors, CD4 and CD8, other surface proteins expressed include receptors for cytokines, growth factors, counterreceptors, and extracellular matrix molecules. To determine the role of integrin adhesion receptors in T cell development, we have expressed a trans-dominant inhibitor of integrin function in the thymus. This inhibitor leads to a block of adhesion to fibronectin due to reduced activation of integrin receptors. This reduced adhesion leads to a partial block in differentiation from CD4−CD8− cells to CD4+CD8+ cells, after the CD25+ stage, suggesting that integrins are important during Lck-mediated differentiation. Furthermore, the overall production of CD4+ cells is reduced compared with that of CD8+ cells without changes in negative selection, suggesting that integrins may be involved in the determination of the fate of the cell as well. These results demonstrate that integrin receptor function is required for proper thymocyte development in vivo.
IL-4 Therapy Prevents the Development of Proteinuria in Active Heymann Nephritis by Inhibition of Tc1 Cells
Authors: Spicer S. Timothy , Ha Hong , Boyd Rochelle A. , He Xiao Y. , Carter Nicole , Tran Giang , Penny Mark J. , Hodgkinson Suzanne J. , Hall Bruce M.
Published: April 2014

The role of IL-4, a key Th2 cytokine, in promoting or inhibiting active Heymann nephritis (HN) was examined. HN is induced by immunization with Fx1A in CFA, and proteinuria in HN is associated with subepithelial IgG and C3 deposition and infiltration of CD8+ T-cytotoxic 1 (Tc1) cells and macrophages into glomeruli, as well as induction of Abs to Crry. Treatment with rIL-4 from the time of Fx1A/CFA immunization stimulated an earlier IgG1 response to Fx1A, induced anti-Crry Abs, and up-regulated IL-4 mRNA in lymphoid tissue, but did not alter proteinuria. Treatment with MRCOx-81, an IL-4-blocking mAb, resulted in greater proteinuria, which suggests endogenous IL-4 regulated the autoimmune response. Delay of rIL-4 treatment until 4 wk post-Fx1A/CFA immunization and just before the onset of proteinuria prevented the development of proteinuria and reduced Tc1 cell infiltrate in glomeruli. Delayed treatment with IL-4 had no effect on titer or isotype of Abs to Fx1A or on Ig, C3, and C9 accumulation in glomeruli. Treatment with rIL-13, a cytokine that alters macrophage function such as rIL-4, but has no direct effect on T or B cell function, reduced glomerular macrophage infiltrate, but did not prevent proteinuria or CD8+ T cell infiltrate. Anti-Crry Abs were paradoxically only induced with rIL-4 therapy, not in HN controls with proteinuria. It was concluded that the rIL-4 effect was probably by inhibition of Tc1 cells, which normally mediate the glomerular injury that results in proteinuria.
Analysis of VpreB Expression During B Lineage Differentiation in λ5-Deficient Mice
Authors: Stephan Robert P. , Elgavish Eynav , Karasuyama Hajime , Kubagawa Hiromi , Cooper Max D.
Published: April 2014

The VpreB/λ5 surrogate L chain complex is an essential component of the pre-B cell receptor, the expression of which serves as an important checkpoint in B cell development. Surrogate L chains also may serve as components of murine pro-B cell receptors whose function is unknown. We have produced two new mAbs, R3 and R5, that recognize a different VpreB epitope than the one recognized by the previously described VP245 anti-mouse VpreB Ab. These Abs were used to confirm the expression of surrogate L chains on wild-type pro-B and pre-B cell lines. Although undetectable on the cell surface, VpreB was found to be normally expressed within B lineage cells of λ5-deficient mice. Nevertheless, VpreB expression was extinguished at the B cell stage of differentiation in these mice. The normal pattern of VpreB expression in λ5-deficient mice excludes an essential role for pro-B and pre-B cell receptors in VpreB regulation.
Induction of TCR Vβ-Specific CD8+ CTLs by TCR Vβ-Derived Peptides Bound to HLA-E
Authors: Li Jianfeng , Goldstein Itamar , Glickman-Nir Eva , Jiang Hong , Chess Leonard
Published: April 2014

Previous studies have identified murine and human regulatory CD8+ T cells specific for TCR-Vβ families expressed on autologous activated CD4+ T cells. In the mouse, these regulatory CD8+ T cells were shown to be restricted by the MHC class Ib molecule, Qa-1. In the present study, we asked whether HLA-E, the human functional equivalent of Qa-1, binds Vβ peptides and whether the HLA-E/Vβ-peptide complex induces and restricts human CD8+ CTLs. We first created stable HLA-E gene transfectants of the C1R cell line (C1R-E). Two putative HLA-E binding nonapeptides identified in human TCR Vβ1 and Vβ2 chains (SLELGDSAL and LLLGPGSGL, respectively) were shown to bind to HLA-E. CD8+ T cells could be primed in vitro by C1R-E cells loaded with the Vβ1 (C1R-E/V1) or Vβ2 (C1R-E/V2) peptide to preferentially kill C1R-E cells loaded with the respective inducing Vβ peptide, compared with targets loaded with the other peptides. Priming CD8+ T cells with untreated C1R-E cells did not induce Vβ-specific CTLs. Of perhaps more physiological relevance was the finding that the CD8+ CTLs primed by C1R-E/V1 also preferentially killed activated autologous TCR Vβ1+. Similar results were observed in reciprocal experiments using C1R-E/V2 for priming. Furthermore, anti-CD8 and anti-MHC class I mAbs inhibited this Vβ-specific killing of C1R-E and CD4+ T cell targets. Taken together, the data provide evidence that certain TCR-Vβ peptides can be presented by HLA-E to further induce Vβ-specific CD8+ CTLs.
Vitamin E-Enhanced IL-2 Production in Old Mice: Naive But Not Memory T Cells Show Increased Cell Division Cycling and IL-2-Producing Capacity
Authors: Adolfsson Oskar , Huber Brigitte T. , Meydani Simin Nikbin
Published: April 2014

Aging is associated with reduced T cell function, as demonstrated by decreased T cell proliferation and IL-2 production. These changes respond to supplemental vitamin E both in animals and humans, in part by the reduction of T cell suppressive PGE2, the production of which by macrophages is increased with age. To evaluate whether vitamin E has a direct PGE2-independent effect on T cell responses, T cells purified from the spleens of young and old mice were preincubated with vitamin E or vehicle control. Activation-induced cell division of T cells from old mice was lower than that by young, and the production of IL-2 following 48-h activation was less by T cells from old mice. There was an age-related decline in both the number of IL-2+ T cells and the amount of IL-2 produced per cell. Despite decreased IL-2 protein at 48 h, the expression of IL-2 mRNA at 6 h and IL-2 protein production at 6 and 16 h was greater by T cells from old mice compared with that of young. Age-related decline in cell division and IL-2 production at 48 h was only observed within the naive T cell subpopulation. Vitamin E increased both cell-dividing and IL-2-producing capacity of naive T cells from old mice, with no effect on memory T cells. These data indicate that naive T cells exhibit the greatest age-related defect and show for the first time that supplemental vitamin E has direct immunoenhancing effect on naive T cells from old mice.
Low-Avidity Self-Specific T Cells Display a Pronounced Expansion Defect That Can Be Overcome by Altered Peptide Ligands
Authors: de Visser Karin E. , Cordaro Tanina A. , Kessels Helmut W. H. G. , Tirion Felicia H. , Schumacher Ton N. M. , Kruisbeek Ada M.
Published: April 2014

Thymic expression of self-Ags results in the deletion of high-avidity self-specific T cells, but, at least for certain Ags, a residual population of self-specific T cells with low-affinity TCRs remains after negative selection. Such self-specific T cells are thought to play a role in the induction of T cell-mediated autoimmunity, but may also be used for the induction of antitumor immunity against self-Ags. In this study, we examine the functional competence of a polyclonal population of self-specific CD8+ T cells. We show that low-affinity interactions between TCR and peptide are associated with selective loss of critical T cell functions. Triggering of low levels of IFN-γ production and cytolytic activity through low-affinity TCRs readily occurs provided high Ag doses are used, but IL-2 production and clonal expansion are severely reduced at all Ag doses. Remarkably, a single peptide variant can form an improved ligand for the highly diverse population of low-avidity self-specific T cells and can improve their proliferative capacity. These data provide insight into the inherent limitations of self-specific T cell responses through low-avidity TCR signals and the effect of modified peptide ligands on self-specific T cell immunity.
Function and Factor Interactions of a Locus Control Region Element in the Mouse T Cell Receptor-α/Dad1 Gene Locus
Authors: Ortiz Benjamin D. , Harrow Faith , Cado Dragana , Santoso Buyung , Winoto Astar
Published: April 2014

Locus control regions (LCRs) refer to cis-acting elements composed of several DNase I hypersensitive sites, which synergize to protect transgenes from integration-site dependent effects in a tissue-specific manner. LCRs have been identified in many immunologically important gene loci, including one between the TCRδ/TCRα gene segments and the ubiquitously expressed Dad1 gene. Expression of a transgene under the control of all the LCR elements is T cell specific. However, a subfragment of this LCR is functional in a wide variety of tissues. How a ubiquitously active element can participate in tissue-restricted LCR activity is not clear. In this study, we localize the ubiquitously active sequences of the TCR-α LCR to an 800-bp region containing a prominent DNase hypersensitive site. In isolation, the activity in this region suppresses position effect transgene silencing in many tissues. A combination of in vivo footprint examination of this element in widely active transgene and EMSAs revealed tissue-unrestricted factor occupancy patterns and binding of several ubiquitously expressed transcription factors. In contrast, tissue-specific, differential protein occupancies at this element were observed in the endogenous locus or full-length LCR transgene. We identified tissue-restricted AML-1 and Elf-1 as proteins that potentially act via this element. These data demonstrate that a widely active LCR module can synergize with other LCR components to produce tissue-specific LCR activity through differential protein occupancy and function and provide evidence to support a role for this LCR module in the regulation of both TCR and Dad1 genes.
Conventional and Surrogate Light Chains Differentially Regulate Ig μ and Dμ Heavy Chain Maturation and Surface Expression
Authors: Fang Terry , Smith Brendan P. , Roman Christopher A. J.
Published: April 2014

Positive selection of precursor (pre-) B cells by Ig membrane μ H chains (μm HC) and counterselection mediated by the truncated HC Dμ depend on the ability of each HC to form a pre-B cell receptor (pre-BCR) signaling complex with the surrogate L chain (SLC) components λ5 and Vpre-B. To better understand how pre-BCR signaling output is determined by its Ig components and the SLC, we investigated the regulation of pre-BCR surface expression and HC secretory maturation in a new nonlymphoid system. We took this approach as a means to distinguish B-lineage-specific effects from pre-BCR-intrinsic properties that may influence these aspects of pre-BCR homeostasis necessary for signaling. As in pre-B cells, the SLC in nonlymphoid cells supported only a limited degree of μm HC maturation and low pre-BCR surface expression levels compared with conventional LCs, indicating that this was due to an intrinsic property of the SLC. We identified the non-Ig region of λ5 as harboring the restrictive activity responsible for this phenotype. This property of λ5 was also evident with Dμ, but the overall SLC- and L chain-dependent requirements for Dμ maturation and surface expression were markedly different from those for μm. Surprisingly, Dμ was modified in an unusual manner that was only dependent on Vpre-B. These results establish a novel function of λ5 in limiting surface pre-BCR levels and reveal biochemical properties of Ig molecules that may underlie the diverse consequences of pre-BCR signaling in vivo by different HCs.
A Novel, Nonclassical MHC Class I Molecule Specific to the Common Chimpanzee
Authors: Adams Erin J. , Cooper Stewart , Parham Peter
Published: April 2014

All expressed human MHC class I genes (HLA-A, -B, -C, -E, -F, and -G) have functional orthologues in the MHC of the common chimpanzee (Pan troglodytes). In contrast, a nonclassical MHC class I gene discovered in the chimpanzee is not present in humans or the other African ape species. In exons and more so in introns, this Patr-AL gene is similar to the expressed A locus in the orangutan, Popy-A, suggesting they are orthologous. Patr-AL/Popy-A last shared a common ancestor with the classical MHC-A locus >20 million years ago. Population analysis revealed little Patr-AL polymorphism: just three allotypes differing only at residues 52 and 91. Patr-AL is expressed in PBMC and B cell lines, but at low level compared with classical MHC class I. The Patr-AL polypeptide is unusually basic, but its glycosylation, association with β2-microglobulin, and antigenicity at the cell surface are like other MHC class I. No Patr-AL-mediated inhibition of polyclonal chimpanzee NK cells was detected. The Patr-AL gene is present in 50% of chimpanzee MHC haplotypes, correlating with presence of a 9.8-kb band in Southern blots. The flanking regions of Patr-AL contain repetitive/retroviral elements not flanking other class I genes. In sequenced HLA class I haplotypes, a similar element is present in the A*2901 haplotype but not the A*0201 or A*0301 haplotypes. This element, 6 kb downstream of A*2901, appears to be the relic of a human gene related to Patr-AL. Patr-AL has characteristics of a class I molecule of innate immunity with potential to provide common chimpanzees with responses unavailable to humans.
Isolated Hypervariable Regions Derived from Streptococcal M Proteins Specifically Bind Human C4b-Binding Protein: Implications for Antigenic Variation
Authors: Morfeldt Eva , Berggård Karin , Persson Jenny , Drakenberg Torbjörn , Johnsson Eskil , Lindahl Erik , Linse Sara , Lindahl Gunnar
Published: April 2014

Antigenic variation in microbial surface proteins represents an apparent paradox, because the variable region must retain an important function, while exhibiting extensive immunological variability. We studied this problem for a group of streptococcal M proteins in which the ∼50-residue hypervariable regions (HVRs) show essentially no residue identity but nevertheless bind the same ligand, the human complement regulator C4b-binding protein (C4BP). Synthetic peptides derived from different HVRs were found to retain the ability to bind C4BP, implying that the HVR corresponds to a distinct ligand-binding domain that can be studied in isolated form. This finding allowed direct characterization of the ligand-binding properties of isolated HVRs and permitted comparisons between different HVRs in the absence of conserved parts of the M proteins. Affinity chromatography of human serum on immobilized peptides showed that they bound C4BP with high specificity and inhibition experiments indicated that different peptides bound to the same site in C4BP. Different C4BP-binding peptides did not exhibit any immunological cross-reactivity, but structural analysis suggested that they have similar folds. These data show that the HVR of streptococcal M protein can exhibit extreme variability in sequence and immunological properties while retaining a highly specific ligand-binding function.
Monoclonal Antibodies with Broad Specificity for Hepatitis C Virus Hypervariable Region 1 Variants Can Recognize Viral Particles
Authors: Cerino Antonella , Meola Annalisa , Segagni Laura , Furione Milena , Marciano Sabrina , Triyatni Miriam , Liang T. Jake , Nicosia Alfredo , Mondelli Mario U.
Published: April 2014

The hypervariable region 1 (HVR1) of the E2 protein of hepatitis C virus (HCV) is a highly heterogeneous sequence that is promiscuously recognized by human sera via binding to amino acid residues with conserved physicochemical properties. We generated a panel of mAbs from mice immunized with HVR1 surrogate peptides (mimotopes) affinity-selected with sera from HCV-infected patients from a phage display library. A high number of specific clones was obtained after immunization with a pool of nine mimotopes, and the resulting mAbs were shown to recognize several 16- and 27-mer peptides derived from natural HVR1 sequences isolated from patients with acute and chronic HCV infection, suggesting that HVR1 mimotopes were efficient antigenic and immunogenic mimics of naturally occurring HCV variants. Moreover, most mAbs were shown to bind HVR1 in the context of a complete soluble form of the E2 glycoprotein, indicating recognition of correctly folded HVR1. In addition, a highly promiscuous mAb was able to specifically capture bona fide viral particles (circulating HCV RNA) as well as rHCV-like particles assembled in insect cells expressing structural viral polypeptides derived from an HCV 1a isolate. These findings demonstrate that it is possible to induce a broadly cross-reactive clonal Ab response to multiple HCV variants. In consideration of the potentially important role of HVR1 in virus binding to cellular receptor(s), such a mechanism could be exploited for induction of neutralizing Abs specific for a large repertoire of viral variants.
Critical Role of Kupffer Cell-Derived IL-10 for Host Defense in Septic Peritonitis
Authors: Emmanuilidis Klaus , Weighardt Heike , Maier Stefan , Gerauer Klaus , Fleischmann Tanja , Zheng Xin X. , Hancock Wayne W. , Holzmann Bernhard , Heidecke Claus-Dieter
Published: April 2014

Intra-abdominal infection in patients following major visceral surgery is associated with high mortality. Using a macrophage depletion technique, we demonstrate that in murine septic peritonitis, Kupffer cells are a major source of systemic IL-10 levels. Kupffer cell-depleted mice were highly susceptible to the lethal effects of septic peritonitis and exhibited an increased bacterial load. Kupffer cell-depleted mice were protected by the administration of an IL-10-Fc fusion protein. Loss of Kupffer cell-derived IL-10 was associated with a weak increase in serum IL-12 levels, whereas TNF, IL-1α, and IL-18 levels were not significantly elevated, suggesting that the loss of Kupffer cell-derived IL-10 did not result in a toxic cytokine release syndrome. Instead, loss of Kupffer cell-derived IL-10 was associated with a reduced splenocyte production of IFN-γ that is required for immune protection in murine septic peritonitis. Therefore, the results suggest that the protective function of IL-10 in septic peritonitis may not be restricted to the anti-inflammatory activities of IL-10.
HSV and Glycoprotein J Inhibit Caspase Activation and Apoptosis Induced by Granzyme B or Fas
Authors: Jerome Keith R. , Chen Zheng , Lang Robin , Torres Monika R. , Hofmeister Joni , Smith Shannon , Fox Richard , Froelich Christopher J. , Corey Lawrence
Published: April 2014

HSV-1 inhibits apoptosis of infected cells, presumably to ensure that the infected cell survives long enough to allow completion of viral replication. Because cytotoxic lymphocytes kill their targets via the induction of apoptosis, protection from apoptosis could constitute a mechanism of immune evasion for HSV. Several HSV genes are involved in the inhibition of apoptosis, including Us5, which encodes glycoprotein J (gJ). Viruses deleted for Us5 showed defects in inhibition of caspase activation after Fas ligation or UV irradiation. Transfected cells expressing the Us5 gene product gJ were protected from Fas- or UV-induced apoptosis, as measured by morphology, caspase activation, membrane permeability changes, or mitochondrial transmembrane potential. In contrast, caspase 3 activation in mitochondria-free cell lysates by granzyme (gr)B was inhibited equivalently by Us5 deletion and rescue viruses, suggesting that gJ is not required for HSV to inhibition this process. However, mitochondria-free lysates from transfected cells expressing Us5/gJ were protected from grB-induced caspase activation, suggesting that Us5/gJ is sufficient to inhibit this process. Transfected cells expressing Us5/gJ were also protected from death induced by incubation with purified grB and perforin. These findings suggest that HSV has a comprehensive set of immune evasion functions that antagonize both Fas ligand- and grB-mediated pathways of CTL-induced apoptosis. The understanding of HSV effects on killing by CTL effector mechanisms may shed light on the incomplete control of HSV infections by the immune system and may allow more rational approaches to the development of immune modulatory treatments for HSV infection.
Abnormal Migration Phenotype of Mitogen-Activated Protein Kinase-Activated Protein Kinase 2−/− Neutrophils in Zigmond Chambers Containing Formyl-Methionyl-Leucyl-Phenylalanine Gradients
Authors: Hannigan Michael O. , Zhan Lijun , Ai Youxi , Kotlyarov Alexey , Gaestel Matthias , Huang Chi-Kuang
Published: April 2014

Time-lapsed video microscopy and confocal imaging were used to study the migration of wild-type (WT) and mitogen-activated protein kinase-activated protein kinase 2 (MK2)−/− mouse neutrophils in Zigmond chambers containing fMLP gradients. Confocal images of polarized WT neutrophils showed an intracellular gradient of phospho-MK2 from the anterior to the posterior region of the neutrophils. Compared with WT neutrophils, MK2−/− neutrophils showed a partial loss of directionality but higher migration speed. Immunoblotting experiments showed a lower protein level of p38 mitogen-activated protein kinase and a loss of fMLP-induced extracellular signal-related kinase phosphorylation in MK2−/− neutrophils. These results suggest that MK2 plays an important role in the regulation of neutrophil migration and may also affect other signaling molecules.
A Novel Long Chain Polyunsaturated Fatty Acid, β-Oxa 21:3n-3, Inhibits T Lymphocyte Proliferation, Cytokine Production, Delayed-Type Hypersensitivity, and Carrageenan-Induced Paw Reaction and Selectively Targets Intracellular Signals
Authors: Costabile Maurizio , Hii Charles S. T. , Robinson Brenton S. , Rathjen Deborah A. , Pitt Michael , Easton Christopher , Miller Robert C. , Poulos Alf , Murray Andrew W. , Ferrante Antonio
Published: April 2014

A novel polyunsaturated fatty acid (PUFA), β-oxa 21:3n-3, containing an oxygen atom in the β position, was chemically synthesized, and found to have more selective biological activity than the n-3 PUFA, docosahexaenoic acid (22:6n-3) on cells of the immune system. Although β-oxa 21:3n-3 was very poor compared with 22:6n-3 at stimulating oxygen radical production in neutrophils, it was more effective at inhibiting human T lymphocyte proliferation (IC50 of 1.9 vs 5.2 μM, respectively). β-Oxa 21:3n-3 also inhibited the production of TNF-β, IFN-γ, and IL-2 by purified human T lymphocytes stimulated with PHA plus PMA, anti-CD3 plus anti-CD28 mAbs, or PMA plus A23187. Metabolism of β-oxa 21:3n-3 via the cyclooxygenase and lipoxygenase pathways was not required for its inhibitory effects. Consistent with its ability to suppress T lymphocyte function, β-oxa 21:3n-3 significantly inhibited the delayed-type hypersensitivity response and carrageenan-induced paw edema in mice. In T lymphocytes, β-oxa 21:3n-3 inhibited the agonist-stimulated translocation of protein kinase C-βI and -ε, but not -α, -βII, or -θ to a particulate fraction, and also inhibited the activation of the extracellular signal-regulated protein kinase, but not c-Jun NH2-terminal kinase and p38. In contrast, 22:6n-3 had no effects on these protein kinase C isozymes. The increase in antiinflammatory activity and loss of unwanted bioaction through the generation of a novel synthetic 22:6n-3 analogue provides evidence for a novel strategy in the development of anti-inflammatory agents by chemically engineering PUFA.