bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification of New HER2/neu-Derived Peptide Epitopes That Can Elicit Specific CTL Against Autologous and Allogeneic Carcinomas and Melanomas
Authors: Rongcun Yang , Salazar-Onfray Flavio , Charo Jehad , Malmberg Karl-Johan , Evrin Kristina , Maes Hubert , Kono Koji , Hising Christina , Petersson Max , Larsson Olle , Lan Li , Appella Ettore , Sette Alessandro , Celis3 Esteban , Kiessling Rolf
Published: December 2022

Twenty-two new HLA-A2.1-binding peptides derived from the protooncogene HER2/neu were identified and analyzed for their capacity to elicit peptide and tumor-specific CTL responses. We used peptide-pulsed autologous DC from the ascites of patients with ovarian carcinomas to induce CTL. Of the 22 tested new HER2/neu-derived epitopes that could bind HLA-A2 with high (IC50 < 50 nM) or intermediate (50 nM < IC50 < 500 nM) affinity, we report the recognition by CTL of at least four novel epitopes, including HER2(9435), HER2(9665), HER2(9689), and HER2(10952), and confirm that of the known HER2 (9369) epitope. These epitopes were able to elicit CTL that specifically killed peptide-sensitized target cells and, most importantly, a HER2/neu-transfected cell line and the autologous tumor cells. We also confirm that HER2/neu is overexpressed in several melanoma lines, and as a new finding, report that some of these lines are sensitive to CTL induced by the HER2 (9369), HER2(9435), and HER2(9689) epitopes. Finally, CTL clones specific for HER2 (9369), HER2(9435), and HER2(9689) epitopes were isolated from tumor-specific CTL lines, further demonstrating the immunodominance of these epitopes. These findings broaden the potential application of HER2/neu-based immunotherapy.
Differential Antitumor Effects of Administration of Recombinant IL-18 or Recombinant IL-12 Are Mediated Primarily by Fas-Fas Ligand- and Perforin-Induced Tumor Apoptosis, Respectively
Authors: Hashimoto Wataru , Osaki Tadashi , Okamura Haruki , Robbins Paul D. , Kurimoto Masashi , Nagata Shigekazu , Lotze Michael T. , Tahara Hideaki
Published: December 2022

Systemic administration of rIL-18 protein to mice significantly suppresses the growth of murine tumor cell lines. The antitumor effect of IL-18 appears to be primarily mediated by asialo GM1+ cells. Since IL-18 enhances Fas ligand (FasL) expression on NK cell lines, the IL-18 antitumor effects could be mediated by FasL-induced cross-linking of Fas and subsequent tumor apoptosis. To address this question, rIL-18 or rIL-12 was administered to animals bearing the CL8-1 melanoma inoculated intradermally into wild type (wt), lymphoproliferation gene (lpr) (Fas deficient), or generalized lymphoproliferative disease gene (gld) (FasL deficient) mice. Although rIL-12 treatment retained significant antitumor effects in gld and lpr mice, those of rIL-18 administration were completely abrogated in gld but not lpr or wt mice. In vitro cytotoxicity was significantly enhanced against NK-sensitive YAC-1 cells and CL8-1 cells by rIL-18 administration to wt mice, but not to gld mice. Furthermore, rIL-18 administration augmented the cytotoxicity of liver lymphocytes harvested from perforin-deficient mice, whereas rIL-12 administration did not. Consistent with the role of this pathway, rIL-18 administration also up-regulates the expression of FasL mRNA in splenocytes. Lysis of CL8-1 cells induced by anti-Fas agonistic Ab was enhanced about 1.4-fold by IFN-γ, a cytokine that is induced by IL-18 in vitro and in vivo. We conclude that the antitumor effect of IL-18 is exerted predominantly through a Fas-dependent pathway. The perforin pathway, however, appears to be the predominant cytolytic pathway mediating IL-12 antitumor effects.
Differential Involvement of the Transcription Factor Blimp-1 in T Cell-Independent and -Dependent B Cell Differentiation to Plasma Cells
Authors: Soro Pilar G. , Morales-A. Patricia , Martínez-M. Jose A. , Morales-A. Susana , Copín Sergio G. , Marcos Miguel A. R. , Gaspar María-Luisa
Published: December 2022

Along humoral immune responses, different stimuli drive the differentiation of B lymphocytes to Ig-secreting plasma cells in discrete microenvironments. The Blimp-1 transcription factor is up-regulated early during the transition of mature B cells to IgM-secreting plasma cells. In the present study, we have examined the requirement of Blimp-1 in plasma cell formation after both T cell-independent (LPS) and -dependent (CD40 + IL-4, Th cell lines) stimulation of spleen B cells. B lymphocyte-induced maturation protein (Blimp-1) was expressed early after in vitro LPS stimulation, mainly in a population of IgM+Syndecan+CD43+ preplasma cells. In contrast, the BSAP transcription factor expressed in mature B cells was down-regulated during the differentiation to plasma cells. Treatment of these cultures with Blimp-1-specific antisense phosphorothioate oligonucleotides suppressed both Blimp-1 protein levels and the emergence of IgM+Syndecan+ cells and plasma cells. However, T-B cell cocultures of spleen B cells from C3H/HeJ (H-2k) mice and syngeneic autoreactive SR.10 Th2 cells submitted to the anti-Blimp-1 therapy did not show any significant reduction in IgM- and IgG1-secreting plasma cell formation. Spleen B cells treated with anti-CD40 mAb + IL-4 differentiated to IgG1-secreting cells without significant transcription of the Blimp-1 gene; anti-Blimp-1 treatment subsequently did not have any effect in the later cultures. Altogether, these results suggest that Blimp-1 transcription factor specifically promotes T cell-independent B cell differentiation to plasma cells, probably at preplasma cell stages. In contrast, T cell-dependent plasma cell formation likely evolves through Blimp-1-independent pathways.
IgG-Mediated Enhancement of Antibody Responses Is Low in Fc Receptor γ Chain-Deficient Mice and Increased in FcγRII-Deficient Mice
Authors: Wernersson Sara , Karlsson Mikael C. I. , Dahlström Jörgen , Mattsson Ragnar , Verbeek J. Sjef , Heyman Birgitta
Published: December 2022

Immunization with IgG/Ag or IgE/Ag complexes leads to a higher production of specific Abs than immunization with Ag alone. The enhancing effect of IgE is exclusively dependent upon the low-affinity receptor for IgE, FcεRII, whereas the mechanism behind IgG-mediated enhancement is unknown. We have investigated whether receptors for the Fc part of IgG are required for responses to IgG/Ag. Mice lacking the γ subunit of Fc receptors (FcRs) (FcRγ−/−), FcγRII (FcγRII−/−), or FcγRIII (FcγRIII−/−) were immunized with BSA-2,4,6-trinitrophenyl (TNP) alone or BSA-TNP complexed to monoclonal TNP-specific IgG1, IgG2a, or IgG2b. As expected, all subclasses enhanced the Ab-response to BSA in wild-type mice. Enhancement was in the same order of magnitude in FcγRIII−/− mice (≤177-fold of controls administered Ag alone), whereas it was abrogated in FcRγ−/− mice and augmented in FcγRII−/− mice (≤5147-fold of controls). The response to IgE/Ag complexes in FcRγ−/− and FcγRII−/− mice was similar to that seen for wild-type mice, demonstrating that non-FcγR-dependent responses were normal. Our observations suggest that IgG/Ag complexes enhance Ab responses via FcγRs. Moreover, they reveal a strong negative regulation of Ab responses to IgG/Ag exerted by FcγRII.
Hypervariable Region 1 Variants Act as TCR Antagonists for Hepatitis C Virus-Specific CD4+ T Cells
Authors: Frasca Loredana , Del Porto Paola , Tuosto Loretta , Marinari Barbara , Scottà Cristiano , Carbonari Maurizio , Nicosia Alfredo , Piccolella Enza
Published: December 2022

In various human viral infections, the appearance of mutated epitopes displaying TCR antagonistic activity has been correlated with the severity and persistence of infection. In hepatitis C virus (HCV) infection, where the virus persistence has been associated with the rapid and substantial Ag modifications occurring during replication, TCR antagonism has been evidenced in CD8+ T cell responses. However, CD4+ T cell antagonism may be another important strategy by which HCV eludes a protective response, because sustained Th responses directed against several HCV Ags are associated with a self-limited course of infection. The data reported here represent the first evidence that variants of the hypervariable region (HVR1) of the putative Envelope 2 protein of HCV can act as powerful TCR antagonists for HVR1-specific CD4+ T cells isolated from HCV-infected individuals. Using classical antagonism assays, we observed strong inhibition of cellular proliferation and cytokine production when the agonist and the antagonist ligands were simultaneously presented by the same APCs. The presence in HVR1 of conserved residues, critical for binding to HLA-DR molecules, supports the function of HVR1 variants as TCR antagonists. In conclusion, our data evidence an antagonism phenomenon, which was achieved by naturally occurring class II-restricted T cell epitopes whose mechanism was addressed in terms of the antagonist capacity to inhibit agonist-mediated TCR down-regulation and early signal transduction.
Activation of the Extracellular Signal-Related Kinase/Mitogen-Activated Protein Kinase Pathway Discriminates CD4 Versus CD8 Lineage Commitment in the Thymus
Authors: Bommhardt Ursula , Basson M. Albert , Krummrei Ulrike , Zamoyska Rose
Published: December 2022

We have investigated the role of the mitogen-activated protein kinase (MAPK) pathway in the differentiation of CD4+ and CD8+ T cells by looking specifically at the effects of inhibitors of MAPK-activating enzyme, MAPK/extracellular signal-related kinase (ERK) kinase (MEK), during the positive selection step from double-positive to single-positive (SP) thymocytes. Using a variety of transgenic/knockout mouse strain combinations that fail to differentiate individual lineages of SP thymocytes together with genetically engineered F(ab′)2 reagents that induce maturation preferentially to either the CD4 or CD8 subpopulations, we show that induction of CD4 differentiation cells is highly sensitive to levels of MEK inhibition that have no effect on CD8 maturation. In addition, the presence of MEK inhibitor is able to modify signals that normally induce CD4 differentiation to instead promote CD8 differentiation. Finally, we show that continuous culture in the presence of inhibitor interferes with TCR up-regulation in SP thymocytes, suggesting that MAPK signaling may be involved in final maturation steps for both lineages. These data indicate that there is discrimination in the biochemical pathways that are necessary to specify CD4 and CD8 lineage commitment and can reconcile previously conflicting reports on the influence of MAPK activation in commitment and maturation of thymocytes.
Activation of the Janus Kinase 3-STAT5a Pathway After CD40 Triggering of Human Monocytes But Not of Resting B Cells
Authors: Revy Patrick , Hivroz Claire , Andreu Georges , Graber Pierre , Martinache Chantal , Fischer Alain , Durandy Anne
Published: December 2022

CD40/CD40 ligand interactions play a key role in the immune responses of B lymphocytes, monocytes, and dendritic cells. The signal transduction events triggered by cross-linking of the CD40 receptor have been widely studied in B cell lines, but little is known about signaling following CD40 stimulation of monocytes and resting tonsillar B cells. Therefore, we studied the CD40 pathway in highly purified human monocytes and resting B cells. After CD40 triggering, a similar activation of the NF-κB (but not of the AP-1) transcription factor complex occurred in both cell preparations. However, the components of the NF-κB complexes were different in monocytes and B cells, because p50 is part of the NF-κB complex induced by CD40 triggering in both monocytes and B cells, whereas p65 was only induced in B cells. In contrast, although the Janus kinase 3 tyrosine kinase was associated with CD40 molecules in both monocytes and resting B cells, Janus kinase 3 phosphorylation induction was observed only in CD40-activated monocytes, with subsequent induction of STAT5a DNA binding activity in the nucleus. These results suggest that the activation signals in human B cells and monocytes differ following CD40 stimulation. This observation is consistent with the detection of normal CD40-induced monocyte activation in patients with CD40 ligand+ hyper IgM syndrome in whom a defect in CD40-induced B cell activation has been reported.
Discoordinate Expression of Invariant Chain and MHC Class II Genes in Class II Transactivator-Transfected Fibroblasts Defective for RFX5
Authors: Peijnenburg Ad , Van Eggermond Marja J. C. A. , Gobin Sam J. P. , Van den Berg Rian , Godthelp Barbara C. , Vossen Jaak M. J. J. , Van den Elsen Peter J.
Published: December 2022

MHC class II deficiency or bare lymphocyte syndrome is a severe combined immunodeficiency caused by defects in MHC-specific transcription factors. In the present study, we show that fibroblasts derived from a recently identified bare lymphocyte syndrome patient, SSI, were mutated for RFX5, one of the DNA-binding components of the RFX complex. Despite the lack of functional RFX5 and resulting MHC class II-deficient phenotype, transfection of exogenous class II transactivator (CIITA) in these fibroblasts can overcome this defect, resulting in the expression of HLA-DR, but not of DP, DQ, and invariant chain. The lack of invariant chain expression correlated with lack of CIITA-mediated transactivation of the invariant chain promoter in transient transfection assays in SSI fibroblast cells. Consequently, these CIITA transfectants lacked Ag-presenting functions.
TCR and CD28 Are Coupled Via ZAP-70 to the Activation of the Vav/Rac-1-/PAK-1/p38 MAPK Signaling Pathway
Authors: Salojin Konstantin V. , Zhang Jian , Delovitch Terry L.
Published: December 2022

CD28 costimulation amplifies TCR-dependent signaling in activated T cells, however, the biochemical mechanism(s) by which this occurs is not precisely understood. The small GTPase Rac-1 controls the catalytic activity of the mitogen-activated protein kinases (MAPKs) and cell cycle progression through G1. Rac-1 activation requires the phospho-tyrosine (p-Tyr)-dependent recruitment of the Vav GDP releasing factor (GRF) to the plasma membrane and assembly of GTPase/GRF complexes, an event critical for Ag receptor-triggered T cell activation. Here, we show that TCR/CD28 costimulation synergistically induces Rac-1 GDP/GTP exchange. Our findings, obtained by using ZAP-70-negative Jurkat T cells, indicate that CD28 costimulation augments TCR-mediated T cell activation by increasing the ZAP-70-mediated Tyr phosphorylation of Vav. This event regulates the Rac-1-associated GTP/GDP exchange activity of Vav and downstream pathway(s) leading to PAK-1 and p38 MAPK activation. CD28 amplifies TCR-induced ZAP-70 activity and association of Vav with ZAP-70 and linker for activation of T cells (LAT). These results favor a model in which ZAP-70 regulates the intersection of the TCR and CD28 signaling pathways, which elicits the coupling of TCR and CD28 to the Rac-1, PAK-1, and p38 MAPK effector molecules.
The Antiviral Activity of HIV-Specific CD8+ CTL Clones Is Limited by Elimination Due to Encounter with HIV-Infected Targets
Authors: M. McKinney Denise , Lewinsohn Deborah A. , Riddell Stanley R. , Greenberg Philip D. , Mosier Donald E.
Published: December 2022

Adoptive immunotherapy of virus infection with viral-specific CTL has shown promise in animal models and human virus infections and is being evaluated as a therapy for established HIV-1 infection. Defining the individual obstacles for success is difficult in human trials. We have therefore examined the localization, persistence, and antiviral activity of HIV-1 gag-specific CTL clones in both HIV-1-infected and uninfected haplotype-matched human (hu)-PBL-SCID mice. Injection of gag-specific clones but not control CTL into HIV-1-infected hosts reduced plasma viremia by >10-fold but failed to eliminate the virus infection from most treated animals. The failure to eradicate virus did not reflect selection of escape variants because the gag epitope remained unmutated in virus isolates obtained after CTL therapy. Injection of carboxyfluorescein diacetate succinimide ester-labeled CTL demonstrated markedly different fates for gag-specific CTL in the presence or absence of HIV-1 infection. HIV-1-specific CTL rapidly disappeared in infected recipients, whereas they were maintained at high numbers in uninfected mice. By contrast, control CTL were long lived in both infected and uninfected recipients. Thus, interaction of CTL with virus-infected target cells in vivo leads not only to target destruction but also to the rapid disappearance of the infused CTL, and it limits the capacity of CTL therapy to eliminate HIV-1 infection.
Lytic Cycle T Cell Epitopes Are Expressed in Two Distinct Phases During MHV-68 Infection
Authors: Liu Luzheng , Flaño Emilio , Usherwood Edward J. , Surman Sherri , Blackman Marcia A. , Woodland David L.
Published: December 2022

Murine herpesvirus-68 (MHV-68) is a type 2 γ herpesvirus that productively infects alveolar epithelial cells during the acute infection and establishes long-term latency in B cells and lung epithelial cells. In C57BL/6 mice, T cells specific for lytic cycle MHV-68 epitope p56/Db dominate the acute phase of the infection, whereas T cells specific for another lytic cycle epitope, p79/Kb, dominate later phases of infection. To further understand this response, we analyzed the kinetics of Ag presentation in vivo using a panel of lacZ-inducible T cell hybridomas specific for several lytic cycle epitopes, including p56/Db and p79/Kb. Two distinct peaks of Ag presentation were observed. The first peak was at day 6 in the mediastinal lymph nodes and correlated with the initial pulmonary lytic infection. The second peak was at day 18 in both the mediastinal lymph nodes and spleen and correlated with the peak of latent infection. Interestingly, the p56 epitope was detected only in the mediastinal lymph nodes at day 6 after infection whereas the p79 epitope was predominantly presented in the spleen at day 18, suggesting that differential epitope presentation drives the switch in T cell responses to this virus. Phenotypic analysis of APCs at day 18 postinfection revealed that dendritic cells, macrophages, and B cells all presented lytic cycle epitopes. Taken together, the data suggest that there is a resurgence of lytic cycle Ags in the spleen, which explains the kinetics and specificity of the T cell response to MHV-68.
IL-12 and NK Cells Are Required for Antigen-Specific Adaptive Immunity Against Malaria Initiated by CD8+ T Cells in thePlasmodium yoeliiModel
Authors: Doolan Denise L. , Hoffman Stephen L.
Published: December 2022

CD8+ T cells have been implicated as critical effector cells in protection against preerythrocytic stage malaria, including the potent protective immunity of mice and humans induced by immunization with radiation-attenuated Plasmodium spp. sporozoites. This immunity is directed against the Plasmodium spp. parasite developing within the host hepatocyte and for a number of years has been presumed to be mediated directly by CD8+ CTL or indirectly by IFN-γ released from CD8+ T cells. In this paper, in BALB/c mice, we establish that after immunization with irradiated sporozoites or DNA vaccines parasite-specific CD8+ T cells trigger a novel mechanism of adaptive immunity that is dependent on T cell- and non-T cell-derived cytokines, in particular IFN-γ and IL-12, and requires NK cells but not CD4+ T cells. The absolute requirement for CD8+ T cells to initiate such an effector mechanism, and the requirement for IL-12 and NK cells in such vaccine-induced protective immunity, are unique and underscore the complexity of the immune responses that protect against malaria and other intracellular pathogens.
IL-2 Mediates Protection Against Abscess Formation in an Experimental Model of Sepsis
Authors: Tzianabos Arthur O. , Russell Pamela R. , Onderdonk Andrew B. , Gibson Frank C. , Cywes Colette , Chan Melvin , Finberg Robert W. , Kasper Dennis L.
Published: December 2022

Little is known regarding the mechanism by which T cells control intraabdominal abscess formation. Treating animals with polysaccharide A (PS A) from Bacteroides fragilis shortly before or after challenge protects against abscess formation subsequent to challenge with different abscess-inducing bacteria. Although bacterial polysaccharides are considered to be T cell-independent Ags, T cells from PS A-treated animals mediate this protective activity. In the present study, we demonstrate that CD4+ T cells transfer PS A-mediated protection against abscess formation, and that a soluble mediator produced by these cells confers this activity. Cytokine mRNA analysis showed that T cells from PS A-treated animals produced transcript for IL-2, IFN-γ, and IL-10, but not for IL-4. The addition of IL-2-specific Ab to T cell lysates taken from PS A-treated animals abrogated the ability to transfer protection, whereas the addition of Abs specific for IFN-γ and IL-10 did not affect protection. Finally, administration of rIL-2 to animals at the time of bacterial challenge prevented abscess formation in a dose-dependent manner. These data demonstrate that PS A-mediated protection against abscess formation is dependent upon a CD4+ T cell-dependent response, and that IL-2 is essential to this immune mechanism.
Neonatal Exposure to Idiotype InducesSchistosoma mansoniEgg Antigen-Specific Cellular and Humoral Immune Responses
Authors: Montesano M. Angela , Colley Daniel G. , Freeman George L. , Secor W. Evan
Published: December 2022

Exposure of neonatal mice to appropriate, cross-reactive Id (CRI) preparations alters immune responsiveness, ameliorates pathology, and prolongs survival of animals upon subsequent Schistosoma mansoni infection. However, because schistosome infections profoundly affect host immunobiology, which responses are effected by neonatal Id exposure alone and which responses are influenced by infection is unclear. To directly examine the schistosome soluble egg Ag (SEA)-specific immune responses altered by CRI exposure, neonatal mice were injected with CRI-expressing (CRI+) SEA-specific Ab preparations, SEA-specific Abs that did not express CRI (CRI−), or normal mouse Ig. At 9 wk of age, only mice that were neonatally exposed to CRI+ anti-SEA Abs displayed significant SEA-specific IgG serum levels and spleen cell proliferative responses. SEA-stimulated spleen cells from these CRI+-exposed mice also produced IFN-γ, although not at significantly higher levels than mice receiving CRI− Id or normal mouse Ig. If CRI+-exposed mice were also injected with SEA at 8 wk of age, the 9-wk IFN-γ responses were significantly higher than those of the other neonatal injection groups. The presence of both CRI and anti-CRI in the sera of animals neonatally injected with CRI, but receiving no exposure to S. mansoni Ags or infection, suggested a functional idiotypic network led to these responses. These data demonstrate that appropriate idiotypic exposure induces B and T cell responsiveness to the Ag recognized by the Id and support the hypothesis that neonatal idiotypic exposure can be an important immunoregulatory factor in schistosomiasis.
IFN-γ Mediates a Novel Antiviral Activity Through Dynamic Modulation of TRAIL and TRAIL Receptor Expression
Authors: Sedger Lisa M. , Shows Donna M. , Blanton Rebecca A. , Peschon Jacques J. , Goodwin Ray G. , Cosman David , Wiley Steven R.
Published: December 2022

TNF-related apoptosis-inducing ligand (TRAIL) is able to kill many transformed cells of diverse tissue types. We show that TRAIL is inducible by IFN-γ, by TNF-α, and by infection with human CMV, and has potent antiviral activity in vitro. CMV infection and IFN-γ also reciprocally modulate TRAIL receptor (TRAIL-R) expression. CMV infection increased the expression of TRAIL-R1 and -R2, whereas IFN-γ down-regulated the expression of TRAIL-Rs on uninfected fibroblasts. Moreover, IFN-γ significantly decreased the basal level of NF-κB activation, a known survival factor that inhibits apoptosis. Thus, TRAIL selectively kills virus-infected cells while leaving uninfected cells intact, and IFN-γ potentiates these effects by dynamic modulation of TRAIL and TRAIL-R expression and by sensitizing cells to apoptosis. The regulation of TRAIL and TRAIL-R expression may represent a general mechanism that contributes to the control of TRAIL-mediated apoptosis.
HIV Type 1 Nef Protein Is a Viral Factor for Leukocyte Recruitment into the Central Nervous System
Authors: Koedel Uwe , Kohleisen Birgit , Sporer Bernd , Lahrtz Fritz , Ovod Vladimir , Fontana Adriano , Erfle Volker , Pfister Hans-Walter
Published: December 2022

Recombinant HIV-1 Nef protein, but not Tat, gp120, and gp160, provoked leukocyte recruitment into the CNS in a rat model. The strong reduction of bioactivity by heat treatment of Nef, and the blocking effect of the mAb 2H12, which recognizes the carboxy-terminal amino acid (aa) residues 171–190 (but not of mAb 3E6, an anti-Nef Ab of the same isotype, which maps the aa sequence 168–175, as well as a mixture of mAbs to CD4) provided evidence for the specificity of the observed Nef effects. Using a modified Boyden chamber technique, Nef exhibited chemotactic activity on mononuclear cells in vitro. Coadministration of the anti-Nef mAb 2H12, as well as treatment of Nef by heat inhibited Nef-induced chemotaxis. Besides soluble Nef, chemotaxis was also induced by a Nef-expressing human astrocytoma cell line, but not by control cells. These data suggest a direct chemotactic activity of soluble Nef. The detection of elevated levels of IL-6, TNF-α, and IFN-γ in rat cerebrospinal fluid 6 h after intracisternal Nef injection hint at the additional involvement of indirect mechanisms in Nef-induced leukocyte migration into rat CNS. These data propose a mechanism by which HIV-1 Nef protein may be essential for AIDS neuropathogenesis, as a mediator of the recruitment of leukocytes that may serve as vehicles of the virus and perpetrators for disease through their production of neurotoxins.
Simultaneous Activation of T Cells and Accessory Cells by a New Class of Intact Bispecific Antibody Results in Efficient Tumor Cell Killing
Authors: Zeidler Reinhard , Reisbach Gilbert , Wollenberg Barbara , Lang Stephan , Chaubal Sarita , Schmitt Bärbel , Lindhofer Horst
Published: December 2022

Bispecific Abs (bsAb) are promising immunological tools for the elimination of tumor cells in minimal residual disease situations. In principle, they target an Ag on tumor cells and recruit one class of effector cell. Because immune reactions in vivo are more complex and are mediated by different classes of effector cell, we argue that conventional bsAb might not yield optimal immune responses at the tumor site. We therefore constructed a bsAb that combines the two potent effector subclasses mouse IgG2a and rat IgG2b. This bispecific molecule not only recruits T cells via its one binding arm, but simultaneously activates FcγR+ accessory cells via its Fc region. We demonstrate here that the activation of both T lymphocytes and accessory cells leads to production of immunomodulating cytokines like IL-1β, IL-2, IL-6, IL-12, and DC-CK1. Thus this new class of bsAb elicits excellent antitumor activity in vitro even without the addition of exogenous IL-2, and therefore represents a totally self-supporting system.
A Monoclonal Antibody Reactive with a 40-kDa Molecule on Fetal Thymocytes and Tumor Cells Blocks Proliferation and Stimulates Aggregation and Apoptosis
Authors: Fernandes Dancella M. , Baird Allison M. , Berg Leslie J. , Rock Kenneth L.
Published: December 2022

E710.2.3 is a murine thymic lymphoma cell line with an immature phenotype (CD4−CD8−) that proliferates in response to thymocytes or PMA when cultured at low density and proliferates spontaneously when grown at high density. To identify functional molecules on this cell line, we screened for mAbs that could block its proliferation. A hamster mAb, DMF10.62.3, inhibited the spontaneous, thymocyte-induced, and PMA-stimulated proliferation of E710.2.3 in vitro and induced these cells to undergo apoptosis. The mAb also caused homotypic aggregation of E710.2.3, which was inhibited by cytochalasin B, trifluoperazine, a combination of sodium azide and 2-deoxyglucose, EDTA, incubation at 4°C, or treatment with paraformaldehyde. The DMF10 62.3 mAb stained a number of immortalized murine and human cell lines and, where tested, blocked their proliferation and caused death to varying extents by apoptosis. The molecule recognized by the mAb DMF10.62.3 was expressed on day 14 fetal thymus Thy1.2-positive cells. However, it was not detected on adult murine thymocytes, splenocytes, or bone marrow cells or on splenic LPS-activated B cells or Con A-activated T cells. The Ab immunoprecipitated a 40-kDa molecule from E710.2.3 that was not glycosylphosphatidylinositol linked. The data suggest that the molecule recognized by DMF62.3 is a novel cell surface molecule that may be involved in cell proliferation and/or cell death.
Multiple Deficiencies Underlie NK Cell Inactivity in Lymphotoxin-α Gene-Targeted Mice
Authors: Smyth Mark J. , Johnstone Ricky W. , Cretney Erika , Haynes Nicole M. , Sedgwick Jonathon D. , Korner Heiner , Poulton Lynn D. , Baxter Alan G.
Published: December 2022

We have evaluated the NK cell antitumor activity in lymphotoxin (LT)-deficient mice. Both NK cell-mediated tumor rejection and protection from experimental metastases were significantly compromised in LT-α-deficient mice. Analysis of LT-α-deficient mice revealed that the absolute number of αβTCR− NK1.1+ NK cells was reduced in bone marrow and thymus, but with overall proportional decreases in other hemopoietic organs. In addition, the antitumor potential of αβTCR− NK1.1+ cells, as determined by their lytic capacity and perforin expression, was reduced 1.5- to 3-fold in LT-α-deficient mice, as compared with wild-type mice. Combined defects in NK cell development and effector function contribute to compromised NK cell antitumor function in LT-α-deficient mice.
Inhibition of Autoimmune T Cell Responses in the DA Rat by Bone Marrow-Derived NK Cells In Vitro: Implications for Autoimmunity
Authors: Smeltz Ronald B. , Wolf Norbert A. , Swanborg Robert H.
Published: December 2022

Regulation of the immune response is critical to homeostasis. While innate immunity can influence the development of adaptive immune responses, its role in regulation is less well understood. Recently, NK cells have been implicated in the control of experimental autoimmune encephalomyelitis, an animal model for multiple sclerosis. In this report, we show that rat bone marrow-derived NK cells exhibited potent inhibitory effects on T cell proliferation to both Con A as well as the central nervous system Ag myelin basic protein. There was also a significant decrease in both IFN-γ and IL-10 production in vitro, whereas levels of the β-chemokine monocyte chemoattractant protein-1 were significantly elevated. Flow cytometry studies suggest that the NK cells may play an important role in regulating both normal and autoimmune T cell responses by exerting a direct effect on activated, autoantigen-specific T cells.
Transactivation of Classical and Nonclassical HLA Class I Genes Through the IFN-Stimulated Response Element
Authors: Gobin Sam J. P. , Zutphen Marlijn van , Woltman Andrea M. , Elsen Peter J. van den
Published: December 2022

The IFN-stimulated response element (ISRE) is an important conserved cis-acting regulatory element in the promoter of MHC class I genes, but displays considerable locus-specific nucleotide variation. In this report, the putative ISREs of classical and nonclassical HLA class I genes were investigated for their contribution to MHC class I transactivation. It is shown that IFN-γ induced MHC class I transactivation through the ISRE of HLA-A, HLA-B, HLA-C, and HLA-F. This is congruent with the binding of IFN regulatory factor-1 to the ISREs of these loci upon IFN-γ treatment. Sp1 was shown to bind to the CG-rich sequences in the ISRE regions of HLA-B, HLA-C, and HLA-G. The putative E box 5′ of the ISRE in most HLA-B alleles was shown to bind the upstream stimulatory factors (USF) 1 and 2. The Sp1 and USF binding sites did not influence IFN-γ-induced transactivation. However, the USF binding site played a suppressive role in the constitutive expression of HLA-B. The locus-specific transcriptional control through the ISRE could be an important mechanism in the differential regulation of classical and nonclassical MHC class I expression, which determines adequate Ag presentation upon pathogenic challenge.
NF-κB Is a Central Regulator of the Intestinal Epithelial Cell Innate Immune Response Induced by Infection with Enteroinvasive Bacteria
Authors: Elewaut Dirk , DiDonato Joseph A. , Mogg Kim Jung , Truong Francis , Eckmann Lars , Kagnoff Martin F.
Published: December 2022

Human intestinal epithelial cells up-regulate the expression of an inflammatory gene program in response to infection with a spectrum of different strains of enteroinvasive bacteria. The conserved nature of this program suggested that diverse signals, which are activated by enteroinvasive bacteria, can be integrated into a common signaling pathway that activates a set of proinflammatory genes in infected host cells. Human intestinal epithelial cell lines, HT-29, Caco-2, and T84, were infected with invasive bacteria that use different strategies to induce their uptake and have different intracellular localizations (i.e., Salmonella dublin, enteroinvasive Escherichia coli, or Yersinia enterocolitica). Infection with each of these bacteria resulted in the activation of TNF receptor associated factors, two recently described serine kinases, IκB kinase (IKK) α and IKKβ, and increased NF-κB DNA binding activity. This was paralleled by partial degradation of IκBα and IκBε in bacteria-infected Caco-2 cells. Mutant proteins that act as superrepressors of IKKβ and IκBα inhibited the up-regulated transcription and expression of downstream targets genes of NF-κB that are key components of the epithelial inflammatory gene program (i.e., IL-8, growth-related oncogene-α, monocyte chemoattractant protein-1, TNF-α, cyclooxygenase-2, nitric oxide synthase-2, ICAM-1) activated by those enteroinvasive bacteria. These studies position NF-κB as a central regulator of the epithelial cell innate immune response to infection with enteroinvasive bacteria.
Glucocorticosteroids Inhibit mRNA Expression for Eotaxin, Eotaxin-2, and Monocyte-Chemotactic Protein-4 in Human Airway Inflammation with Eosinophilia
Authors: Jahnsen Frode L. , Haye Rolf , Gran Einar , Brandtzaeg Per , Johansen Finn-Eirik
Published: December 2022

How eosinophils are preferentially recruited to inflammatory sites remains elusive, but increasing evidence suggests that chemokines that bind to the CCR3 participate in this process. In this study, we investigated the transcript levels and chemotactic activity of CCR3-binding chemokines in nasal polyps, a disorder often showing prominent eosinophilia. We found that mRNA expression for eotaxin, eotaxin-2, and monocyte-chemotactic protein-4 was significantly increased in nasal polyps compared with turbinate mucosa from the same patients, or histologically normal nasal mucosa from control subjects. Interestingly, the novel CCR3-specific chemokine, eotaxin-2, showed the highest transcript levels. Consistent with these mRNA data, polyp tissue fluid exhibited strong chemotactic activity for eosinophils that was significantly inhibited by a blocking Ab against CCR3. When patients were treated systemically with glucocorticosteroids, the mRNA levels in the polyps were reduced to that found in turbinate mucosa for all chemokines. Together, these findings suggested an important role for CCR3-binding chemokines in eosinophil recruitment to nasal polyps. Such chemokines, therefore, most likely contribute significantly in the pathogenesis of eosinophil-related disorders; and the reduced chemokine expression observed after steroid treatment might reflect, at least in part, how steroids inhibit tissue accumulation of eosinophils.
Characterization of Fractalkine in Rat Brain Cells: Migratory and Activation Signals for CX3CR-1-Expressing Microglia
Authors: Maciejewski-Lenoir Dominique , Chen Shizhong , Feng Lilli , Maki Richard , Bacon Kevin B.
Published: December 2022

Molecular analyses of the chemokine fractalkine and its receptor CX3C-R1 in the rat brain have revealed a striking polarization: fractalkine is expressed constitutively in neurons and is up-regulated by TNF-α and IL-1β in astrocytes. Expression of its specific receptor, CX3C-R1, is restricted to astrocytes and microglia. We have analyzed the functional correlates of this expression and demonstrate that fractalkine induces microglial cell migration and activation. However, the activity of this chemokine on astrocytes may also be highly relevant in inducing astrocyte-microglia cell interactions through cytokine/mediator release leading to microglial activation.
Identification of T Cell Determinants on Human Type II Collagen Recognized by HLA-DQ8 and HLA-DQ6 Transgenic Mice
Authors: Krco Christopher J. , Watanabe Shohei , Harders Jerry , Griffths Marie M. , Luthra Harvinder , David Chella S.
Published: December 2022

HLA-DQA1*0301 and HLA-DQB1*0302 genes encoding the HLA-DQ8 molecule and HLA-DQA1*0103 and HLA-DQB1*0601 genes encoding the HLA-DQ6 molecule were introduced into H-2Aβo knockout mice. Three lines of transgenic mice were established: HLA-DQ8, HLA-DQ6, and HLA-DQ8β6α. HLA-DQ8 mice are susceptible to collagen-induced arthritis, while HLA-DQ6 mice are resistant. HLA-DQ8β6α mice develop polychrondritis in addition to arthritis. Transgenic mice were primed and challenged with individual synthetic peptides representing human type II collagen. A total of 101 synthetic peptides were tested in each transgenic line of mice. HLA-DQ8 mice responded to 15 synthetic peptides representing all cyanogen bromide fragments. In contrast, HLA-DQ6 mice responded to a subset of the peptides recognized by HLA-DQ8 T cells. HLA-DQ8β6α mice, although exhibiting diminished responses to the majority of HLA-DQ8-restricted determinants, elicited enhanced responses to two peptides. In addition, HLA-DQ8β6α mice respond to two unique peptide determinants contained within cyanogen bromide fragments CB10 and CB11 showing the significance of mixed isotype dimers in the immune response. The determinants recognized by the HLA-DQ transgenic mice are distinct from those previously identified using conventional laboratory mice. These results suggest that human class II transgenic mice offer a means of identifying human class II-restricted epitopes associated with potential human autoantigens.
Differential Presentation of Glutamic Acid Decarboxylase 65 (GAD65) T Cell Epitopes Among HLA-DRB1*0401-Positive Individuals
Authors: Reijonen Helena , Elliott John F. , van Endert Peter , Nepom Gerald
Published: December 2022

Glutamic acid decarboxylase 65 (GAD65) is one of the major autoantigens in type 1 diabetes. We investigated whether there is variation in the processing of GAD65 epitopes between individuals with similar HLA backgrounds and whether the processing characteristics of certain immunogenic epitopes are different in distinct APC subpopulations. Using DR401-restricted T cell hybridomas specific for two immunogenic GAD65 epitopes (115–127 and 274–286), we demonstrate an epitope-specific presentation pattern in human B-lymphoblastoid cell lines (B-LCL). When pulsed with the GAD protein, some DRB1*0401-positive B-LCL, which presented GAD65 274–286 epitope efficiently, were unable to present the GAD65 115–127 epitope. However, all B-LCL presented synthetic peptides corresponding to either GAD epitope. In addition, when pulsed with human serum albumin, all cell lines gave equal stimulation of a DR4-restricted human serum albumin-specific T hybridoma. GAD65-transfected cell lines displayed the same presentation phenotype, showing that lack of the presentation of the 115–127 epitope was not due to inefficient uptake of the protein. Blood mononuclear adherent cells, B cells, or dendritic cells derived from the same individual displayed the same presentation pattern as observed in B cell lines, suggesting that the defect most likely is genetically determined. Therefore, individual differences in Ag processing may result in the presentation of distinct set of peptides derived from an autoantigen such as GAD65. This may be an important mechanism for the deviation of the immune response either into a regulatory pathway or into an inflammatory autoimmune reactivity.
Impact of Cytokine Administration on the Generation of Antitumor Reactivity in Patients with Metastatic Melanoma Receiving a Peptide Vaccine
Authors: Rosenberg Steven A. , Yang James C. , Schwartzentruber Douglas J. , Hwu Patrick , Marincola Francesco M. , Topalian Suzanne L. , Restifo Nicholas P. , Sznol Mario , Schwarz Susan L. , Spiess Paul J. , Wunderlich John R. , Seipp Claudia A. , Einhorn Jan H. , Rogers-Freezer Linda , White Donald E.
Published: December 2022

Patients with metastatic melanoma were immunized with an immunodominant peptide derived from the gp100 melanoma-melanocyte differentiation Ag that was modified to increase binding to HLA-A+0201. A total of 10 of 11 patients who received the g209–2M peptide alone developed precursors reactive with the native g209 peptide, compared with only 5 of 16 patients who received g209–2M peptide plus IL-2 (p2 = 0.005). Peptide reactivity closely correlated with the recognition of HLA-A+0201 melanoma cells (p < 0.001). The decrease in immune reactivity when peptide was administered with IL-2 appeared specific for the immunizing peptide, since reactivity to an influenza peptide resulting from prior exposure was not affected. Preexisting antitumor precursors did not decrease when peptide plus IL-2 was administered. The administration of GM-CSF or IL-12 also resulted in a decrease in circulating precursors compared with the administration of peptide alone, though not as great a decrease as that seen with IL-2. Immunization with peptide plus IL-2 did, however, appear to have clinical impact since 6 of the 16 patients (38%) that received peptide plus IL-2 had objective cancer regressions. It thus appeared possible that immunization with peptide plus IL-2 resulted in sequestering or apoptotic destruction of newly activated immune cells at the tumor site. These represent the first detailed studies of the impact of immunization with tumor peptides in conjunction with a variety of cytokines in patients with metastatic cancer.
Blockade of CD28 During In Vitro Activation of Encephalitogenic T Cells or After Disease Onset Ameliorates Experimental Autoimmune Encephalomyelitis
Authors: Perrin Peter J. , June Carl H. , Maldonado Jairo H. , Ratts Robert B. , Racke Michael K.
Published: December 2022

Previous studies have shown complex roles for the B7 receptors in providing both positive and negative regulation of experimental autoimmune encephalomyelitis (EAE). B7 blockade can ameliorate clinical EAE by indirectly interfering with CD28 signaling. However, B7 blockade can also result in disease exacerbation, presumably by interfering with regulatory B7:CTLA-4 interactions. Therefore, we have directly targeted T cell CD28 with specific mAbs both during initial Ag priming and after the onset of clinical signs of EAE. We found that CD28 blockade ameliorated EAE during the efferent and afferent limbs of the immune response. Disease amelioration at disease onset was associated with suppression of TNF-α production. Finally, Ab blockade of T cell CD28 during the first disease episode resulted in significant attenuation of the subsequent disease course, with no significant relapses. In contrast to previous studies targeting APC B7 with CTLA4-Ig, reagents targeting CD28 can block ongoing disease. Therefore, the present results suggest a clinically relevant therapeutic scenario for human diseases, such as multiple sclerosis.
Cutting Edge: Inflammatory Signaling byBorrelia burgdorferiLipoproteins Is Mediated by Toll-Like Receptor 2
Authors: Hirschfeld Matthew , Kirschning Carsten J. , Schwandner Ralf , Wesche Holger , Weis John H. , Wooten R. Mark , Weis Janis J.
Published: December 2022

The agent of Lyme disease, Borrelia burgdorferi, produces membrane lipoproteins possessing potent inflammatory properties linked to disease pathology. The recent association of toll-like receptors (TLR) 2 and 4 with LPS responses prompted the examination of TLR involvement in lipoprotein signaling. The ability of human cell lines to respond to lipoproteins was correlated with the expression of TLR2. Transfection of TLR2 into cell lines conferred responsiveness to lipoproteins, lipopeptides, and sonicated B. burgdorferi, as measured by nuclear translocation of NF-κB and cytokine production. The physiological importance of this interaction was demonstrated by the 10-fold greater sensitivity of TLR2-transfected cells to lipoproteins than LPS. Futhermore, TLR2-dependant signaling by lipoproteins was facilitated by CD14. These data indicate that TLR2 facilitates the inflammatory events associated with Lyme arthritis. In addition, the widespread expression of lipoproteins by other bacterial species suggests that this interaction may have broad implications in microbial inflammation and pathogenesis.
Reduced IL-4-, Lipopolysaccharide-, and IFN-γ-Induced MHC Class II Expression in Mice Lacking Class II Transactivator Due to Targeted Deletion of the GTP-Binding Domain
Authors: Itoh-Lindstrom Yoshie , Piskurich Janet F. , Felix Nathan J. , Wang Ying , Brickey W. June , Platt Jeffrey L. , Koller Beverly H. , Ting Jenny P.-Y.
Published: December 2022

Class II transactivator (CIITA) is an unusual transcriptional coactivator in that it contains a functionally important, GTP-binding consensus domain. To assess the functional role of the GTP-binding domain of CIITA in vivo, we have generated knockout mice that bear a mutation in the CIITA gene spanning the GTP-binding domain. Upon analysis, these mice show no detectable CIITA mRNA; hence, they represent mice with deleted CIITA rather than mice with defects in the GTP-binding domain only. In these knockout mice, MHC class II expression is nearly eliminated, although a faint RT-PCR signal is visible in spleen, lymph node, and thymus, suggestive of the presence of CIITA-independent regulation of MHC class II expression. Invariant chain expression is also greatly reduced, but to a lesser extent than MHC class II. Serum IgM is not decreased, but the serum IgG level is greatly reduced, further confirming the absence of MHC class II Ag-dependent Ig class switching. Induction of MHC class II expression by IL-4 or LPS was absent on B cells, and Mac-1+ cells showed no detectable induction of MHC class II by either IL-4, LPS, or IFN-γ. These findings demonstrate a requirement for CIITA in IFN-γ-, IL-4-, and endotoxin-induced MHC class II expression as well as the possibility of rare CIITA-independent MHC class II expression.