PubMed Health⌕ Search

Biomedical subjects

Ralph M Steinman

Publications and source records attributed to Ralph M Steinman.

At least 19 recordsLinked to original sources
PubMed2007

DEC-205 receptor on dendritic cells mediates presentation of HIV gag protein to CD8+ T cells in a spectrum of human MHC I haplotypes.

Optimal HIV vaccines should elicit CD8+ T cells specific for HIV proteins presented on MHC class I products, because these T cells contribute to host resistance to viruses. We had previously found that the targeting of antigen to dendritic cells (DCs) in mice efficiently induces CD8+ T cell responses. To extend this finding to humans, we introduced the HIV p24 gag protein into a mAb that targets DEC-205/CD205, an endocytic receptor of DCs. We then assessed cross-presentation, which is the processing of nonreplicating internalized antigen onto MHC class I for recognition by CD8+ T cells. Low doses of alphaDEC-gag, but not control Ig-gag, stimulated proliferation and IFN-gamma production by CD8+ T cells isolated from the blood of HIV-infected donors. alphaCD205 fusion mAb was more effective for cross-presentation than alphaCD209/DC-SIGN, another abundant DC uptake receptor. Presentation was diverse, because we identified eight different gag peptides that were recognized via DEC-205 in 11 individuals studied consecutively. Our results, based on humans with highly polymorphic MHC products, reveal that DCs and DEC-205 can cross-present several different peptides from a single protein. Because of the consistency in eliciting CD8+ T cell responses, these data support the testing of alphaDEC-205 fusion mAb as a protein-based vaccine.

Amino Acid Sequence↗
PubMed2007

Dendritic cell-expanded, islet-specific CD4+ CD25+ CD62L+ regulatory T cells restore normoglycemia in diabetic NOD mice.

Most treatments that prevent autoimmune diabetes in nonobese diabetic (NOD) mice require intervention at early pathogenic stages, when insulitis is first developing. We tested whether dendritic cell (DC)-expanded, islet antigen-specific CD4+ CD25+ suppressor T cells could treat diabetes at later stages of disease, when most of the insulin-producing islet beta cells had been destroyed by infiltrating lymphocytes. CD4+ CD25+ CD62L+ regulatory T cells (T reg cells) from BDC2.5 T cell receptor transgenic mice were expanded with antigen-pulsed DCs and IL-2, and were then injected into NOD mice. A single dose of as few as 5x10(4) of these islet-specific T reg cells blocked diabetes development in prediabetic 13-wk-old NOD mice. The T reg cells also induced long-lasting reversal of hyperglycemia in 50% of mice in which overt diabetes had developed. Successfully treated diabetic mice had similar responses to glucose challenge compared with nondiabetic NOD mice. The successfully treated mice retained diabetogenic T cells, but also had substantially increased Foxp3+ cells in draining pancreatic lymph nodes. However, these Foxp3+ cells were derived from the recipient mice and not the injected T reg cells, suggesting a role for endogenous T reg cells in maintaining tolerance after treatment. Therefore, inoculation of DC-expanded, antigen-specific suppressor T cells has considerable efficacy in ameliorating ongoing diabetes in NOD mice.

Animals↗
PubMed2007

Differential antigen processing by dendritic cell subsets in vivo.

Dendritic cells (DCs) process and present self and foreign antigens to induce tolerance or immunity. In vitro models suggest that induction of immunity is controlled by regulating the presentation of antigen, but little is known about how DCs control antigen presentation in vivo. To examine antigen processing and presentation in vivo, we specifically targeted antigens to two major subsets of DCs by using chimeric monoclonal antibodies. Unlike CD8+ DCs that express the cell surface protein CD205, CD8- DCs, which are positive for the 33D1 antigen, are specialized for presentation on major histocompatibility complex (MHC) class II. This difference in antigen processing is intrinsic to the DC subsets and is associated with increased expression of proteins involved in MHC processing.

Amino Acid Sequence↗
PubMed2007

A single vaccination with attenuated SIVmac 239 via the tonsillar route confers partial protection against challenge with SIVmac 251 at a distant mucosal site, the rectum.

Elucidating the mechanisms that protect monkeys previously immunized with attenuated SIV (SIVDeltanef) against challenge infection with pathogenic virus may reveal new strategies for the development of an effective HIV vaccine. Here we show that a single atraumatic application of SIVDeltanef to the tonsils of four rhesus macaques conferred protection against SIVmac251 applied intrarectally 26 weeks later. While this protection was not complete, i.e., challenge virus could be isolated from all immunized animals, it was reflected by significantly lower viral loads in the blood (weeks 2-16 after challenge, p < 0.01) and considerably lower loads in lymphoid organs, and more stable peripheral CD4 counts in a proportion of the immunized animals as compared to four non-immunized, SIVmac251-infected control monkeys. SIV-specific humoral as well as systemic and mucosal T cell responses were detected in the immunized animals, but there was no correlation between their magnitude of expression and the level of protection. Analyses of leukocyte subsets in these animals at necropsy (24 weeks after challenge) did not reveal a significantly enhanced proportion of gamma/delta T cells in the tissues of protected monkeys. Therefore, tonsillar application of attenuated SIV induces protection in some animals against a superinfection with wild-type SIV distant at a distant mucosal site.

Animals↗
PubMed2006

Dendritic cell targeting of survivin protein in a xenogeneic form elicits strong CD4+ T cell immunity to mouse survivin.

To determine whether strong CD4+ T cell immunity could be induced to a nonmutated self protein that is important for tumorigenesis, we selectively targeted the xenogeneic form of survivin, a survival protein overexpressed in tumors, to maturing dendritic cells in lymphoid tissues. Dendritic cell targeting via the DEC205 receptor in the presence of anti-CD40 and poly(I:C) as maturation stimuli, induced strong human and mouse survivin-specific CD4+ T cell responses, as determined by IFN-gamma, TNF-alpha, and IL-2 production, as well as the development of lytic MHC class II-restricted T cells and memory. Immunity was enhanced further by depletion of CD25+foxp3+ cells before vaccination. anti-DEC205-human survivin was superior in inducing CD4+ T cell responses relative to other approaches involving survivin plasmid DNA or survivin peptides with adjuvants. However, we were unable to induce CD8+ T cell immunity to survivin by two doses of DEC205-targeted survivin or the other strategies. Therefore, significant CD4+ T cell immunity to a self protein that is overexpressed in most human cancers can be induced by DEC205 targeting of the Ag in its xenogeneic form to maturing DCs.

Animals↗
PubMed2006

APOBEC3G/3F mediates intrinsic resistance of monocyte-derived dendritic cells to HIV-1 infection.

HIV-1 infects immature dendritic cells (iDCs), but infection is inefficient compared with activated CD4+ T cells and only involves a small subset of iDCs. We analyzed whether this could be attributed to specific cellular restrictions during the viral life cycle. To study env-independent restriction to HIV-1 infection, we used a single-round infection assay with HIV-1 pseudotyped with vesicular stomatitis virus G protein (HIV-VSVG). Small interfering RNA-mediated depletion of APOBEC3G/3F (A3G/3F), but not TRIM5alpha, enhanced HIV-1 infection of iDCs, indicating that A3G/3F controls the sensitivity of iDCs to HIV-1 infection. Furthermore, sequences of HIV reverse transcripts revealed G-to-A hypermutation of HIV genomes during iDC infection, demonstrating A3G/3F cytidine deaminase activity in iDCs. When we separated the fraction of iDCs that was susceptible to HIV, we found the cells to be deficient in A3G messenger RNA and protein. We also noted that during DC maturation, which further reduces susceptibility to infection, A3G levels increased. These findings highlight a role for A3G/3F in explaining the resistance of most DCs to HIV-1 infection, as well as the susceptibility of a fraction of iDCs. An increase in the A3G/3F-mediated intrinsic resistance of iDCs could result in a block of HIV infection at its mucosal point of entry.

APOBEC-3G Deaminase↗
PubMed2006

Glycolipid alpha-C-galactosylceramide is a distinct inducer of dendritic cell function during innate and adaptive immune responses of mice.

alpha-Galactosylceramide (alpha-GalCer) is the prototype compound for studying the presentation of glycolipids on CD1d molecules to natural killer T (NKT) lymphocytes. A single i.v. dose of glycolipid triggers a cascade of events involving the production of several cytokines over the course of a day, a short-lived activation of NKT and natural killer (NK) cells, and a more prolonged adaptive T cell immune response if certain antigens are given together with alpha-GalCer. We find that a recently described analogue, alpha-C-galactosylceramide (alpha-C-GalCer), more potently induces these innate and adaptive immune responses in mice. alpha-C-GalCer acts as a more effective trigger for IL-12 and IFN-gamma production, although it minimally elicits IL-4 and TNF-alpha release into the serum. Also, alpha-C-GalCer better mobilizes NKT and natural killer cells to resist B16 melanoma. To help understand these effects, we find that alpha-C-GalCer binds more stably to dendritic cells than alpha-GalCer and that dendritic cells loaded with alpha-C-GalCer induce larger and more long lasting NKT cell responses in vivo. When glycolipid is targeted to dendritic cells in spleen together with antigens in dying cells, such as irradiated tumor cells, alpha-C-GalCer is active as an adjuvant for T cell-mediated immunity at lower doses, just 20 ng per mouse, where it is also able to up-regulate the required CD40L costimulatory molecule on NKT cells. Therefore, alpha-C-GalCer represents a glycolipid that binds more stably to dendritic cells and acts as a more effective link between innate and adaptive immunity in vivo.

Animals↗
PubMed2006

Expansion of FOXP3high regulatory T cells by human dendritic cells (DCs) in vitro and after injection of cytokine-matured DCs in myeloma patients.

CD4(+)CD25(+)FOXP3(+) regulatory T cells (Treg's) play an important role in the maintenance of immune tolerance. The mechanisms controlling the induction and maintenance of Treg's in humans need to be defined. We find that human myeloid dendritic cells (DCs) are superior to other antigen presenting cells for the maintenance of FOXP3(+) Treg's in culture. Coculture of DCs with autologous T cells leads to an increase in both the number of Treg's, as well as the expression of FOXP3 protein per cell both in healthy donors and myeloma patients. DC-mediated expansion of FOXP3(high) Treg's is enhanced by endogenous but not exogenous interleukin-2 (IL-2), and DC-T-cell contact, including the CD80/CD86 membrane costimulatory molecules. DCs also stimulate the formation of Treg's from CD25(-) T cells. The efficacy of induction of Treg's by DCs depends on the nature of the DC maturation stimulus, with inflammatory cytokine-treated DCs (Cyt-DCs) being the most effective Treg inducers. DC-induced Treg's from both healthy donors and patients with myeloma are functional and effectively suppress T-cell responses. A single injection of cytokine-matured DCs led to rapid enhancement of FOXP3(+) Treg's in vivo in 3 of 3 myeloma patients. These data reveal a role for DCs in increasing the number of functional FOXP3(high) Treg's in humans.

Adoptive Transfer↗
PubMed2006

Immunogenicity and efficacy of immunodeficiency virus-like particles pseudotyped with the G protein of vesicular stomatitis virus.

Vaccination with exogenous antigens such as recombinant viral proteins, immunodeficiency virus-derived whole inactivated virus particles, or virus-like particles (VLP) has generally failed to provide sufficient protection in animal models for AIDS. Pseudotyping VLPs with the vesicular stomatitis virus G protein (VSV-G), which is known to mediate entry into dendritic cells, might allow more efficient stimulation of immune responses. Therefore, we pseudotyped noninfectious immunodeficiency virus-like particles with VSV-G and carried out a preliminary screen of their immunogenicity and vaccination efficacy. Incorporation of VSV-G into HIV-1 VLPs led to hundred-fold higher antibody titers to HIV-1 Gag and enhancement of T cell responses in mice. Repeated vaccination of rhesus monkeys for 65 weeks with VSV-G pseudotyped simian immunodeficiency virus (SIV)-like particles (VLP[G]) provided initial evidence for efficient suppression of viral load after mucosal challenge with the SIVmac239 virus. Challenge of monkeys after a 28 week vaccination regimen with VLP[G] led to a reduction in peak viremia, but persistent suppression of viral load was not achieved. Due to limitations in the number of animals available for this study, improved efficacy of VSV-G pseudotyped VLPs in nonhuman primates could not be demonstrated. However, mouse experiments revealed that pseudotyping of VLPs with fusion-competent VSV-G clearly improves their immunogenicity. Additional strategies, particularly adjuvants, should be considered to provide greater protection against a challenge with pathogenic immunodeficiency virus.

AIDS Vaccines↗
PubMed2006

A dominant complement fixation pathway for pneumococcal polysaccharides initiated by SIGN-R1 interacting with C1q.

The intricate system of serum complement proteins provides resistance to infection. A pivotal step in the complement pathway is the assembly of a C3 convertase, which digests the C3 complement component to form microbial binding C3 fragments recognized by leukocytes. The spleen and C3 provide resistance against blood-borne S. pneumoniae infection. To better understand the mechanisms involved, we studied SIGN-R1, a lectin that captures microbial polysaccharides in spleen. Surprisingly, conditional SIGN-R1 knockout mice developed deficits in C3 catabolism when given S. pneumoniae or its capsular polysaccharide intravenously. There were marked reductions in proteolysis of serum C3, deposition of C3 on organisms within SIGN-R1(+) spleen macrophages, and formation of C3 ligands. We found that SIGN-R1 directly bound the complement C1 subcomponent, C1q, and assembled a C3 convertase, but without the traditional requirement for either antibody or factor B. The transmembrane lectin SIGN-R1 therefore contributes to innate resistance by an unusual C3 activation pathway.

Animals↗
PubMed2006

Antigen targeting to dendritic cells elicits long-lived T cell help for antibody responses.

Resistance to several prevalent infectious diseases requires both cellular and humoral immune responses. T cell immunity is initiated by mature dendritic cells (DCs) in lymphoid organs, whereas humoral responses to most antigens require further collaboration between primed, antigen-specific helper T cells and naive or memory B cells. To determine whether antigens delivered to DCs in lymphoid organs induce T cell help for antibody responses, we targeted a carrier protein, ovalbumin (OVA), to DCs in the presence of a maturation stimulus and assayed for antibodies to a hapten, (4-hydroxy-3-nitrophenyl) acetyl (NP), after boosting with OVA-NP. A single DC-targeted immunization elicited long-lived T cell helper responses to the carrier protein, leading to large numbers of antibody-secreting cells and high titers of high-affinity antihapten immunoglobulin Gs. Small doses of DC-targeted OVA induced higher titers and a broader spectrum of anti-NP antibody isotypes than large doses of OVA in alum adjuvant. Similar results were obtained when the circumsporozoite protein of Plasmodium yoelii was delivered to DCs. We conclude that antigen targeting to DCs combined with a maturation stimulus produces broad-based and long-lived T cell help for humoral immune responses.

Adjuvants, Immunologic↗
PubMed2006

Intensified and protective CD4+ T cell immunity in mice with anti-dendritic cell HIV gag fusion antibody vaccine.

Current human immunodeficiency virus (HIV) vaccine approaches emphasize prime boost strategies comprising multiple doses of DNA vaccine and recombinant viral vectors. We are developing a protein-based approach that directly harnesses principles for generating T cell immunity. Vaccine is delivered to maturing dendritic cells in lymphoid tissue by engineering protein antigen into an antibody to DEC-205, a receptor for antigen presentation. Here we characterize the CD4+ T cell immune response to HIV gag and compare efficacy with other vaccine strategies in a single dose. DEC-205-targeted HIV gag p24 or p41 induces stronger CD4+ T cell immunity relative to high doses of gag protein, HIV gag plasmid DNA, or recombinant adenovirus-gag. High frequencies of interferon (IFN)-gamma- and interleukin 2-producing CD4+ T cells are elicited, including double cytokine-producing cells. In addition, the response is broad because the primed mice respond to an array of peptides in different major histocompatibility complex haplotypes. Long-lived T cell memory is observed. After subcutaneous vaccination, CD4+ and IFN-gamma-dependent protection develops to a challenge with recombinant vaccinia-gag virus at a mucosal surface, the airway. We suggest that a DEC-targeted vaccine, in part because of an unusually strong and protective CD4+ T cell response, will improve vaccine efficacy as a stand-alone approach or with other modalities.

AIDS Vaccines↗
PubMed2006

The interactions of dendritic cells with antigen-specific, regulatory T cells that suppress autoimmunity.

Dendritic cells (DCs) are important for several aspects of the development and function of CD4(+) CD25(+) regulatory T cells (Tregs), which are critical for maintaining peripheral tolerance and preventing autoimmunity. In cultures from human thymus, dendritic cells (DCs) conditioned with thymic stromal lymphopoietin (TSLP) mediate the production of Tregs from CD4(+) CD25(-) thymocytes. In cultures from mouse lymphoid organs, CD86-rich DCs induce the proliferation and improved suppressive function of antigen-specific Tregs. DC-expanded, antigen-specific Tregs show greatly enhanced efficacy relative to polyclonal populations in blocking experimental autoimmunity. In several animal models including NOD diabetes, Tregs directed to one autoantigen are able to block autoimmunity induced by multiple antigens from the target organ. Distinct states of DC differentiation or maturation are likely to be important for the emerging roles of DCs in the biology of Tregs, particularly the control of autoimmunity in an antigen-dependent manner.

Animals↗
PubMed2006

Effective expansion of alloantigen-specific Foxp3+ CD25+ CD4+ regulatory T cells by dendritic cells during the mixed leukocyte reaction.

Thymic-derived CD25+ CD4+ T regulatory cells (Tregs) suppress immune responses, including transplantation. Here we evaluated the ability of dendritic cells (DCs) to expand alloantigen-specific Tregs in the mixed leukocyte reaction (MLR) that develops from polyclonal populations of T cells. The allogeneic DCs, when supplemented with IL-2 in the cultures, were much more effective than bulk spleen cells in expanding the numbers of Tregs. Likewise, DCs and not spleen cells were effective in sustaining expression of the transcription factor Foxp3 in Tregs, but neither IL-2 nor CD80/86 was required for this effect in the cultures. On a per-cell basis, the DC-expanded, but not unexpanded, Tregs were more potent suppressors of a fresh MLR by CD25- CD4+ T cells. Suppression was 3- to 10-fold more active for MLRs induced by the original alloantigens than for third-party stimulators. When DC-expanded Tregs were introduced into sublethally irradiated hosts, the T cells suppressed graft-versus-host-disease induced by CD25- CD4+ T cells. Again, suppression was more active against the same mouse strain that provided the DCs to expand the Tregs. Therefore, alloantigen-selected Tregs are more effective suppressors of responses to major transplantation antigens, and these Tregs can be expanded from a polyclonal repertoire by DCs.

Animals↗
PubMed2006

HIV-1 selectively infects a subset of nonmaturing BDCA1-positive dendritic cells in human blood.

The infection of cultured monocyte-derived dendritic cells (DCs) with HIV-1 involves CD4 and CCR5 receptors, while transmission to T cells is enhanced at least in part by the lectin DC-SIGN/CD209. In the present study, we studied BDCA-1+ myeloid DCs isolated directly from human blood. These cells express CD4 and low levels of CCR5 and CXCR4 coreceptors, but not DC-SIGN. The myeloid DCs replicate two R5 viruses, BaL and YU2, and transfer infection to activated T cells. The virus productively infects a small fraction of the blood DCs that fail to mature in culture, as indicated by the maturation markers CD83 and DC-LAMP/CD208, and the expression of high CD86 and MHC class II, in contrast to many noninfected DCs. A greater proportion of BDCA-1+ DCs are infected when the virus is pseudotyped with the vesicular stomatitis envelope VSV-G (5-15%), as compared with the R5 virus (0.3-3.5%), indicating that HIV-1 coreceptors may limit the susceptibility of DCs to become infected, or the endocytic route of viral entry used by HIV/vesicular stomatitis virus enhances infectivity. When infected and noninfected cells are purified by cell sorting, the former uniformly express HIV p24 gag and are virtually inactive as stimulators of the allogeneic MLR, in contrast to potent stimulation by noninfected DCs from the same cultures. These results point to two roles for a small fraction of blood DCs in HIV-1 pathogenesis: to support productive infection and to evade the direct induction of T cell-mediated immunity.

Antigens, Surface↗
PubMed2006

Dendritic cells expand antigen-specific Foxp3+ CD25+ CD4+ regulatory T cells including suppressors of alloreactivity.

Thymic derived naturally occurring CD25+ CD4+ T regulatory cells (Tregs) suppress immune responses, including transplantation. Here we discuss the capacity of dendritic cells (DCs) to expand antigen-specific Tregs, particularly polyclonal Tregs directed to alloantigens. Initial studies have shown that mature DCs are specialized antigen-presenting cells (APCs) for expanding antigen-specific CD25+ CD4+ Tregs from TCR transgenic mice. When triggered by specific antigen, these Tregs act back on immature DCs to block the upregulation of CD80 and CD86 costimulatory molecules. More recently, DCs have been used to expand alloantigen-specific CD25+ CD4+ Tregs from the polyclonal repertoire in the presence of interleukin-2 (IL-2). Allogeneic DCs are much more effective than allogeneic spleen cells for expanding CD25+ CD4+ Tregs. The DC-expanded Tregs continue to express high levels of Foxp3, even without supplemental IL-2, whereas spleen cells poorly sustain Foxp3 expression. When suppressive activity is tested, relatively small numbers of DC-expanded CD25+ CD4+ Tregs exert antigen-specific suppression in the mixed leukocyte reaction (MLR), blocking immune responses to the original stimulating strain 10 times more effectively than to third party stimulating cells. DC-expanded Tregs also retard graft versus host disease (GVHD) across full major histocompatibility complex (MHC) barriers. In vitro and in vivo, the alloantigen-specific CD25+ CD4+ Tregs are much more effective suppressors of transplantation reactions than polyclonal populations. We suggest that the expansion of Tregs from a polyclonal repertoire via antigen-presenting DCs will provide a means for antigen-specific control of unwanted immune reactions.

Animals↗
PubMed2005

Increase in TNF-alpha and inducible nitric oxide synthase-expressing dendritic cells in psoriasis and reduction with efalizumab (anti-CD11a).

We find that CD11c(+) cells with many markers of dendritic cells (DCs) are a major cell type in the skin lesions of psoriasis. These CD11c(+) cells, which are evident in both epidermis and dermis, are the sites for the expression of two mediators of inflammation, inducible nitric oxide synthase (iNOS) and TNF-alpha in diseased skin. These cells express HLA-DR, CD40, and CD86, lack the Langerin and CD14 markers of Langerhans cells and monocytes, respectively, and to a significant extent express the DC maturation markers DC-LAMP and CD83. Treatment of psoriasis with efalizumab (anti-CD11a, Raptiva) strongly reduces infiltration by these DCs in patients responding to this agent. Disease activity after therapy was more related to DC infiltrates and iNOS mRNA levels than T cell infiltrates, and CD11c(+) cells responded more quickly to therapy than epidermal keratinocytes. Our results suggest that a type of DC, which resembles murine "Tip-DCs" that can accumulate during infection, has proinflammatory effects in psoriasis through nitric oxide and TNF-alpha production, and can be an important target for suppressive therapies.

Antibodies, Monoclonal↗