Biomedical subjects

Benjamin M Chain

Publications and source records attributed to Benjamin M Chain.

At least 19 recordsLinked to original sources

PubMed2006

beta1-Integrins determine the dendritic morphology which enhances DC-SIGN-mediated particle capture by dendritic cells.

The morphology of antigen-presenting dendritic cells (DCs) is characterized by the presence of numerous long dendrites. The formation of these processes is shown to require the interaction between the beta1-integrin (CD29) on the surface of the DCs and fibronectin in the extracellular matrix. This interaction occurs at focal contacts formed at the tips of dendrites, which contain high concentrations of the beta1-integrins, actin and the cytoskeletal proteins vinculin, paxillin and talin. Dendrites contain an extensive microtubule (MT) network, and are retracted in the presence of the MT inhibitor colchicine, suggesting that MTs are essential for dendrite stability. The dendritic morphology is shown to contribute directly to an enhanced ability to capture dendritic cell specific ICAM-3 grabbing nonintegrin (DC-SIGN)-coated beads. Time-lapse photography demonstrates that dendrites are highly dynamic structures, with cells extending and retracting multiple dendrites in different directions over a 3-h period. This motility increases the area scanned by an individual DC by over 2-fold. The unusual combination of a dendritic morphology and high motility is likely to play a major role in the efficient function of DCs as sentinels of the immune system.

Actins↗

PubMed2006

Mannose-pepstatin conjugates as targeted inhibitors of antigen processing.

The molecular details of antigen processing, including the identity of the enzymes involved, their intracellular location and their substrate specificity, are still incompletely understood. Selective inhibition of proteolytic antigen processing enzymes such as cathepsins D and E, using small molecular inhibitors such as pepstatin, has proven to be a valuable tool in investigating these pathways. However, pepstatin is poorly soluble in water and has limited access to the antigen processing compartment in antigen presenting cells. We have synthesised mannose-pepstatin conjugates, and neomannosylated BSA-pepstatin conjugates, as tools for the in vivo study of the antigen processing pathway. Conjugation to mannose and to neomannosylated BSA substantially improved the solubility of the conjugates relative to pepstatin. The mannose-pepstatin conjugates showed no reduction in inhibition of cathepsin E, whereas the neomannosylated BSA-pepstatin conjugates showed some loss of inhibition, probably due to steric factors. However, a neomannosylated BSA-pepstatin conjugate incorporating a cleavable disulfide linkage between the pepstatin and the BSA showed the best uptake to dendritic cells and the best inhibition of antigen processing.

Animals↗

PubMed2006

Hypochlorous acid enhances immunogenicity and uptake of allogeneic ovarian tumor cells by dendritic cells to cross-prime tumor-specific T cells.

BACKGROUND: Ovarian cancer commonly relapses after remission and new strategies to target microscopic residual diseases are required. One approach is to activate tumor-specific cytotoxic T cells with dendritic cells loaded with tumor cells. In order to enhance their immunogenicity, ovarian tumor cells (SK-OV-3, which express two well-characterized antigens HER-2/neu and MUC-1) were killed by oxidation with hypochlorous acid (HOCl). RESULTS: Treatment for 1 h with 60 microM HOCl was found to induce necrosis in all SK-OV-3 cells. Oxidized, but not live, SK-OV-3 was rapidly taken up by monocyte-derived dendritic cells, and induced partial dendritic cell maturation. Dendritic cells cultured from HLA-A2 healthy volunteers were loaded with oxidized SK-OV-3 (HLA-A2-) and co-cultured with autologous T cells. Responding T cells were tested for specificity after a further round of antigen stimulation. In ELISPOT assays, T cells produced interferon-gamma (IFN-gamma) in response to the immunizing cellular antigen, and also to peptides coding for MUC-1 and HER-2/neu HLA-A2 restricted epitopes, demonstrating efficient cross-presentation of cell-associated antigens. In contrast, no responses were seen after priming with heat-killed or HCl-killed SK-OV-3, indicating that HOCl oxidation and not cell death/necrosis per se enhanced the immunogenicity of SK-OV-3. Finally, T cells stimulated with oxidized SK-OV-3 showed no cross-reaction to oxidized melanoma cells, nor vice versa, demonstrating that the response was tumor-type specific. CONCLUSIONS: Immunization with oxidized ovarian tumor cell lines may represent an improved therapeutic strategy to stimulate a polyclonal anti-tumor cellular immune response and hence extend remission in ovarian cancer.

Cell Line, Tumor↗

PubMed2006

Monocyte derived dendritic cells from HIV-1 infected individuals partially reconstitute CD4 T-cell responses.

OBJECTIVES: The study tests the hypothesis that monocyte derived dendritic cells from HIV-1 infected individuals are normal and can restore impaired CD4 T-cell antigen specific responses. DESIGN: Monocyte derived dendritic cells were isolated from individuals at three different stages of HIV-1 infection with a wide spectrum of viral load and CD4 T-cell counts, and from healthy volunteers. The cell surface phenotype and allogeneic stimulatory potential of these dendritic cells was documented. CD4 T-cell responses to HIV p24, tetanus toxoid and purified protein derivative were measured using either unfractionated peripheral blood mononuclear cells, or purified dendritic cell/T-cell cultures. RESULTS: Dendritic cells from all three HIV-1 infected groups did not differ from each other or from healthy volunteers in terms of cell surface phenotype or allogeneic stimulatory potential using T cells from healthy volunteers. Dendritic cells from immunosuppressed antiretroviral naive individuals enhanced the autologous recall proliferative responses both to HIV-1 p24, and third party antigens tetanus toxoid and purified protein derivative, both in terms of the proportion of responding individuals, and median proliferation. CONCLUSION: Antigen presentation by dendritic cells partially restores impaired antigen specific CD4 T-cell responses associated with HIV-1 infection. Immunization strategies which target dendritic cells may therefore offer significant advantages in the ability to stimulate HIV-specific protective immune responses.

Adult↗

PubMed2006

Modulation of dendritic cell differentiation and cytokine secretion by the hydatid cyst fluid of Echinococcus granulosus.

Chronic infection by Echinococcus granulosus results in establishment of fluid-filled cysts (hydatid cysts) in liver or lungs of infected hosts, which can escape destruction by the host immune system for long periods. This study explores the modulation by hydatid cyst fluid of the in vitro human monocyte to dendritic cell (DC) transition induced by granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4). Addition of the fluid to adherent peripheral blood monocytes cultured in GM-CSF/IL-4 stimulates release of prostaglandin E2 (PGE2) and IL-6. Exposure of differentiating DC to the fluid during the 7-day culture in GM-CSF/IL-4 impairs their subsequent ability to secrete IL-12, IL-6 or PGE2 in response to lipopolysaccharide (LPS) stimulation. This inhibition is not dependent on the initial release of PGE2. The presence of hydatid cyst fluid also modulates the phenotype of the cells generated during culture, resulting in increased CD14 expression and decreased expression of CD1a. Finally, hydatid fluid can stimulate predifferentiated DC to mature, as evidenced by release of IL-12 and IL-6, and by up-regulation of class II major histocompatibility complex and CD86. The possible role of dendritic cell modulation in regulating the host immune response to hydatid cysts is discussed.

Animals↗

PubMed2005

Immunization with a lentiviral vector stimulates both CD4 and CD8 T cell responses to an ovalbumin transgene.

Lentiviral vectors encoding antigens are promising vaccine candidates because they transduce dendritic cells (DC) in vivo and prime CTL responses. Here we examine their stimulation of antigen-specific CD4(+) T cells, critical for protective immunity against tumors or infectious disease. We constructed lentiviral vectors (lentivectors) expressing ovalbumin, which was secreted (OVA), cytoplasmic (OVAcyt), or fused to either invariant chain (Ii-OVA) or transferrin receptor (TfR-OVA) sequences, targeting the MHC class II presentation pathway. Murine DC infected with the various lentivectors could stimulate OT-I (CD8(+), OVA TCR transgenic) T cells and all except OVAcyt could also stimulate OT-II (CD4(+), OVA TCR transgenic) T cells in vitro. Direct injection of the OVA-, Ii-OVA-, or TfR-OVA-expressing vectors into mice resulted in a CD4(+) T cell response, as shown by expansion of adoptively transferred OT-II T cells and upregulation of CD44 on these cells. The Ii-OVA vector was the most potent inducer of IFN-gamma-secreting CD4(+) and CD8(+) T cells and was the only vector to protect mice completely from challenge with OVA-expressing tumor cells. Therefore directly injected lentivectors can stimulate CD4(+) T cells; both CD4(+) and CD8(+) responses can be enhanced by targeting the antigen to the MHC class II pathway.

Animals↗

PubMed2005

Variability and conservation in hepatitis B virus core protein.

BACKGROUND: Hepatitis B core protein (HBVc) has been extensively studied from both a structural and immunological point of view, but the evolutionary forces driving sequence variation within core are incompletely understood. RESULTS: In this study, the observed variation in HBVc protein sequence has been examined in a collection of a large number of HBVc protein sequences from public sequence repositories. An alignment of several hundred sequences was carried out, and used to analyse the distribution of polymorphisms along the HBVc. Polymorphisms were found at 44 out of 185 amino acid positions analysed and were clustered predominantly in those parts of HBVc forming the outer surface and spike on intact capsid. The relationship between HBVc diversity and HBV genotype was examined. The position of variable amino acids along the sequence was examined in terms of the structural constraints of capsid and envelope assembly, and also in terms of immunological recognition by T and B cells. CONCLUSION: Over three quarters of amino acids within the HBVc sequence are non-polymorphic, and variation is focused to a few amino acids. Phylogenetic analysis suggests that core protein specific forces constrain its diversity within the context of overall HBV genome evolution. As a consequence, core protein is not a reliable predictor of virus genotype. The structural requirements of capsid assembly are likely to play a major role in limiting diversity. The phylogenetic analysis further suggests that immunological selection does not play a major role in driving HBVc diversity.

Amino Acid Sequence↗

PubMed2005

JNK activation limits dendritic cell maturation in response to reactive oxygen species by the induction of apoptosis.

Dendritic cells (DC) sense infection in their local microenvironment and respond appropriately in order to induce T cell immunity. This response is mediated in part via the mitogen-activated protein kinase (MAPK) pathways. Hydrogen peroxide is present frequently in the inflammatory DC milieu and is known to activate MAPK. Therefore this study examines the role of hydrogen peroxide, both alone and in combination with lipopolysaccharide (LPS), in the regulation of activation of two key MAPK, p38 and JNK, regulation of phenotype, and regulation of apoptosis in human monocyte-derived DC. At low concentrations, hydrogen peroxide activates p38, but does not alter DC phenotype. At higher concentrations, hydrogen peroxide activates both p38 and JNK. Activation of JNK, which is associated with inhibition of tyrosine phosphatases in DC, is linked to the induction of DC apoptosis. An upstream JNK inhibitor (CEP11004) and a competitive JNK inhibitor (SP600125) both partially protected the DC from the proapoptotic effects of hydrogen peroxide. Unexpectedly, hydrogen peroxide and LPS synergize in inducing JNK activation and DC apoptosis. JNK-mediated apoptosis may limit damaging immune responses against neoepitopes generated by modification of self-antigens by reactive oxygen species present at sites of inflammation.

Apoptosis↗

PubMed2005

The expression and function of cathepsin E in dendritic cells.

Cathepsin E is an aspartic proteinase that has been implicated in Ag processing within the class II MHC pathway. In this study, we document the presence of cathepsin E message and protein in human myeloid dendritic cells, the preeminent APCs of the immune system. Cathepsin E is found in a perinuclear compartment, which is likely to form part of the endoplasmic reticulum, and also a peripheral compartment just beneath the cell membrane, with a similar distribution to that of Texas Red-dextran within 2 min of endocytosis. To investigate the function of cathepsin E in processing, a new soluble targeted inhibitor was synthesized by linking the microbial aspartic proteinase inhibitor pepstatin to mannosylated BSA via a cleavable disulfide linker. This inhibitor was shown to block cathepsin D/E activity in cell-free assays and within dendritic cells. The inhibitor blocked the ability of dendritic cells from wild-type as well as cathepsin D-deficient mice to present intact OVA, but not an OVA-derived peptide, to cognate T cells. The data therefore support the hypothesis that cathepsin E has an important nonredundant role in the class II MHC Ag processing pathway within dendritic cells.

Animals↗

PubMed2005

LIGHTing up dendritic cell activation: Immune regulation and viral exploitation.

The maturation state of dendritic cells (DC) is regulated by a variety of factors. These include ligands expressed by T cells, such as members of the TNF superfamily. Recent studies have highlighted the role of one such molecule, LIGHT, as a positive regulator of DC biology, promoting the maturation of these cells through the activation of NF-kappaB pathways. In addition, HSV-1 envelope glycoproteins can also bind the LIGHT receptor, herpes virus entry mediator (HVEM), and activate similar downstream signalling pathways in DC. The consequence of this host-viral interaction may be a novel pathway of viral immune evasion.

Dendritic Cells↗

PubMed2005

Is hepatitis C virus infection of dendritic cells a mechanism facilitating viral persistence?

More than 170 million people worldwide are chronically infected with hepatitis C virus (HCV), which is a major cause of chronic hepatitis, cirrhosis, and hepatocellular carcinoma. Impaired T-cell reactivity to HCV, a hallmark of inefficient adaptive immunity, is believed to be responsible for the high propensity of HCV to cause chronic infection. Dendritic cells are the most potent antigen-presenting cells and many viruses affect various dendritic cell functions. Data suggest that such changes induced by HCV may have an important role in viral persistence. HCV has been shown to bind to dendritic cells, although viral replication within these cells occurs at a very low level. Dendritic cells from people with chronic HCV infection are impaired in their capacity to stimulate T cells. This impairment may be a consequence of changes in the expression of major histocompatibility complex and costimulatory molecules on its surface, as well as in the production of cytokines such as interleukin 12. In addition, hepatic dendritic cells may be affected by the tolerogenic microenvironment of the liver, possibly generating dendritic cells that promote regulatory T cells, which suppress the cellular immune response mounted against HCV.

Dendritic Cells↗

PubMed2005

Dendritic cells in viral pathogenesis: protective or defective?

Dendritic cells (DC) are potent antigen-presenting cells that are critical in the initiation of immune responses to control and/or eliminate viral infections. Recent studies have investigated the effects of virus infection on the biology of DC. This review summarizes these changes, focusing on both the DC parameters affected and the viral factors involved. In addition, the central role of DC biology in the pathogenesis of several viral families, including herpesviruses, paramyxoviruses and retroviruses, is explored. The field of pathogen recognition by DC is addressed, focusing on its role in protecting the host from viral infection, as well as the ability of viruses to exploit such host receptor ligation and signalling to their replicative advantage. The hypothesis is proposed that virus and host have evolved a symbiotic relationship to ensure both viral transmission and host survival.

Cell Survival↗

PubMed2004

The use of targeted microbeads for quantitative analysis of the phagocytic properties of human monocyte-derived dendritic cells.

The relationship between phagocytic capacity and morphology of dendritic cells (DCs) has not been investigated previously. Therefore, in order to approach this question, we have developed a novel assay, which is described here. The model of dendritic cells (DCs) used was based upon cytokine-induced differentiation of peripheral blood mononuclear cells, followed by culture on a fibronectin substratum. Under these conditions, standard current methods of quantifying phagocytosis are not applicable, as they rely upon flow cytometric analysis of fluid phase cells; and for adherent cells, quantitative efficiency of uptake is very difficult to measure. Furthermore, for both fluid phase and adherent cells, it is difficult to discriminate between internal and externally bound probe, and degradation of internalised probes can lead to artefacts. Therefore, in this study, these technical issues have been overcome by a simple and flexible assay. Phycoerythrin (PE)-conjugated antibodies are used to target microbeads to the DCs. Following an appropriate incubation period, secondary staining with fluorescein isothiocyanate (FITC)-conjugated antibody allows discrimination between internal and externally bound beads. Microscopic visualisation allows individual beads to be studied easily and thus phagocytosis quantified, whilst permitting parallel examination of morphological parameters. In particular, the relationship between bead uptake and the nature and distribution of the dendritic processes can be evaluated.

Antibodies↗

PubMed2004

Cynaropicrin, a sesquiterpene lactone, as a new strong regulator of CD29 and CD98 functions.

Cynaropicrin is a sesquiterpene lactone displaying immunomodulatory effects on the production of cytokine and nitric oxide from macrophages/monocytes. In this study we have examined inhibitory effect of cynaropicrin on activation of major adhesion molecules [CD29 (beta1 integrins), CD43, and CD98] on the cells assessed by U937 (promonocytic cells) homotypic aggregation. Cynaropicrin potently blocked CD29 (beta1 integrins)- and CD98-induced homotypic aggregation with IC(50) values of 3.46 and 2.98 microM, respectively, without displaying cytotoxicity. Similarly, flow cytometric analysis exhibited that cynaropicrin down-regulated strikingly surface level of CD29 and CD147, a functional regulator of CD98, but not CD43. More importantly, cynaropicrin inhibition was linked to blockade of extracellular signal-related kinase (ERK) activation and distinct from other enzyme inhibitors including rottlerin, propranolol, forskolin, and chloroquine, but not cytochalasin B. Therefore, our finding is the first demonstration that cynaropicrin may be a potent functional regulator of CD29 and CD98 via interrupting ERK activation which may be linked to cytoskeleton rearrangement, suggesting further application to CD29- and CD98-mediated diseases such as virus-induced chronic inflammation, and invasion, migration, and metastasis of leukocyte cancer cells.

Antigens, CD↗

PubMed2004

The host response to herpes simplex virus infection.

PURPOSE OF REVIEW: Infection with herpes simplex virus remains a significant cause of disease. The host immune system plays an important role in containing viral replication, and there has been considerable progress in defining which components of immunity are key to the resolution of infection. Nevertheless, effective immunoprophylaxis or immunotherapy has not yet been achieved. RECENT FINDINGS: Recent work has focused on understanding the early events leading to the herpes simplex virus-specific immune response, in particular on the role of antigen-presenting dendritic cells. Herpes simplex virus has evolved a number of ways of interfering with antigen presentation by dendritic cells, thus presumably impeding or delaying the host immune response. Nevertheless, herpes simplex virus triggers strong cellular and humoral immunity. The ability of dendritic cells to take up dead or dying infected cells and cross-present them to cognate T cells may be the key to resolving this apparent paradox. Interaction between dendritic cell subsets, and particularly the virus-induced release of type I interferons may be essential to drive efficient antigen cross-presentation and subsequent T-cell activation. SUMMARY: A greater understanding of the importance of dendritic cells in driving viral immunity, and of the ligands that activate these cells and the cytokines they secrete, has provided novel vaccination strategies. The delivery of immunomodulatory genes together with viral antigens, for example by DNA vaccination, may harness the full potential of dendritic cells, and achieve the goal of effective immunological control of herpes simplex virus.

Dendritic Cells↗

PubMed2004

Structural relationships and cellular tropism of staphylococcal superantigen-like proteins.

The staphylococcal superantigen-like proteins (SSLs) are a family of polymorphic paralogs encoded in the Staphylococcus aureus genome whose function is unknown. The crystal structure of SSL7 was determined and compared to that of SSL5 and that of a classical superantigen, streptococcal pyrogenic exotoxin. Although the overall architecture of the superantigen family is retained in both SSL7 and SSL5, there are significant differences in the structures which suggest that the characteristic major histocompatibility complex binding site of superantigens has been lost. To complement these data, the abilities of SSL7 and a closely related paralog, SSL9, to interact with cells of the immune system were investigated. In populations of human white blood cells, both SSLs interacted selectively with monocytes via specific saturable but separate binding sites, which led to rapid uptake of the SSLs. In addition, SSLs were rapidly taken up by dendritic cells, but not by macrophages, into the same endosomal compartment as dextran. The ability of these secreted proteins to target antigen-presenting cells may enhance a misplaced antibody response against the proteins, which may facilitate bacterial colonization rather than contribute to host protection. Like classical superantigens, therefore, SSLs may distract the host's immune system, but they may do so via entirely different molecular mechanisms.

Bacterial Proteins↗

PubMed2003

Current issues in antigen presentation--focus on the dendritic cell.

A personal view of some of the major current issues in antigen presentation, which focuses on the biology of the dendritic cell. The question of dendritic cell heterogeneity is discussed and the inclusion of the "plasmacytoid" dendritic cell in this family is questioned. Defining the molecules involved in dendrite formation may ultimately provide the only reliable signature of the dendritic cell. The role of the dendritic cell as a key link between innate and adaptive immunity is here revisited, and the direct and indirect interactions between dendritic cells and microorganisms are discussed. Reactive oxygen species may be key modulators of dendritic cell function, although it remains unclear whether direct or indirect mechanisms are more important. Study of the cell biology of antigen processing reveals a tightly regulated complex system involving multiple proteinases, adapted to the most efficient presentation of invading microorganisms and playing a key role in shaping the T cell repertoire.

Animals↗

PubMed2003

Regulation of CD43-induced U937 homotypic aggregation.

CD43 (leukosialin, sialophorin), a prominent component of the hemopoietic cell surface, has an enigmatic role in cell-cell interaction. The observation that CD43 ligation triggers homotypic aggregation of monoblastoid U937 cells has permitted analysis of this: CD43-induced aggregation was distinguishable from CD29- (also known as beta1 integrin) or CD98- (also known as 4F2, or fusion-related protein 1) induced aggregation, with different energy requirements and with partial dependence on beta2 integrins. Previous studies have focused on the role of CD43 ligation in tyrosine phosphorylation. However, in the homotypic adhesion assay, although there is initial tyrosine phosphorylation, protein tyrosine kinase inhibitors did not block aggregation. Therefore, other signaling pathways were examined. CD43 ligation induced protein tyrosine dephosphorylation, and protein tyrosine phosphatase inhibitors blocked aggregation. Activation of MAP kinases was not necessary. Cytoskeletal inhibitors amplified aggregation. Protein kinase C (PKC) inhibitors amplified aggregation, implicating PKC as a negative regulator. CD43 ligation up-regulated surface adhesion molecules and enhanced CD29- and CD98-induced aggregation. Thus, CD43 participation in cell-cell adhesion is under stringent control, involving both surface events and several different intracellular signaling pathways, acting together to regulate the process. These mechanisms add a further dimension to the potential role of CD43 in tissue immune responses.

Antigens, CD↗