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Christopher C Dascher

Publications and source records attributed to Christopher C Dascher.

At least 19 recordsLinked to original sources
PubMed2006

BCG vaccine elicits both T-cell mediated and humoral immune responses directed against mycobacterial lipid components.

The universe of antigens recognized by alphabeta T cells has recently been expanded to include not only major histocompatibility complex (MHC)-presented protein antigens but also CD1-presented lipid antigens. The significance of lipid-reactive T cells in host defense has been appreciated, using the guinea pig model of human tuberculosis. Here, we show that immunization with Mycobacterium bovis bacillus Calmette-Guerin (BCG), the commonly used anti-tuberculosis vaccine, induces activation of guinea pig cytotoxic T cells recognizing BCG lipids in the context of CD1 molecules. Further, BCG-immunized, but not mock-immunized, guinea pigs mount IgG antibody responses directed against lipoarabinomannnan, an essential cell wall lipid component of mycobacteria. These observations emphasize the ability of BCG to activate the host adaptive immunity to mycobacteria-derived lipids, which could potentially contribute to protection against tuberculosis.

Animals↗
PubMed2005

Mycobacterium tuberculosis regulates CD1 antigen presentation pathways through TLR-2.

Mycobacterium tuberculosis remains a major pathogen of worldwide importance, which releases lipid Ags that are presented to human T cells during the course of tuberculosis infections. Here we report that cellular infection with live M. tuberculosis or exposure to mycobacterial cell wall products converted CD1- myeloid precursors into competent APCs that expressed group 1 CD1 proteins (CD1a, CD1b, and CD1c). The appearance of group 1 CD1 proteins at the surface of infected or activated cells occurred via transcriptional regulation, and new CD1 protein synthesis and was accompanied by down-regulation of CD1d transcripts and protein. Isolation of CD1-inducing factors from M. tuberculosis using normal phase chromatography, as well as the use of purified natural and synthetic compounds, showed that this process involved polar lipids that signaled through TLR-2, and we found that TLR-2 was necessary for the up-regulation of CD1 protein expression. Thus, mycobacterial cell wall lipids provide two distinct signals for the activation of lipid-reactive T cells: lipid Ags that activate T cell receptors and lipid adjuvants that activate APCs through TLR-2. These dual activation signals may represent a system for selectively promoting the presentation of exogenous foreign lipids by those myeloid APCs, which come into direct contact with pathogens.

Animals↗
PubMed2005

The Mycobacterium tuberculosis serine/threonine kinases PknA and PknB: substrate identification and regulation of cell shape.

The Mycobacterium tuberculosis genome contains 11 serine/threonine kinase genes including two, pknA and pknB, that are part of an operon encoding genes involved in cell shape control and cell wall synthesis. Here we demonstrate that pknA and pknB are predominantly expressed during exponential growth, and that overexpression of these kinases slows growth and alters cell morphology. We determined the preferred substrate motifs of PknA and PknB, and identified three in vivo substrates of these kinases: PknB; Wag31, an ortholog of the cell division protein DivIVA; and Rv1422, a conserved protein of unknown function. Expression of different alleles of wag31 in vivo alters cell shape, in a manner dependent on the phosphoacceptor residue in the protein produced. Partial depletion of pknA or pknB results in narrow, elongated cells. These data indicate that signal transduction mediated by these kinases is a novel mechanism for the regulation of cell shape in mycobacteria, one that may be conserved among gram-positive bacteria.

Amino Acid Sequence↗
PubMed2005

Characterization of two avian MHC-like genes reveals an ancient origin of the CD1 family.

Many of the genes that comprise the vertebrate adaptive immune system are conserved across wide evolutionary time scales. Most notably, homologs of the mammalian MHC gene family have been found in virtually all jawed vertebrates, including sharks, bony fishes, reptiles, and birds. The CD1 family of antigen-presenting molecules are related to the MHC class I family but have evolved to bind and present lipid antigens to T cells. Here, we describe two highly divergent nonclassical MHC class I genes found in the chicken (Gallus gallus) that have sequence homology to the mammalian CD1 family of proteins. One of the chicken CD1 genes expresses a full-length transcript, whereas the other has multiple splice variants. Both Southern blot and single nucleotide polymorphism analysis indicates that chicken CD1 is relatively nonpolymorphic. Moreover, cross-hybridizing bands are present in other bird species, suggesting broad conservation in the avian class. Northern analysis of chicken tissue shows a high level of CD1 expression in the bursa and spleen. In addition, molecular modeling predicts that the potential antigen-binding pocket is probably hydrophobic, a universal characteristic of CD1 molecules. Genomic analysis indicates that the CD1 genes are located on chicken chromosome 16 and maps to within 200 kb of the chicken MHC B locus, suggesting that CD1 genes diverged from classical MHC genes while still linked to the major histocompatibility complex locus. The existence of CD1 genes in an avian species suggests that the origin of CD1 extends deep into the evolutionary history of terrestrial vertebrates.

Amino Acid Sequence↗
PubMed2005

Alternative bisphosphonate targets and mechanisms of action.

As the number of bisphosphonates continues to increase, they have found widespread use in an increasing number of clinical conditions. Ongoing examination of their targets and mechanisms of action has revealed that this surprisingly diverse class of drugs has effects beyond those first described for osteoclasts. These additional targets include osteoblasts, osteocytes, angiogenesis, and gammadelta T lymphocytes of the human immune system. The immune system effects are specifically targeted to gammadelta T cells and are reminiscent of the effects seen after ingestion of tea beverage. BP effects on such alternate targets may explain not only their antiresorptive effect, but also their effect on bone quality, tumors, and microbes.

Animals↗
PubMed2005

Subcellular localization of mycobacteria in tissues and detection of lipid antigens in organelles using cryo-techniques for light and electron microscopy.

The survival of intracellular pathogens within a host is determined by microbial evasion, which can be partially attributed to their subcellular trafficking strategies. Microscopic techniques have become increasingly important in understanding the cell biology of microbial infections. These recently developed techniques can be used for the subcellular localization of antigens not only in cultured cells but also within tissues such as Mycobacterium tuberculosis in lung and Mycobacterium leprae in skin. High-resolution immunofluorescence microscopy can be used in combination with cryo-immunogold electron microscopy using consecutive cryo-sections on the same tissue block forming a direct connection between the two microscopy techniques. The detection of mycobacterial lipid antigens in situ at an ultrastructural level is currently a challenge, but new modifications can be used to address this. These methods might be of interest to microbiologists and cell biologists who study host-pathogen interactions.

Antigens, Bacterial↗
PubMed2004

Cadherin-11 provides specific cellular adhesion between fibroblast-like synoviocytes.

Cadherins are integral membrane proteins expressed in tissue-restricted patterns that mediate homophilic intercellular adhesion. During development, they orchestrate tissue morphogenesis and, in the adult, they determine tissue integrity and architecture. The synovial lining is a condensation of fibroblast-like synoviocytes (FLS) and macrophages one to three cells thick. These cells are embedded within the extracellular matrix, but the structure is neither an epithelium nor an endothelium. Previously, the basis for organization of the synovium into a tissue was unknown. Here, we cloned cadherin-11 from human rheumatoid arthritis (RA)-derived FLS. We developed L cell transfectants expressing cadherin-11, cadherin-11 fusion proteins, and anti-cadherin-11 mAb. Cadherin-11 was found to be expressed mainly in the synovial lining by immunohistologic staining of human synovium. FLS adhered to cadherin-11-Fc, and transfection of cadherin-11 conferred the formation of tissue-like sheets and lining-like structures upon fibroblasts in vitro. These findings support a key role for cadherin-11 in the specific adhesion of FLS and in synovial tissue organization and behavior in health and RA.

Animals↗
PubMed2004

Identification of guinea pig gammadelta T cells and characterization during pulmonary tuberculosis.

Guinea pigs are an alternative small animal model for many disease studies. Here we describe a pan-gammadelta monoclonal antibody (anti-TCRdelta1) specific for the constant region of human T cell receptor delta chains that cross-reacts with a subpopulation of guinea pig (Cavia porcellus) lymphocytes. The phenotype and distribution of this subpopulation is consistent with the guinea pig gammadelta T cell subset. FACS analysis of fresh PBMC and splenocytes from naïve guinea pigs revealed the presence of a subset of cells that stained with the anti-TCRdelta1 mAb. The relative percentage of anti-TCRdelta1 positive cells in PBMC and tissues is similar to that described for gammadelta T cells in other species. Immunohistochemistry of tissues also revealed a distribution of anti-TCRdelta1 positive cells consistent with gammadelta T cells. These data are further supported by staining of a polyclonal guinea pig T cell line that became progressively CD4 and CD8 negative in long-term culture. Analysis of PBMC from guinea pigs following aerosol infection with virulent Mycobacterium tuberculosis revealed no apparent changes in the steady-state percentage of blood gammadelta+ T cells. Taken together, these data suggest that the anti-TCRdelta1 antibody recognizes the gammadelta T cell subset in guinea pigs. This reagent may be useful for examining gammadelta T cells in various disease models where the guinea pig is a more desirable model for study.

Animals↗
PubMed2004

Protection elicited by a double leucine and pantothenate auxotroph of Mycobacterium tuberculosis in guinea pigs.

We developed a live, fully attenuated Mycobacterium tuberculosis vaccine candidate strain with two independent attenuating auxotrophic mutations in leucine and pantothenate biosynthesis. The deltaleuD deltapanCD double auxotroph is fully attenuated in the SCID mouse model and highly immunogenic and protective in the extremely sensitive guinea pig tuberculosis model, reducing both bacterial burden and disease pathology.

Animals↗
PubMed2004

Transcription regulation by the Mycobacterium tuberculosis alternative sigma factor SigD and its role in virulence.

Mycobacterium tuberculosis, an obligate mammalian pathogen, adapts to its host during the course of infection via the regulation of gene expression. Of the regulators of transcription that play a role in this response, several alternative sigma factors of M. tuberculosis have been shown to control gene expression in response to stresses, and some of these are required for virulence or persistence in vivo. For this study, we examined the role of the alternative sigma factor SigD in M. tuberculosis gene expression and virulence. Using microarray analysis, we identified several genes whose expression was altered in a strain with a sigD deletion. A small number of these genes, including sigD itself, the gene encoding the autocrine growth factor RpfC, and a gene of unknown function, Rv1815, appear to be directly regulated by this sigma factor. By identifying the in vivo promoters of these genes, we have determined a consensus promoter sequence that is putatively recognized by SigD. The expression of several genes encoding PE-PGRS proteins, part of a large family of related genes of unknown function, was significantly increased in the sigD mutant. We found that the expression of sigD is stable throughout log phase and stationary phase but that it declines rapidly with oxygen depletion. In a mouse infection model, the sigD mutant strain was attenuated, with differences in survival and the inflammatory response in the lung between mice infected with the mutant and those infected with the wild type.

Anaerobiosis↗
PubMed2003

Characterization of a divergent non-classical MHC class I gene in sharks.

Sharks are the most ancient group of vertebrates known to possess members of the major histocompatibility complex (MHC) gene family. For this reason, sharks provide a unique opportunity to gain insight into the evolution of the vertebrate immune system through comparative analysis. Two genes encoding proteins related to the MHC class I gene family were isolated from splenic cDNA derived from spiny dogfish shark ( Squalus acanthias). The genes have been designated MhcSqac-UAA*01 and MhcSqac-UAA*NC1. Comparative analysis demonstrates that the Sqac-UAA*01 protein sequence clusters with classical MHC class I of several shark species and has structural elements common to most classical MHC class I molecules. In contrast, Sqac-UAA*NC1 is highly divergent from all vertebrate classical MHC class I proteins, including the Sqac-UAA *01 sequence and those of other shark species. Although Sqac-UAA*NC1 is clearly related to the MHC class I gene family, no orthologous genes from other species were identified due to the high degree of sequence divergence. In fact, the Sqac NC1 protein sequence is the most divergent MHC class-I-like protein identified thus far in any shark species. This high degree of divergence is similar in magnitude to some of the MHC class-I-related genes found in mammals, such as MICA or CD1. These data support the existence of a class of highly divergent non-classical MHC class I genes in the most primitive vertebrates known to possess homologues of the MHC and other components of the adaptive immune system.

Amino Acid Sequence↗
PubMed2003

Epidermal Langerhans cells efficiently mediate CD1a-dependent presentation of microbial lipid antigens to T cells.

Langerhans cells are a critical component of skin immunity, capable of capturing protein antigens in the epidermis and presenting them to specific T cells in the context of major histocompatibility complex class II molecules. Recently, a major histocompatibility complex independent pathway of lipid antigen presentation has been identified and is mediated by molecules of the CD1 family (CD1a, CD1b, CD1c, and CD1d). Because Langerhans cells are professional antigen-presenting cells and express CD1a molecules prominently, we hypothesized that Langerhans cells might play a role in T cell responses directed against not only peptide antigens but also lipid antigens. Here, we show that freshly isolated immature Langerhans cells as well as mature Langerhans cells that have migrated from the epidermis are efficient in presenting foreign microbial lipid antigens to specific T cells whereas dermal dendritic cells express much less CD1a molecules and function inefficiently. Further, we found that Langerhans cells migrating from epidermal sheets that were exposed to microbial lipid antigens expressed lipid-antigen-loaded CD1a molecules on the cell surface, resulting in activation of specific T cells. These results underscore an outstanding ability of Langerhans cells to mediate CD1a-dependent lipid antigen presentation. Thus, Langerhans-cell-mediated skin immunity may involve T cell recognition of both peptide and lipid antigens.

Antigen Presentation↗
PubMed2003

Immunization with a mycobacterial lipid vaccine improves pulmonary pathology in the guinea pig model of tuberculosis.

Lipids and glycolipid molecules derived from Mycobacterium tuberculosis can be presented to T cells by CD1 cell-surface molecules in humans. These lipid-specific T cells are cytolytic, secrete pro-inflammatory cytokines and have bactericidal activity. Here, we describe studies in which lipids from M. tuberculosis were incorporated into liposomes with adjuvant and tested as vaccines in a guinea pig aerosol tuberculosis challenge model. Animals vaccinated with mycobacterial lipids showed reduced bacterial burdens in the lung and spleen at 4 weeks after infection. In addition, the lungs of lipid-vaccinated animals also had significantly less pathology, with granulomatous lesions being smaller and more lymphocytic. In contrast, animals receiving only vehicle control immunizations had granulomatous lesions that were larger and often contained caseous necrotic centers. Quantification of histopathology by morphometric analysis revealed that the overall percentage of lung occupied by diseased tissue was significantly smaller in lipid-vaccinated animals as compared to vehicle control animals. In addition, the mean area of individual granulomatous lesions was found to be significantly smaller in both lipid- and bacillus Calmette-Guerin-vaccinated guinea pigs. These data support an important role for lipid antigens in the immune response to M. tuberculosis infection, potentially through the generation of CD1-restricted T cells. Immunogenic lipids thus represent a novel class of antigens that might be included to enhance the protective effects of subunit vaccine formulations.

Adjuvants, Immunologic↗
PubMed2003

Regulation of tumor necrosis factor alpha gene expression by mycobacteria involves the assembly of a unique enhanceosome dependent on the coactivator proteins CBP/p300.

Tumor necrosis factor alpha (TNF-alpha) plays an important role in host containment of infection by Mycobacterium tuberculosis, one of the leading causes of death by an infectious agent globally. Using the pathogenic M. tuberculosis strain H37Rv, we present evidence that upon stimulation of monocytic cells by M. tuberculosis a unique TNF-alpha enhanceosome is formed, and it is distinct from the TNF-alpha enhanceosome that forms in T cells stimulated by antigen engagement or virus infection. A distinct set of activators including ATF-2, c-jun, Ets, Sp1, Egr-1 and the coactivator proteins CBP/p300 are recruited to the TNF-alpha promoter after stimulation with M. tuberculosis. Furthermore, the formation of this enhanceosome is dependent on inducer-specific helical phasing relationships between transcription factor binding sites. We also show that the transcriptional activity of CBP/p300 is potentiated by mycobacterial stimulation of monocytes. The identification of TNF-alpha regulatory elements and coactivators involved in M. tuberculosis-stimulated gene expression thus provides potential selective molecular targets in the modulation of TNF-alpha gene expression in the setting of mycobacterial infection.

Activating Transcription Factor 2↗
PubMed2003

CD1 antigen presentation and infectious disease.

Taken together, the data generated thus far strongly suggest that CD1 plays a role in the immune response against various infections (table 1). For obvious reasons, the data gathered thus far using model infection systems have focused primarily on the mouse and therefore only examine the role of CD1d. This leaves an important gap in our understanding of the CD1 antigen presentation pathway given the potential role of CD1a, CD1b and CD1c for contributing to antimicrobial immunity. The functional dichotomy between group 1 and group 2 CD1 isoforms obviously requires further analysis. However, we propose that the group 1 CD1 (CD1a, CD1b, CD1c) antigen presentation pathway is closer to the traditional adaptive immune response mechanisms with the capacity to present unique foreign antigens to specific T cells. This broadens the universe antigens that T cells can use to target pathogens and provides important antimicrobial effector mechanisms that may be critical for combating some types of infections. Lipid antigens may also provide a more effective means of targeting intracellular pathogens by T cells since CD1 is able to sample almost all of the intracellular reservoirs that are exploited by this class of pathogen and may provide an important component of the cytotoxic T cell response [80]. On the other hand, the group 2 CD1 protein (CD1d) may be more intermediate in terms of lying functionally between the innate and adaptive immune systems. The activation of CD1d-restricted T cells may, therefore, help bridge the temporal gap between the onset of innate immunity and the purely adaptive responses typified by the MHC-restricted T cells. Hence, the CD1d-restricted [table: see text] T cells are primed for rapid high-level cytokine release. In addition, the interaction of CD1d-restricted T cells with CD1d on DCs can trigger the release of IL-4 and GM-CSF to promote maturation of tissue-resident DC at the site of infection. The maturation of tissue DC would lead to migration of the activated DC to regional lymph nodes and initiation of MHC-restricted T cell responses. Subsequent IL-12 production by the DC in response to CD1d-mediated T cell stimulation could then drive IFN-gamma production by CD1d-restricted T cells and influence the polarization of the T cell response to infection. In addition, early bursts of IFN-gamma by CD1d-restricted T cells could also upregulate antimicrobial activity in macrophages and activate other important effector cells such as NK cells prior to MHC-restricted T cell responses. In the constant struggle between the microbial pathogen and its host, the evolutionary balance almost always favors the microbe. The rapid rate of evolution and adaptation of the microbe accounts for most of this advantage. Hence, it is not surprising that the host immune system has evolved a complex set of pathways, in addition to the MHC, that are able to recognize and target the unique molecular signatures of infectious microorganisms. The lipid antigens presented by CD1 add to this array and thus provide a further layer of immune defense to the host for combating pathogens.

Animals↗
PubMed2002

Conservation of CD1 intracellular trafficking patterns between mammalian species.

Dendritic cells (DC) are potent APCs that sample Ags from the surrounding environment and present them to naive T cells using cell surface Ag-presenting molecules. The DC in both lymphoid and nonlymphoid tissues express high levels of CD1, a cell surface glycoprotein capable of presenting lipids and glycolipids to T cells. Distinct group 1 CD1 isoforms (CD1a, -b, -c) in man are known to traffic to different parts of the endocytic system where microbial Ags may be sampled. Guinea pigs are the only known rodent species that express the group 1 CD1 proteins. Therefore, we examined the expression and trafficking of guinea pig CD1 (gpCD1) isoforms on isolated DC. Confocal microscopy using mAbs specific for individual gpCD1 isoforms revealed differential trafficking of two distinct CD1b isoforms within DC. Colocalization of MHC class II was observed with the gpCD1b1 isoform, consistent with localization in the late endosomes of DC. In contrast, the gpCD1b3 isoform lacks an endosomal sorting motif and remains on the cell surface. Following incubation with Mycobacterium tuberculosis lipoarabinomannan, colocalization of endocytosed lipoarabinomannan with the gpCD1b1 isoform was observed but not with the gpCD1b3 isoform, which remained primarily on the cell surface. These data demonstrate that guinea pig DC express CD1 isoforms with unique trafficking patterns that recapitulate the patterns seen for human CD1 isoforms. This suggests evolutionary pressure for a conserved mechanism in mammals that allows CD1 to sample lipid Ags from various subcompartments of the endocytic system.

Animals↗
PubMed2002

Induction of CD1-restricted immune responses in guinea pigs by immunization with mycobacterial lipid antigens.

Group 1 CD1 molecules have been shown to present lipid and glycolipid Ags of mycobacteria to human T cells. However, a suitable animal model for the investigation of this component of antimycobacterial immunity has not yet been established. Previously, we found that guinea pigs express multiple isoforms of group 1 CD1 proteins that are homologous to human CD1b and CD1c. In this study, we show that CD1-restricted T cell responses can be generated in guinea pigs following immunization with lipid Ags from Mycobacterium tuberculosis. Splenic T cells from lipid Ag-immunized guinea pigs showed strong proliferative responses to total lipid Ags and partially purified glycolipid fractions from M. tuberculosis. These lipid Ag-reactive T cells were enriched in CD4-negative T cell fractions and showed cytotoxic activity against CD1-expressing guinea pig bone marrow-derived dendritic cells pulsed with M. tuberculosis lipid Ags. Using guinea pig cell lines transfected with individual CD1 isoforms as target cells in cytotoxic T cell assays, we found that guinea pig CD1b and CD1c molecules presented M. tuberculosis glycolipid Ags to T cells raised by mycobacterial lipid immunization. These results were confirmed using a T cell line derived from M. tuberculosis lipid Ag-immunized guinea pigs, which also showed CD1-restricted responses and cytolytic activity. Our results demonstrate that CD1-restricted responses against microbial glycolipid Ags can be generated in vivo by specific immunization and provide support for the use of the guinea pig as a relevant small animal model for the study of CD1-restricted immune responses to mycobacterial pathogens.

Animals↗