PubMed Health⌕ Search

Biomedical subjects

D Askew

Publications and source records attributed to D Askew.

At least 19 recordsLinked to original sources
PubMed2001

Systemic deficits in transporter for antigen presentation (TAP)-1 or proteasome subunit LMP2 have little or no effect on tumor incidence.

Some tumor cells have deficits in class I MHC antigen processing, suggesting that T cells exert selective pressure on tumor cells. Previous studies have not revealed increased tumor incidence in mice with deficits in T-cell immunity, including mice lacking TAP1 (a subunit of the transporter for antigen presentation) or LMP2 (a regulated subunit of the 20S proteasome). The incidence of spontaneous tumors in these mice, however, is too low to assess differences in host resistance to tumors. To increase tumor incidence and better assess the role of systemic expression of TAP1 and LMP2 in responses to tumors, TAP1-/- and LMP2-/- mice were bred with p53-/- mice to create TAP1-/-p53-/- and LMP2-/-p53-/- double knockout mice. Lymphomas and sarcomas (malignant fibrous histiocytoma and angiosarcoma) occurred with high incidence in all p53-deficient populations. Tumor incidence and death rate were similar in TAP1-/-p53-/- mice and closely matched control TAP1+/+p53-/- mice. Tumor incidence and death rate were slightly accelerated in LMP2-/-p53-/- mice relative to control LMP2+/+p53-/- mice, but the biological significance of this difference was unclear. The relative incidence of lymphomas vs. sarcomas was not significantly altered by variation in TAP1 or LMP2. In conclusion, systemic absence of TAP1 did not alter tumor incidence, while absence of LMP2 was associated with only a slight acceleration of tumor incidence of uncertain significance. These observations are consistent with other evidence that normal T-cell responses do not effectively limit tumorigenesis. Even though T cells can attack some tumor cells, the ability of tumors to alter their immunogenicity and evade T-cell surveillance may render the native immune system ineffective at providing a rate-limiting barrier to tumorigenesis and preventing cancer.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2000

CpG DNA induces maturation of dendritic cells with distinct effects on nascent and recycling MHC-II antigen-processing mechanisms.

Murine bone marrow cultured with GM-CSF produced dendritic cells (DCs) expressing MHC class II (MHC-II) but little CD40, CD80, or CD86. Oligodeoxynucleotides (ODN) containing CpG motifs enhanced DC maturation, increased MHC-II expression, and induced high levels of CD40, CD80, and CD86. When added with Ag to DCs for 24 h, CpG ODN enhanced Ag processing, and the half-life of peptide:MHC-II complexes was increased. However, Ag processing was only transiently enhanced, and exposure of DCs to CpG ODN for 48 h blocked processing of hen egg lysozyme (HEL) to HEL(48-61):I-A(k) complexes. Processing of this epitope required newly synthesized MHC-II and was blocked by brefeldin A (BFA), suggesting that reduced MHC-II synthesis could explain decreased processing. Real-time quantitative PCR confirmed that CpG ODN decreased I-A(beta)(k) mRNA in DCs. In contrast, RNase(42-56):I-A(k) complexes were generated via a different processing mechanism that involved recycling MHC-II and was partially resistant to BFA. Processing of RNase(42-56):I-A(k) persisted, although at reduced levels, after CpG-induced maturation of DCs, and this residual processing by mature DCs was completely resistant to BFA. Changes in endocytosis, which was transiently enhanced and subsequently suppressed by CpG ODN, may affect Ag processing by both nascent and recycling MHC-II mechanisms. In summary, CpG ODN induce DC maturation, transiently increase Ag processing, and increase the half-life of peptide-MHC-II complexes to sustain subsequent presentation. Processing mechanisms that require nascent MHC-II are subsequently lost, but those that use recycling MHC-II persist even in fully mature DCs.

Adjuvants, Immunologic↗
PubMed1999

CpG oligodeoxynucleotides down-regulate macrophage class II MHC antigen processing.

Unmethylated CpG motifs in bacterial DNA or short oligodeoxynucleotides (ODN) stimulate cells of the immune system and provide adjuvant activity. CpG DNA directly activates macrophages to secrete IL-12 and TNF-alpha and increases transcription of various genes, but its effects on macrophage Ag processing remain uncertain. The effects of CpG ODN on class II MHC (MHC-II) Ag processing and presentation were examined using peritoneal macrophages that were cultured for 18 h with CpG ODN and then pulsed with protein Ags. T cell hybridomas were used to detect presentation of specific peptide:MHC-II complexes. Both CpG ODN and LPS inhibited processing of bovine RNase and hen egg lysozyme. Presentation of exogenous peptides was inhibited to a lesser degree. Treatment of macrophages for 18 h with CpG ODN decreased surface MHC-II expression, as measured by flow cytometry. Furthermore, Northern blot analysis revealed that treatment with CpG ODN decreased I-Ak mRNA. Endocytosis by macrophages, as measured by uptake of fluorescent dextran, was not altered by treatment with CpG ODN. The inhibitory effect of CpG ODN on Ag processing was seen after prolonged (18 h) treatment of macrophages, but not after short treatment (e.g., 2 h) with CpG ODN and protein Ag. Enhancement of macrophage Ag processing was not seen at any time point of CpG ODN exposure, in contrast to data from other studies with dendritic cells. In summary, exposure of macrophages to CpG ODN results in a decrease in macrophage Ag processing and presentation, which is largely mediated by a decrease in synthesis of MHC-II molecules.

Adjuvants, Immunologic↗
PubMed1999

Phagocytic antigen processing and effects of microbial products on antigen processing and T-cell responses.

Processing of exogenous antigens and microbes involves contributions by multiple different endocytic and phagocytic compartments. During the processing of soluble antigens, different endocytic compartments have been demonstrated to use distinct antigen-processing mechanisms and to process distinct sets of antigenic epitopes. Processing of particulate and microbial antigens involves phagocytosis and functions contributed by phagocytic compartments. Recent data from our laboratory demonstrate that phagosomes containing antigen-conjugated latex beads are fully competent class II MHC (MHC-II) antigen-processing organelles, which generate peptide:MHC-II complexes. In addition, phagocytosed antigen enters an alternate class I MHC (MHC-I) processing pathway that results in loading of peptides derived from exogenous antigens onto MHC-I molecules, in contrast to the cytosolic antigen source utilized by the conventional MHC-I antigen-processing pathway. Antigen processing and other immune response mechanisms may be activated or inhibited by microbial components to the benefit of either the host or the pathogen. For example, antigen processing and T-cell responses (e.g. Th1 vs Th2 differentiation) are modulated by multiple distinct microbial components, including lipopolysaccharide, cholera toxin, heat labile enterotoxin of Escherichia coli, DNA containing CpG motifs (found in prokaryotic and invertebrate DNA but not mammalian DNA) and components of Mycobacterium tuberculosis.

Animals↗
PubMed1998

Mouse resident microglia: isolation and characterization of immunoregulatory properties with naïve CD4+ and CD8+ T-cells.

We describe a non-enzymatic procedure designed to isolate in high purity resident microglia from the brains of normal mice. This procedure allowed for the characterization of the cells without concern that their surface features had been enzymatically altered during tissue processing. A cell population was obtained and judged to consist primarily of microglia because essentially all the cells were Mac-1+, Mac-3+, F4/80+, CD44+, CD54+, and CD86+, and they expressed CD45 with a mean fluorescence intensity value of about one-half that of tissue macrophages. The cells also expressed marginal levels of MHC class I, CD14, CD40, and CD80, but lacked detectable MHC class II, CD4, CD8, CD45R, and CD102 molecules. Molecular phenotyping revealed that the purified microglial population contained mRNA transcripts encoding the receptor for colony stimulating factor-1 (CSF-1), the macrophage growth factor, and contained few, if any, transcripts for glial fibrillary acidic protein, an astrocyte-specific marker. Ex vivo, the microglia constitutively stimulated, in the mixed leukocyte reaction, the proliferation of naive allogeneic CD8+, but not CD4+ T-cells. However, they failed to present a protein antigen to naive antigen-specific CD4+ T-cells unless pretreated with interferon-gamma, a response that was inhibited by antibodies to CD86. Agar-cloning experiments confirmed that normal mouse brain contains a CSF-1-responsive cell that gave rise to cells with identical immunophenotypic characteristics as freshly isolated resident microglia. Moreover, the microglial progenitor cell located in a density fraction that was different from that containing the mature resident microglia.

Animals↗
PubMed1997

The genes encoding the peripheral cannabinoid receptor and alpha-L-fucosidase are located near a newly identified common virus integration site, Evi11.

A new common region of virus integration, Evi11, has been identified in two retrovirally induced murine myeloid leukemia cell lines, NFS107 and NFS78. By interspecific backcross analysis, it was shown that Evi11 is located at the distal end of mouse chromosome 4, in a region that shows homology with human 1p36. The genes encoding the peripheral cannabinoid receptor (Cnr2) and alpha-L-fucosidase (Fuca1) were identified near the integration site by using a novel exon trapping system. Cnr2 is suggested to be the target gene for viral interference in Evi11, since proviruses are integrated in the first intron of Cnr2 and retroviral integrations alter mRNA expression of Cnr2 in NFS107 and NFS78. In addition, proviral integrations were demonstrated within the 3' untranslated region of Cnr2 in five independent newly derived CasBrM-MuLV (mouse murine leukemia virus) tumors, CSL13, CSL14, CSL16, CSL27, and CSL97. The Cnr2 gene encodes a seven-transmembrane G-protein-coupled receptor which is normally expressed in hematopoietic tissues. Our data suggest that the peripheral cannabinoid receptor gene might be involved in leukemogenesis as a result of aberrant expression of Cnr2 due to retroviral integration in Evi11.

3T3 Cells↗
PubMed1996

Alloantigen presentation to naive CD8+ T cells by mouse microglia: evidence for a distinct phenotype based on expression of surface-associated and soluble costimulatory molecules.

We previously showed that approximately one-third of mouse primary microglial clones derived from individual precursor cells residing in normal brain constitutively present alloantigens (alloAgs) to naive CD8+ T cells (Moore et al.: J Neuroimmunol 41:203, 1992). To understand the basis for this alloAg presenting (alloAgP) activity, we developed a panel of microglial cell lines that were characterized by patterns of alloAgP activity similar to that of the primary clones. Flow cytometric analysis revealed that microglia with and without alloAgP activity expressed similar levels of major histocompatibility complex class I molecules; however, CD80 (B7-1) and CD86 (B7-2) expression was primarily restricted to the alloAgP- cell lines. Monoclonal antibody (Mab) to CD80 only partially blocked the proliferative response of allogeneic CD8+ T cells cocultured with the presenting cell lines, whereas Mab to CD86 completely inhibited the response, indicating a significant role for this molecule in T-cell activation. Using an immunoassay, recombinant mouse cytokines, cytokine-specific Mabs, and the reverse transcriptase-polymerase chain reaction to detect specific cytokine mRNAs, we found the synthesis of interleukin (IL)-1 alpha, IL-6, IL-12, and tumor necrosis factor-alpha (TNF-alpha) to be restricted to the alloAgP- cell lines. Costimulatory roles were then identified for these molecules. We conclude that the ability to present alloAg is a property of a subset of microglia that constitutively express CD86 and secrete costimulatory cytokines that promote the expansion of the alloAg-stimulated CD8+ T cells.

Animals↗
PubMed1995

Mouse microglial cell lines differing in constitutive and interferon-gamma-inducible antigen-presenting activities for naive and memory CD4+ and CD8+ T cells.

We developed a panel of non-virus transformed cell lines derived from individual microglial precursors residing in the brains of normal mice. These colony stimulating factor-1-dependent cell lines are B7-1+ (CD80), Mac-1+, Mac-2+, Mac-3+, CD45+, MHC class I+, colony stimulating factor-1 receptor+, and they ingest antibody-coated particles. However, the cell lines differ in their expression of B7-2 (CD86), F4/80, Ly-6C and MHC class II molecules. They also differ in their ability to constitutively process and present antigens to naive CD4+ and CD8+ T cells, memory CD4+ and CD8+, and in the manner by which interferon gamma modulates their antigen-presenting activities. These cell lines should be valuable as models for studies on the immunobiology of the microglia.

Adjuvants, Immunologic↗
PubMed1995

A subset of splenic macrophages process and present native antigen to naive antigen-specific CD4+ T-cells from mice transgenic for an alpha beta T-cell receptor.

The progeny of individual macrophage precursors from mouse spleen were examined for their ability to constitutively process and present native pigeon cytochrome c or a peptide fragment of this antigen to naive CD4+ T-cells from mice transgenic for a V alpha 11/V beta 3 TCR that recognizes an epitope in the antigen fragment. The results show that constitutive Ag processing and presentation is a stable characteristic restricted to the progeny of approximately 20% of splenic macrophage precursors. This property does not appear to be randomly acquired, but to reflect the ability of certain macrophages to produce IL-12.

Animals↗
PubMed1995

Relative contributions of voluntary apnoea, exposure to cold and face immersion in water to diving bradycardia in humans.

1. Diving or face immersion bradycardia is a well recognized but incompletely understood reflux which occurs in man and other mammals. 2. In order to investigate the contributions made by voluntary apnoea, face immersion in water and cold exposure, 18 normal subjects were exposed to these challenges separately and in various combination. 3. Tested individually, cold and apnoea caused significant reductions in heart beat (P < 0.01 and 0.002, respectively). Face immersion in thermoneutral water had no effect on heart rate. 4. The bradycardic effect of apnoea at maximal inspiration may be due to stimulation of pulmonary stretch receptors. 5. Cold exposure and voluntary apnoea applied simultaneously caused a summative effect but when tested with face immersion in water there was a synergistic response greater than the sum of individual responses. 6. The results confirm the bradycardic effect of apnoea and cold exposure, whereas immersion in thermoneutral water had little effect, a finding which has been disputed in the literature.

Adult↗
PubMed1994

Macrophage priming and activation during fibrosarcoma growth: expression of c-myb, c-myc, c-fos, and c-fms.

Macrophages (M phi)3 function by a two-step process that includes priming (induction of cytokine and enzyme mRNA) and activation (production of effector molecules). The initial steps in M phi priming involve the expression of certain proto-oncogenes that regulate expression of other genes. Because tumor growth primes M phi to produce several suppressor monokines, we determined if cancer induced M phi expression of these proto-oncogenes. Unstimulated peritoneal M phi from tumor-bearing hosts (TBH) constitutively expressed the proto-oncogenes c-fms, c-fos, c-myc, and c-myb, whereas normal host (NH) M phi had little or no expression of these proto-oncogenes. When M phi were given a 24-h adherence priming stimulus, NH M phi expressed c-fms and c-fos at levels equivalent to TBH M phi constitutive expression. Adherence had little or no effect on c-fms and c-fos expression in TBH M phi or on NH and TBH M phi c-myc expression. c-myb expression was not induced in NH M phi during adherence and was strongly decreased in TBH M phi. Activation with a 1-h lipopolysaccharide-treatment increased NH and TBH M phi expression of c-fms, c-fos, and c-myc, with higher expression of these proto-oncogenes in TBH M phi. Activation failed to induce c-myb expression in NH M phi and completely inhibited expression in TBH M phi. Because c-fms, c-fos, and c-myc are normally expressed early during M phi activation, our results suggest that tumor growth primes M phi by inducing expression of these proto-oncogenes. c-myb is expressed in immature M phi and is downregulated during M phi activation. These observations explain why NH M phi expression of c-myb was not induced and are consistent with reports that suggest TBH M phi have not reached full developmental maturity. The induction of M phi proto-oncogene expression during cancer may put M phi in a primed state, which leads to earlier and stronger production of adverse suppressor and cytotoxic molecules.

Animals↗
PubMed1993

Alloantigen presentation by individual clones of mouse splenic macrophages. Selective expression of IL-1 alpha in response to CD8+ T cell-derived IFN-gamma defines the alloantigen-presenting phenotype.

Approximately one-third of mouse splenic macrophage (M theta) progenitors yield progeny that constitutively present MHC class I alloantigen to naive T cells, a response that is restricted to CD8+ T cells and is elicited in a CD4+ Th cell-independent manner. In addition, both the alloantigen-presenting (alloAP+) and nonpresenting (alloAP-) M theta subsets constitutively express similar levels of MHC class I molecules, and their alloAP phenotypes are unaffected by IFN-gamma, which enhances the expression of both class I and II MHC molecules. We therefore postulated the restricted expression of costimulator molecules to account for the alloAP+ phenotype. Using cytokine-specific antibodies, recombinant mouse cytokines, and polymerase chain reaction analyses (to detect specific cytokine mRNA transcripts), we identified the putative costimulators as IL-1 alpha, IL-6, and TNF-alpha. TNF-alpha transcripts were present in both the alloAP+ and alloAP- M theta subsets, but IL-1 alpha and IL-6 were not constitutively expressed by the alloAP+ subset of M theta; rather, they were induced by IFN-gamma, which was released from naive CD8+ T cells only during coculture with alloAP+ M theta. Although IFN-gamma induced IL-6 gene transcription in both alloAP+ and alloAP- M theta subsets, it induced IL-1 alpha transcripts only in the alloAP+ subset. Finally, CD8+ T cells exposed to alloAP- M theta were unresponsive when subsequently cultured with alloAP+ M theta. We conclude that the ability of some M theta to elicit IFN-gamma from CD8+ T cells and to respond to this cytokine by producing IL-1 alpha defines the alloAP phenotype of the cell population, and that alloAP- M theta induce a state of alloantigen-specific tolerance in naive CD8+ T cells.

Animals↗
PubMed1993

Tumor growth and adherence change the expression of macrophage Mac-2.

The galactose-specific animal lectin, Mac-2, has been identified in macrophage (M phi) membrane, cytoplasmic, and nuclear fractions. Flow cytometric analyses showed that there is a decrease in membrane Mac-2 during tumor growth. After 24-h adherence there was an increase in the number of normal host (NH) and tumor-bearing host (TBH) Mac-2+ M phi. Immunoblot analyses of NH and TBH M phi identified changes in the subcellular localization of Mac-2. The increase in nuclear Mac-2 during tumor growth, and after prolonged adherence of NH and TBH M phi, correlates with an increase in M phi entering the late G1 phases of the cell cycle. Northern blot analyses showed an increase in Mac-2 mRNA during tumor growth, and an increase in NH and TBH M phi after 24-h adherence. Tumor growth is able to manipulate the immune system through M phi by causing a down-regulation in membrane Mac-2 and an up-regulation in intracellular Mac-2. NH and TBH M phi respond to adherence by expressing increased membrane and nuclear Mac-2, but TBH M phi response is lower.

Animals↗
PubMed1993

Fibrosarcoma-induced increase in macrophage tumor necrosis factor alpha synthesis suppresses T cell responses.

Tumors down-regulate T cell responses partly by increasing macrophage (m phi) production of the suppressive molecule prostaglandin E2 (PGE2). Because tumor growth increases m phi tumor necrosis factor alpha (TNF-alpha) production and TNF-alpha stimulates m phi PGE2 synthesis, we examined the contribution of TNF-alpha to fibrosarcoma-induced m phi-mediated suppression of alloreactive CD4+ T cell proliferation. We showed that tumor-bearing host (TBH) m phi s express high levels of TNF-alpha mRNA, which leads to increased lipopolysaccharide-induced TNF-alpha production. Tumor cells were directly involved in m phi TNF-alpha synthesis because fibrosarcoma cells induced normal host (NH) m phi s to produce TNF-alpha. Addition of TBH m phi s to allogeneic mixed lymphocyte reaction (MLR) cultures suppressed CD4+ T cell proliferation more than NH m phi s. The neutralization of endogenous TNF-alpha activity with anti-TNF-alpha antibody (Ab) treatment reversed TBH, but not NH, m phi-mediated suppression. Conversely, exogenous TNF-alpha increased NH or TBH m phi-mediated suppression but stimulated T cell proliferation without m phi s. Kinetic treatment of MLR cultures with anti-TNF-alpha Ab or TNF-alpha showed that TNF-alpha production and activity occurred at the beginning of T cell proliferation. When arachidonic acid metabolite synthesis was inhibited, TNF-alpha-induced suppression was blocked in NH m phi-containing cultures and completely reversed in TBH m phi-containing cultures. A PGE2-specific enzyme-linked immunosorbent assay showed that TNF-alpha addition increased PGE2 production in NH m phi-containing cultures to that of TBH m phi-containing cultures. Exogenous PGE2 did not affect the TNF-alpha enhancement of T cell proliferation without m phi s. Therefore, suppression induced by TNF-alpha was caused by increased m phi PGE2 production and not by TNF-alpha in concert with PGE2. Even though TNF-alpha is known to enhance lymphocyte proliferation, we show that in the presence of m phi s, the main TNF-alpha producers, TNF-alpha suppresses T cell proliferation. Perhaps increased TNF-alpha production during pathological states, such as cancer, triggers the initial stages of suppression.

Animals↗
PubMed1993

Tumor-induced modulation of macrophage class II MHC molecule mRNA expression.

Class II MHC protein expression in macrophages (M phi) is reduced during tumor growth. Because regulation of class II MHC proteins occurs during transcription, tumor growth may suppress class II MHC protein expression by suppressing mRNA. The decrease in class II mRNA may result from (i) a decrease in M phi responsiveness to an inducing agent, such as interferon-gamma (IFN-gamma), or (ii) an increase in M phi sensitivity to suppressing agents, such as prostaglandin E2 (PGE2). To determine how tumors induce suppression of class II mRNA, M phi were cultured in the presence of IFN-gamma with or without other factors, and Northern blot analyses were performed. Unstimulated normal host (NH) or tumor-bearing host (TBH) M phi do not express detectable class II mRNA. The addition of IFN-gamma induces class II mRNA expression in NH and TBH M phi, but class II mRNA expression is significantly lower in TBH M phi. Kinetic studies suggested that NH M phi class II mRNA is induced faster and in greater amounts than TBH M phi class II mRNA. There is a decrease in M phi class II mRNA stability during tumor growth that may account for the decreased induction by IFN-gamma. Lipopolysaccharide (LPS) suppresses class II mRNA induction in both NH and TBH IFN-gamma-treated M phi, but TBH M phi are more sensitive to its suppression. PGE2 and tumor-necrosis factor-alpha (TNF-alpha), two factors produced by LPS-stimulated M phi, were tested for their ability to modulate class II mRNA expression in NH and TBH IFN-gamma-treated M phi. PGE2 suppressed class II mRNA expression in both NH and TBH M phi. The addition of TNF-alpha to IFN-gamma-treated M phi suppressed class II mRNA in NH M phi but, surprisingly, had an additive effect on IFN-gamma-induced class II mRNA expression. TNF-alpha did not induce class II mRNA expression in TBH M phi in the absence of IFN-gamma. The cause of the reduced class II mRNA expression during tumor growth is a decreased response to IFN-gamma and an increased sensitivity to PGE2. This change may cause the observed suppression mediated by TBH M phi.

Animals↗
PubMed1991

Functional dissection of the paired segmentation gene in Drosophila embryos.

An ectopic expression assay in Drosophila embryos was used to investigate the roles of pair-rule segmentation genes in the spatial regulation of the segment-polarity gene, engrailed (en). It is hypothesized that the regions of overlap in expression of two genes, paired (prd) and even-skipped (eve), define the odd-numbered en expression stripes. Consistent with this combinatorial model, ectopic expression of prd caused these en stripes to be expanded posteriorly. Surprisingly, however, ectopic expression of a prd gene with a deletion of the conserved paired box resulted in loss of these odd-numbered en stripes. This dominant negative effect is a phenocopy of en expression in prd embryos and suggests that the paired box is necessary for normal prd- function. A similar deletion of odd-numbered en stripes was also observed after ectopic expression of a chimeric fushi tarazu (ftz) gene containing a substituted prd gene homeo box; in addition, in these embryos, the even-numbered en stripes were expanded anteriorly, as observed when the unaltered ftz gene is ectopically expressed. These effects suggest that the chimeric protein may have DNA or protein targets of both the normal Ftz and Prd proteins.

Animals↗