Biomedical subjects

C A Hunter

Publications and source records attributed to C A Hunter.

At least 19 recordsLinked to original sources

PubMed2001

Cutting edge: systemic inhibition of angiogenesis underlies resistance to tumors during acute toxoplasmosis.

The ability of various infections to suppress neoplastic growth has been well documented. This phenomenon has been traditionally attributed to infection-induced concomitant, cell-mediated antitumor immunity. We found that infection with Toxoplasma gondii effectively blocked neoplastic growth of a nonimmunogenic B16.F10 melanoma. Moreover, this effect was independent of cytotoxic T or NK cells, production of NO by macrophages, or the function of the cytokines IL-12 and TNF-alpha. These findings suggested that antitumor cytotoxicity was not the primary mechanism of resistance. However, infection was accompanied by strong, systemic suppression of angiogenesis, both in a model system and inside the nascent tumor. This suppression resulted in severe hypoxia and avascular necrosis that are incompatible with progressive neoplastic growth. Our results identify the suppression of tumor neovascularization as a novel mechanism critical for infection-induced resistance to tumors.

Acute Disease↗

PubMed2001

A role for CD44 in the production of IFN-gamma and immunopathology during infection with Toxoplasma gondii.

The interaction of activated CD44 with its ligand, low m.w. hyaluronan, is involved in inflammation, but no role has been identified for this interaction in the regulation of an immune response to infection. In these studies, infection of C57BL/6 mice with Toxoplasma gondii resulted in increased expression of CD44 on T cells, B cells, NK cells, and macrophages, and a small percentage of CD4(+) T cells express an activated form of CD44. Administration of anti-CD44 to infected mice prevented the development of a CD4(+) T cell-dependent, infection-induced inflammatory response in the small intestine characterized by the overproduction of IFN-gamma. The protective effect of anti-CD44 treatment was associated with reduced production of IFN-gamma, but not IL-12, in vivo and in vitro. Furthermore, the addition of low m.w. hyaluronan to cultures of splenocytes or purified CD4(+) T cells from infected mice resulted in the production of high levels of IFN-gamma, which was dependent on IL-12 and TCR stimulation. Together, these results identify a novel role for CD44 in the regulation of IFN-gamma production by CD4(+) T cells during infection and demonstrate a role for CD44 in the regulation of infection-induced immune pathology.

Adjuvants, Immunologic↗

PubMed2001

Bystander activation of CD8+ T cells contributes to the rapid production of IFN-gamma in response to bacterial pathogens.

The bacterium Burkholderia pseudomallei causes a life-threatening disease called melioidosis. In vivo experiments in mice have identified that a rapid IFN-gamma response is essential for host survival. To identify the cellular sources of IFN-gamma, spleen cells from uninfected mice were stimulated with B. pseudomallei in vitro and assayed by ELISA and flow cytometry. Costaining for intracellular IFN-gamma vs cell surface markers demonstrated that NK cells and, more surprisingly, CD8(+) T cells were the dominant sources of IFN-gamma. IFN-gamma(+) NK cells were detectable after 5 h and IFN-gamma(+) CD8(+) T cells within 15 h after addition of bacteria. IFN-gamma production by both cell populations was inhibited by coincubation with neutralizing mAb to IL-12 or IL-18, while a mAb to TNF had much less effect. Three-color flow cytometry showed that IFN-gamma-producing CD8(+) T cells were of the CD44(high) phenotype. The preferential activation of NK cells and CD8(+) T cells, rather than CD4(+) T cells, was also observed in response to Listeria monocytogenes or a combination of IL-12 and IL-18 both in vitro and in vivo. This rapid mechanism of CD8(+) T cell activation may be an important component of innate immunity to intracellular pathogens.

Animals↗

PubMed2001

Interleukin-10 does not contribute to the pathogenesis of a virulent strain of Toxoplasma gondii.

Interleukin (IL)-10 is an inhibitor of cell mediated immunity and an antagonist of the development of protective immune responses associated with resistance to T. gondii. These observations led to the hypothesis that the production of IL-10 could contribute to the ability of T. gondii to replicate and survive in an immune competent host. To determine whether the production of IL-10 affects the ability of the RH strain of T. gondii to cause a lethal infection in mice, we compared the immune response to RH in IL-10+/+ and IL-10-/- BALB/c mice. Both groups of mice produced comparable amounts of IL-12 and interferon (IFN)-gamma and had similar mortality curves and parasite burdens. The use of green fluorescent protein-labelled parasites allowed us to infect IL-10+/+ and IL-10-/- mice and use a fluorescence-activated cell sorter to distinguish infected and uninfected populations of macrophages and compare their expression of CD80, CD86 and major histocompatibility complex (MHC) class II. Although infected cells expressed higher overall levels of these molecules than uninfected cells, there were no differences between cells isolated from IL-10+/+ and IL-10-/- mice. Taken together, these results indicate that IL-10 is not required for the virulence of the RH strain of T. gondii, nor is it involved in the regulation of the CD80, CD86 and MHC class II molecules during RH-infection.

Animals↗

PubMed2001

Dichloroacetate improves cardiac efficiency after ischemia independent of changes in mitochondrial proton leak.

Dichloroacetate (DCA) is a pyruvate dehydrogenase activator that increases cardiac efficiency during reperfusion of ischemic hearts. We determined whether DCA increases efficiency of mitochondrial ATP production by measuring proton leak in mitochondria from isolated working rat hearts subjected to 30 min of ischemia and 60 min of reperfusion. In untreated hearts, cardiac work and efficiency decreased during reperfusion to 26% and 40% of preischemic values, respectively. Membrane potential was significantly lower in mitochondria from reperfused (175.6 +/- 2.2 mV) versus aerobic (185.8 +/- 3.1 mV) hearts. DCA (1 mM added at reperfusion) improved recovery of cardiac work (1.9-fold) and efficiency (1.5-fold) but had no effect on mitochondrial membrane potential (170.6 +/- 2.9 mV). At the maximal attainable membrane potential, O(2) consumption (nmol O(2) x mg(-1) x min(-1)) did not differ between untreated or DCA-treated hearts (128.3 +/- 7.5 and 120.6 +/- 7.6, respectively) but was significantly greater than aerobic hearts (76.6 +/- 7.6). During reperfusion, DCA increased glucose oxidation 2.5-fold and decreased H(+) production from glucose metabolism to 53% of untreated hearts. Because H(+) production decreases cardiac efficiency, we suggest that DCA increases cardiac efficiency during reperfusion of ischemic hearts by increasing the efficiency of ATP use and not by increasing the efficiency of ATP production.

Adenosine Triphosphate↗

PubMed2000

Identification of a role for NF-kappa B2 in the regulation of apoptosis and in maintenance of T cell-mediated immunity to Toxoplasma gondii.

The NF-kappaB family of transcription factors are involved in the regulation of innate and adaptive immune functions associated with resistance to infection. To assess the role of NF-kappaB(2) in the regulation of cell-mediated immunity, mice deficient in the NF-kappaB(2) gene (NF-kappaB(2)(-/-)) were challenged with the intracellular parasite Toxoplasma gondii. Resistance to this opportunistic pathogen is dependent on the production of IL-12, which is required for the development of innate NK cell and adaptive T cell responses dominated by the production of IFN-gamma necessary to control replication of this parasite. Although wild-type controls were resistant to T. gondii, NF-kappaB(2)(-/-) mice developed severe toxoplasmic encephalitis and succumbed to disease between 3 and 10 wk following infection. However, NF-kappaB(2) was not required for the ability of macrophages to produce IL-12 or to inhibit parasite replication and during the acute stage of infection, NF-kappaB(2)(-/-) mice had no defect in their ability to produce IL-12 or IFN-gamma and infection-induced NK cell responses appeared normal. In contrast, during the chronic phase of the infection, susceptibility of NF-kappaB(2)(-/-) mice to toxoplasmic encephalitis was associated with a reduced capacity of their splenocytes to produce IFN-gamma associated with a loss of CD4(+) and CD8(+) T cells. This loss of T cells correlated with increased levels of apoptosis and with elevated expression of the pro-apoptotic molecule Fas by T cells from infected NF-kappaB(2)(-/-) mice. Together, these results suggest a role for NF-kappaB(2) in the regulation of lymphocyte apoptosis and a unique role for this transcription factor in maintenance of T cell responses required for long-term resistance to T. gondii.

Animals↗

PubMed2000

Identification of STAT4-dependent and independent mechanisms of resistance to Toxoplasma gondii.

The capacity of IL-12 to stimulate T and NK cell production of IFN-gamma is required for resistance to Toxoplasma gondii. To identify the transcription factors involved in this mechanism of resistance, mice deficient in STAT4, a protein involved in IL-12 signaling, were infected with T. gondii and their immune responses were analyzed. STAT4-/- mice were unable to control parasite replication and died during the acute phase of infection, whereas wild-type mice controlled parasite replication and survived this challenge. The susceptibility of STAT4-/- mice to toxoplasmosis correlated with a defect in their ability to produce IFN-gamma in response to infection, whereas administration of IFN-gamma to these mice inhibited parasite replication and delayed time to death. Interestingly, analysis of infected STAT4-/- mice revealed that these mice did produce low levels of IFN-gamma during infection, and the ability of splenocytes from infected or uninfected STAT4-/- mice to produce IFN-gamma was enhanced by the addition of IL-2 plus IL-18. Moreover, administration of IL-2 plus IL-18 to STAT4-/- mice resulted in elevated serum levels of IFN-gamma associated with a decreased parasite burden and delayed time to death. In vivo depletion studies demonstrated that the ability of IL-2 plus IL-18 to mediate STAT4-independent resistance to T. gondii is dependent on NK cell production of IFN-gamma. Together, these studies identify STAT4 as an important transcription factor required for development of the innate NK and adaptive T cell responses necessary for resistance to T. gondii. However, other signaling pathways can be used to bypass STAT4-dependent production of IFN-gamma and enhance innate resistance to T. gondii.

Adjuvants, Immunologic↗

PubMed2000

Self-assembly of oligomeric porphyrin rings.

A cobalt porphyrin equipped with two different but geometrically complementary pyridine ligands self-assembles to form an unusually stable complex with approximately 12 porphyrin monomers arranged in a macrocyclic array.

Cobalt↗

PubMed2000

Sequence-dependent DNA structure: dinucleotide conformational maps.

We have used a computational model to calculate the potential energy surface for dinucleotide steps in double helical DNA as a function of the two principal degrees of freedom, slide and shift. By using a virtual bond to model the constraints imposed by the sugar-phosphate backbone, twist, roll, tilt and rise can be simultaneously optimised for any given values of slide and shift. Thus we have been able to construct complete conformational maps for all step types. For some steps, the maps agree well with experimental data from X-ray crystal structures, but other steps appear to be strongly perturbed by the effects of context (conformational coupling with the neighbouring steps). The optimised values of twist and roll show sequence-dependent variations consistent with the crystal structure data. The conformational maps allow us to construct adiabatic paths, and hence calculate the flexibility of each step with respect to slide and shift. Again the results agree well with the available experimental assignments of flexibility: YR steps, CA/TG and CG, are the most flexible and RR steps, such as AA, the least flexible.

Base Pairing↗

PubMed2000

Sequence-dependent DNA structure: tetranucleotide conformational maps.

A database of X-ray crystal structures of double helical DNA oligomers has been used to analyse the role of the sugar-phosphate backbone in coupling the conformational properties of neighbouring dinucleotide steps. The base step parameters which are most strongly coupled to the backbone degrees of freedom are slide and shift, and these are the two dinucleotide step parameters which show strong correlations along a sequence: the value of slide follows the values in the neighbouring steps, whereas shift tends to alternate. This conformational coupling is mediated by the shared furanose rings at the step junctions: a change in the value of slide causes a change in the mean value of the same strand 3' and 5'-chi torsion angle, and a change in the mean value of the 3' and 5' sugar pseudo-rotation phase angle, P; a change in the value of shift causes a difference between the same strand 3' and 5'-chi in A-DNA and a difference between the 3' and 5'-P in B-DNA. We have used a database of tetranucleotide X-ray crystal structures to parameterise a simple model for the coupling of slide and shift. Using this junction model together with our dinucleotide step potential energy maps described previously, we can in principle calculate the structure of any DNA oligomer. The parameterisation indicates that the rotational step parameters are accurate to within 5 degrees, and the translational step parameters are accurate to within 0.5 A. The model has been used to study the potential energy surfaces of all possible tetranucleotide sequences, and the calculations agree well with the experimental data from X-ray crystal structures. Some dinucleotide steps are context independent (AA/TT, AT and TA), because the conformational properties of all possible neighbouring steps are compatible. When the conformational properties of the neighbours are not compatible, the behaviour of a step cannot be understood at the dinucleotide level. Thus the conformations of CG, GC and GG/CC are all strongly context dependent. The remaining mixed sequence steps show weakly context-dependent behaviour. The approach allows the calculation of the relative stability and flexibility of tetranucleotide sequences, and the results indicate why TATA is used as an origin of replication. Clear predictions are made about sequences which have not yet been characterised crystallographically. In particular, poly(CCA).poly(TGG) is predicted to have an unusual structure which lies between the C and D-DNA polymorphs.

Base Pairing↗

PubMed2000

Cytokines and T cells in host defense.

Soluble and cell-bound ligands profoundly influence the differentiative fate of lymphocytes during an immune response. Recent advances have been made in understanding the role of cytokine signals and costimulatory signals in the regulation of T cell responses associated with resistance or susceptibility to infection. There has also been recent progress in defining the requirements for the generation and maintenance of immunologic memory, a critical component of adaptive immunity.

Animals↗

PubMed2000

Interleukin-18 (IL-18) enhances innate IL-12-mediated resistance to Toxoplasma gondii.

Innate resistance to Toxoplasma gondii is dependent on the ability of interleukin-12 (IL-12) to stimulate natural killer (NK) cell production of gamma interferon (IFN-gamma). Since IL-18 is a potent enhancer of IL-12-induced production of IFN-gamma by NK cells, SCID mice (which lack an adaptive immune response) were used to assess the role of IL-18 in innate resistance to T. gondii. Administration of anti-IL-18 to SCID mice infected with T. gondii resulted in an early reduction in serum levels of IFN-gamma but did not significantly decrease resistance to this infection. In contrast, administration of exogenous IL-18 to infected SCID mice resulted in increased production of IFN-gamma, reduced parasite burden, and a delay in time to death. The protective effects of IL-18 treatment correlated with increased NK cell numbers and cytotoxic activity at the local site of administration and with elevated levels of inducible nitrous oxide synthose in the spleens of treated mice. In addition, in vivo depletion studies demonstrated that the ability of exogenous IL-18 to enhance resistance to T. gondii was dependent on IL-12, IFN-gamma, and NK cells. Together, these studies demonstrate that although endogenous IL-18 appears to have a limited role in innate resistance to T. gondii, treatment with IL-18 can augment NK cell-mediated immunity to this pathogen.

Animals↗

PubMed2000

The CD40/CD40 ligand interaction is required for resistance to toxoplasmic encephalitis.

Since the CD40/CD40 ligand (CD40L) interaction is involved in the regulation of macrophage production of interleukin 12 (IL-12) and T-cell production of gamma interferon (IFN-gamma), effector cell functions associated with resistance to Toxoplasma gondii, the role of CD40L in immunity to this parasite was assessed. Infection of C57BL/6 mice with T. gondii results in an upregulation of CD40 expression on accessory cell populations at local sites of infection as well as in lymphoid tissues. Splenocytes from C57BL/6 mice infected with T. gondii for 5 days produced high levels of IL-12 and IFN-gamma when stimulated with toxoplasma lysate antigen, and blocking CD40L did not significantly alter the production of IFN-gamma or IL-12 by these cells. Similar results were observed with splenocytes and mononuclear cells isolated from the brains of chronically infected mice. Interestingly, although CD40L(-/-) mice infected with T. gondii produced less IL-12 than wild-type mice, they produced comparable levels of IFN-gamma but succumbed to toxoplasmic encephalitis 4 to 5 weeks after infection. The inability of CD40L(-/-) mice to control parasite replication in the brain correlated with the ability of soluble CD40L, in combination with IFN-gamma, to activate macrophages in vitro to control replication of T. gondii. Together, these results identify an important role for the CD40/CD40L interaction in resistance to T. gondii. However, this interaction may be more important in the control of parasite replication in the brain rather than the generation of protective T-cell responses during toxoplasmosis.

Animals↗

PubMed2000

Blockade of costimulation prevents infection-induced immunopathology in interleukin-10-deficient mice.

Interleukin-10 (IL-10) is associated with inhibition of cell-mediated immunity and downregulation of the expression of costimulatory molecules required for T-cell activation. When IL-10-deficient (IL-10KO) mice are infected with Toxoplasma gondii, they succumb to a T-cell-mediated shock-like reaction characterized by the overproduction of IL-12 and gamma interferon (IFN-gamma) associated with widespread necrosis of the liver. Since costimulation is critical for T-cell activation, we investigated the role of the CD28-B7 and CD40-CD40 ligand (CD40L) interactions in this infection-induced immunopathology. Our studies show that infection of mice with T. gondii resulted in increased expression of B7 and CD40 that was similar in wild-type and IL-10KO mice. In vivo blockade of the CD28-B7 or CD40-CD40L interactions following infection of IL-10KO mice with T. gondii did not affect serum levels of IFN-gamma or IL-12, nor did it prevent death in these mice. However, when both pathways were blocked, the IL-10KO mice survived the acute phase of infection and had reduced serum levels of IFN-gamma and alanine transaminase as well as decreased expression of inducible nitric oxide synthase in the liver and spleen. Analysis of parasite-specific recall responses from infected IL-10KO mice revealed that blockade of the CD40-CD40L interaction had minimal effects on cytokine production, whereas blockade of the CD28-B7 interaction resulted in decreased production of IFN-gamma but not IL-12. Further reduction of IFN-gamma production was observed when both costimulatory pathways were blocked. Together, these results demonstrate that the CD28-B7 and CD40-CD40L interactions are involved in the development of infection-induced immunopathology in the absence of IL-10.

Abatacept↗

PubMed1999

Aluminium hydroxide adjuvant initiates strong antigen-specific Th2 responses in the absence of IL-4- or IL-13-mediated signaling.

Previous studies demonstrate that aluminium hydroxide adjuvant (alum) produces increased Th1 responses in IL-4-deficient mice compared with wild-type animals, although the continued production of IL-5 by spleen cells from these mice also indicates that Th2 responses are induced. In the present study, we demonstrate that alum can induce Th2-associated IL-4 and IL-5 production in the absence of IL-4 signaling in mice deficient in either IL-4Ralpha or Stat6. The Th2 responses observed could not be due to IL-13 as IL-13 responses are also impaired in IL-4Ralpha- and Stat6-deficient mice. We also detected higher levels of IL-4 in IL-4Ralpha gene-deficient, though not Stat6-deficient, mice compared with their wild-type counterparts. The increased levels of IL-4 could be explained by the IL-4R being unavailable to neutralize this cytokine in IL-4Ralpha-deficient mice. While levels of IL-5 production in IL-4Ralpha- or Stat6-deficient mice were similar to IL-4-deficient and wild-type mice, other type 2-associated responses, which are largely or wholly IL-4 dependent, such as the production of IgG1 or IgE Abs, were either reduced or absent. We conclude that alum adjuvants can induce IL-4 production and Th2 responses independently of IL-4 or IL-13, negating the requirement for an early source of IL-4 in the Th2 response induced by this adjuvant.

Adjuvants, Immunologic↗

PubMed1999

The NF-kappa B family member RelB is required for innate and adaptive immunity to Toxoplasma gondii.

The NF-kappa B family of transcription factors are associated with the regulation of innate and adaptive immunity to infection. Infection of C57BL/6 mice with Toxoplasma gondii resulted in up-regulation of NF-kappa B activity that included the NF-kappa B family member RelB. To assess the role of RelB in the regulation of the immune response to this infection, we challenged RelB-deficient mice (RelB-/-) and wild-type (WT) littermate controls with T. gondii. Although WT controls were resistant to T. gondii, RelB-/- mice succumbed 10-15 days after infection. Examination of accessory cell functions associated with resistance to T. gondii revealed that RelB-/- macrophages stimulated with IFN-gamma plus LPS or TNF-alpha produced IL-12 as well as reactive nitrogen intermediates and inhibited parasite replication similar to WT macrophages. Analysis of the systemic responses of RelB-/- and WT mice revealed that infected mice had similar serum levels of IL-12. However, RelB-/- mice challenged with T. gondii produced negligible levels of IFN-gamma and had reduced NK cell activity compared with WT mice. Similarly, splenocytes from uninfected RelB-/- mice stimulated with polyclonal stimuli were deficient in their ability to produce IFN-gamma. Together, our results demonstrate that RelB is essential for the development of innate NK and adaptive T cell responses that lead to the production of IFN-gamma and resistance to T. gondii.

Animals↗

PubMed1999

Role of CD28 in the generation of effector and memory responses required for resistance to Toxoplasma gondii.

CD28 deficient (CD28-/-) mice were used to study the role of costimulation in the T cell-mediated, IFN-gamma-dependent mechanism of resistance to Toxoplasma gondii. These mice were resistant to infection with the ME49 strain of T. gondii. Analysis of the immune response of acutely infected CD28-/- mice revealed that IL-12 was required for T cell production of IFN-gamma and this was independent of the CD40/CD40 ligand interaction. A similar mechanism of IL-12-dependent, CD28/B7 independent production of IFN-gamma by T cells was also observed in wild-type mice. Interestingly, although chronically infected wild-type mice were resistant to rechallenge with the virulent RH strain of T. gondii, chronically infected CD28-/- mice were susceptible to rechallenge with the RH strain. This deficiency in the protective memory response by CD28-/- mice correlated with a lack of IL-2 and IFN-gamma in recall responses and reduced numbers of CD4+ T cells expressing a memory phenotype. Together, our findings demonstrate that CD28 is not required for the development of a protective T cell response to T. gondii, but CD28 is required for an optimal secondary immune response.

Animals↗

PubMed1999

The CD28/B7 interaction is not required for resistance to Toxoplasma gondii in the brain but contributes to the development of immunopathology.

Infection of C57BL/6 mice with Toxoplasma gondii leads to chronic encephalitis characterized by infiltration into the brain of T cells that produce IFN-gamma and mediate resistance to the parasite. Our studies revealed that expression of B7.1 and B7.2 was up-regulated in brains of mice with toxoplasmic encephalitis (TE). Because CD28/B7-mediated costimulation is important for T cell activation, we assessed the contribution of this interaction to the production of IFN-gamma by T cells from brains and spleens of mice with TE. Stimulation of splenocytes with Toxoplasma Ag or anti-CD3 mAb resulted in production of IFN-gamma, which was inhibited by 90% in the presence of CTLA4-Ig, an antagonist of B7 stimulation. However, production of IFN-gamma by T cells from the brains of these mice was only slightly reduced (20%) by the addition of CTLA4-Ig. To address the role of the CD28/B7 interaction during TE, we compared the development of disease in C57BL/6 wild-type (wt) and CD28-/- mice. Although the parasite burden was similar in wt and CD28-/- mice, CD28-/- mice developed less severe encephalitis and survived longer than wt mice. Ex vivo recall responses revealed that mononuclear cells isolated from the brains of chronically infected CD28-/- mice produced less IFN-gamma than wt cells, and this correlated with reduced numbers of intracerebral CD4+ T cells in CD28-/- mice compared with wt mice. Taken together, our data show that resistance to T. gondii in the brain is independent of CD28 and suggest a role for CD28 in development of immune-mediated pathology during TE.

Animals↗