Progress 09/01/02 to 08/31/08
Outputs
OUTPUTS: project terminated 8/31/2008. nothing reported PARTICIPANTS: Not relevant to this project. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
nothing reported
Publications
- No publications reported this period
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Progress 01/01/06 to 12/31/06
Outputs
We are identifying candidate immunomodulatory proteins in Aedes aegypti saliva by tryptic peptide fingerprinting following chromatofocussing and anion exchange HPLC. Expression of a possible macrophage inhibitory protein using the DES system (Invitrogen) in Drosophila S2 cells in progress. In addition, we are exploring the use of a novel technique for expressing antibodies, using a DNA vaccination strategy in Swiss-Webster mice. We have begun to examine the hypothesis that the mosquito salivary immunomodulator has a fitness benefit for the mosquito by minimizing the adverse consequences of host seroconversion. The fitness impact of host seroconversion was examined in four mosquito species. Seroconversion is associated with a reduction in production of total and viable eggs, and with a decrease in blood meal volume. Cross-reaction occurs between related mosquito species. The effect is dependent on host antibodies, as there is no fitness decline when mosquitoes feed on
B-cell knockout mice that cannot produce antibodies. This effect might be exploited to develop vaccinations that can reduce mosquito populations. Preliminary experiments with the blackfly, Simulium vitattum, indicate the saliva contains an immunomodulatory activity, reducing T-cell proliferation without increasing T-cell death by necrosis or apoptosis. As this fly is a vector of vesicular stomatitis virus (VSV), this immunomodulatory activity of saliva might be an appropriate target for manipulation to decrease VSV transmission.
Impacts
Knowledge of the effect of vector saliva on physiological and immunological defenses at the point of entry of pathogens is critical to understanding the transmission process. The immunomodulatory effect of saliva is likely to impact the effect of anti-pathogen vaccines, especially at the point of introduction of the pathogen when control should be easiest. Further, identification of immunomodulatory proteins can suggest novel targets for vaccine or drug intervention, resulting in local conditions in the skin less favorable to pathogen transmission. In particular, we are targeting the vectors of both human (Yellow Fever, Dengue Fever) and veterinary (Vesicular Stomatitis) diseases, with the goal of developing novel transmission-blocking vaccines.
Publications
- No publications reported this period
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Progress 01/01/05 to 12/31/05
Outputs
Studies of the immunomodulatory effect of mosquito saliva have been extended to consider effects on murine macrophages, and on aspects of human peripheral blood monocytic cell (PBMC) function. Aedes aegypti saliva inhibits phagocytosis, MHC class I and II presentation, NO production, and cytokine (especially IL-12) secretion from macrophages. These effects compromise the ability of macrophages to control infectious agents, and they alter the ability of these cells to drive acquired immune responses through interaction with both CD4 and CD8 helper T-cells. Indeed saliva-treated macrophages and dendritic cells drive a weak Th2-type response in CD4 T-cells following stimulation with ovalbumin, whereas control macrophages drive a Th1 response. In addition, saliva induces apoptosis in CD4, CD8, and gamma-delta T-cells, and in B-cells. Altogether these effects of saliva result in depletion of T- and B-cells at the bite site, and alteration of macrophage function compromises
innate immune responses and conditions a subsequent systemic Th2 response in the host. These immunomodulatory and immunosuppressive effects are likely to facilitate transmission of viruses, including Dengue and Yellow Fever viruses, by this mosquito. Similar effects are produced by saliva of other Aedes and Ochlerotatus species, but not by Culex pipiens (at least not in mammalian cells). Chromatofocussing HPLC indicates that the pI of the immunomodulatory protein is 5.2, and size-exclusion chromatography indicates an estimated size of >300,000 Da for the native protein. In human PBMCs, mitogen-stimulated CD4 and CD8 T-cell proliferation is inhibited, but these cells are about 10-fold less sensitive than their murine counterparts. Aedes aegypti saliva stimulates IL-12 secretion, and inhibits IL-2, from PBMCs and dendritic cells. Consequently net interferon-gamma secretion is not affected. T-cell secretion of several Th2-type cytokines, including IL-4, IL-5, and IL-13 is decreased in a
dose-dependent manner. IL-10 secretion from PBMCs and from monocytes was stimulated at low saliva concentrations and inhibited at higher concentrations, but secretion of this cytokine by neutrophils and dendritic cells was inhibited at all saliva concentrations. Effects on tumor necrosis factor alpha secretion was also complex, with stimulation by low concentrations of saliva and inhibition by higher concentrations seen in monocytes, while secretion from neutrophils and dendritic cells was inhibited at all saliva concentrations. Overall saliva of this mosquito produces a more inflammatory response in humans, compared to an anti-inflammatory effect in murine models. The effect of saliva of two tsetse flies, Glossina morsitans morsitans and G. palpalis, was also examined. These vectors of African trypanosomiasis weakly inhibit secretion of Th1-type cytokines, including IL-2 and interferon-gamma, and they upregulate secretion of the Th2-type cytokine IL-4 and IL-10 by 2- and 5-fold
respectively. This strong pro-Th2 effect is likely to alter the ability of the host immune response to manage trypanosome infections at the time of inoculation into the vertebrate host.
Impacts
Knowledge of the effect of vector saliva on physiological and immunological defenses at the point of entry of pathogens is critical to understanding the transmission process. The immunomodulatory effect of saliva is likely to impact the effect of anti-pathogen vaccines, especially at the point of introduction of the pathogen when control should be easiest. Further, identification of immunomodulatory proteins can suggest novel targets for vaccine or drug intervention, resulting in local conditions in the skin less favorable to pathogen transmission.
Publications
- Donald Champagne and Heather Wasserman. (2005) Contrasting effects of Aedes aegypti saliva on murine and human immune effector cells. American Journal of Tropical Medicine and Hygiene, 75(6) Supplement pg 48.
- Donald Champagne, Irene Kasumba, Changyun Hu, Sandra Abrajim, and Serap Aksoy. (2005). Saliva of the tsetse flies Glossina morsitans morsitans and Glossina palpalis modulates antigen-stimulated immune effector cell functions in a mouse model. American Journal of Tropical Medicine and Hygiene, 75(6) Supplement pg 212.
- Champagne, D.E. (2006) Antihemostatic molecules from saliva of blood-feeding arthropods. Pathophysiology of Haemostasis and Thrombosis (in press).
- Wasserman, H.A., Abrajim, S., Champagne D.E. (2006. Modulation of murine macrophage function by saliva of the Yellow Fever mosquito, Aedes aegypti. Immunity and Infection (in review).
- Wasserman, H.A., Champagne, D.E. (2006). Immunomodulatory effects of saliva of the Yellow Fever mosquito, Aedes aegypti, on human peripheral blood monocytes. American Journal of Tropical Medicine and Hygiene (in review).
- Champagne, DE., Wasserman, HA., Abrajim, S. (2006). Effects of saliva of the malaria vector Anopheles stephensi on murine and human immune cell function. American Journal of Tropical Medicine and Hygiene (in review)
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Progress 01/01/04 to 12/31/04
Outputs
In the past year we have extended our examination of the immunomodulatory effects of Aedes aegypti and Anopheles stephensi saliva to include effects on antigen-presenting cells, including macrophages and dendritic cells. Effects on these cells include decreased secretion of IL-12 and THF-alpha, decreased phagocytosis, and decreased presentation of both MHC class I and MHC class II. These effects could severely compromise the ability of these cells to function in both innate and acquired immune responses. We have also characterized the spectrum of cells that become apoptotic in the presence of Aedes aegypti saliva, which includes CD4, CD8, and gamma-delta T-cells, B-cells, and neutrophils, but not macrophages or dendritic cells. Finally we have examined the effect of Aedes aegypti saliva on human PBMCs, finding effects that are broadly comparable to the results of the mouse experiments, except for and elevation (rather than a decrease) in IL-12 and TNF-alpha
secretion.
Impacts
Transmission of pathogens and parasites by mosquitoes remains a significant source of veterinary and human disease. Understanding the immunomodulatory activity of mosquito saliva may lead to improvements in vaccines, or lead to novel strategies for blocking pathogen transmission.
Publications
- Champagne, D.E., Wasserman, H.A., Kumar, S. and Singh, S. (2004). Pharmacological and Immunological Properties of Saliva of the Blood-Feeding Insects Rhodnius prolixus and Aedes aegypti. Physiological Entomology, 29:269-277.
- Wasserman, H.A., Singh, S., and Champagne, D.E. (2004). Saliva of the Yellow Fever mosquito, Aedes aegypti, modulates murine lymphocyte function. Parasite Immunology 26: 295-306.
- Wanasen, N., Nussenzveig, R.H., Champagne, D.E., Soong, L., and Higgs, S. (2004). Differential modulation of murine host immune response by salivary gland extracts from the mosquitoes Aedes aegypti and Culex quinquefasciatus. Medical and Veterinary Entomology 18: 191-199.
- Champagne, D.E. (2004). Antihemostatic strategies of blood-feeding arthropods. Current Drug Targets: Cardiovascular and Haematologic Disorders, 4:375-396.
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Progress 01/01/03 to 12/31/03
Outputs
In this past year we have concentrated on comparing the immunomodulatory activity of salivary gland extract (SGE) from three mosquito species: Aedes aegypti, Anopheles stephensi, and Culex pipiens quinquifasciatus. At concentrations between 0.15 and 0.6 salivary gland pair (SGP)/ml, Aedes aegypti SGE inhibits CD4 and CD8 T-cell proliferation and secretion of Th1 type cytokines in response to antigen challenge. At concentrations above this, secretion of both Th1 and Th2 cytokines is inhibited. The latter effect is due to aopotosis induced in T- and B-cells, gamma-delta-T-cells, and neutrophils, but not in antigen-presenting cells including macrophages and dendritic cells. The effect of Anopheles stephensi SGE is less pronounced, with modest (40-50%) suppression of T-cell proliferation and cytokine secretion at concentrations equivalent to 15 SGP/ml. Culex pipiens quinquifasciatus SGE has no effect at concentrations up to 2 SGP/ml. Work in the coming year will focus on
characterization of the immunomodulatory component of Aedes aegypti saliva.
Impacts
Transmission of pathogens and parasites by mosquitoes remains a significant source of veterinary and human disease. Understanding the immunomodulatory activity of mosquito saliva may lead to improvements in vaccines, or lead to novel strategies for blocking pathogen transmission.
Publications
- Nanchaya Wanasen, Roberto H. Nussenzveig, Donald E. Champagne, Lynn Soong, Stephen Higgs. 2004. The modulation of the murine host immune response by salivary gland extracts from Aedes aegypti and Culex quinquefasciatus mosquitoes. Medical and Veterinary Parasitology, in press.
- Champagne, D.E. 2004. Anti-hemostatic strategies of blood-feeding arthropods. Current Drug Targets, in press.
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Progress 01/01/02 to 12/31/02
Outputs
In earlier work we showed that saliva or salivary gland extracts from Aedes aegypti inhibited proliferation and cytokine sectretion from murine T cells. We have now shown that this is due to the presence of an apotosis-inducing activity, indicated by increased binding of the apoptsis marker Annexin V, upregulation of the Fas receptor, and activation of PARP. We are further defining the spectrum of cell types affected by this activity; initial results indicate that T- and B-cells are depleted, but macrophages and other antigen-presenting cells are resistant. This activity was not seen with extracts from Anopheles stephensi salivary glands, and the effect of these extracts on T-cell derived cytokines is more subtle, with only a 25-30% reduction following exposure to as much as 3 salivary gland pair equivalents/ml.
Impacts
This work suggests an important potential impact of Aedes aegypti feeding on the immune environment at the bite site. If immune effector cells are depleted but macrophages are left intact, viruses with a trophism for macrophages would get a "free ride" in the initial stages of the infection process. This suggests novel targets for inhibiting virus transmission.
Publications
- No publications reported this period
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