Jeffrey M. Jordan

Prior Exposure to Uninfected Mosquitoes Enhances Mortality in Naturally-Transmitted West Nile Virus Infection

Background The global emergence of West Nile virus (WNV) has highlighted the importance of mosquito-borne viruses. These are inoculated in vector saliva into the vertebrate skin and circulatory system. Arthropod-borne (arbo)viruses such as WNV are transmitted to vertebrates as an infectious mosquito probes the skin for blood, depositing the virus and saliva into the skin and circulation. Growing evidence has demonstrated that arthropod, and recently mosquito, saliva can have a profound effect on pathogen transmission efficiency, pathogenesis, and disease course. A potentially important aspect of natural infections that has been ignored is that in nature vertebrates are typically exposed to the feeding of uninfected mosquitoes prior to the mosquito that transmits WNV. The possibility that pre-exposure to mosquito saliva might modulate WNV infection was explored. Principal Findings Here we report that sensitization to mosquito saliva exacerbates viral infection. Prior exposure of mice to mosquito feeding resulted in increased mortality following WNV infection. This aggravated disease course was associated with enhanced early viral replication, increased interleukin-10 expression, and elevated influx of WNV-susceptible cell types to the inoculation site. This exacerbated disease course was mimicked by passive transfer of mosquito-sensitized serum. Significance This is the first report that sensitization to arthropod saliva can exacerbate arthropod-borne infection, contrary to previous studies with parasite and bacteria infections. This research suggests that in addition to the seroreactivity of the host to virus, it is important to take into account the immune response to vector feeding.

An Intradermal Environment Promotes a Protective Type-1 Response against Lethal Systemic Monocytotropic Ehrlichial Infection

ABSTRACT Immune responses against monocytotropic ehrlichiosis during infection with a strain of Ehrlichia from Ixodes ovatus (IOE) were evaluated using a model that closely reproduces the pathology and immunity associated with tick-transmitted human monocytotropic ehrlichiosis. C57BL/6 mice were inoculated intradermally or intraperitoneally with high-dose highly virulent IOE or intraperitoneally with mildly virulent Ehrlichia muris. Intradermal (i.d.) infection with IOE established mild, self-limited disease associated with minimal hepatic apoptosis, and all mice survived past 30 days. Intraperitoneal (i.p.) infection with IOE resulted in acute, severe toxic shock-like syndrome and severe multifocal hepatic apoptosis and necrosis, and all mice succumbed to disease. Compared to i.p. infection with IOE, intradermally infected mice had a 100- to 1,000-fold lower bacterial load in the spleen with limited dissemination. Compared to mice infected intraperitoneally with IOE, i.d. infection stimulated a stronger protective type-1 cell-mediated response on day 7 of infection, characterized by increased percentages of both CD4+ and CD8+ splenic T cells, generation of a greater number of IOE-specific, gamma interferon-producing CD4+ Th1 cells, and higher levels of tumor necrosis factor (TNF-α) in the spleen but lower concentrations of serum TNF-α and interleukin-10. These data suggest that under the conditions of natural route of challenge (i.e., i.d. inoculation), the immune response has the capacity to confer complete protection against monocytotropic ehrlichiosis, which is associated with a strong cell-mediated type-1 response and decreased systemic production of pro- and anti-inflammatory cytokines.

The Absence of Toll-Like Receptor 4 Signaling in C3H/HeJ Mice Predisposes Them to Overwhelming Rickettsial Infection and Decreased Protective Th1 Responses

ABSTRACT The importance of toll-like receptor 4 (TLR4) in immunity to rickettsiae remains elusive. To investigate the role of TLR4 in protection against rickettsioses, we utilized C3H/HeJ mice, which are naturally defective in TLR4 signaling, and compared the responses of C3H/HeN and C3H/HeJ mice following intravenous inoculation with Rickettsia conorii. Mice genetically defective in TLR4 signaling developed overwhelming, fatal rickettsial infections when given an inoculum that was nonfatal for TLR4-competent mice. In addition, mice lacking the ability to signal through TLR4 had significantly greater rickettsial burdens in vivo. Moreover, we observed greater concentrations of the cytokines interleukin 6 (IL-6), tumor necrosis factor alpha, IL-12p40, IL-12p70, and IL-17 in the sera of mice with intact TLR4 function as well as significantly greater quantities of activated CD4+ and CD8+ T lymphocytes. Additionally, we also observed that Th17 cells were present only in TLR4-competent mice, suggesting an important role for TLR4 ligation in the activation of this subset. In agreement with these data, we also observed significantly greater percentages of immunosuppressive regulatory T cells in the spleen during infection in TLR4-defective mice. Together, these data demonstrate that, while rickettsiae do not contain endotoxic lipopolysaccharide, they nevertheless initiate TLR4-specific immune responses, and these responses are important in protection.

Rickettsiae-Stimulated Dendritic Cells Mediate Protection against Lethal Rickettsial Challenge in an Animal Model of Spotted Fever Rickettsiosis

The role played by dendritic cells (DCs), initiators and orchestrators of the immune response, remains unclear in rickettsial infections. To investigate their importance in rickettsioses, we analyzed the responses of murine bone marrow-derived DCs (BMDCs) after rickettsial stimulation in vitro and their protective role in vivo. Rickettsia conorii stimulation of BMDCs caused significant maturation and production of proinflammatory cytokines. Transfer of rickettsiae-stimulated DCs protected mice from lethal rickettsial challenge by limiting rickettsial proliferation in vivo, whereas partial protection was observed in mice receiving lipopolysaccharide (LPS)-stimulated DCs. Immunity to R. conorii after transfer of DCs was associated with up-regulation of CD40, CD80, CD86, and major histocompatibility complex class II; with production of IL-2, IL-12, and IL-23; and with production of antigen-specific interferon- gamma in T cells. Taken together, our data suggest that a vigorous proinflammatory response in DCs is associated with protective immunity to rickettsiae and that generation of antigen-specific immunity is crucial to complete protection.

Rickettsiae stimulate dendritic cells through Toll-like receptor 4, leading to enhanced NK cell activation in vivo.

Adoptive transfer of Toll-like receptor (TLR) 4-stimulated dendritic cells (DCs) induces protective immunity against an ordinarily lethal rickettsial challenge, but the mechanism underlying this protection remains elusive. Therefore, we sought to determine the importance of TLR4 in early immunity to rickettsiae in vivo, particularly that conferred by TLR4-stimulated DCs. Rickettsial growth proceeded logarithmically in mice lacking TLR4 function, whereas in TLR4-competent mice rickettsial growth manifested a lag phase early, suggesting that TLR4 may initiate innate rickettsial immunity. TLR4-competent mice produced significant amounts of interferon (IFN)-gamma on day 1 of Rickettsia conorii infection, which was associated with significant expansion of the population of activated NK cells. Moreover, NK cells from TLR4-competent mice produced significantly higher levels of IFN-gamma and had greater cytotoxic activity than did those from TLR4-deficient mice. Last, adoptive transfer of rickettsiae-exposed, TLR4-stimulated DCs activated NK cells in vivo. Together, these data reveal an important role for DCs in recognizing rickettsiae through TLR4 and inducing early antirickettsial immunity.

T Cells Mediate Cross-Protective Immunity between Spotted Fever Group Rickettsiae and Typhus Group Rickettsiae

Rickettsioses are severe infections caused by obligately intracellular bacteria that preferentially infect the endothelium lining the vasculature. The causative agents, rickettsiae, have been divided according to biological, genetic, and antigenic parameters into 2 main groups: spotted fever and typhus. They have not been thought to stimulate cross-reactive protective immune responses; however, in this study, we show that, in relevant animal models that mimic human rickettsial infections, there is reciprocal immunological cross-protection between spotted fever group and typhus group rickettsiae. Furthermore, we present evidence that T cells are responsible for this cross-immunity and that cross-stimulation of T cells also occurs in humans.

Rickettsiae Stimulate Dentric Cells Through Toll-Like Receptor 4, Leading To Enhanced NK Cell Activation In Vivo

Adoptive transfer of Toll-like receptor (TLR) 4-stimulated dendritic cells (DCs) induces protective immunity against an ordinarily lethal rickettsial challenge, but the mechanism underlying this protection remains elusive. Therefore, we sought to determine the importance of TLR4 in early immunity to rickettsiae in vivo, particularly that conferred by TLR4-stimulated DCs. Rickettsial growth proceeded logarithmically in mice lacking TLR4 function, whereas in TLR4-competent mice rickettsial growth manifested a lag phase early, suggesting that TLR4 may initiate innate rickettsial immunity. TLR4-competent mice produced significant amounts of interferon (IFN)-gamma on day 1 of Rickettsia conorii infection, which was associated with significant expansion of the population of activated NK cells. Moreover, NK cells from TLR4-competent mice produced significantly higher levels of IFN-gamma and had greater cytotoxic activity than did those from TLR4-deficient mice. Last, adoptive transfer of rickettsiae-exposed, TLR4-stimulated DCs activated NK cells in vivo. Together, these data reveal an important role for DCs in recognizing rickettsiae through TLR4 and inducing early antirickettsial immunity.

Hyperthermia and Chemotherapy: The Science

Cancer ranks second only to heart disease as the greatest cause of mortality in the United States. Despite significant strides in diagnosis and therapeutics, clinical treatment of metastatic cancer still yields a poor prognosis. Basic science research is increasing our knowledge of the mechanisms by which chemotherapeutic and hyperthermic modalities exert their antitumor activity. Herein, we describe the current state of knowledge of these treatment modalities and the basic science behind their clinical usefulness. Moreover, we identify gaps in our understanding of the basic science of these treatments. Basic science is primed to exert a substantial role in extending our therapeutic options through the clarification of current therapeutic mechanisms, as well as the generation of novel treatments.

Rickettsiae stimulate dendritic cells through TLR4, leading to enhanced NK cell activation in vivo

Adoptive transfer of Toll-like receptor (TLR) 4-stimulated dendritic cells (DCs) induces protective immunity against an ordinarily lethal rickettsial challenge, but the mechanism underlying this protection remains elusive. Therefore, we sought to determine the importance of TLR4 in early immunity to rickettsiae in vivo, particularly that conferred by TLR4-stimulated DCs. Rickettsial growth proceeded logarithmically in mice lacking TLR4 function, whereas in TLR4-competent mice rickettsial growth manifested a lag phase early, suggesting that TLR4 may initiate innate rickettsial immunity. TLR4-competent mice produced significant amounts of interferon (IFN)-gamma on day 1 of Rickettsia conorii infection, which was associated with significant expansion of the population of activated NK cells. Moreover, NK cells from TLR4-competent mice produced significantly higher levels of IFN-gamma and had greater cytotoxic activity than did those from TLR4-deficient mice. Last, adoptive transfer of rickettsiae-exposed, TLR4-stimulated DCs activated NK cells in vivo. Together, these data reveal an important role for DCs in recognizing rickettsiae through TLR4 and inducing early antirickettsial immunity.

Tissue Culture Models

The use of tissue cultures as a research tool to investigate the pathophysiological bases of diseases has become essential in the current ag of molecular biomedical research. Although it will always be necessary to translate and validate the observations seen in vitro to the patient or animal, the ability to investigate the role(s) of individual variables free from confounders is paramount toward increasing our understanding of the physiology and their role in disease. Additionally, it is not feasible to conduct certain research in humans because of ethical constraints, yet investigators may still be interested in the physiological response in human tissues; in vitro characterization of human tissue is an acceptable choice. Tissue culture techniques have been utilized extensively to investigate questions pertaining to physiology and disease. The isolation and propagation of human epithelial cells has allowed investigators to begin to characterize the interactions and reactions that occur in response to various stimuli. Moreover, the culture of other human tissue has allowed researchers to investigate pathological cascades as well as other physiological responses. Combining cultured endothelial cells and leukocytes together in vitro under laminar flow conditions has helped elucidate the critical interactions that occur in rolling and emigration of leukocytes during the inflammatory response. Cultured embryonic stem cells that had been subjected to various growth conditions have advanced our understanding of cellular differentiation and growth.