Heralded Distribution of Single-Photon Path Entanglement.

We report the experimental realization of heralded distribution of single-photon path entanglement at telecommunication wavelengths in a repeater-like architecture. The entanglement is established upon detection of a single photon, originating from one of two spontaneous parametric down-conversion photon pair sources, after erasing the photon's which-path information. In order to certify the entanglement, we use an entanglement witness which does not rely on postselection. We herald entanglement between two locations, separated by a total distance of 2 km of optical fiber, at a rate of 1.6 kHz. This work paves the way towards high-rate and practical quantum repeater architectures.

Association between i.v. thrombolysis volume and door-to-needle times in acute ischemic stroke.

Centralization of intravenous thrombolysis (IVT) for acute ischemic stroke in high-volume centers is believed to improve the door-to-needle times (DNT), but limited data support this assumption. We examined the association between DNT and IVT volume in a large Dutch province. We identified consecutive patients treated with IVT between January 2009 and 2013. Based on annualized IVT volume, hospitals were categorized as low-volume (≤ 24), medium-volume (25-49) or high-volume (≥ 50). In logistic regression analysis, low-volume hospitals were used as reference category. Of 17,332 stroke patients from 11 participating hospitals, 1962 received IVT (11.3 %). We excluded 140 patients because of unknown DNT (n = 86) or in-hospital stroke (n = 54). There were two low-volume (total 101 patients), five medium-volume (747 patients) and four high-volume hospitals (974 patients). Median DNT was shorter in high-volume hospitals (30 min) than in medium-volume (42 min, p < 0.001) and low-volume hospitals (38 min, p < 0.001). Patients admitted to high-volume hospitals had a higher chance of DNT < 30 min (adjusted OR 3.13, 95 % CI 1.70-5.75), lower risk of symptomatic intracerebral hemorrhage (adjusted OR 0.39, 95 % CI 0.16-0.92), and a lower mortality risk (adjusted OR 0.45, 95 % CI 0.21-1.01), compared to low-volume centers. There was no difference in DNT between low- and medium-volume hospitals. Onset-to-needle times (ONT) did not differ between the groups. Hospitals in this Dutch province generally achieved short DNTs. Despite this overall good performance, higher IVT volumes were associated with shorter DNTs and lower complication risks. The ONT was not associated with IVT volume.

Landau's quasiparticle mapping: Fermi liquid approach and Luttinger liquid behavior.

A continuous unitary transformation is introduced which realizes Landau's mapping of the elementary excitations (quasiparticles) of an interacting Fermi liquid system to those of the system without interaction. The conservation of the number of quasiparticles is important. The transformation is performed numerically for a one-dimensional system, i.e., the worst case for a Fermi liquid approach. Yet evidence for Luttinger liquid behavior is found. Such an approach may open a route to a unified description of Fermi and Luttinger liquids on all energy scales.

Helicobacter hepaticus-induced colitis in interleukin-10-deficient mice: cytokine requirements for the induction and maintenance of intestinal inflammation.

We have previously shown that specific-pathogen-free interleukin-10 (IL-10)-deficient (IL-10 KO) mice reconstituted with Helicobacter hepaticus develop severe colitis associated with a Th1-type cytokine response. In the present study, we formally demonstrate that IL-12 is crucial for disease induction, because mice deficient for both IL-10 and IL-12 p40 show no intestinal pathology following H. hepaticus infection. By using monoclonal antibodies (MAbs) to IL-12, gamma interferon (IFN-gamma), and tumor necrosis factor alpha (TNF-alpha), we have further analyzed the role of these cytokines in the maintenance of the Th1 response and inflammation in IL-10 KO mice with established H. hepaticus-induced colitis. Treatment of infected colitic IL-10 KO mice with anti-IL-12 p40 resulted in markedly reduced intestinal inflammation, colonic IFN-gamma, TNF-alpha, and inducible nitric oxide synthase (iNOS) mRNA levels, and H. hepaticus-specific IFN-gamma secretion by mesenteric lymph node (MLN) cells compared to the findings in control MAb-treated mice. Moreover, the diminished pathology was associated with decreased numbers of colonic CD3(+) T cells and significantly reduced frequencies of Helicobacter-reactive CD4(+) Th1 cells in MLN. In contrast, anti-IFN-gamma and/or anti-TNF-alpha had no effect on intestinal inflammation in IL-10 KO mice with established colitis. Using IL-10/IFN-gamma double-deficient mice, we further show that IFN-gamma is not required for the development of colitis following H. hepaticus infection. MLN cells from infected IL-10/IFN-gamma KO animals secreted elevated amounts of IL-12 and TNF-alpha following bacterial antigen stimulation, indicating alternative pathways of disease induction. Taken together, our results demonstrate a crucial role for IL-12 in both inducing and sustaining intestinal inflammation through recruitment and maintenance of a pool of pathogenic Th1 cells.

Recombinant attenuated Toxoplasma gondii expressing the Plasmodium yoelii circumsporozoite protein provides highly effective priming for CD8+ T cell-dependent protective immunity against malaria.

The protozoan parasite Toxoplasma gondii elicits strong cell-mediated immunity against itself as well as nonspecific resistance against other pathogens and tumors. For this reason, we asked whether recombinant Toxoplasma could be utilized as an effective vaccine vehicle for inducing immunity against heterologous microbial infections. The circumsporozoite protein (PyCSP) of Plasmodium yoelii was engineered into a T. gondii temperature-sensitive strain (ts-4), a mutant that induces complete protection against virulent Toxoplasma challenge. When administered to mice in a single dose, a recombinant ts-4 (CSC3) that both secretes and expresses surface PyCSP induced strong anti-CSP Ab responses, with an isotype distribution pattern similar to that stimulated by the T. gondii carrier. When challenged with P. yoelii sporozoites during the first month after CSC3 vaccination, these animals displayed substantial levels of nonspecific resistance attributable entirely to the T. gondii carrier. Nevertheless, after the nonspecific protection had waned, high levels (up to 79%) of specific immunity against sporozoite challenge were achieved by boosting the animals with recombinant vaccinia virus expressing PyCSP. These CSC3-primed PyCSP-vaccinia-boosted mice displayed high frequencies of splenic PyCSP-specific IFN-gamma-producing cells, as well as CD8+ T cell-dependent cytolytic activity. In vivo depletion of CD8+ lymphocytes at the time of challenge completely ablated protective immunity in the T. gondii-primed/vaccinia-boosted animals, while neutralization of IFN-gamma or IL-12 caused a partial but significant reduction in resistance. Together these findings establish the efficacy of recombinant attenuated Toxoplasma as a vaccine vehicle for priming CD8+-dependent cell-mediated immunity.

Host resistance and immune deviation in pigeon cytochrome c T-cell receptor transgenic mice infected with Toxoplasma gondii.

Resistance to Toxoplasma gondii has been shown to be mediated by gamma interferon (IFN-gamma) produced by NK, CD4(+), and CD8(+) T cells. While studies of SCID mice have implicated NK cells as the source of the cytokine in acute infection, several lines of evidence suggest that IFN-gamma production by CD4(+) T lymphocytes also plays an important role in controlling early parasite growth. To evaluate whether this function is due to nonspecific as opposed to T-cell receptor (TCR)-dependent stimulation by the parasite, we have examined the resistance to T. gondii infection of pigeon cytochrome c transgenic (PCC-Tg) Rag-2(-/-) mice in which all CD4(+) T lymphocytes are unreactive with the protozoan. When inoculated with the ME49 strain, PCC-Tg animals exhibited only temporary control of acute infection and succumbed by day 17. Intracellular cytokine staining by flow cytometry revealed that, in contrast to infected nontransgenic controls, infected PCC-Tg animals failed to develop IFN-gamma-producing CD4(+) T cells. Moreover, the CD4(+) lymphocytes from these mice showed no evidence of activation as judged by lack of upregulated expression of CD44 or CD69. Nevertheless, when acutely infected transgenic mice were primed by PCC injection, the lymphokine responses measured after in vitro antigen restimulation displayed a strong Th1 bias which was shown to be dependent on endogenous interleukin 12 (IL-12). The above findings argue that, while T. gondii-induced IL-12 cannot trigger IFN-gamma production by CD4(+) T cells in the absence of TCR ligation, the pathogen is able to nonspecifically promote Th1 responses against nonparasite antigens, an effect that may explain the immunostimulatory properties of T. gondii infection.

Single cell analysis reveals that IL-4 receptor/Stat6 signaling is not required for the in vivo or in vitro development of CD4+ lymphocytes with a Th2 cytokine profile.

The concept that IL-4 is the primary signal for Th2 lymphocyte differentiation has recently been put in doubt by studies in which the production of Th2-associated cytokines was detected in mice deficient in IL-4 synthesis or IL-4R triggering. In this study, we formally demonstrate by single cell analysis that CD4+ lymphocytes with a classical Th2 phenotype (IL-4+, IL-5+, IFN-gamma-, IL-2-) develop in significant numbers in helminth-infected mice deficient in either IL-4R alpha-chain or Stat6. While an expanded population of Th1 (IL-4-, IL-5-, IFN-gamma+, IL-2+) lymphocytes was observed in the same animals, surprisingly, cells with a mixed Th0 cytokine pattern were rare. The cytokine production phenotypes of the Th1 and Th2 subpopulations generated in infected Stat6-deficient mice were unaffected by in vitro neutralization of endogenous IL-4 or IFN-gamma. Nevertheless, while addition of exogenous rIL-12 resulted in transitory IFN-gamma production by Th2 lymphocytes from both wild-type and Stat6-deficient mice, IL-4 synthesis was preserved in the former, but temporarily ablated in the latter cells. Importantly, IL-4+ IFN-gamma- and IL-4- IFN-gamma+ populations similar to those arising in helminth-infected Stat6-deficient mice could also be generated in vitro by repetitive polyclonal stimulation of CD4+CD62Lhigh lymphocytes from uninfected mice of the same strain. Together, the results of these single cell analysis experiments demonstrate that IL-4R/Stat6 signaling, while influencing the final frequency of Th2 lymphocytes, is not essential for Th2 cell development, and suggest that this pathway has a previously unrecognized function in stabilizing Th2 populations once they have emerged.

Schistosome-infected IL-4 receptor knockout (KO) mice, in contrast to IL-4 KO mice, fail to develop granulomatous pathology while maintaining the same lymphokine expression profile.

Th2 lymphocytes have been postulated to play a major role in the immunopathology induced by Schistosoma mansoni infection. Nevertheless, infected IL-4 knockout (KO) and wild-type (wt) mice develop egg granulomas comparable in size. To further investigate the function of the Th2 response in egg pathology we studied IL-4Ralpha-deficient mice, which are nonresponsive to both IL-4 and IL-13. In striking contrast to IL-4 KO animals, infected IL-4Ralpha KO mice developed only minimal hepatic granulomas and fibrosis despite the presence of CD3+ T cells in the residual egg lesions. Moreover, liver lymphokine mRNA levels in these animals and IL-4 KO mice were equivalent. In addition, infected IL-4Ralpha-deficient, IL-4-deficient, and wt animals developed similar egg Ag-specific IgG Ab titers, arguing that CD4-dependent Th activity is intact in KO mice. As expected, IFN-gamma secretion was strongly up-regulated in mesenteric lymph node cultures from both groups of deficient animals, a change reflected in increased serum IgG2a and IgG2b Ab levels. Surprisingly, Th2 cytokine production in infected IL-4Ralpha KO mice was not abolished but was only reduced and resembled that previously documented in IL-4 KO animals. This residual Th2 response is likely to explain the ability of IL-4 KO mice to generate egg granulomas, which cannot be formed in IL-4Ralpha-deficient animals because of their lack of responsiveness to the same cytokine ligands. Taken together, these findings argue that tissue pathology in schistosomiasis requires, in addition to egg-specific CD4+ lymphocytes, a previously unrecognized IL-4Ralpha+ non-T cell effector population.

Optimal vaccination against Schistosoma mansoni requires the induction of both B cell- and IFN-gamma-dependent effector mechanisms.

Mice immunized with radiation-attenuated cercariae of Schistosoma mansoni display resistance to challenge infection, which increases with multiple boosting. Protection in animals receiving a single vaccination is thought to involve a primarily cell-mediated, IFN-gamma-dependent mechanism, while humoral immunity has been shown to contribute to challenge rejection in multiply (three times) immunized mice. To better understand the respective contribution of the B lymphocyte- and IFN-gamma-dependent effector arms in host resistance, we compared vaccine-induced immunity in B cell-deficient (muMT) and IFN-gamma knockout (GKO) animals. Unexpectedly, after a single vaccination, B cell knockout (KO) mice displayed reduced protection against challenge infection, although they developed a normal IFN-gamma-dominated cytokine response. This defect in resistance was equivalent to that displayed by GKO animals. Moreover, whereas two additional vaccinations significantly increased the level of immunity in wild-type mice, the protection in B cell KO animals remained unchanged. In contrast, multiple vaccination resulted in increased but, nevertheless, defective resistance in GKO mice. Since FcR gamma KO mice, which lack functional FcgammaRI, FcgammaRIII, and FcepsilonRI, show no defects in vaccine-induced resistance after immunization either one or three times, the B cell-dependent mechanism of protection involved does not appear to require FcR signaling. Together, these findings indicate that effective vaccination against schistosomes depends on the simultaneous induction of both humoral and cell-mediated immunity, a conclusion that may explain the limited success of most subunit vaccine protocols designed to preferentially induce either B cell- or IFN-gamma-dependent protective mechanisms.

IFN-gamma, IL-12, and TNF-alpha are required to maintain reduced liver pathology in mice vaccinated with Schistosoma mansoni eggs and IL-12.

The development of hepatic fibrosis and portal hypertension is the principal cause of morbidity and mortality in schistosomiasis mansoni. Nevertheless, relatively little is known about the mechanisms that lead to excessive collagen deposition during infection with Schistosoma mansoni. In the murine model, infection leads to significant egg-induced granuloma formation, tissue eosinophilia, and hepatic fibrosis. The pathology has been linked to dominant type 2 cytokine expression, and our recent studies showed that sensitizing animals to egg Ags in combination with IL-12, before infection, led to a highly significant reduction in egg-induced immunopathology. In this study, we demonstrate that in contrast with egg/IL-12-sensitized animals that showed marked decreases in pathology, mice similarly sensitized but depleted of IFN-gamma, IL-12, or TNF-alpha at the time of egg laying developed granulomas that were similar to the non-IL-12-treated control group. Although all three anti-cytokine-treated groups exhibited a dominant type 1 response in lymph node cells restimulated ex vivo, the expression of type 2 cytokine mRNA was markedly restored at the site of granuloma formation, which suggests that all three cytokines are required to maintain the suppressed type 2 pattern. Moreover, egg/IL-12-sensitized mice depleted of IFN-gamma or IL-12 displayed a partial reduction in IFN-gamma production, suggesting that multiple type 1 cytokines were required to maintain polarized type 1 responses to chronic type 2-inducing stimuli. Together, these data reveal key roles for IFN-gamma, IL-12, and TNF-alpha in the protective effects mediated by this IL-12-based vaccine to prevent pathology.

Helicobacter hepaticus triggers colitis in specific-pathogen-free interleukin-10 (IL-10)-deficient mice through an IL-12- and gamma interferon-dependent mechanism.

Mice rendered deficient in interleukin-10 (IL-10) by gene targeting (IL-10(-/-) mice) develop chronic enterocolitis resembling human inflammatory bowel disease (IBD) when maintained in conventional animal facilities. However, they display a minimal and delayed intestinal inflammatory response when reared under specific-pathogen-free (SPF) conditions, suggesting the involvement of a microbial component in pathogenesis. We show here that experimental infection with a single bacterial agent, Helicobacter hepaticus, induces chronic colitis in SPF-reared IL-10(-/-) mice and that the disease is accompanied by a type 1 cytokine response (gamma interferon [IFN-gamma], tumor necrosis factor alpha, and nitric oxide) detected by restimulation of spleen and mesenteric lymph node cells with a soluble H. hepaticus antigen (Ag) preparation. In contrast, wild-type (WT) animals infected with the same bacteria did not develop disease and produced IL-10 as the dominant cytokine in response to Helicobacter Ag. Strong H. hepaticus-reactive antibody responses as measured by Ag-specific total immunoglobulin G (IgG), IgG1, IgG2a, IgG2b, IgG3, and IgA were observed in both WT and IL-10(-/-) mice. In vivo neutralization of IFN-gamma or IL-12 resulted in a significant reduction of intestinal inflammation in H. hepaticus-infected IL-10(-/-) mice, suggesting an important role for these cytokines in the development of colitis in the model. Taken together, these microbial reconstitution experiments formally establish that a defined bacterial agent can serve as the immunological target in the development of large bowel inflammation in IL-10(-/-) mice and argue that in nonimmunocompromised hosts IL-10 stimulated in response to intestinal flora is important in preventing IBD.

Inducible nitric oxide synthase-deficient mice develop enhanced type 1 cytokine-associated cellular and humoral immune responses after vaccination with attenuated Schistosoma mansoni cercariae but display partially reduced resistance.

High levels of nitric oxide (NO) are produced by inducible nitric oxide synthase (iNOS) in response to activating signals from Th1-associated cytokines and play an important role in cytotoxicity and cytostasis against many pathogenic microorganisms. In addition to its direct effector function, NO serves as a potent immunoregulatory factor. NO produced by gamma interferon-activated macrophages immobilizes and kills Schistosoma mansoni larvae, and several studies have indicated a role for this pathway in protective immunity against this parasite. The potential regulatory influence of NO in immunity to S. mansoni is less well understood. In this study, we have used iNOS-deficient mice to determine the role of NO in mice vaccinated with irradiated cercariae of S. mansoni. We show by enzyme-linked immunosorbent assay and reverse transcriptase PCR analysis that vaccinated iNOS-deficient mice develop exacerbated type 1 cytokine responses in the lungs, the site where resistance to infection is primarily manifested. In addition, parasite-specific immunoglobulin G2a (IgG2a) and IgG2b antibody responses were significantly increased in vaccinated iNOS-deficient animals and total IgE antibody levels in serum were decreased relative to those in wild-type controls. Surprisingly, since resistance in this vaccine model is largely Th1 dependent and since Th1-related cellular and humoral immune responses were found to be exacerbated in vaccinated iNOS-deficient mice, vaccine-elicited protective immunity against challenge infection was found to be reduced. These findings demonstrate that iNOS plays a paradoxical role in immunity to S. mansoni, both in the effector mechanism of resistance and in the down regulation of the type 1 cytokine response, which is ultimately required for NO production.

Failure of P strain mice to respond to vaccination against schistosomiasis correlates with impaired production of IL-12 and up-regulation of Th2 cytokines that inhibit macrophage activation.

In contrast to most inbred strains, P mice fail to develop significant resistance to Schistosoma mansoni infection as a result of vaccination with either radiation-attenuated cercariae or schistosome antigens plus Bacillus Calmette Guérin, and this failure correlates with defects in macrophage larvicidal activity. Supernatant fluids from antigen-treated in vitro cultures of splenocytes from vaccinated P mice demonstrate less macrophage stimulatory activity than do supernatants from cells of vaccine-responsive strains such as C57BL/6. This is not due either to diminished production of the macrophage-activating cytokine IFN-gamma by P mice, or to a lesser responsiveness of macrophages from P mice to activation by IFN-gamma. Rather, P splenocytes produce two-to threefold higher amounts of IL-4 and IL-10, cytokines which down-regulate the cytotoxic potential of IFN-gamma-treated macrophages. Thus, the macrophage-activating potential of cytokine preparations from vaccinated P mice can be completely recovered by in vitro treatment with antibodies to IL-4 or IL-10. Moreover, lower levels of IL-12, a cytokine involved in promoting development of Th1 responses, are produced by splenocytes from P mice as compared to C57BL/6 counterparts. These studies indicate that a genetic predisposition toward an impaired production of IL-12 and an increased production of down-regulatory Th2 cytokines correlate with low response to vaccination against S. mansoni.

IL-10 regulates liver pathology in acute murine Schistosomiasis mansoni but is not required for immune down-modulation of chronic disease.

We have used IL-10 gene knockout mice (IL-10T) to examine the role of endogenous IL-10 in the down-modulation of hepatic granuloma formation and lymphocyte responses that occurs in chronic infection with the helminth parasite Schistosoma mansoni. Although IL-10-deficient animals showed 20 to 30% mortality between 8 and 14 wk postinfection, they displayed no alterations in their susceptibility to infection and produced similar numbers of eggs as their wild-type littermates. The IL-10T mice displayed a significant increase in hepatic granuloma size at the acute stage of infection, which was associated with increased IFN-gamma, IL-2, IL-1beta, and TNF-alpha mRNA expression in liver and elevated Th1-type cytokine production by lymphoid cells. Despite developing an enhanced Th1-type cytokine response, the IL-10T mice showed no consistent decrease in their Th2-type cytokine profile. Surprisingly, although granulomatous inflammation was enhanced at the acute stage of infection, the livers of IL-10T mice displayed no significant increase in fibrosis and underwent normal immune down-modulation at the chronic stage of infection. Moreover, the down-modulated state could be induced in IL-10T mice by sensitizing the animals to schistosome eggs before infection, further demonstrating that the major down-regulatory mechanism is not dependent upon IL-10. We conclude that while IL-10 plays an important role in controlling acute granulomatous inflammation, it plays no essential role in the process of immune down-modulation in chronic schistosome infection.

CD4+ T cell-mediated granulomatous pathology in schistosomiasis is downregulated by a B cell-dependent mechanism requiring Fc receptor signaling.

The effector functions of CD4+ T lymphocytes are generally thought to be controlled by distinct populations of regulatory T cells and their soluble products. The role of B cells in the regulation of CD4-dependent host responses is less well understood. Hepatic egg granuloma formation and fibrosis in murine schistosomiasis are dependent on CD4+ lymphocytes, and previous studies have implicated CD8+ T cells or cross-regulatory cytokines produced by T helper (Th) lymphocytes as controlling elements of this pathologic process. In this report, we demonstrate that B cell-deficient (muMT) mice exposed to Schistosoma mansoni develop augmented tissue pathology and, more importantly, fail to undergo the spontaneous downmodulation in disease normally observed during late stages of infection. Unexpectedly, B cell deficiency did not significantly alter T cell proliferative response or cause a shift in the Th1/Th2 balance. Since schistosome-infected Fc receptor-deficient (FcR gamma chain knockout) mice display the same exacerbated egg pathology as that observed in infected muMT mice, the B cell- dependent regulatory mechanism revealed by these experiments appears to require receptor-mediated cell triggering. Together, the data demonstrate that humoral immune response/FcR interactions can play a major role in negatively controlling inflammatory disease induced by CD4+ T cells.

Adsorption to aluminum hydroxide promotes the activity of IL-12 as an adjuvant for antibody as well as type 1 cytokine responses to HIV-1 gp120.

A series of protocols were tested to examine the adjuvant effects of IL-12 on humoral and type 1 cytokine responses elicited in mice by recombinant gp120 envelope protein from HIV-1. This Ag fails to induce detectable Ab responses when administered s.c. alone, but stimulates low Ab levels when combined with aluminum hydroxide (alum). Moreover, when i.p. injected rIL-12 was included in the immunization, no increase in Ab production was observed. Importantly, optimal gp120 Ab responses were achieved by immunizing mice s.c. with gp120 and rIL-12 simultaneously coadsorbed to alum. These animals displayed a highly polarized, type 1 cytokine profile, with the emergence of anti-gp120 Ig belonging to the IgG2 and IgG3 isotypes. In addition, a major increase occurred in Ab of the IgG1 subclass. The superior adjuvant activity of alum-adsorbed IL-12 compared with that of the free cytokine correlated with the prolonged detection of IFN-gamma in the sera of animals immunized using the former procedure. In related experiments, in vitro neutralization of IL-12 was shown to inhibit IFN-gamma production by spleen cells from mice immunized with gp120 plus alum, but not by splenocytes from mice primed in the presence of IL-12, suggesting that the latter protocol induces a stable type 1 phenotype. These studies demonstrate that presentation of IL-12 on alum enhances its immunomodulatory effects and establish a protocol for the use of the cytokine as an adjuvant for simultaneously promoting both humoral Ab and type 1 cytokine responses.

Fc epsilonRI-deficient mice infected with Schistosoma mansoni mount normal Th2-type responses while displaying enhanced liver pathology.

The IgE/Fc epsilonRI interaction is postulated to play an important role in resistance to helminths both at the level of anti-parasitic effector cell function and in the initiation of Th2 responses through IL-4 produced by Fc epsilonRI+ non-B, non-T (NBNT) cells. To formally evaluate the role of IgE/Fc epsilonRI signaling in the host response to helminths we studied Schistosoma mansoni infection in Fc epsilonRI knockout (KO) mice. Infected wild-type (wt) and KO animals showed comparable adult worm and tissue egg burdens, arguing against a role for Fc epsilonRI interactions in host resistance. Significantly, NBNT cells from infected KO, in contrast to wt animals, did not secrete IL-4 when stimulated with anti-IgE Ab or soluble parasite Ag. Nevertheless, serum IgE levels and Th2 cytokine production profiles were comparable in both strains of mice, demonstrating that the Ag-dependent stimulation of IL-4 secretion by NBNT cells is not essential for helminth-induced Th2 differentiation. However, when stimulated with low Ag doses, splenocytes from infected Fc epsilonRI-deficient mice produced less IL-4 in vitro than similar cultures from infected wt animals, an effect attributable to their defective NBNT cell function. Moreover, infected KO mice showed enhanced egg granuloma formation and hepatic fibrosis, revealing that the IgE/Fc epsilonRI interaction, while not essential for Th2 response development or resistance to primary infection, plays a significant role in down-regulating host pathology.

Perforin-mediated cytolysis plays a limited role in host resistance to Toxoplasma gondii.

Resistance of perforin knockout (PKO) mice to infection with Toxoplasma gondii was assessed in models of acute infection and during chronic disease. PKO mice vaccinated with the attenuated mutant, ts-4, displayed severely defective CTL responses against tachyzoite-infected targets. Lysis of the NK target, YAC-1, was also severely impaired in PKO mice following ts-4 vaccination. In contrast, wild-type mice developed high levels of CTL and NK lytic activity after ts-4 vaccination. Despite severely defective lytic activity, vaccinated PKO animals were completely resistant to challenge with the virulent strain RH, which normally causes a lethal acute infection. Resistance was attributable to production of IFN-gamma, which remained unimpaired in the PKO animals. In contrast, when PKO mice were infected with low virulence parasite strain ME49, which progresses to the cyst-forming stage after passage through an acute phase, accelerated mortality was observed beginning at 75 days postinfection. A three- to fourfold increase in brain cyst numbers was also found by day 30 in infected PKO animals. Nevertheless, the PKO strain produced normal levels of IFN-gamma after ME49 infection, ruling out impaired production of the latter cytokine as a cause of increased susceptibility. Together, these results show that perforin-dependent cytolytic function is not required for host resistance to lethal acute infection in preimmunized animals, but that the latter activity contributes to the control of infection during the chronic stage.

Unimpaired down-modulation of the hepatic granulomatous response in CD8 T-cell- and gamma interferon-deficient mice chronically infected with Schistosoma mansoni.

The granulomatous response to schistosome eggs is a CD4 T-cell-dependent, Th2-cytokine-dominated immunopathologic response. As infection proceeds to chronicity, both granuloma formation and egg-induced cytokine production become downregulated, and previous experiments have implicated CD8 T cells in this process. One mechanism by which CD8 T cells could suppress immunopathology is through the production of the counterregulatory cytokine gamma interferon (IFN-gamma), but no in vivo evidence exists to directly support this hypothesis. In this study, we analyzed hepatic granuloma formation and egg-induced cytokine production in Schistosoma mansoni-infected gene knockout mice deficient in either CD8 lymphocytes or IFN-gamma. Surprisingly, we found that neither immunologic component plays an essential function in the control of granuloma and cytokine responses during either the acute or chronic stage of infection. Thus, other mechanisms may be more important in the regulation of immunopathology in schistosomiasis.

Developmental differences determine larval susceptibility to nitric oxide-mediated killing in a murine model of vaccination against Schistosoma mansoni.

A persistent paradox in our understanding of protective immunity against Schistosoma mansoni infection in animals vaccinated with attenuated parasites has been that attrition of challenge parasites occurs during migration through the lungs in vivo, although parasites recovered from the lungs appear to be relatively resistant to cytotoxic effector mechanisms in vitro. We have compared the susceptibilities of different stages of larvae to killing by nitric oxide (NO), which was previously shown to be involved in the larvicidal function of cytokine-activated cytotoxic effector cells. Lung-stage larvae obtained 1 week after infection were not killed in vitro by NO generated either by a chemical NO donor or by activated cells. In contrast, parasites obtained from the portal system of control mice or from the lungs of vaccinated mice 2.5 weeks following challenge infection were killed by NO. As previously shown for mammalian cell targets, the effects of NO in susceptible larval stages may involve enzymes required for aerobic energy metabolism, since similar cytotoxicity was demonstrated by chemical inhibitors of the citric acid cycle or mitochondrial respiration. Taken together with previous observations of enhanced Th1 activity and expression of NO synthase in the lungs of vaccinated mice at 2.5 weeks after challenge infection, these observations elucidate the immune mechanism of vaccine-induced resistance to S. mansoni infection. Moreover, they suggest that conversion to a less metabolically active state may allow pathogens to escape the effects of the important effector molecule NO.