Immune Response of Cattle Infected with African Trypanosomes

Trypanosomosis is the most economically important disease constraint to livestock productivity in sub-Saharan Africa and has significant negative impact in other parts of the world. Livestock are an integral component of farming systems and thus contribute significantly to food and economic security in developing countries. Current methods of control for trypanosomosis are inadequate to prevent the enormous socioeconomic losses resulting from this disease. A vaccine has been viewed as the most desirable control option. However, the complexity of the parasite's antigenic repertoire made development of a vaccine based on the variable surface glycoprotein coat unlikely. As a result, research is now focused on identifying invariant trypanosome components as potential targets for interrupting infection or infection-mediated disease. Immunosuppression appears to be a nearly universal feature of infection with African trypanosomes and thus may represent an essential element of the host-parasite relationship, possibly by reducing the host's ability to mount a protective immune response. Antibody, T cell and macrophage/monocyte responses of infected cattle are depressed in both trypanosusceptible and trypanotolerant breeds of cattle. This review describes the specific T cell and monocyte/macrophage functions that are altered in trypanosome-infected cattle and compares these disorders with those that have been described in the murine model of trypanosomosis. The identification of parasite factors that induce immunosuppression and the mechanisms that mediate depressed immune responses might suggest novel disease intervention strategies.

Whole blood assay to access T cell-immune responses to Mycobacterium tuberculosis antigens in healthy Brazilian individuals

The production of interferon gamma (IFNgamma) guarantees effective T cell-mediated immunity against Mycobacterium tuberculosis infection. In the present study, we simply compare the in vitro immune responses to Mycobacterium antigens in terms of IFNg production in a total of 10 healthy Brazilian volunteers. Whole blood and mononuclear cells were cultivated in parallel with PPD, Ag85B, and M. bovis hsp65, and five-days supernatants were harvested for cytokine detection by ELISA. The inter-assay result was that the overall profile of agreement in response to antigens was highly correlated (r² = 0.9266; p = 0.0102). Potential analysis is in current progress to dictate the usefulness of this method to access the immune responses also in tuberculosis patients and its contacts.

Schistosoma mansoni infection modulates the immune response against allergic and auto-immune diseases

Chronic Schistosoma mansoni infection leads to a type 2-immune response with increased production of interleukin (IL-10). Evidence indicates chronic exposure to S. mansoni down regulates the type 1 immune response and prevents the onset of Th1-mediated diseases such as multiple sclerosis, diabetes mellitus and Cronh's disease. Furthermore, our own studies have revealed that chronic exposure to S. mansoni also down regulates atopic disease, Th2-mediated diseases. Our studies show an inverse association between the skin prick test reactivity and infection with S. mansoni and show the severity of asthma is reduced in subjects living in an endemic area of S. mansoni. Moreover, we hypothesize the mechanisms involved in the modulation of inflammatory response in atopic individuals, is likely dependent on IL-10 production, an anti-inflammatory cytokine elevated during helminth infections. Patients with asthma and helminth infections produced less IL-5 than patients with asthma without helminth infections, and this down regulation could, in part, be mediated by IL-10. In conclusion, helminthic infections, through induction of regulatory mechanisms, such as IL-10 production, are able to modulate the inflammatory immune response involved in the pathology of auto-immune and allergic disease.

Improving the production of the denatured recombinant N-terminal domain of rhoptry-associated protein 2 from a Plasmodium falciparum target in the pathology of anemia in falciparum malaria

Rhoptry-associated protein 2 (RAP2) is known to be discharged from rhoptry onto the membrane surface of infected and uninfected erythrocytes (UEs) ex vivo and in vitro and this information provides new insights into the understanding of the pathology of severe anemia in falciparum malaria. In this study, a hexahistidine-tagged recombinant protein corresponding to residues 5-190 of the N-terminal of Plasmodium falciparum RAP2 (rN-RAP2) was produced using a new method of solubilization and purification. Expression was induced with D-lactose, a less expensive alternative inducer to the more common isopropyl-²-D-thio-galactopyranosidase. The recombinant protein was purified using two types of commercially-available affinity columns, iminodiacetic and nitrilotriacetic. rN-RAP2 had immunogenic potential, since it induced high titers of anti-RAP2 antibodies in mice. These antibodies recognized full-length RAP2 prepared from Triton X-100 extracts from two strains of P. falciparum. In fact, the antibody recognized a 29-kDa product of RAP2 cleavage as well as 82 and 70-kDa products of RAP1 cleavage. These results indicate that the two antigens share sequence epitopes. Our expressed protein fragment was shown to contain a functional epitope that is also present in rhoptry-derived ring surface protein 2 which attaches to the surface of both infected and UEs and erythroid precursor cells in the bone marrow of malaria patients. Serum from malaria patients who developed anemia during infection recognized rN-RAP2, suggesting that this protein fragment may be important for epidemiological studies investigating whether immune responses to RAP2 exacerbate hemolysis in falciparum malaria patients.

The effects of nitric oxide on the immune system during Trypanosoma cruzi infection

Trypanosoma cruzi infection triggers substantial production of nitric oxide (NO), which has been shown to have protective and toxic effects on the host's immune system. Sensing of trypomastigotes by phagocytes activates the inducible NO-synthase (NOS2) pathway, which produces NO and is largely responsible for macrophage-mediated killing of T. cruzi. NO is also responsible for modulating virtually all steps of innate and adaptive immunity. However, NO can also cause oxidative stress, which is especially damaging to the host due to increased tissue damage. The cytokines IFN-³ and TNF-±, as well as chemokines, are strong inducers of NOS2 and are produced in large amounts during T. cruzi acute infection. Conversely, TGF-² and IL-10 negatively regulate NO production. Here we discuss the recent evidence describing the mechanisms by which NO is able to exert its antimicrobial and immune regulatory effects, the mechanisms involved in the oxidative stress response during infection and the implications of NO for the development of therapeutic strategies against T. cruzi.

Schistosomiasis differentially affects vasoconstrictor responses: up-regulation of 5-HT receptor-mediated aorta contraction

Schistosomiasis, classified by the World Health Organization as a neglected tropical disease, is an intravascular parasitic disease associated to a chronic inflammatory state. Evidence implicating inflammation in vascular dysfunction continues to mount, which, broadly defined, reflects a failure in the control of intracellular Ca2+ and consequently, vascular contraction. Therefore, we measured aorta contraction induced by 5-hydroxytryptamine (5-HT) and endothelin-1 (ET-1), two important regulators of vascular contraction. Isometric aortic contractions were determined in control and Schistosoma mansoni-infected mice. In the infected animals, 5-HT induced a 50% higher contraction in relation to controls and we also observed an increased contraction in response to Ca2+ mobilisation from sarcoplasmic reticulum. Nevertheless, Rho kinase inhibition reduced the contraction in response to 5-HT equally in both groups, discarding an increase of the contractile machinery sensitivity to Ca2+. Furthermore, no alteration was observed for contractions induced by ET-1 in both groups. Our data suggest that an immune-vascular interaction occurs in schistosomiasis, altering vascular contraction outside the mesenteric portal system. More importantly, it affects distinct intracellular signalling involved in aorta contraction, in this case increasing 5-HT receptor signalling.

Induction of cell-mediated immunity during early stages of infection with intracellular protozoa

Toxoplasma gondii and Trypanosoma cruzi are intracellular parasites which, as part of their life cycle, induce a potent cell-mediated immunity (CMI) maintained by Th1 lymphocytes and IFN-g. In both cases, induction of a strong CMI is thought to protect the host against rapid parasite multiplication and consequent pathology and lethality during the acute phase of infection. However, the parasitic infection is not eliminated by the immune system and the vertebrate host serves as a parasite reservoir. In contrast, Leishmania sp, which is a slow growing parasite, appears to evade induction of CMI during early stages of infection as a strategy for surviving in a hostile environment (i.e., inside the macrophages which are their obligatory niche in the vertebrate host). Recent reports show that the initiation of IL-12 synthesis by macrophages during these parasitic infections is a key event in regulating CMI and disease outcome. The studies reviewed here indicate that activation/inhibition of distinct signaling pathways and certain macrophage functions by intracellular protozoa are important events in inducing/modulating the immune response of their vertebrate hosts, allowing parasite and host survival and therefore maintaining parasite life cycles.

Precipitated immune complexes of IgM induce the generation of reactive oxygen species by rabbit polymorphonuclear leucocytes

We report that immune complexes of IgM (ICIgM) antibodies and ovalbumin in the form of a precipitate from the equivalence zone induce the generation of reactive oxygen species by rabbit blood polymorphonuclear leucocytes (PMN), as measured by the chemiluminescence (CL) production in the presence of luminol. The kinetics of CL generation induced by ICIgM is quite different from that induced by precipitated immune complexes of IgG (ICIgG): the maximum rate of CL production for ICIgM occurs around 14 min, whereas for ICIgG it occurs about 5 min after incubation with the cells. Also the triggering of the process requires a higher concentration of ICIgM than of ICIgG. Evidence is presented that these effects are not mediated by interaction of the antigen (ovalbumin) with the cell, since immune precipitates of ovalbumin and the F(ab')2 fragment had no effect. Our observations that precipitated ICIgM can also be an effective stimulus for CL generation and thus for O2- production reveal a new functional capability of PMN. These results may have implications for the understanding of the participation of ICIgM (as well as of ICIgG) in inflammatory reactions mediated by PMN in immune complex diseases, and in the mechanisms of defense against microbes and other non-self agents.

High frequency of the CCR5delta32 variant among individuals from an admixed Brazilian population with sickle cell anemia

Homozygous sickle cell disease (SCD) has a wide spectrum of clinical manifestations. In Brazil, the main cause of death of individuals with SCD is recurrent infection. The CCR5delta32 allele, which confers relative resistance to macrophage-tropic HIV virus infection, probably has reached its frequency and world distribution due to other pathogens that target macrophage in European populations. In the present investigation a relatively higher prevalence (5.1%) of the CCR5delta32 allele was identified, by PCR amplification using specific primers, in 79 SCD patients when compared to healthy controls (1.3%) with the same ethnic background (Afro-Brazilians). Based on a hypothesis that considers SCD as a chronic inflammatory condition, and since the CCR5 chemokine receptor is involved in directing a Th1-type immune response, we suggest that a Th1/Th2 balance can influence the morbidity of SCD. If the presence of the null CCR5delta32 allele results in a reduction of the chronic inflammation state present in SCD patients, this could lead to differential survival of SCD individuals who are carriers of the CCR5delta32 allele. This differential survival could be due to the development of less severe infections and consequently reduced or less severe vaso-occlusive crises.

Changes in red blood cell osmotic fragility induced by total plasma and plasma fractions obtained from rats bearing progressive and regressive variants of the Walker 256 tumor

Two variants (A and B) of the widely employed Walker 256 rat tumor cells are known. When inoculated sc, the A variant produces solid, invasive, highly metastasizing tumors that cause severe systemic effects and death. We have obtained a regressive variant (AR) whose sc growth is slower, resulting in 70-80% regression followed by development of immunity against A and AR variants. Simultaneously with the beginning of tumor regression, a temporary anemia developed (~8 days duration), accompanied by marked splenomegaly (~300%) and changes in red blood cell osmotic fragility, with mean corpuscular fragility increasing from 4.1 to 6.5 g/l NaCl. The possibility was raised that plasma factors associated with the immune response induced these changes. In the present study, we identify and compare the osmotic fragility increasing activity of plasma fractions obtained from A and AR tumor bearers at different stages of tumor development. The results showed that by day 4 compounds precipitating in 60% (NH4)2SO4 and able to increase red blood cell osmotic fragility appeared in the plasma of A and AR tumor bearers. Later, these compounds disappeared from the plasma of A tumor bearers but slightly increased in the plasma of AR tumor bearers. Furthermore, by day 10, compounds precipitating between 60 and 80% (NH4)2SO4 and with similar effects appeared only in plasma of AR tumor bearers. The salt solubility, production kinetics and hemolytic activity of these compounds resemble those of the immunoglobulins. This, together with their preferential increase in rats bearing the AR variant, suggest their association with an immune response against this tumor.

A model of chronic IgE-mediated food allergy in ovalbumin-sensitized mice

Food allergy is most frequently the result of IgE-mediated hypersensitivity reactions. Here, we describe a chronic model in which some of the intestinal and systemic consequences of continuous egg white solution ingestion by ovalbumin-sensitized eight-week-old BALB/c mice, 6 animals per group, of both sexes, were investigated. There was a 20% loss of body weight that began one week after antigen exposure and persisted throughout the experiment (3 weeks). The sensitization procedure induced the production of anti-ovalbumin IgG1 and IgE, which were enhanced by oral antigen exposure (129% for IgG1 and 164% for IgE, compared to sensitization values). Intestinal changes were determined by jejunum edema at 6 h (45% Evans blue extravasation) and by a significant eosinophil infiltration with a peak at 48 h. By day 21 of continuous antigen exposure, histological findings were mild, with mast cell hyperplasia (100%) and increased mucus production (483%). Altogether, our data clearly demonstrate that, although immune stimulation was persistently occurring in response to continuous oral antigen exposure, regulatory mechanisms were occurring in the intestinal mucosa, preventing overt pathology. The experimental model described here reproduces the clinical and pathological changes of mild chronic food allergy and may be useful for mechanistic studies of this common clinical condition.

Secondary prevention of type 1 diabetes mellitus: stopping immune destruction and promoting ß-cell regeneration

Type 1 diabetes mellitus results from a cell-mediated autoimmune attack against pancreatic ß-cells. Traditional treatments involve numerous daily insulin dosages/injections and rigorous glucose control. Many efforts toward the identification of ß-cell precursors have been made not only with the aim of understanding the physiology of islet regeneration, but also as an alternative way to produce ß-cells to be used in protocols of islet transplantation. In this review, we summarize the most recent studies related to precursor cells implicated in the regeneration process. These include embryonic stem cells, pancreas-derived multipotent precursors, pancreatic ductal cells, hematopoietic stem cells, mesenchymal stem cells, hepatic oval cells, and mature ß-cells. There is controversial evidence of the potential of these cell sources to regenerate ß-cell mass in diabetic patients. However, clinical trials using embryonic stem cells, umbilical cord blood or adult bone marrow stem cells are under way. The results of various immunosuppressive regimens aiming at blocking autoimmunity against pancreatic ß-cells and promoting ß-cell preservation are also analyzed. Most of these regimens provide transient and partial effect on insulin requirements, but new regimens are beginning to be tested. Our own clinical trial combines a high dose immunosuppression with mobilized peripheral blood hematopoietic stem cell transplantation in early-onset type 1 diabetes mellitus.

Combating oncogene activation associated with retrovirus-mediated gene therapy of X-linked severe combined immunodeficiency

A successful gene therapy clinical trial that also encountered serious adverse effects has sparked extensive study and debate about the future directions for retrovirus-mediated interventions. Treatment of X-linked severe combined immunodeficiency with an oncoretrovirus harboring a normal copy of the gc gene was applied in two clinical trials, essentially curing 13 of 16 infants, restoring a normal immune system without the need for additional immune-related therapies. Approximately 3 years after their gene therapy, tragically, 3 of these children, all from the same trial, developed leukemia as a result of this experimental treatment. The current understanding of the mechanism behind this leukemogenesis involves three critical and cooperating factors, i.e., viral integration, oncogene activation, and the function of the therapeutic gene. In this review, we will explore the causes of this unwanted event and some of the possibilities for reducing the risk of its reoccurrence.

Attenuation of phosphamidon-induced oxidative stress and immune dysfunction in rats treated with N-acetylcysteine

The effect of N-acetylcysteine, a thiolic antioxidant, on attenuation of phosphamidon-induced oxidative stress and immune dysfunction was evaluated in adult male Wistar rats weighing 200-250 g. Rats were divided into four groups, 8 animals/group, and treated with phosphamidon, N-acetylcysteine or the combination of both for 28 days. Oral administration of phosphamidon (1.74 mg/kg), an organophosphate insecticide, increased serum malondialdehyde (3.83 ± 0.18 vs 2.91 ± 0.24 nmol/mL; P < 0.05) and decreased erythrocyte superoxide dismutase (567.8 ± 24.36 vs 749.16 ± 102.61 U/gHb; P < 0.05), catalase activity (1.86 ± 0.18 vs 2.43 ± 0.08 U/gHb; P < 0.05) and whole blood glutathione levels (1.25 ± 0.21 vs 2.28 ± 0.08 mg/gHb; P < 0.05) showing phosphamidon-induced oxidative stress. Phosphamidon exposure markedly suppressed humoral immune response as assessed by antibody titer to ovalbumin (4.71 ± 0.51 vs 8.00 ± 0.12 -log2; P < 0.05), and cell-mediated immune response as assessed by leukocyte migration inhibition (25.24 ± 1.04 vs 70.8 ± 1.09%; P < 0.05) and macrophage migration inhibition (20.38 ± 0.99 vs 67.16 ± 5.30%; P < 0.05) response. Phosphamidon exposure decreased IFN-у levels (40.7 ± 3.21 vs 55.84 ± 3.02 pg/mL; P < 0.05) suggesting a profound effect of phosphamidon on cell-mediated immune response. A phosphamidon-induced increase in TNF-α level (64.19 ± 6.0 vs 23.16 ± 4.0 pg/mL; P < 0.05) suggests a contributory role of immunocytes in oxidative stress. Co-administration of N-acetylcysteine (3.5 mmol/kg, orally) with phosphamidon attenuated the adverse effects of phosphamidon. These findings suggest that oral N-acetylcysteine treatment exerts protective effect and attenuates free radical injury and immune dysfunction caused by subchronic phosphamidon exposure.

Evasion of immune responses by Trypanosoma cruzi, the etiological agent of Chagas disease

Infection with the protozoan parasite Trypanosoma cruzi leads to Chagas disease, which affects millions of people in Latin America. Infection with T. cruzi cannot be eliminated by the immune system. A better understanding of immune evasion mechanisms is required in order to develop more effective vaccines. During the acute phase, parasites replicate extensively and release immunomodulatory molecules that delay parasite-specific responses mediated by T cells. This immune evasion allows the parasite to spread in the host. In the chronic phase, parasite evasion relies on its replication strategy of hijacking the TGF-β signaling pathway involved in inflammation and tissue regeneration. In this article, the mechanisms of immune evasion described for T. cruzi are reviewed.