Antigens from Rhipicephalus sanguineus ticks elicit potent cell-mediated immune responses in resistant but not in susceptible animals

In the present study we compared the immunological reactions between Rhipicephalus sanguineus tick-infested susceptible (dogs and mice) and tick-resistant hosts (guinea pigs), elucidating some of the components of efficient protective responses against ticks. We found that T-cells from guinea pigs infested with adult ticks proliferate vigorously in the presence of concanavalin A (ConA), whereas ConA-induced cell proliferation of tick-infested mice and dogs was significantly decreased at 43.1 and 94.0%, respectively, compared to non-infested controls. Moreover, cells from mice and dogs submitted to one or three successive infestations did not exhibit a T-cell proliferative response to tick antigens, whilst cells from thrice tick-infested guinea pigs, when cultured with either a tick extract or tick saliva, displayed a significant increase in cell proliferation. Also, we evaluated the response of tick-infested mice to a cutaneous hypersensitivity test induced by a tick extract. Tick-infested mice developed a significant immediate reaction, whereby a 29.9% increase in the footpad thickness was observed. No delayed-type hypersensitivity (DTH) reaction was detected. Finally, the differential cell count at the tick attachment site in repeatedly infested mice exhibited a 6.6- and 4.1-fold increase in the percentage of eosinophils and neutrophils, respectively, compared to non-infested animals, while a decrease of 77.0-40.9 in the percentage of mononuclear cells was observed. The results of the cutaneous hypersensitivity test and the cellular counts at the tick feeding site for mice support the view that tick-infested mice develop an immune response to R. sanguineus ticks very similar to dogs, the natural host of this species of tick, but very different from guinea pigs (resistant host), which develop a DTH reaction in addition to a basophil and mononuclear cell infiltration at the tick-attachment site. In conclusion, saliva introduced during tick infestations reduces the ability of a susceptible animal host to respond to tick antigens that could stimulate a protective immune response. As a consequence, the animals present a lack of DTH response and disturbed cellular migration to tick feeding site, which can represent a deficient response against ticks. © 2003 Elsevier B.V. All rights reserved.

Immune regulatory effect of pHSP65 DNA therapy in pulmonary tuberculosis: Activation of CD8+ cells, interferon-γ recovery and reduction of lung injury

A DNA vaccine based on the heat-shock protein 65 Mycobacterium leprae gene (pHSP65) presented a prophylactic and therapeutic effect in an experimental model of tuberculosis. In this paper, we addressed the question of which protective mechanisms are activated in Mycobacterium tuberculosis-infected mice after immune therapy with pHSP65. We evaluated activation of the cellular immune response in the lungs of infected mice 30 days after infection (initiation of immune therapy) and in those of uninfected mice. After 70 days (end of immune therapy), the immune responses of infected untreated mice, infected pHSP65-treated mice and infected pCDNA3-treated mice were also evaluated. Our results show that the most significant effect of pHSP65 was the stimulation of CD8+ lung cell activation, interferon-γ recovery and reduction of lung injury. There was also partial restoration of the production of tumour necrosis factor-α. Treatment with pcDNA3 vector also induced an immune stimulatory effect. However, only infected pHSP65-treated mice were able to produce significant levels of interferon-γ and to restrict the growth of bacilli.

Immune Response to the Rhodococcus equi infection in high and low antibody-producing mice (selection IV-A)

Rhodococcus equi is a Gram-positive, facultative intracellular bacterium which infects macrophages and causes rhodococcal pneumonia and enteritis in foals. Recently, this agent has been recognized as an opportunistic pathogen for immunocompromised humans. Several murine experimental models have been used to study R. equi infection. High (H IV-A) and Low (L IV-A) antibody (Ab)-producers mice were obtained by bi-directional genetic selections for their ability to produce antibodies against sheep and human erythrocytes (Selection IV-A). These lines maintain their phenotypes of high and low responders also for other antigens than those of selection (multispeciflc effect). A higher macrophage activity in L IV-A mice has been described for several intracellular infectious agents, which could be responsible for their intense macrophage antigens (Ag)-handling and low Ab production. Due to these differences, L IV-A mice were found to exhibit a better performance to trigger an effective immune response towards intracellular pathogens. The objective of this work was to characterize the immune response of Selection IV-A against R. equi. H IV-A and L IV-A mice were infected with 2.0 × 10 6 CFU of ATCC 33701 +R. equi by intravenous route. With regards to bacterial clearance and survival assays, L IV-A mice were more resistant than H IV-A mice to virulent R. equi. L IV-A mice presented a higher hydrogen peroxide (H 2O 2) and nitric oxide (NO) endogenous production by splenic macrophages than H IV-A mice. L IV-A expressed the most intense cellular response, available by the Delayed-Type Hypersensitivity (DTH) reaction, which activated macrophages and produced more H 2O 2 and NO. The three times higher specific antibodies titres in H IV-A indicated that Selection IV-A maintained the multispecific effect and the polygenic control of humoral and cellular responses also to R. equi.

Immune reconstitution in HIV-1 infected patients treated for two years with highly active antiretroviral therapy

The aim of this paper was to evaluate the immune reconstitution of HIV-1 patients subjected to highly active antiretroviral therapy (HAART) for two years or more according to CD 45RA and CD 45RO cell count; determination of IL-2, IFN-γ, IL-4, IL-10 and TNF-α serum levels; CD 4 + T and CD 8 + T lymphocyte count; and plasma viral load (VL) determination. For this purpose, a cross sectional study was carried out in the Tropical Diseases Area, Botucatu School of Medicine, São Paulo State University, UNESP, Botucatu, São Paulo, Brazil. Between June 2001 and April 2002, 37 HIV-1 infected patients were evaluated, 13 with treatment indication but untreated (G1), 9 subjected to HAART for 5-7 months (G2), and 15 treated for two years or more (G3); both treated groups used medication regularly and without failure. Forty-nine normal individuals were studied as controls (GC-1 and GC-2). There was a tendency (p<0.10) for the predominance of two nucleoside reverse transcriptase inhibitors (NRTI) associated with one non-nucleoside reverse transcriptase inhibitor (NNRTI) regimen in G2; and two NRTI associated with a protease inhibitor (PI) in G3. Statistical differences between groups were seen for CD 45RA (G1<[G3=GC-2]; p<0.05) and CD 45RO (G1[G2=G3]; p<0.001), TNF-α serum determination ([G1>G3; G2=intermediate]>GC-1; p<0.001), IL-2 (G1<[G2=G3=GC-1]; p<0.01), IFN-γ ([G1=GC-1]>[GC-2=G3]; p<0.001), IL-4 and IL-10 ([G1=G2=G3]>GC-1; p<0.001), serum cytokine profiles, with a higher proportion of subtype 2 in G1 and mature subtype 0 in G2 and G3 (p<0.005). There was no statistical difference for CD 8 + T lymphocyte counts (G1=G2=G3; p<0.50). Consistency was seen between positive correlations of profile 1 definer cytokines (IL-2 and IFN-γ), CD 45RA and CD 45RO cells, and CD 4 + T lymphocyte counts and between positive correlations of profile 2 definer cytokines (IL-4 and IL-10) with TNF-α, and VL. The negative correlations were also consistent as they expressed the inverse of the positives. The variables with the highest number of correlations were IL-2, IFN-γ, and VL, followed by CD 45RA and CD 45RO cells, and IL-10. The variables with the lowest number of correlations were CD 4 + T and CD 8 + T lymphocytes. The results express the partial but important immune reconstitution in HIV-1 infected individuals with the interference of HAART and the importance of cytokines especially IL-2 and IFN-γ, and CD 45RA and CD 45RO cells as surrogate markers of this reconstitution.

Artificial immune systems applied to optimal capacitor placement in radial distribution networks

The capacitor placement problem for radial distribution networks aims to determine capacitor types, sizes, locations and control scheme. This is a combinatorial problem that can be formulated as a mixed integer nonlinear program. The paper presents an algorithm inspired in artificial immune systems and developed for this specific problem. A good performance was obtained through experimental tests applied to known systems. © 2006 IEEE.

Pro- and anti-inflammatory cytokines produced by human monocytes challenged in vitro with Paracoccidioides brasiliensis

Monocytes and macrophages play a central role in innate and adaptive immune response against systemic fungal infections. Imbalances in suppressor or stimulatory cytokine secretion caused by these cells may influence disease development, microorganism death, and the nature of the adaptive immune response. This study analyzed the monocyte cytokine profiles of healthy individuals challenged with high and low virulent strains of P. brasiliensis and mRNA cytokine expression kinetics by reverse transcription polymerase chain reaction (RT-PCR). Peripheral blood monocytes from healthy volunteers were cultured in vitro with and without virulent (Pb18) or low virulence (Pb265) strains from P. brasiliensis viable yeast cells. Interleukin-1 beta (IL-1β), IL-6, IL-8, IL-10, tumor necrosis factor-alpha (TNF-α), and transforming growth factor-beta (TGF-β1) were measured in culture supernatants by enzyme immunoassay (ELISA), and mRNA cytokine expression was determined by RT-PCR at 0, 4, 8, 12, 18 and 48 hr. Both P. brasiliensis strains induced monocyte production of IL-1β, IL-6, IL-10 and TNF-α. Pb18 induced higher levels of IL-1β, IL-6, and IL-10 than Pb265. IL-8 and TGF-β1 levels were not significantly different from those cultured without stimulus. The mRNA cytokine expression was similar to supernatant cytokines measured by ELISA. In vitro monocyte challenge with virulent P. brasiliensis strain induces earlier and higher levels of pro- and anti-inflammatory cytokines than low virulence strain.

Kaposi's sarcoma-associated herpesvirus latency-associated nuclear antigen 1 mimics Epstein-Barr virus EBNA1 immune evasion through central repeat domain effects on protein processing

Kaposi's sarcoma-associated herpesvirus (KSHV/human herpesvirus 8 [HHV8]) and Epstein-Barr virus (EBV/HHV4) are distantly related gammaherpesviruses causing tumors in humans. KSHV latency-associated nuclear antigen 1 (LANA1) is functionally similar to the EBV nuclear antigen-1 (EBNA1) protein expressed during viral latency, although they have no amino acid similarities. EBNA1 escapes cytotoxic lymphocyte (CTL) antigen processing by inhibiting its own proteosomal degradation and retarding its own synthesis to reduce defective ribosomal product processing. We show here that the LANA1 QED-rich central repeat (CR) region, particularly the CR2CR3 subdomain, also retards LANA1 synthesis and markedly enhances LANA1 stability in vitro and in vivo. LANA1 isoforms have half-lives greater than 24 h, and fusion of the LANA1 CR2CR3 domain to a destabilized heterologous protein markedly decreases protein turnover. Unlike EBNA1, the LANA1 CR2CR3 subdomain retards translation regardless of whether it is fused to the 5′ or 3′ end of a heterologous gene construct. Manipulation of sequence order, orientation, and composition of the CR2 and CR3 subdomains suggests that specific peptide sequences rather than RNA structures are responsible for synthesis retardation. Although mechanistic differences exist between LANA1 and EBNA1, the primary structures of both proteins have evolved to minimize provoking CTL immune responses. Simple strategies to eliminate these viral inhibitory regions may markedly improve vaccine effectiveness by maximizing CTL responses. Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Immune response to Mycobacterium bovis-an5 infection in genetically selected mice (selection IV-A)

Mice genetically selected for high (H) and low (L) antibody production (Selection IV-A) were used as murine experimental model. The aim of the present work was to evaluate the macrophagic activity and to characterize the immune response in Mycobacterium bovis-AN5 infected mice (3×10 7 bacteria). The response profile previously observed in such strains was not similar to that obtained during M. bovis infection; however, it corroborated works carried out using Selection I, which is very similar to Selection IV-A regarding infection by M. tuberculosis and Bacillus Calmette-Guérin (BCG). Considering bacterial recovery, LIV-A mice showed higher control of the infectious process in the lungs than in the spleen, whereas HIV-A mice presented more resistance in the spleen. With respect to macrophagic activity, hydrogen peroxide (H2O 2) was probably not involved in the infection control since there was an inhibition in the production of this metabolite. Nitric oxide (NO) and TNF-α production seemed to be important in the control of bacterial replication and varied according to the strain, period and organ. Evaluation of the antibody production indicated that the multi-specific effect commonly observed in these strains was not the same in the response to M. bovis. Antibody concentrations were higher in LIV-A than in HIV-A mice at the beginning of the infection, being similar afterwards. Such data were compared with delayed-type hypersensitivity (DTH), which was more intense in HIV-A than in LIV-A mice, indicating that antibody production is independent of the capability to trigger DTH reactions and that cellular and humoral responses to M. bovis antigens show a polygenic control and an independent quantitative genetic regulation. Differences were observed among organs and metabolites, suggesting that different mechanisms play an important role in this infection in natural heterogeneous populations, indicating that NO, TNF-α and Th1 cytokines are involved in the infection control.

Genetic vaccine for tuberculosis (pVAXhsp65) primes neonate mice for a strong immune response at the adult stage

Background: Vaccination of neonates is generally difficult due to the immaturity of the immune system and consequent higher susceptibility to tolerance induction. Genetic immunization has been described as an alternative to trigger a stronger immune response in neonates, including significant Th1 polarization. In this investigation we analysed the potential use of a genetic vaccine containing the heat shock protein (hsp65) from Mycobacterium leprae (pVAXhsp65) against tuberculosis (TB) in neonate mice. Aspects as antigen production, genomic integration and immunogenicity were evaluated. Methods: Hsp65 message and genomic integration were evaluated by RT-PCR and Southern blot, respectively. Immunogenicity of pVAXhsp65 alone or combined with BCG was analysed by specific induction of antibodies and cytokines, both quantified by ELISA. Results: This DNA vaccine was transcribed by muscular cells of neonate mice without integration into the cellular genome. Even though this vaccine was not strongly immunogenic when entirely administered (three doses) during early animal's life, it was not tolerogenic. In addition, pVAXhsp65 and BCG were equally able to prime newborn mice for a strong and mixed immune response (Th1 + Th2) to pVAXhsp65 boosters administered later, at the adult life. Conclusion: These results suggest that pVAXhsp65 can be safely used as a priming stimulus in neonate animals in prime-boost similar strategies to control TB. However, priming with BCG or pVAXhsp65, directed the ensuing immune response triggered by an heterologous or homologous booster, to a mixed Th1/Th2 pattern of response. Measures as introduction of IL-12 or GM-CSF genes in the vaccine construct or even IL-4 neutralization, are probably required to increase the priming towards Th1 polarization to ensure control of tuberculosis infection. © 2007 Pelizon et al; licensee BioMed Central Ltd.

Citotoxicity and immune response induced by organopalladium(II) compounds in mice bearing Ehrlich ascites tumour

Cyclometallated palladium(II) complexes are reactive inorganic compounds employed in several biological studies because of their antitumour potential and interaction with immune system. In the present study, the immune and citotoxic response induced by two organopalladated complexes: [{Pd(N,C-dmba)} 2(μ-NCS) 2] (1), [Pd(C-dmba)(NCS)(dppp)] (2) [dmba = N,N′-dimethylbenzylamine, dppp = 1,3-bis(diphenylphosphino)propane] and cisplatin (cis-DDP), as standard, were investigated in mice bearing Ehrlich ascites tumour. The mice were divided into five groups and inoculated with the compounds (1) or (2) or cisplatin, or only vehicle or phosphate-buffered saline (PBS). Many parameters were evaluated, such as tumour cell percentage in the peritoneal exsudate, levels of seric nitric oxide (NO) and tumour necrosis factor-alpha (TNF-α) and increase in life span. Analysis of all data revealed, for compound (2), an activity similar to that presented by cisplatin, resulting in increased life span, lower levels of seric TNF-α and increase in NO production. ©2007 Sociedade Brasileira de Química.

Occurrence and expression of p53 suppressor gene and c-Myc oncogene in dog eyelid tumors

Purpose: To detect the occurrence and expression of the suppressor gene p53 and of the oncogene c-Myc in eyelid tumors of dogs using the PCR, RT-PCR, PCR-ELISA and RT-PCR-ELISA techniques. These genes have not been described in dog eyelid tumors before. Methods: Nine samples of eyelid or third eyelid epithelial tumors were obtained from the archives of the Department of Veterinary Pathology. Tumor diagnosis was confirmed by evaluation of hematoxylin-eosin stained sections, and immunohistochemistry for cytokeratin AE1/AE3 and vimentin V9. A canine mammary tumor was used for positive control. Agarose gel electrophoresis, PCR-ELISA and RT-PCR-ELISA were used to detect p53 and c-Myc genes. Results: The occurrence of p53 was detected in most of the eyelid tumors and third eyelid tumors studied (88.8%, n = 8) and was expressed in 75% of the positive samples, as indicated by ELISA. The c-Myc gene was found in 77.7% (n = 7) of the samples and was expressed in eight samples. Conclusions: Eyelid and third eyelid tumors of dogs express both the p53 and the c-Myc genes as shown by PCR and RT-PCR. However, PCR ELISA and RT-PCR ELISA were more efficient in assessing occurrence and expression of these genes because they identified amplified products that were not detected by agarose gel electrophoresis. © 2010 American College of Veterinary Ophthalmologists.

Modeling for performance analysis of electromagnetic zero-sequence suppressor

The purpose of this work is to present a frequency domain model to demonstrate the operation of an electromagnetic arrangement for controlling the injection of zero-sequence currents in the electrical system. Considering the diversity of sequential distribution of harmonic components of a current, the device proposed can be used in the process of mitigation of zero-sequence components. This device, here called electromagnetic suppressor, consists of a blocker and filter both electromagnetic, whose joint operation can provide paths of high and low impedances that can be conveniently adjusted in order to search for a desired performance. This study presents physical considerations, mathematical modeling and computer simulations that clearly demonstrate the viability of this application as a more viable alternative in the conception of filtering systems. The performance analysis is based on the frequency response of harmonic transmittances. The efficacy of this technique in direct actions to maximize the harmonic mitigation process is demonstrated. ©2010 IEEE.

A malware detection system inspired on the human immune system

Malicious programs (malware) can cause severe damage on computer systems and data. The mechanism that the human immune system uses to detect and protect from organisms that threaten the human body is efficient and can be adapted to detect malware attacks. In this paper we propose a system to perform malware distributed collection, analysis and detection, this last inspired by the human immune system. After collecting malware samples from Internet, they are dynamically analyzed so as to provide execution traces at the operating system level and network flows that are used to create a behavioral model and to generate a detection signature. Those signatures serve as input to a malware detector, acting as the antibodies in the antigen detection process. This allows us to understand the malware attack and aids in the infection removal procedures. © 2012 Springer-Verlag.

Experimental analysis of an electromagnetic zero-sequence suppressor

The intent of this paper is to present contributions focused on the analysis and development of harmonic attenuator devices. Among these, highlights here the so-called electromagnetic zero-sequence suppressor. This arrangement consists of a filter and a blocker, both electromagnetic, whose combined operation provides paths for low and high impedance, respectively, which can be conveniently adjusted to the desired performance. In this context, here are present results related to experimental studies that show the behavior of the equipment in front of different operating conditions. The tests were performed on a low-power prototype (1kVA/220V) and the analysis results show the main motivator aspects for the use of these devices. © 2012 IEEE.

Role of TLR-2 and fungal surface antigens on innate immune response against Sporothrix schenckii

Sporotrichosis is an infection caused by the dimorphic fungus Sporothrix schenckii. Toll-like receptors (TLRs) play an important role in immunity, since they bind to pathogen surface antigens and initiate the immune response. However, little is known about the role of TLR-2 and fungal surface antigens in the recognition of S. schenckii and in the subsequent immune response. This study aimed to evaluate the involvement of TLR-2 and fungal surface soluble (SolAg) and lipidic (LipAg) antigens in phagocytosis of S. schenckii and production of immune mediators by macrophages obtained from WT and TLR-2 -/- animals. The results showed that TLR-2-/- animals had had statistical lower percentage of macrophages with internalized yeasts compared to WT. SolAg and LipAg impaired phagocytosis and immunological mediator production for both WT and TLR-2-/-. The absence of TLR-2 led to lower production of the cytokines TNF, IL-1β, IL-12 and IL-10 compared to WT animals. These results suggest a new insight in relation to how the immune system, through TLR-2, recognizes and induces the production of mediators in response to the fungus S. schenckii. Copyright © Informa Healthcare USA, Inc.