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We assessed the effect of sodium butyrate (SB) as a histone deacetylase inhibitor (HDACi) on Toll-like receptor 4 (TLR4) gene expression levels, in low TLR4 expressing (HCT116) and high TLR4 expressing (SW480) colorectal cancer cells. The cytotoxic effect of SB was assessed by culturing SW480 and HCT116 cell lines using a broad spectrum of times and concentrations of SB. The MTT assay was done to check the cytotoxic properties of different SB concentrations. Gene expression levels of TLR4 was then evaluated for non-cytotoxic SB concentrations. Morphological analysis and MTT assay confirmed that SB concentrations equal to or less than 5mM were not cytotoxic for both cell lines. At 5mM concentration of SB in SW480 cell line and 1mM concentration of SB in HCT116 cell line, TLR4 gene expression level significantly increased from 24 to 48 hrs and decreased significantly from 48 to 72 hrs with an "early increased and late decreased pattern". At 1mM concentration of SB in SW480 cell line and 5mM concentration of SB in HCT116 cell line, TLR4 expression had a "gradually increased pattern". This study focuses on the dose-time-effect of SB in the pathogenesis of colorectal cancer. SB alters the expression level of TLR4 in colorectal cancer cells. This effect may depend on the cell type, treatment duration and SB concentration. The alterations in TLR4 expression may be due to the direct effect of SB on TLR4 and/or the expression changes of in other genes which may indirectly affect the TLR4 expression. Abbreviations: TLR4: Toll-like receptor 4; HDACi: histone deacetylase inhibitor; SB: sodium Butyrate; CRC: colorectal cancer; SCFA: short-chain fatty acid; hrs: hours.

Background Visceral leishmaniasis (VL), caused by Leishmania infantum, is a systemic parasitic disease and presents a global health problem which can be fatal if not diagnosed and treated. Dogs are the main hosts and provide reservoirs for the transmission of the disease to humans. Methods In this study, the gene encoding a 21-kDa protein was cloned and expressed as a fusion protein in Escherichia coli strain BL21 (DE3) for developing a rapid immunochromatographic test (ICT) to identify infected dogs. The expression of the recombinant 21-kDa protein (r21) was investigated using SDS-PAGE and Western blot methods. The purified r21-kDa protein was spotted onto ICT strips and tested by sera from experimentally infected, naturally infected and uninfected dogs. Results The SDS-PAGE and Western blot methods showed the successful expression of r21-kDa protein. The ICT strip test revealed that the r21-kDa protein was detected by the sera of experimentally and naturally infected dogs. The specificity tests also confirmed no cross-reactivity with animals infected with Trypanosoma cruzi, Toxoplasma gondii and Ehrlichia canis. Conclusions Based on these findings, the new r21-kDa protein may be a suitable target for developing a new simple, specific and rapid serological method to detect VL in infected dogs.

Antibodies are still widely used in several programs including early research, imaging, Targeting drug delivery system, Affinity chromatography, flowcytometry technic, diagnosis and treatment. Purification of antibody is a standard approach for detection of infection agent in different species. The reservoir hosts for Leishmania infantum are Dogs and they have active role in the transmission of leishmania to humans by the bite of a sand fly belonging to genus Phlebotomus and Lutzomiya. Consequently, elimination of dogs in endemic areas and vaccination of dogs contributes to reduction of the human and canine VL cases. Serological antibody tests such as IFAT (Indirect Fluorescent Antbody Test), DFAT (Direct Fluorescent Antbody Test), ELISA (Enzyme–Linked Immunosorbent Assay), PCR (Polymerase chain Reaction Assay) have been extensively used to investigate canine infection with L. infantum. In this study we produced and purified polyclonal antibody against attenuated and wild type leishmania infantum in dogs. Anti-leishmania in dog serums precipitated with ammonium sulphate. The IgG recovered from ammonium sulphate precipitation was subject to ion exchange chromatography (IEC) and the purity of IgG was confirmed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) under reduced condition. The purity of proteins were above 95% and then purified IgG was conjugated with FITC. We determined optimum titer of dog IgG by observation parasites under fluorescent microscope. The optimum dilution of prepared FITC conjugated dog IgG was 1: 400. This polyclonal antibody can be used for other applications in research, diagnosis and clinic.

Rheumatoid arthritis (RA) is a systemic autoimmune disease predominantly involving the synovial joints and affects up to 1 of adults worldwide. The aim of the present study was to determine whether the interleukin-1 receptor (IL1R)-associated kinase (IRAK1) rs3027898 gene polymorphism confers susceptibility to RA in a sample of patients from Iran. This gene encodes IRAK1, one of two putative serine/threonine kinases that associates with IL1R upon stimulation. IRAK1 is partially responsible for IL-1-induced upregulation of the transcription factor, nuclear factor-kappa B. The present case-control study was performed on 120 patients with RA and 120 healthy individuals. Genomic DNA was extracted from whole blood, and the gene polymorphism was evaluated using a tetra-primer amplification refractory mutation system-polymerase chain reaction method. The results demonstrated that there was no association between IRAK1 rs3027898 CA genotype and the risk of RA in women (odds ratio= 0.72, 95 confidence interval= 0.41-1.49; P= 0.446). Further studies with larger sample sizes and different ethnicities are required to validate the present findings.

Background: Different background of immunity responses determine resistance or susceptibility of certain mouse strains to Leishmania major infection. Some have been well known previously. Antimicrobial peptides (AMPs) such as cathelicidins and defensins are unique fragments of innate immunity system with well-known effects against the invasion pathogens. Despite their outstanding roles and being of extensive cases of cutaneous leishmaniasis (CL) caused by L. major, they have been less studied in Leishmania fields. The aim of present study was to determine whether these components play a role in the protection of skin against Leishmania infections. Methods: The animal model of Leishmania infection was established by the subcutaneous inoculation of 5×106(parasites/ml) from the stationary phase of L. major promastigotes to BALB/c and C57BL/6 mice from January 2016 to August 2016 in Kerman Province, southeast of Iran. After 1, 3 and 7 d of post-infection (PI), the samples needed for detecting of the mRNA levels of mouse beta-defensin (mBD)-1, mBD2, mBD3, mBD4, mBD6, cathlin-related antimicrobial peptide (CRAMP), interleukin (IL)-10, IL-12 and parasite load were taken under standard methods. Results: The findings related to cytokines profiles in BALB/c (↑IL-10, ↓IL-12) and C57BL/6 mouse strains (↓IL-10, ↑IL-12) demonstrated that immunity system has been accurately activated during CL caused by L. Major parasites. We also observed a significant up-regulation of all aforementioned AMPs genes in BALB/c mice at selected times compared to another strain. Conclusion: CL occurred in BALB/c mice in spite of the fact that the expression of AMPs was higher than the other strain. AMPs genes are well expressed to provide defense against the parasites that have increased and escaped from immunity system but cannot create an absolute protection.

Cutaneous Leishmaniasis is a serious public health problem, typically affecting poor populations with limited access to health care. Control is largely dependent on chemotherapies that are inefficient, costly and challenging to deliver. Vaccination is an attractive and feasible alternative because long-term protection is typical in patients who recover from the disease. No human vaccine is yet approved for use, but several candidates are at various stages of testing. Live attenuated parasites, which stimulate long-term immune protection, have potential as effective vaccines, and their challenges relating to safety, formulation and delivery can be overcome. Here we review current data on the potential of live attenuated Leishmania vaccines and discuss possible routes to regulatory approval.

Background Resistance to antimonials is a fundamental determinant of treatment failure in anthroponotic cutaneous leishmaniasis (ACL). Detection of reliable molecular markers to distinguish unresponsive and responsive parasites is critical for consolidating strategies to monitor drug efficacy. Methods We analysed the expression of five major antimony resistance-associated genes that is aquaglyceroporin1 (AQP1), γ-glutamylcysteine synthetase (γ-GCS), multidrug resistance protein A (MRPA), trypanothione reductase (TR) and thiol-dependent reductase 1 (TDR1), in unresponsive and responsive Leishmania tropica field isolates by quantitative real-time PCR in comparison with sensitive and resistant reference strains. Results Gene expression analysis showed the down-regulation of AQP1, γ-GCS and TDR1 by a factor of 1.9, 1.7 and 3.55, respectively, in unresponsive isolates vs. responsive ones. The average RNA expression level of MRPA increased by a factor of 1.9 in the unresponsive group. Isolates exhibited a strong positive linear correlation between gene expression of AQP1 and γ-GCS. A negative correlation between the AQP1 and γ-GCS expression level and lesion duration in responsive patients indicated the potential role in diagnosing drug-unresponsive parasites in endemic areas of ACL. Conclusion In cases of inconclusive outcomes of resistance tests in clinical isolates, expression analysis of a set of influential genes can be beneficial to identify distinctive biomarkers between antimony-unresponsive and responsive parasites.

Melanoma differentiation-associated protein 5 (MDA5) and retinoic acid-inducible gene 1 (RIG-1) as the pattern recognition receptors play important roles in viral mRNA recognition. Chronic HBV-infected (CHB) patients are unable to properly respond to hepatitis B virus (HBV). Therefore, the aim of the present study was to evaluate the mRNA levels of MDA5 and RIG-1 in the peripheral blood immune cells of CHB patients in comparison to healthy controls. In this cross-sectional study, the mRNA levels of MDA5 and RIG-1 were examined in 60 CHB patients and 60 healthy controls using the real-time polymerase chain reaction (PCR) technique. Our results showed that mRNA levels of MDA5 and RIG-1 were significantly decreased and increased, respectively, in CHB patients when compared to healthy controls. Our results also revealed that mRNA levels of MDA5 and RIG-1 were not altered among CHB patients with various states of e-antigen of hepatitis B and HBV-DNA viral loads. According to the results presented here, it may be concluded that downregulation of MDA5 may be a responsible mechanism from several reasons, which leads to HBV persistence in CHB patients.

The regulatory T (Treg) cells play a major role in the control of the autoimmunity and inflammation, and IL-35 has been described as an immunosuppressive cytokine that is mainly produced by CD4+ FOXP3+ Treg cells. The aim of this study was to evaluate the serum levels of IL-35 and a single nucleotide polymorphism (SNP), rs3761548, in FOXP3 gene in patients with multiple sclerosis. The blood samples were collected from 140 multiple sclerosis (MS) patients (including 51 untreated and 89 treated patients) and 140 healthy subjects as a control group. The serum levels of IL-35 were measured by ELISA. The DNA was analyzed for SNP rs3761548 in FOXP3 gene using SSP-PCR. There was no significant difference between untreated MS patients and control group regarding the mean serum levels of IL-35, although this parameter was higher in untreated patients. However, the mean serum level of IL-35 in treated MS patients was significantly higher than that in the control group (P

An attenuated line of Leishmania infantum (the H-line), developed through exposure to gentamicin, has been shown to protect dogs against canine visceral leishmaniasis. A specific diagnostic test to differentiate dogs vaccinated with the attenuated line from dogs infected with L. infantum wild-type (L. infantum WT) could be a valuable tool in evaluating the effectiveness of canine vaccination. In this study, 28 healthy dogs were allocated into four groups. In Group I and Group II (eight dogs per group), dogs were immunized subcutaneously (s.c.) with L. infantum H-line, and the dogs of Group II challenged s.c. with L. infantum WT, at 2 months post-immunization. In Group III, eight animals were challenged s.c. with L. infantum WT, and four dogs of Group IV were injected s.c. with PBS. We found that sera from vaccinated dogs recognize a 21 kDa antigen of promastigotes of L. infantum H-line but not of L. infantum WT, whereas sera from unvaccinated dogs challenged with L. infantum WT, recognized a 21 kDa antigen of promastigotes of L. infantum WT but not of L. infantum H-line. Sera from dogs challenged with L. infantum WT with prior vaccination with L. infantum H-line, recognized a 21 kDa antigen of both L. infantum WT and L. infantum H-line. These results suggest that the Western blot analysis of antibodies against 21 kDa antigens of L. infantum H-line and WT may be a useful technique for distinguishing between dogs vaccinated with L. infantum H-line and dogs naturally infected with L. infantum WT.

Chemokines play an important role in the autoimmune diseases. The aim of this study was to investigate the levels of CCL20 and a polymorphism [-786C > T (rs6749704)] in the chemokine gene in patients with multiple sclerosis (MS). The blood samples were collected from 135 MS patients and 135 healthy subjects as a control group. The patients have relapsing-remitting (RRMS; n = 65), primary progressive (PPMS; n = 47), secondary progressive (SPMS; n = 35) or progressive relapsing (PRMS; n = 14) patterns. The serum levels of CCL20 were measured by ELISA. The DNA was analyzed for CCL20 polymorphism using PCR–RLFP. The mean serum levels of CCL20 in the MS group were significantly higher than in the healthy group (P

Chemokines play a major role in autoimmune diseases such as multiple sclerosis (MS). Gender also affects the susceptibility and course of MS. The aim of this study was to investigate the serum levels of the macrophage-derived chemokine (CCL22) in women and men patients with MS. Blood samples were collected from 135 healthy subjects (35 men and 100 women) and 135 MS patients (29 men and 136 women; 47 newly diagnosed and 88 treated patients and have relapsing-remitting (RRMS; n = 65), secondary progressive (SPMS; n = 37), primary progressive (PPMS; n = 19), or progressive relapsing (PRMS; n = 14) patterns). The serum levels of CCL22 were measured by ELISA. The difference of the mean serum levels of CCL22 between the newly diagnosed MS men and healthy men was not significant, but in newly diagnosed MS women, the mean serum levels of CCL22 were significantly lower than those in treated MS women and healthy women (P

Pentavalent antimony compounds are expensive, toxic and drug resistance is prevalent, whereas the plant extract derivatives are safe. In the present study, the effect of methanolic and aqueous extracts of Eucalyptus camaldulensis on the promastigotes of Leishmania major was evaluated. The methanolic and aqueous extracts of E. camaldulensis leaves were prepared. The compounds were dried and powdered. Serial dilutions of the extracts and control drugs in phosphate buffer solution were prepared. The stationary phase promastigotes of L. major were incubated to the methanolic and aqueous extractions in vitro. Tartar emetic was used as the positive control drug. After 72 h of incubation the activity of the extracts was measured, using MTT method. The IC50 values (50 % inhibitory concentration) were 586.2 ± 47.6 and 1,108.6 ± 51.9 μg/ml for methanolic and aqueous extracts, respectively, whereas it was 32.5 ± 6.8 μg/ml for tartar emetic. The results indicated that the methanolic extract was more effective than aqueous extract, although there was no significant difference. The extracts were less effective as compared to the control drug. Further investigation is required to evaluate these extracts on clinical stage in macrophage-amastigote model.

Hepatitis B virus (HBV) which includes, fulminant, acute, chronic, asymptomatic, and occult HBV infection is the most prevalent virus that leads to human liver diseases. Chronic, asymptomatic, and occult infection can induce further sever diseases such as hepatocellular carcinoma (HCC) and cirrhosis of the liver. The underlying mechanisms that allow progression of the prolonged forms of the infection and subsequent HCC or cirrhosis of the liver are yet to be clarified. However, many researchers have suggested that immunological and genetic parameters may play important roles in the etiology of hepatitis B. Transforming growth factor beta (TGF-β) is an important cytokine with dual regulatory functions in the immune system and in the responses against viral infections. However, the pathways and mechanisms controlling these are not fully understood. The crucial roles of TGF-β in the development of Th17 and T regulatory lymphocytes, the main cell types involved in autoimmunity and destructive immune related diseases, have been documented and this provides insights into TGF-β function during hepatitis infection and subsequent HCC and cirrhosis of the liver. Recent findings also confirm that TGF-β directly alters hepatocyte function during hepatitis B, hence, the aim of this review is to address the current data regarding the association and status of TGF-β with hepatitis B infection and its related disorders including HCC and cirrhosis of the liver.

Rheumatoid arthritis (RA) is a systemic autoimmune disease predominantly involving the synovial joints and affects up to 1% of adults worldwide. The aim of the present study was to determine whether the interleukin-1 receptor (IL1R)-associated kinase (IRAK1) rs3027898 gene polymorphism confers susceptibility to RA in a sample of patients from Iran. This gene encodes IRAK1, one of two putative serine/threonine kinases that associates with IL1R upon stimulation. IRAK1 is partially responsible for IL-1-induced upregulation of the transcription factor, nuclear factor-κB. The present case-control study was performed on 120 patients with RA and 120 healthy individuals. Genomic DNA was extracted from whole blood, and the gene polymorphism was evaluated using a tetra-primer amplification refractory mutation system-polymerase chain reaction method. The results demonstrated that there was no association between IRAK1 rs3027898 CA genotype and the risk of RA in women (odds ratio=0.72, 95% confidence interval=0.41–1.49; P=0.446). Further studies with larger sample sizes and different ethnicities are required to validate the present findings.

In this study we aimed to evaluate the possible association of PTPN22 rs2476601 as well as epidermal growth factor receptor (EGFR) rs17337023 gene polymorphism and rheumatoid arthritis (RA) in a sample of Iranian population. This case-control study was performed on 120 patients with RA and 120 healthy subjects. Genomic DNA was extracted from whole blood and PTPN22 rs2476601 and EGFR rs17337023 polymorphisms were determined using tetra amplification refractory mutation system-polymerase chain reaction (T-ARMS-PCR). The results showed that PTPN22 rs2476601 CT genotype as well as rs2476601 T allele was a risk factor for susceptibility to RA (OR=5.89 95%CI = 1.78-19.48, P = 0.004 and OR = 4.78, 95%CI = 1.59-14.35, P = 0.003, respectively). We also found that EGFR rs17337023 AT and rs17337023 TT genotypes were risk factor for susceptibility to RA (OR = 9.94 95%CI = 3.65-26.73, P < 0.001 and OR = 3.66, 95%CI = 1.46-9.15, P = 0.005, respectively). In addition the EGFR rs17337023 T allele was a risk for predisposition to RA (OR = 1.56, 95%CI=1.06-2.30, P = 0.030). In conclusion, we found an association between PTPN22 rs2476601 and EGFR rs17337023 polymorphisms and the risk of RA in a sample of Iranian population.

We have previously described an attenuated line of Leishmania infantum (H-line), selected by culturing promastigotes in vitro in the presence of gentamicin. To elucidate the molecular basis for this attenuation, we undertook a comparative proteomic analysis using multiplex 2-dimensional (2D) difference gel electrophoresis. Eighteen proteins that showed significant and reproducible changes in expression were identified. Many of these were components of the thiol-redox control system in Leishmania and this observation, validated by Western blot, prompted us to investigate the sensitivity of the attenuated line to oxidative stress. The attenuated line was found to be significantly more susceptible to hydrogen peroxide, a change which may explain the loss of virulence. In a direct assay of trypanothione-dependent peroxidase activity, hydrogen peroxide metabolism in the H-line was significantly lower than in wild type. Furthermore, trypanothione reductase activity was significantly lower in the H-line, suggesting that gentamicin selection may result in pleiotropic affects on thiol metabolism in Leishmania. A putative RNA-binding protein was very strongly up-regulated in the attenuated line, suggesting a possible target for gentamicin in Leishmania.

An attenuated line of Leishmania infantum (L. infantum H-line) has been established by culturing promastigotes in vitro under gentamicin pressure. Here, we show that L. infantum H-line induced significantly higher levels of IFN-γ and lower levels of IL-10 compared with those in dogs infected with L. infantum wild type (WT). Anti-Leishmania-specific total IgG, IgG1, and IgG2 antibodies were present in the serum of all infected dogs, with levels of IgG2 subclass highest in the sera of dogs inoculated with L. infantum H-line. Relatively high levels of IgG1 were found in the sera of dogs infected with L. infantum WT. Six of seven dogs immunized intradermally (i.d.) with the attenuated line later showed a positive skin test to leishmanin, whereas the dogs infected with L. infantum WT did not. No clinical abnormalities were observed, and no parasites found in the visceral organs of the dogs inoculated intravenously (i.v.) with L. infantum H-line over 24 months post-inoculation. Dogs which had been immunized with L. infantum H-line i.d. 12 months previously were protected against challenge with L. infantum WT. These data suggest that the L. infantum H-line was safe and induced a protection which is correlated with cellular immunity in dogs.

Aims: Quantum dots are optical nanocrystals whose in vitro and in vivo use in molecular imaging is expanding rapidly. In comparison with organic fluorophores, quantum dots exhibit desirable properties, such as multiwavelength fluorescence emission, excellent brightness and resistance to photobleaching. Their electron-dense, metallic cores suggest utility in other clinical imaging modalities. Methods: Core-shell zinc sulfide-cadmium telluride quantum dots were studied by magnetic resonance and computed tomography phantoms. Quantum dots were also injected into rat brain, as well as intravenously, using convection-enhanced delivery, prior to animal imaging. Results: Computed tomography studies suggest that current formulations of quantum dots might be imaged in vivo in animals. Conclusions: Used in conjunction with optical imaging techniques, quantum dots have the potential to function as multimodal imaging platforms in vivo. The ability to detect an optical nanoparticle preoperatively with clinical imaging modality offers a distinct advantage to clinicians engaged in image-guided surgical applications.

Clostridium perfringens type D infects ruminants and causes the enterotoxemia disease by e-toxin. A mutated e-toxin gene lacking toxicity was designed, synthesized, and cloned into the pT1NX vector and electroporated into Lactobacillus casei competent cells to yield LC-pT1NX-e recombinant strain. BALB/c mice, immunized orally with this strain, highly induced mucosal, humoral, and cell-mediated immune responses and developed a protection against 200 MLD/ml of the activated e-toxin. This study showed that the LC-pT1NX-e could be a promising vaccine candidate against the enterotoxemia disease.

Objective The identification of neoplastic tissue within normal brain during biopsy and tumor resection remains a problem in the operative management of gliomas. A variety of nanoparticles are phagocytized by macrophages in vivo. This feature may allow optical nanoparticles, such as quantum dots, to colocalize with brain tumors and serve as an optical aid in the surgical resection or biopsy of brain tumors. Methods Male Fisher rats (Charles River Labs, Wilmington, MA) were implanted intracranially with C6 gliosarcoma cell lines to establish tumors. Two weeks after the implantation of tumors, 705-nm emission Qdot ITK Amino(PEG) Quantum Dots (Quantum Dot Corp., Hayward, CA) were injected via the tail vein at doses of 3 to 17 nmol. The animals were sacrificed 24 hours after the injection of quantum dots and their tissues were examined. Results Quantum dots are avidly phagocytized by macrophages and are taken up by the liver, spleen, and lymph nodes. A dose-response relationship was noted. At low doses, the majority of the quantum dots are sequestered in the liver, spleen, and lymph nodes. At higher doses, increasing quantities of quantum dots are noted within the experimental brain tumors. Macrophages and microglia colocalize with glioma cells, carrying the quantum dot and thereby optically outlining the tumor. Excitation with blue or ultraviolet wavelengths stimulates the quantum dots, which give off a deep red fluorescence detectable with charge-coupled device cameras, optical spectroscopy units, and in dark-field fluorescence microscopy. Conclusion Quantum dots are optical nanoparticles that, when delivered in nanomole doses, are phagocytized by the macrophages and microglia that infiltrate experimental gliomas. The optical signal may be detected, allowing for improved identification and visualization of tumors, potentially augmenting brain tumor biopsy and resection.

The stimulator of interferon genes (STING) induces the activation of interferon regulatory factor 3 (IRF3) in response to intracellular viral double-stranded (ds) DNA. The aim of this study was to evaluate mRNA levels of STING and its downstream transcription factor, IRF3, in the isolated peripheral blood mononuclear cells (PBMCs) of patients with chronic HBV (CHB) infection.This study was performed on 60 healthy controls and 60 CHB patients. The mRNA levels of STING and IRF3 were determined using Real-Time polymerase chain reaction (PCR) techniques. The SPSS software version 18 was used to analyze raw data.The results revealed that mRNA levels of STING were significantly decreased in CHB patients in comparison to healthy controls (P = 0.013). Our results also revealed that expression levels of IRF3 were not different between CHB patients and healthy controls (P = 0.828).In the present study, we found that CHB patients were unable to express appropriate levels of STING. Thus, it may result in impairment of HBV-DNA recognition and subsequently disruption of immune responses. These results suggest a plausible mechanism which may partially define the fact that immune responses are impaired in CHB patients.

An attenuated line of Leishmania infantum (L. infantum H-line) has been established by culturing promastigotes in vitro under gentamicin pressure. A vaccine trial was conducted using 103 naive dogs from a leishmaniosis non-endemic area (55 vaccinated and 48 unvaccinated) brought into an endemic area of southeast Iran. No local and/or general indications of disease were observed in the vaccinated dogs immediately after vaccination. The efficacy of the vaccine was evaluated after 24 months (4 sandfly transmission seasons) by serological, parasitological analyses and clinical examination. In western blot analysis of antibodies to L. infantum antigens, sera from 10 out of 31 (32.2%) unvaccinated dogs, but none of the sera from vaccinated dogs which were seropositive at100, recognized the 21 kDa antigen of L. infantum wild-type (WT). Nine out of 31 (29%) unvaccinated dogs, but none of vaccinated dogs, were positive for the presence of Leishmania DNA. One out of 46 (2.2%) vaccinated dogs and 9 out of 31 (29%) unvaccinated dogs developed clinical signs of disease. These results suggest that gentamicin-attenuated L. infantum induced a significant and strong protective effect against canine visceral leishmaniosis in the endemic area.

A method for attenuation of Leishmania species by culturing in vitro under gentamicin pressure has been used successfully with Leishmania mexicana, L. major, L. infantum, and L. donovani. The attenuated lines invaded but were unable to survive within bone marrow-derived macrophages in vitro, whereas wild-type parasites survived and multiplied. The attenuated lines of L. mexicana and L. major both failed to induce cutaneous lesions in the majority of BALB/c mice over a minimum 12-week observation period after subcutaneous injection of stationary phase parasites. The attenuated line of L. mexicana retained its properties in gentamicin-free medium over 40 subcultures. The attenuated lines of L. mexicana and L. major both induced significant protection in mice against challenge with wild-type parasites.

An attenuated line of Leishmania infantum (L. infantum H-line) has been established by culturing promastigotes in vitro under gentamicin pressure. A vaccine trial was conducted using 103 naive dogs from a leishmaniosis non-endemic area (55 vaccinated and 48 unvaccinated) brought into an endemic area of southeast Iran. No local and/or general indications of disease were observed in the vaccinated dogs immediately after vaccination. The efficacy of the vaccine was evaluated after 24 months (4 sandfly transmission seasons) by serological, parasitological analyses and clinical examination. In western blot analysis of antibodies to L. infantum antigens, sera from 10 out of 31 (32.2%) unvaccinated dogs, but none of the sera from vaccinated dogs which were seropositive at >100, recognized the 21 kDa antigen of L. infantum wild-type (WT). Nine out of 31 (29%) unvaccinated dogs, but none of vaccinated dogs, were positive for the presence of Leishmania DNA. One out of 46 (2.2%) vaccinated dogs and 9 out of 31 (29%) unvaccinated dogs developed clinical signs of disease. These results suggest that gentamicin-attenuated L. infantum induced a significant and strong protective effect against canine visceral leishmaniosis in the endemic area., Author Summary A 24 month vaccine trial was conducted using 103 leishmania free dogs in an area of southeast Iran endemic for visceral leishmaniosis. The dogs were vaccinated with gentamicin-attenuated line of Leishmania infantum. No local and/or general indications of disease were observed in the vaccinated dogs immediately after vaccination with an attenuated line of Leishmania infantum. Nine out of 31 (29%) unvaccinated dogs, but none of those vaccinated, were positive for presence of Leishmania DNA by PCR. In western blots, sera from 10 out of 31 (32.2%) unvaccinated dogs, but none of the sera from vaccinated dogs, recognized the 21 kDa antigen of Leishmania infantum wild-type. One out of 46 (2.2%) vaccinated dogs and 9 out of 31 (29%) unvaccinated dogs developed clinical signs of disease. The attenuated Leishmania infantum induced a significant and strong protective effect against Leishmania infantum infection in the field.

The objective of this paper was to test the hypothesis that testosterone (T) raises blood pressure (BP), which is associated with increased coronary adventitial collagen, whereas the hemodynamic force of BP increases the coronary media:lumen ratio. Five treatment groups of spontaneously hypertensive rat (SHR) were established (n = 8-10 per group): controls; hydralazine (HYZ); castration; castration + HYZ; and castration + HYZ + T + captopril. At 12 weeks of age, the castrate + HYZ group was divided so that the mean BP was the same in both groups (162 mmHg). Both groups continued to receive HYZ treatment; however one group received T implants. Also, at 12 weeks of age the castrate + HYZ + T + captopril group received T implants. BP in the HYZ group was reduced compared with controls (192 mmHg vs 218 mmHg, p0.01). Castration lowered BP to 170 mmHg (p0.01) compared with controls. However, T implants increased BP by 15 mmHg (p0.02) in the castrate + HYZ group and by 44 mmHg in the castrate + HYZ + captopril group (p0.01). Captopril in combination with HYZ significantly reduced BP compared with controls but T replacement increased BP and coronary collagen deposition in spite of HYZ and captopril treatment.

Previous studies from our laboratory have demonstrated that the Y chromosome from the spontaneously hypertensive rat (SHR) is responsible for a significant portion of the elevated blood pressure and also produces an earlier pubertal rise in plasma testosterone. We performed the following studies to determine whether the SHR Y chromosome raises blood pressure by sympathetic nervous system responses as measured by adrenal chromogranin A and plasma and tissue catecholamines. Male SHR from The University of Akron colony were studied from 5 to 20 weeks of age. Blood pressure was measured by tail-cuff, tail artery cannulation, and aortic telemetry (Data Sciences); acute (air stress) and chronic (territorial colony) social stressors were compared; blood was collected for determination of plasma catecholamines; and adrenal glands were analyzed at 15 weeks for catecholamines. Rats with the SHR Y chromosome had higher blood pressure and plasma norepinephrine than those with the normotensive Wistar-Kyoto (WKY) Y chromosome. However, the SHR Y chromosome did not significantly change responsiveness to acute or chronic stressors. Phentolamine and clonidine prevented the stress responses. Adrenal chromogranin A levels were elevated 37% and 40% and adrenal norepinephrine content 29% and 100% at 4 and 10 weeks of age, respectively, in rats with an SHR Y chromosome compared with WKY. Chemical sympathectomy normalized blood pressure in all strains and significantly reduced norepinephrine (36% to 41%) in all strains except in WKY, which already had a normal blood pressure. In conclusion, the SHR Y chromosome appears to increase the chronic sympathetic nervous system. A potential mechanism could be a Y locus that influences chronic sympathetic nervous system activity, which may reinforce neurohumoral factors and structural components of the vessel wall, accelerating the development of hypertension.

SUMMARYThe impact of immunization with gentamicin-attenuatedLeishmania infantum(H-line) on the immunophenotypic profile of popliteal lymph node (PLN) and peripheral blood mononuclear cells (PBMCs) of dogs was assessed by flow cytometry and immunohistochemistry. Compared with the dogs infected withL. infantumwild-type (Group WT), there was a significantly higher percentage of CD4+, CD44+T cells and CD14+, MHC-II+cells and a lower percentage of CD4+CD25+regulatory T cells in PLN of the immunized dogs withL. infantumH-line (Group H). The percentage of CD4+and CD8+T cells in PBMCs of immunized dogs was higher than that in dogs of Group WT. The CD4:CD8 ratio in PLN of dogs of Group H was significantly higher than that in dogs of Group WT. A significantly higher percentage of CD21+B cells and a lower percentage of CD79b+cells were found in PLN of the immunized dogs compared with dogs of Group WT. Immunohistochemical investigation showed no parasites in the PLN of immunized dogs whereas there were parasites in the PLN of 60% of dogs infected withL. infantumWT. In this study, the immunophenotypic profile of mononuclear cells of the immunized dogs correlates with cellular immunity.

In this study, different positively charged niosomal formulations containing sorbitan esters, cholesterol and cetyl trimethyl ammonium bromide were prepared by film hydration method for the entrapment of autoclaved Leishmania major (ALM). Size distribution pattern and stability of niosomes were investigated by laser light scattering method and ALM encapsulation per cent was measured by the bicinchoninic acid method. Finally, the selected formulation was used for the induction of the immune response against cutaneous leishmaniasis in BALB/c mice. Size distribution curves of all the formulations followed a log-normal pattern and the mean volume diameter was in the range 7.57-15.80 µm. The mean volume diameters were significantly increased by adding Tween to Span formulations (p 0.05). The percentage of ALM entrapped in all formulations varied between 14.88% and 36.65%. In contrast to ALM, in vivo studies showed that the niosomes containing ALM have a moderate effect in the prevention of cutaneous leishmaniasis in BALB/c mice.

One of the most critical issues in the wireless sensor networks is the network's nodes limited availability of energy. The network lifetime strictly depends on its energy efficiency. Clustering is a method to make the consumed energy efficient. LEACH is one of the fundamental clustering algorithms. In this paper we have proposed an algorithm which extends LEACH by considering number of neighbors and remaining of the energy of each node to be a cluster head. We have also proposed to use gateway nodes in each cluster to enable the cluster head to reduce its transmission radius within the cluster to achieve better power saving. We analyze and compared the proposed algorithm along with the LEACH, LEACH-c and MIMO the most recent extend over the LEACH and show that the proposed algorithm can prolong the lifetime of the network.

The aim of this study was to assess the potential adverse effects of the chronic use of surgically implanted radiotelemetry transmitters and femoral catheters on embryo-fetal development in rats.Two groups of 10 female rats were implanted with femoral catheters and radiotelemetry transmitters. Ten additional rats were implanted with femoral catheters alone. The females were then time mated with male rats and pregnancy confirmed. Saline was continuously infused from gestation days 6-17. Saline infusion was replaced with 0.8 μg/ml angiotensin II in one group of animals, on gestation days 6, 9, 12, and 15 in order to demonstrate the validity of the system to detect changes in hemodynamic function by a pharmacologically active agent. Embryo-fetal development was assessed by examination of the number and location of the fetuses, early and late resorptions, and number of implantation sites. Parameters evaluated included postimplantation loss, live litter size, mean fetal body weights, and fetal sex ratios.Intrauterine survival was unaffected in all groups. Mean numbers of corpora lutea and implantation sites and the mean litter proportions of preimplantation loss were similar across all groups. Infusion of angiotensin II elicited the expected hemodynamic response, i.e., increased arterial blood pressure with a concomitant decrease in heart rate.Radiotelemetry can be used in combination with continuous intravenous infusion as a viable methodology for detecting test article induced changes in cardiovascular function in pregnant rats, with no effects on intrauterine growth and survival or fetal morphology.

Our laboratory has shown that the Y chromosome has a significant effect on blood pressure in the spontaneously hypertensive rat (SHR) model of hypertension and that the testes and androgen receptor contribute to the blood pressure rise. As an extension of our research, we have developed two new rat strains, SHR/a and SHR/y (F11) to study the Y chromosome. The objectives of the following research were 1) to study the blood pressure of rats with an SHR Y chromosome in a normotensive genetic background (SHR/y) or a normotensive Y chromosome in an SHR genetic background (SHR/a), 2) to determine the effect of male sex phenotype on the blood pressure of these rats, 3) to determine if testosterone replacement in castrated rats would restore blood pressure, and 4) to determine whether the Y chromosome from the SHR/y strain when crossed with a normotensive female can induce hypertension in androgen receptor-deficient male offspring. Blood pressure of male SHR/y rats was significantly higher than that of normotensive Wistar-Kyoto males (p < 0.01), and SHR/a males had significantly lower blood pressure compared with that of the parent SHR strain (p = 0.05). Testosterone replacement in castrated rats of both strains (SHR/a and SHR/y) restored blood pressure to control levels. Normotensive female King-Holtzman rats heterozygous for the testicular feminization gene were crossed with F11 SHR/a and SHR/y males.(ABSTRACT TRUNCATED AT 250 WORDS)

This paper, presents a learning automata based algorithm (LABTA) to determine an appropriate time interval for topology control, while capturing network updates and minimizing algorithm overhead. This algorithm is application dependent and it is possible to consider other parameters like delay, etc. In this paper, we describe how to merge parameters and compute the fitness. We have implemented the proposed algorithm with both of standard and distributed models of the learning automata. Simulation results show that our proposed algorithm outperforms periodic topology control. In addition, a comparison can be also found between two most well known topology control protocols; LMST and K Neigh.

SUMMARYAn attenuated line of Leishmania major (L. major H-line) has been established by culturing promastigotes in vitro under gentamicin pressure. A modification of the previously described method for the generation of attenuated L. major is described, giving rise to attenuated parasites after 8 rather than 12 subpassages. No lesions developed in BALB/c mice infected with L. major H-line, whereas L. major wild-type (WT) induced a Th2 like response with progressive lesions. Analysis of splenocyte IFN-γ and IL-4 production following stimulation with promastigotes shows that the L. major H-line preferentially induces Th1-like responses and possibly down-regulates Th2 responses in BALB/c mice. L. major H-line parasites remained localized in the skin and draining lymph node, whereas L. major WT parasites disseminated into the visceral organs of BALB/c mice. Mice infected with L. major H-line acquired some resistance against L. major WT. These results show that the attenuated cell line of L. major is not only avirulent but that it may also modulate the host immune response.

Nowadays systems have became more complex and faults can have more influence on them. There are some cases where we can not monitor or repair, like satellites. Also some systems have a crucial role and they can not fail at all. So however our components are fault tolerant but more fault tolerance is needed. On the other hand, the processing power has increased and the interconnection network between them has became the bottleneck. In this paper we study some switching techniques in the interconnection networks. Then an improved switching technique is proposed. Our proposed technique tries to provide more fault tolerance while keeping performance too. finally, this techniques are compared.

The clinicopathological changes following infection with an attenuated line of Leishmania infantum (L. infantum H-line) were evaluated in mixed breed dogs. Two groups of dogs were infected intravenously (i.v.) or intradermally (i.d.) with L. infantum H-line and two control groups were infected i.v. or i.d. with L. infantum wild-type (L. infantum WT). None of the dogs, which were infected i.v. or i.d. with L. infantum H-line, showed any abnormalities during the observation period. In contrast, two out of three dogs, which were infected i.v. with L. infantum WT, developed clinical signs of disease. In addition, no histopathological changes were seen in the liver and spleen of the dogs infected with the attenuated line of parasite, whereas the histopathological changes in the two dogs infected i.v. with L. infantum WT were severe in form and manifested by infiltration of high numbers of inflammatory cells. No promastigotes were found in cultures set up from spleens and livers of dogs infected with L. infantum H-line at 12 months post-infection, whereas promastigotes were seen in the spleen and liver cultures from 2 dogs infected i.v. with L. infantum WT. Serum levels of total IgG anti-Leishmania antibody were raised in all dogs. The antibody level in the serum of dogs infected i.v. with L. infantum WT was higher than that in dogs infected with L. infantum H-line. These results show no clinicopathological abnormalities in the dogs infected with gentamicin-attenuated L. infantum H-line. Moreover, L. infantum H-line induced IgG anti-Leishmania antibody in the dogs.

Introduction: Brain tumor margin detection remains a challenging problem in the operative resection of gliomas. A novel nanoparticle, a PEGylated quantum dot, has been shown to be phagocytized by macrophages in vivo . This feature may allow quantum dots to co-localize with brain tumors and serve as an optical aid in the surgical resection of brain tumors. Methods: Sprague-Daly rats were injected intracranially with C6 gliosarcoma cell lines to establish tumors. Two weeks after implantation of brain tumors, PEGylated quantum dots emitting at 705 nm (PEG-705 QD) were injected via the tail vein. Twenty-four hours post PEG-705 QD injection, the animals were sacrificed and their tissues examined. Results: PEGylated quantum dots are avidly phagocytized by macrophages and are taken up by liver, spleen and lymph nodes. Macrophages and microglia co-localize with glioma cells, carrying the optical nanoparticle, the quantum dot. Excitation of the PEG-705 quantum dots gives off a deep red fluorescence detectable with charge coupled device (CCD) cameras, optical spectroscopy units, and in dark field fluorescence microscopy. Conclusions: PEG-705QDs co-localize with brain tumors and may serve as an optical adjunct to aid in the operative resection of gliomas. The particles may be visualized in surgery with CCD cameras or detected by optical spectroscopy.

An attenuated line of Leishmania mexicana (the L. mexicana H-line) has been established by culturing in vitro under gentamicin pressure. BALB/c mice infected with the L. mexicana H-line developed a CD4(+)Th1-like response, indicated by the cytokine profile of their splenocytes stimulated by L. mexicana wild-type (WT) promastigotes. This profile is sustained after these mice are challenged with L. mexicana WT. Control mice infected with L. mexicana WT alone developed a CD4(+)Th2-like cytokine profile. In mice immunized with L. mexicana H-line and then challenged with WT-line, were eliminated when immunizing H-line parasites persisted in the skin and draining popliteal lymph nodes (PLNs). In experiments in which mice were inoculated with attenuated and WT parasites at the same time, either at the same site or on separate sides of the mouse, growth of the WT parasites was significantly contained and controlled, indicating a possible therapeutic role for the attenuated parasites.

The objective of this study was to compare strain and gender differences in kidney and heart norepinephrine (NE) content and turnover rate in normotensive Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR, SHR/a, and SHR/y). Our laboratory has shown that the Y chromosome has a significant effect on blood pressure in the SHR model of hypertension through the use of two new rat stains, SHR/a and SHR/y, to study the Y chromosome. SHR/a have a SHR autosomal genetic background with a WKY Y chromosome, whereas the SHR/y rats have a WKY autosomal genetic background with a SHR Y chromosome. Tissues were homogenized after α-methyl-dl- p-tyrosine injection and analyzed for NE. The male kidney NE content was significantly lower in the WKY compared with the SHR, SHR/y, and SHR/a. Kidney and heart NE content was significantly higher in females compared with males in all strains except the SHR/y. The WKY and SHR/y females had significantly lower kidney NE turnover rates, and the SHR and SHR/a females had significantly higher kidney NE turnover rates than strain-matched males. This study suggests both a strain and gender difference in sympathetic nervous system activity through noradrenergic neurotransmission.

This study reports the concentration of collagen and its hydroxypyridinoline crosslinks, collagen fibril organization in the dorsal aortas, and systolic blood pressure during the progression of atherosclerosis in Japanese quail selected for cholesterol-induced atherosclerosis. The quail were placed on either a control or 0.5% cholesterol-added diet at approximately 16 weeks of age. The concentration of total collagen did not change in the control arteries during the course of the study, whereas at 5 and 10 weeks of cholesterol feeding, collagen levels decreased in the cholesterol-fed birds. Hydroxypyridinoline concentration increased during the duration of the study in the cholesterol-fed birds and by 15 and 20 weeks of cholesterol feeding, levels were significantly increased over those observed in the control arteries. Transmission electron microscopy showed changes in the organization of collagen fibrils. Increased systolic blood pressure was noted beginning at 10 weeks of cholesterol feeding, which is suggestive of other systemic changes induced by hypercholesterolemia. These results demonstrated remodeling of the collagen component of the dorsal aorta extracellular matrix during the progression of atherosclerosis and are suggestive of other systemic cardiovascular system changes.

The hypothesis was tested that the sympathetic nervous system (SNS) developmentally influences circulating testosterone (T), systolic blood pressure (SBP) and cardio-renal pathology in SHR/y animals. A sympathoplegic drug, guanethidine, and an antibody to nerve growth factor were administered to WKY and borderline hypertensive SHR/y male rats (n = 20/group) for the first 3 weeks of life; control groups (n = 20/group) received saline. SBP, serum T and luteinizing hormone (LH) were measured. SBP in the WKY and SHR/y sympathectomy (sympx) groups decreased 10mmHg (p0.001) and 50mmHg (p0.001), respectively, when compared to their control groups. Serum T levels in the sympx WKY group were lower (p0.01) than those in controls, and the rise of T typically observed in SHR/y from weeks 6-8 was delayed in the sympx SHR/y group, similar to the pattern in WKY. Serum LH levels were increased in the sympx WKY group, but not in the SHR/y group. Sympx caused a greater reduction in renal glomerular changes (p0.01), coronary artery collagen deposition (p0.01) and myocardial fibrosis (p0.01) in SHR/y than WKY rats. In conclusion, the SHR Y chromosome has a locus that enhances SNS activity, which can raise SBP and result in renal and cardiovascular tissue damage.