Regulation of stem cell factor expression in inflammation and asthma

Stem cell factor (SCF) is a major mast cell growth factor, which could be involved in the local increase of mast cell number in the asthmatic airways. In vivo, SCF expression increases in asthmatic patients and this is reversed after treatment with glucocorticoids. In vitro in human lung fibroblasts in culture, IL-1beta, a pro-inflammatory cytokine, confirms this increased SCF mRNA and protein expression implying the MAP kinases p38 and ERK1/2 very early post-treatment, and glucocorticoids confirm this decrease. Surprisingly, glucocorticoids potentiate the IL-1beta-enhanced SCF expression at short term treatment, implying increased SCF mRNA stability and SCF gene transcription rate. This potentiation involves p38 and ERK1/2. Transfection experiments with the SCF promoter including intron1 also confirm this increase and decrease of SCF expression by IL-1beta and glucocorticoids, and the potentiation by glucocorticoids of the IL-1beta-induced SCF expression. Deletion of the GRE or kappaB sites abolishes this potentiation, and the effect of IL-1beta or glucocorticoids alone. DNA binding of GR and NF-kappaB are also demonstrated for these effects. In conclusion, this review concerns new mechanisms of regulation of SCF expression in inflammation that could lead to potential therapeutic strategy allowing to control mast cell number in the asthmatic airways.

Role of protease-activated receptor-2 in inflammation, and its possible implications as a putative mediator of periodontitis

Proteinase-activated receptor-2 (PAR2) belongs to a novel subfamily of G-protein-coupled receptors with seven-transmembrane domains. This receptor is widely distributed throughout the body and seems to be importantly involved in inflammatory processes. PAR2 can be activated by serine proteases such as trypsin, mast cell tryptase, and bacterial proteases, such as gingipain produced by Porphyromonas gingivalis. This review describes the current stage of knowledge of the possible mechanisms that link PAR2 activation with periodontal disease, and proposes future therapeutic strategies to modulate the host response in the treatment of periodontitis.

Trypanosoma cruzi strains isolated from human, vector, and animal reservoir in the same endemic region in Mexico and typed as T. cruzi I, discrete typing unit 1 exhibit considerable biological diversity

In this study, three strains of Trypanosoma cruzi were isolated at the same time and in the same endemic region in Mexico from a human patient with chronic chagasic cardiomyopathy (RyC-H); vector (Triatoma barberi) (RyC-V); and rodent reservoir (Peromyscus peromyscus) (RyC-R). The three strains were characterized by multilocus enzyme electrophoresis, random amplified polymorphic DNA, and by pathological profiles in experimental animals (biodemes). Based on the analysis of genetic markers the three parasite strains were typed as belonging to T. cruzi I major group, discrete typing unit 1. The pathological profile of RyC-H and RyC-V strains indicated medium virulence and low mortality and, accordingly, the strains should be considered as belonging to biodeme Type III. On the other hand, the parasites from RyC-R strain induced more severe inflammatory processes and high mortality (> 40%) and were considered as belonging to biodeme Type II. The relationship between genotypes and biological characteristics in T. cruzi strains is still debated and not clearly understood. An expert committee recommended in 1999 that Biodeme Type III would correspond to T. cruzi I group, whereas Biodeme Type II, to T. cruzi II group. Our findings suggest that, at least for Mexican isolates, this correlation does not stand and that biological characteristics such as pathogenicity and virulence could be determined by factors different from those identified in the genotypic characterization

Genetic variability of hepatitis A virus strain HAF-203 isolated in Brazil and expression of the VP1 gene in Escherichia coli

The hepatitis A virus (HAV) HAF-203 strain was isolated from an acute case of HAV infection. The primary isolation of HAF-203 in Brazil and its adaptation to the FRhK-4 cell lineage allowed the production of large amounts of viral particles enabling molecular characterization of the first HAV isolate in Brazil. The aim of our study was to determine the nucleotide sequence of the HAF-203 strain genome, compare it to other HAV genomes and highlight its genetic variability. The complete nucleotide sequence of the HAF-203 strain (7472 nucleotides) was compared to those obtained earlier by others for other HAV isolates. These analyses revealed 19 HAF-specific nucleotide sequence differences with 10 amino acid substitutions. Most of the non-conservative changes were located at VP1, 2C, and 3D genes, but the 3B region was the most variable. The availability of HAF-203 complementary DNA was useful for the production of the recombinant VP1 protein, which is a major determinant of viral infectivity. This recombinant protein was shown by enzyme-linked immunoassay and blotting, to be immunogenic and resemble the native protein, therefore suggesting its value as a reagent for incorporation into diagnostic tests.

Leishmania (Viannia) lainsoni: occurrence of intracellular promastigote forms in vivo and in vitro

Experimental chronic (45-day-old) skin lesion in hamster hind foot induced by Leishmania (Viannia) lainsoni infection showed the presence of promastigote forms in the tissue, inside parasitophorous vacuoles, as assessed by transmission electron microscopy. Experimental in vitro interaction (24 and 48 h) between Leishmania (V.)lainsoni and J774-G8 macrophage cells also demonstrated the same profile. This morphological aspect is unusual, since in this parasite genus only amastigote forms have been described as the resistant and obligate intracellular forms.

Plasmodium vivax thrombospondin related adhesion protein: immunogenicity and protective efficacy in rodents and Aotus monkeys

The thrombospondin related adhesion protein (TRAP) is a malaria pre-erythrocytic antigen currently pursued as malaria vaccine candidate to Plasmodium falciparum. In this study, a long synthetic peptide (LSP) representing a P. vivax TRAP fragment involved in hepatocyte invasion was formulated in both Freund and Montanide ISA 720 adjutants and administered by IM and subcutaneous routes to BALB/c mice and Aotus monkeys. We measured specific humoral immune responses in both animal species and performed a sporozoite challenge in Aotus monkeys to assess the protective efficacy of the vaccine. After immunization both mice and Aotus seroconverted as shown by ELISA, and the specific anti-peptide antibodies cross reacted with the parasite in IFAT assays. Only two out of six immunized animals became infected after P. vivax sporozoite challenge as compared with four out of six animals from the control group. These results suggest that this TRAP fragment has protective potential against P. vivax malaria and deserves further studies as vaccine candidate.

The involvement of tetA and tetE tetracycline resistance genes in plasmid and chromosomal resistance of Aeromonas in Brazilian strains

This study analyzed the involvement of tetA and tetE genes in the tetracycline resistance of 16 strains of genus Aeromonas, isolated from clinical and food sources. Polymerase chain reactions revealed that 37.5% of the samples were positive for tetA, and also 37.5% were tetE positive. One isolate was positive for both genes. Only the isolate A. caviae 5.2 had its resistance associated to the presence of a plasmid, pSS2. The molecular characterization of pSS2 involved the construction of its restriction map and the determination of its size. The digestion of pSS2 with HindIII originated two fragments (A and B) that were cloned separately into the pUC18 vector. The tetA gene was shown to be located on the HindIII-A fragment by PCR. After transforming a tetracycline-sensitive strain with pSS2, the transformants expressed the resistance phenotype and harbored a plasmid whose size was identical to that of pSS2. The results confirmed the association between pSS2 and the tetracycline resistance phenotype, and suggest a feasible dissemination of tetA and tetE among strains of Aeromonas. This study suggests the spreading tetA and tetE genes in Aeromonas in Brazil and describes a resistance plasmid that probably contributes to the dissemination of the resistance.

Access to diagnosis and treatment of Chagas disease/infection in endemic and non-endemic countries in the XXI century

In this article, Médicos Sin Fronteras (MSF) Spain faces the challenge of selecting, piecing together, and conveying in the clearest possible way, the main lessons learnt over the course of the last seven years in the world of medical care for Chagas disease. More than two thousand children under the age of 14 have been treated; the majority of whom come from rural Latin American areas with difficult access. It is based on these lessons learnt, through mistakes and successes, that MSF advocates that medical care for patients with Chagas disease be a reality, in a manner which is inclusive (not exclusive), integrated (with medical, psychological, social, and educational components), and in which the patient is actively followed. This must be a multi-disease approach with permanent quality controls in place based on primary health care (PHC). Rapid diagnostic tests and new medications should be available, as well as therapeutic plans and patient management (including side effects) with standardised flows for medical care for patients within PHC in relation to secondary and tertiary level, inclusive of epidemiological surveillance systems.

Molecular identification of Rickettsia felis in ticks and fleas from an endemic area for Brazilian Spotted Fever

Rickettsioses are arthropod-borne diseases caused by parasites from the Order Rickettsiales. The most prevalent rickettsial disease in Brazil is Brazilian Spotted Fever (BSF). This work intends the molecular detection of those agents in ectoparasites from an endemic area of BSF in the state of Espírito Santo. A total of 502 ectoparasites, among them Amblyomma cajennense, Amblyomma dubitatum (A. cooperi), Riphicephalus sanguineus, Anocentor nitens and Ctenocephalides felis, was collected from domestic animals and the environment and separated in 152 lots according to the origin. Rickettsia sp. was detected in pools of all collected species by amplification of 17kDa protein-encoding gene fragments. The products of PCR amplification of three samples were sequenced, and Rickettsia felis was identified in R. sanguineus and C. felis. These results confirm the presence of Rickettsia felis in areas previously known as endemic for BSF, disease caused by Rickettsia rickettsii. Moreover, they show the needing of further studies for deeper knowledge of R. felis-spotted fever epidemiology and differentiation of these diseases in Brazil.

In vitro and in vivo activity of meglumine antimoniate produced at Farmanguinhos-Fiocruz, Brazil, against Leishmania (Leishmania) amazonensis, L (L.) chagasi and L (Viannia) braziliensis

The leishmanicidal activity of four batches of meglumine antimoniate, produced in Farmanguinhos-Fiocruz, Brazil (TAMs), was assessed and compared to Glucantime®-Aventis Pharma Ltda. Using the amastigote-like in vitro model, the active concentrations of Sb v varied from 10µg/ml to 300 µg/ml for L. (L.) chagasi and from 50µg/ml to 300µg/ml for L. (L.) amazonensis, with no statistically significant differences among the four batches of TAMs and Glucantime®. The inhibitory concentrations (IC50) determined by the amastigote-infected macrophage model for TAM01/03 and Glucantime® were, respectively: 26.3µg/ml and 127.6µg/ml for L. chagasi, 15.4µg /ml and 22.9µg/ml for L. amazonensis, and 12.1µg/ml and 24.2µg/ml for L. (V.) braziliensis. The activities of the four batches of TAMs were confirmed in an in vivo model by assessing, during eight weeks skin lesions caused by L. braziliensis in hamster that were treated with 20mg Sb v/Kg/day for 30 consecutive days. The meglumine antimoniate produced by Farmanguinhos was as effective as the reference drug, Glucantime®-Aventis, against three species of Leishmania that are of medical importance in Brazil.

IgA1 levels in milk and serum samples from intestinal parasite-infected or normal puerperae

In this study, IgA1 levels in the milk and serum of puerperae were compared and a correlation was established between the levels of this immunoglobulin and the occurrence of parasitism. Eighty-three paired milk and serum samples were obtained from puerperal and IgA1 levels were analyzed. In addition, the presence of intestinal parasites in stool samples from these puerperae was determined. Twelve puerperae tested positive for intestinal parasites and all their samples presented an IgA1 ELISA Index > 1. There was a correlation between serum and milk IgA1 levels and puerperae with any parasite in their stool (r = 0.6723; p = 0.0166). This finding may reinforce the importance of breast-feeding for the protection of neonates.

Mutations in rpoB and katG genes in Mycobacterium isolates from the Southeast of Mexico

The most frequent mutations associated with rifampin and isoniazid resistance in Mycobacterium are the substitutions at codons 531 and 315 in the rpoB and katG genes, respectively. Hence, the aim of this study was to characterize these mutations in Mycobacterium isolates from patients suspected to be infected with drug-resistant (DR) pulmonary tuberculosis (TB) in Veracruz, Mexico. Drug susceptibility testing of 25 clinical isolates revealed that five were susceptible while 20 (80%) were DR (15% of the annual prevalence for Veracruz). Of the DR isolates, 15 (75%) were resistant to rifampin, 17 (85%) to isoniazid and 15 (75%) were resistant to both drugs (MDR). Sequencing analysis performed in the isolates showed that 14 (93%) had mutations in the rpoB gene; seven of these (47%) exhibited a mutation at 531 (S[L). Ten (58%) of the 20 resistant isolates showed mutations in katG; nine (52%) of these 10 exhibited a mutation at 315 (S[T). In conclusion, the DR profile of the isolates suggests a significant number of different DR-TB strains with a low frequency of mutation at codons 531 and 315 in rpoB and katG, respectively. This result leads us to consider different regions of the same genes, as well as other genes for further analysis, which is important if a genetic-based diagnosis of DR-TB is to be developed for this region.

Effect of untreated bed nets on blood-fed Phlebotomus argentipes in kala-azar endemic foci in Nepal and India

Observational studies in the Indian subcontinent have shown that untreated nets may be protective against visceral leishmaniasis (VL). In this study, we evaluated the effect of untreated nets on the blood feeding rates of Phlebotomus argentipes as well as the human blood index (HBI) in VL endemic villages in India and Nepal. The study had a "before and after intervention" design in 58 households in six clusters. The use of untreated nets reduced the blood feeding rate by 85% (95% CI 76.5-91.1%) and the HBI by 42.2% (95% CI 11.1-62.5%). These results provide circumstantial evidence that untreated nets may provide some degree of personal protection against sand fly bites.

Experimental chemotherapy for Chagas disease: 15 years of research contributions from in vivo and in vitro studies

Chagas disease, which is caused by the intracellular parasite Trypanosoma cruzi, is a neglected illness with 12-14 million reported cases in endemic geographic regions of Latin America. While the disease still represents an important public health problem in these affected areas, the available therapy, which was introduced more than four decades ago, is far from ideal due to its substantial toxicity, its limited effects on different parasite stocks, and its poor activity during the chronic phase of the disease. For the past 15 years, our group, in collaboration with research groups focused on medicinal chemistry, has been working on experimental chemotherapies for Chagas disease, investigating the biological activity, toxicity, selectivity and cellular targets of different classes of compounds on T. cruzi. In this report, we present an overview of these in vitro and in vivo studies, focusing on the most promising classes of compounds with the aim of contributing to the current knowledge of the treatment of Chagas disease and aiding in the development of a new arsenal of candidates with anti-T. cruzi efficacy.