Hyperbaric oxygen augments susceptibility to C. difficile infection by impairing gut microbiota ability to stimulate the HIF-1α-IL-22 axis in ILC3.

Hyperbaric oxygen (HBO) therapy is a well-established method for improving tissue oxygenation and is typically used for the treatment of various inflammatory conditions, including infectious diseases. However, its effect on the intestinal mucosa, a microenvironment known to be physiologically hypoxic, remains unclear. Here, we demonstrated that daily treatment with hyperbaric oxygen affects gut microbiome composition, worsening antibiotic-induced dysbiosis. Accordingly, HBO-treated mice were more susceptible to Clostridioides difficile infection (CDI), an enteric pathogen highly associated with antibiotic-induced colitis. These observations were closely linked with a decline in the level of microbiota-derived short-chain fatty acids (SCFAs). Butyrate, a SCFA produced primarily by anaerobic microbial species, mitigated HBO-induced susceptibility to CDI and increased epithelial barrier integrity by improving group 3 innate lymphoid cell (ILC3) responses. Mice displaying tissue-specific deletion of HIF-1 in RORγt-positive cells exhibited no protective effect of butyrate during CDI. In contrast, the reinforcement of HIF-1 signaling in RORγt-positive cells through the conditional deletion of VHL mitigated disease outcome, even after HBO therapy. Taken together, we conclude that HBO induces intestinal dysbiosis and impairs the production of SCFAs affecting the HIF-1α-IL-22 axis in ILC3 and worsening the response of mice to subsequent C. difficile infection.

Monitoring Leishmania infantum Infections in Female Lutzomyia longipalpis by Using DNA Extraction on Cation Exchange Paper and PCR Pool Testing

Visceral leishmaniasis remains a serious public health issue, and Brazil was among the seven countries with the highest prevalence of this disease worldwide. The measures to control this disease are not easily developed, and the improvement of its diagnosis, surveillance, and control is still needed. This study aimed to carry out the polymerase chain reaction (PCR) diagnosis of Leishmania infantum in vector samples in some municipalities of the State of São Paulo, which included two municipalities with human disease transmission and two with dog transmission only. Vectors were collected in traps with luminous bait. Next, they were killed at -4 °C and kept in 70% alcohol. Groups of ten female insects (pools) were mashed on cation exchange paper (fine cellulose phosphate with 18 µEq/cm² ionic exchange capacity) for DNA extraction. The PCR was carried out to identify the natural infection of the Leishmania genus in female Lutzomyia longipalpis (Lu. Longipalpis). Out of the 3,880 Lu. longipalpis phlebotomines, 1060 were female and 2820 were male (3:1). The method used to extract the DNA in pools of ten phlebotomines and the PCR resulted in sensitivity, specificity, practicality, and faster analyses when compared to the individual analysis method. The procedure described can be used on a large scale in the leishmaniasis epidemiological surveillance, enabling a higher number of analyses and the optimization of human resources because the traditional diagnostic method is carried out via desiccation of the insect digestive system and microscopic examination, which is time-demanding and there is the need of manual skills.

Aspectos evolutivos dos fenômenos imunopatológicos com ênfase na COVID-19 / Evolutionary aspects of immunopathological phenomena with emphasis on COVID-19

A seleção natural é o principal mecanismo da evolução das espécies, e favorece fenótipos com defesas imunes efetivas contra patógenos. Entretanto, há uma grande variação das respostas imunes entre os indivíduos da espécie humana e a ocorrência de fenômenos imunopatológicos. A infecção com o vírus da família Coronaviridae, SARS-CoV-2, responsável pela doença conhecida como COVID-19, induz a respostas imunes inflamatórias exacerbadas e à tempestade de citocinas, nos casos graves. Nesta revisão discutiremos, à luz da Evolução, esse aparente paradoxo entre as respostas imunes, e os três principais fatores que contribuem para a manutenção dos fenótipos hiperativos: o custo-benefício das respostas imunes, a coevolução e a história de vida da espécie.

Natural selection is the main mechanism by which species evolve, and it favors phenotypes associated with an effective immune defense against pathogens. However, human immune responses and the occurrence of immunopathological phenomena vary considerably from individual to individual. Infection with SARSCoV- 2, a virus of the Coronaviridae family causing the disease known as COVID-19, induces exacerbated inflammatory immune responses and cytokine storm in severe cases. In this review, we discuss, in the light of Evolution, this apparent paradox between the immune responses and the 3 main factors contributing to the maintenance of hyperactive phenotypes: the cost-effectiveness of immune responses, coevolution, and the life history of the species.

Evaluation of prime and prime-boost immunization strategies in BALB/c mice inoculated with Leishmania infantum transfected with toxic plasmids.

The etiologic agents of visceral leishmaniasis are Leishmania infantum and Leishmania donovani. Despite the variety of drugs available to treat leishmaniasis, most lead to serious adverse effects, and resistance to these drugs has been reported. Currently, no leishmaniasis vaccine is available for humans. That is why the group developed transgenic L. infantum promastigote lines, which express toxic proteins after differentiation into amastigotes. That is why group developed the pFL-AMA plasmid and transfected it into L. Infantum promastigotes. This plasmid was expressed only in the amastigote form of the parasite. Sequences encoding toxic proteins (active bovine trypsin and egg avidin) were inserted in this plasmid, and the transfected parasites died after the differentiation process. In this study, two immunization protocols were performed in BALB/c mice: prime and prime-boost immunization prior to challenge with the wild-type L. infantum (WT). The parasite burdens in the spleen, liver, and bone marrow were evaluated to verify immunological protection. Histopathological analysis of the spleen and liver and the humoral immune response were also performed. The data showed that the parasite burden was reduced in prime-boosted mice in the spleen, liver, and bone marrow, indicating that mice immunized with two doses of the transfected parasites were satisfactorily protected. High levels of IgG, IgG1, and IgG2a antibodies were observed, as well as the presence of anti-inflammatory cytokine Interleukine-10 and pro-inflammatory cytokine Tumor Necrosis Factor-α (TNF-α) and Interferon-γ (IFN - γ) suggesting a Th1/Th2 mix response, in addition to the presence of multinucleated giant cells in the spleen and lymphocyte infiltration in the liver. Therefore, L. infantum transfected with a toxic plasmid is an excellent vaccine candidate against visceral leishmaniasis and the application of a boost before the challenge promotes greater protection against WT L. infantum infection.

Virulence and DNA sequence analysis of Cronobacter spp. isolated from infant cereals.

Cronobacter spp. is an opportunistic pathogen that causes severe infections, affecting newborns and infants, and is also an emerging cause of hospital-acquired infection in elderly populations. These infections are mainly associated with the consumption of infant formulas, even though these bacteria have been isolated from other foods as well. Cronobacter spp. invades epithelial cells and escapes the immune response mechanisms, multiplying inside macrophages. However, the pathogenesis and virulence factors of these bacteria have not been fully elucidated and need to be further studied. Therefore, this study aimed to evaluate the ability of Cronobacter spp. strains isolated from infant cereals to invade and survive within macrophages, investigate the virulence phenotype using the Galleria mellonella model, and identify possible genes involved in bacterial pathogenesis through pan-genome analysis. All the isolates were able to invade macrophages and the survival of bacteria decreased over a 72 h period, with bacterial cell counts reaching up to 106 CFU/ml. Cronobacter sakazakii isolate 112 exhibited a similar mortality rate (40-70%) to the ATCC BAA 894 strain (Cronobacter sakazakii) in G. mellonella assay. In addition, some unique virulence genes (isolate 7, ada_2, tcmA_1, acrB_3; isolate 78, ampC_2, rihC_1 and isolate 112, fimH, ylpA, gtrA) were identified within isolates with the invasive profile in the in vivo and in vitro assays. Furthermore, isolates from different species were grouped into seven distinct clusters in the pan-genome analysis. The most virulent isolates (7, 78, and 112) were grouped in distinct subclusters in the cladogram. This work revealed potential Cronobacter spp. pathogenic strains recovered from infant cereals.

Leishmania (Viannia) braziliensis replicates in mouse bone marrow.

Leishmaniasis is a neglected disease caused by species of the protozoan Leishmania. Leishmania (Viannia) braziliensis causes the cutaneous and mucocutaneous forms of the disease. Experimental cutaneous infection of mice is one of the most important preclinical research models of leishmaniasis. Here, we investigated the course of infection in mice inoculated with two reference strains of L. (V.) braziliensis (MHOM/BR/00/BA788 strain [BA] and MHOM/BR/94/H-3227 strain [CE]). Although both parasite strains induced detectable footpad lesions, BA-infected mice developed small non-ulcerated lesions that self-healed, whereas CE-infected mice developed small non-ulcerated lesions that did not regress. The parasites were detected in the footpad lesions, lymph nodes draining the site of inoculation, spleen, and bone marrow of mice infected with BA or CE parasites at 6 and 25 weeks post-inoculation. These data indicate that L (V.) braziliensis-infected mice harbor parasites that spread, even when these animals do not display overt lesions. In addition, this is the first report of the presence of the parasite in the bone marrow of mice inoculated with L. (V.) braziliensis.

Leishmania and its relationships with bacteria.

Leishmaniasis is a zoonotic and neglected disease, which represents an important public health problem worldwide. Different species of Leishmania are associated with different manifestations, and a practical problem that can worsen the condition of hosts infected with Leishmania is the secondary infection caused by bacteria. This review aims to examine the importance and prevalence of bacteria co-infection during leishmaniasis and the nature of this ecological relationship. In the cases discussed in this review, the facilitation phenomenon, defined as any interaction where the action of one organism has a beneficial effect on an organism of another species, was considered in the Leishmania-bacteria interaction, as well as the effects on one another and their consequences for the host.

Improved efficacy of meglumine antimoniate incorporated in anionic liposomes against Leishmania infantum infecting canine macrophages.

OBJECTIVES: Leishmaniasis is a zoonotic disease and several drugs have been used in the treatment, including meglumine antimoniate (AME). The chemotherapy reaches clinical cure but does not eliminate parasites, contributing to drug resistance. To improve AME efficacy we incorporated it in anionic liposomes. The antiparasitic activity and intracellular localization were investigated in canine macrophages infected with Leishmania infantum. METHODS: Liposomes (L-AME) is composed of egg phosphatidylcholine, cholesterol, palmitoyl oleoyl phosphatidyl serine and α-tocopherol (4 : 3 : 0.4 : 0.07 mol%) plus AME. L-AME size, polydispersity, zeta potential and morphology were analysed as well as antileishmanial activity and intracellular localization in DH82 macrophages. KEY FINDINGS: Liposomes (360 nm) zeta potential range from -40 to -65 mV, had 23% encapsulation efficiency and were stable for 180 days at 4°C. Free AME was cytotoxic towards L. infantum infected macrophages (ID50 = 0.012 M) while L-AME did not reduce cell viability. L-AME colocalized with parasites inside macrophages in a time-dependent manner, and reduced the percentage of infected cells and the number of intracellular parasites, decreasing the infection index (75-80%) twice as compared with AME treatment. CONCLUSIONS: Liposomal AME is a promising delivery system for treating visceral leishmaniasis, improving meglumine efficacy against L. infantum and minimizing its cytotoxicity towards canine macrophages.

Exploring TERRA during Leishmania major developmental cycle and continuous in vitro passages.

Telomeres from different eukaryotes, including trypanosomatids, are transcribed into TERRA noncoding RNAs, crucial in regulating chromatin deposition and telomere length. TERRA is transcribed from the C-rich subtelomeric strand towards the 3'-ends of the telomeric array. Using bioinformatics, we confirmed the presence of subtelomeric splice acceptor sites at all L. major chromosome ends. Splice leader sequences positioned 5' upstream of L. major chromosomes subtelomeres were then mapped using SL-RNA-Seq libraries constructed from three independent parasite life stages and helped confirm TERRA expression from several chromosomes ends. Northern blots and RT-qPCR validated the results showing that L. major TERRA is processed by trans-splicing and polyadenylation coupled reactions. The number of transcripts varied with the parasite's life stage and continuous passages, being more abundant in the infective forms. However, no putative subtelomeric promoters involved in TERRA's transcriptional regulation were detected. In contrast, the observed changes in parasite's telomere length during development, suggest that differences in telomeric base J levels may control TERRA transcription in L. major. Also, TERRA-R loops' detection, mainly in the infective forms, was suggestive of TERRA's involvement in telomere protection. Therefore, Leishmania TERRA shares conserved features with other eukaryotes and advances new telomere specific functions in a Public Health-impacting parasite.

Phenotype evaluation of human and canine isolates of Leishmania infantum.

Human visceral leishmaniasis (VL) and canine leishmaniasis (CanL) in countries of South and Central America are caused by Leishmania infantum and has been endemic in Brazil for several years. The parasite biodiversity as well as the pharmacologic properties of drugs and the host species, are involved in the efficacy or inefficacy of leishmaniasis treatments. Although there are substantial number of reports describing the genetic characterization of the clinical field isolates of L. infantum,the phenotypic parameters have been less studied. In this study isolates from human and canine leishmaniasis (Hum1 and Can1) obtained in Campinas, São Paulo state, Brazil were identified as L. infantum. The Hum1 and Can1 isolates exhibited typical promastigote growth pattern. Regarding morphological features Can1 isolate differed in cell size. The infectivity in vitro of both isolatesis lower compared to the reference strain of L. infantum. Moreover, the in vivo infectivity of the three parasites is similar in Balb/c mice. The Hum1 isolate is more sensitive to leishmanial drugs (amphotericin B, miltefosine and glucantime) than the Can1 isolate when inside human macrophages, but not when inside canine macrophages. These findings indicated that L. infantum isolates differs in some phenotypic characteristics.

Leishmania infantum transfected with toxic plasmid induces protection in mice infected with wild type L. infantum or L. amazonensis.

Leishmania infantum infection may cause visceral leishmaniasis (VL), a fatal disease having worldwide distribution, that may be silent or asymptomatic. The latter indicates that immunity is naturally developed in some individuals, and, therefore, a vaccine against VL would be possible. Molecular mechanisms of gene expression are being understood in Leishmania, and this knowledge may be useful for vaccine development. The aim of this study was developing an attenuated strain by regulating the expression of toxic proteins in a stage specific manner. For that purpose, the 3' UTR of an amastin gene, known by its increased expression in the amastigote phase, was selected for direct the expression of exogenous proteins. This construct (pFL-AMA), firstly, was proved effective for the expression of mCherry specifically in the intracellular form of L. infantum, as demonstrated by fluorescence microscopy, flow cytometry and Western blotting. Afterwards, mCherry coding sequence was replaced, in the pFL-AMA plasmid, by either egg avidin or the active form of bovine trypsin. Viability of transfected parasites was evaluated in promastigote axenic cultures and in in vitro infection of macrophages. Both lines of transfected parasites showed a limited capacity to multiply inside macrophages. BALB/c mice were inoculated intraperitoneally (i.p.) with a single dose consisting of 2 × 106L. infantum promastigotes transfected with plasmids bearing the toxic genes. After 10 weeks post-inoculation, no parasites were recovered by limiting dilution in either liver or spleen, but a specific immunological response was detected. The immunization with transfected parasites induced cellular and humoral immune responses with activation of TCD4+, TCD8+ and B cells, having a TH1-type response with increased levels of pro-inflammatory cytokines such as IFN-γ, TNF-α and IL-6. In parallel groups of mice, a challenge consisting on 1 × 106 virulent parasites of either L. infantum (inoculated i.p.) or L. amazonensis subcutaneously (s.c.) was performed. Vaccinated mice, challenged with L. infantum, showed lower parasite burdens in liver, spleen and bone marrow than infected mice with WT L. infantum (non-vaccinated); similarly, vaccinated mice developed smaller footpad inflammation than control group. These data support this strategy as an efficient immunization system aimed to the development of vaccines against different forms of leishmaniasis.

Granulomas in parasitic diseases: the good and the bad.

Parasitic diseases affect more than one billion people worldwide, and most of them are chronic conditions in which the treatment and prevention are difficult. The appearance of granulomas, defined as organized and compact structures of macrophages and other immune cells, during various parasitic diseases is frequent, since these structures will only form when individual immune cells do not control the invading agent. Th2-typering various parasitic diseases are frequent, since these structures will only form when individual immune cells do not control the invading agent. The characterization of granulomas in different parasitic diseases, as well as recent findings in this field, is discussed in this review, in order to understand the significance of the granuloma and its modulation in the host-parasite interaction and in the immune, pathological, and parasitological aspects of this interaction. The parasitic granulomatous diseases granulomatous amebic encephalitis, toxoplasmosis, leishmaniasis, neurocysticercosis, and schistosomiasis mansoni are discussed as well as the mechanistic and dynamical aspects of the infectious granulomas.

Tandem Mass Tag Proteomic Analysis of in Vitro and in Vivo Models of Cutaneous Leishmaniasis Reveals Parasite-Specific and Nonspecific Modulation of Proteins in the Host.

Cutaneous leishmaniasis, the most common form of leishmaniasis, is endemic in several regions of the world, and if not treated properly, it can cause disfiguring scars on the skin. Leishmania spp. infection causes an inflammatory response in its host, and it modulates the host metabolism differently depending on the Leishmania species. Since Leishmania spp. has begun to develop resistance against current therapies, we believe efforts to identify new possibilities for treatment are critical for future control of the disease. Proteomics approaches such as isobaric labeling yield accurate relative quantification of protein abundances and, when combined with chemometrics/statistical analysis, provide robust information about protein modulation across biological conditions. Using a mass spectrometry-based proteomics approach and tandem mass tag labeling, we have investigated protein modulation in murine macrophages (in vitro model) and skin biopsies after exposure to Leishmania spp. (in vivo murine model). Infections induced by L. amazonensis (endemic in the New World) and L. major (endemic in the Old World) were compared to an inflammation model to search for Leishmania-specific and nonspecific protein modulation in the host. After protein extracts obtained from in vitro and in vivo experiments were digested, the resulting peptides were labeled with isobaric tags and analyzed by liquid chromatography-MS (LC-MS). Several proteins that were found to be changed upon infection with Leishmania spp. provide interesting candidates for further investigation into disease mechanism and development of possible immunotherapies.

Comparative Proteomic Analysis of Murine Cutaneous Lesions Induced by Leishmania amazonensis or Leishmania major.

Cutaneous leishmaniasisis is the most common clinical form of leishmaniasis and one of the most relevant neglected diseases. It is known that the progress of the disease is species specific and the host's immune response plays an important role in its outcome. However, the pathways that lead to parasite clearance or survival remain unknown. In this work, skin tissue from mice experimentally infected with L. amazonensis, one of the causative agents of cutaneous leishmaniasis in the Amazon region, L. major, another causative agent of cutaneous leishmaniasis in Africa, the Middle East, China, and India, or lipopolysaccharides from Escherichia coli as an inflammation model were investigated using label-free proteomics to unveil Leishmania-specific protein alterations. Proteomics is a powerful tool to investigate host-pathogen relationships to address biological questions. In this work, proteins from mice skin biopsies were identified and quantified using nano-LC coupled with tandem mass spectrometry analyses. Integrated Proteomics Pipeline was used for peptide/protein identification and quantification. Western blot was used for validation of protein quantification by mass spectrometry, and protein pathways were predicted using Ingenuity Pathway Analysis. In this proteomics study, several proteins were pointed out as hypothetical targets to guide future studies on Leishmania-specific modulation of proteins in the host. We identified hundreds of exclusively modulated proteins after Leishmania spp. infection and 17 proteins that were differentially modulated in the host after L. amazonensis or L. major infection.

Evaluation of antileishmanial drugs activities in an ex vivo model of leishmaniasis.

Leishmaniasis is a poverty-related disease, the chemotherapy of which is based on few drugs. The in vitro macrophage-amastigote model using mouse peritoneal cells, human-monocyte transformed macrophages and immortalized cell lines have been used to test new and safe antileishmanial drugs. Considering the differences for drug sensitivities between these Leishmania infected cells, the efficacy of amphotericin B, pentavalent antimonial, miltefosine and resveratrol was evaluated in a recently developed ex vivo culture of macrophages isolated from mouse lesion induced by L. amazonensis (CD11b+F4/80+CD68+CD14+) compared with infected peritoneal macrophages (CD11b+F4/80+CD68+CD14+). The results show that IC50 values of amphotericin B, miltefosine and pentavalent antimonial for parasites in lesional and peritoneal macrophages were similar, although high doses of these compounds did not result in total clearance of parasites in lesional cells (amphotericin B), peritoneal cells (miltefosine) and both cell cultures (pentavalent antimonial). Amastigotes infecting lesional macrophages were more resistant to resveratrol as compared to parasites in peritoneal macrophages. The cytoxicity of miltefosine and resveratrol was higher in infected peritoneal macrophages than in lesional cells. These data suggest that the antileishmanial effect and citotoxicity of some anti leishmanial compounds are dependent of macrophage source and mouse peritoneal macrophages loaded with amastigotes do not represent the lesion cell.

Label-Free Proteomic Analysis Reveals Parasite-Specific Protein Alterations in Macrophages Following Leishmania amazonensis, Leishmania major, or Leishmania infantum Infection.

Leishmania is an obligate intracellular parasite known to modulate the host cell to survive and proliferate. However, the complexity of host-parasite interactions remains unclear. Also, the outcome of the disease has been recognized to be species-specific and dependent on the host's immune responses. Proteomics has emerged as a powerful tool to investigate the host-pathogen interface, allowing us to deepen our knowledge about infectious diseases. Quantification of the relative amount of proteins in a sample can be achieved using label-free proteomics, and for the first time, we have used it to quantify Leishmania-specific protein alterations in macrophages. Protein extracts were obtained and digested, and peptides were identified and quantified using nano-LC coupled with tandem mass spectrometry analyses. Protein expression was validated by Western blot analysis. Integrated Proteomics Pipeline was used for peptide/protein identification and for quantification and data processing. Ingenuity Pathway Analysis was used for network analysis. In this work, we investigated how this intracellular parasite modulates protein expression on a host macrophage by comparing three different Leishmania species- L. amazonensis, one of the causative agents of cutaneous disease in the Amazon region; L. major, another causative agent of cutaneous leishmaniasis in Africa, the Middle East, China, and India; L. infantum, the causative agent of visceral leishmaniasis affecting humans and dogs in Latin America-and lipopolysaccharide stimulated macrophages as an in vitro inflammation model. Our results revealed that Leishmania infection downregulates apoptosis pathways while upregulating the activation of phagocytes/leukocytes and lipid accumulation.

In vitro immunotoxicological assessment of a potent microbicidal nanocomposite based on graphene oxide and silver nanoparticles.

Graphene oxide (GO) and silver nanoparticles (AgNPs) can be formed into a hybrid nanomaterial, known as GOAg nanocomposite, which presents high antibacterial activity. The successful translation of this nanomaterial into medical use depends on critical information about its toxicological profile. In keeping with a Safe-by-design approach, we evaluated the immunotoxicity of GOAg using J774 and primary murine macrophages. The interaction between GOAg and macrophages was investigated with a scanning electron microscope (SEM). High-throughput technologies were employed to evaluate cell viability, apoptosis/necrosis, mitochondrial depolarization and lipid peroxidation. The inflammogenicity of nanomaterials was predicted after quantification of the cytokines IL-1ß, TNF-α and IL-10 before and after stimulation with interferon-γ (IFN-γ). The ratio between CD80 and CD206 macrophage populations were also estimated. In addition, the production of nitric oxide (NO) was investigated. SEM surveys revealed the potential of GOAg to induce frustrated phagocytosis. GOAg induced a dose-dependent mitochondrial depolarization, apoptosis and lipid peroxidation to J774 macrophages. GOAg toxicity was not modified in an inflammatory microenvironment, but its toxicity was within the range of concentrations used in bacterial inactivation. GOAg did not induce primary macrophages to significantly produce inflammatory cytokines, and previous macrophage stimulation did not enhance GOAg inflammogenicity. Additionally, the pristine nanomaterials and GOAg do not shift macrophages polarization towards M1. Sublethal concentrations of GOAg did not impair macrophages NO production. Finally, we suggest options for improvement of GOAg nanocomposite in ways that may help minimize its possible adverse outcomes to human health.

Hyperbaric oxygen affects endothelial progenitor cells proliferation in vitro.

Hyperbaric oxygen is a clinical treatment that contributes to wound healing by increasing fibroblasts proliferation, collagen synthesis, and production of growth factors, inducing angiogenesis and inhibiting antimicrobial activity. It also has been shown that hyperbaric oxygen treatment (HBO), through the activation of nitric oxide synthase promotes an increase in the nitric oxide levels that may improve endothelial progenitor cells (EPC) mobilization from bone marrow to the peripheral blood and stimulates the vessel healing process. However, cellular mechanisms involved in cell proliferation and activation of EPC after HBO treatment remain unknown. Therefore, the present work aimed to analyze the effect of HBO on the proliferation of pre-treated bone marrow-derived EPC with TNF-alpha. Also, we investigated the expression of ICAM and eNOS by immunochemistry, the production of reactive species of oxygen and performed an in vitro wound healing. Although 1h of HBO treatment did not alter the rate of in vitro wound closure or cell proliferation, it increased eNOS expression and decreased ICAM expression and reactive oxygen species production in cells pre-treated with TNF-alpha. These results indicate that HBO can decrease the inflammatory response in endothelial cells mediated by TNF-alpha, and thus, promote vascular recovery after injury.

A cell-culturing system for the study of interaction between macrophages infected with leishmania amazonensis

The cell culture insert system is a culturing system for the study of contact-independent cellular communication. Leishmaniasis is a neglect tropical disease with no vaccines and the available drugs present toxic side effects. Studies on Leishmania interaction with host macrophages aim to develop strategies for parasite control and drug development. The purpose of this study was to evaluate the effects of interaction between non-infected and L. amazonensis-infected human macrophages, by using the cell culture system. The results showed that the infection index was reduced by 56.2% as compared to controls only when infected macrophages were inserted on both sides of the Transwell membranes. An improvement in macrophage viability was also observed in this cell culture. The levels of interleukin-1ß, an inflammatory cytokine, and nitric oxide, a microbicidal molecule, did not increase in L. amazonensis-infected macrophage cultures in the Transwell system; thus other soluble factors were responsible for parasite control

Expression on hypoxia-inducible factor-1α in human tegumentary leishmaniosis caused by Leishmania braziliensis

Several immune markers have been studied in controlling American tegumentary leishmaniosis based on mouse models. However, there is a lack of studies regarding human tegumentary leishmaniosis caused by Leishmania braziliensis. In this study, hypoxia-inducible factor-1α was found to be an important effector element in the localized control of human cutaneous and mucocutaneous lesions.