Identification of serologically active Helicobacter pylori antigens related to alterations in serum pepsinogen levels / Identificación de antígenos de Helicobacter pylori serológicamente activos relacionados con alteraciones en los niveles de pepsinógenos en suero

Resumen El adenocarcinoma gástrico se asocia con la infección por Helicobacter pylori. La transición a un proceso de carcinogénesis está precedida por atrofia glandular, y los niveles séricos de pepsinógeno I y II (PGI y PGII) se correlacionan con este tipo de lesiones gástricas. El objetivo del trabajo fue estudiar posibles asociaciones de los niveles de pepsinógenos (PG) en suero en relación con la frecuencia de actividad serológica hacia antígenos de H. pylori. Se utilizaron muestras de suero de pacientes con patología gástrica asociada a H. pylori (n = 26) y de individuos asintomáticos como controles (n = 37). Los antígenos seroactivos se identificaron mediante inmunoblot utilizando un extracto proteico de H. pylori. Los títulos de anticuerpos anti-H. pylori y la concentración de PG en suero se determinaron por ELISA. De los 31 antígenos seroactivos identificados, 9 presentaron una frecuencia diferencial entre ambos grupos (116,7; 68,8; 61,9; 54,9; 45,6; 38,3; 36,5; 33,8 y 30,1 kDa) y solo 3 se relacionaron con niveles alterados de PG en suero. En el grupo control, la seropositividad del antígeno de 33,8 kDa se relacionó con un aumento de PGII, mientras que el antígeno de 68,8kDa se relacionó con valores normales de PG (PGII disminuido y PGI/PGII elevado), sugiriendo que la seropositividad a este antígeno podría ser un factor protector frente a patologías gástricas. La seropositividad del antígeno de 54,9 kDa se relacionó con valores alterados de PG indicadores de inflamación y atrofia gástrica (aumento de PGII y disminución de PGI/PGII). La identificación de alteraciones séricas en los niveles de pepsinógeno relacionadas con la seropositividad a los antígenos de 33,8; 54,9 y 68,8 kDa de H. pylori sienta un precedente para futuros estudios como posibles biomarcadores serológicos pronósticos.

Identification of serologically active Helicobacter pylori antigens related to alterations in serum pepsinogen levels.

Gastric adenocarcinoma is associated with Helicobacter pylori infection. The transition to a carcinogenic process is preceded by glandular atrophy and serum levels of pepsinogen I and II (PGI and PGII) correlate with this type of gastric lesions. Possible associations of serum PG levels in relation to the frequency of serological activity against H. pylori antigens were studied. Serum samples from patients with gastric pathology associated with H. pylori (n=26) and asymptomatic individuals as controls (n=37) were used. Seroactive antigens were identified by immunoblot using a protein extract of H. pylori. The antibody titers anti-H. pylori and the concentration of PGs in serum was determined by ELISA. Thirty-one seroactive antigens were identified, nine of which exhibited a differential frequency between both groups (116.7, 68.8, 61.9, 54.9, 45.6, 38.3, 36.5, 33.8 and 30.1kDa) and only 3 were related to altered levels of PGs in serum. In the control group, the seropositivity of the 33.8kDa antigen was related to an increase in PGII, while the 68.8kDa antigen was related to normal PG values (decreased PGII and elevated PGI/PGII levels) indicating that seropositivity to this antigen could be a protective factor to gastric pathology. The seropositivity of the 54.9kDa antigen was related to altered values of PGs indicative of inflammation and gastric atrophy (increased in PGII and decreased in PGI/PGII). The identification of serum alterations in pepsinogen levels related to seropositivity to H. pylori 33.8, 54.9 and 68.8kDa antigens sets a precedent for further study as possible prognostic serological biomarkers.

Enzyme activity and expression of catalases in response to oxidative stress in Sporothrix schenckii.

Sporothrix schenckii is a dimorphic fungus, pathogenic to humans and animals, which is usually infective in the yeast form. Reactive oxygen species (ROS) play an important role in the host's defense, damaging the pathogen's DNA, proteins, and lipids. To prevent oxidative damage, the ROS are detoxified by pathogen-derived antioxidant enzymes such as catalases (CATs). In this work, we analyzed the activity and expression level of three S. schenckii genes, designated as CAT1, CAT2, and CAT3, that putatively encoded for three isoforms of monofunctional CAT with a predicted molecular weight of 57.6, 56.2, and 81.4 kDa, respectively. Our results demonstrate that oxidative stress induced by exogenous H2O2 leads to an altered lipid peroxidation, modifying CAT activity and the expression levels of the CAT genes, being CAT1 and CAT3 the genes with the highest expression in response to the oxidizing agent. These results show that CAT isoforms in S. schenckii can be regulated in response to oxidative stress and might help to control ROS homeostasis in the fungus-host interaction.

The Role of Macrophages in the Host's Defense against Sporothrix schenckii.

The role of immune cells associated with sporotrichosis caused by Sporothrix schenckii is not yet fully clarified. Macrophages through pattern recognition receptors (PRRs) can recognize pathogen-associated molecular patterns (PAMPs) of Sporothrix, engulf it, activate respiratory burst, and secrete pro-inflammatory or anti-inflammatory biological mediators to control infection. It is important to consider that the characteristics associated with S. schenckii and/or the host may influence macrophage polarization (M1/M2), cell recruitment, and the type of immune response (1, 2, and 17). Currently, with the use of new monocyte-macrophage cell lines, it is possible to evaluate different host-pathogen interaction processes, which allows for the proposal of new mechanisms in human sporotrichosis. Therefore, in order to contribute to the understanding of these host-pathogen interactions, the aim of this review is to summarize and discuss the immune responses induced by macrophage-S. schenckii interactions, as well as the PRRs and PAMPs involved during the recognition of S. schenckii that favor the immune evasion by the fungus.

Development of Nano-Antifungal Therapy for Systemic and Endemic Mycoses.

Fungal mycoses have become an important health and environmental concern due to the numerous deleterious side effects on the well-being of plants and humans. Antifungal therapy is limited, expensive, and unspecific (causes toxic effects), thus, more efficient alternatives need to be developed. In this work, Copper (I) Iodide (CuI) nanomaterials (NMs) were synthesized and fully characterized, aiming to develop efficient antifungal agents. The bioactivity of CuI NMs was evaluated using Sporothrix schenckii and Candida albicans as model organisms. CuI NMs were prepared as powders and as colloidal suspensions by a two-step reaction: first, the CuI2 controlled precipitation, followed by hydrazine reduction. Biopolymers (Arabic gum and chitosan) were used as surfactants to control the size of the CuI materials and to enhance its antifungal activity. The materials (powders and colloids) were characterized by SEM-EDX and AFM. The materials exhibit a hierarchical 3D shell morphology composed of ordered nanostructures. Excellent antifungal activity is shown by the NMs against pathogenic fungal strains, due to the simultaneous and multiple mechanisms of the composites to combat fungi. The minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) of CuI-AG and CuI-Chitosan are below 50 µg/mL (with 5 h of exposition). Optical and Atomic Force Microscopy (AFM) analyses demonstrate the capability of the materials to disrupt biofilm formation. AFM also demonstrates the ability of the materials to adhere and penetrate fungal cells, followed by their lysis and death. Following the concept of safe by design, the biocompatibility of the materials was tested. The hemolytic activity of the materials was evaluated using red blood cells. Our results indicate that the materials show an excellent antifungal activity at lower doses of hemolytic disruption.

Development of a sustainable photocatalytic process for air purification.

Nowadays, air pollution has become a global menace being responsible of a significant increase on the morbidity and mortality of human beings. In view of this, sustainable and efficient technologies for air purification are being sought. Air purification by photocatalytic treatment has received a lot of attention due to the unspecific and high oxidation capacity of the catalyst; however still some variables must be optimized to assure practical applications. In this work, visible light active TiO2-Cu2+@perlite and Ag@TiO2-Cu2+/perlite supported materials were fabricated. TiO2-Cu2+ (2 at. %) were synthesized using a sol-gel procedure followed of the impregnation of the support by immersion. For Ag@TiO2-Cu2+, silver deposition was conducted by chemical reduction using sodium citrate and sodium borohydride. The materials (powders and supported materials) were characterized by Scanning Electron Microscopy (SEM) to demonstrate their small size and adherence to the substrate. A prototype of a photocatalytic air purifier was built. The efficacy of the prototype was evaluated for the disinfection of indoor air (dentistry clinics). The photo-catalyst was activated using visible and UVA low-cost high-energy LEDs. The antibacterial activity of the air filter was evaluated. Ag@TiO2-Cu2+ exerts better air disinfection activity at lower doses in comparison to TiO2-Cu2+. Bacterial growth inhibitions up to 99% were achieved for both, Gram-negative and Gram-positive bacteria. The incorporation of Ag and Cu to TiO2 improves the antibacterial activity of the materials due to enhanced photocatalytic activity and the synergic activity of TiO2 and dopant elements (Ag, Cu) to inhibit microorganism's growth.

Morphological changes and phagocytic activity during the interaction of human neutrophils with Sporothrix schenckii: An in vitro model.

Histopathological studies of human sporotrichosis lesions show pyogenic and granulomatous processes in which polymorphonuclear neutrophils (PMNs) play a central role. Few studies regarding the events associated with the interaction of human PMNs with Sporothrix schenckii have been made despite their importance in the clinical manifestations of the disease. In this study, human PMNs were co-cultured with conidia or yeast cells of S. schenckii to compare the phagocytic activity and morphological changes that could provide a clearer insight into the role of these phagocytes in the initial phase of sporotrichosis. PMNs showed increased cell size and separation of the nuclear lobes after phagocytosis. Through Scanning Electron Microscopy (SEM) analysis, an increase in cells with flattened filaments and vesicles on their surface was observed. Phagocytosed conidia showed a significant increase in width and size. The phagocytic activity was greater against yeasts than with conidia, but the viability of both S. schenckii cellular morphotypes was not drastically affected even after 2 h of co-culture. In conclusion, morphological changes in PMNs suggest that S. schenckii induces processes that may favor proinflammatory events. These phagocytes show a high ability to bind or ingest S. schenckii cells without affecting their viability. Morphological changes recorded in ingested conidia, suggest that this fungus could make the dimorphic switching in PMNs.

Immune Response Induced by an Immunodominant 60 kDa Glycoprotein of the Cell Wall of Sporothrix schenckii in Two Mice Strains with Experimental Sporotrichosis.

Cell wall (CW) components of fungus Sporothrix schenckii are the major inductors antigens of immune responses. The immunodominant 60 kDa glycoprotein (gp60) has been shown to be associated with the virulence of this fungus but its role in experimental sporotrichosis is unknown. In this work, the immunological effects of CW-purified gp60 were investigated in a model of experimental subcutaneous sporotrichosis in normal and gp60-preimmunized C57BL/6 and BALB/c mice strains which were then infected with S. schenckii conidia. Results showed that both mice strains use different cytokine profiles in order to fight S. schenckii infection; C57BL/6 mice seem to use a Th17 response while BALB/c mice tend to depend on a Th1 profile. Preimmunization with gp60 showed a downregulatory effect on the immune response since cytokines levels were diminished in both strains. There were no significant differences in the magnitude of dorsoplantar inflammation between gp60-preimmunized and nonimmunized mice of both strains. However, skin lesions due to the infection in gp60-preimmunized mice were more severe in BALB/c than in C57BL/6 mice, suggesting that the antigen exerts a higher downregulatory effect on the Th1 response.

Molecular Components of the Sporothrix schenckii Complex that Induce Immune Response.

Sporotrichosis is a fungal disease caused by the Sporothrix schenckii complex that includes species such as S. brasiliensis, S. schenckii sensu stricto, S. globosa, S. luriei, S. mexicana, and S. pallida, which exhibit different potentially antigenic molecular components. The immune response of susceptible hosts to control infection and disease caused by these fungi has been little studied. Besides, the fungus-host interaction induces the activation of different types of immune response. This mini-review analyzes and discusses existing reports on the identification and functional characterization of molecules from species of the S. schenckii complex with clinical relevance, and the mechanisms that mediate the type and magnitude of the immune response in experimental models in vivo and in vitro. This knowledge is expected to contribute to the development of protective and therapeutic strategies against sporotrichosis and other mycoses.

Sporothrix schenckii yeasts induce ERK pathway activation and secretion of IL-6 and TNF-α in rat mast cells, but no degranulation.

Sporothrix schenckii is a dimorphic fungus that causes sporotrichosis, a subcutaneous mycosis found throughout the world in humans and other mammals. After contact with conidia, transition to the yeast stage is required for establishment of infection. Mast cells are one of the first components of the immune system to make contact with invading pathogens. They release potent mediators that are decisive in initiating and directing the course of immune and inflammatory responses in the host. It remains unknown whether or not yeast cells of S. schenckii activate mast cells. Our aim in this study was to evaluate the in vitro response of mast cells to S. schenckii yeasts cells. Mast cells became activated after interaction with the yeasts, although exocytosis of preformed mediators was not stimulated. Sporothrix schenckii yeasts induced the release of early response cytokines such as tumor necrosis factor alpha (TNF-α) and interleukin (IL)-6 and activation of the extracellular signal-regulated kinase (ERK) signaling pathway in mast cells. As TNF-α and IL-6 are considered crucial mediators in the defense of the host against fungal disease, the release of both mediators from mast cells may contribute to the overall response of the host immune system during S. schenckii infection.