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Stanhopea tigrina Bateman ex Lindl. (Orchidaceae) is an orchid endemic to Mexico, known as "Calavera" or "calaverita", in the Huasteca Potosina (central region of Mexico). This plant species is used for the folk treatment of mental disorders and urological kidney disorders, according to the ethnomedicinal information obtained in this study. Ethanolic extracts of leaves (HE) and pseudobulb (PE) were obtained by microwave-assisted extraction (MAE). Gas Chromatography coupled with Mass Spectrometry (GC-MS) was used to carry out the chemical characterization of HE and PE. The pharmacological effects (antioxidant, diuretic, anxiolytic, locomotor, hypnotic, and sedative) of HE and PE were evaluated. The possible mechanism of action of the anxiolytic-like activity induced by HE was assessed using inhibitors of the GABAergic, adrenergic, and serotonergic systems. The possible mechanism of the diuretic action of HE was assessed using prostaglandin inhibitory antagonists and nitric oxide synthase (NOS) blockers. HE at 50 and 100 mg/kg exerted anxiolytic-like activity without inducing hypnosis or sedation. Flumazenil, prazosin, and ketanserin inhibited the anxiolytic-like activity shown by HE, which suggests the participation of GABA, α1-adrenergic receptors, and 5-HT2 receptors, respectively. The diuretic effect was reversed by the non-selective NOS inhibitor L-NAME, which caused the reduction in nitric oxide (NO). These results demonstrate that the ethanolic extract of S. tigrina leaves exhibited anxiolytic-like activity and diuretic effects without inducing hypnosis or sedation. This work validates the medicinal uses of this orchid species.

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Candida albicans is an opportunistic fungal pathogen that may cause invasive infections in immunocompromised patients, disseminating through the bloodstream to other organs. In the heart, the initial step prior to invasion is the adhesion of the fungus to endothelial cells. Being the fungal cell wall's outermost structure and the first to come in contact with host cells, it greatly modulates the interplay that later will derive in the colonization of the host tissue. In this work, we studied the functional contribution of N-linked and O-linked mannans of the cell wall of C. albicans to the interaction with the coronary endothelium. An isolated rat heart model was used to assess cardiac parameters related to vascular and inotropic effects in response to phenylephrine (Phe), acetylcholine (aCh) and angiotensin II (Ang II) when treatments consisting of: (1) live and heat-killed (HK) C. albicans wild-type yeasts; (2) live C. albicans pmr1Δ yeasts (displaying shorter N-linked and O-linked mannans); (3) live C. albicans without N-linked and O-linked mannans; and (4) isolated N-linked and O-linked mannans were administered to the heart. Our results showed that C. albicans WT alters heart coronary perfusion pressure (vascular effect) and left ventricular pressure (inotropic effect) parameters in response to Phe and Ang II but not aCh, and these effects can be reversed by mannose. Similar results were observed when isolated cell walls, live C. albicans without N-linked mannans or isolated O-linked mannans were perfused into the heart. In contrast, C. albicans HK, C. albicans pmr1Δ, C. albicans without O-linked mannans or isolated N-linked mannans were not able to alter the CPP and LVP in response to the same agonists. Taken together, our data suggest that C. albicans interaction occurs with specific receptors on coronary endothelium and that O-linked mannan contributes to a greater extent to this interaction. Further studies are necessary to elucidate why specific receptors preferentially interact with this fungal cell wall structure.

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Giardiasis is one of the most common gastrointestinal infections worldwide, mainly in developing countries. The etiological agent is the Giardia lamblia parasite. Giardiasis mainly affects children and immunocompromised people, causing symptoms such as diarrhea, dehydration, abdominal cramps, nausea, and malnutrition. In order to develop an effective vaccine against giardiasis, it is necessary to understand the host-Giardia interactions, the immunological mechanisms involved in protection against infection, and to characterize the parasite antigens that activate the host immune system. In this study, we identify and characterize potential T-cell and B-cell epitopes of Giardia immunogenic proteins by immunoinformatic approaches, and we discuss the potential role of those epitopes to stimulate the host´s immune system. We selected the main immunogenic and protective proteins of Giardia experimentally investigated. We predicted T-cell and B-cell epitopes using immunoinformatic tools (NetMHCII and BCPREDS). Variable surface proteins (VSPs), structural (giardins), metabolic, and cyst wall proteins were identified as the more relevant immunogens of G. lamblia. We described the protein sequences with the highest affinity to bind MHC class II molecules from mouse (I-Ak and I-Ad) and human (DRB1*03:01 and DRB1*13:01) alleles, as well as we selected promiscuous epitopes, which bind to the most common range of MHC class II molecules in human population. In addition, we identified the presence of conserved epitopes within the main protein families (giardins, VSP, CWP) of Giardia. To our knowledge, this is the first in silico study that analyze immunogenic proteins of G. lamblia by combining bioinformatics strategies to identify potential T-cell and B-cell epitopes, which can be potential candidates in the development of peptide-based vaccines. The bioinformatics analysis demonstrated in this study provides a deeper understanding of the Giardia immunogens that bind to critical molecules of the host immune system, such as MHC class II and antibodies, as well as strategies to rational design of peptide-based vaccine against giardiasis.

Assuntos

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Bioremediation technology is one of the most profitable and sustainable strategies for remediating soils contaminated with hydrocarbons. This study focuses on assessing the influence of biostimulation and bioaugmentation with Pseudomonas fluorescens to contribute to the removal of total petroleum hydrocarbons (TPHs) of a soil. Laboratory studies were carried out (measurements of emitted CO2, surface tension, and residual TPH) to select the best bioaugmentation and biostimulation treatment. The sources of C, N, and P were glucose-yeast extract, NH4Cl-NaNO3, and K2HPO4-K3PO4, respectively. The effect of culture conditions on the reduction of TPH and respiratory activity was evaluated through a factorial design, 23, in a solid culture system. After 80 days of incubation, it was observed that treatments of yeast extract-NH4Cl-K2HPO4 (Y4) and glucose-NaNO3-K3PO4 (Y5) presented a higher level of TPH removal (20.91% and 20.00% degradation of TPH, respectively). Biostimulation favors the production of biosurfactants, indirectly measured by the change in surface tension in the soil extracts. The treatments Y4 and Y5 showed a lower change value of the surface tension (23.15 and 23.30 mN·m-1 at 25 °C). A positive correlation was determined between the change in surface tension and the removal of TPH; hence there was a contribution of the biosurfactants produced to the removal of hydrocarbons.

Assuntos

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We have shown that angiotensin II (Ang II) does not diffuse across the vessel wall, remaining intravascularly confined and acting solely on the coronary endothelial luminal membrane (CELM) receptors. A sustained intracoronary infusion of Ang II causes transient coronary vasoconstriction (desensitization) due to membrane internalization of CELM Ang II type 1 receptors (CELM-AT1R). In contrast, sustained intracoronary infusion of a non-diffusible polymer of Ang II (Ang II-Pol, 15,000 kDa) causes a sustained vasoconstriction by preventing CELM-AT1R internalization. In addition, a sustained intracoronary infusion of Ang II leads to a depressed response following a secondary Ang II administration (tachyphylaxis) that is reversed by Ang II-Pol. These findings led us to hypothesize that the rate of desensitization, tachyphylaxis, and AT1R internalization were dependent on Ang II-Pol molecular weight. To test this hypothesis, we synthesized Ang II-Pols of the following molecular weights (in kDa): 1.3, 2.7, 11, 47, 527, 3270 and 15,000. Vasoconstriction was measured following intracoronary infusion of Ang II-Pols in Langendorff-perfused guinea pig hearts at constant flow. The CELM protein fraction was extracted using the silica pellicle technique at different time points in order to determine the rate of AT1R internalization following each Ang II-Pol infusion. CELM-AT1R density was quantified by Western blot. We found that the rate of desensitization and the tachyphylaxis effect varied inversely with the molecular weight of the Ang II-Pols. Inversely proportional to the molecular weight of Ang II-Pol the CELM-AT1R density decreases over time. These results indicate that the mechanism responsible for the decreased rate of desensitization and tachyphylaxis by higher molecular weight Ang II polymers is due to reduction in the rate of CELM-AT1R internalization. These Ang II polymers would be valuable tools for studying the relationship between AT1R internalization and physiological effects.

Assuntos

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Diverse intracoronary hormones cause their cardiac effects solely via activation of their coronary endothelial luminal membrane (CELM) receptors. To test this hypothesis for Ang II, we synthesized: a) two large polymers of Ang II (Ang II-POL) and Losartan (Los-POL) which act only in the CELM's AT1R because they cannot cross the endothelial barrier and b) biotin-labeled Ang II (Ang II-Biotin) and Ang II-POL-Biotin to be identified by microscopy in tissues. Sustained coronary perfusion of Ang II (potentially diffusible) or Ang II-POL caused a positive inotropic effect (PIE) and an increase in coronary perfusion pressure (CPP) of equal magnitude that were blocked by Losartan and Los-POL. However, Ang II effects, in contrast to Ang II-POL effects, were transient due to desensitization and resulted in tachyphylaxis to a second administration of Ang II or Ang II-POL. Furthermore, if Ang II and Ang II-POL acted differently on the same receptor; a competition of effects would be expected. This was demonstrated by infusing simultaneously a molar ratio of Ang II:Ang II-POL. As this molar ratio decreased, Ang II-induced desensitization and tachyphylaxis decreased. Intravascularly-administered Ang II-Biotin and Ang II-POL-Biotin remained bound and confined to the endothelium. Our results support the hypothesis and indicate intravascular Ang II, not by mass exchange with the interstitium, but by an action restricted to the CELM's AT1R, causes release of endothelial chemical messengers that exert physiological effects and modulate the effects and metabolism of paracrine Ang II. Endocrine Ang II controls and communicates with its paracrine counterparts solely through endothelial receptors.

Assuntos

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Coronary blood flow applied to the endothelial lumen modulates parenchymal functions via paracrine effectors, but the mechanism of flow sensation is unknown. We and others have demonstrated that coronary endothelial luminal membrane (CELM) oligosaccharides and lectins are involved in flow detection, and we proposed that cardiac effects of coronary flow result from a reversible flow-modulated lectin-oligosaccharide interaction. Recently, glycosylated and amiloride-sensitive Na(+)/Ca(++) channels (ENaCs) have been proposed to be involved in the flow-induced endothelial responses. Because N-acetylglucosamine (GlcNac) is one of the main components of glycocalyx oligosaccharides (i.e., hyaluronan [-4GlcUAbeta1-3GlcNAcbeta1-](n)), the aim of this article is to isolate and define CELM GlcNac-binding lectins and determine their role in cardiac and vascular flow-induced effects. For this purpose, we synthesized a 460-kDa GlcNac polymer (GlcNac-Pol) with high affinity toward GlcNac-recognizing lectins. In the heart, intracoronary administration of GlcNac-Pol upon binding to CELM diminishes the flow-dependent positive inotropic and dromotropic effects. Furthermore, GlcNac-Pol was used as an affinity probe to isolate CELM GlcNac-Pol-recognizing lectins and at least 35 individual lectinic peptides were identified, one of them the beta-ENaC channel. Some of these lectins could participate in flow sensing and in GlcNac-Pol-induced effects. We also adopted a flow-responsive and well-accepted model of endothelial-parenchymal paracrine interaction: isolated blood vessels perfused at controlled flow rates. We established that flow-induced vasodilatation (FIV) is blocked by endothelial luminal membrane (ELM) bound GlcNac-Pol, nitro-l-arginine methyl ester and indomethacin, amiloride, and hyaluronidase. The effect of hyaluronidase was reversed by infusion of soluble hyaluronan. These results indicate that GlcNac-Pol inhibits FIV by competing and displacing intrinsic hyaluronan bound to a lectinic structure such as the amiloride-sensitive ENaC. Nitric oxide and prostaglandins are the putative paracrine mediators of FIV.

Assuntos

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Diverse intracoronary agonists cause cardiac effects while acting on coronary endothelial luminal membrane (CELM) receptor. Our data show: a) the presence of AT(1)R in isolated CELM and in all cardiac cell types and b) sustained intracoronary infusions of Ang II-POL, a large sized molecule (approximately 15,000 kDa) confined to the vessel lumen that can only act on CELM's AT(1)R or Ang II (approximately 1 kDa); both exert the same maximum positive inotropic (PIE) and coronary constriction (CPP). The effects of these two agonists are blocked by Losartan and by Sar-POL; a large size antagonist (approximately 15,000 kDa) that acts only on CELM. Ang II effects are transient due to desensitization and cause tachyphylaxis to Ang II and toward Ang II-POL suggesting that both Ang II and Ang II-POL act on the same receptor group. In contrast, Ang II-POL effects are sustained and do not cause tachyphylaxis. The results show that intravascular Ang II and Ang II-POL act differentially by an unknown mechanism on CELM's AT(1)R and suggest that intravascular Ang II and Ang II-POL cause PIE and CCP by activation limited to CELM's AT(1)R through an unknown mechanism that is space-confined to the CELM's AT(1)R.

Assuntos