Extracellular Vesicles Secreted by Acanthamoeba culbertsoni Have COX and Proteolytic Activity and Induce Hemolysis.

Several species of Acanthamoeba genus are potential pathogens and etiological agents of several diseases. The pathogenic mechanisms carried out by these amoebae in different target tissues have been documented, evidencing the relevant role of contact-dependent mechanisms. With the purpose of describing the pathogenic processes carried out by these protozoans more precisely, we considered it important to determine the emission of extracellular vesicles (EVs) as part of the contact-independent pathogenicity mechanisms of A. culbertsoni, a highly pathogenic strain. Through transmission electronic microscopy (TEM) and nanoparticle tracking analysis (NTA), EVs were characterized. EVs showed lipid membrane and a size between 60 and 855 nm. The secretion of large vesicles was corroborated by confocal and TEM microscopy. The SDS-PAGE of EVs showed proteins of 45 to 200 kDa. Antigenic recognition was determined by Western Blot, and the internalization of EVs by trophozoites was observed through Dil-labeled EVs. In addition, some EVs biological characteristics were determined, such as proteolytic, hemolytic and COX activity. Furthermore, we highlighted the presence of leishmanolysin in trophozites and EVs. These results suggest that EVs are part of a contact-independent mechanism, which, together with contact-dependent ones, allow for a better understanding of the pathogenicity carried out by Acanthamoeba culbertsoni.

Chiranthodendron pentadactylon Larreat (Sterculiaceae), a Potential Nephroprotector against Oxidative Damage Provoked by STZ-Induced Hyperglycemia in Rats.

BACKGROUND: Chiranthodendron pentadactylon, known in Mexico as the "tree of the little hands", flower's infusion is used to treat kidney failure associated with diseases such as diabetes. The aim of this work is to evaluate the antioxidant effect of the methanolic extract of its flowers on oxidative damage in kidneys caused by streptozotocin in rats. METHODS: The extract phytochemical profile was performed with HPLC. Antioxidant potential in vitro was determined with DPPH and total phenolic tests; antioxidant evaluation in vivo was performed in diabetic rats administered daily via the intragastric route (100 and 200 mg/kg) for 6 weeks; serum glucose/creatinine, food/water consumption, and urinary volume were measured. Relative weight, protein/DNA ratios and oxidative stress were measured in renal tissue. RESULTS: The extract showed 20.53% of total phenolic content and IC50 of 18.05 µg/mL in DPPH, and this was associated with ferulic acid, phloretin and α-amyrin. Both doses showed a moderate decrease in the protein/DNA ratio in renal tissue, and the same behavior was observed for total urinary protein loss and serum creatinine, while the best antioxidant effect was exerted by a lower dose, which increased catalase activity and decreased lipid peroxidation in the kidneys. CONCLUSIONS: Results demonstrated that C. pentadactylon methanolic flower's extract improves renal function through antioxidant mechanisms during experimental diabetes.

Antihypertensive, antidyslipidemic, and renoprotective effects of Bursera simaruba on metabolic syndrome.

Objectives: Metabolic syndrome is associated with the development of chronic kidney disease. Bursera simaruba "chaca" is a medicinal plant used in Mexico for hypertension and empirical therapy. In this study, were examined the effects of ethanol extract of B. simaruba on metabolic syndrome. Materials and Methods: For induction of metabolic syndrome, 20% fructose was used, and it was administered in the water and food to male Wistar rats for 12 weeks, after administering ethanol extract of B. simaruba intragastrically (100 and 200 mg/kg/day) for 6 weeks, blood pressure was determined. In plasma, glucose, cholesterol, triglycerides, angiotensin II, oxide nitric, and angiotensin 1-7 were quantified. In the kidney was performed histological study and the activity of anti-oxidant enzymes was quantified. Results: Rats with metabolic syndrome developed obesity, arterial hypertension, dyslipidemia, and kidney damage characterized by proliferative glomerulonephritis, necrosis, and reduced activity of anti-oxidant enzymes. These alterations were significantly ameliorated by ethanol extract of B. simaruba. Conclusion: The ethanolic extract of B. simaruba showed antidyslipidemic, antihypertensive, anti-oxidant, and renoprotective effects.

Mecanismos involucrados en la enfermedad renal relacionada con la obesidad / Mechanisms involved in renal illness related to obesity

RESUMEN Introducción: En las últimas tres décadas la prevalencia de adultos con sobrepeso y obesidad ha aumentado sustancialmente en todo el mundo. La acumulación de grasa abdominal y visceral está altamente correlacionada con efectos adversos renales, a través de la acumulación de tejido adiposo dentro y alrededor de los riñones y la acumulación intrarrenal de matriz extracelular. Esta glomerulopatía relacionada con la obesidad se caracteriza en etapas tempranas por hipertrofia glomerular con o sin glomeruloesclerosis segmentaria focal secundaria. Objetivo: Revisar los mecanismos involucrados en la enfermedad renal relacionada con la obesidad. Métodos: Se realizó una búsqueda bibliográfica en PubMed y Google Académico de 2013 a 2017 sobre este tema. Conclusiones: Los efectos nocivos de la obesidad en la vasculatura renal se deben al aumento de la presión arterial, inflamación, hiperglucemia, lipotoxicidad causada por un exceso de metabolismo no-β-oxidativo de los ácidos grasos, estrés oxidativo y activación de múltiples sistemas neurohumorales. El exceso de grasa visceral es una fuente de citocinas y otros factores que crean un medio de estrés oxidativo e inflamación que contribuyen a la disfunción endotelial, la rigidez vascular y al desarrollo de la aterosclerosis. La hiperfiltración e hipertrofia glomerular causan el daño renal asociado a la obesidad. La hiperfiltración compensatoria, que preserva el filtrado glomerular, se acompaña de expansión del volumen glomerular, incremento en los componentes de la matriz glomerular, células endoteliales y mesangiales. Tras la inflamación ocurre la neovascularización, mecanismo compensatorio para mantener la perfusión de los tejidos lesionados o isquémicos(AU)

ABSTRACT Introduction: The prevalence of overweight and obese adults has increased substantially worldwide, in the last three decades. The accumulation of abdominal and visceral fat is highly correlated with renal adverse effects, through the accumulation of adipose tissue in and around the kidneys and the intrarenal accumulation of extracellular matrix. This obesity-related glomerulopathy is characterized in early stages by glomerular hypertrophy with or without secondary focal segmental glomerulosclerosis. Objective: To review the mechanisms involved in the obesity related renal illness. Methods: A bibliographical search on this topic was carried out in PubMed and Academic Google databases from 2013 to 2017. Conclusions: The harmful effects of obesity on the renal vasculature are due to increased blood pressure, inflammation, hyperglycemia, lipotoxicity caused by an excess of non-β-oxidative metabolism of fatty acids, oxidative stress and activation of multiple neurohumoral systems. Excess visceral fat is a source of cytokines and other factors that create a means of oxidative stress and inflammation contributing to endothelial dysfunction, vascular stiffness and the development of atherosclerosis. Hyperfiltration and glomerular hypertrophy cause kidney damage associated with obesity. Compensatory hyperfiltration, which preserves the glomerular filtration rate, is accompanied by glomerular volume expansion, an increase in the components of the glomerular matrix, endothelial and mesangial cells. Following inflammation, neovascularization occurs, a compensatory mechanism to maintain perfusion of injured or ischemic tissues(AU)

Mecanismos de inducción de la matriz extracelular en la nefropatía diabética / Mechanisms of induction of the extracellular matrix in diabetic nephropathy

La nefropatía diabética es una complicación grave en la diabetes mellitus. Sus principales cambios morfológicos típicos se deben al aumento de la cantidad de proteínas de la matriz extracelular. Los productos finales de glicación avanzada, resultado de la hiperglucemia, estimulan la producción de proteínas de la matriz extracelular en las células mesangiales, lo que resulta en la glomeruloesclerosis. Se revisan las alteraciones de las vías metabólicas que inducen la producción de factores que aumentan la síntesis de proteínas de la matriz extracelular y su acumulación durante el desarrollo de la nefropatía diabética. La glucosa intracelular elevada induce un aumento de angiotensina II y activación de proteína cinasa C, que a su vez, aumentan varios factores de crecimiento, como el transformante 1, el endotelial vascular, el de tejido conectivo, el epidérmico y el derivado de plaquetas, que llevan al incremento en la síntesis de proteínas de la matriz extracelular renal, como el colágeno, la fibronectina, la entactina y la laminina, lo que engrosará las membranas basales y expandirá progresivamente la matriz mesangial glomerular. Las metaloproteinasas de la matriz, que modulan la cantidad de proteínas de la matriz extracelular, son, a su vez, reguladas por los inhibidores tisulares de las metaloproteinasas(AU)

Diabetic nephropathy is a serious complication in diabetes mellitus. Its main and typical morphological changes are caused by the rise of the amount of the extracellulary matrix proteins. The final products of advanced glycation resulting from hyperglycemia stimulate the production of the extracellulary matrix proteins in the mesangial cells which leads to glomerulosclerosis. This article reviewed the alterations in the metabolic pathways that induce the production of factors capable of increasing the extracellulary matrix protein synthesis and their accumulation during the development of diabetic nephropathy. Elevated intracellular glucose leads to increased angiotensin II and C-kinase protein activation which, in turn, increase the number of several growth factors such as B1 transforming, vascular endothelial, connective tissue, epidermal and platelet-derived factors. All the above-mentioned causes more synthesis of renal extracellulary matrix proteins such as collagen, fibronectin, entactin and laminin which will thicken the basal membranes and will progresively extend the glomerular mesangial matrix. The matrix metalloproteins in charge of modulating the amount of proteins in the said matrix are then regulated by the tisular inhibitors of the metalloproteinases(AU)

Spasmolytic effect of Alternanthera repens on isolated rat ileum.

Abstract Context: Alternanthera repens (L.) Kuntze (Amaranthaceae) is widely used in Mexican traditional medicine for the treatment of gastrointestinal disorders that are mainly related to diarrhea. Objective: The aim of the present study was to investigate the spasmolytic effect of hexane (Hx), methanol (Me) and aqueous (Aq) extracts as well as chromatographic Me fractions (F1-F6) of A. repens in rat ileum. Materials and methods: Dried and powdered aerial parts were used to obtain the extracts. The rat ileum preparations were incubated in Tyrode's solution gassed (95% O2-5% CO2) at 37 °C. The effect on the contractile response of isolated ileum was evaluated by obtaining cumulative concentration-response curves to CaCl2, KCl, 5-HT and acetylcholine in the absence and presence of different doses of Aq (0.56-2.09 mg/mL), Me (0.24-0.91 mg/mL) and Hx (0.24-0.91 mg/mL) extracts, as well as six Me fractions of 0.66 mg/mL (F1 to F6). Results: The A. repens Me (0.24 mg/mL) caused an inhibitory response of the Ca2+-induced contractions, with IC50 values of 0.18 ± 0.061 and 0.67 ± 0.061 mM in the presence and absence of the Me, respectively. Me fractions F2 to F4 presented a significant inhibitory effect (F(3,8) = 60.17, p = 0.0001), causing a reduction in the CaCl2-induced contractions and shifting the Ca2+ (0.39 to 1.81 mM) concentration-response curves to the right. With respect to the effect on 5-HT-induced contractions, IC50 values Hx extract (0.24 mg/mL) were 5.44 ± 0.08 × 10-6 M and 3.38 ± 0.07 × 10-6 M in the presence and absence of the Hx, respectively. Discussion and conclusion: The spasmolytic effects induced by Me and Me fractions of A. repens may involve a serotonergic and Ca2+ influx blockade mechanisms, which may justify the use of A. repens extracts as an effective traditional treatment against diarrhea.

Inositol phosphoceramide synthase is a regulator of intracellular levels of diacylglycerol and ceramide during the G1 to S transition in Saccharomyces cerevisiae.

We recently reported that DAG (diacylglycerol) generated during sphingomyelin synthesis plays an important role in protein kinase C activation and cell proliferation in Madin-Darby canine kidney cells [Cerbon and Lopez-Sanchez (2003) Biochem. J. 373, 917-924]. In yeast cells, IPC (inositol phosphoceramide) synthase catalyses the transfer of phosphoinositol from phosphatidylinositol to ceramide to form IPC and generates DAG. In the present study, we found that, during the G1 to S transition after N2-starvation, there was a significant increase in the synthesis of IPC accompanied by a progressive increase (up to 6-fold) in the level of DAG. The increased DAG levels coincided with decrements in ceramide and sphingoid base levels, conditions that are adequate for the activation of putative protein kinase C required for the G1 to S transition and proliferation of yeast cells. To separate the role of DAG generated during IPC synthesis from that originating from other sources, we utilized beta-chloroalanine and myriocin, inhibitors of serine:palmitoyl-CoA transferase, the first committed step in sphingolipid synthesis, to avoid accumulation of sphingolipid intermediates. When the synthesis of sphingolipids was inhibited, DAG accumulation was significantly decreased and the G1 to S transition was blocked; such blockage was avoided by metabolic complementation with phytosphingosine. The DAG/ceramide ratio was 0.27 and it changed to 2.0 during growth re-initiation, suggesting that the synthesis of phosphosphingolipids could act to switch growth arrest (increased ceramide) to a mitogenic signal (increased DAG), and that this signalling process is preserved in yeast and mammalian cells.

In vitro antibradykinin activity of Aloe barbadensis gel.

When bradykinin-induced contraction of the isolated rat ileum was tested in the presence of Aloe barbadensis Mill. (Liliaceae) gel (fraction F-1) and with the fraction obtained by precipitation of the F-1 with 55% ammonium sulfate (F-55), the maximal responses to bradykinin were reduced by 10 and 22%, respectively. Furthermore, purification of the F-55 by filtration through a column of Sephacryl (S-500-HR) yielded the F-SH fraction, which inhibited the bradykinin effect by 60%. Purification of the F-SH fraction, by filtration through a column of Sephadex G-100, brought about four new fractions: F-GA, F-GB, F-GC, and F-GD. F-GB was the only one that showed the bradykinin inhibition effect (67%). Clearly, Aloe barbadensis gel contains a material that inhibits the bradykinin effect, which might explain the anti-inflammatory properties of Aloe barbadensis.