Exploring porcine kidney explants as a model for the study of nephrotoxins and the therapeutic potential of phytic acid.

The use of in vivo models to assess nephrotoxicity has faced ethical limitations. A viable alternative is the ex vivo model that combines the 3 R principles with the preservation of tissue histology. Here, we established a gentamicin nephrotoxicity model using pigs` kidney explants and investigated the effect of phytic acid (IP6) against gentamicin- induced nephrotoxicity. A total of 360 kidney explants were divided into control, gentamicin (10 mM), IP6 (5 mM), and gentamicin+IP6 groups. The activity of gammaglutamyltransferase (GGT), creatinine levels, histological assessment, oxidative stress, and inflammatory cytokine expression were analyzed. Exposure to gentamicin induced an increase in GGT activity, creatinine levels, lesion score, lipoperoxidation and IL-8 expression. Explants exposed to IP6 remained like the control. The addition of IP6 to gentamicin prevented tissue damage, increasing the antioxidant status and gene expression of IL-10. This model proved to be an adequate experimental approach for identifying nephrotoxins and potential products to modulate the toxicity.

Histopathological and ultrastructural findings induced by heat-inactivated Lactobacillus plantarum and the culture supernatant on the intestinal mucosa of piglets: an ex vivo approach / Achados histopatológicos e ultraestruturais induzidos por Lactobacillus plantarum inativado pelo calor na mucosa intestinal de leitões: estudo ex-vivo

In the present study, histological, morphometrical and ultrastructural analysis were performed to investigate intestinal mucosa changes in piglets jejunal explants exposed to two concentration of heat-inactivated Lactobacillus plantarum and their respective culture supernatants. Jejunal explants were incubated for 4 hours in DMEM culture medium with a) only culture medium (control group), b) heat-inactivated Lactobacillus plantarum strain1 - LP1 (1.1 x 108CFU/ml), c) heat-inactivated Lactobacillus plantarum strain2 - LP2 (2.0 x 109CFU/ml), d) heat-inactivated Lactobacillus plantarum strain1 culture supernatant (CS1), and e) heat-inactivated Lactobacillus plantarum strain2 culture supernatant (CS2). Explants exposed to heat-inactivated L. plantarum strain 1 and 2 showed multifocal to difuse villi atrophy, villi apical necrosis and enterocyte flattening. Morphological assessment revealed similar results with bacterial adhesion to mucus and intestinal epithelial cells and, morphometric analysis showed a decreased villi height compared to the control group. Alterations in explants treated with the culture supernatant of both strains include mild villi atrophy and mild enterocyte apical necrosis. Morphological assesment reveled numerous well delineated villi and, morphometric analysis showed a significant increase in villi height compared to the control group. In general, exposure to the culture supernatants improved the intestinal morphology.(AU)

No presente estudo, foram realizadas análises histológica, morfométrica e ultraestrutural para investigar as alterações da mucosa intestinal em explantes jejunais de leitões expostos a duas cepas e concentrações de Lactobacillus plantarum inativado pelo calor e seus sobrenadantes de cultura. Os explantes jejunais foram incubados durante quatro horas, em meio de cultura DMEM com: a) meio de cultura (grupo controle); b) Lactobacillus plantarum, cepa 1 - LP1 (1,1 x 108CFU/mL); c) Lactobacillus plantarum, LP2 (2,0 x 109CFU/mL); d) sobrenadante da cultura do Lactobacillus plantarum, cepa 1 (SC1); e e) sobrenadante da cultura do Lactobacillus plantarum, cepa 2 (SC2). Os explantes expostos às cepas 1 e 2 do L. plantarum inativado pelo calor mostraram atrofia difusa de vilosidades, necrose apical das vilosidades e achatamento de enterócitos. A avaliação morfológica revelou resultados semelhantes, com adesão bacteriana ao muco e às células epiteliais intestinais, e a análise morfométrica mostrou uma diminuição da altura das vilosidades em relação ao grupo controle. Alterações nos explantes tratados com o sobrenadante da cultura de ambas as cepas caracterizaram-se por atrofia leve das vilosidades e necrose apical leve dos enterócitos. A avaliação morfológica revelou vilosidades bem delineadas, e a análise morfométrica mostrou um aumento significativo na altura das vilosidades em comparação ao grupo controle. Em geral, a exposição aos sobrenadantes da cultura melhora a morfologia intestinal.(AU)

Purely-elastic flow instabilities and elastic turbulence in microfluidic cross-slot devices.

We experimentally investigate the dynamics of viscoelastic fluid flows in cross-slot microgeometries under creeping flow conditions. We focus on the unsteady flow regime observed at high Weissenberg numbers (Wi) with the purpose of understanding the underlying flow signature of elastic turbulence. The effects of the device aspect ratio and fluid rheology on the unsteady flow state are investigated. Visualization of the flow patterns and time-resolved micro-particle image velocimetry were carried out to study the fluid flow behavior for a wide range of Weissenberg numbers. A periodic flow behavior is observed at low Weissenberg numbers followed by a more complex dynamics as Wi increases, eventually leading to the onset of elastic turbulence for very high Weissenberg numbers.

Lactobacillus plantarum culture supernatants improve intestinal tissue exposed to deoxynivalenol.

In the present study, histological, morphometrical and ultrastructural analysis were performed to investigate intestinal mucosa changes in piglets exposed to deoxynivalenol alone or associated with two strains of Lactobacillus plantarum and the respective culture supernatants. Jejunal explants were incubated for 4h in culture medium with a) only culture medium (DMEM, control group), b) deoxynivalenol (DON, 10µM), c) heat-inactivated Lactobacillus plantarum strain1 - LP1 (1.1×108 CFU/ml) plus DON, d) heat-inactivated Lactobacillus plantarum strain2-LP2 (2.0×109 CFU/ml) plus DON, e) heat-inactivated Lactobacillus plantarum strain1 culture supernatant (CS1) plus DON, and f) heat-inactivated Lactobacillus plantarum strain1 culture supernatant (CS1) plus DON. Explants exposed to DON and DON plus LP1 and LP2 showed a significant increase in histological changes (mainly villi atrophy and apical necrosis) and a significant decrease in villi height when compared to unexposed explants. However, explants treated with CS1+DON and CS2+DON remained similar to the control group both in histological and morphometrical aspects. DON also induced a significant decrease in goblet cell density compared to control whereas CS1+DON treatment induced an increase in the number of goblet cells in comparison to DON explants. In addition, ultrastructural assessment showed control, CS1+DON and CS2+DON explants with well delineated finger shape villi, meanwhile DON-treated, LP1+DON and LP2+DON explants showed a severe villi atrophy with leukocytes exudation on the intestinal surface. Taken together, our results indicate that the culture supernatant treatment reduced the toxic effects induced by DON on intestinal tissue and may contribute as an alternative strategy to reduce mycotoxin toxicity.

Purely elastic flow instabilities in microscale cross-slot devices.

We present an experimental investigation of viscoelastic fluid flow in a cross-slot microgeometry under low Reynolds number flow conditions. By using several viscoelastic fluids, we investigate the effects of the microchannel bounding walls and the polymer solution concentration on the flow patterns. We demonstrate that for concentrated polymer solutions, the flow undergoes a bifurcation above a critical Weissenberg number (Wi) at which the flow becomes asymmetric but remains steady. The appearance of this elastic instability depends on the channel aspect ratio, defined as the ratio between the depth and the width of the channels. At high aspect ratios, when bounding wall effects are reduced, two types of elastic instabilities were observed, one in which the flow becomes asymmetric and steady, followed by a second instability at higher Wi, in which the flow becomes time-dependent. When the aspect ratio decreases, the bounding walls have a stabilizing effect, preventing the occurrence of steady asymmetric flow and postponing the transition to unsteady flow to higher Wi. For less concentrated solutions, the first elastic instability to steady asymmetric flow is absent and only the time-dependent flow instability is observed.

Analytical and numerical study of the electro-osmotic annular flow of viscoelastic fluids.

In this work we present semi-analytical solutions for the electro-osmotic annular flow of viscoelastic fluids modeled by the Linear and Exponential PTT models. The viscoelastic fluid flows in the axial direction between two concentric cylinders under the combined influences of electrokinetic and pressure forcings. The analysis invokes the Debye-Hückel approximation and includes the limit case of pure electro-osmotic flow. The solution is valid for both no slip and slip velocity at the walls and the chosen slip boundary condition is the linear Navier slip velocity model. The combined effects of fluid rheology, electro-osmotic and pressure gradient forcings on the fluid velocity distribution are also discussed.

Analytical solution of two-fluid electro-osmotic flows of viscoelastic fluids.

This paper presents an analytical model that describes a two-fluid electro-osmotic flow of stratified fluids with Newtonian or viscoelastic rheological behavior. This is the principle of operation of an electro-osmotic two-fluid pump as proposed by Brask et al. [Tech. Proc. Nanotech., 1, 190-193, 2003], in which an electrically non-conducting fluid is transported by the interfacial dragging viscous force of a conducting fluid that is driven by electro-osmosis. The electric potential in the conducting fluid and the analytical steady flow solution of the two-fluid electro-osmotic stratified flow in a planar microchannel are presented by assuming a planar interface between the two immiscible fluids with Newtonian or viscoelastic rheological behavior. The effects of fluid rheology, shear viscosity ratio, holdup and interfacial zeta potential are analyzed to show the viability of this technique, where an enhancement of the flow rate is observed as the shear-thinning effects are increased.

Shear viscosity and nonlinear behavior of whole blood under large amplitude oscillatory shear.

We investigated experimentally the rheological behavior of whole human blood subjected to large amplitude oscillatory shear under strain control to assess its nonlinear viscoelastic response. In these rheological tests, the shear stress response presented higher harmonic contributions, revealing the nonlinear behavior of human blood that is associated with changes in its internal microstructure. For the rheological conditions investigated, intra-cycle strain-stiffening and intra-cycle shear-thinning behavior of the human blood samples were observed and quantified based on the Lissajous-Bowditch plots. The results demonstrated that the dissipative nature of whole blood is more intense than its elastic component. We also assessed the effect of adding EDTA anticoagulant on the shear viscosity of whole blood subjected to steady shear flow. We found that the use of anticoagulant in appropriate concentrations did not influence the shear viscosity and that blood samples without anticoagulant preserved their rheological characteristics approximately for up to 8 minutes before coagulation became significant.

Extensional flow of blood analog solutions in microfluidic devices.

In this study, we show the importance of extensional rheology, in addition to the shear rheology, in the choice of blood analog solutions intended to be used in vitro for mimicking the microcirculatory system. For this purpose, we compare the flow of a Newtonian fluid and two well-established viscoelastic blood analog polymer solutions through microfluidic channels containing both hyperbolic and abrupt contractions∕expansions. The hyperbolic shape was selected in order to impose a nearly constant strain rate at the centerline of the microchannels and achieve a quasihomogeneous and strong extensional flow often found in features of the human microcirculatory system such as stenoses. The two blood analog fluids used are aqueous solutions of a polyacrylamide (125 ppm w∕w) and of a xanthan gum (500 ppm w∕w), which were characterized rheologically in steady-shear flow using a rotational rheometer and in extension using a capillary breakup extensional rheometer (CaBER). Both blood analogs exhibit a shear-thinning behavior similar to that of whole human blood, but their relaxation times, obtained from CaBER experiments, are substantially different (by one order of magnitude). Visualizations of the flow patterns using streak photography, measurements of the velocity field using microparticle image velocimetry, and pressure-drop measurements were carried out experimentally for a wide range of flow rates. The experimental results were also compared with the numerical simulations of the flow of a Newtonian fluid and a generalized Newtonian fluid with shear-thinning behavior. Our results show that the flow patterns of the two blood analog solutions are considerably different, despite their similar shear rheology. Furthermore, we demonstrate that the elastic properties of the fluid have a major impact on the flow characteristics, with the polyacrylamide solution exhibiting a much stronger elastic character. As such, these properties must be taken into account in the choice or development of analog fluids that are adequate to replicate blood behavior at the microscale.

Cardiac and pulmonary alterations in symptomatic and asymptomatic dogs infected naturally with Leishmania (Leishmania) chagasi.

Fifteen symptomatic and seven asymptomatic dogs infected naturally with Leishmania chagasi were examined in order to identify the presence of parasites and changes in heart and lung. Histopathological, cytological, and immunohistochemical analyses were performed on samples of heart and lung tissues. An inflammatory reaction characterized by inflammatory mononuclear, perivascular and intermuscular infiltrates was observed in both symptomatic and asymptomatic animals on histopathological analysis of the heart. In the lung, there was thickening of the alveolar septa due to congestion, edema, inflammatory infiltrate, and fibroblast proliferation. A focal reaction was observed although a diffuse reaction was present in both groups. On cytological examination, heart and lung imprints revealed amastigotes in two symptomatic animals and heart imprints were found in 1 asymptomatic dog. Immunoperoxidase staining showed amastigotes in the lung and heart of only 1 of 6 symptomatic animals examined. Within the ethical principles and limits of this research, it can be inferred that the study of heart and lung alterations in canine visceral leishmaniasis is increasingly important for understanding the problem related to humans. Dogs with visceral leishmaniasis were a good experimental model, since infection was caused by the same agent and the animals developed clinical, pathological and immunological alterations similar to those observed in humans.

Cardiac and pulmonary alterations in symptomatic and asymptomatic dogs infectednaturally with Leishmania (Leishmania) chagasi

Fifteen symptomatic and seven asymptomatic dogs infected naturally with Leishmania chagasi were examined in order to identify the presence of parasites and changes in heart and lung. Histopathological, cytological, and immunohistochemical analyses were performed on samples of heart and lung tissues. An inflammatory reaction characterized by inflammatory mononuclear, perivascular and intermuscular infiltrates was observed in both symptomatic and asymptomatic animals on histopathological analysis of the heart. In the lung, there was thickening of the alveolar septa due to congestion, edema, inflammatory infiltrate, and fibroblast proliferation. A focal reaction was observed although a diffuse reaction was present in both groups. On cytological examination, heart and lung imprints revealed amastigotes in two symptomatic animals and heart imprints were found in 1 asymptomatic dog. Immunoperoxidase staining showed amastigotes in the lung and heart of only 1 of 6 symptomatic animals examined. Within the ethical principles and limits of this research, it can be inferred that the study of heart and lung alterations in canine visceral leishmaniasis is increasingly important for understanding the problem related to humans. Dogs with visceral leishmaniasis were a good experimental model, since infection was caused by the same agent and the animals developed clinical, pathological and immunological alterations similar to those observed in humans.

Steady viscoelastic fluid flow between parallel plates under electro-osmotic forces: Phan-Thien-Tanner model.

The electro-osmotic flow of a viscoelastic fluid between parallel plates is investigated analytically. The rheology of the fluid is described by the Phan-Thien-Tanner model. This model uses the Gordon-Schowalter convected derivative, which leads to a non-zero second normal stress difference in pure shear flow. A nonlinear Poisson-Boltzmann equation governing the electrical double-layer field and a body force generated by the applied electrical potential field are included in the analysis. Results are presented for the velocity and stress component profiles in the microchannel for different parametric values that characterize this flow. Equations for the critical shear rates and maximum electrical potential that can be applied to maintain a steady fully developed flow are derived and discussed.

Antinociceptive effects of the essential oil of Alpinia zerumbet on mice.

Alpinia zerumbet (Pers.) Burtt. et Smith is an aromatic plant that is distributed widely in the tropical and sub-tropical regions of the world. In Brazil, where A. zerumbet is called "colonia", it is used widely in folk medicine for the treatment of various diseases, including hypertension. In the present study, the antinociceptive effects of the orally administered essential oil of A. zerumbet (EOAz) were evaluated in male Swiss mice (20-25 g each). In the acetic acid-induced writhing test, EOAz (30, 100 and 300 mg/kg body wt.; n = 10, n = 13 and n = 15, respectively) was effective at all doses. In the hot-plate test, EOAz significantly increased the latency at doses of 100 and 300 mg/kg body wt., but not at 30 mg/kg body wt., at all observation times up to the 180th min (n = 10 for each dose). In the formalin test, EOAz significantly reduced paw licking time in the second phase of the test at 100 mg/kg body wt. (n = 10), but decreased it in both phases at 300 mg/kg body wt. (n = 10). At 30 mg/kg body wt., the effect of EOAz did not differ from control values in either phase of the formalin test (n = 10). Pretreatment with naloxone (5 mg/kgbodywt., i.p.) caused a significant reversal of the analgesic effect of 300 mg/kg body wt. EOAz (n = 8) that was complete for the first phase, but only partial for the second phase of the formalin test. The data show that orally administered OEAz promotes a dose-dependent antinociceptive effect, with a mechanism of action which probably involves the participation of opiate receptors.

Fatal cerebral hemorrhage and acute renal failure after young Bothrops jararacussu snake bite.

An unusual case of a patient developing severe coagulopathy disorder and a clinical picture of cerebral hemorrhage and acute renal failure after young Bothrops jararacussu snake bite is reported. The mechanisms of snake venom-induced injury are discussed and similar cases in literature are revised and compared. The use of bothropic-Crotalus antivenom in severe B. jararacussu envenomation is discussed.

Process-scale reversed-phase high-performance liquid chromatography purification of LL-E19020 alpha, a growth promoting antibiotic produced by Streptomyces lydicus ssp. tanzanius.

LL-E19020 alpha is a novel antibiotic produced by fermentation of the soil microorganism Streptomyces lydicus ssp. tanzanius. The compound is highly effective in inducing increases in weight gain and feed conversion efficiency in livestock. In order to obtain kilogram quantities of the material for field trials, pilot plant scale fermentations (up to 7500 l) were carried out. The antibiotic was recovered from the fermentation broth by solvent extraction. The resultant crude extract was subjected to reversed-phase (C18) chromatography on a process-scale high-performance liquid chromatography (HPLC) unit. The heart of the instrumentation is the Millipore Kiloprep chromatograph with the standard 12-l cartridge column. The laboratory housing the chromatograph has been specifically designed for this work. Tanks for mobile phase preparation are mounted on load cells for precise measurement of components. In this explosion-proof laboratory, all solvent handling areas are well ventilated and a separate breathing air system is provided for the operators. For the purification of the LL-E19020 antibiotics, the mobile phase consisted of a gradient of acetonitrile in 0.1 M ammonium acetate at pH 4.5. The effluent was monitored by UV absorbance at 325 nm. Fractions were collected across the peaks of interest and these were analyzed by analytical HPLC. The maximum yield of LL-E19020 alpha obtained in a single run was approximately 100 g. The antibiotic was recovered from the mobile phase by extraction with methylene chloride. The methylene chloride phase was concentrated under reduced pressure to yield a gummy residue which was finally freeze-dried from tertiary butanol to yield an off-white solid suitable for blending with various feed components.