Modulatory effect of inulin with soya isoflavones on plasma lipid profile and liver SCD-18 index in rats with induced type-2 diabetes mellitus.

Obesity and type-2 diabetes are often associated with nonalcoholic fatty liver disease (NAFLD). Soya isoflavones act as antidiabetic agents and protect against NAFLD. There are data suggesting that inulin may increase the plasma concentration and effect of soya isoflavones. The aim of the present study was to compare the effect of soya isoflavones, as opposed to the effect of soya isoflavones with inulin, on plasma lipid profile, liver morphology, and liver fatty acids in rats with induced type-2 diabetes mellitus. Data were collected on thirty-six male Sprague-Dawley rats divided into control and diabetic groups. Animals in the diabetic (DM) group were on a high-fat diet and were injected with low doses of streptozotocin. Animals in the control groups were fed a regular diet and were injected with a buffer. After the injections, the animals were divided into three groups of nondiabetic rats (nDM)-controls (c-nDM), rats treated with isoflavones (IS-nDM), and rats treated with isoflavones plus inulin (IS+IN-nDM)-and three parallel diabetic (DM) subgroups: controls (c-DM), rats treated with isoflavone (IS-DM), and rats treated with isoflavones plus inulin (IS+IN-DM). Hepatic steatosis and fibrosis were examined using hematoxylin-eosin staining and Mallory's trichrome methods respectively. Liver fatty acids were extracted and analyzed by gas chromatography. A lipid blood test was performed. The study showed significant changes in liver fatty acids, liver morphology, and plasma lipid profile. The estimated SCD-18 index significantly decreased in both the control and DM groups after isoflavone supplementation. The level of liver steatosis and fibrosis also decreased after isoflavone supplementation in the DM groups. The plasma lipid profile showed increased levels of HDL-C after isoflavone supplementation in the DM groups. These results support the protective use of isoflavones in liver steatosis and as beneficial to plasma lipid profile in individuals with diabetes. A novelty of this work is its comparison of supplementation using soya isoflavones with supplementation using both soya isoflavones and inulin. Surprisingly, additional supplementation with inulin modulates the positive effect of isoflavones.

Nanoporous silica nanoparticles functionalized with a fluorescent turn-on spirorhodamineamide as pH indicators.

We prepared water soluble, biocompatible fluorescent turn-on pH nanosensors and characterized their behavior as a function of changes in pH. The response relies on a halochromic reaction of a spirorhodamineamide derived from the bright and highly chemically and photo-stable rhodamine 6G, encapsulated in core/nanoporous shell silica nanoparticles. The fluorescent sensors displayed a fast response in the pH range of intracellular compartments. The encapsulation conferred solubility in aqueous environments and biocompatibility. We assessed the two main properties of the sensor, namely the useful pH range and the kinetics of the response, and compared them to those of the free probe. We found that such properties are strongly dependent on the functionalization and position in the silica matrix relative to the core/shell structure. Finally, we demonstrated the cellular uptake of the nanosensors, and their localization in lysosomes of living cells, by fluorescence confocal microscopy.

The inflammatory effect of infection with Hymenolepis diminuta via the increased expression and activity of COX-1 and COX-2 in the rat jejunum and colon.

The aim of this study was to determine whether Hymenolepis diminuta may affect the expression and activity of cyclooxygenase 1 (COX-1) and cyclooxygenase 2 (COX-2), resulting in the altered levels of their main products - prostaglandins (PGE2) and thromboxane B2 (TXB2). The study used the same experimental model as in our previous studies in which we had observed changes in the transepithelial ion transport, tight junctions and in the indicators of oxidative stress, in both small and large intestines of rats infected with H. diminuta. In this paper, we investigated not only the site of immediate presence of the tapeworm (jejunum), but also a distant site (colon). Inflammation related to H. diminuta infection is associated with the increased expression and activation of cyclooxygenase (COX), enzyme responsible for the synthesis of PGE2 and TXB2, local hormones contributing to the enhanced inflammatory reaction in the jejunum and colon in the infected rats. The increased COX expression and activity is probably caused by the increased levels of free radicals and the weakening of the host's antioxidant defense induced by the presence of the parasite. Our immunohistochemical analysis showed that H. diminuta infection affected not only the intensity of the immunodetection of COX but also the enzyme protein localization within intestinal epithelial cells - from the entire cytoplasm to apical/basal regions of cells, or even to the nucleus.

Changes in the Immune System of Female Wistar Rats After Exposure to Immunosuppressive Treatment During Pregnancy.

This experimental study assessed the impact of medications frequently used after kidney transplantation on the immune system of pregnant female Wistar rats. The study evaluates medications, both approved and contraindicated during pregnancy in common therapeutic combinations. The study was conducted on 32 female Wistar rats, subjected to immunosuppressive regimens most commonly used in therapy of human kidney transplant recipients (cyclosporine A, mycophenolate mofetil and prednisone; tacrolimus, mycophenolate mofetil and prednisone; and cyclosporine A, everolimus and prednisone). The animals received drugs by oral gavage 2 weeks before pregnancy and at 3 weeks of pregnancy. We found drug regimen-dependent differences in cytometry from spleen. Many subpopulations of lymphocytes were suppressed in rats treated with cyclosporine A, mycophenolate mofetil and prednisone and tacrolimus, mycophenolate mofetil and prednisone; the number of NK cells was increased in group of rats treated with cyclosporine A, everolimus and prednisone. We also found changes in histological examination of thymus and spleen of all treated dams. In cytokine assay, we noticed increasing levels of IL-17 with increasing doses of concanavalin A in control group and in group of dams treated with cyclosporine A, mycophenolate mofetil and prednisone. This increase was blocked in rats treated with tacrolimus, mycophenolate mofetil and prednisone and cyclosporine A, everolimus and prednisone. Qualitative, quantitative and morphological changes of immune system in pharmacologically immunosuppressed females have been observed. Thymus structure, spleen composition and splenocytes IL-17 production were mostly affected in drug regimen-dependent manner.

Hyaluronan degrading silica nanoparticles for skin cancer therapy.

We report the first nanoformulation of Hyaluronidase (Hyal) and its enhanced adjuvant effect over the free enzyme. Hyaluronic acid (HA) degrading enzyme Hyal was immobilized on 250 nm silica nanoparticles (SiNP) maintaining specific activity of the enzyme via the layer-by-layer self-assembly technique. This process was characterized by dynamic light scattering (DLS), zeta potential, infrared and UV-Vis spectroscopy, transmission electron microscopy (TEM) and enzymatic activity measurements. The nanoparticles were tested in vivo as adjuvants of carboplatin (CP), peritumorally injected in A375 human melanoma bearing mice and compared with the non-immobilized enzyme, on the basis of equal enzymatic activity. Alcian Blue staining of A375 tumors indicated large overexpression of hyaluronan. At the end of the experiment, tumor volume reduction with SiNP-immobilized Hyal was significantly enhanced compared to non-immobilized Hyal. Field emission scanning electron microscopy (FE-SEM) images together with energy dispersive X-ray spectroscopy (EDS) spectra confirmed the presence of SiNP on the tumor. We mean a proof of concept: this extracellular matrix (ECM) degrading enzyme, immobilized on SiNP, is a more effective local adjuvant of cancer drugs than the non-immobilized enzyme. This could prove useful in future therapies using other or a combination of ECM degrading enzymes.

Controlled deposition of silver nanoparticles in mesoporous single- or multilayer thin films: from tuned pore filling to selective spatial location of nanometric objects.

Silver nanoparticle assemblies are embedded within mesoporous oxide thin films by an in situ mild reduction leading to nanoparticle-mesoporous oxide thin-film composites (NP@MOTF). A quantitative method based on X-ray reflectivity is developed and validated with energy dispersive spectroscopy in order to assess pore filling. The use of dilute formaldehyde solutions leads to control over the formation of silver nanoparticles within mesoporous titania films. Inclusion of silver nanoparticles in mesoporous silica requires more drastic conditions. This difference in reactivity can be exploited to selectively synthesize nanoparticles in a predetermined layer of a multilayered mesoporous stack leading to complex 1D-ordered multilayers with precise spatial location of nanometric objects. The metal oxide nanocomposites synthesized have potential applications in catalysis, optical devices, surface-enhanced Raman scattering, and metal enhancement fluorescence.

Layer-by-layer self-assembly of glucose oxidase and Os(Bpy)2CIPyCH2NH-poly(allylamine) bioelectrode.

The uptake of glucose oxidase (GOx) onto a polycationic redox polymer (PAA-Os)-modified surface, by adsorption from dilute aqueous GOx solutions, was followed by the quartz crystal microbalance (QCM) and shows double exponential kinetics. The electrochemistry of the layer-by-layer-deposited redox-active polymer was followed by cyclic voltammetry in glucose-free solutions, and the enzyme catalysis mediated by the redox polymer was studied in beta-D-glucose-containing solutions. AFM studies of the different layers showed the existence of large two dimension enzyme aggregates on the osmium polymer for 1 microM GOx and less aggregation for 50 nM GOx solutions. When the short alkanethiol, 2,2'-diaminoethyldisulfide was preadsorbed onto gold, a monoexponential adsorption law was observed, and single GOx enzyme molecules could be seen on the surface where the enzyme was adsorbed from 50 nM GOx in water.

Layer-by-layer electrostatic deposition of biomolecules on surfaces for molecular recognition, redox mediation and signal generation.

Layer-by-layer supramolecular structures composed of alternate layers of negatively charged enzymes and cationic redox polyelectrolyte have been assembled. Glucose oxidase (GOx), lactate oxidase (LOx) and soybean peroxidase (SBP) have been electrically wired to the underlying electrode by means of poly(allylamine) with [Os(bpy)2ClPyCOH]+ covalently attached (PAA-Os) in organized structures with high spatial resolution. Biotinylated glucose oxidase has also been used to assemble step-by-step on antibiotin goat immunoglobulin (IgG) layers and the enzyme was electrically wired by PAA-Os. These spatially organized multilayers with mono- and bienzymatic schemes can work efficiently in molecular recognition, redox mediation and generation of an electrical signal. The concentration of redox mediator integrated into the multilayers, obtained from the voltammetric charge and an estimation of the layer thickness, exceeds by 100-fold the amount of deposited enzyme assessed by quartz crystal microbalance. Differences in GOx electrical wiring efficiency have been detected with the different assembling strategies. The surface concentration of electrically wired enzyme represents a small proportion of all the enzyme molecules present in the multilayers which can be oxidized by the soluble mediator [Os(bpy)2Cl PyCOOH]Cl. This proportion, as well as the rate of FADH2 oxidation by PAA-Os, increases with the number of electrically wired enzyme layers and with the spatial accessibility of the Os moiety to the enzyme active center.

Effect of ionic strength on the behavior of amperometric enzyme electrodes mediated by redox hydrogels.

The electron-transfer behavior of electroactive hydrogels formed by cross-linking ferrocene poly(allylamine) (Fc-PAA) and glucose oxidase is investigated as a function of electrolyte ionic strength using several techniques. Cyclic voltammetry and electrochemical impedance spectroscopy show that the quantity cD(e)(1/2) increases with electrolyte concentration. Enhancement of enzyme catalysis for the oxidation of glucose mediated by Fc-PAA is also apparent at higher KNO(3) concentration. The electroactive redox center concentration, c, and the diffusion coefficient due to electron hopping in the gel, D(e), are independently measured by chronoamperometry at ultramicroelectrodes. Larger electrolyte ionic strength induces an increase in electroactive redox center concentration while D(e) slightly decreases. These results are rationalized in terms of the electrostatic interactions within the redox gel backbone due to water and ion exchange with the external electrolyte, producing swelling and shrinking of the hydrogel.