Examining metastatic behavior within 3D bioprinted vasculature for the validation of a 3D computational flow model.

Understanding the dynamics of circulating tumor cell (CTC) behavior within the vasculature has remained an elusive goal in cancer biology. To elucidate the contribution of hydrodynamics in determining sites of CTC vascular colonization, the physical forces affecting these cells must be evaluated in a highly controlled manner. To this end, we have bioprinted endothelialized vascular beds and perfused these constructs with metastatic mammary gland cells under physiological flow rates. By pairing these in vitro devices with an advanced computational flow model, we found that the bioprinted analog was readily capable of evaluating the accuracy and integrated complexity of a computational flow model, while also highlighting the discrete contribution of hydrodynamics in vascular colonization. This intersection of these two technologies, bioprinting and computational simulation, is a key demonstration in the establishment of an experimentation pipeline for the understanding of complex biophysical events.

Preparation and characterization of metallomicelles of Ru(II). Cytotoxic activity and use as vector.

The use of nanovectors in several medicinal treatments has reached a great importance in the last decade. Some drugs need to be protected to increase their lifetimes in the blood flow, to avoid degradation, to be delivered into target cells or to decrease their side effects. The goal of this work was to design and prepare nanovectors formed by novel surfactants derived from the [Ru(bpy)3]2+ complex. These amphiphilic molecules are assembled to form metallomicelles which can act as pharmaceutical agents and, at the same time, as nanovectors for several drugs. TEM images showed a structural transition from spherical to elongated micelles when the surfactant concentration increased. Fluorescence microscopy confirmed the internalization of these metallomicelles into diverse cell lines and cytotoxicity assays demonstrated specificity for some human cancer cells. The encapsulation of various antibiotics was carried out as well as a thorough study about the DNA condensation by the metallomicelles. To the best of our knowledge, applications of these metallomicelles have not been shown in the literature yet.

Cooperative interaction between metallosurfactants, derived from the [Ru(2,2'-bpy)3](2+) complex, and DNA.

With the idea of improving and advancing the design and preparation of new reagents based on cationic surfactants for gene therapy, two luminescent metallosurfactants derived from the [Ru(2,2'-bpy)3](2+) complex were synthesized. Their interaction with DNA and the effect they exert on the conformation of the polynucleotide were studied by using different techniques. The equilibrium binding constants, Kb, of the two surfactants to DNA were obtained at different molar ratios X=[surfactant]/[DNA]. The observed sigmoidal dependence of Kb on X confirms the cooperative character of the binding. After the addition of a determined surfactant concentration, the condensation of the polymer was observed. The amount of surfactant needed to produce this conformational change is lower for the double stranded surfactant than for the single chain surfactant due to a stronger hydrophobic interaction. The addition of α-cyclodextrin molecules to the metallosurfactant/DNA solutions results in polynucleotide decompaction, which confirms the importance of the hydrophobic interactions in the condensation of the polynucleotide. Results also show the importance of choosing both a proper system to study and the most seeming measuring technique to use. It is demonstrated that, in some cases, the use of several techniques is desirable to obtain reliable and accurate results.

Binding of DNA by a dinitro-diester calix[4]arene: denaturation and condensation of DNA.

A study of a dinitro-diester calix[4]arene (5,17-(3-nitrobenzylideneamino)-11,23-di-tert-butyl-25,27-diethoxycarbonyl methyleneoxy-26,28-dihydroxycalix[4]arene) interaction with calf-thymus DNA was carried out using several techniques. The measurements were done at various molar ratios X=[calixarene]/[DNA]. Results show diverse changes in the DNA conformation depending on the X value. Thus, at low macrocycle concentrations, the calixarene binds to the polynucleotide. This interaction, mainly in groove mode, weakens the hydrogen bonds between base pairs of the helix inducing denaturation of the double strands, as well as condensation of the macromolecule, from an extended coil state to a globular state. An opposite effect is observed at X molar ratios higher than 0.07. The de-condensation of DNA happens, that is, the transition from a compact state to a more extended conformation, probably due to the stacking of calixarene molecules in the solution. Results also show the importance of making a proper choice of the system under consideration.

Conformational changes of DNA in the presence of 12-s-12 gemini surfactants (s=2 and 10). Role of the spacer's length in the interaction surfactant-polynucleotide.

A multifaceted study on the interaction of calf-thymus DNA with two different cationic gemini surfactants alkanediyl-α-ω-bis(dodecyldimethyl-amonium)bromide, 12-s-12,2Br(-) (with s=2, G2, and 10, G10) was carried out. The measurements were done at different molar ratios X=[surfactant]/[DNA]. Results show two different conformational changes in DNA: a first compaction of the polynucleotide corresponding to a partial conformational (not total) change of DNA from an extended coil state to a globular state that happens at the lower molar ratio X. A second change corresponds to a breaking of the partial condensation, that is, the transition from the compacted state to a new more extended conformation (for the higher X values) different to the initial extension. According to circular dichroism spectra and dynamic light scattering measurements, this new state of DNA seems to be similar to a ψ-phase. Measurements confirm that interactions involved in the compaction are different to those previously obtained for the analog surfactant CTAB. X values at which the conformational changes happen depend on the length of the spacer in the surfactant along with the charge of the polar heads.

Interaction between monomers of two surfactants derived from the [Ru(2,2'-bpy)(3)](2+) complex and α, ß and γ-cyclodextrins: formation of [2]- and [3]-pseudorotaxanes.

Two new surfactants derived from the tris(2,2'-bipyridine) ruthenium(II) complex, [Ru(2,2'-bpy)(3)](2+), were synthesized and characterized: the double-tailed [Ru(2,2'-bipy)2(4,4'-(C(11)H(23))(2)-2,2'-bipy)](2+) surfactant (RuC11) and the mono-tailed [Ru(2,2'-bipy)(2)(4-(CH(3))-4'-(C(13)H(27))-2,2'-bipy)](2+) surfactant (RuC13). The main characteristic of these species is the presence of an inorganic complex as the polar head of the surfactant with interesting luminescence properties, which were used to study the interaction of these cationic surfactants with α-, ß- and γ-cyclodextrins (CD). The results showed the formation of [2]- and [3]-pseudorotaxanes. The binding constant values as well as the stoichiometry of the complexes formed were obtained; the results were confirmed, from a qualitative point of view, with NMR spectra.

Radical scavenging ability of polyphenolic compounds towards DPPH free radical.

Free radical scavenging activity of different polyphenolic compounds commonly present in wine has been evaluated using DPPH method. The experiments were performed with different amounts of phenols within the linear interval of response and with an excess of DPPH in all cases. In these conditions, for most of the compounds tested, the reaction was biphasic. Total stoichiometry values n confirm the implication of more than one step in the process. Flavan-3-ol compounds showed the highest values, especially procyanidins B1 (9.8) and B2 (9.1). In this family, n values coincide with the number of hydroxyl groups available. EC(50) and TEC(50) parameters have been calculated. EC(50) values are extremely diverse, being the procyanidins B1 and B2 the most potent scavenging compounds and resveratrol the less one. TEC(50) considers the rate of reaction towards the free radical. (+)-Catechin and (-)-epicatechin are the phenolic compounds that need more time to react. In contrast, caftaric and caffeic acids are the phenolic acids that react more rapidly. Antioxidant efficacy (AE) is a parameter that combines both factors. Compounds as kaempferol, with a high EC(50) value, could be considered as an antioxidant with low relevance, but instead shows the highest AE value of the phenolic compounds tested, due to its fast rate of reaction, what is of great biological importance.

First Report of Fusarium oxysporum f. sp. basilici in Sweet Basil Cultivation in Spain.

Recently, sweet basil (Ocimum basilicum L.) grown in greenhouses was introduced in Almería, Spain. It is typically cultivated in soil or perlite soilless culture. During the last 6 years, the following symptoms were observed sequentially in basil cultivation: yellowing and wilting of apical tips, wilting and necrosis of leaves and petioles, stunting, black lines along stems and petioles, and discoloration and necrosis of the xylem. Symptoms began at the apex and progressed to the plant base. Within 4 months of planting, symptoms developed in more than 14% of soil-cultivated plants, and in more than 13% of perlite-cultivated plants. Isolations from diseased xylem revealed the presence of Fusarium oxysporum. Inoculations were performed with a fungal suspension (104 CFU/ml) on basil cv. Genovesa, by drenching plants grown in sterile substrate or dipping the roots and transplanting plants into sterile substrate. Of 30 isolates, 80% were pathogenic and resulted in symptoms of the disease described above. The pathogen was reisolated from all inoculated plants. Inoculating Melissa officinalis L., Salvia officinalis L., Origanum majorana L., Mentha piperita L., Satureja hortensis L., and Thymus vulgaris L tested specificity of F. oxysporum. This test utilized the same methods used for basil. None of these species developed symptoms. Results indicated that symptoms of the disease on basil were caused by F. oxysporum f. sp. basilici. Since cultivation of basil is relatively new to Almería, it was necessary to determine the source of the inoculum. Accordingly, 3,200 seeds from Germany and Italy, the primary source of seed in Almería, were analyzed. F. oxysporum was isolated from 0.5% of the seeds. Following methods used earlier, all isolates were inoculated on basil. Fifty percent of the isolates reproduced the disease symptoms. The results suggest that the seeds from Germany and Italy were the source of the inoculum, and to our knowledge, introduced the disease into the growing basil cultures of Almería.

Scavenger receptor BI promotes high density lipoprotein-mediated cellular cholesterol efflux.

Scavenger receptor BI (SR-BI) binds high density lipoproteins (HDL) with high affinity and mediates the selective uptake of HDL cholesteryl ester. We examined the potential role of SR-BI in mediating cellular cholesterol efflux. In Chinese hamster ovary cells stably transfected with murine SR-BI, overexpression of SR-BI resulted in a 3-4-fold stimulation of initial cholesterol efflux rates. Efflux rates correlated with SR-BI expression in cells and HDL concentration in the medium. When incubated with synthetic cholesterol-free HDL, SR-BI-transfected cells showed approximately 3-fold increases in initial rates of efflux compared with control cells, indicating that SR-BI expression enhances net cholesterol efflux mediated by discoidal HDL. In six different cell types, including cultured macrophages, the rate of efflux of cholesterol mediated by HDL or serum was well correlated with cellular SR-BI expression level. In addition, in situ hybridization experiments revealed that SR-BI mRNA was expressed in the thickened intima of atheromatous aorta of apolipoprotein E knockout mice. Thus, SR-BI is an authentic HDL receptor mediating cellular cholesterol efflux. SR-BI may facilitate the initial steps of HDL-mediated cholesterol efflux in the arterial wall as well as later steps of reverse cholesterol transport involving uptake of HDL cholesterol in the liver.