DNA-protein noncovalent cross-linking: ruthenium dipyridophenazine biotin complex for the assembly of proteins and gold nanoparticles on DNA templates

We report the first example of a small molecule that can noncovalently cross-link DNA with streptavidin and streptavidin-labeled materials. Molecule 1 possesses a ruthenium dipyridophenazine DNA-intercalating moiety and a biotin unit; these two units are adequately separated to ensure efficient cross-linking of DNA with protein. Complex 1 is essentially nonemissive in aqueous solution and when bound to streptavidin, however, its luminescence is turned "on" when it binds to DNA. We have used these properties to establish that this complex can simultaneously bind to DNA and streptavidin, and can thus bring these two biomolecules together. We also synthesized a related molecule, 3, in which the biotin and DNA-intercalating moieties are covalently bound. While complex 3 can intercalate into DNA through a threading mechanism, luminescence experiments show that it cannot simultaneously bind DNA and streptavidin, most likely due to the proximity of its two molecular-recognition units. The cross-linking ability of molecule 1 was used to template the assembly of streptavidin molecules on circular plasmid DNA, as visualized by atomic force microscopy. In addition, using 1, we show the organization of discrete groups of gold nanoparticles labeled with streptavidin on a linear DNA template of finite size, with transmission electron microscopy. In these experiments the DNA template acted as a "molecular ruler" that dictated the number of particles in the assembly.     

In Vitro Photodynamic Activity of N‐Methylated and N‐Oxidised Tripyridyl Porphyrins with Long Alkyl Chains and Their Inhibitory Activity in Sphingolipid Metabolism

A series of N‐methylated and N‐oxidised tripyridyl porphyrins were synthesised, characterised, and their PDT activity was studied with six cell lines. All the tested porphyrins with a long alkyl chain, except one, were more efficient for PDT than an N‐methylated hydrophilic porphyrin and N‐oxidised porphyrin without the long alkyl chain. Generally, N‐methylated tripyridyl porphyrins were more active than those N‐oxidised, but IC₅₀ values for phototoxicity of two N‐oxides, named TOPyP3‐C₁₇H₃₃O and TOPyP3‐C₁₇H₃₅ , were still in the nanomolar concentration range for most of the tested cell lines. However, TOPyP3‐C₁₇H₃₅ did not show phototoxicity on human foreskin fibroblast cells. Two methylated amphiphilic porphyrins, named TMPyP3‐C₁₇H₃₃ and TMPyP4‐C₁₇H_{35 ,} showed significant dark toxicity, whereas none of the oxidopyridyl porphyrins were toxic without light activation. The selected photosensitisers were shown to be apoptosis inducers, and had inhibitory effects on the clonogenic growth of HCT116 and HeLa cells. All three N‐methylated amphiphilic porphyrins significantly reduced the migratory potential of HCT116 cells. Porphyrins TMPyP3‐C₁₇H₃₅ and TOPyP3‐C₁₇H₃₅ reduced the activity of acid ceramidase, whereas TOPyP3‐C₁₇H₃₃O had a significant inhibitory effect on sphingosine kinase 1 activity in HeLa cells. Compounds with this dual activity were shown to be the most promising photosensitisers, with potential to treat invasive cancers.     

The Effects of Cryogenic Storage on Human Dental Pulp Stem Cells

Dental pulp stem cells (DPSCs) are a type of easily accessible adult mesenchymal stem cell. Due to their ease of access, DPSCs show great promise in regenerative medicine. However, the tooth extractions from which DPSCs can be obtained are usually performed at a period of life when donors would have no therapeutic need of them. For this reason, it is imperative that successful stem cell storage techniques are employed so that these cells remain viable for future use. Any such techniques must result in high post-thaw stem cell recovery without compromising stemness, proliferation, or multipotency. Uncontrolled-rate freezing is not a technically or financially demanding technique compared to expensive and laborious controlled-rate freezing techniques. This study was aimed at observing the effect of uncontrolled-rate freezing on DPSCs stored for 6 and 12 months. Dimethyl sulfoxide at a concentration of 10% was used as a cryoprotective agent. Various features such as shape, proliferation capacity, phenotype, and multipotency were studied after DPSC thawing. The DPSCs did not compromise their stemness, viability, proliferation, or differentiating capabilities, even after one year of cryopreservation at −80 °C. After thawing, they retained their stemness markers and low-level expression of hematopoietic markers. We observed a size reduction in recovery DPSCs after one year of storage. This observation indicates that DPSCs can be successfully used in potential clinical applications, even after a year of uncontrolled cryopreservation.     

Multivariate multinomial T2 control chart using fuzzy approach

Quality of a product is often measured through various quality characteristics generally correlated. Multivariate control charts are a response to the need for quality control in such situations. If quality characteristics are qualitative, it sometimes happens that the product quality is defined by linguistic variables – where quality levels are represented by some specific words – and product units are classified into several linguistic forms categories, depending on the degree of fulfilment of expectations, creating a situation of fuzzy classifications. This study first reviews the concepts found in the literature on the development of fuzzy multivariate control charts. We propose a method to control these fuzzy quality evaluations, with correlated multiple attributes quality characteristics, through the use of a Hotelling T² control chart.     

Benchmarking of the pre/post-combustion chemical absorption for the CO2 capture

The aim of the article was to compare the pre- and post-combustion CO₂ capture process employing the chemical absorption technology. The integration of the chemical absorption process before or after the coal combustion has an impact on the power plant efficiency because, in both cases, the thermal energy consumption for solvent regeneration is provided by the steam extracted from the low pressure steam turbine. The solvent used in this study for the CO₂ capture was monoethanolamine (MEA) with a weight concentration of 30%. In the case of the pre-combustion integration, the coal gasification was analysed for different ratios air/fuel (A/F) in order to determine its influences on the syngas composition and consequently on the low heating value (LHV). The LHV maximum value (28 MJ/kg) was obtained for an A/F ratio of 0.5 kg_air/kg_fuel, for which the carbon dioxide concentration in the syngas was the highest (17.26%). But, considering the carbon dioxide capture, the useful energy (the difference between the thermal energy available with the syngas fuel and the thermal energy required for solvent regeneration) was minimal. The maximum value (61.59 MJ) for the useful energy was obtained for an A/F ratio of 4 kg_air/kg_fuel. Also, in both cases, the chemical absorption pre- and post-combustion process, the power plant efficiency decreases with the growth of the L/G ratio. In the case of the pre-combustion process, considering the CO₂ capture efficiency of 90%, the L/G ratio obtained was of 2.55 mol_solvent/mol_syngas and the heat required for the solvent regeneration was of 2.18 GJ/tCO₂. In the case of the post-combustion CO₂ capture, for the same value of the CO₂ capture efficiency, the L/G ratio obtained was of 1.13 mol_solvent/mol_{flue gas} and the heat required was of 2.80 GJ/tCO₂. However, the integration of the CO₂ capture process in the power plant leads to reducing the global efficiency to 30% in the pre-combustion case and to 38% to the post-combustion case.     

Immunohistochemical Expression Pattern of FGFR1, FGFR2, RIP5, and HIP2 in Developing and Postnatal Kidneys of Dab1−/− (yotari) Mice

This study aimed to explore how Dab1 gene functional silencing influences the spatial and temporal expression patterns of fibroblast growth factor receptor 1 (FGFR1), fibroblast growth factor receptor 2 (FGFR2), receptor-interacting protein kinase 5 (RIP5), and huntingtin-interacting protein 2 (HIP2) in the developing and postnatal kidneys of the yotari mice as potential determinants of normal kidney formation and function. Dab1^−/− animal kidneys exhibit diminished FGFR1/FGFR2 expression in all examined developmental stages, whereas RIP5 cell immunoreactivity demonstrated negligible variation. The HIP2 expression revealed a discernible difference during the postnatal period, where we noted a significant decrease in almost all the observed kidney structures of yotari animals. An extracellular signal-regulated kinase (Erk1/2) and mammalian target of rapamycin (mTOR) expression in yotari kidneys decreased in embryonic and postnatal developmental phases for which we can hypothesize that the Erk1/2 signaling pathway in the yotari mice kidneys is dependent on Reelin with Dab1 only partially implicated in Reelin-mediated MEK/Erk1/2 activation. The impairment of FGFR1 and FGFR2 expression suggests the involvement of the observed markers in generating the CAKUT phenotype resulting in renal hypoplasia. Our study demonstrates the critical role of HIP2 in reducing cell death throughout nephrogenesis and maturation in wild-type mice and indicates a possible connection between decreased HIP2 expression in postnatal kidney structures and observed podocyte injury in yotari. Our results emphasize the crucial function of the examined markers throughout normal kidney development and their potential participation in kidney pathology and diagnostics, where they might serve as biomarkers and therapeutic targets.     

Designing gluten-free,anthocyanins-enriched cookies on scientific basis

The essential treatment of the coeliac disease is a strict lifelong gluten-free diet based on the avoidance of gluten, thus requiring new perspectives for identifying technological and functional alternatives for gluten-free products, with similar functionality and efficiency. Therefore, the objective of this study was to explore the potential of purple corn flour (PCF) as a functional and technological alternative to design new gluten-free, anthocyanins-enriched cookies. The PCF was characterised in terms of phytochemical and antioxidant activity, yielding an extract with a total anthocyanin content of 14.94 ± 0.68 mg cyanidin-3-O-glucoside (C3G) per g dry weight (DW). The results of the thermo-mechanical tests indicated that composite flour consisting of rice flour (RF, 75%) and PCF (25%) is suitable for obtaining gluten-free cookies. Tests showed increased anthocyanin content in cookies with 75% addition of PCF, up to 6.99 ± 0.20 mg C3G per 100 g DW, yielding an antioxidant activity of 18.46 ± 0.18 mm Trolox equivalents per g DW. The ELISA test confirmed the absence of prolamins recognised by the monoclonal antibody R5. The sensory analysis revealed that the substitution of RF with PCF had no significant influence on overall acceptability. The results of the shelf-life tests performed on the cookies showed slight variation in phytochemicals in both variants, influencing the antioxidant activity. The obtained results may bring potential benefits to people on restricted diets and coeliac consumers, which can benefit from a contribution of bioactives, with significant health benefits.     

Influence Of Trehalose Addition On Instrumental Textural Properties Of Strawberry Pastes

Objective of this study was investigation of influence of trehalose addition (3, 5 and 10%) on instrumental textural properties of evaporated and freeze-dried strawberry pastes. As a control, strawberry paste without addition of trehalose was used. Addition of trehalose had influence on both, hardness and stickiness of the samples, but it is very important to define the amount of trehalose in order to achieve desirable properties. With addition of trehalose hardness of both, evaporated and freeze-dried samples decreased, from 29.89 N to 14.72 N for evaporated samples and from 184.83 N to 44.95 N in freeze-dried samples. Also, from those results it could be seen that freeze-dried samples were much harder than evaporated ones, regardless of trehalose addition. While hardness of samples decreased with addition of trehalose, stickiness increased. Stickiness values were from 0.016 N to 0.0102 N for evaporated samples, and from 0.066 N to 0.018 N for freeze-dried samples. Since evaporated samples had lower values for stickiness they were stickier than freeze-dried ones.     

Photodynamic Inactivation of Legionella pneumophila Biofilm Formation by Cationic Tetra- and Tripyridylporphyrins in Waters of Different Hardness

The bacterium Legionella pneumophila is still one of the probable causes of waterborne diseases, causing serious respiratory illnesses. In the aquatic systems, L. pneumophila exists inside free-living amoebae or can form biofilms. Currently developed disinfection methods are not sufficient for complete eradication of L. pneumophila biofilms in water systems of interest. Photodynamic inactivation (PDI) is a method that results in an antimicrobial effect by using a combination of light and a photosensitizer (PS). In this work, the effect of PDI in waters of natural origin and of different hardness, as a treatment against L. pneumophila biofilm, was investigated. Three cationic tripyridylporphyrins, which were previously described as efficient agents against L. pneumophila alone, were used as PSs. We studied how differences in water hardness affect the PSs’ stability, the production of singlet oxygen, and the PDI activity on L. pneumophila adhesion and biofilm formation and in biofilm destruction. Amphiphilic porphyrin showed a stronger tendency for aggregation in hard and soft water, but its production of singlet oxygen was higher in comparison to tri- and tetracationic hydrophilic porphyrins that were stable in all water samples. All three studied porphyrins were shown to be effective as PDI agents against the adhesion of the L. pneumophila to polystyrene, against biofilm formation, and in the destruction of the formed biofilm, in their micromolar concentrations. However, a higher number of dissolved ions, i.e., water hardness, generally reduced somewhat the PDI activity of all the porphyrins at all tested biofilm growth stages.