MV-mimicking micelles loaded with PEG-serine-ACP nanoparticles to achieve biomimetic intra/extra fibrillar mineralization of collagen in vitro

Since their discovery, matrix vesicles (MVs) containing minerals have received considerable attention for their role in the mineralization of bone, dentin and calcified cartilage. Additionally, MVs' association with collagen fibrils, which serve as the scaffold for calcification in the organic matrix, has been repeatedly highlighted. The primary purpose of the present study was to establish a MVs–mimicking model (PEG-S-ACP/micelle) in vitro for studying the exact mechanism of MVs-mediated extra/intra fibrillar mineralization of collagen in vivo. In this study, high-concentration serine was used to stabilize the amorphous calcium phosphate (S-ACP), which was subsequently mixed with polyethylene glycol (PEG) to form PEG-S-ACP nanoparticles. The nanoparticles were loaded in the polysorbate 80 micelle through a micelle self-assembly process in an aqueous environment. This MVs–mimicking model is referred to as the PEG-S-ACP/micelle model. By adjusting the pH and surface tension of the PEG-S-ACP/micelle, two forms of minerals (crystalline mineral nodules and ACP nanoparticles) were released to achieve the extrafibrillar and intrafibrillar mineralization, respectively. This in vitro mineralization process reproduced the mineral nodules mediating in vivo extrafibrillar mineralization and provided key insights into a possible mechanism of biomineralization by which in vivo intrafibrillar mineralization could be induced by ACP nanoparticles released from MVs. Also, the PEG-S-ACP/micelle model provides a promising methodology to prepare mineralized collagen scaffolds for repairing bone defects in bone tissue engineering.     

Molecular Semiotics toward the Emergence of Life

Molecular imprints of organisms serving as both the agents and the products of the underlying sign activities are quantum mechanical in their origins. In particular, molecules in any reaction networks constituting a biological organism are semiotic or context-dependent in the sense that their activities reside within the proper coordination of the entire networks. The origin of life could have been related to a specific aspect of molecular semiotics, especially in the transition from molecules as the physical symbols of material units to molecules as the semiotic signs having the capacity of pointing to something else other than the molecules themselves. Quantum mechanical underpinning of the molecular imprints leading to the emergence of life is in the appraisal of the material capacities of both coherent assimilation and decoherent dissociation already latent in the imprints. One empirical evidence suggesting the likelihood of both coherent assimilation and decoherent dissociation in prebiotic settings could have been found in synthetic chemical reactions running in hydrothermal circulation of seawater through hot vents in the Haedean ocean on the primitive Earth.     

Cloning of genes for bacteriophage T5 stable RNAs

One EcoRI-generated fragment (440 basepairs) and two EcoRI/HindIII fragments (220 and 960 basepairs) from the deletion region of T5 phage have been inserted into the phage λ XIII and the plasmid pBR322 as vectors. Recombinant DNA molecules were studied by hybridization with in vivo ³²P-labeled T5 4–5 S RNAs on nitrocellulose filters. Two-dimensional polyacrylamide gel electrophoretic fractionation and fingerprint analysis of the RNAs eluted from the filters were carried out to identify RNAs coded by cloned fragments. For the accurate localization of the genes for these RNAs, RNA-DNA hybrids were treated with T1 and pancreatic RNAases, and the eluted RNA fragments stable against RNAase action were electrophoresed. It was shown that the EcoRI¹440 fragment contains the gene for tRNA 10 (tRNA^Asp), the EcoRI/HindIII¹220 fragment contains the gene for RNA III (107 bases) and parts of the genes for RNA I (107 bases) and tRNA 12 (tRNA^His), and the EcoRI/HindIII¹960 fragment contains only a part of the gene for tRNA 9 (tRNA^Gln). The arrangement of these genes on the physical map of T5 phage was as follows: -tRNA^Gln-tRNA^His-RNA III-RNA I-…-tRNA^Asp.     

Measurement of 7-dehydrocholesterol and cholesterol in hair can be used in the diagnosis of Smith-Lemli-Opitz syndrome

7-dehydrocholesterol (7-DHC) and cholesterol (CHOL) are biomarkers of Smith-Lemli-Opitz Syndrome (SLOS), a congenital autosomal recessive disorder characterized by elevated 7-DHC level in patients. Hair samples have been shown to have great diagnostic and research value, which has long been neglected in the SLOS field. In this study, we sought to investigate the feasibility of using hair for SLOS diagnosis. In the presence of antioxidants (2,6-ditert-butyl-4-methylphenol and triphenylphosphine), hair samples were completely pulverized and extracted by micro-pulverized extraction in alkaline solution or in n-hexane. After microwave-assisted derivatization with N,O-Bis(trimethylsilyl)trifluoroacetamide, the analytes were measured by GC-MS. We found that the limits of determination for 7-DHC and CHOL were 10 ng/mg and 8 ng/mg, respectively. In addition, good linearity was obtained in the range of 50–4000 ng/mg and 30–6000 ng/mg for 7-DHC and CHOL, respectively, which fully meets the requirement for SLOS diagnosis and related research. Finally, by applying the proposed method to real hair samples collected from 14 healthy infants and two suspected SLOS patients, we confirmed the feasibility of hair analysis as a diagnostic tool for SLOS. In conclusion, we present an optimized and validated analytical method for the simultaneous determination of two SLOS biomarkers using human hair.     

Activity estimation and biokinetic analysis of 99mTc-DMSA in renal infant patients using a gamma camera

Biokinetic data from the administration of radiopharmaceuticals is essential in nuclear medicine dosimetry. It has particular significance in children, as their metabolism is very different from adults. Biokinetic models for paediatric patients could therefore need to be adapted to better reflect their absorption, retention and excretion functions, when compared to adults. Obtaining quality in vivo infant or paediatric biokinetic data is then essential to improve the available reference models, which in turn can lead to the optimization of paediatric procedures and protocols in clinical practice.This study analyses the biokinetic behaviour of ^99mTc-dimercaptosuccinic acid (DMSA), in 8 infants aged 4 months to 2 years old, through an imaging study using a gamma camera, and compares the obtained values with those obtained with the reference ICRP biokinetic model. The in vivo data was treated using an adapted methodology from the MIRD 16 pamphlet. Activity curves for the liver, the kidney and the whole body, were built, and new effective absorption, retention and excretion half-lives were estimated, and compared with the reference biokinetic parameters of ICRP 128. The obtained residence time in the kidneys of 2.56 h, has a deviation of 30.8% to the ICRP 128 value of 3.70 h. The obtained maximum uptake in the kidneys was of 0.22/A₀, which compares to the value of 0.31/A₀ for ICRP.The obtained biokinetic parameters were used to estimate the absorbed dose. The obtained dose values are smaller than the reference ICRP 128 ones by 32.1% in the kidneys, and 18.4% in the liver.     

The Circadian Clock Gene Circuit Controls Protein and Phosphoprotein Rhythms in Arabidopsis thaliana

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Comparative study of the antioxidant activity of the essential oils of five plants against the H2O2 induced stress in Saccharomyces cerevisiae

The purpose of this work was to investigate the protective effect of five essential oils (EOs); Rosmarinus officinalis, Thymus vulgaris, Origanum compactum Benth., Eucalyptus globulus Labill. and Ocimum basilicum L.; against oxidative stress induced by hydrogen peroxide in Saccharomyces cerevisiae. The chemical composition of the EOs was analyzed by gas chromatography (GC) and gas chromatography-mass spectrometry (GC/MS). The in vitro antioxidant activity was evaluated and the protective effect of EOs was investigated. Yeast cells were pretreated with different concentrations of EOs (6.25–25 µg/ml) for an hour then incubated with H₂O₂ (2 mM) for an additional hour. Cell viability, antioxidants (Catalase, Superoxide dismutase and Glutathione reductase) and metabolic (Succinate dehydrogenase) enzymes, as well as the level of lipid peroxidation (LPO) and protein carbonyl content (PCO) were evaluated. The chemical composition of EOs has shown the difference qualitatively and quantitatively. Indeed, O. compactum mainly contained Carvacrol, O. basilicum was mainly composed of Linalool, T. vulgaris was rich in thymol, R. officinalis had high α-Pinene amount and for E. globulus, eucalyptol was the major compound. The EOs of basil, oregano and thyme were found to possess the highest amount of total phenolic compounds. Moreover, they have shown the best protective effect on yeast cells against oxidative stress induced by H₂O₂. In addition, in a dose dependent manner of EOs in yeast medium, treated cells had lower levels of LPO, lower antioxidant and metabolic enzymes activity than cells exposed to H₂O₂ only. The cell viability was also improved. It seems that the studied EOs are efficient natural antioxidants, which can be exploited to protect against damages and serious diseases related to oxidative stress.     

Comparison of Different Standard Methods to Evaluate the Total Concentrations of Heavy Metals in Waste Rock

The mining industry is a very rapidly increasing production sector in Finland. Many new mines have been opened recently and others are in the planning phase. Mining legislation has been recently revised and compilation of an ID-directive (Industrial Emission Directive, 2010) required to implement environmental law revision is in progress. The aim of this study was to compare different digestion methods recommended by the Finnish Environmental Administration for threshold and guideline value determinations in the Finnish PIMA Decree (214/2007) and Government Decree (591/2006) concerning recovery of certain wastes in earth construction. Altogether six methods were tested, of which five were microwave-assisted acid digestion methods: ISO 11466, EPA Method 3051A (aqua region), EPA Method 3051A (HNO₃), EPA Method 3051A (HNO₃/HCl), and SFS-EN 13656, while one was a fusion method with lithium tetraborate (ASTM C 1391 - 95). The results showed that concentrations of heavy metals (As, Cd, Co, Cu, Ni, Pb, Sb, V, and Zn) can be determined reproducibly with standard methods, but the chemical method used can have a significant impact on the results. This conclusion is very important, because the measured heavy metal values can affect the classification and processing of waste materials, and thereby can significantly impact the cost of treatment.