Statistical properties of contact vectors

We study the statistical properties of contact vectors, a construct to characterize a protein's structure. The contact vector of an N-residue protein is a list of N integers n_i, representing the number of residues in contact with residue i. We study analytically (at mean-field level) and numerically the amount of structural information contained in a contact vector. Analytical calculations reveal that a large variance in the contact numbers reduces the degeneracy of the mapping between contact vectors and structures. Exact enumeration for lengths up to N=16 on the three dimensional cubic lattice indicates that the growth rate of number of contact vectors as a function of N is only 3% less than that for contact maps. In particular, for compact structures we present numerical evidence that, practically, each contact vector corresponds to only a handful of structures. We discuss how this information can be used for better structure prediction.Comment: 20 pages, 6 figure

THE ROLE OF DISPERSIVE FORCES IN THE ONSET OF SUPERRADIANT BEATING

Superradiance is a stochastic process which shows macroscopic shot to shot fluctuations in some of the properties of the emitted pulse. These fluctuations may have microscopic and quantum-mechanical origin. The aim of this paper is to discuss a simple and hopefully intriguing example of one of the approaches used to deal with the statistical properties of this process

Prevention of UVA-induced oxidative damage in human dermal fibroblasts by new UV filters, assessed using a novel in vitro experimental system.

BACKGROUND: UVA rays present in sunlight are able to reach the dermal skin layer generating reactive oxygen species (ROS) responsible for oxidative damage, alterations in gene expression, DNA damage, leading to cell inflammation, photo-ageing/-carcinogenesis. Sunscreens contain UV filters as active ingredients that absorb/reflect/dissipate UV radiation: their efficiency depends on their spectral profile and photostability which should then be reflected in biological protection of underlying skin. METHODS: A set of new UV filters was synthesized, and the most photostable one was compared to BMDBM, a widely used UVA filter. Cultured human dermal fibroblasts were exposed to UVA radiation which was filtered by a base cream containing or not UV filters placed above cell culture wells. The endpoints measured were: cell viability (MTT assay), ROS generation (DCFH-DA assay), mitochondrial function (JC-1 assay), DNA integrity (Comet assay) and gene expression (MMP-1, COL1A1) by RT-qPCR. RESULTS: The new UV filter resulted more efficient than BMDBM in preserving cell viability, mitochondrial functionality and oxidative DNA damage, despite similar inhibition levels of intracellular ROS. Moreover, expression of genes involved in dermal photoageing were positively affected by the filtering action of the tested molecules. CONCLUSIONS: The experimental model proposed was able to validate the efficacy of the new UV filter, taking into account important cellular events related to UV-induced intracellular oxidative stress, often underestimated in the assessments of these compounds. GENERAL SIGNIFICANCE: The model may be used to compare the actual biological protection of commercial sunscreens and suncare products aside from their SPF and UVA-PF values

Reference gene validation for qPCR on normoxia- and hypoxia-cultured human dermal fibroblasts exposed to UVA: is β-actin a reliable normalizer for photoaging studies?

Data normalization of gene expression on human dermal fibroblasts (HDF) exposed to UVA has commonly been done using either GAPDH or β-actin as reference genes without any validation of their expression stability. Since this aspect, important for accurate normalization, has been overlooked, we aimed to establish a suitable set of reference genes for studies on UVA-treated HDF cultured under both standard atmospheric oxygen tension (normoxia, 21%) and under a physiological, low oxygen tension for these cells (hypoxia, 5%). The stability of six commonly used reference genes was assessed using the geNorm and NormFinder softwares subsequent to reverse-transcription quantitative real-time PCR (RT-qPCR). GAPDH/SDHA were found to be the most stable genes under normoxia, while SDHA/TBP or HPRT1/β2M were the most stable ones under hypoxia in HDF exposed to 18 J/cm(2) UVA. β-Actin was always the most unstable reference gene. To emphasize the importance of selecting the most stably expressed reference genes for obtaining reliable results, mRNA expression levels of MMP-1 and COL1A1 were analyzed vs the best reference genes and the worst one. These reference genes are hence recommended for future qPCR analyses in studies concerning photo-damage on UVA-treated HDF

A novel Real Time PCR strategy to detect SOD3 SNP using LNA probes.

Extracellular superoxide dismutase (SOD3) is the primary enzymatic antioxidant defence of the vascular wall. The physiopathological role of SOD3 has been examined in vascular-related diseases, atherosclerosis, hypertension, diabetes, ischaemia-reperfusion injury, lung disease, various inflammatory conditions, and neurological diseases. An important single nucleotide polymorphism (SNP), nt.760 G>C of the SOD3 gene (rs#1799895) leads to the amino acid substitution Arg(213)Gly (R213G) in the center of the heparin-binding domain and consequently to a lowered affinity for the endothelium. This mutation, which occurs with a relatively high frequency in the population (4\% of Swedish, 3\% of Australian and 6\% of Japanese people), is associated with decreased tissue antioxidant defences and increased risk of ischaemic heart disease. The identification of patients carrying this mutation is therefore of great interest in order to highlight lowered antioxidant defences at a vascular level which could lead to increased susceptibility toward coronary artery disease and atherogenesis. Here we describe a method to detect the 760 G>C single nucleotide polymorphism based on Real Time PCR strategy using locked nucleic acid (LNA) probes. This technique, a modification of classic TaqMan probes SNP genotyping, amplifies and detects the mutation in a single reaction tube. Moreover, the implementation of LNA probes remarkably increases the specificity of the reaction. The proposed method enables unambigous and rapid discrimination of wild type and mutant genotype both in plasmid and genomic DNA samples. In light of the role of SOD3 polymorphism, the genotyping of 760 G>C mutant has important clinical implications. The proposed assay combines rapidity, high specificity, can be easily automated and overall reduces labor and cost of analyses. Moreover, identification of patients with lowered vascular antioxidant defences could address pharmacogenomical approaches to the therapy of cardiovascular diseases

Prolonged coenzyme Q10 treatment in Down syndrome patients: effect on DNA damage

Oxidative stress is known to play a relevant role in Down syndrome (DS) and its effects are documented from embryonic life. Oxidative DNA damage has been shown to be significantly elevated in Down syndrome patients, and this has been indicated as an early event promoting neurodegeneration and Alzheimer type dementia. The aim of this study was to investigate the efficacy of coenzyme Q(10) (CoQ(10)) in delaying the effect of oxidative damage in these patients. In our previous study we demonstrated a mild protective effect of CoQ(10) on DNA, although the treatment was unable to modify the overall extent of oxidative damage at the patient level. Possible limitations of the previous study were: time of treatment (6 months) or spectrum of DNA lesions detected. In order to overcome these limitations we planned a continuation of the trial aimed at evaluating the effects of CoQ(10) following a prolonged treatment. Our results highlight an age-specific reduction in the percentage of cells showing the highest amount of oxidized bases, indicating a potential role of CoQ(10) in modulating DNA repair mechanisms

Vitamin MK-7 enhances vitamin D3-induced osteogenesis in hMSCs: modulation of key effectors in mineralization and vascularization

The osteoblast is the bone-forming cell and is derived from mesenchymal stem cells (MSCs). Osteo-inductive substances could represent a useful therapeutic approach during the fracture repair process. The aim of this work was to evaluate the effects of vitamin MK-7, alone or in association with vitamin D3, in differentiating human MSCs (hMSCs) in vitro along the osteoblastic lineage. In particular, primary endpoints of the study include gene and protein markers of osteoblast differentiation. Considering genes involved in bone formation and mineralization, our data show that vitamin MK-7 enhances vitamin D3 gene induction of osteocalcin (OC). Among genes related to cell growth and differentiation, a specific effect of vitamin MK-7 was observed for growth differentiation factor-10 (GDF10) and insulin-like growth factor 1 (IGF1), the latter being also involved in the induction of vascular endothelial growth factors (VEGFA). Accordingly, vitamin co-supplementation greatly affected VEGFA and its receptor fms-related tyrosine kinase 1 (FLT1), a key factor in both angiogenic and osteogenic processes. These results stress the relevance of MK-7 and D3 co-supplementation in the bone-healing process as able to modulate the expression of genes involved in both mineralization and angiogenesis. Moreover, at the protein level co-association of vitamins might provide an optimal balance between induction and carboxylation of osteocalcin, essential for its functionality in the extracellular matrix (ECM). Our results may provide hints for therapeutic application of hMSCs in bone disease, clarifying mechanisms involved in stem cell-mediated bone development, and they also highlight the relevance of co-supplementation strategies, since single supplementations might result in a suboptimal effect