Conformational altered p53 affects neuronal function: relevance for the response to toxic insult and growth-associated protein 43 expression

The role of p53 in neurodegenerative diseases is essentially associated with neuronal death. Recently an alternative point of view is emerging, as altered p53 conformation and impaired protein function have been found in fibroblasts and blood cells derived from Alzheimer's disease patients. Here, using stable transfected SH-SY5Y cells overexpressing APP751wt (SY5Y-APP) we demonstrated that the expression of an unfolded p53 conformation compromised neuronal functionality. In particular, these cells showed (i) augmented expression of amyloid precursor protein (APP) and its metabolites, including the C-terminal fragments C99 and C83 and β-amyloid peptide (ii) high levels of oxidative markers, such as 4-hydroxy-2-nonenal Michael-adducts and 3-nitro-tyrosine and (iii) altered p53 conformation, mainly due to nitration of its tyrosine residues. The consequences of high-unfolded p53 expression resulted in loss of p53 pro-apoptotic activity, and reduction of growth-associated protein 43 (GAP-43) mRNA and protein levels. The role of unfolded p53 in cell death resistance and lack of GAP-43 transcription was demonstrated by ZnCl(2) treatment. Zinc supplementation reverted p53 wild-type tertiary structure, increased cells sensitivity to acute cytotoxic injury and GAP-43 levels in SY5Y-APP clone

Conformational altered p53 as an early marker of oxidative stress in Alzheimer\u27s disease

In order to study oxidative stress in peripheral cells of Alzheimer\u27s disease (AD) patients, immortalized lymphocytes derived from two peculiar cohorts of patients, referring to early onset AD (EOSAD) and subjects harboured AD related mutation (ADmut), were used. Oxidative stress was evaluated measuring i) the typical oxidative markers, such as HNE Michel adducts, 3 Nitro-Tyrosine residues and protein carbonyl on protein extracts, ii) and the antioxidant capacity, following the enzymatic kinetic of superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GRD). We found that the signs of oxidative stress, measured as oxidative marker levels, were evident only in ADmut but not in EOSAD patients. However, oxidative imbalance in EOSAD as well as ADmut lymphocytes was underlined by a reduced SOD activity and GRD activity in both pathological groups in comparison with cells derived from healthy subjects. Furthermore, a redox modulated p53 protein was found conformational altered in both EOSAD and ADmut B lymphocytes in comparison with control cells. This conformational altered p53 isoform, named unfolded p53 , was recognized by the use of two specific conformational anti-p53 antibodies. Immunoprecipitation experiments, performed with the monoclonal antibodies PAb1620 (that recognizes p53wt) and PAb240 (that is direct towards unfolded p53), and followed by the immunoblotting with anti-4-hydroxynonenal (HNE) and anti- 3-nitrotyrosine (3NT) antibodies, showed a preferential increase of nitrated tyrosine residues in unfolded p53 isoform comparing to p53 wt protein, in both ADmut and EOSAD. In addition, a correlation between unfolded p53 and SOD activity was further found. Thus this study suggests that ROS/RNS contributed to change of p53 tertiary structure and that unfolded p53 can be considered as an early marker of oxidative imbalance in these patients

Conformational Altered p53 as an Early Marker of Oxidative Stress in Alzheimer's Disease

In order to study oxidative stress in peripheral cells of Alzheimer's disease (AD) patients, immortalized lymphocytes derived from two peculiar cohorts of patients, referring to early onset AD (EOSAD) and subjects harboured AD related mutation (ADmut), were used. Oxidative stress was evaluated measuring i) the typical oxidative markers, such as HNE Michel adducts, 3 Nitro-Tyrosine residues and protein carbonyl on protein extracts, ii) and the antioxidant capacity, following the enzymatic kinetic of superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GRD). We found that the signs of oxidative stress, measured as oxidative marker levels, were evident only in ADmut but not in EOSAD patients. However, oxidative imbalance in EOSAD as well as ADmut lymphocytes was underlined by a reduced SOD activity and GRD activity in both pathological groups in comparison with cells derived from healthy subjects. Furthermore, a redox modulated p53 protein was found conformational altered in both EOSAD and ADmut B lymphocytes in comparison with control cells. This conformational altered p53 isoform, named “unfolded p53”, was recognized by the use of two specific conformational anti-p53 antibodies. Immunoprecipitation experiments, performed with the monoclonal antibodies PAb1620 (that recognizes p53wt) and PAb240 (that is direct towards unfolded p53), and followed by the immunoblotting with anti-4-hydroxynonenal (HNE) and anti- 3-nitrotyrosine (3NT) antibodies, showed a preferential increase of nitrated tyrosine residues in unfolded p53 isoform comparing to p53 wt protein, in both ADmut and EOSAD. In addition, a correlation between unfolded p53 and SOD activity was further found. Thus this study suggests that ROS/RNS contributed to change of p53 tertiary structure and that unfolded p53 can be considered as an early marker of oxidative imbalance in these patients

Keep them alive! Design and Evaluation of the “Community Fostering Reference Model”

Firms host online communities for commercial purposes, for example in order to integrate customers into ideation for new product development. The success of these firm-hosted online communities depends entirely on the cooperation of a high number of customers that constantly produce valuable knowledge for firms. However, in practice, the majority of successfully implemented communities suffers from stagnation and even a decrease of member activities over time. Literature provides numerous guidelines on how to build and launch these online communities. While these models describe the initial steps of acquiring and activating a community base from scratch very well and explicitly, they neglect continuous member activation and acquistion after a successful launch. Against this background, the authors propose the Community Fostering Reference Model (CoFoRM), which represents a set of general procedures and instruments to continuously foster member activity. In this paper, the authors present the theory-driven design as well as the evaluation of the CoFoRM in a practical use setting. The evaluation results reveal that the CoFoRM represents a valuable instrument in the daily working routine of community managers, since it efficiently helps activating community members especially in the late phases of a community’s LifeCycle

"Diffusion of web-based product innovation"

Customers have proven to be a precious source of new solutions in various product categories and through their direct involvement in the innovation process, companies can better anticipate market changes. The Web can greatly simplify these activities by making it easier to manage systematic interactions with a select group of customers at a low cost. However, empirical evidence regarding the actual diffusion of Web-based tools supporting collaborative innovation remains weak. After reviewing the past findings on web-based tools for customer integration, we provide data on an exploratory analysis of over 200 brand and corporate sites. With our results we highlight the tools that are mainly used by firms; we also identify some relevant industrial and firm-specificities. We conclude by providing managerial implications of our findings and some directions of future research

Il ruolo del Web ai fini del coinvolgimento del cliente nei processi di innovazione: teoria e prassi a confronto

L'articolo esplora la diffusione degli strumenti Web-based per coinvolgere il consumatore nei processi di innovazione gestiti dall'impresa in cinque diversi settor

A new testing-apparatus simulating sliding between brackets and orthodontic wires

it's very important to test the sliding effects in orthodontic phases. it's necessary obtain a good sliding to prevent collateral effects in teeth movement. this study creates in vitro the simulating movement to test different kinds of bkt frictio

Ascorbic acid rescues cardiomyocyte development in Fgfr1-/- murineembryonic stem cells

Fibroblast growth factor receptor 1 (Fgfr1) gene knockout impairs cardiomyocyte differentiation in murine embryonic stem cells (mESC). Here, various chemical compounds able to enhance cardiomyocyte differentiation in mESC were tested for their ability to rescue the cardiomyogenic potential of Fgfr1-/- mESC. Among them, only the reduced form of vitC, L-ascorbic acid, was able to recover beating cell differentiation in Fgfr1-/- mESC. The appearance of contracting cells was paralleled by the expression of early and late cardiac gene markers, thus suggesting their identity as cardiomyocytes. In the attempt to elucidate the mechanism of action of vitC on Fgfr1-/- mESC, we analyzed several parameters related to the intracellular redox state, such as reactive oxygen species content, Nox4 expression, and superoxide dismutase activity. The results did not show any relationship between the antioxidant capacity of vitC and cardiomyocyte differentiation in Fgfr1-/- mESC. No correlation was found also for the ability of vitC to modulate the expression of pluripotency genes. Prolyl hydroxylases (PHD) are members of the iron- and 2-oxoglutarate-dependent dioxygenase family, that direct hypoxia inducible factor (HIF)-1α to degradation in normoxic conditions. To test the hypothesis that vitC was acting by maintaining iron ion in the reduced form (Fe2+), we initially evaluated HIF-1α levels in the undifferentiated state of mESC lines. Western blot analysis of isolated nuclei showed a higher HIF-1α content in undifferentiated Fgfr1- /- mESC versus the heterozygous counterpart. Unexpectedly, vitC treatment similarly up regulates HIF- 1α content in both cell lines, thus preventing any conclusion from this result. However, treatment of Fgfr1-/- mESC with both Fe2+ or the HIF inhibitor CAY10585 were able to rescue cardiomyocyte differentiation. In conclusion, our data point to a crucial role for HIF-1α modulation in Fgfr1-dependent cardiomyogenesis