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Background and objectives: Promoting healthy behaviors throughout life is an essential prevention tool against chronic disease onset and progression in adulthood. This study aims to explore the lifestyles of an Italian academic community to plan targeted initiatives to promote and improve healthy lifestyles in students and employees with consequent impact on their entourage. Methods: A sample of 8715 adults (mean age = 26 years; range = 18-76; 30% male) participated in an online survey designed to evaluate the associations among lifestyle profiles (including diet, physical activity, cigarette smoking, and alcohol consumption), sociodemographic factors (age, gender, and academic role), and Body Mass Index (BMI). Lifestyle profiles were identified through cluster analysis, and a multinomial logistic regression was then performed to relate clusters obtained to sociodemographic variables and BMI. Results: Data show that older age was associated with the probability of belonging to the clusters with the worst behavior in smoking and alcohol consumption, but with the healthiest diet. The younger the age, the greater the probability of belonging to the more physically active cluster. Men were more likely than women to belong to the lifestyle profile with the highest alcohol consumption and the highest physical activity. Lower BMI was associated with the highest amount of physical activity profile. Conclusion: This study shed light on factors associated with different co-occurring health-related behaviors that should be considered in planning effective communication strategies promoting adherence to health claims.

Objectives: The aim of this work was to evaluate the activation of oxidative stress and inflammation in mice exposed to ultrafine particles (UFPs), and to elucidate putative physiopathological correlations with neurodegeneration. Materials: UFPs derived from two different anthropogenic sources were used: DEP, from EURO 4 diesel engine emission, and BC, from pellet boiler biomass emission (ENEA). Method: Male BALB/c mice were submitted to single or repeated intratracheal instillation of DEP and BC (50µg); in parallel, control mice were always considered (sham). Cortex, cerebellum and hippocampus from sham and treated mice were screened for markers of inflammation and oxidative stress (iNOS, HO-1), endoplasmic reticulum stress (Hsp70), a putative pro-carcinogenic marker (Cyp1B1) and markers related to the onset of Alzheimer Disease (AD) (P-Tau, Tau, P-APP, APP, BACE1). Results: After both single and repeated DEP and BC instillation, we observed an increase in iNOS, HO-1 and Hsp70 levels in all districts, while Cyp1B1 and markers of neurodegeneration changed only after repeated treatment. Discussion: The single DEP and BC instillation induced oxidative stress, endoplasmic reticulum stress and inflammation, while typical AD-related proteins seemed not affected. The repeated instillation was able to sustain oxidative stress and inflammation; moreover, it induced increase of BACE1 levels and changes in Tau and APP total levels and phosphorylation. Finally, BC seemed to be less effective than DEP. Conclusions: Our results suggest UFPs or inflammatory mediators translocation from lungs, and their effects on the brain, after both acute and repeated DEP and BC treatment. Grant by Fondazione Cariplo. Genc S., Zadeoglulari Z., Fuss S.H., Genc K. The adverse effects of air pollution on the nervous system. J Toxicol. 2012. 2012:782462. Levesque S., Surace M.J., McDonald J., Block M.L. Air pollution & the brain: Subchronic diesel exhaust exposure causes neuroinflammation and elevates early markers of neurodegenerative disease. J Neuroinflammation. 2011. 8:105. Block M.L. and Calderón-Garcidueñas L. Air pollution: mechanisms of neuroinflammation and CNS disease. Trends Neurosci. 2009. 9: 506–516.

Objectives: Understanding the ischemic injury contribution to Alzheimer’s disease (AD) onset, pinpointing to post-translational modifications, turnover alterations, secretion of proteins and networks identified as AD hallmarks. Materials: Neuronal cultures prepared from hippocampi of E18-E19 embryos from pregnant Sprague Dawley rats. 5%CO2: 95%N2 gaseous mixtures (Sapio); hypoxia chamber (Billups–Rothenberg). Methods: DIV8 neurons were subjected to oxygen and glucose deprivation (OGD). After medium replacement with a glucose-free balanced salt solution (ogR), cells were incubated for 3 hours in the chamber saturated with the gaseous mixture. The restoration solution (glucose and B27) was administrated to neurons (ogR) for one hour (R1h) or overnight (R16h). Results: We observed Tau and amyloid precursor protein (APP) post-translational and protein level modifications after OGD/ogR. Tau cleavage and secretion occurs at R1h through microvesicles (MVs) population, including LC3 and/or LAMP1 positive vesicles, marker of autophagy-mediated secretion (exophagy). In MVs extract we also identified α- and β- C-terminal fragments (CTFs) of APP, and the peptidyl prolyl cis/trans isomerase Pin1. Discussion: MVs represent an intercellular communication delivering multiple cargo with beneficial or harmful messages [1]. Thus, differences in MVs contents after OGD/ogR suggest that ischemia leads to a robust upheaval with the autophagic mechanisms activation resultant in neuron-to-neuron transfer of material with neurodegenerative potential. Conclusions: OGD treatment leads to mis-processed protein unconventional secretion also including exophagy pathway. These proteins represent seeds for protein aggregation according to the “prion-like” propagation mechanism [2] that may represent an early event in AD and proteinopathies.

In recent years particulate matter (PM) has been found to trigger several mechanisms responsible for neuroinflammation, and accumulating evidence points to neuroinflammation as an active participant in the progression of neurodegenerative diseases. This project aimed to assess whether, in a model of neuronal cells, PM, specifically ultrafine particles (UFP), induces oxidative stress and inflammation. We performed in-vitro analyses using HT22 cells, an immortalized mouse hippocampus cell line. HT22 cells have been treated for 3h and 24h with DEP SRM1650b (Standard Reference Material, National Institute of Standards and Technology, USA), a diesel particulate with aerodynamic diameter of 0,18 µm, suspended at different concentrations (10 µg/ml, 25 µg/ml, 50 µg/ml, 100 µg/ml) in culture medium. The following parameters have been measured: cell viability, oxidative stress related toxicity and activation of inflammation pathways. MTT viability assay demonstrated that DEP does not induce a significant cell death neither at 3h nor at 24h of treatment, for all the doses used. After 3h of DEP treatment, protein analysis revealed the presence of oxidative stress; in fact DEP induced a significant dose-dependent increase of HO-1 levels, a stress related protein which catalysed heme degradation. The increase of iNOS protein levels suggests the activation of inflammation pathway, although we need further studies to confirm it. Moreover, we observed an increase in Cyp1B1 levels, a cytochrome of the P450 superfamily involved in the activation of many xenobiotics and in polycyclic aromatic hydrocarbons metabolism, and in HSP70 levels, a protein activated in cellular stress conditions. After 24h of DEP treatment, analysis showed that HO-1 and HSP70 protein levels are still high, although the dose function increase is more pronounced at 3h, whereas Cyp1B1 and iNOS return almost to control levels. Based on the data so far available, we can hypothesize that exposure of neuronal cells to DEP results in the induction of oxidative stress and in the activation of inflammation pathway.

Objectives. To understand brain ischemia injury contribution to Alzheimer’s disease (AD) onset, looking for post-translational modifications, turnover alterations and AD hallmarks secretion such as tau and amyloid precursor protein (APP), associated to autophagy activation. Methods. Rat cultured hippocampal neurons are subjected to oxygen glucose deprivation (OGD) followed by normal culture condition restoration (R), mimicking an ischemia/reperfusion (I/R) event. Afterwards AD hallmarks and autophagy markers expression and secretion are analyzed in the cells lysate and in microvescicles (MVs) isolated from culture medium. Confocal microscopy is employed to evaluate the potential re- and co-localization of tau and autophagic markers after treatment. Results. In our cellular model subjected to OGD/R, we observed tau and APP post-translational and protein level modifications in parallel to the autophagic markers (LC3II and beclin-1) expression alterations. Moreover, OGD/R treatment elicits a time-dependent increase in tau and APP C-terminal fragments (CTF) secretion by means of a heterogeneous MV population, including a group of LC3II positive vescicles. Preliminary data obtained by confocal microscopy suggest that LC3 might undergo re-localization after OGD/R treatment. Conclusions. The obtained results suggest that OGD treatment leads to multiple unconventional secretion mechanisms for tau and APP CTFs, in which autophagy activation seems to partially outcome in the exophagy secretory pathway. Although exophagy might be a defensive mechanism to prevent intracellular accumulation of misfolded proteins under conventional autophagy alteration, the secretion of misprocessed proteins has been recently associated to dendritic degeneration and neuronal death becoming a possible link to Alzheimer’s disease.

Alzheimer's disease (AD) is a neurodegenerative illness affecting the elderly population, characterized by plaques of Aβ42 aggregates, neurofibrillary tangles and neuronal loss (Allsop, 2000). In AD vascular factors could precede the neurodegenerative process (de la Torre, 2002, 2008); Aβ42 accumulation in the cerebral capillary may be a consequence of a local production in the vascular domain (Natté et al., 1999). Air pollution has been associated with central nervous system (CNS) diseases. Inhaled UFPs (< 100 nm) could easily translocate cross the air-blood barrier, reach the bloodstream and be distributed to the cardiovascular system or the CNS (Oberdorster et al., 2002), thus affecting systemic microvasculature (Nurkiewicz et al., 2011). hCMEC/D3 and HT22 cells have been exposed to different DEP concentrations for different times. The following parameters have been measured: cell viability, oxidative stress and inflammation markers (HO-1, iNOS, Cyp1b1, COX-2, TNFalpha, IL1beta, IL8, VEGF), tight junction proteins (claudin-5, occludin), an amyloidogenic processing marker (BACE-1), besides an AD marker (Tau). In both cell lines, none of the concentrations induced cytotoxicity. In hCMEC/D3, DEP caused increases in HO-1, COX-2 and BACE-1 levels; moreover, the lower dose elicit a significant VEGF release. In HT22, after 3h all the concentrations caused an increase in HO-1, iNOS, HSP70 and Cyp1b1, whereas after 24h iNOS and Cyp1B1 return almost to control levels. Finally, after 24h a decrease in Tau levels has been found. In conclusion, all the parameters, except cytotoxicity, were differently affected in hCMEC/D3 and HT22, confirming the major susceptibility of neurons to toxic insults. Supported by Fondazione Cariplo.

In recent years particulate matter (PM) has been found to trigger several mechanisms responsible for neuroinflammation, an active participant in the progression of neurodegenerative diseases (Block and Calderón-Garcidueñas, 2009; Levesque et al., 2011). This project aimed to assess whether, in a model of neuronal cells, PM, specifically ultrafine particles (UFP), induces oxidative stress and inflammation. HT22 cells, an immortalized mouse hippocampus cell line, have been treated for 3h and 24h with DEP SRM1650b (Standard Reference Material, National Institute of Standards and Technology, USA), a diesel particulate with mean aerodynamic diameter of 0,18 µm, suspended at different concentrations (10 µg/ml, 25 µg/ml, 50 µg/ml, 100 µg/ml) in culture medium. The following parameters have been measured: cell viability, oxidative stress related toxicity and activation of inflammation pathways. MTT viability assay demonstrated that DEP does not induce a significant cell death neither at 3h nor at 24h of treatment, for all the doses used. After 3h of DEP treatment, protein analysis revealed the presence of oxidative stress; in fact DEP induced a significant dose-dependent increase of HO-1 levels, a stress related protein which catalyse heme degradation. The increase of iNOS protein levels suggests the activation of inflammation pathway. Moreover, we observed an increase in Cyp1B1 levels, a cytochrome of the P450 superfamily involved in polycyclic aromatic hydrocarbons metabolism, and in HSP70 levels, a protein activated in cellular stress conditions. After 24h of DEP treatment, analysis showed that HO-1 and HSP70 protein levels are still high, whereas Cyp1B1 and iNOS return almost to control levels. Indeed, we can hypothesize that exposure of neuronal cells to DEP results in oxidative stress and inflammation induction. In in-progress experiments, BALB/c mice will be submitted to intratracheal instillation of single or repeated doses of DEP, and effects on the SNC system (in particular on hippocampus, cortex and cerebellum) will be studied. Figure 1. Protein analyses after 3h of DEP treatment. * p