Combination of Coenzyme Q10 Intake and Moderate Physical Activity Counteracts Mitochondrial Dysfunctions in a SAMP8 Mouse Model.

Aging skeletal muscles are characterized by a progressive decline in muscle mass and muscular strength. Such muscular dysfunctions are usually associated with structural and functional alterations of skeletal muscle mitochondria. The senescence-accelerated mouse-prone 8 (SAMP8) model, characterized by premature aging and high degree of oxidative stress, was used to investigate whether a combined intervention with mild physical exercise and ubiquinol supplementation was able to improve mitochondrial function and preserve skeletal muscle health during aging. 5-month-old SAMP8 mice, in a presarcopenia phase, have been randomly divided into 4 groups (n = 10): untreated controls and mice treated for two months with either physical exercise (0.5 km/h, on a 5% inclination, for 30 min, 5/7 days per week), ubiquinol 10 (500 mg/kg/day), or a combination of exercise and ubiquinol. Two months of physical exercise significantly increased mitochondrial damage in the muscles of exercised mice when compared to controls. On the contrary, ubiquinol and physical exercise combination significantly improved the overall status of the skeletal muscle, preserving mitochondrial ultrastructure and limiting mitochondrial depolarization induced by physical exercise alone. Accordingly, combination treatment while promoting mitochondrial biogenesis lowered autophagy and caspase 3-dependent apoptosis. In conclusion, the present study shows that ubiquinol supplementation counteracts the deleterious effects of physical exercise-derived ROS improving mitochondrial functionality in an oxidative stress model, such as SAMP8 in the presarcopenia phase.

Modification of translation factor aIF5A from Sulfolobus solfataricus.

Eukaryotic eIF5A and its bacterial orthologue EF-P are translation elongation factors whose task is to rescue ribosomes from stalling during the synthesis of proteins bearing particular sequences such as polyproline stretches. Both proteins are characterized by unique post-translational modifications, hypusination and lysinylation, respectively, which are essential for their function. An orthologue is present in all Archaea but its function is poorly understood. Here, we show that aIF5A of the crenarchaeum Sulfolobus solfataricus is hypusinated and forms a stable complex with deoxyhypusine synthase, the first enzyme of the hypusination pathway. The recombinant enzyme is able to modify its substrate in vitro resulting in deoxyhypusinated aIF5A. Moreover, with the aim to identify the enzyme involved in the second modification step, i.e. hypusination, a set of proteins interacting with aIF5A was identified.

Is the adjuvanted influenza vaccine more effective than the trivalent inactivated vaccine in the elderly population? Results of a case-control study.

INTRODUCTION: Influenza illness is an important public health problem and annual vaccination is globally recommended for high risk populations. OBJECTIVE: The aim was to evaluate and compare the effectiveness of influenza vaccines in reducing hospitalizations for influenza or pneumonia during two influenza seasons in the elderly. METHODS: A case-control study was performed, using administrative database of the Local Health Unit Roma-A (LHU RM-A). The included subjects were at least 65 years old and residing in one of the four districts of the LHU. The cases were hospitalized for influenza or pneumonia during influenza season in the years 2010-2011 and 2011-2012. The controls were hospitalized in the same period, but not for influenza or pneumonia. The subjects were immunized with the trivalent inactivated influenza vaccine (TIV) in the first influenza season (2010-2011) and with the adjuvanted influenza vaccine MF59 (ATIV) in the second season (2011-2012). RESULTS: A total of 269 cases and 1247 controls were included for the 2010-2011 influenza season, and 365 cases and 1227 controls were selected for the 2011-2012 season. Up to 63.6% cases and 53.5% controls in the 2010-2011 season and 78.6% of cases and 64.1% of controls in the 2011-2012 season have not been vaccinated. Female gender and high educational level were protective factors for hospitalization. Subjects over 75 years were at high risk of hospitalization compared to 65-74 years olds. Influenza vaccination reduced significantly hospitalization in both seasons. In subjects with 65-74 years TIV was more effective than ATIV; vice versa for those over 75 years old. DISCUSSION AND CONCLUSION: TIV and ATIV reduce hospitalization for influenza or pneumonia with a variable degree of protection in different age groups. In particular, ATIV is more effective in individuals over 75 years old.

Action against vaccine-preventable infectious diseases and tuberculosis in Nomad Camps: the experience of a Local Health Unit in Rome.

In Italy the highest incidence of Tuberculosis (TB) cases is in young adult migrants. In 2011, the sanitarystaff of the Local Health Unit (ASL) Roma A promoted a vaccination campaign conducting several public health interventions in Nomad Camps. After notification of a case of TB in the Camp of Via Salaria, out of 357 Mantoux skin tests performed, 93 were positive (26%); subsequently, 5 subjects with radiographic positivity were hospitalized. The vaccination campaign was carried out to prevent the spread of infectious diseases in immigrant communities at high risk of contagion and to avoid the consequent transmission in the host country. As a result, vaccinations coverage among the residents of the Camps increased: 367 vaccinated subjects (30% more than previous year) and 612 administered vaccinations.

The liposome-protein corona in mice and humans and its implications for in vivo delivery.

As soon as nanomaterials, such as nanoparticles (NPs), are injected into a physiological environment a rich coating of biomolecules known as the "protein corona" rapidly covers them. This protein dress is the main factor, which affects the interaction of NPs with living systems. While the relationship between NP features and the biomolecule corona has been extensively investigated, whether and how changes in the physiological environment affect the NP-protein corona remains under-investigated. This is one of the most important steps in translating results in animal models to the clinic. Here we investigated thoroughly the biological identity of lipid NPs (size, charge, aggregation state and composition of the corona) after incubation with human plasma (HP) and mouse plasma (MP) by dynamic light scattering, micro-electrophoresis and nano-liquid chromatography tandem mass spectrometry (nanoLC/MS-MS). Specifically, we used two different liposomal formulations: the first one was made of polyethyleneglycol (PEG)-coated 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), while the second one was made of 30% of DOTAP, 50% of neutral saturated 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) and 20% cholesterol. The temporal evolution and complexity of the NP-protein corona was found to be strongly dependent on the biological environment. In MP, liposomes were more negatively charged, less enriched in opsonins and appreciably more enriched in apolipoproteins than their counterparts in HP. Collectively, our results suggest that the biological identities of NPs in mice and humans can be markedly different from each other. Relevance of results to in vivo applications is discussed.

Mechanistic understanding of gene delivery mediated by highly efficient multicomponent envelope-type nanoparticle systems.

We packaged condensed DNA/protamine particles in multicomponent envelope-type nanoparticle systems (MENS) combining different molar fractions of the cationic lipids 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and 3ß-[N-(N,N-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) and the zwitterionic lipids dioleoylphosphocholine (DOPC) and dioleoylphosphatidylethanolamine (DOPE). Dynamic light scattering (DLS) and microelectrophoresis allowed us to identify the cationic lipid/DNA charge ratio at which MENS are small sized and positively charged, while synchrotron small-angle X-ray scattering (SAXS) and atomic force microscopy (AFM) revealed that MENS are well-shaped DNA/protamine particles covered by a lipid monobilayer. Transfection efficiency (TE) experiments indicate that a nanoparticle formulation, termed MENS-3, was not cytotoxic and highly efficient to transfect Chinese hamster ovary (CHO) cells. To rationalize TE, we performed a quantitative investigation of cell uptake, intracellular trafficking, endosomal escape, and final fate by laser scanning confocal microscopy (LSCM). We found that fluid-phase macropinocytosis is the only endocytosis pathway used by MENS-3. Once taken up by the cell, complexes that are actively transported by microtubules frequently fuse with lysosomes, while purely diffusing systems do not. Indeed, spatiotemporal image correlation spectroscopy (STICS) clarified that MENS-3 mostly exploit diffusion to move in the cytosol of CHO cells, thus explaining the high TE levels observed. Also, MENS-3 exhibited a marked endosomal rupture ability resulting in extraordinary DNA release. The lipid-dependent and structure-dependent TE boost suggests that efficient transfection requires both the membrane-fusogenic activity of the nanocarrier envelope and the employment of lipid species with intrinsic endosomal rupture ability.

Screening of frailty in elderly patients with disability by the means of Marigliano-Cacciafesta polypathology scale (MCPS) and Canadian Study of Health and Aging (CSHA) scales.

Frailty is an age-related condition, characterized by a decreased homeostatic reserve and increased vulnerability to stressful events, with high risk of adverse outcomes. The aim of this study was to compare the evaluation of the frailty by the means of the MCPS and the Rockwood criteria. We enrolled 98 patients (mean age ± standard deviation, m ± SD, 80.7 ± 7.0 years) and 20 controls (82.7 ± 3.4 ys), who attended our outpatient clinic for the evaluation of disability and the renewal of driving license, respectively. The multidisciplinary geriatric assessment (MGA) was performed including the administration of the following scales for frailty: MCPS scale (range 0-245), CSHA-Rules-Based Definition of Frailty (CSHA-RBDF) (range 0-3) and CSHA-Clinical Frailty Scale (CSHA-CFS) (range 0-7). The patients and controls showed MCPS=52.39 ± 11.36 and 4.6 ± 3.28, CSHA-RBDF=2.27 ± 0.62 and 0.10 ± 0.44, CSHA-CFS=6.22 ± 0.75 and 2.95 ± 0.51, respectively (p<0.000001). Frailty scores were higher in female than in male (p=0.065 for CSHA-RDBF and p<0.05 for CSHA-CFS). The MCPS scores were significantly related to both CSHA-RDBF (r=0.753, p<0.001) and CSHA-CFS scores (r=0.793, p<0.001). The frailty scales were significantly related to disability, cognitive impairment and polypathology. In conclusion, the frail patient may be a carrier of multiple chronic pathologies and/or of physical/cognitive decline. The frail patient has to be considered the elective geriatric patient, characterized by a continuous multidimensional care requirement. MCPS is an useful tool for the frailty screening and to set up a tailored program of geriatric rehabilitation, in order to prevent or reduce the development of frailty-related complications.

Cadmium, lead, and chromium in large game: a local-scale exposure assessment for hunters consuming meat and liver of wild boar.

Heavy metals are ubiquitous in soil, water, and air. Their entrance into the food chain is an important environmental issue that entails risks to humans. Several reports indicate that game meat can be an important source of heavy metals, particularly because of the increasing consumption of game meat, mainly by hunters. We performed an exposure assessment of hunters and members of their households, both adults and children, who consumed wild boar (WB) meat and offal. We estimated the amount of cadmium, lead, and chromium in the tissues of WB hunted in six areas within Viterbo Province (Italy) and gathered data on WB meat and offal consumption by conducting specific diet surveys in the same areas. The exposure to cadmium, lead, and chromium was simulated with specifically developed Monte Carlo simulation models. Cadmium and lead levels in WB liver and meat harvested in Viterbo Province (Italy) were similar to or lower than the values reported in other studies. However, some samples contained these metals at levels greater then the EU limits set for domestic animals. The chromium content of meat or liver cannot be evaluated against any regulatory limit, but our results suggest that the amounts of this metal found in WB products may reflect a moderate environmental load. Our survey of the hunter population confirmed that their consumption of WB meat and liver was greater than that of the general Italian population. This level of consumption was comparable with other European studies. Consumption of WB products contributes significantly to cadmium and lead exposure of both adults and children. More specifically, consumption of the WB liver contributed significantly to total cadmium and lead exposure of members of the households of WB hunters. As a general rule, liver consumption should be kept to a minimum, especially for children living in these hunter households. The exposure to chromium estimated for this population of hunters may be considered to be safe. However, a specific and complete assessment of chromium speciation in relevant dietary and environmental situations should be conducted.

Role of temperature-independent lipoplex-cell membrane interactions in the efficiency boost of multicomponent lipoplexes.

Multicomponent lipoplexes have recently emerged as especially promising transfection candidates, as they are from 10 to 100 times more efficient than binary complexes usually employed for gene delivery purposes. Previously, we investigated a number of chemical-physical properties of DNA-lipid complexes that were proposed to affect transfection efficiency (TE) of lipoplexes, such as nanoscale structure, size, surface potential, DNA-protection ability and DNA release from complexes upon interaction with cellular lipids. Although some minor differences between multicomponent and binary lipoplexes were found, they did not correlate clearly with efficiency. Instead, here we show that a marked difference between the cell internalization mechanism of binary and multicomponent lipoplexes does exist. Multicomponent lipoplexes significantly transfect cells at 4 °C, when endocytosis does not take place suggesting that they can enter cells via a temperature-independent mechanism. Confocal fluorescence microscopy experiments showed the existence of a correlation between endosomal escape and TE. Multicomponent lipoplexes exhibited a distinctive ability of endosomal escape and release DNA into the nucleus, whereas, poorly efficient binary lipoplexes exhibited minor, if any, endosomal rupture ability and remained confined in perinuclear late endosomes. Stopped-flow mixing measurements showed that the fusion rates of multicomponent cationic liposomes with anionic vesicles, used as model systems of cell membranes, were definitely shorter than those of binary liposomes. As either lipoplex uptake and endosomal escape involve fusion between lipoplex and cellular membranes, we suggest that a mechanism of lipoplex-cellular membrane interaction, driven by lipid mixing between cationic and anionic cellular lipids, does explain the TE boost of multicomponent lipoplexes.

Evaluation of safety and efficacy of DNA vaccines against bovine herpesvirus-1 (BoHV-1) in calves.

Four DNA vaccines against BoHV-1 were evaluated for their efficacy in calves. Twelve animals were divided into four groups which were injected with four different DNA vaccines: pVAX-tgD (Vaccine A); pVAX-tgD co-immunised with pVAX-48CpG (Vaccine B); pVAX-UbiLacI-tgD-L (Vaccine C); pVAX-UbiLacI-tgD-L co-immunised with pVAX-48CpG (Vaccine D). Three additional calves were given the plasmid vector and served as controls. Ninety days after the first vaccination all calves were challenge infected with BoHV-1. All animals developed a severe form of infections bovine rhinotracheitis. Only the calves given the pVAX-tgD co-immunised with pVAX-48CpG (Vaccine B) developed humoral antibodies against BoHV-1 between 56 and 90 days after the first vaccination, whereas in calves of other groups and in the controls, antibodies appeared only after the infection. In the calves vaccinated with either pVAX-tgD (Vaccine A) or pVAX-tgD combined with pVAX-48CpG (Vaccine B), BoHV-1-specific IFN-γ secreting cells were detected in PBMCs 90 days after the first vaccination and their number increased after challenge exposure. In the other groups the IFN-γ secreting cells were detected after virus infection and at low values.

Self-administrated test based on the Marigliano-Cacciafesta Polypathological Scale (MCPS), as a screening tool for early identification of frailty in the elderly: a cohort study.

The aim of our study was to screen people of advanced age for "frailty", and illnesses, in order to prevent future disabilities through interventions made globally and individually to these patients. Applying the items such as those of the Marigliano-Cacciafesta Polypathological Scale (MCPS), we utilized a completely multi-dimensional evaluation. We have elaborated a series of simple and comprehensive questions enabling all participants to answer independently and easily. The results of the testing were successful. Within their limits, all the participants considered the test as a valuable instrument to assess their weaknesses. The test was especially efficient when it came to identifying problem areas in the psycho-physical state and frailty of elderly patients, as well as some loss of their own autonomy. These studies successfully tested a cohort of elderly people with similar symptoms.

New frontiers of cognitive rehabilitation in geriatric age: the Mozart Effect (ME).

The ME was described for the first time in 1993. Subsequently other studies with similar designs were performed. The present study, therefore, proposes: (i) to verify the existence of the benefits of exposure to music in elderly subjects with mild cognitive impairment (MCI), (ii) to explore whether it is possible to find any lasting improvement after training, conducted for a long period of time, with such musical pieces, in the measurable cognitive performances. The study we conducted showed that the ME is present in geriatric patients with MCI; the influence on spatial-temporal abilities remains constant in time if the stimulation is maintained. The continuation of our study will consist of increasing the number of individuals examined and in having them listen to music during the study of ECG rhythms and during the acquisition of cerebral functional magnetic resonance imaging (fMRI), and, at the same time, testing them by neuropsychometric methods.

A multi-DNA preventive vaccine for p53/Neu-driven cancer syndrome.

The highly aggressive cancer syndrome of female mice carrying a p53 knockout allele and a rat HER-2/neu (Neu) transgene (BALB-p53Neu) can be prevented by a cell vaccine presenting three components: Neu, interleukin (IL)-12 production, and allogeneic major histocompatibility complex (MHC) alleles (Triplex cell vaccine). Here we tested a second-generation Triplex DNA-based vaccine (Tri-DNA), consisting of the combination of three gene components (a transmembrane-extracellular domain fragment of the Neu gene, IL-12 genes, and the H-2D(q) allogeneic MHC gene), carried by separate plasmids. The Tri-DNA vaccine was at least as effective as the Triplex cell vaccine for cancer immunoprevention, giving a similar delay in the onset of mammary cancer and complete protection from salivary cancer. Both vaccines induced anti-Neu antibodies of the murine IgG2a isotype at similar levels. The Tri-DNA vaccine gave more restricted immunostimulation, consisting of a fully helper T cell type 1 (Th1)-polarized response, with effective production of interferon (IFN)-gamma in response to the vaccine but no spontaneous production, and no induction of anti-Neu IgG3 antibodies. On the other hand, the Triplex cell vaccine induced both Th1 and Th2 cytokines, a strong increase in spontaneous IFN-gamma production, and high levels of IgG3 antibodies recognizing Neu-positive syngeneic cells. In conclusion, the Tri-DNA vaccine is as effective as Triplex cell vaccine, exploiting a more restricted immune stimulation.

The evaluation of the presence of comorbidity by the Marigliano-Cacciafesta polypathology scale (MCPS) and the cumulative illness rating scale (CIRS) in elderly subjects with disability.

Multiple factors associated with the frailty syndrome may be involved in the appearance of disability, including the presence of comorbidity. The CIRS is commonly used for the evaluation of comorbidity, consisting of two parts: the comorbidity index (CI) and the severity index (SI). A multidimensional scale, the MCPS, has been recently developed, predicting the risk to develop disability. Fifty-nine subjects were examined by a structured multidimensional geriatric assessment. The MCPS and CIRS was significantly correlated (r=0.410; p<0.01 with the CI; and r=0.443, p<0.001 with the SI). The patients were divided in two groups, according to the MCPS score. The mean activities of daily living (ADL) and instrumental activities of daily living (IADL), as well as the corrected mini-mental state examination (MMSE) score (+/-S.E.M.) were: 3.19+/-0.26; 0.28+/-0.04 and 24.00+/-1.14 in moderate-severe polypathology (n=21); 2.16+/-0.22; 0.13+/-0.02 and 21.23+/-0.72 in severe polypathology (n=38) (p<0.001, p<0.01 and p<0.05), respectively. The MCPS score was correlated with the main indices of disability. In conclusion, we found that the MCPS is a useful tool in order to quantify and classify the presence of comorbidity, with results significantly related to that obtained with the CIRS. The MCPS offers an important stratification of the patients on the base of a well-established classification, not supplied by the CIRS.

Exaggerated morning blood pressure surge and cardiovascular events. A 5-year longitudinal study in normotensive and well-controlled hypertensive elderly.

Cardiovascular events (CE) occur most frequently in the morning hours in hypertensive subjects. We studied the association between the morning blood pressure (BP) surge and CE in prognosis of 10 normotensive and 32 well-controlled hypertensive elderly, in whom ambulatory BP monitoring was performed and who were followed prospectively for 5 years. The morning surge (MS) of BP was calculated as mean systolic BP during 2h after awakening--mean systolic BP during 1h that included the lowest sleep BP. During an average of 60 months, five CE occurred. When the patients were divided into two groups according to MS, those in the top terzile (MS group; MS> or =34 mmHg, n=14) had a higher prevalence of CE (5 versus 0, p=0.001) during the follow-up period, than the others (non-MS group; MS<34 mmHg, n=28). The logistic regression analysis showed the MS sleep-trough surge as predictive variable of CE (odds ratio, OR=0.794, p=0.022). In conclusion, in older normotensives and well-controlled hypertensives, a higher BP MS is associated with vascular risk independently of clinical and ambulatory BP. Reduction of the MS could thus be a therapeutic target for preventing vascular events also in non-hypertensive patients.

The Marigliano-Cacciafesta polypathology scale (MCPS): a tool for predicting the risk of developing disability.

The purpose of the work was to verify whether our MCPS can be a tool for predicting the risk of developing disabilities. We considered 45 elderly subjects divided into three groups of 15 subjects each. Group 1 consisted of cases with a "moderate-severe" degree of polypathology, with no associated condition of disability evaluated by means of the activities of daily living (ADL). Group 2 contained cases with a "moderate" degree of polypathology (with no associated condition of ADL disability). The Group 3 was the control group with a "mild" degree of polypathology (with no disability associated with ADL). All subjects were re-evaluated after 6 and 12 months. Both Groups 1 and 2 of cases over time developed greater disabilities, compared to the control Group 3; in particular, the subjects with "moderate-severe" polypathology were more disabled after 12 months.

CCL16 enhances the CD8+ and CD4+ T cell reactivity to human HER-2 elicited by dendritic cells loaded with rat ortholog HER-2.

T cells from HLA-A2+ healthy donors were co-cultured with autologous dendritic cells (DC) loaded with apoptotic tumor cells expressing rat neu, and were induced to mature by tumor necrosis factor (TNF)alpha and interleukin (IL)-1beta (mDC(neu)) or by the CCL16 chemokine (CCL16/mDC(neu)). Priming by CCL16/mDC(neu) induces a larger population of T cells that express cytoplasmatic interferon (IFN)gamma, TNFalpha, perforin and granzyme B compared to those primed by mDC(neu). T cells primed by CCL16/mDC(neu) release IFNgamma in response to human HER-2+ cells and kill human HER-2+ target cells more efficiently than those primed by mDC(neu). Our results show that both the loading of DC with xenogeneic rat neu and their maturation by CCL16 are two issues of critical importance for the elicitation of an effective response to human HER-2 in T cells from normal donors.

DNA immunization using constant-current electroporation affords long-term protection from autochthonous mammary carcinomas in cancer-prone transgenic mice.

A recently developed, adaptive constant-current electroporation technique was used to immunize mice with an intramuscular injection of plasmid coding for the extracellular and transmembrane domains of the product of the rat neu(664V-E) oncogene protein. In wild-type BALB/c mice, plasmid electroporation at lower current settings elicits higher antibody titers, a strong cytotoxic response and completely protects all mice vaccinated with 10, 25 and 50 microg of plasmid against a lethal challenge of rat neu+ carcinoma cells. BALB/c mice transgenic for the transforming rat neu(664V-E) (ErbB-2, Her-2/neu) oncogene (BALB-neuT(664V-E)) develop an invasive mammary gland carcinoma by 20 weeks of age. Remarkably, when transgenic BALB-neuT(664V-E) mice were vaccinated at a 10- week interval with 50 microg of plasmid with 0.2 A electroporation, mice remained tumor free for more than a year. A single administration of plasmid associated with electroporation was enough to markedly delay carcinogenesis progression in mice with multiple microscopic invasive carcinomas, and keep about 50% of mice tumor free at one year of age. Thus, vaccination using a clinically relevant dose of plasmid encoding the extracellular and transmembrane domains of the neu oncogene delivered by electroporation prevents long-term tumor formation. These improvements in the efficacy of this cancer vaccine regimen vastly increase its chances for clinical success.

The Marigliano-Cacciafesta polypathological scale: a tool for assessing fragility.

The aim of our studies was to establish a standard method of assessment that allows an early identification of frailty in the elderly, i.e., to predict who are at risk of developing disabilities, in order to be able to intervene with preventive global and individualized measures. A new multidimensional scale called Marigliano-Cacciafesta polypathological scale (MCPS) was used on 180 elderly people, together with the Barthel index (BI), the global evaluation functional index (GEFI), the geriatric depression scale (GDS), the mini mental state examination (MMSE), the mini nutritional assessment (MNA), and the Tinetti test. A strongly significant statistical correlation was found between the MCPS and the nutritional state, mood level, motor functionality, level of disability and global functionality. As the fragile patients are at a risk to develop disabilities, we think that our scale can be a significant contribution to the multidimensional geriatric assessment (MGA), aimed at identifying and quantifying the parameter of fragility of each patient, an information which should be known, if we intend to introduce preventive measures.