Extended Cleavage Specificity of two Hematopoietic Serine Proteases from a Ray-Finned Fish, the Spotted Gar (Lepisosteus oculatus).

The extended cleavage specificities of two hematopoietic serine proteases originating from the ray-finned fish, the spotted gar (Lepisosteus oculatus), have been characterized using substrate phage display. The preference for particular amino acids at and surrounding the cleavage site was further validated using a panel of recombinant substrates. For one of the enzymes, the gar granzyme G, a strict preference for the aromatic amino acid Tyr was observed at the cleavable P1 position. Using a set of recombinant substrates showed that the gar granzyme G had a high selectivity for Tyr but a lower activity for cleaving after Phe but not after Trp. Instead, the second enzyme, gar DDN1, showed a high preference for Leu in the P1 position of substrates. This latter enzyme also showed a high preference for Pro in the P2 position and Arg in both P4 and P5 positions. The selectivity for the two Arg residues in positions P4 and P5 suggests a highly specific substrate selectivity of this enzyme. The screening of the gar proteome with the consensus sequences obtained by substrate phage display for these two proteases resulted in a very diverse set of potential targets. Due to this diversity, a clear candidate for a specific immune function of these two enzymes cannot yet be identified. Antisera developed against the recombinant gar enzymes were used to study their tissue distribution. Tissue sections from juvenile fish showed the expression of both proteases in cells in Peyer's patch-like structures in the intestinal region, indicating they may be expressed in T or NK cells. However, due to the lack of antibodies to specific surface markers in the gar, it has not been possible to specify the exact cellular origin. A marked difference in abundance was observed for the two proteases where gar DDN1 was expressed at higher levels than gar granzyme G. However, both appear to be expressed in the same or similar cells, having a lymphocyte-like appearance.

First-principles predictions for shear viscosity of air components at high temperature.

The direct molecular simulation (DMS) method is used to obtain shear viscosity data for non-reacting air and its components by simulating isothermal, plane Poiseuille subsonic flows. Shear viscosity is estimated at several temperatures, from 273 K to 10 000 K, by fitting the DMS velocity profiles using the analytic solution of the Navier-Stokes equations for this simple canonical flow. The ab initio potential energy surfaces (PESs) that describe the various atomic-level interactions are the only input in the simulations. Molecules involved in a collision within the flow can occupy any rovibrational state that is allowed by the effective diatomic potential. For molecular nitrogen, oxygen, and air at standard condition molar composition, the DMS shear viscosity predictions are in excellent agreement with the experimental data that are available up to about 2000 K. The results for pure molecular nitrogen and pure molecular oxygen also agree very well with previously published quasi-classical trajectory (QCT) calculations based on the same PESs. It is further shown that the ab initio shear viscosity data are generally lower than the corresponding values used in popular computational fluid dynamics codes, over a wide temperature range. Finally, Wilke's mixing rule is demonstrated to accurately predict the DMS air viscosity results from the pure molecular components data up to 4000 K.

Comparison of potential energy surfaces and computed rate coefficients for N2 dissociation.

Comparisons are made between potential energy surfaces (PES) for N2 + N and N2 + N2 collisions and between rate coefficients for N2 dissociation that were computed using the quasiclassical trajectory method (QCT) on these PESs. For N2 + N we compare the Laganà's empirical LEPS surface with one from NASA Ames Research Center based on ab initio quantum chemistry calculations. For N2 + N2 we compare two ab initio PESs (from NASA Ames and from the University of Minnesota). These use different methods for computing the ground state electronic energy for N4, but give similar results. Thermal N2 dissociation rate coefficients, for the 10,000K-30,000K temperature range, have been computed using each PES and the results are in excellent agreement. Quasi-stationary state (QSS) rate coefficients using both PESs have been computed at these temperatures using the Direct Molecular Simulation of Schwartzentruber and coworkers. The QSS rate coefficients are up to a factor of 5 lower than the thermal ones and the thermal and QSS values bracket the results of shock-tube experiments. We conclude that the combination of ab initio quantum chemistry PESs and QCT calculations provides an attractive approach for the determination of accurate high-temperature rate coefficients for use in aerothermodynamics modeling.

An improved potential energy surface and multi-temperature quasiclassical trajectory calculations of N2 + N2 dissociation reactions.

Accurate modeling of high-temperature hypersonic flows in the atmosphere requires consideration of collision-induced dissociation of molecular species and energy transfer between the translational and internal modes of the gas molecules. Here, we describe a study of the N2 + N2⟶N2 + 2N and N2 + N2⟶4N nitrogen dissociation reactions using the quasiclassical trajectory (QCT) method. The simulations used a new potential energy surface for the N4 system; the surface is an improved version of one that was presented previously. In the QCT calculations, initial conditions were determined based on a two-temperature model that approximately separates the translational-rotational temperature from the vibrational temperature of the N2 diatoms. Five values from 8000 K to 30,000 K were considered for each of the two temperatures. Over 2.4 × 10(9) trajectories were calculated. We present results for ensemble-averaged dissociation rate constants as functions of the translational-rotational temperature T and the vibrational temperature T(v). The rate constant depends more strongly on T when T(v) is low, and it depends more strongly on T(v) when T is low. Quasibound reactant states contribute significantly to the rate constants, as do exchange processes at higher temperatures. We discuss two sets of runs in detail: an equilibrium test set in which T = T(v) and a nonequilibrium test set in which T(v) < T. In the equilibrium test set, high-v and moderately-low-j molecules contribute most significantly to the overall dissociation rate, and this state specificity becomes stronger as the temperature decreases. Dissociating trajectories tend to result in a major loss of vibrational energy and a minor loss of rotational energy. In the nonequilibrium test set, as T(v) decreases while T is fixed, higher-j molecules contribute more significantly to the dissociation rate, dissociating trajectories tend to result in a greater rotational energy loss, and the dissociation probability's dependence on v weakens. In this way, as T(v) decreases, rotational energy appears to compensate for the decline in average vibrational energy in promoting dissociation. In both the equilibrium and nonequilibrium test sets, in every case, the average total internal energy loss in the dissociating trajectories is between 10.2 and 11.0 eV, slightly larger than the equilibrium potential energy change of N2 dissociation.

Complete Percutaneous Revascularization in Patients Aged ≥85 Years With Acute Coronary Syndrome and Multivessel Coronary Artery Disease.

Multivessel coronary disease is frequent in older patients who underwent coronary angiography for acute coronary syndrome. Whether a complete revascularization or a culprit-only approach is preferable in these patients is still debated. We included consecutive patients aged ≥85 years, presenting with acute coronary syndrome and showing multivessel coronary disease at coronary angiography. Patients were grouped according to complete (residual SYNTAX score [RSS] 0 to 8) or incomplete (RSS >8) revascularization. Primary end point was the rate of major adverse cardiovascular events (MACEs, the composite of cardiovascular death, re-myocardial infarction [re-MI], clinically driven percutaneous coronary intervention, and rehospitalization because of cardiac disease) at 2 years follow-up. A total of 166 patients met the criteria for enrollment; 108 patients had a final RSS 0 to 8 (complete revascularization) and 58 patients had a final RSS >8 (incomplete revascularization). The rate of MACE was reduced in patients who underwent complete revascularization (35.2% vs 51.7%, p = 0.039, adjusted hazard ratio 0.60, 95% confidence interval 0.37 to 0.98, p = 0.04), a difference mainly driven by a reduction in re-MI (8.3% vs 19.0%, p = 0.045), clinically driven percutaneous coronary intervention (2.8% vs 19.0%, p <0.001), and rehospitalization for cardiac disease (9.3% vs 24.1%, p = 0.009). Other independent predictors of MACE were active malignancy, previous MI, left ventricle ejection fraction <35% (increasing risk of events), and radial access (reducing risk of events). In conclusion, in patients aged ≥85 years, a complete revascularization is associated with a better prognosis, especially in terms of nonfatal events.

Recovery, Molecular Characterization, and Ampelographic Assessment of Marginal Grapevine Germplasm from Southern Umbria (Central Italy).

The protection of grapevine biodiversity and the safeguarding of genetic variability are certainly primary and topical objectives for wine research, especially in territories historically devoted to viticulture. To assess the autochthonous germplasm of three different districts of Southern Umbria (Central Italy), the plant material of 70 grapevines retrieved from reforested land plots or old vineyards was collected, and their genetic identity was investigated using 13 microsatellite markers (SSR). The results revealed the presence of 39 unique genotypes, divided into 24 already-known cultivars and 15 never-reported SSR profiles. Most of the grapevine accessions were then vegetatively propagated and cultivated in a vineyard collection both to be protected from extinction and to be evaluated at the ampelographic level. Overall, this work emphasizes the need for recovering the threatened genetic variability that characterizes minor neglected grapevine cultivars or biotypes of Southern Umbria germplasm, and the requirement to revalue and exploit the more valuable genetic resources to enhance the local agri-food economy.

Molecular dynamics simulation of O2 sticking on Pt(111) using the ab initio based ReaxFF reactive force field.

The molecular dynamics technique with the ab initio based classical reactive force field ReaxFF is used to study the adsorption dynamics of O(2) on Pt(111) for both normal and oblique impacts. Overall, good quantitative agreement with the experimental data is found at low incident energies. Specifically, our simulations reproduce the characteristic minimum of the trapping probability at kinetic incident energies around 0.1 eV. This feature is determined by the presence of a physisorption well in the ReaxFF potential energy surface (PES) and the progressive suppression of a steering mechanism when increasing the translational kinetic energy (or the molecule's rotational energy) because of steric hindrance. In the energy range between 0.1 and 0.4 eV, the sticking probability increases, similar to molecular beam sticking data. For very energetic impacts (above 0.4 eV), ReaxFF predicts sticking probabilities lower than experimental sticking data by almost a factor of 3 due to an overall less attractive ReaxFF PES compared to experiments and density functional theory. For oblique impacts, the trapping probability is reduced by the nonzero parallel momentum because of the PES corrugation and does not scale with the total incident kinetic energy. Furthermore, our simulations predict quasispecular (slightly supraspecular) distributions of angles of reflection, in accordance with molecular beam experiments. Increasing the beam energy (between 1.2 and 1.7 eV) causes the angular distributions to broaden and to exhibit a tail toward the surface normal because molecules have enough momentum to get very near the surface and thus probe more corrugated repulsive regions of the PES.

Constructing feed-forward artificial neural networks to fit potential energy surfaces for molecular simulation of high-temperature gas flows.

Kinetic rates for thermochemical nonequilibrium models are generally computed from quasiclassical trajectory (QCT) calculations on accurate ab initio potential energy surfaces (PES). In this article, we use a feed-forward artificial neural network (ANN) to fit existing single-point energies for N_{2}+N_{2} interactions [Bender et al., J. Chem. Phys. 143, 054304 (2015)JCPSA60021-960610.1063/1.4927571] to construct a PES suitable for molecular simulation of high-temperature gas flows. We then perform detailed comparisons with a widely used N_{4} PES that was built using the permutation invariant polynomials (PIP) method. Specific physical considerations in the construction of the ANN for this application are detailed. Translation, rotation, and permutation invariance are precisely satisfied by mapping the interatomic distances onto a set of permutation invariant inputs, known as fundamental invariants (FI) that generate the permutation invariant polynomial ring. The diatomic energy is imposed by decomposing the total potential energy into a sum of a two-body and a many-body energy contribution. To obtain the correct dynamical behavior with the most basic, yet computationally efficient ANN, spurious long-distance interactions must be removed to avoid incorrect physical behavior at the dissociation threshold. We use a simple apodization function to smoothly taper off to zero any residual many-body interaction at large separations. Both accuracy and performance of the FI-ANN PES are assessed. QCT calculations are used to compute dissociation probabilities and vibrational energy distributions at various equilibrium temperatures. Excellent agreement with the results obtained from the PIP PES is found. For our test case, the ANN PES is also significantly more computationally efficient than the PIP PES at comparable root-mean-square error levels.

Active and Healthy Ageing as a Wicked Problem: The Contribution of a Multidisciplinary Research University.

The quest for an active and healthy ageing can be considered a "wicked problem." It is a social and cultural problem, which is difficult to solve because of incomplete, changing, and contradictory requirements. These problems are tough to manage because of their social complexity. They are a group of linked problems embedded in the structure of the communities in which they occur. First, they require the knowledge of the social and cultural context in which they occur. They can be solved only by understanding of what people do and why they do it. Second, they require a multidisciplinary approach. Wicked problems can have different solutions, so it is critical to capture the full range of possibilities and interpretations. Thus, we suggest that Università Cattolica del Sacro Cuore (UCSC) is well suited for accepting and managing this challenge because of its applied research orientation, multidisciplinary approach, and integrated vision. After presenting the research activity of UCSC, we describe a possible "systems thinking" strategy to consider the complexity and interdependence of active ageing and healthy living.

[Management of acute coronary syndromes in elderly patients: a single-center experience]. / La gestione della sindrome coronarica acuta nell'anziano: l'esperienza di un singolo centro.

Demographic studies show an ever increasing number of subjects >65 years among the Italian population. In particular, subjects aged >75 years represent nowadays 9% of the Italian population and are expected to exceed 14% by 2030. In the district of Crema (Italy), subjects aged >75 years are 16 000, and are expected to reach 25 000 by 2030. This phenomenon will result in an increasing number of elderly patients with acute coronary syndromes (ACS) admitted to our hospital. It is therefore necessary to define appropriate guidelines for clinical management of this subset of patients, in order to pursue a reduction in mortality rates and rehospitalizations, while maintaining an acceptable quality of life. These strategies are not adequately supported by international guidelines or randomized studies, where advanced age often represents an exclusion criterion. In our Cardiology Unit, hemodynamic evaluation is performed in all patients >75 years presenting with ST-elevation myocardial infarction (STEMI), ACS, positive troponin I, recurrent episodes of heart failure associated with clinical and instrumental suspicion of ischemia, or in survivors of cardiac arrest without neurological deficit. In all patients, presence of comorbidities is also evaluated, in particular renal insufficiency, disabling cerebrovascular disease, and residual quality of life. From February 2011 to February 2012, 974 coronary angiographies and 692 coronary angioplasties (PTCA) were performed. Of these, 194 procedures were performed in patients ≥75 years, the remaining in younger patients. The reason for hospitalization was ACS in 71% of elderly patients versus 53% in younger patients. In elderly patients, drug-eluting stents were used in 30% of the procedures. PTCA was successful in 98% of cases. Main complications included major bleeding (0.8%), access site-related bleeding (all femoral) (2%), and arterial rupture (n = 1). From a technical viewpoint, the radial access was predominantly used (78%), single plain old balloon angioplasty was performed in 12% of the procedures, with tirofiban administration in 42% of cases. Periprocedural and in-hospital mortality was 5%, mainly occurring in patients with STEMI and/or diffuse coronary artery disease. During follow-up, 80% of patients achieved a fairly good quality of life; 15% of patients required rehospitalization for cardiovascular reasons. In conclusion, patient's age should not be regarded as the main key factor for decision-making when evaluating different clinical approaches to patients with ACS.

Italian Euromelanoma Day Screening Campaign (2005-2007) and the planning of melanoma screening strategies.

Although no study has definitively shown that unfocused screening of skin cancer is effective, many campaigns have been organized with the aim of increasing awareness on melanoma risk factors. The objective of this study was to analyse the results of the Skin Cancer Screening Day in Italy during the period 2005-2007, to determine the priorities for melanoma control plans in a Mediterranean country. A total of 5002 patients were screened by dermatologists in 31 cities. Individuals who considered themselves to have many naevi and those with a family history of melanoma showed a higher number of common and atypical naevi. Ten melanomas, 20 basal cell carcinomas and two squamous cell carcinomas were histopathologically confirmed. Our observations provide the following suggestions for melanoma prevention strategies: (a) an unfocused campaign is suitable to inform the public about the importance of self-examination of the skin, but is not useful to identify a larger number of melanomas; and (b) melanoma screening campaigns should focus on a selected population, which meets rigorous risk criteria to maintain higher cost-effectiveness. The financial support to effective melanoma screening programmes could be increased, especially in southern populations where lower levels of self-surveillance and socioeconomic conditions represent risk factors for late identification of melanoma.

[Coronary angioplasty of anomalous coronary arteries]. / L'angioplastica coronarica nelle anomalie coronariche complesse.

BACKGROUND: The aim of this study was to evaluate the incidence of coronary artery anomalies in a homogeneous population undergoing coronary angiography, and to assess acute and follow-up results of coronary angioplasty (PTCA). METHODS: From September 2001 to May 2010, 23 complex coronary artery anomalies were observed among 6300 patients undergoing coronary angiography (0.36%) at our cath lab. Mean age of the study population (20 males, 3 females) was 61.8 years (range 35-79 years). Seventeen patients presented with acute coronary syndrome associated with severe coronary stenosis, and 6 patients showed angina and ventricular arrhythmias associated with inducible ischemia on stress testing and no coronary artery lesions. PTCA was performed in 16 patients, 9 of whom with an anomalous origin of the coronary artery. RESULTS: The most frequent coronary artery anomaly was the anomalous origin of the right coronary artery from the left coronary cusp, running between the aorta and the pulmonary artery (n = 10, 8 males, 2 females; PTCA of the anomalous coronary artery performed in 4 cases). Eight patients showed an anomalous origin of the circumflex artery arising from the right coronary artery with a retroaortic course (PTCA of the anomalous coronary artery performed in 4 cases), and 2 patients (1 male, 1 female) an anomalous origin of the left main coronary artery from the right coronary cusp with a septal course. The remaining 3 patients showed an isolated anomalous origin of the left coronary artery from the right coronary artery with interarterial course, the left anterior descending artery from the right coronary artery, and the left anterior descending artery from the right coronary cusp, respectively. CONCLUSIONS: PTCA proved to be effective in the treatment of coronary artery anomalies showing no procedure-related complications and stable results over time. The high number of coronary artery anomalies in our study patients may help estimating the actual incidence of these anomalies in the Italian population by providing a rationale for implementing a national registry.

Right coronary artery aneurysm: percutaneous treatment with graft-coated stent during the acute phase of myocardial infarction.

We report a case of acute myocardial infarction due to acute thrombosis of the right coronary artery just before a large atherosclerotic aneurysm. The patient was treated with primary percutaneous coronary angioplasty (PCA) and deployment of graft-coated stent with optimal final result. Patients with atherosclerotic coronary aneurysms usually show the same cardiovascular risk factors and the same clinical presentation of patients with atherosclerotic obstructive coronary artery disease, but with an increased risk of endovascular thrombosis and consequently more frequent episodes of distal coronary embolism. Furthermore, they may develop other specific complications, such as rapid aneurysm enlargement and rupture leading to cardiac tamponade. In conclusion, our report shows that percutaneous approach to coronary aneurysms with exclusion of aneurismal lumen by placement of graft-coated stent is a feasible and safety procedure even during the acute phase of myocardial infarction, and it may probably reduce the risk of subsequent distal embolization, improving myocardial perfusion.

[24-hour primary angioplasty service for acute myocardial infarction at the Crema Hospital: considerations after one year of experience]. / L'angioplastica primaria 24/24 ore nell'infarto miocardico acuto a Crema: considerazioni dopo un anno di esperienza.

The aim of this contribution is to draw considerations on the first year of activity of a cath lab on call 24/24 h for primary coronary angioplasty (PTCA) (April 2007-April 2008) of a small hospital without surgical backup. We performed 152 primary PTCA in 156 patients presenting to the emergency room with ST-elevation myocardial infarction. The mean time from arrival to the emergency room to the insertion of the arterial introducer was 48 min. Patient management included a 12-lead ECG and family history, the telephone call to the hospital cardiologist and then to the cath lab operator and nurse. Procedural success was obtained in 97% of cases. Mortality was 6%. Other complications occurred in 4% of patients. The no-reflow phenomenon occurred in 4% of patients. We used 1.9 stents/patient. Direct stenting was performed in 43% of cases. In 14% of lesions, the procedure was concluded with only balloon angioplasty. The radial access was used in 15% of patients, whereas the femoral access was used in the remainder. Drug-eluting stents were implanted in 1% only of patients with acute myocardial infarction. Glycoprotein IIb/IIIa inhibitors and clopidogrel were extensively used prior to the procedure. Our data, although obtained in a smaller population sample and referring to a relatively short period of time, are similar to those of the American registry on PTCA. In conclusion, the results obtained in our center with the invasive management of acute myocardial infarction appear consistent with those of other centers and encourage us to continue along our chosen path.

[Outsourced service management of the catheterization laboratory in the cardiology department: lights and shadows]. / La gestione in service del laboratorio di elettrofisiologia ed emodinamica: luci ed ombre.

BACKGROUND: The aim of this study was to compare the economic impact and results achieved by recourse to outsourced management of the procedures carried out in the electrophysiology and catheterization laboratory of the Department of Cardiology of the Crema Hospital with the in-house setting up and operation of the same activities. METHODS: The comparison between the two possible options, "make" or "buy in", was made using the methodology of advanced direct costing, which provides for the allocation of only direct fixed and variable costs to clinical procedures, the subject of calculation. In addition to the financial evaluation, the quality variables showing the advantages and limitations of outsourcing in terms of organizational improvements, streamlining of the organizational structure, operational efficiency and improvement of the quality of service, were examined. RESULTS: The financial evaluation from 2002 to 2006 came out in favor of "make" as opposed to "buy in". Income derived from diagnosis-related-group payments for the more than 4000 procedures carried out was Euro26.239.034,96. On the basis of the economical evaluation the second contribution margin was slightly inferior with the "buy in" than with the "make" hypothesis. Specifically, it is Euro16.397.669,96 in the "buy in" and Euro16.753.579,16 in the "make" hypothesis, with a difference of Euro355.909,20 (-2%). CONCLUSIONS: The economic advantage lies with the "make" alternative compared with "buy in", nevertheless, outsourcing offers greater operational efficiency, better cost control, setting up of the laboratory within a very short time, simplified administration (single point of contact) and an opportunity to concentrate on core business. However, there are limitations due to greater dependence on the supplier, not all the equipment provided for under the contract was used, and loss of management know-how in non-core business areas.