Metabolomic profile of patients with left ventricular assist devices: a pilot study

Background: Metabolomic profiling has important diagnostic and prognostic value in heart failure (HF). We investigated whether left ventricular assist device (LVAD) support has an impact on the metabolomic profile of chronic HF patients and if specific metabolic patterns are associated with the development of adverse events. Methods: We applied untargeted metabolomics to detect and analyze molecules such as amino acids, sugars, fatty acids and other metabolites in plasma samples collected from thirty-three patients implanted with a continuous-flow LVAD. Data were analyzed at baseline, i.e., before implantation of the LVAD, and at long-term follow-up. Results: Our results reveal significant changes in the metabolomic profile after LVAD implant compared to baseline. In detail, we observed a pre-implant reduction in amino acid metabolism (aminoacyl-tRNA biosynthesis) and increased galactose metabolism, which reversed over the course of support [median follow-up 187 days (63–334 days)]. These changes were associated with improved patient functional capacity driven by LVAD therapy, according to NYHA functional classification of HF (NYHA class I-II: pre-implant =0% of the patients; post-implant =97% of the patients; P<0.001). Moreover, patients who developed adverse thromboembolic events (n=4, 13%) showed a pre-operative metabolomic fingerprint mainly associated with alterations of fatty acid biosynthesis and mitochondrial beta-oxidation of short-chain saturated fatty acids. Conclusions: Our data provide preliminary evidence that LVAD therapy is associated with changes in the metabolomic profile of HF and suggest the potential use of metabolomics as a new tool to stratify LVAD patients in regard to the risk of adverse events

The comparison of the proteomic profile of periodontal pocket and of corresponding gingival crevicular fluid to study periodontal disease biomarkers: feasibility study. biomarkers: feasibility study

Aim: Periodontitis is a set of inflammatory disorders characterized by periodontal attachment loss by periodontal pocket development, leading to tooth loss if remain untreated. The etiology and progress of periodontal disease is complex and remains mostly unknown. So, periodontal disease therapy has considerable limitations. The easy, reliable and correct early detection and control of the disease, markedly reduces biological and social costs. However, the diagnosis of periodontitis is established exclusively by clinical criteria based on probing to assess periodontal pockets, which are the pathognomonic expression of periodontal disease. The -omic sciences acquired substantial significance of late years and, in particular, proteomic seemed to be the more promising in this initial stage. Most proteomic analysis regarding periodontal diseases have been performed on saliva, crevicular fluid samples, peripheral blood or periodontal plaque samples which are more easily to harvest than the tissue of the periodontal pocket. However, they failed to provide reliable results for clinical applications. On the contrary, very few studies were directly performed on the periodontal pocket. So, the aim of this study was to compare the proteomic profile of interproximal pocket tissues with that of GCF, and to analyze if they show a significant similarity in the proteomic profile. Methods: in this preliminary study, we enrolled 3 healthy subjects affected by severe periodontitis needing of periodontal surgery. Immediately before the surgery, GCF samples were taken by means of filter paper strips positioned in the gingival sulcus correspondent to periodontal pockets. Then, periodontal pocket tissue, harvested during surgery, was adequately stored for proteomic analyses. All samples were immediately frozen at \u201380\ub0C and maintained until further analysis. Tissue samples were mechanically disrupted and incubated in lysis buffer, while GCF was obtained incubating the collecting paper in phosphate buffered. In both cases, after centrifugation, the supernatant was precipitated in cold acetone overnight and protein content were pelleted by centrifugation and then dissolved in a rehydration buffer. Mono-dimensional gel electrophoresis was used to separate protein content. After staining gel images were acquired and compared. Liquid chromatography coupled to mass spectrometry (LC-MS/MS) analysis was performed to allow protein spot identification. Results: 1-DE gels from periodontal pocket tissue and the correspondent GCF was analyzed by software Quantity One. Almost the same qualitative protein expression profile in pocket tissue and GCF was found from each patient. However, no statistical significant correlation between the quantitative proteomic profile of pocket tissue and GCF was found. Only one band (that of K immunoglobulin) resulted statistically significant between GCF and pocket tissue proteome in all patients. Conclusions To date, this is the first study comparing the proteome of periodontal pocket tissue and corresponding GCF. The periodontal pocket and the GCF are similar as proteomic networks, but the protein network of the periodontal pocket does not influence significantly the GCF protein network and the other way around. So, with the limitations of this study, the preliminary results seem to indicate that the GCF does not seem suitable to study on the pathogenesis of periodontal disease explaining the reason for the failure of studies based only on GCF to control the periodontal disease in real-time

Monophasic and Biphasic Electrical Stimulation Induces a Precardiac Differentiation in Progenitor Cells Isolated from Human Heart

Electrical stimulation (ES) of cells has been shown to induce a variety of responses, such as cytoskeleton rearrangements, migration, proliferation, and differentiation. In this study, we have investigated whether monophasic and biphasic pulsed ES could exert any effect on the proliferation and differentiation of human cardiac progenitor cells (hCPCs) isolated from human heart fragments. Cells were cultured under continuous exposure to monophasic or biphasic ES with fixed cycles for 1 or 3 days. Results indicate that neither stimulation protocol affected cell viability, while the cell shape became more elongated and reoriented more perpendicular to the electric field direction. Moreover, the biphasic ES clearly induced the upregulation of early cardiac transcription factors, MEF2D, GATA-4, and Nkx2.5, as well as the de novo expression of the late cardiac sarcomeric proteins, troponin T, cardiac alpha actinin, and SERCA 2a. Both treatments increased the expression of connexin 43 and its relocation to the cell membrane, but biphasic ES was faster and more effective. Finally, when hCPCs were exposed to both monophasic and biphasic ES, they expressed de novo the mRNA of the voltage-dependent calcium channel Cav 3.1(α(1G)) subunit, which is peculiar of the developing heart. Taken together, these results show that ES alone is able to set the conditions for early differentiation of adult hCPCs toward a cardiac phenotype

Isolated slaughterhouse liver as model for normothermic perfusion after warm and cold ischemia: single case report

AbstractLiver transplantation is an ultimate procedure in patients suffering end-stage liver diseases. In these last years the donation after cardiac death (DCD) has increased the pool of potential liver donors. Different studies and procedures are involved in the prevention of the main ischemic problems during the reconditioning and resuscitation of the marginal livers. Normothermic extracorporeal liver perfusion (NELP) avoids prolonged cold storage damage that is the main cause of steatosis and biliary tract ischemia in transplanted patiens. Different porcine models have been studied and developed to understand the ischemia mechanism and to select the better technique for NELP.We conducted our study using a DCD pig liver model collected from slaughterhouse. Using extracorporeal membrane oxygenation, 2000 ml of total fluid containing autologous blood, lidocaine, heparin, antibiotics, glucose 10 % solution and flunixin, the NELP was achieved. The liver was perfused over 7 hours after 48 hours of cold storage (4C°), using Eurocollins solution. During the liver withdrawal in the slaughterhouse 20 minutes were waited to simulate the warm ischemia (WI) time. Histological samples, swab for bacterial grow, blood sample, temperature and pulse oximetry saturation were collected to assess the liver viability and function. These analyses revealed stable metabolism throughout perfusion identifying a cycles 2 hours length, coinciding with recovery of oxygen uptake rates to fresh liver, as described in literature.In summary the preliminary established model of isolated hemoperfused slatherhouse liver reveals the important role of the relation between cold storage and normothermic perfusion. Moreover this preliminary study justifies further investigation of the optimization of the treatment protocols and perfusion media

Microfluidic flow-based platforms for induction and analysis of dynamic shear-mediated platelet activation - Initial validation versus the standardized hemodynamic shearing device

A microfluidic flow-based platform (μFP), able to stimulate platelets via exposure of shear stress patterns pertinent to cardiovascular devices and prostheses, was compared to the Hemodynamic Shearing Device (HSD) - a state-of-the-art bench-top system for exposure of platelets to defined levels and patterns of shear. Platelets were exposed to time-varying shear stress patterns in the two systems; in detail, platelets were recirculated in the μFP or stimulated in the HSD to replicate comparable exposure time. Shear-mediated platelet activation was evaluated via (i) the platelet activity state assay, allowing the measurement of platelet-mediated thrombin generation and associated prothrombotic tendencies, (ii) scanning electron microscopy to evaluate morphological changes of sheared platelets, and (iii) flow cytometry for the determination of platelet phosphatidylserine exposure as a marker of shear activation. The results revealed good matching and comparability between the two systems, with similar trends of platelet activation, formation of microaggregates, and analogous trends of activation marker exposure for both the HSD and microfluidic-stimulated samples. These findings support future translation of the microfluidic platform as a Point-of-Care facsimile system for the diagnosis of thrombotic risk in patients implanted with cardiovascular devices