Analysis of the efficacy of EPIONE therapies to treat phantom limb pain

Indiana University-Purdue University Indianapolis (IUPUI)The primary objectives of this thesis are (1) to discuss the current understanding of phenomena associated with, proposed mechanisms of, and suggested treatments for amputation related pain, (2) to describe the software developed for analyzing results of a clinical study for the treatment of phantom limb pain (PLP), (3) to discuss the methods for a multi-center trial by the EPIONE consortium along with presenting preliminary results, and (4) to discuss the methods and results of a case study involving a new therapy modality for alleviating PLP. Each objective has been expanded into a chapter as described below. Chapter 1 serves as a literature review introducing the topic of amputation, associ- ated phenomena, and proposed mechanisms. The chapter also discusses the currently available treatments and the instruments used to measure PLP. Key topics include the definition of PLP, the prevalence of PLP, current treatment options for PLP, and experimental measurement of PLP. The final objective of this chapter is to introduce topics related to the investigation paradigm utilized for the studies following in Chapter 4 and Chapter 5. Therefore, a minor emphasis has been put on surface electrical stimulation (SES) and operant conditioning. As with any multi-center clinical study, coordination is key. Chapter 2 introduces the common clinical protocol (CCP) and methods of analysis for the clinical trials conducted by the EPIONE consortium. In order to analyze results in an automated fashion, a software tool was developed. This tool, the EPIONE Extraction Program (EEP) along with its extension the Group Analysis Module (GAM), is the focus of Chapter 3. A high-level overview of the requirements, process flow, and software testing are described. This chapter also discusses the methods of analysis for several self-report instruments used to determine effect size in Chapter 4 and Chapter 5. The outputs of the software tools make up the results presented and described in these chapters. In addition to the details included in Chapter 3, supplemental information is available in Appendix A and Appendix B, which are the detailed User Guides for the EEP and GAM. Chapter 4 reviews the pilot study data conducted by the EPIONE consortium. The primary and two secondary instruments used for analysis are discussed. This chapter provides a brief overview of results from the group. Each clinical site used slightly different variations of a common clinical protocol to better understand what effectively drives alleviation of PLP and to allow comparison of results. The work done at Indiana University - Purdue University Indianapolis (IUPUI) represents a small part of several other universities involved in the EPIONE consortium. Chapter 5 focuses on a case study at IUPUI with a more in-depth review of data collected throughout the study period. Using SES, we seek to reverse cortical reorganization by giving meaningful stimuli through existing circuitry. In this chapter the present work is discussed by introducing a case study in detail with an analysis of psychophysical data

Targeting the MET oncogene by concomitant inhibition of receptor and ligand via an antibody-“decoy” strategy

MET, a master gene sustaining "invasive growth," is a relevant target for cancer precision therapy. In the vast majority of tumors, wild-type MET behaves as a "stress-response" gene and relies on the ligand (HGF) to sustain cell "scattering," invasive growth and apoptosis protection (oncogene "expedience"). In this context, concomitant targeting of MET and HGF could be crucial to reach effective inhibition. To test this hypothesis, we combined an anti-MET antibody (MvDN30) inducing "shedding" (i.e., removal of MET from the cell surface), with a "decoy" (i.e., the soluble extracellular domain of the MET receptor) endowed with HGF-sequestering ability. To avoid antibody/decoy interaction-and subsequent neutralization-we identified a single aminoacid in the extracellular domain of MET-lysine 842-that is critical for MvDN30 binding and engineered the corresponding recombinant decoyMET (K842E). DecoyMET(K842E) retains the ability to bind HGF with high affinity and inhibits HGF-induced MET phosphorylation. In HGF-dependent cellular models, MvDN30 antibody and decoyMET(K842E) used in combination cooperate in restraining invasive growth, and synergize in blocking cancer cell "scattering." The antibody and the decoy unbridle apoptosis of colon cancer stem cells grown in vitro as spheroids. In a preclinical model, built by orthotopic transplantation of a human pancreatic carcinoma in SCID mice engineered to express human HGF, concomitant treatment with antibody and decoy significantly reduces metastatic spread. The data reported indicate that vertical targeting of the MET/HGF axis results in powerful inhibition of ligand-dependent MET activation, providing proof of concept in favor of combined target therapy of MET "expedience.

The tyrosine-phosphorylated hepatocyte growth factor/scatter factor receptor associates with phosphatidylinositol 3-kinase.

The receptor for hepatocyte growth factor, also known as scatter factor (HGF/SF), has recently been identified as the 190-kDa heterodimeric tyrosine kinase encoded by the MET proto-oncogene (p190MET). The signaling pathway(s) triggered by HGF/SF are unknown. In A549 cells, a lung epithelial cell line, nanomolar concentrations of HGF/SF induced tyrosine phosphorylation of the p190MET receptor. The autophosphorylated receptor coprecipitated with phosphatidylinositol 3-kinase (PI 3-kinase) activity. In GTL16 cells, a cell line derived from a gastric carcinoma, the p190MET receptor, overexpressed and constitutively phosphorylated on tyrosine, coprecipitated with PI 3-kinase activity and with the 85-kDa PI 3-kinase subunit. In these cells activation of protein kinase C or the increase of intracellular [Ca2+] inhibits tyrosine phosphorylation of the p190MET receptor as well as the association with both PI 3-kinase activity and the 85-kDa subunit of the enzyme. In an in vitro assay, tyrosine phosphorylation of the immobilized p190MET receptor was required for binding of PI 3-kinase from cell lysates. These data strongly suggest that the signaling pathway activated by the HGF/SF receptor includes generation of D-3-phosphorylated inositol phospholipids

Definition of an indicator assessing the impact of a dam on the downstream river landscape

The increasing number of water withdrawals in Alpine regions represents a significant threat to aquatic ecosystems and river landscape (riverscape). To assess their sustainability, the impacts on river ecological status and landscape features need to be quantified with appropriate indicators. However, assessment of landscape attributes is a complex challenge, due to the lack of standardized methods. Moreover, few metrics quantifying the impacts of water withdrawal on downstream riverscape perception are available in the scientific literature. In this paper, a new indicator, named Landscape Protection Level (LPL), aimed at assessing the effects of water withdrawals on the river landscape, is presented. The indicator has been developed in Aosta Valley (NW Italian Alps), where the river network is heavily exploited by hundreds of withdrawals for hydropower production and irrigation, and it has been included in a multi-criteria analysis (MCA) procedure to assess the sustainability of water withdrawal licenses in relation to different flow release scenarios. The LPL indicator is based on three parameters, Constraint Factor, Release Factor, and Visual Elements Factor, quantifying the presence of landscape protection constraints, the ratio of flow released downstream of the dam to the available river discharge, and the impact on the visual perception of the bypassed stretch, respectively. Its application in four real case studies of existing hydropower plants is presented and discussed in the paper, demonstrating the indicator applicability to assess both specific release values and flow release scenarios varying over the year. Results are analyzed by highlighting the main strengths and weaknesses of the indicator and proposing some suggestions for future improvements. In particular, the reactiveness of the indicator, the representativeness of the stakeholders’ interests, the transparency of the indicator calculation procedure, and the time required for data collection and processing are discussed. Finally, future activities aimed at further improving the indicator applicability and transferability to different river contexts are proposed

The long-lasting protective effect of HGF in cardiomyoblasts exposed to doxorubicin requires a positive feed-forward loop mediated by ERK1,2-TIMP1-STAT3

Previous studies showed that the hepatocyte growth factor (HGF)–Met receptor axis plays long-lasting cardioprotection against doxorubicin anti-cancer therapy. Here, we explored the mechanism(s) underlying the HGF protective effect. DNA damage was monitored by histone H2AX phosphorylation and apoptosis by proteolytic cleavage of caspase 3. In doxorubicin-treated H9c2 cardiomyoblasts, the long-lasting cardioprotection is mediated by activation of the Ras/Raf/Mek/Erk (extracellular signal-regulated kinase 1,2) signaling pathway and requires Stat3 (signal transducer and activator of transcription 3) activation. The HGF protection was abrogated by the Erk1,2 inhibitor, PD98059. This translated into reduced Y705 phosphorylation and impaired nuclear translocation of Stat3, showing crosstalk between Erk1,2 and Stat3 signaling. An array of 29 cytokines, known to activate Stat3, was interrogated to identify the molecule(s) linking the two pathways. The analysis showed a selective increase in expression of the tissue inhibitor of metalloproteinases-1 (Timp1). Consistently, inhibition in cardiomyoblasts of Timp1 translation by siRNAs blunted both Stat3 activation and the cardioprotective effect of HGF. Thus, Timp1 is responsible for the generation of a feed-forward loop of Stat3 activation and helps cardiomyocytes to survive during the genotoxic stress induced by anthracyclines

Minimally invasive mitral valve surgery: State-of-the-art and our experience

The minimally invasive approach is becoming the standard-of-care for surgery of the mitral valve. As any less invasive strategy, it entails an increased surgical complexity. Standard-of-care mitral repair using the totally videoscopic approach is indeed reproducible; however, few specific data on patients with complex mitral valve disease are available in the published literature. The purpose of the present paper is to provide an overview of the current state-of-the-art in minimally invasive cardiac surgery, and a summary of recent evidence on the topic, with particular regard to the surgical techniques and comparisons with conventional surgery. The experience of the GVM Care and Research network in the field is also briefly reported