Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field

Incorporation of the emotional indicators of the patient journey into healthcare organization management

Abstract Background In recent years, attempts have been made to incorporate patients' experiences into healthcare processes, to complement clinical indicators, with what are known as patient‐reported outcome measures (PROMs) and patient‐reported experience measures (PREMs). While the research into PROMs is more developed, the application of PREMs faces some difficulties. The incorporation of emotional indicators into assessments of the experience is an area that remains to be explored. Objectives This study proposes a new technique to analyse the emotions experienced by patients during the care process, examines how these emotions influence their satisfaction and propose that if healthcare services focus more on patients' emotions, they can improve the effectiveness of the sector. Methods The first, qualitative stage, gathered data from patients to design a patient journey (PJ). The PJ was then reproduced as a video. In a subsequent, quantitative stage, the video was shown to experimental participants, and their emotions were measured through facial expression analysis and a questionnaire. Results A new technique to gather emotional data showed that the emotions patients experience do not affect their satisfaction with their clinical care or the physical aspects of the process. However, their emotions did affect their satisfaction with people and organizations. Conclusions The importance of the emotional component of patients' experiences was underlined. Therefore, healthcare organizations should take account of this dimension, as well as the cognitive, to increase patient satisfaction and improve their care processes. Understanding the impact of the emotions identified at the subconscious level can help improve the patient experience. A new methodology was applied that may help health professionals to collect emotional data about patients' experiences and to develop PREMs. Patient/Public Contribution Patients were involved in all stages of this research. In the exploratory phase, some helped define the touchpoints of the PJ. The data from the subsequent experimental phase were collected from another group, and the emotions they experienced were identified through the analysis of their facial expressions. Based on the results of this study, a working group including patients has been established to work on improvements in the PJ

Osseointegration of TI6Al4V dental implants modified by thermal oxidation in osteoporotic rabbits

[Background]: In this work, the effect of the heat treatment on Ti6Al4V implants and topical administration of growth hormone to address a better osseointegration in osteoporotic patients has been analysed. [Methods]: The osseointegration process of Ti6Al4V implants modified by oxidation treatment at 700 °C for 1 h and the influence of local administration of growth hormone (GH) in osteoporotic female rabbits after 15 and 30 days of implantation have been studied. Bone response was analysed through densitometric and histomorphometric studies. Characterization of the surface was provided by scanning electron microscopy. [Results]: The oxidation treatment promotes the formation of an oxide scale grown on the Ti6Al4V implants that alters the nanoroughness of the surface. Bone mineral density (BMD) increases from 0.347 ± 0.014 (commercial) to 0.383 ± 0.012 g cm−2 (modified), and bone-to-implant contact (BIC) goes from 48.01 ± 14.78 (commercial) to 55.37 ± 15.31 (modified) after 30 days of implantation. [Conclusions]: The oxidation treatment on the Ti6Al4V dental implants enhances the early bone formation at the longest periods of implantation. No significant differences in the BMD and BIC results in healthy and osteoporotic rabbits were revealed with respect to the local administration of GH in the implantation site.The authors acknowledge the source of funding received from Ministerio de Economia y Competitividad (Spain) through the MAT2011-29152-C02-01 project.Peer Reviewe

Novel Cancer Chemotherapy Hits by Molecular Topology: Dual Akt and Beta-Catenin Inhibitors

<div><p>Background and Purpose</p><p>Colorectal and prostate cancers are two of the most common types and cause of a high rate of deaths worldwide. Therefore, any strategy to stop or at least slacken the development and progression of malignant cells is an important therapeutic choice. The aim of the present work is the identification of novel cancer chemotherapy agents. Nowadays, many different drug discovery approaches are available, but this paper focuses on Molecular Topology, which has already demonstrated its extraordinary efficacy in this field, particularly in the identification of new <i>hit</i> and <i>lead</i> compounds against cancer. This methodology uses the graph theoretical formalism to numerically characterize molecular structures through the so called topological indices. Once obtained a specific framework, it allows the construction of complex mathematical models that can be used to predict physical, chemical or biological properties of compounds. In addition, Molecular Topology is highly efficient in selecting and designing new <i>hit</i> and <i>lead</i> drugs. According to the aforementioned, Molecular Topology has been applied here for the construction of specific Akt/mTOR and β-catenin inhibition mathematical models in order to identify and select novel antitumor agents.</p><p>Experimental Approach</p><p>Based on the results obtained by the selected mathematical models, six novel potential inhibitors of the Akt/mTOR and β-catenin pathways were identified. These compounds were then tested <i>in vitro</i> to confirm their biological activity.</p><p>Conclusion and Implications</p><p>Five of the selected compounds, CAS n° 256378-54-8 (Inhibitor n°1), 663203-38-1 (Inhibitor n°2), 247079-73-8 (Inhibitor n°3), 689769-86-6 (Inhibitor n°4) and 431925-096 (Inhibitor n°6) gave positive responses and resulted to be active for Akt/mTOR and/or β-catenin inhibition. This study confirms once again the Molecular Topology’s reliability and efficacy to find out novel drugs in the field of cancer.</p></div

Pharmacological distribution diagram for natural Akt inhibitors obtained using the DF₁ (the black colour represents Akt inhibitors and the white colour, the compounds without Akt inhibition activity).

<p>Pharmacological distribution diagram for natural Akt inhibitors obtained using the DF₁ (the black colour represents Akt inhibitors and the white colour, the compounds without Akt inhibition activity).</p

Pharmacological distribution diagram for Akt inhibitors obtained using the DF₂ (the <i>black colour</i> represents Akt inhibitors and the <i>white colour</i>, the compounds without Akt inhibition activity).

<p>Pharmacological distribution diagram for Akt inhibitors obtained using the DF₂ (the <i>black colour</i> represents Akt inhibitors and the <i>white colour</i>, the compounds without Akt inhibition activity).</p

Effect of of selected anti-cancer agents on Akt/mTOR signaling pathway.

<p>PC-3 cells were treated with vehicle (C) or 50 μM of the selected compounds and proteins were detected by Western blot. Upper panel, a representative image of three different experiments. Lower panel, densitometric values represented as the mean ± S.D. of the three experiments.</p

Effect of selected anti-cancer agents on β-catenin cellular distribution in PC-3 cells.

<p>PC-3 cells were seeded on glass coverslips and pre-treated for 30 min with 50 μM of the inhibitors and then co-treated with 40 ng/ml Wnt3 (Wnt) for 4 h. Cells were stained with polyclonal antibody anti-beta-catenin followed by Alexa-Fluor488-conjugated anti rabbit IgG as described in methods. Confocal image shown is representative of two experiments.</p