Dalhousie University

About: Dalhousie University is a education organization based out in Halifax, Nova Scotia, Canada. It is known for research contribution in the topics: Population & Health care. The organization has 25660 authors who have published 58465 publications receiving 2082403 citations. The organization is also known as: Dalhousie College & The Governors of Dalhousie College and University.

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

Abstract: In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. For example, a key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process versus those that measure flux through the autophagy pathway (i.e., the complete process including the amount and rate of cargo sequestered and degraded). In particular, a block in macroautophagy that results in autophagosome accumulation must be differentiated from stimuli that increase autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. It is worth emphasizing here that lysosomal digestion is a stage of autophagy and evaluating its competence is a crucial part of the evaluation of autophagic flux, or complete autophagy. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation, it is imperative to target by gene knockout or RNA interference more than one autophagy-related protein. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways implying that not all Atg proteins can be used as a specific marker for an autophagic process. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular assays, we hope to encourage technical innovation in the field.

Randomized Assessment of Rapid Endovascular Treatment of Ischemic Stroke

Abstract: Among patients with a proximal vessel occlusion in the anterior circulation, 60 to 80% of patients die within 90 days after stroke onset or do not regain functional independence despite alteplase treatment. We evaluated rapid endovascular treatment in addition to standard care in patients with acute ischemic stroke with a small infarct core, a proximal intracranial arterial occlusion, and moderate-to-good collateral circulation. Methods We randomly assigned participants to receive standard care (control group) or standard care plus endovascular treatment with the use of available thrombectomy devices (intervention group). Patients with a proximal intracranial occlusion in the anterior circulation were included up to 12 hours after symptom onset. Patients with a large infarct core or poor collateral circulation on computed tomography (CT) and CT angiography were excluded. Workflow times were measured against predetermined targets. The primary outcome was the score on the modified Rankin scale (range, 0 [no symptoms] to 6 [death]) at 90 days. A proportional odds model was used to calculate the common odds ratio as a measure of the likelihood that the intervention would lead to lower scores on the modified Rankin scale than would control care (shift analysis). Results The trial was stopped early because of efficacy. At 22 centers worldwide, 316 participants were enrolled, of whom 238 received intravenous alteplase (120 in the intervention group and 118 in the control group). In the intervention group, the median time from study CT of the head to first reperfusion was 84 minutes. The rate of functional independence (90-day modified Rankin score of 0 to 2) was increased with the intervention (53.0%, vs. 29.3% in the control group; P<0.001). The primary outcome favored the intervention (common odds ratio, 2.6; 95% confidence interval, 1.7 to 3.8; P<0.001), and the intervention was associated with reduced mortality (10.4%, vs. 19.0% in the control group; P = 0.04). Symptomatic intracerebral hemorrhage occurred in 3.6% of participants in intervention group and 2.7% of participants in control group (P = 0.75). Conclusions Among patients with acute ischemic stroke with a proximal vessel occlusion, a small infarct core, and moderate-to-good collateral circulation, rapid endovascular treatment improved functional outcomes and reduced mortality. (Funded by Covidien and others; ESCAPE ClinicalTrials.gov number, NCT01778335.)

Guidelines for the use and interpretation of assays for monitoring autophagy

Abstract: In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field.