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- Klionsky, Daniel J., et al.
(författare)
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Guidelines for the use and interpretation of assays for monitoring autophagy
- 2012
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Ingår i: Autophagy. - : Informa UK Limited. - 1554-8635 .- 1554-8627. ; 8:4, s. 445-544
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Forskningsöversikt (refereegranskat)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.
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- 2019
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Tidskriftsartikel (refereegranskat)
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| 7. |
- Cheng, Ran, et al.
(författare)
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In Vitro and in Vivo Evaluation of C-11-Labeled Azetidinecarboxylates for Imaging Monoacylglycerol Lipase by PET Imaging Studies
- 2018
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Ingår i: Journal of Medicinal Chemistry. - : American Chemical Society (ACS). - 0022-2623 .- 1520-4804. ; 61:6, s. 2278-2291
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Tidskriftsartikel (refereegranskat)abstract
- Monoacylglycerol lipase (MAGL) is the principle enzyme for metabolizing endogenous cannabinoid ligand 2-arachidonoyglycerol (2-AG). Blockade of MAGL increases 2-AG levels, resulting in subsequent activation of the endocannabinoid system, and has emerged as a novel therapeutic strategy to treat drug addiction, inflammation, and neurodegenerative diseases. Herein we report a new series of MAGL inhibitors, which were radiolabeled by site-specific labeling technologies, including C-11-carbonylation and spirocyclic iodonium ylide (SCIDY) radio fluorination. The lead compound [C-11]10 (MAGL-0519) demonstrated high specific binding and selectivity in vitro and in vivo. We also observed unexpected washout kinetics with these irreversible radiotracers, in which in vivo evidence for turnover of the covalent residue was unveiled between MAGL and azetidine carboxylates. This work may lead to new directions for drug discovery and PET tracer development based on azetidine carboxylate inhibitor scaffold.
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| 8. |
- Li, Zhijian, et al.
(författare)
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Systematic exploration of essential yeast gene function with temperature-sensitive mutants.
- 2011
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Ingår i: Nature biotechnology. - : Springer Science and Business Media LLC. - 1546-1696 .- 1087-0156. ; 29:4, s. 361-7
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Tidskriftsartikel (refereegranskat)abstract
- Conditional temperature-sensitive (ts) mutations are valuable reagents for studying essential genes in the yeast Saccharomyces cerevisiae. We constructed 787 ts strains, covering 497 (∼45%) of the 1,101 essential yeast genes, with ∼30% of the genes represented by multiple alleles. All of the alleles are integrated into their native genomic locus in the S288C common reference strain and are linked to a kanMX selectable marker, allowing further genetic manipulation by synthetic genetic array (SGA)-based, high-throughput methods. We show two such manipulations: barcoding of 440 strains, which enables chemical-genetic suppression analysis, and the construction of arrays of strains carrying different fluorescent markers of subcellular structure, which enables quantitative analysis of phenotypes using high-content screening. Quantitative analysis of a GFP-tubulin marker identified roles for cohesin and condensin genes in spindle disassembly. This mutant collection should facilitate a wide range of systematic studies aimed at understanding the functions of essential genes.
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| 9. |
- Lin, Zhen, et al.
(författare)
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Outcomes after readmission at the index or nonindex hospital following acute myocardial infarction complicated by cardiogenic shock
- 2021
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Ingår i: Clinical Cardiology. - : John Wiley & Sons. - 0160-9289 .- 1932-8737. ; 44:2, s. 200-209
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Tidskriftsartikel (refereegranskat)abstract
- Little is known about the prevalence and outcomes of readmission to nonindex hospitals after an admission for acute myocardial infarction complicated by cardiogenic shock (AMI-CS). We aimed to determine the rate of nonindex readmissions following AMI-CS and to evaluate its association with clinical factors, hospitalization cost, length of stay (LOS), and in-hospital mortality rates.HYPOTHESIS: Nonindex readmission may lead to worse in-hospital outcomes.METHODS: We reviewed the data of inpatients with AMI-CS between 2010 and 2017 using the National Readmission Database. The survey analytical methods recommended by the Healthcare Cost and Utilization Project were used for national estimates. Multiple regression models were used to evaluate the predictors of nonindex readmission, and its association with hospitalization cost, LOS, and in-hospital mortality rates.RESULTS: Of 238 349 patients with AMI-CS, 28028 (11.76%) had an unplanned readmission within 30 days. Of these patients, 7423 (26.48%) were readmitted to nonindex hospitals. Compared with index readmission, nonindex readmission was associated with higher hospitalization costs (p < .0001), longer LOS (p < .0001), and increased in-hospital mortality rates (p = .0016). Patients who had a history of percutaneous coronary intervention, received intubation/mechanical ventilation, or left against medical advice during the initial admission had greater odds of a nonindex readmission.CONCLUSIONS: Over one-fourth of readmissions following AMI-CS were to nonindex hospitals. These admissions were associated with higher hospitalization costs, longer LOS, and higher in-hospital mortality rates. Further studies are needed to evaluate whether a continuity of care plan in the acute hospital setting can improve outcomes after AMI-CS.
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