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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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- Klionsky, Daniel J., et al.
(författare)
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Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes
- 2008
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Ingår i: Autophagy. - : Landes Bioscience. - 1554-8627 .- 1554-8635. ; 4:2, s. 151-175
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Forskningsöversikt (refereegranskat)abstract
- Research in autophagy continues to accelerate,1 and as a result many new scientists are entering the field. Accordingly, it is important to establish a standard set of criteria for monitoring macroautophagy in different organisms. Recent reviews have described the range of assays that have been used for this purpose.2,3 There are many useful and convenient methods that can be used to monitor macroautophagy in yeast, but relatively few in other model systems, and there is much confusion regarding acceptable methods to measure macroautophagy in higher eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers of autophagosomes versus those that measure flux through the autophagy pathway; thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from fully functional autophagy that includes delivery to, and degradation within, lysosomes (in most higher eukaryotes) or the vacuole (in plants and fungi). Here, we present a set of guidelines for the selection and interpretation of the methods that can be used by investigators who are attempting to examine macroautophagy and related processes, as well as by reviewers who need to provide realistic and reasonable critiques of papers that investigate these processes. This set of guidelines is 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 verify an autophagic response.
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- Tellez, Helio Fernandez, et al.
(författare)
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Exercise during Short-Term and Long-Term Continuous Exposure to Hypoxia Exacerbates Sleep-Related Periodic Breathing
- 2016
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Ingår i: Sleep. - : Associated Professional Sleep Societies. - 0161-8105 .- 1550-9109. ; 39:4, s. 773-783
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Tidskriftsartikel (refereegranskat)abstract
- Study Objectives: Exposure to hypoxia elevates chemosensitivity, which can lead to periodic breathing. Exercise impacts gas exchange, altering chemosensitivity; however, interactions between sleep, exercise and chronic hypoxic exposure have not been examined. This study investigated whether exercise exacerbates sleep-related periodic breathing in hypoxia. Methods: Two experimental phases. Short-Term Phase: a laboratory controlled, group-design study in which 16 active, healthy men (age: 25 +/- 3 y, height: 1.79 +/- 0.06 m, mass: 74 +/- 8 kg) were confined to a normobaric hypoxic environment (FIO2 = 0.139 +/- 0.003, 4,000 m) for 10 days, after random assignment to a sedentary (control, CON) or cycle-exercise group (EX). Long-Term Phase: conducted at the Concordia Antarctic Research Station (3,800 m equivalent at the Equator) where 14 men (age: 36 +/- 9 y, height: 1.77 +/- 0.09 m, mass: 75 +/- 10 kg) lived for 12-14 months, continuously confined. Participants were stratified post hoc based on self-reported physical activity levels. We quantified apnea-hypopnea index (AHI) and physical activity variables. Results: Short-Term Phase: mean AHI scores were significantly elevated in the EX group compared to CON (Night1 = CON: 39 +/- 51, EX: 91 +/- 59; Night10 = CON: 32 +/- 32, EX: 92 +/- 48; P = 0.046). Long-Term Phase: AHI was correlated to mean exercise time (R-2 = 0.4857; P = 0.008) and the coefficient of variation in night oxyhemoglobin saturation (SpO(2); R-2 = 0.3062; P = 0.049). Conclusions: Data indicate that exercise (physical activity) per se affects night SpO(2) concentrations and AHI after a minimum of two bouts of moderateintensity hypoxic exercise, while habitual physical activity in hypobaric hypoxic confinement affects breathing during sleep, up to 13+ months' duration
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