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

Nurses' perceptions of aids and obstacles to the provision of optimal end of life care in ICU

Contains fulltext : 172380.pdf (publisher's version ) (Open Access

Azaaldol Condensation of a Lithium Enolate Solvated by <i>N,N,N′,N′</i>-Tetramethylethylenediamine: Dimer-Based 1,2-Addition to Imines

The lithium enolate of <i>tert</i>-amylacetate solvated by <i>N,N,N′,N′</i>-tetramethylethylenediamine (TMEDA) is shown to be a doubly chelated dimer. Adding the dimeric enolate to 4-fluorobenzaldehyde-<i>N</i>-phenylimine affords an N-lithiated β-amino ester shown to be monomeric using ⁶Li and ¹⁵N NMR spectroscopies. Rate studies using ¹⁹F NMR spectroscopy reveal reaction orders consistent with a transition structure of stoichiometry [(ROLi)₂(TMEDA)₂(imine)]^⧧. Density functional theory computations explore several possible dimer-based transition structures with monodentate and bidentate coordination of TMEDA. Supporting rate studies using <i>trans-N,N,N′,N′</i>-1,2-tetramethylcyclohexanediamine showing analogous rates and rate law suggest that TMEDA is fully chelated