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

Rapid Method for Small Grain and Corn Flour Authentication Using GC/EI-MS and Multivariate Analysis

The aim of this study was the application of the gas chromatography-mass spectrometry system (GC/EI-MS) system and multivariate data analysis to investigate the possibility of chemical differentiation between small grain flour (wheat, barley, oat, triticale, rye) and corn flour samples. All cereal flour samples were first defatted with hexane, after which the extraction with ethanol was performed. Extracted simple sugars (monosaccharides, disaccharides, trisaccharides, and sugar alcohols) were analyzed in the form of their corresponding trimethylsilyl oximes. Peaks of simple sugar derivatives were selected in total ion current (TIC) chromatograms by monitoring exclusively the following characteristic abundant ions: 204, 217, and 361 m/z. The total surface areas under the selected peaks were subjected to multivariate analysis. Applying principal coordinate analysis and hierarchical cluster analysis to obtained data, samples of corn flour could be very clearly distinguished from all samples of small grain flour, which presented a weaker separation among each other. This method circumvents common analytical procedures by excluding simple sugar identifications, quantitative analysis, the use of analytical standards, and calibration curves. Results are applicable in the quality assurance of mixed flour on the market, considering the increased popularity of their consumption in human nutrition

Antibacterial properties and healing effects of Melipona scutellaris honey in MRSA-infected wounds of rats

ABSTRACT PURPOSE : To investigate the antimicrobial, immunological and healing effects of Melipona scutellaris honey on infected wounds of rat skin. METHODS: Twenty four Wistar rats were distributed in four groups (6-each). The uninfected skin wounds of group I rats were treated daily with saline for 7 days. Uninfected wounds (group II) rats were treated with honey. In group III (treated with saline) and group IV (treated with honey) wounds were inoculated with MRSA ATTC43300. The first bacterial culture was performed 24 hours later. In the 7th day new culture was done, and wound biopsies were used for cytokines dosage and histopathology. RESULTS: In group I and III rats the CFU/g count of S. aureus in wounds was zero. In group II rats the CFU/g counts in the wound tissue were significantly higher than in wounds of group IV rats. The density histopathological parameters and the expression of TNF-α, IL1-β, Il-6 were significantly higher on wounds of group IV then in the other groups. CONCLUSION: Honey of Melipona scutellaris was effective in the management of infected wounds, by significant bacterial growth inhibition, enhancement of cytokine expression, and positively influenced the wound repair