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

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 fl ux 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, defi ned as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (inmost 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 fi eld 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 delete or knock down more than one autophagy-related gene. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways so 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 autophagy assays, we hope to encourage technical innovation in the field

Isolation and Characterization of a Slowly Milk-Coagulating Variant of Lactobacillus helveticus Deficient in Purine Biosynthesis

A slowly milk-coagulating variant (Fmc(−)) of Lactobacillus helveticus CRL 1062, designated S1, was isolated and characterized. Strain S1 possessed all the known essential components required to utilize casein as a nitrogen source, which include functional proteinase and peptidase activities as well as functional amino acid, di- and tripeptide, and oligopeptide transport systems. The amino acid requirements of strain S1 were similar to those of the parental strain. However, on a purine-free, chemically defined medium, the growth rate of the Fmc(−) strain was threefold lower than that of the wild-type strain. L. helveticus S1 was found to be defective in IMP dehydrogenase activity and therefore was deficient in the ability to synthesize XMP and GMP. This conclusion was further supported by the observation that the addition of guanine or xanthine to milk, a substrate poor in purine compounds, restored the Fmc(+) phenotype of L. helveticus S1

Panoramic maps collection

Collection contains about 1,700 idealized schematic views of towns and cities; a wealth of detail in setting, streets, and buildings. The majority of items presented here are documented in "Panoramic maps of cities in the United States and Canada", second edition (1984), by John R. Hébert and Patrick E. Dempsey.Title from HTML header (viewed on Aug. 8, 2003)

Crystal and Electronic Structures of Complex Bismuth Iodides <i>A</i>₃Bi₂I₉ (<i>A</i> = K, Rb, Cs) Related to Perovskite: Aiding the Rational Design of Photovoltaics

Ternary bismuth halides form an interesting functional materials class in the context of the closely related Pb halide perovskite photovoltaics, especially given the significantly reduced toxicity of Bi when compared with Pb. The compounds <i>A</i>₃Bi₂I₉ (<i>A</i> = K, Rb, Cs) examined here crystallize in two different structure types: the layered defect-perovskite K₃Bi₂I₉ type, and the Cs₃Cr₂Cl₉ type. The latter structure type features isolated Bi₂I₉^3– anions. Here, the crystal structures of the ternary iodides are redetermined and a corrected structural model for Rb₃Bi₂I₉, as established by single crystal X-ray diffraction and solid state ⁸⁷Rb NMR spectroscopy and supported by density functional theory (DFT) calculations is presented. A variety of facile preparation techniques for single crystals, bulk materials, as well as solution-processed thin films are described. The optical properties and electronic structures are investigated experimentally by optical absorption and ultraviolet photoemission spectroscopy and computationally by DFT calculations. Absolute band positions of the valence and conduction bands of these semiconductors, with excellent agreement of experimental and calculated values, are reported, constituting a useful input for the rational interface design of efficient electronic and optoelectronic devices. The different structural connectivity in the two different structure types, somewhat surprisingly, appears to not impact band positions or band gaps in a significant manner. Computed dielectric properties, including the finding of anomalously large Born effective charge tensors on Bi³⁺, suggest proximal structural instabilities arising from the Bi³⁺ 6s² lone pair. These anomalous Born effective charges are promising for defect screening and effective charge carrier transport. The structural, electronic, and optical properties of the complex bismuth iodides are to some extent similar to the related lead iodide perovskites. The deeper valence band positions in the complex bismuth iodides point to the need for different choices of hole transport materials for Bi-iodide based solar cell architectures

Association of diet quality indices with serum and metabolic biomarkers in participants of the ORISCAV-LUX-2 study

Purpose: Diet quality is a critical modifiable factor related to health, including the risk of cardiometabolic complications. Rather than assessing the intake of individual food items, it is more meaningful to examine overall dietary patterns. This study investigated the adherence to common dietary indices and their association with serum/metabolic parameters of disease risk. Methods: Dietary intakes of the general adult population (n = 1404, 25–79 years) were assessed by a validated food-frequency questionnaire (174 items). The French ANSES-Ciqual food composition database was used to compute nutrient intakes. Seven indicators were calculated to investigate participants’ diet quality: the Alternative Healthy Eating Index (AHEI), Dietary Approaches to Stop Hypertension Score (DASH-S), Mediterranean Diet Score (MDS), Diet Quality Index-International (DQI-I), Dietary Inflammatory Index (DII), Dietary Antioxidant Index (DAI), and Naturally Nutrient-Rich Score (NNRS). Various serum/metabolic parameters were used in the validity and association analyses, including markers of inflammation, blood glucose, and blood lipid status. Results: Following linear regression models adjusted for confounders, the DASH-S was significantly associated with most metabolic parameters (14, e.g., inversely with blood pressure, triglycerides, urinary sodium, uric acid, and positively with serum vitamin D), followed by the DQI-I (13, e.g., total cholesterol, apo-A/B, uric acid, and blood pressure) and the AHEI (11, e.g., apo-A, uric acid, serum vitamin D, diastolic blood pressure and vascular age). Conclusion: Food-group-based indices, including DASH-S, DQI-I, and AHEI, were good predictors for serum/metabolic parameters, while nutrient-based indices, such as the DAI or NNRS, were less related to biological markers and, thus, less suitable to reflect diet quality in a general population

A New Dietary Inflammatory Index Predicts Interval Changes in Serum High-Sensitivity C-Reactive Protein1–3

Inflammation is associated with a number of chronic conditions, such as cancer and cardiovascular disease. Reducing inflammation may help prevent or treat these conditions. Diet has consistently been shown to modulate inflammation. To facilitate research into the inflammatory effect of diet on health in humans, we sought to develop and validate an Inflammatory Index designed to assess the inflammatory potential of individuals' diets. An Inflammatory Index was developed based on the results of an extensive literature search. Using data from a longitudinal observational study that carefully measured diet and the inflammatory marker, serum high-sensitivity (hs) C-reactive protein (CRP), in ∼600 adults for 1 y, we conducted analyses to test the effect of Inflammatory Index score on hs-CRP as a continuous and dichotomous (≤3 mg/L, >3 mg/L) indicator of inflammatory response, while controlling for important potential confounders. Results based on continuous measures of hs-CRP suggested that an increasing Inflammatory Index score (representing movement toward an antiinflammatory diet) was associated with a decrease in hs-CRP. Analyses using hs-CRP as a dichotomous variable showed that an antiinflammatory diet was associated with a decrease in the odds of an elevated hs-CRP (P = 0.049). The results are consistent with the ability of the Inflammatory Index to predict hs-CRP and provide additional evidence that diet plays a role in the regulation of inflammation, even after careful control of a wide variety of potential confounders