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

Omics-based molecular techniques in oral pathology centred cancer: Prospect and challenges in Africa

: The completion of the human genome project and the accomplished milestones in the human proteome project; as well as the progress made so far in computational bioinformatics and “big data” processing have contributed immensely to individualized/personalized medicine in the developed world.At the dawn of precision medicine, various omics-based therapies and bioengineering can now be applied accurately for the diagnosis, prognosis, treatment, and risk stratifcation of cancer in a manner that was hitherto not thought possible. The widespread introduction of genomics and other omics-based approaches into the postgraduate training curriculum of diverse medical and dental specialties, including pathology has improved the profciency of practitioners in the use of novel molecular signatures in patient management. In addition, intricate details about disease disparity among diferent human populations are beginning to emerge. This would facilitate the use of tailor-made novel theranostic methods based on emerging molecular evidences

Sequence-specific binding of normal serum immunoglobulin M to exposed protein C-termini

Both the timely clearance of degraded endogenous structures and the presence of secreted natural immunoglobulin M (IgM) are needed to avoid autoimmunity. These requirements may be causally related provided that natural IgM preferentially reacts with degraded antigens and, by activating complement, mediates their non-inflammatory clearance through complement receptors. We have previously shown that normal serum IgM reacts in vivo and in vitro with virtually all randomly generated C-terminal peptides displayed on T7 phage. The resultant multivalent IgM–peptide complexes activate complement and are detected by a loss of phage infectivity. A striking feature of these reactions is that different C-terminal peptides (≈ 3–4 amino acids) specifically react with different ‘C-terminal’ IgM (C-IgM) antibodies. This suggests that degraded supramolecular structures, expressing elevated levels of identical C-termini as a result of proteolysis, denaturation and abnormal exposure of repetitive protein constituents, may be preferential targets of C-IgM-mediated complement activation in the physiological environment. The specificity of C-IgM–peptide reactions is much higher than one would expect, assuming that normal serum IgM mostly comprises polyspecific natural antibodies. However, it is possible that polyspecific IgM is not adequately registered by our ‘functional’ phage-inactivation assays. In this study, we resolve the issue of C-IgM specificity by directly characterizing the binding reactivity of normal serum IgM with phage-displayed C-terminal peptides

Biological characterization of clones derived from the edmonston strain of measles virus in comparison with schwarz and CAM-70 vaccine strains

Four virus clones were derived from the Edmonston strain of measles virus by repeated plaque purification. These clones were compared with the vaccine strains Schwarz and CAM-70 in terms of biological activities including plaque formation, hemagglutination, hemolysis and replication in Vero cells and chick embryo fibroblasts (CEF). Two clones of intermediate plaque yielded mixed plaque populations on subcultivation whereas the other two, showing small and large plaque sizes, showed stable plaque phenotypes. The vaccine strains showed consistent homogeneous plaque populations. All the Edmonston clones showed agglutination of monkey erythrocytes in isotonic solution while both vaccine strains hemagglutinated only in the presence of high salt concentrations. Variation in the hemolytic activity was observed among the four clones but no hemolytic activity was detected for the vaccine virus strains. Vaccine strains replicated efficiently both in Vero cells and CEF. All four clones showed efficient replication in Vero cells but different replication profiles in CEF. Two of them replicated efficiently, one was of intermediate efficiency and the other showed no replication in CEF. Two of the clones showed characteristics similar to vaccine strains. One in terms of size and homogeneity of plaques, the other for a low hemolytic activity and both for the efficiency of propagation in CEF