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The last decade has seen a sharp increase in the number of scientific publications describing physiological and pathological functions of extracellular vesicles (EVs), a collective term covering various subtypes of cell-released, membranous structures, called exosomes, microvesicles, microparticles, ectosomes, oncosomes, apoptotic bodies, and many other names. However, specific issues arise when working with these entities, whose size and amount often make them difficult to obtain as relatively pure preparations, and to characterize properly. The International Society for Extracellular Vesicles (ISEV) proposed Minimal Information for Studies of Extracellular Vesicles (“MISEV”) guidelines for the field in 2014. We now update these “MISEV2014” guidelines based on evolution of the collective knowledge in the last four years. An important point to consider is that ascribing a specific function to EVs in general, or to subtypes of EVs, requires reporting of specific information beyond mere description of function in a crude, potentially contaminated, and heterogeneous preparation. For example, claims that exosomes are endowed with exquisite and specific activities remain difficult to support experimentally, given our still limited knowledge of their specific molecular machineries of biogenesis and release, as compared with other biophysically similar EVs. The MISEV2018 guidelines include tables and outlines of suggested protocols and steps to follow to document specific EV-associated functional activities. Finally, a checklist is provided with summaries of key points.
Men and women differ substantially regarding height, weight, and body fat. Interestingly, previous work detecting genetic effects for waist-to-hip ratio, to assess body fat distribution, has found that many of these showed sex-differences. However, systematic searches for sex-differences in genetic effects have not yet been conducted. Therefore, we undertook a genome-wide search for sexually dimorphic genetic effects for anthropometric traits including 133,723 individuals in a large meta-analysis and followed promising variants in further 137,052 individuals, including a total of 94 studies. We identified seven loci with significant sex-difference including four previously established (near GRB14/COBLL1, LYPLAL1/SLC30A10, VEGFA, ADAMTS9) and three novel anthropometric trait loci (near MAP3K1, HSD17B4, PPARG), all of which were significant in women, but not in men. Of interest is that sex-difference was only observed for waist phenotypes, but not for height or body-mass-index. We found no evidence for sex-differences with opposite effect direction for men and women. The PPARG locus is of specific interest due to its link to diabetes genetics and therapy. Our findings demonstrate the importance of investigating sex differences, which may lead to a better understanding of disease mechanisms with a potential relevance to treatment options.
High-performance liquid chromatography (HPLC) has proved extremely versatile over the past 25 yr for the isolation and punfication of peptides varying widely in their sources, quantity and complexity. This article covers the major modes of HPLC utilized for peptides (size-exclusion, ion-exchange, and reversed-phase), as well as demonstrating the potential of a novel mixed-mode hydrophilic interaction/cation-exchange approach developed in this laboratory. In addition to the value of these HPLC modes for peptide separations, the value of various HPLC techniques for structural characterization of peptides and proteins will be addressed, e.g., assessment of oligomerization state of peptideslproteins by sizeexclusion chromatography and monitoring the hydrophilicitykydrophobicity of amphipathic cr-helical peptides, a vital precursor Tor the development of novel antimicrobial peptides. The value of capillary electrophoresis for peptide separations is also demonstrated. Preparative reversed-phase chromatography purification protocols for sample loads of up to 200 mg on analytical columns and instrumentation are introduced for both peptides and recombinant proteins. Key Words: Peptides; proteins; size-exclusion chromatography (SEC); anion-exchange chromatography (AEX); cation-exchange chromatography (CEX); mixed-mode hydrophilic interaction chromatography (HIL1C)/cation-exchange chromatography (CEX); reversed-phase high-performance liquid chromatography (RP-HPLC); preparative RP-HPLC of peptides and proteins; amino acid side-chain hydrophilicitylhydrophobicity coefficients; amino acid U-helical propensity values; amino acid side-chain stability coefficients
