Porphyromonas gingivalis gingipains cause defective macrophage migration towards apoptotic cells and inhibit phagocytosis of primary apoptotic neutrophils:gingipains, apoptotic cell removal & inflammation

Periodontal disease is a prevalent chronic inflammatory condition characterised by an aberrant host response to a pathogenic plaque biofilm resulting in local tissue damage and frustrated healing that can result in tooth loss. Cysteine proteases (gingipains) from the key periodontal pathogen Porphyromonas gingivalis have been implicated in periodontal disease pathogenesis by inhibiting inflammation resolution and are linked with systemic chronic inflammatory conditions such as rheumatoid arthritis. Efficient clearance of apoptotic cells is essential for the resolution of inflammation and tissue restoration. Here we sought to characterise the innate immune clearance of apoptotic cells and its modulation by gingipains. We examined the capacity of gingipain-treated macrophages to migrate towards and phagocytose apoptotic cells. Lysine gingipain treatment of macrophages impaired macrophage migration towards apoptotic neutrophils. Furthermore, lysine gingipain treatment reduced surface expression levels of CD14, a key macrophage receptor for apoptotic cells, which resulted in reduced macrophage interactions with apoptotic cells. Additionally, whilst apoptotic cells and their derived secretome were shown to inhibit TNF-α induced expression by P.gingivalis LPS, we demonstrated that gingipain preparations induced a rapid inflammatory response in macrophages that was resistant to the anti-inflammatory effects of apoptotic cells or their secretome. Taken together these data indicate that P.gingivalis may promote the chronic inflammation seen in periodontal disease patients by multiple mechanisms including rapid, potent gingipain-mediated inflammation coupled with receptor cleavage leading to defective clearance of apoptotic cells and reduced anti-inflammatory responses. Thus gingipains represent a potential therapeutic target for intervention in the management of chronic periodontal disease

Assessment of higher order cognitive skills in undergraduate education: modified essay or multiple choice questions? Research paper

Background: Reliable and valid written tests of higher cognitive function are difficult to produce, particularly for the assessment of clinical problem solving. Modified Essay Questions (MEQs) are often used to assess these higher order abilities in preference to other forms of assessment, including multiple-choice questions (MCQs). MEQs often form a vital component of end-of-course assessments in higher education. It is not clear how effectively these questions assess higher order cognitive skills. This study was designed to assess the effectiveness of the MEQ to measure higher-order cognitive skills in an undergraduate institution. Methods: An analysis of multiple-choice questions and modified essay questions (MEQs) used for summative assessment in a clinical undergraduate curriculum was undertaken. A total of 50 MCQs and 139 stages of MEQs were examined, which came from three exams run over two years. The effectiveness of the questions was determined by two assessors and was defined by the questions ability to measure higher cognitive skills, as determined by a modification of Bloom's taxonomy, and its quality as determined by the presence of item writing flaws. Results: Over 50% of all of the MEQs tested factual recall. This was similar to the percentage of MCQs testing factual recall. The modified essay question failed in its role of consistently assessing higher cognitive skills whereas the MCQ frequently tested more than mere recall of knowledge. Conclusion: Construction of MEQs, which will assess higher order cognitive skills cannot be assumed to be a simple task. Well-constructed MCQs should be considered a satisfactory replacement for MEQs if the MEQs cannot be designed to adequately test higher order skills. Such MCQs are capable of withstanding the intellectual and statistical scrutiny imposed by a high stakes exit examination.Edward J Palmer, Peter G Devit

Developmentally regulated GTP binding protein 1 (DRG1) controls microtubule dynamics

The mitotic spindle, essential for segregating the sister chromatids into the two evolving daughter cells, is composed of highly dynamic cytoskeletal filaments, the microtubules. The dynamics of microtubules are regulated by numerous microtubule associated proteins. We identify here Developmentally regulated GTP binding protein 1 (DRG1) as a microtubule binding protein with diverse microtubule-associated functions. In vitro, DRG1 can diffuse on microtubules, promote their polymerization, drive microtubule formation into bundles, and stabilize microtubules. HeLa cells with reduced DRG1 levels show delayed progression from prophase to anaphase because spindle formation is slowed down. To perform its microtubule-associated functions, DRG1, although being a GTPase, does not require GTP hydrolysis. However, all domains are required as truncated versions show none of the mentioned activities besides microtubule binding

Current understanding of the relationship between cervical manipulation and stroke: what does it mean for the chiropractic profession?

The understanding of the relationship between cervical manipulative therapy (CMT) and vertebral artery dissection and stroke (VADS) has evolved considerably over the years. In the beginning the relationship was seen as simple cause-effect, in which CMT was seen to cause VADS in certain susceptible individuals. This was perceived as extremely rare by chiropractic physicians, but as far more common by neurologists and others. Recent evidence has clarified the relationship considerably, and suggests that the relationship is not causal, but that patients with VADS often have initial symptoms which cause them to seek care from a chiropractic physician and have a stroke some time after, independent of the chiropractic visit

Transbilayer Phospholipid Movements in ABCA1-Deficient Cells

Tangier disease is an inherited disorder that results in a deficiency in circulating levels of HDL. Although the disease is known to be caused by mutations in the ABCA1 gene, the mechanism by which lesions in the ABCA1 ATPase effect this outcome is not known. The inability of ABCA1 knockout mice (ABCA1−/−) to load cholesterol and phospholipids onto apoA1 led to a proposal that ABCA1 mediates the transbilayer externalization of phospholipids, an activity integral not only to the formation of HDL particles but also to another, distinct process: the recognition and clearance of apoptotic cells by macrophages. Expression of phosphatidylserine (PS) on the surface of both macrophages and their apoptotic targets is required for efficient engulfment of the apoptotic cells, and it has been proposed that ABCA1 is required for transbilayer externalization of PS to the surface of both cell types. To determine whether ABCA1 is responsible for any of the catalytic activities known to control transbilayer phospholipid movements, these activities were measured in cells from ABCA1−/− mice and from Tangier individuals as well as ABCA1-expressing HeLa cells. Phospholipid movements in either normal or apoptotic lymphocytes or in macrophages were not inhibited when cells from knockout and wildtype mice or immortalized cells from Tangier individuals vs normal individuals were compared. Exposure of PS on the surface of normal thymocytes, apoptotic thymocytes and elicited peritoneal macrophages from wildtype and knockout mice or B lymphocytes from normal and Tangier individuals, as measured by annexin V binding, was also unchanged. No evidence was found of ABCA1-stimulated active PS export, and spontaneous PS movement to the outer leaflet in the presence or absence of apoA1 was unaffected by the presence or absence of ABCA1. Normal or Tangier B lymphocytes and macrophages were also identical in their ability to serve as targets or phagocytes, respectively, in apoptotic cell clearance assays. No evidence was found to support the suggestion that ABCA1 is involved in transport to the macrophage cell surface of annexins I and II, known to enhance phagocytosis of apoptotic cells. These results show that mutations in ABCA1 do not measurably reduce the rate of transbilayer movements of phospholipids in either the engulfing macrophage or the apoptotic target, thus discounting catalysis of transbilayer movements of phospholipids as the mechanism by which ABCA1 facilitates loading of phospholipids and cholesterol onto apoA1

Minimal information for studies of extracellular vesicles 2018 (MISEV2018): a position statement of the International Society for Extracellular Vesicles and update of the MISEV2014 guidelines

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