A High Proliferation Rate is Critical for Reproducible and Standardized Embryoid Body Formation from Laminin-521-Based Human Pluripotent Stem Cell Cultures

When aiming for homogenous embryoid body (EB) differentiation, the use of equal-sized EBs is required to avoid a size-induced differentiation bias. In this study we developed an efficient and standardized EB formation protocol for human pluripotent stem cells (hPSC) cultured in a laminin-521-based xeno-free system. As the cell proliferation rate of the cells growing on laminin-521 strongly affected the efficiency of aggregate formation, we found that recently passaged cells, as well as the addition of ROCK inhibitor, were essential for reproducible EB formation from hPSC single-cell suspensions. EBs could be obtained in a variety of differentiation media, in 96-well round-bottom plates and in hanging drops. Gene expression studies on differentially sized EBs from three individual human embryonic stem cell lines demonstrated that the medium used for differentiation influenced the differentiation outcome to a much greater extent than the number of cells used for the initial EB formation. Our findings give a new insight into factors that influence the EB formation and differentiation process. This optimized method allows us to easily manipulate EB formation and provide an excellent starting point for downstream EB-based differentiation protocols.     

Myoklonus – eine Orientierungshilfe

The puzzling variety of myoclonus is a diagnostic and therapeutic challenge. Early in the diagnostic work-up of myoclonus, electrophysiologic investigations should clarify whether it is myoclonus or not, and whether there are topographic clues, clinical or electrophysiological, to the generating structure. Such clues would influence both the laboratory strategies or neuroimaging investigations aimed at the underlying disease and the selection of drugs to suppress myoclonus.     

Synkopen

Syncope occurs in approximately 25% of the population at least once during their lifetime. It therefore represents the most frequent differential diagnosis of an epileptic seizure. Depending on the prognosis and therapy, syncopes can be divided in cardiac syncope, syncope due to orthostatic hypotension and reflex syncope, of which vasovagal (neurocardiogenic) syncope is the most frequent type. Vasovagal syncope typically occurs after prolonged standing, the sight of blood, pain and medical procedures. The pathophysiology is not well understood. Genetic factors play a role as well, mainly following a complex mode of inheritance. Syncope can usually be differentiated from an epileptic seizure based on the medical history. Typical vasovagal triggers, a typical prodrome and pallor suggest syncope, whereas cyanosis, tongue biting and a prolonged postictal state favor an epileptic seizure. In cases of doubt or if cardiac syncope is suspected additional diagnostic tests may be necessary. Due to the increased mortality, patients with cardiac syncope need urgent cardiological treatment. The aim of the treatment of vasovagal syncope is to avoid additional events as this type is not associated with increased mortality. This can be achieved by behavioral changes, e.g. avoiding the trigger situation and lying down or tensing of arm and leg musculature during the prodrome to avoid further reduction of blood pressure. There is only sparse evidence for pharmacotherapeutic options. Cardiac pacemakers mostly have no effect. The implantation of a pacemaker seems to be beneficial only in patients over 40 years old with frequent vasovagal syncope, asystole and negative tilt table test.     

Early Impacts of a Healthy Food Distribution Program on the Availability and Price of Fresh Fruits and Vegetables in Small Retail Venues in Los Angeles

Healthy food distribution programs that allow small retailers to purchase fresh fruits and vegetables at wholesale prices may increase the profitability of selling produce. While promising, little is known about how these programs affect the availability of fresh fruits and vegetables in underserved communities. This study examined the impacts of a healthy food distribution program in Los Angeles County over its first year of operation (August 2015–2016). Assessment methods included: (1) a brief survey examining the characteristics, purchasing habits, and attitudes of stores entering the program; (2) longitudinal tracking of sales data examining changes in the volume and variety of fruits and vegetables distributed through the program; and (3) the collection of comparison price data from wholesale market databases and local grocery stores. Seventeen stores participated in the program over the study period. One-fourth of survey respondents reported no recent experience selling produce. Analysis of sales data showed that, on average, the total volume of produce distributed through the program increased by six pounds per week over the study period (95% confidence limit: 4.50, 7.50); trends varied by store and produce type. Produce prices offered through the program approximated those at wholesale markets, and were lower than prices at full-service grocers. Results suggest that healthy food distribution programs may reduce certain supply-side barriers to offering fresh produce in small retail venues. While promising, more work is needed to understand the impacts of such programs on in-store environments and consumer behaviors.     

Diagnosis of Paroxysmal Nocturnal Hemoglobinuria: Recent Advances

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired clonal hematopoietic stem cell disorder with its protean clinical manifestations. This is due to partial or complete absence of ‘glycophosphatidyl-inositol-anchor proteins’ (GPI-AP). The main aim of this review is to highlight various diagnostic modalities available, basic principle of each test and recent advances in the diagnosis of PNH. Recently among various tests available, the flow cytometry has become ‘the gold standard’ for PNH testing. In order to overcome the difficulties encountered by the testing and research laboratories throughout the world, International Clinical Cytometry Society has come up with guidelines regarding the indications for testing, protocol for sample collection, processing, panel of antibodies as well as gating strategies to be used, how to interpret the test and reporting format to be used. It is essential to test at least two GPI-linked markers on at least two different lineages particularly on red cells and granulocytes/monocytes. The fluorescent aerolysin combined with other monoclonal antibodies in multicolour flow cytometry offered an improved assay not only for diagnosis but also for monitoring of PNH clones. It is equally important to diagnose this rare entity with high index of suspicion.     

Bubble removal with the use of a vacuum pressure generated by a converging-diverging nozzle

Bubbles are an intrinsic problem in microfluidic devices and they can appear during the initial filling of the device or during operation. This report presents a generalizable technique to extract bubbles from microfluidic networks using an adjacent microfluidic negative pressure network over the entire microfluidic channel network design. We implement this technique by superimposing a network of parallel microchannels with a vacuum microfluidic channel and characterize the bubble extraction rates as a function of negative pressure applied. In addition, we generate negative pressure via a converging-diverging (CD) nozzle, which only requires inlet gas pressure to operate. Air bubbles generated during the initial liquid filling of the microfluidic network are removed within seconds and their volume extraction rate is calculated. This miniaturized vacuum source can achieve a vacuum pressure of 7.23 psi which corresponds to a bubble extraction rate of 9.84 pL/s, in the microfluidic channels we characterized. Finally, as proof of concept it is shown that the bubble removal system enables bubble removal on difficult to fill microfluidic channels such as circular or triangular shaped channels. This method can be easily integrated into many microfluidic experimental protocols.