L1CAM is a transmembrane protein expressed on neurons that was presumed to be found on neuron-derived extracellular vesicles (NDEVs) in human biofluids. We developed a panel of single-molecule array ...assays to evaluate the use of L1CAM for NDEV isolation. We demonstrate that L1CAM is not associated with extracellular vesicles in human plasma or cerebrospinal fluid and therefore recommend against its use as a marker in NDEV isolation protocols.
Several programmable transcription factors exist based on the versatile Cas9 protein, yet their relative potency and effectiveness across various cell types and species remain unexplored. Here, we ...compare Cas9 activator systems and examine their ability to induce robust gene expression in several human, mouse, and fly cell lines. We also explore the potential for improved activation through the combination of the most potent activator systems, and we assess the role of cooperativity in maximizing gene expression.
Glioblastomas represent the most aggressive glioma grade and are associated with a poor patient prognosis. The current standard of care, consisting of surgery, radiation and chemotherapy, only ...results in a median survival of 14 months, underscoring the importance of developing effective new therapeutic strategies. Among the challenges in treating glioblastomas are primary resistance and the rapid emergence of recurrent disease, which can result from tumor cell-intrinsic mechanisms in addition to tumor microenvironment (TME)-mediated extrinsic resistance. Using a PDGF-B-driven proneural glioma mouse model, we assessed a panel of tyrosine kinase inhibitors with different selectivity profiles. We found that PLX3397, an inhibitor of colony stimulating factor-1 receptor (CSF-1R), blocks glioma progression, markedly suppresses tumor cell proliferation and reduces tumor grade. By contrast, the multi-targeted tyrosine kinase inhibitors dovitinib and vatalanib, which directly target tumor cells, exert minimal anti-tumoral effects in vivo, despite killing glioma cells in vitro, suggesting a TME-mediated resistance mechanism may be involved. Interestingly, PLX3397 interferes with tumor-mediated education of macrophages and consequently restores the sensitivity of glioma cells to tyrosine kinase inhibitors in vivo in preclinical combination trials. Our findings thus demonstrate that microenvironmental alteration by CSF-1R blockade renders tumor cells more susceptible to receptor tyrosine kinase inhibition in a preclinical glioblastoma model, which may have important translational relevance.
Background
To facilitate efficient screening and reduce the length of comprehensive self‐report batteries, a four‐item short form of the Pain Catastrophizing Scale (PCS) and a two‐item short form of ...the Pain Self‐Efficacy Questionnaire (PSEQ) have been developed and evaluated in samples of patients with arm and upper extremity pain.
Aims
The first aim of this study was to evaluate these short forms in a heterogeneous sample of patients seeking treatment for chronic musculoskeletal pain, using a priori criteria for determining adequate internal consistency, construct validity and sensitivity to change. In addition, the findings of past studies were used to identify items suitable for new and potentially stronger short forms of these measures.
Method
Data were provided by 280 patients who completed the original PCS and PSEQ as part of an interdisciplinary rehabilitation programme.
Results
The previously developed four‐item PCS and the newly developed six‐item short form of the PCS both met the internal consistency and construct validity criteria. They did not meet the criterion regarding sensitivity to change. However, similar to what was obtained using the original PCS, large effect sizes were found when using these short forms to examine pre‐treatment to post‐treatment changes in catastrophizing. For the PSEQ, the new four‐item short form was clearly superior to the other alternatives and met all three criteria.
Conclusion
The strongest short forms of the PCS and PSEQ could facilitate the assessment of pain catastrophizing and self‐efficacy in situations in which the use of the longer original measures is not feasible.
The release of RNA-containing extracellular vesicles (EV) into the extracellular milieu has been demonstrated in a multitude of different in vitro cell systems and in a variety of body fluids. ...RNA-containing EV are in the limelight for their capacity to communicate genetically encoded messages to other cells, their suitability as candidate biomarkers for diseases, and their use as therapeutic agents. Although EV-RNA has attracted enormous interest from basic researchers, clinicians, and industry, we currently have limited knowledge on which mechanisms drive and regulate RNA incorporation into EV and on how RNA-encoded messages affect signalling processes in EV-targeted cells. Moreover, EV-RNA research faces various technical challenges, such as standardisation of EV isolation methods, optimisation of methodologies to isolate and characterise minute quantities of RNA found in EV, and development of approaches to demonstrate functional transfer of EV-RNA in vivo. These topics were discussed at the 2015 EV-RNA workshop of the International Society for Extracellular Vesicles. This position paper was written by the participants of the workshop not only to give an overview of the current state of knowledge in the field, but also to clarify that our incomplete knowledge - of the nature of EV(-RNA)s and of how to effectively and reliably study them - currently prohibits the implementation of gold standards in EV-RNA research. In addition, this paper creates awareness of possibilities and limitations of currently used strategies to investigate EV-RNA and calls for caution in interpretation of the obtained data.
The nanostructure of engineered bioscaffolds has a profound impact on cell response, yet its understanding remains incomplete as cells interact with a highly complex interfacial layer rather than the ...material itself. For bioactive glass scaffolds, this layer comprises of silica gel, hydroxyapatite (HA)/carbonated hydroxyapatite (CHA), and absorbed proteins-all in varying micro/nano structure, composition, and concentration. Here, we examined the response of MC3T3-E1 pre-osteoblast cells to 30 mol% CaO-70 mol% SiO
porous bioactive glass monoliths that differed only in nanopore size (6-44 nm) yet resulted in the formation of HA/CHA layers with significantly different microstructures. We report that cell response, as quantified by cell attachment and morphology, does not correlate with nanopore size, nor HA/CHO layer micro/nano morphology, or absorbed protein amount (bovine serum albumin, BSA), but with BSA's secondary conformation as indicated by its β-sheet/α-helix ratio. Our results suggest that the β-sheet structure in BSA interacts electrostatically with the HA/CHA interfacial layer and activates the RGD sequence of absorbed adhesion proteins, such as fibronectin and vitronectin, thus significantly enhancing the attachment of cells. These findings provide new insight into the interaction of cells with the scaffolds' interfacial layer, which is vital for the continued development of engineered tissue scaffolds.
A novel method of hit time and hit position reconstruction in scintillator detectors is described. The method is based on comparison of detector signals with results stored in a library of ...synchronized model signals registered for a set of well-defined positions of scintillation points. The hit position is reconstructed as the one corresponding to the signal from the library which is most similar to the measurement signal. The time of the interaction is determined as a relative time between the measured signal and the most similar one in the library. A degree of similarity of measured and model signals is defined as the distance between points representing the measurement- and model-signal in the multi-dimensional measurement space. Novelty of the method lies also in the proposed way of synchronization of model signals enabling direct determination of the difference between time-of-flights (TOF) of annihilation quanta from the annihilation point to the detectors. The introduced method was validated using experimental data obtained by means of the double strip prototype of the J-PET detector and 22Na sodium isotope as a source of annihilation gamma quanta. The detector was built out from plastic scintillator strips with dimensions of 5mm×19mm×300mm, optically connected at both sides to photomultipliers, from which signals were sampled by means of the Serial Data Analyzer. Using the introduced method, the spatial and TOF resolution of about 1.3cm (σ) and 125ps (σ) were established, respectively.
Gap junctions (GJs) are the only known cellular structures that allow a direct cell-to-cell transfer of signaling molecules by forming densely packed arrays or "plaques" of hydrophilic channels that ...bridge the apposing membranes of neighboring cells. The crucial role of GJ-mediated intercellular communication (GJIC) for all aspects of multicellular life, including coordination of development, tissue function, and cell homeostasis, has been well documented. Assembly and degradation of these membrane channels is a complex process that includes biosynthesis of the connexin (Cx) subunit proteins (innexins in invertebrates) on endoplasmic reticulum (ER) membranes, oligomerization of compatible subunits into hexameric hemichannels (connexons), delivery of the connexons to the plasma membrane (PM), head-on docking of compatible connexons in the extracellular space at distinct locations, arrangement of channels into dynamic spatially and temporally organized GJ channel plaques, as well as internalization of GJs into the cytoplasm followed by their degradation. Clearly, precise modulation of GJIC, biosynthesis, and degradation are crucial for accurate function, and much research currently addresses how these fundamental processes are regulated. Here, we review posttranslational protein modifications (e.g., phosphorylation and ubiquitination) and the binding of protein partners (e.g., the scaffolding protein ZO-1) known to regulate GJ biosynthesis, internalization, and degradation. We also look closely at the atomic resolution structure of a GJ channel, since the structure harbors vital cues relevant to GJ biosynthesis and turnover.
This book examines international dance performances in New York City in the 1940s as sites in which dance artists and audiences contested what it meant to practice globalism in mid-twentieth-century ...America. Debates over globalism in dance proxied larger cultural struggles over how to reconcile the nation’s new role as a global superpower. In dance as in cultural politics, Americans labored over how to realize diversity while honoring difference and manage dueling impulses toward globalism, on the one hand, and isolationism, on the other.
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