Piezo1 activation attenuates thrombin-induced blebbing in breast cancer cells.

Cancer cells exploit a variety of migration modes to leave primary tumors and establish metastases, including amoeboid cell migration, which is typically reliant on bleb formation. Here we demonstrate that thrombin induces dynamic blebbing in the MDA-MB-231 breast cancer cell line and confirm that protease-activated receptor 1 (PAR1) activation is sufficient to induce this effect. Cell confinement has been implicated as a driving force in bleb-based migration. Unexpectedly, we found that gentle contact compression, exerted using a custom built 'cell press' to mechanically stimulate cells, reduced thrombin-induced blebbing. Thrombin-induced blebbing was similarly attenuated using the small molecule Yoda1, an agonist of the mechanosensitive Ca2+ channel Piezo1, and this attenuation was impaired in Piezo1-depleted cells. Additionally, Piezo1 activation suppressed thrombin-induced phosphorylation of ezrin, radixin and moesin (ERM) proteins, which are implicated in the blebbing process. Our results provide mechanistic insights into Piezo1 activation as a suppressor of dynamic blebbing, specifically that which is induced by thrombin.

Well-Plate µFASP for Proteomic Analysis of Single Pancreatic Islets.

Filter-aided sample preparation (FASP) is widely used in bottom-up proteomics for tryptic digestion. However, the sample recovery yield of this method is limited by the amount of the starting material. While ∼100 ng of digested protein is sufficient for thorough protein identification, proteomic information gets lost with a protein content <10 µg due to incomplete peptide recovery from the filter. We developed and optimized a flexible well-plate µFASP device and protocol that is suitable for an ∼1 µg protein sample. In 1 µg of HeLa digest, we identified 1295 ± 10 proteins with µFASP followed by analysis with liquid chromatography-mass spectrometry. In contrast, only 524 ± 5 proteins were identified with the standard FASP protocol, while 1395 ± 4 proteins were identified in 20 µg after standard FASP as a benchmark. Furthermore, we conducted a combined peptidomic and proteomic study of single pancreatic islets with well-plate µFASP. Here, we separated neuropeptides and digested the remaining on-filter proteins for bottom-up proteomic analysis. Our results indicate inter-islet heterogeneity for the expression of proteins involved in glucose catabolism, pancreatic hormone processing, and secreted peptide hormones. We consider our method to provide a useful tool for proteomic characterization of samples where the biological material is scarce. All proteomic data are available under DOI: 10.6019/PXD029039.

Prevention of posterior wall puncture with a self-made needle block.

Vascular access is one of the most commonly performed invasive procedures in medicine. Ultrasound-guided vascular access has been shown to improve patient safety, decrease associated complications and increase first attempt success rates, however, the risk for a posterior venous wall puncture (PVWP) still exists. To reduce this complication, needle guides have been used, though, current methods have limited accessibility and generalizability. Thus, the aim of this article is to describe how a self-made needle block constructed with materials present in a central line kit can reduce the incidence of PVWP and its associated complications in novice POCUS users.

An open source extrusion bioprinter based on the E3D motion system and tool changer to enable FRESH and multimaterial bioprinting.

Bioprinting is increasingly used to create complex tissue constructs for an array of research applications, and there are also increasing efforts to print tissues for transplantation. Bioprinting may also prove valuable in the context of drug screening for personalized medicine for treatment of diseases such as cancer. However, the rapidly expanding bioprinting research field is currently limited by access to bioprinters. To increase the availability of bioprinting technologies we present here an open source extrusion bioprinter based on the E3D motion system and tool changer to enable high-resolution multimaterial bioprinting. As proof of concept, the bioprinter is used to create collagen constructs using freeform reversible embedding of suspended hydrogels (FRESH) methodology, as well as multimaterial constructs composed of distinct sections of laminin and collagen. Data is presented demonstrating that the bioprinted constructs support growth of cells either seeded onto printed constructs or included in the bioink prior to bioprinting. This open source bioprinter is easily adapted for different bioprinting applications, and additional tools can be incorporated to increase the capabilities of the system.

Integrating First Responder Hands-on Training into Medical School Curricula-a Perspective from Medical Students.

An investigation of the effectiveness of an emergency-responder-based program entitled "First Five Minutes" in teaching medical students the necessary assessments and procedures upon arrival to a medical crisis situation in a timely and cost-effective manner. The "First Five" includes scene safety, primary/secondary survey, airway management, basic life support (BLS)/cardiopulmonary resuscitation (CPR), and hemorrhage control.

NUrF-Optimization of in situ UV-vis and fluorescence and autonomous characterization techniques with small-angle neutron scattering instrumentation.

We have designed, built, and validated a (quasi)-simultaneous measurement platform called NUrF, which consists of neutron small-angle scattering, UV-visible, fluorescence, and densitometry techniques. In this contribution, we illustrate the concept and benefits of the NUrF setup combined with high-performance liquid chromatography pumps to automate the preparation and measurement of a mixture series of Brij35 nonionic surfactants with perfluorononanoic acid in the presence of a reporter fluorophore (pyrene).