The morphology and cytoskeletal structure of fibroblasts, endothelial cells, neutrophils are documented for cells cultured on surfaces with stiffness ranging from 2 to 55,000 Pa that have been laminated fibronectin or collagen as adhesive ligand. When grown in sparse culture no cell-cell contacts, fibroblasts show an abrupt change spread area occurs at a range around 3,000 Pa. No actin stress fibers seen soft surfaces, the appearance is complete coincident which they spread. Upregulation α5...

The mechanical properties of substrates underlying cells can have profound effects on cell structure and function. To examine the effect substrate deformability neuronal growth, protein-laminated polyacrylamide gels were prepared with differing amounts bisacrylamide to generate varying elastic moduli ranging from 500 5500 dyne/cm2. Mouse spinal cord primary plated allowed grow extend neurites for several weeks in culture. While neurons grew well gels, glia, which are normally co-cultured...

This paper describes a gradient-generating microfluidic platform for optimizing proliferation and differentiation of neural stem cells (NSCs) in culture. Microfluidic technology has great potential to improve cell (SC) cultures, whose promise cell–based therapies is limited by the inability precisely control their behavior Compared traditional culture tools, platforms should provide much greater over microenvironment rapid optimization media composition using relatively small numbers cells....

Neural stem cells are multipotent with the ability to differentiate into neurons, astrocytes, and oligodendrocytes. Lineage specification is strongly sensitive mechanical properties of cellular environment. However, molecular pathways transducing matrix cues intracellular signaling linked lineage remain unclear. We found that mechanically gated ion channel Piezo1 expressed by brain-derived human neural stem/progenitor responsible for a induced ionic current. activity triggered traction...

Alternative polyadenylation (APA) of mRNAs has emerged as an important mechanism for post-transcriptional gene regulation in higher eukaryotes. Although microarrays have recently been used to characterize APA globally, they a number serious limitations that prevents comprehensive and highly quantitative analysis. To better its regulation, we developed deep sequencing-based method called P oly( A ) S ite Seq uencing (PAS-Seq) quantitatively profiling RNA at the transcriptome level. PAS-Seq...

Modeling the in vivo microenvironment typically involves placing cells a three-dimensional (3D) extracellular matrix (ECM) physiologically relevant context with respect to other cells. The mechanical and chemical features of 3D microenvironments play important roles tissue engineering, tumor growth metastasis, defining stem cell niches, it is increasingly recognized that behave much differently when surrounded by ECM than anchored 2D substrate. To create more closely mimic settings, here we...

The earliest step in creating the cerebral cortex is specification of neuroepithelium to a cortical fate. Using mouse genetic mosaics and timed inactivations, we demonstrated that Lhx2 acts as classic selector gene essential intrinsic determinant identity. activity restricted an early critical period when stem cells comprise neuroepithelium, where it cell-autonomously specify identity suppress alternative fates spatially dependent manner. Laterally, null adopt antihem identity, whereas...

Piezo channels transduce mechanical stimuli into electrical and chemical signals to powerfully influence development, tissue homeostasis, regeneration. Studies on Piezo1 have largely focused transduction of "outside-in" forces, its response internal, cell-generated forces remains poorly understood. Here, using measurements endogenous activity traction in native cellular conditions, we show that generate spatially-restricted Piezo1-mediated Ca

Abstract Deciphering the factors that regulate human neural stem cells will greatly aid in their use as models of development and therapeutic agents. The extracellular matrix (ECM) is a component cell niches vivo regulates multiple functions diverse types, yet little known about its effects on stem/precursor (NSPCs). We therefore plated NSPCs four different substrates (poly‐L‐ornithine, fibronectin, laminin, matrigel) compared responses with those mouse NSPCs. Compared other substrates,...

A novel dielectrophoresis switching with vertical electrodes in the sidewall of microchannels for multiplexed objects has been designed, fabricated and tested. With appropriate electrode design, lateral DEP force can be generated so that one dynamically position particulates along width channel. set interdigitated is used generation non-uniform electrical fields to generate negative forces repel beads/cells from sidewalls. countering another patterned on opposing sidewall. These adjusted by...

Doublecortin (DCX) is a microtubule-associated protein required for neuronal migration to the cerebral cortex. DCAMKL1 consists of an N terminus that 65% similar DCX throughout entire length DCX, but also contains additional 360 amino acid C-terminal domain encoding putative Ca 2+ /calmodulin-dependent kinase. The homology suggested may regulate microtubules, as well mediate phosphorylation-dependent signal transduction pathway. Here we show expressed CNS and PNS in migrating populations...

The relatively new field of stem cell biology is hampered by a lack sufficient means to accurately determine the phenotype cells. Cell-type-specific markers, such as surface proteins used for flow cytometry or fluorescence-activated sorting, are limited and often recognize multiple members lineage. We sought develop complementary approach that would be less dependent on identification particular markers subpopulations cells instead measure their overall character. tested whether microfluidic...

In the stem cell field there is a lack of non invasive and fast methods to identify cell's metabolic state, differentiation state cell-lineage commitment. Here we describe label-free method that uses NADH as an intrinsic biomarker Phasor approach Fluorescence Lifetime microscopy measure fingerprint cells. We show different states are related stages bias neuronal glial fate, prior expression lineage markers. Our data demonstrate FLIM signature distinguishes non-invasively neurons from...

Human neural stem/progenitor cells (hNSPCs) are good candidates for treating central nervous system (CNS) trauma since they secrete beneficial trophic factors and differentiate into mature CNS cells; however, many die after transplantation. This cell death can be ameliorated by inclusion of a biomaterial scaffold, making identification optimal scaffolds hNSPCs critical research focus. We investigated the properties fibrin-based their effects on found that fibrin generated from salmon...

Abstract This paper presents a novel design and separation strategy for lateral flow‐through of cells/particles in microfluidics by dual frequency coupled dielectrophoresis (DEP) forces enabled vertical interdigitated electrodes embedded the channel sidewalls. Unlike field‐flow‐fractionation‐DEP separations microfluidics, which utilize planar on microchannel floor to generate DEP force balance gravitational separate objects at different height locations, is sidewall that are used non‐uniform...

Conducting detailed cellular analysis of complex biological samples poses challenges in cell sorting and recovery for downstream analysis. Label-free microfluidics provide a promising solution these applications. In this work, we investigate particle manipulation on two label-free microdevice designs using cDEP to enrich E. coli from whole human blood mimic infection workflows. is still growing source bacteremia, sepsis, other infections modern countries, affecting millions patients...

The Arp2/3 complex and filamin A (FLNa) branch actin filaments. To define the role of these actin-binding proteins in cellular architecture, we compared morphology FLNa-deficient human melanoma (M2) cells three stable derivatives expressing normal FLNa concentrations. All cell lines contain similar amounts complex. Serum addition causes serum-starved M2 to extend flat protrusions transiently; thereafter, turn into spherical blebs do not crawl. short-lived lamellae a dense mat long filaments...

Neuronal cytoskeletal elements such as neurofilaments, F-actin, and microtubules are actively translocated by an yet unidentified mechanism. This report describes a novel interaction between neurofilaments microtubule motor proteins that mediates the translocation of along in vitro. Native purified from spinal cord transported at rates 100-1000 nm/s to both plus minus ends. motion requires ATP is partially inhibited vanadate, consistent with activity neurofilament-bound molecular motors....

Zinc is an essential trace element participating in diverse biological processes. Cellular zinc levels are strictly controlled by two families of transport proteins: ZIP channels (SLC39A) and ZnT transporters (SLC30A). increase cytosolic importing into cells or releasing from intracellular stores such as the ER. Among all 14 human members family, ZIP7 a gatekeeper release stores, requiring post-translational activation phosphorylation on residues S275 S276, resulting multiple downstream...

Distinguishing human neural stem/progenitor cell (huNSPC) populations that will predominantly generate neurons from those produce glia is currently hampered by a lack of sufficient type-specific surface markers predictive fate potential. This limits investigation lineage-biased progenitors and their potential use as therapeutic agents. A live-cell biophysical label-free measure would solve this problem obviating the need for specific markers.We used dielectrophoresis (DEP) to analyze...

Abstract Neural stem and progenitor cell (NSPC) fate is strongly influenced by mechanotransduction as modulation of substrate stiffness affects lineage choice. Other types mechanical stimuli, such stretch (tensile strain), occur during CNS development trauma, but their consequences for NSPC differentiation have not been reported. We delivered a 10% static equibiaxial to NSPCs examined effects on differentiation. found specifically impacts into oligodendrocytes, neurons or astrocytes this...