New views inside cells with a digital imaging microscope

1991 ◽  
Vol 49 ◽  
pp. 240-241
Author(s):  
F. S. Fay ◽  
Kevin Fogarty ◽  
Richard Tuft ◽  
Walter Carrington
Keyword(s):  
Digital Imaging ◽  
Cell Biology ◽  
Cell Function ◽  
Single Cells ◽  
Low Noise ◽  
Molecular Composition ◽  
Specific Cell ◽  
Wide Field ◽  
High Quantum Efficiency ◽  
Cell Functions

Many current questions in cell biology revolve around questions regarding how changes in cell function are caused by changes in their molecular composition. Given that cells are highly organized structures often carrying out diverse functions in different compartments, it follows that changes in specific cell functions must involve highly localized changes in molecular composition.We have been involved in the development of the digital imaging microscope as a tool to investigate the distribution of molecules inside single living cells. The system measures fluorescence of probes that are highly fluorescent and specific for a molecule or ion of interest and utilizes a wide-field rather than a confocal microscope to produce an image of fluorescence in a single cell. The image is captured by a high quantum efficiency, low noise cooled CCD, thereby providing ultrahigh efficiency in the acquisition of fluorescent images. By utilizing very powerful light sources, the system is capable of generating an image with good signal-to-noise ratio in a millisecond or less, thereby allowing one to follow extremely rapid changes in molecular or ion distribution in single cells.

Surfaces Designed to Control the Projected Area and Shape of Individual Cells

1999 ◽  
Vol 121 (1) ◽  
pp. 40-48 ◽  
Author(s):  
C. H. Thomas ◽  
J.-B. Lhoest ◽  
D. G. Castner ◽  
C. D. McFarland ◽  
K. E. Healy
Keyword(s):  
Mammalian Cells ◽  
Cell Function ◽  
Single Cells ◽  
Polymer Network ◽  
Interpenetrating Polymer Network ◽  
Specific Cell ◽  
Cell Binding ◽  
Projected Area ◽  
Cell Functions ◽  
Spatially Resolved

Materials with spatially resolved surface chemistry were designed to isolate individual mammalian cells to determine the influence of projected area on specific cell functions (e.g., proliferation, cytoskeletal organization). Surfaces were fabricated using a photolithographic process resulting in islands of cell binding N-(2-aminoethyl)-3-aminopropyl-trimethoxysilane (EDS) separated by a nonadhesive interpenetrating polymer network [poly (acrylamide-co-ethylene glycol); P(AAm-co-EG)]. The surfaces contained over 3800 adhesive islands/cm2, allowing for isolation of single cells with projected areas ranging from 100 μm2 to 10,000 μm2. These surfaces provide a useful tool for researching how cell morphology and mechanical forces affect cell function.

scholarly journals A flexible ontology for inference of emergent whole cell function from relationships between subcellular processes

2017 ◽  
Author(s):  
Jens Hansen ◽  
David Meretzky ◽  
Simeneh Woldesenbet ◽  
Gustavo Stolovitzky ◽  
Ravi Iyengar
Keyword(s):  
Cell Biology ◽  
Cell Function ◽  
Biological Knowledge ◽  
Dependent Manner ◽  
Whole Cell ◽  
Cell Functions ◽  
Cell Responses ◽  
Growing Cell ◽  
Cell Ontology ◽  
Context Specific

AbstractWhole cell responses arise from coordinated interactions between diverse human gene products functioning within various pathways underlying sub-cellular processes (SCP). Lower level SCPs interact to form higher level SCPs, often in a context specific manner to give rise to whole cell function. We sought to determine if capturing such relationships enables us to describe the emergence of whole cell functions from interacting SCPs. We developed the “Molecular Biology of the Cell” ontology based on standard cell biology and biochemistry textbooks and review articles. Currently, our ontology contains 5,385 genes, 753 SCPs and 19,180 expertly curated gene-SCP associations. Our algorithm to populate the SCPs with genes enables extension of the ontology on demand and the adaption of the ontology to the continuously growing cell biological knowledge. Since whole cell responses most often arise from the coordinated activity of multiple SCPs, we developed a dynamic enrichment algorithm that flexibly predicts SCP-SCP relationships beyond the current taxonomy. This algorithm enables us to identify interactions between SCPs as a basis for higher order function in a context dependent manner, allowing us to provide a detailed description of how SCPs together can give rise to whole cell functions. We conclude that this ontology can, from omics data sets, enable the development of detailed multidimensional SCP networks for predictive modeling of emergent whole cell functions.

New views inside cells with the digital imaging microscope

1992 ◽  
Vol 50 (1) ◽  
pp. 560-561
Author(s):  
F.S. Fay ◽  
E.D.W. Moore ◽  
D. Elliot ◽  
M. Rosbash ◽  
K. Carter ◽  
...  
Keyword(s):  
Digital Imaging ◽  
Cell Function ◽  
Single Cells ◽  
Image Feature ◽  
Fluorescent Indicators ◽  
Interactive Analysis ◽  
Molecular Distribution ◽  
Versatile Tool ◽  
Molecular Specificity ◽  
Characteristic Features

The digital imaging microscope has been developed as a tool for understanding changes in molecular distribution underlying changes in cell function. It has evolved principally to meet the needs and opportunities afforded cell biologists to analyze molecular distribution in single cells with an ever increasing range of bright fluorescent indicators having a high degree of molecular specificity. The operation of the digital imaging microscope usually involves four basic steps: 1. image acquisition; 2) image restoration, to reverse the distortion introduced by the optics; 3) image feature extraction, to identify and isolate important characteristic features of an image, and; 4) visualization and interactive analysis of the image. These procedures differ depending on the nature of the specimen and the questions being asked. It is this malleability of the digital imaging microscope that makes it such a powerful and versatile tool.

scholarly journals The Nuclear Option: Evidence Implicating the Cell Nucleus in Mechanotransduction

2017 ◽  
Vol 139 (2) ◽  
Author(s):  
Spencer E. Szczesny ◽  
Robert L. Mauck
Keyword(s):  
Nuclear Envelope ◽  
Mathematical Models ◽  
Cell Nucleus ◽  
Cell Biology ◽  
Cell Function ◽  
Mechanical Stimuli ◽  
Cell Functions ◽  
External Forces ◽  
Nuclear Remodeling

Biophysical stimuli presented to cells via microenvironmental properties (e.g., alignment and stiffness) or external forces have a significant impact on cell function and behavior. Recently, the cell nucleus has been identified as a mechanosensitive organelle that contributes to the perception and response to mechanical stimuli. However, the specific mechanotransduction mechanisms that mediate these effects have not been clearly established. Here, we offer a comprehensive review of the evidence supporting (and refuting) three hypothetical nuclear mechanotransduction mechanisms: physical reorganization of chromatin, signaling at the nuclear envelope, and altered cytoskeletal structure/tension due to nuclear remodeling. Our goal is to provide a reference detailing the progress that has been made and the areas that still require investigation regarding the role of nuclear mechanotransduction in cell biology. Additionally, we will briefly discuss the role that mathematical models of cell mechanics can play in testing these hypotheses and in elucidating how biophysical stimulation of the nucleus drives changes in cell behavior. While force-induced alterations in signaling pathways involving lamina-associated polypeptides (LAPs) (e.g., emerin and histone deacetylase 3 (HDAC3)) and transcription factors (TFs) located at the nuclear envelope currently appear to be the most clearly supported mechanism of nuclear mechanotransduction, additional work is required to examine this process in detail and to more fully test alternative mechanisms. The combination of sophisticated experimental techniques and advanced mathematical models is necessary to enhance our understanding of the role of the nucleus in the mechanotransduction processes driving numerous critical cell functions.

scholarly journals An engineered CRISPR/Cas9 mouse line for simultaneous readout of lineage histories and gene expression profiles in single cells

2019 ◽  
Author(s):  
Sarah Bowling ◽  
Duluxan Sritharan ◽  
Fernando G. Osorio ◽  
Maximilian Nguyen ◽  
Priscilla Cheung ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cell Biology ◽  
Cell Function ◽  
Fetal Liver ◽  
Expression Profiles ◽  
Single Cells ◽  
Gene Expression Profiles ◽  
Mouse Line ◽  
Hematopoietic Stem

AbstractTracing the lineage history of cells is key to answering diverse and fundamental questions in biology. Particularly in the context of stem cell biology, analysis of single cell lineages in their native state has elucidated novel fates and highlighted heterogeneity of function. Coupling of such ancestry information with other molecular readouts represents an important goal in the field. Here, we describe the CARLIN (for CRISPR Array Repair LINeage tracing) mouse line and corresponding analysis tools that can be used to simultaneously interrogate the lineage and transcriptomic information of single cells in vivo. This model exploits CRISPR technology to generate up to 44,000 transcribed barcodes in an inducible fashion at any point during development or adulthood, is compatible with sequential barcoding, and is fully genetically defined. We have used CARLIN to identify intrinsic biases in the activity of fetal liver hematopoietic stem cell (HSC) clones and to uncover a previously unappreciated clonal bottleneck in the response of HSCs to injury. CARLIN also allows the unbiased identification of transcriptional signatures based on in vivo stem cell function without a need for markers or cell sorting.

scholarly journals A New Take on Prion Protein Dynamics in Cellular Trafficking

2020 ◽  
Vol 21 (20) ◽  
pp. 7763
Author(s):  
Rodrigo Nunes Alves ◽  
Rebeca Piatniczka Iglesia ◽  
Mariana Brandão Prado ◽  
Maria Isabel Melo Escobar ◽  
Jacqueline Marcia Boccacino ◽  
...  
Keyword(s):  
Prion Protein ◽  
Cell Function ◽  
Cellular Prion Protein ◽  
Specific Cell ◽  
Membrane Domains ◽  
Cellular Processes ◽  
Cell Functions ◽  
Heterogeneous Membrane ◽  
Cell Compartments ◽  
Distinct Cell

The mobility of cellular prion protein (PrPC) in specific cell membrane domains and among distinct cell compartments dictates its molecular interactions and directs its cell function. PrPC works in concert with several partners to organize signaling platforms implicated in various cellular processes. The scaffold property of PrPC is able to gather a molecular repertoire to create heterogeneous membrane domains that favor endocytic events. Dynamic trafficking of PrPC through multiple pathways, in a well-orchestrated mechanism of intra and extracellular vesicular transport, defines its functional plasticity, and also assists the conversion and spreading of its infectious isoform associated with neurodegenerative diseases. In this review, we highlight how PrPC traffics across intra- and extracellular compartments and the consequences of this dynamic transport in governing cell functions and contributing to prion disease pathogenesis.

scholarly journals New Technologies for Use in Toxicology Studies: Monitoring the Effects of Xenobiotics on Immune Function

1990 ◽  
Vol 9 (3) ◽  
pp. 303-317 ◽  
Author(s):  
J. Paul Robinson ◽  
R.W. Pfeifer
Keyword(s):  
Flow Cytometry ◽  
New Technologies ◽  
Single Cells ◽  
Specific Cell ◽  
Kinetic Measurements ◽  
New Developments ◽  
Cell Functions ◽  
Alternative Approach ◽  
In Vitro Systems

New developments in flow cytometry are now being applied in toxicology studies. There are several reasons for using this technology. First, techniques are well characterized to measure functional parameters of single cells. Such measurements can be directly related to perturbations by xenobiotics, cell-mediated immune responses, or trauma. Second, there is a clear indication for movement toward in vitro systems as highly objective assessments of toxicologic interactions. By measuring specific cell functions at the single cell level, it is possible to define a range of normal responses. More importantly, a multiparametric analysis can be performed with flow cytometry and parameters can be directly related to one another. Furthermore, kinetic measurements can be made, providing vital clues to the mechanisms of actions of drugs or chemicals on functions of specific cell populations. Major advantages of this approach are that studies can be performed on very small volumes of blood, body fluid, or cell culture lines and it is not necessary to isolate pure populations of cells to perform these assays. We believe that this alternative approach in toxicology will provide valuable information unobtainable by traditional means.

Metabolic Regulation and Related Molecular Mechanisms in Various Stem Cell Functions

2020 ◽  
Vol 15 (6) ◽  
pp. 531-546 ◽  
Author(s):  
Hwa-Yong Lee ◽  
In-Sun Hong
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Oxidative Phosphorylation ◽  
Metabolic Pathways ◽  
Critical Role ◽  
The Self ◽  
Specific Cell ◽  
Differentiation Potential ◽  
Self Renewal ◽  
Cell Functions

Recent studies on the mechanisms that link metabolic changes with stem cell fate have deepened our understanding of how specific metabolic pathways can regulate various stem cell functions during the development of an organism. Although it was originally thought to be merely a consequence of the specific cell state, metabolism is currently known to play a critical role in regulating the self-renewal capacity, differentiation potential, and quiescence of stem cells. Many studies in recent years have revealed that metabolic pathways regulate various stem cell behaviors (e.g., selfrenewal, migration, and differentiation) by modulating energy production through glycolysis or oxidative phosphorylation and by regulating the generation of metabolites, which can modulate multiple signaling pathways. Therefore, a more comprehensive understanding of stem cell metabolism could allow us to establish optimal culture conditions and differentiation methods that would increase stem cell expansion and function for cell-based therapies. However, little is known about how metabolic pathways regulate various stem cell functions. In this context, we review the current advances in metabolic research that have revealed functional roles for mitochondrial oxidative phosphorylation, anaerobic glycolysis, and oxidative stress during the self-renewal, differentiation and aging of various adult stem cell types. These approaches could provide novel strategies for the development of metabolic or pharmacological therapies to promote the regenerative potential of stem cells and subsequently promote their therapeutic utility.

II. One size fits all: nonclassical MHC molecules fulfill multiple roles in epithelial cell function

1998 ◽  
Vol 274 (2) ◽  
pp. G227-G231 ◽  
Author(s):  
Richard S. Blumberg
Keyword(s):  
Epithelial Cell ◽  
Mhc Class I ◽  
Cell Biology ◽  
Cell Function ◽  
Surface Glycoprotein ◽  
Class I ◽  
Human Major Histocompatibility Complex ◽  
Cell Surface Glycoprotein ◽  
Mhc Molecules ◽  
Nonclassical Mhc

The human major histocompatibility complex (MHC) on chromosome 6 encodes three classical class I genes: human leukocyte antigen-A (HLA-A), HLA-B, and HLA-C. These polymorphic genes encode a 43- to 45-kDa cell surface glycoprotein that, in association with the 12-kDa β2-microglobulin molecule, functions in the presentation of nine amino acid peptides to the T cell receptor of CD8-bearing T lymphocytes and killer inhibitory receptors on natural killer cells. In addition to these ubiquitously expressed polymorphic proteins, the human genome also encodes a number of nonclassical MHC class I-like, or class Ib, genes that in general encode nonpolymorphic molecules involved in a variety of specific immunologic functions. Many of these genes, including CD1, the neonatal Fc receptor for immunoglobulin G, HLA-G, the MHC class I chain-related gene A, and Hfe, are prominently displayed on epithelial cells, suggesting an important role in epithelial cell biology.

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