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Cells
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Innovative Methods to Monitor Single Live Cells
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Innovative Methods to Monitor Single Live Cells

A special issue of Cells (ISSN 2073-4409).

Deadline for manuscript submissions: closed (30 April 2018) | Viewed by 46778

Special Issue Editors

Dr. Myong-Hee Sung

E-Mail Website
Guest Editor
Laboratory of Molecular Biology and Immunology, National Institute on Aging, National Institutes of Health, 251 Bayview Boulevard, Baltimore, MD 21224, USA
Interests: systems biology; epigenomics; quantitative live microscopy; transcription
Dr. Erik Martin

E-Mail
Assistant Guest Editor
Laboratory of Molecular Biology and Immunology, National Institute on Aging, National Institutes of Health, 251 Bayview Boulevard, Baltimore, MD 21224, USA
Interests: quantitative live microscopy; systems biology; cancer biology; aging; therapeutics

Special Issue Information

Dear Colleagues,

In this Special Issue, we are assembling a collection of recent methods that enable non-invasive monitoring of biological processes in single cells. Following the same cell over time and acquiring molecular information without disrupting its physiology is arguably one of the most under-served areas in the current technical toolbox of molecular biology. Although significant strides have been made in the past several years, it is still challenging to learn and employ methods that allow real-time measurements of abundance, localization, or interactions of specific molecules inside cells, particularly if the relevant biological process unfolds over hours or days. The cell systems biology community stands to benefit from rigorous and transparent discussions about successful applications and pitfalls of available techniques. We hope to address the need in this issue with a latest set of advances in live-microscopy approaches and related methods.

Dr. Myong-Hee Sung
Dr. Erik Martin
Guest Editors

Manuscript Submission Information

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All submissions that pass pre-check are peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. Cells is an international peer-reviewed open access semimonthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. The Article Processing Charge (APC) for publication in this open access journal is 2700 CHF (Swiss Francs). Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Keywords

  • single cell analysis
  • non-invasive monitoring
  • cell tracking
  • quantitative microscopy
  • real-time assay
  • imaging tools and probes
  • in vivo microscopy
  • automated image analysis software

Published Papers (7 papers)

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Research

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17 pages, 2473 KiB  
Article
Tracking Cell Recruitment and Behavior within the Tumor Microenvironment Using Advanced Intravital Imaging Approaches
by Madison Turk, Victor Naumenko, Douglas J. Mahoney and Craig N. Jenne
Cells 2018, 7(7), 69; https://doi.org/10.3390/cells7070069 - 03 Jul 2018
Cited by 12 | Viewed by 5995
Abstract
Recent advances in imaging technology have made it possible to track cellular recruitment and behavior within the vasculature of living animals in real-time. Using approaches such as resonant scanning confocal and multiphoton intravital microscopy (IVM), we are now able to observe cells within [...] Read more.
Recent advances in imaging technology have made it possible to track cellular recruitment and behavior within the vasculature of living animals in real-time. Using approaches such as resonant scanning confocal and multiphoton intravital microscopy (IVM), we are now able to observe cells within the intact tumor microenvironment of a mouse. We are able to follow these cells for extended periods of time (hours) and can characterize how specific cell types (T cells, neutrophils, monocytes) interact with the tumor vasculature and cancer cells. This approach provides greater insight into specific cellular behaviors and cell–cell interactions than conventional techniques such as histology and flow cytometry. In this report, we describe the surgical preparation of animals to expose the tumor and both resonant scanning confocal and multiphoton imaging approaches used to track leukocyte recruitment, adhesion, and behavior within the tumor microenvironment. We present techniques for the measurement and quantification of leukocyte behavior within the bloodstream and tumor interstitium. The use of IVM to study leukocyte behavior within the tumor microenvironment provides key information not attainable with other approaches, that will help shape the development of better, more effective anticancer drugs and therapeutic approaches. Full article
(This article belongs to the Special Issue Innovative Methods to Monitor Single Live Cells)
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Review

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14 pages, 3821 KiB  
Review
Real-Time Determination of the Cell-Cycle Position of Individual Cells within Live Tumors Using FUCCI Cell-Cycle Imaging
by Shuya Yano and Robert M. Hoffman
Cells 2018, 7(10), 168; https://doi.org/10.3390/cells7100168 - 14 Oct 2018
Cited by 18 | Viewed by 6447
Abstract
Most cytotoxic agents have limited efficacy for solid cancers. Cell-cycle phase analysis at the single-cell level in solid tumors has shown that the majority of cancer cells in tumors is not cycling and is therefore resistant to cytotoxic chemotherapy. Intravital cell-cycle imaging within [...] Read more.
Most cytotoxic agents have limited efficacy for solid cancers. Cell-cycle phase analysis at the single-cell level in solid tumors has shown that the majority of cancer cells in tumors is not cycling and is therefore resistant to cytotoxic chemotherapy. Intravital cell-cycle imaging within tumors demonstrated the cell-cycle position and distribution of cancer cells within a tumor, and cell-cycle dynamics during chemotherapy. Understanding cell-cycle dynamics within tumors should provide important insights into novel treatment strategies. Full article
(This article belongs to the Special Issue Innovative Methods to Monitor Single Live Cells)
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Graphical abstract

19 pages, 1054 KiB  
Review
Real-Time Imaging of Retinal Ganglion Cell Apoptosis
by Timothy E. Yap, Piero Donna, Melanie T. Almonte and Maria Francesca Cordeiro
Cells 2018, 7(6), 60; https://doi.org/10.3390/cells7060060 - 15 Jun 2018
Cited by 32 | Viewed by 6614
Abstract
Monitoring real-time apoptosis in-vivo is an unmet need of neurodegeneration science, both in clinical and research settings. For patients, earlier diagnosis before the onset of symptoms provides a window of time in which to instigate treatment. For researchers, being able to objectively monitor [...] Read more.
Monitoring real-time apoptosis in-vivo is an unmet need of neurodegeneration science, both in clinical and research settings. For patients, earlier diagnosis before the onset of symptoms provides a window of time in which to instigate treatment. For researchers, being able to objectively monitor the rates of underlying degenerative processes at a cellular level provides a biomarker with which to test novel therapeutics. The DARC (Detection of Apoptosing Retinal Cells) project has developed a minimally invasive method using fluorescent annexin A5 to detect rates of apoptosis in retinal ganglion cells, the key pathological process in glaucoma. Numerous animal studies have used DARC to show efficacy of novel, pressure-independent treatment strategies in models of glaucoma and other conditions where retinal apoptosis is reported, including Alzheimer’s disease. This may forge exciting new links in the clinical science of treating both cognitive and visual decline. Human trials are now underway, successfully demonstrating the safety and efficacy of the technique to differentiate patients with progressive neurodegeneration from healthy individuals. We review the current perspectives on retinal ganglion cell apoptosis, the way in which this can be imaged, and the exciting advantages that these future methods hold in store. Full article
(This article belongs to the Special Issue Innovative Methods to Monitor Single Live Cells)
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21 pages, 3895 KiB  
Review
Nanopipettes as Monitoring Probes for the Single Living Cell: State of the Art and Future Directions in Molecular Biology
by Gonca Bulbul, Gepoliano Chaves, Joseph Olivier, Rifat Emrah Ozel and Nader Pourmand
Cells 2018, 7(6), 55; https://doi.org/10.3390/cells7060055 - 06 Jun 2018
Cited by 26 | Viewed by 7693
Abstract
Examining the behavior of a single cell within its natural environment is valuable for understanding both the biological processes that control the function of cells and how injury or disease lead to pathological change of their function. Single-cell analysis can reveal information regarding [...] Read more.
Examining the behavior of a single cell within its natural environment is valuable for understanding both the biological processes that control the function of cells and how injury or disease lead to pathological change of their function. Single-cell analysis can reveal information regarding the causes of genetic changes, and it can contribute to studies on the molecular basis of cell transformation and proliferation. By contrast, whole tissue biopsies can only yield information on a statistical average of several processes occurring in a population of different cells. Electrowetting within a nanopipette provides a nanobiopsy platform for the extraction of cellular material from single living cells. Additionally, functionalized nanopipette sensing probes can differentiate analytes based on their size, shape or charge density, making the technology uniquely suited to sensing changes in single-cell dynamics. In this review, we highlight the potential of nanopipette technology as a non-destructive analytical tool to monitor single living cells, with particular attention to integration into applications in molecular biology. Full article
(This article belongs to the Special Issue Innovative Methods to Monitor Single Live Cells)
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Graphical abstract

28 pages, 3182 KiB  
Review
Fluorescent, Bioluminescent, and Optogenetic Approaches to Study Excitable Physiology in the Single Cardiomyocyte
by Connor N. Broyles, Paul Robinson and Matthew J. Daniels
Cells 2018, 7(6), 51; https://doi.org/10.3390/cells7060051 - 31 May 2018
Cited by 31 | Viewed by 9970
Abstract
This review briefly summarizes the single cell application of classical chemical dyes used to visualize cardiomyocyte physiology and their undesirable toxicities which have the potential to confound experimental observations. We will discuss, in detail, the more recent iterative development of fluorescent and bioluminescent [...] Read more.
This review briefly summarizes the single cell application of classical chemical dyes used to visualize cardiomyocyte physiology and their undesirable toxicities which have the potential to confound experimental observations. We will discuss, in detail, the more recent iterative development of fluorescent and bioluminescent protein-based indicators and their emerging application to cardiomyocytes. We will discuss the integration of optical control strategies (optogenetics) to augment the standard imaging approach. This will be done in the context of potential applications, and barriers, of these technologies to disease modelling, drug toxicity, and drug discovery efforts at the single-cell scale. Full article
(This article belongs to the Special Issue Innovative Methods to Monitor Single Live Cells)
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Other

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9 pages, 1256 KiB  
Perspective
Challenges of Decoding Transcription Factor Dynamics in Terms of Gene Regulation
by Erik W. Martin and Myong-Hee Sung
Cells 2018, 7(9), 132; https://doi.org/10.3390/cells7090132 - 07 Sep 2018
Cited by 11 | Viewed by 4046
Abstract
Technological advances are continually improving our ability to obtain more accurate views about the inner workings of biological systems. One such rapidly evolving area is single cell biology, and in particular gene expression and its regulation by transcription factors in response to intrinsic [...] Read more.
Technological advances are continually improving our ability to obtain more accurate views about the inner workings of biological systems. One such rapidly evolving area is single cell biology, and in particular gene expression and its regulation by transcription factors in response to intrinsic and extrinsic factors. Regarding the study of transcription factors, we discuss some of the promises and pitfalls associated with investigating how individual cells regulate gene expression through modulation of transcription factor activities. Specifically, we discuss four leading experimental approaches, the data that can be obtained from each, and important considerations that investigators should be aware of when drawing conclusions from such data. Full article
(This article belongs to the Special Issue Innovative Methods to Monitor Single Live Cells)
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11 pages, 823 KiB  
Protocol
A Caspase-3 Reporter for Fluorescence Lifetime Imaging of Single-Cell Apoptosis
by Johanna M. Buschhaus, Brock Humphries, Kathryn E. Luker and Gary D. Luker
Cells 2018, 7(6), 57; https://doi.org/10.3390/cells7060057 - 13 Jun 2018
Cited by 15 | Viewed by 5477
Abstract
Fluorescence lifetime imaging (FLIM) is a powerful imaging modality used to gather fluorescent reporter data independent of intracellular reporter intensity or imaging depth. We applied this technique to image real-time activation of a reporter for the proteolytic enzyme, caspase-3, in response to apoptotic [...] Read more.
Fluorescence lifetime imaging (FLIM) is a powerful imaging modality used to gather fluorescent reporter data independent of intracellular reporter intensity or imaging depth. We applied this technique to image real-time activation of a reporter for the proteolytic enzyme, caspase-3, in response to apoptotic cell death. This caspase-3 reporter activity provides valuable insight into cancer cell responsiveness to therapy and overall viability at a single-cell scale. Cleavage of a aspartate-glutamate-valine-aspartate (DEVD) linkage sequence alters Förster resonance energy transfer (FRET) within the reporter, affecting its lifetime. Cellular apoptosis was quantified in multiple environments ranging from 2D flat and 3D spheroid cell culture systems to in vivo murine mammary tumor xenografts. We evaluated cell-by-cell apoptotic responses to multiple pharmacological and genetic methods of interest involved in cancer cell death. Within this article, we describe methods for measuring caspase-3 activation at single-cell resolution in various complex environments using FLIM. Full article
(This article belongs to the Special Issue Innovative Methods to Monitor Single Live Cells)
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