scholarly journals Disease Modeling with 3D Cell-Based Assays Using a Novel Flowchip System and High-Content Imaging

2021 ◽  
pp. 247263032110006
Author(s):  
Evan F. Cromwell ◽  
Michelle Leung ◽  
Matthew Hammer ◽  
Anthony Thai ◽  
Rashmi Rajendra ◽  
...  
Keyword(s):  
Disease Modeling ◽  
Three Dimensional ◽  
Assay System ◽  
Biological Research ◽  
Functional Evaluation ◽  
High Content Imaging ◽  
Wide Range ◽  
Sample Chamber ◽  
Cytotoxicity Effects

There is an increasing interest in using three-dimensional (3D) cell structures for modeling tumors, organs, and tissue to accelerate translational research. We describe here a novel automated organoid assay system (the Pu·MA System) combined with microfluidic-based flowchips that can facilitate 3D cell-based assays. The flowchip is composed of sample wells, which contain organoids, connected to additional multiple wells that can hold various assay reagents. Organoids are positioned in a protected chamber in sample wells, and fluids are exchanged from side reservoirs using pressure-driven flow. Media exchange, sample staining, wash steps, and other processes can be performed without disruption to or loss of 3D sample. The bottom of the sample chamber is thin, optically clear plastic compatible with high-content imaging (HCI). The whole system can be kept in an incubator, allowing long-term cellular assays to be performed. We present two examples of use of the system for biological research. In the first example, cytotoxicity effects of anticancer drugs were evaluated on HeLa and HepG2 spheroids using HCI and vascular endothelial growth factor expression. In the second application, the flowchip system was used for the functional evaluation of Ca2+ oscillations in neurospheroids. Neurospheres were incubated with neuroactive compounds, and neuronal activity was assessed using Ca2+-sensitive dyes and fast kinetic fluorescence imaging. This novel assay system using microfluidics enables automation of 3D cell-based cultures that mimic in vivo conditions, performs multidosing protocols and multiple media exchanges, provides gentle handling of spheroids and organoids, and allows a wide range of assay detection modalities.

Epithelial Organotypic Cultures: A Viable Model to Address Mechanisms of Carcinogenesis by Epitheliotropic Viruses

2018 ◽  
Vol 18 (4) ◽  
pp. 246-255 ◽  
Author(s):  
Lara Termini ◽  
Enrique Boccardo
Keyword(s):  
Three Dimensional ◽  
Cell Types ◽  
Experimental Models ◽  
Biological Research ◽  
Specific Gene ◽  
Specific Cell ◽  
Organotypic Cultures ◽  
Skin Physiology

In vitro culture of primary or established cell lines is one of the leading techniques in many areas of basic biological research. The use of pure or highly enriched cultures of specific cell types obtained from different tissues and genetics backgrounds has greatly contributed to our current understanding of normal and pathological cellular processes. Cells in culture are easily propagated generating an almost endless source of material for experimentation. Besides, they can be manipulated to achieve gene silencing, gene overexpression and genome editing turning possible the dissection of specific gene functions and signaling pathways. However, monolayer and suspension cultures of cells do not reproduce the cell type diversity, cell-cell contacts, cell-matrix interactions and differentiation pathways typical of the three-dimensional environment of tissues and organs from where they were originated. Therefore, different experimental animal models have been developed and applied to address these and other complex issues in vivo. However, these systems are costly and time consuming. Most importantly the use of animals in scientific research poses moral and ethical concerns facing a steadily increasing opposition from different sectors of the society. Therefore, there is an urgent need for the development of alternative in vitro experimental models that accurately reproduce the events observed in vivo to reduce the use of animals. Organotypic cultures combine the flexibility of traditional culture systems with the possibility of culturing different cell types in a 3D environment that reproduces both the structure and the physiology of the parental organ. Here we present a summarized description of the use of epithelial organotypic for the study of skin physiology, human papillomavirus biology and associated tumorigenesis.

Functional implications of supercontracting muscle in the chameleon tongue retractors

2001 ◽  
Vol 204 (21) ◽  
pp. 3621-3627 ◽  
Author(s):  
Anthony Herrel ◽  
Jay J. Meyers ◽  
Peter Aerts ◽  
Kiisa C. Nishikawa
Keyword(s):  
Prey Capture ◽  
Three Dimensional ◽  
Force Production ◽  
Retractor Muscle ◽  
Muscle Physiology ◽  
Large Prey ◽  
Wide Range ◽  
Ambush Predators ◽  
Full Body

SUMMARYChameleons capture prey items using a ballistic tongue projection mechanism that is unique among lizards. During prey capture, the tongue can be projected up to two full body lengths and may extend up to 600 % of its resting length. Being ambush predators, chameleons eat infrequently and take relatively large prey. The extreme tongue elongation (sixfold) and the need to be able to retract fairly heavy prey at any given distance from the mouth are likely to place constraints on the tongue retractor muscles. The data examined here show that in vivo retractor force production is almost constant for a wide range of projection distances. An examination of muscle physiology and of the ultrastructure of the tongue retractor muscle shows that this is the result (i) of active hyoid retraction, (ii) of large muscle filament overlap at maximal tongue extension and (iii) of the supercontractile properties of the tongue retractor muscles. We suggest that the chameleon tongue retractor muscles may have evolved supercontractile properties to enable a substantial force to be produced over a wide range of tongue projection distances. This enables chameleons successfully to retract even large prey from a variety of distances in their complex three-dimensional habitat.

scholarly journals Genomics Analysis of Metabolic Pathways of Human Stem Cell-Derived Microglia-Like Cells and the Integrated Cortical Spheroids

2019 ◽  
Vol 2019 ◽  
pp. 1-21 ◽  
Author(s):  
Julie Bejoy ◽  
Xuegang Yuan ◽  
Liqing Song ◽  
Thien Hua ◽  
Richard Jeske ◽  
...  
Keyword(s):  
Brain Tissue ◽  
Metabolic Pathways ◽  
Disease Modeling ◽  
Aerobic Glycolysis ◽  
Three Dimensional ◽  
Initiation Factor ◽  
Human Brain Tissue ◽  
P53 Signaling

Brain spheroids or organoids derived from human pluripotent stem cells (hiPSCs) are still not capable of completely recapitulating in vivo human brain tissue, and one of the limitations is lack of microglia. To add built-in immune function, coculture of the dorsal forebrain spheroids with isogenic microglia-like cells (D-MG) was performed in our study. The three-dimensional D-MG spheroids were analyzed for their transcriptome and compared with isogenic microglia-like cells (MG). Cortical spheroids containing microglia-like cells displayed different metabolic programming, which may affect the associated phenotype. The expression of genes related to glycolysis and hypoxia signaling was increased in cocultured D-MG spheroids, indicating the metabolic shift to aerobic glycolysis, which is in favor of M1 polarization of microglia-like cells. In addition, the metabolic pathways and the signaling pathways involved in cell proliferation, cell death, PIK3/AKT/mTOR signaling, eukaryotic initiation factor 2 pathway, and Wnt and Notch pathways were analyzed. The results demonstrate the activation of mTOR and p53 signaling, increased expression of Notch ligands, and the repression of NF-κB and canonical Wnt pathways, as well as the lower expression of cell cycle genes in the cocultured D-MG spheroids. This analysis indicates that physiological 3-D microenvironment may reshape the immunity of in vitro cortical spheroids and better recapitulate in vivo brain tissue function for disease modeling and drug screening.

Multifunctional graphene oxide for bioimaging: emphasis on biological research

2017 ◽  
Vol 9 (2) ◽  
Author(s):  
Do Won Hwang ◽  
Byung Hee Hong ◽  
Dong Soo Lee
Keyword(s):  
Graphene Oxide ◽  
Targeted Delivery ◽  
Near Infrared ◽  
Biological Research ◽  
Natural Interaction ◽  
Drug Delivery Carrier ◽  
Wide Range ◽  
Surface Enhanced Raman

AbstractGraphene oxide (GO) nanomaterials offer a wide range of bioimaging applicability. Almost complete quenching ability of fluorescence by GO and natural interaction of GO with single stranded nucleic acid made GO a useful and intriguing multifunctional nanoplatform both as a biosensor for in vitro microplate diagnostics and as a drug delivery carrier for targeted delivery. GO’s large surface area and strong near infrared absorbance contribute to enhancement of a therapeutic effect with abundant loading of drugs for possible photothermal and photodynamic therapy. Bioimaging capability of GO made it a good theranostic tool, while enabling tracing in vivo pharmacokinetics during concurrent treatment. Fluorescence, either signal on or off, Raman and surface-enhanced Raman scattering (SERs), photoacoustic, and radionuclide imaging modalities can be used for theranostic purposes using GO nanomaterials. In this review, we highlight current applications of GO for bioimaging that are classified into in vitro microplate, in vitro cellular and in vivo bioimaging.

scholarly journals Recent Advances in Aptasensor for Cytokine Detection: A Review

Sensors ◽  
2021 ◽  
Vol 21 (24) ◽  
pp. 8491
Author(s):  
Jinmyeong Kim ◽  
Seungwoo Noh ◽  
Jeong Ah Park ◽  
Sang-Chan Park ◽  
Seong Jun Park ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Physiological Role ◽  
Dimensional Structure ◽  
Biological Research ◽  
Immune Modulators ◽  
Production Methods ◽  
Cytokine Detection ◽  
Cell Signal Transduction ◽  
Excessive Secretion

Cytokines are proteins secreted by immune cells. They promote cell signal transduction and are involved in cell replication, death, and recovery. Cytokines are immune modulators, but their excessive secretion causes uncontrolled inflammation that attacks normal cells. Considering the properties of cytokines, monitoring the secretion of cytokines in vivo is of great value for medical and biological research. In this review, we offer a report on recent studies for cytokine detection, especially studies on aptasensors using aptamers. Aptamers are single strand nucleic acids that form a stable three-dimensional structure and have been receiving attention due to various characteristics such as simple production methods, low molecular weight, and ease of modification while performing a physiological role similar to antibodies.

scholarly journals The Science and Engineering of Stem Cell–Derived Organoids-Examples from Hepatic, Biliary, and Pancreatic Tissues

2020 ◽  
Author(s):  
Ogechi Ogoke ◽  
Mitchell Maloy ◽  
Natesh Parashurama
Keyword(s):  
Stem Cell ◽  
Cellular Therapy ◽  
Disease Modeling ◽  
Historical Context ◽  
Three Dimensional ◽  
Self Organization ◽  
Wide Range ◽  
Clinical Interest ◽  
And Function ◽  
Better Than

Organoid engineering promises to revolutionize medicine with wide ranging applications of scientific, engineering, and clinical interest, including precision and personalized medicine, gene editing, drug development, disease modeling, cellular therapy, and a basic understanding of human development. Organoids are a three-dimensional (3D), miniature, caricature of a target organ, are initiated with stem/progenitor cells, and are extremely promising tools to model organ function. The biological basis for organoids is that they foster stem cell-self renewal, differentiation, and self-organization, recapitulating tissue structure or function better than 2D systems. In this review, we first discuss the importance of epithelial organs and the general properties of epithelial cells to provide context for the liver, pancreas, and gall bladder and rationale for organoid cultures. Next, we develop a general framework to understand self-organization, tissue hierarchy, and organoid cultivation. For each of these areas, we provide historical context, and review both a wide range of biological and/or biophysical/mathematic perspectives that enhances understanding of organoids. Next, we review existing techniques and progress in hepatobiliary and pancreatic organoid engineering. To do this, we review organoids from both primary tissues, cell lines, and stem cells, and introduce engineering studies when applicable. Noninvasive assessment of 1 organoids can reveal underlying biology and enable improved assays for growth, metabolism, and function. Applications of organoid for cell therapy are also discussed. Taken together, we establish a broad strong scientific foundation for organoids and provide an in-depth review of hepatic, biliary and pancreatic organoids.

scholarly journals Biobanking of human gut organoids for translational research

2021 ◽  
Author(s):  
Francesca Perrone ◽  
Matthias Zilbauer
Keyword(s):  
Translational Research ◽  
Three Dimensional ◽  
Short Review ◽  
Human Gut ◽  
Exciting Field ◽  
Wide Range ◽  
Culture Models ◽  
Scientific Expertise ◽  
Key Aspects

AbstractThe development of human organoid culture models has led to unprecedented opportunities to generate self-organizing, three-dimensional miniature organs that closely mimic in vivo conditions. The ability to expand, culture, and bank such organoids now provide researchers with the opportunity to generate next-generation living biobanks, which will substantially contribute to translational research in a wide range of areas, including drug discovery and testing, regenerative medicine as well as the development of a personalized treatment approach. However, compared to traditional tissue repositories, the generation of a living organoid biobank requires a much higher level of coordination, additional resources, and scientific expertise. In this short review, we discuss the opportunities and challenges associated with the generation of a living organoid biobank. Focusing on human intestinal organoids, we highlight some of the key aspects that need to be considered and provide an outlook for future development in this exciting field.

scholarly journals Optical Coherence Tomography for Three-Dimensional Imaging in the Biomedical Field: A Review

2021 ◽  
Vol 9 ◽  
Author(s):  
Shu Zheng ◽  
Yanru Bai ◽  
Zihao Xu ◽  
Pengfei Liu ◽  
Guangjian Ni
Keyword(s):  
Optical Coherence Tomography ◽  
High Speed ◽  
Near Infrared ◽  
Polarization State ◽  
Three Dimensional ◽  
Biological Research ◽  
Light Sources ◽  
Optical Coherence ◽  
Biomedical Field

Optical coherence tomography (OCT) has become a novel approach to noninvasive imaging in the past three decades, bringing a significant potential to biological research and medical biopsy in situ, particularly in three-dimensional (3D) in vivo conditions. Specifically, OCT systems using broad bandwidth sources, mainly centered at near-infrared-II, allow significantly higher imaging depth, as well as maintain a high-resolution and better signal-to-noise ratio than the traditional microscope, which avoids the scattering blur and thus obtains more details from delicate biological structures not just limited to the surface. Furthermore, OCT systems combined the spectrometer with novel light sources, such as multiplexed superluminescent diodes or ultra-broadband supercontinuum laser sources, to obtain sub-micron resolution imaging with high-speed achieve widespread clinical applications. Besides improving OCT performance, the functional extensions of OCT with other designs and instrumentations, taking polarization state or birefringence into account, have further improved OCT properties and functions. We summarized the conventional principle of OCT systems, including time-domain OCT, Fourier-domain OCT, and several typical OCT extensions, compared their different components and properties, and analyzed factors that affect OCT performance. We also reviewed current applications of OCT in the biomedical field, especially in hearing science, discussed existing limitations and challenges, and looked forward to future development, which may provide a guideline for those with 3D in vivo imaging desires.

scholarly journals Determining protein structures using genetics

2018 ◽  
Author(s):  
Jörn M. Schmiedel ◽  
Ben Lehner
Keyword(s):  
Structure Determination ◽  
Low Cost ◽  
Protein Structures ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Biological Research ◽  
X Ray Crystallography ◽  
Close Relationship ◽  
And Function

SummaryDetermining the three dimensional structures of macromolecules is a major goal of biological research because of the close relationship between structure and function. Structure determination usually relies on physical techniques including x-ray crystallography, NMR spectroscopy and cryo-electron microscopy. Here we present a method that allows the high-resolution three-dimensional structure of a biological macromolecule to be determined only from measurements of the activity of mutant variants of the molecule. This genetic approach to structure determination relies on the quantification of genetic interactions (epistasis) between mutations and the discrimination of direct from indirect interactions. This provides a new experimental strategy for structure determination, with the potential to reveal functional and in vivo structural conformations at low cost and high throughput.

High Voltage Electron Microscopy of Cultured Muscle Cells

1976 ◽  
Vol 34 ◽  
pp. 76-77
Author(s):  
Jerry W. Shay
Keyword(s):  
High Voltage ◽  
Three Dimensional ◽  
Muscle Differentiation ◽  
Biological Research ◽  
Muscular Tissue ◽  
High Voltage Electron Microscopy ◽  
Voltage Electron ◽  
High Voltage Electron

It is known from earlier observations that the in vitro rat L6 muscle cell line exhibits many of the features characteristic of in vivo muscle differentiation. The appearance of multinucleated cells, the development of highly organized contractile proteins and the phenomenon of muscle contraction are all easily recognizable features unique to muscular tissue. The L6 myoblasts fuse in a rather homogeneous and synchronous fashion to form myotubes and offer an excellent in vitro system to study the general mechanisms of muscle differentiation.The availability of the high voltage electron microscope (H.V.E.M.) in recent years for biological research has brought about a new interest in the three dimensional organization of components within whole cells, and the present study on L6 cells was undertaken, using the Jeol-1000 facility, at the University of Colorado in Boulder.

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