Gastrointestinal organoid cultures for functional evaluation of oncogenic loci.

2015 ◽  
Vol 33 (3_suppl) ◽  
pp. 85-85
Author(s):  
Ameen Abdulla Salahudeen ◽  
Xingnan Li ◽  
Michael Cantrell ◽  
Calvin Jay Kuo
Keyword(s):  
Digestive Tract ◽  
Cancer Biology ◽  
Three Dimensional ◽  
Functional Evaluation ◽  
3 Dimensional ◽  
Upper Digestive Tract ◽  
Conditional Allele ◽  
Sequencing Studies ◽  
Control Virus

85 Background: Novel in vitro methods surpassing limitations of current gastrointestinal cancer models such as gastric and esophagus cancer are required to functionally validate putative oncogenic loci discovered by genome sequencing efforts. The in vitro culture of primary, non-transformed tissues as three-dimensional organoids that accurately recapitulate organ structure and physiology has diverse applications including cancer biology. Methods: Mouse wild type, or p53flox/flox in tandem with lox-stop-lox KRASG12D upper digestive tract tissue containing epithelial and mesenchymal components were cultured in an air-liquid-interface and subjected to adenovirus expressing either immunoglobulin Fc (control) or GFP tagged Cre recombinase. Results: 3-dimensional organoids were generated with histological adherence to normal tissue architecture including that seen in esophagus and were able to be maintained in long term culture. Organoids exposed to GFP tagged Cre adenovirus demonstrated green fluorescence not seen in organoids exposed to control virus. Conditional allele organoids that were exposed to Cre adenovirus demonstrated increased rate of growth compared to controls. Histology of these rapidly growing organoids demonstrated cellular features consistent with dysplasia. Conclusions: 3-dimensional organoids can be generated from upper digestive tract tissues, can undergo adenoviral mediated transfection to achieve oncogenic gene expression or inactivation resulting in dysplastic morphology. 3-dimensional organoids are therefore an attractive model to study or identify candidate oncogenic loci identified by recent genomic sequencing studies.

3D Culture Modelling: An Emerging Approach for Translational Cancer Research in Sarcomas

2020 ◽  
Vol 27 (29) ◽  
pp. 4778-4788 ◽  
Author(s):  
Victoria Heredia-Soto ◽  
Andrés Redondo ◽  
José Juan Pozo Kreilinger ◽  
Virginia Martínez-Marín ◽  
Alberto Berjón ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
Cancer Biology ◽  
Soft Tissues ◽  
Three Dimensional ◽  
3D Culture ◽  
Resistance Mechanisms ◽  
In Vitro Models ◽  
Tumour Spheroids ◽  
Current Therapies

Sarcomas are tumours of mesenchymal origin, which can arise in bone or soft tissues. They are rare but frequently quite aggressive and with a poor outcome. New approaches are needed to characterise these tumours and their resistance mechanisms to current therapies, responsible for tumour recurrence and treatment failure. This review is focused on the potential of three-dimensional (3D) in vitro models, including multicellular tumour spheroids (MCTS) and organoids, and the latest data about their utility for the study on important properties for tumour development. The use of spheroids as a particularly valuable alternative for compound high throughput screening (HTS) in different areas of cancer biology is also discussed, which enables the identification of new therapeutic opportunities in commonly resistant tumours.

scholarly journals The Role of Pre-Clinical 3-Dimensional Models of Osteosarcoma

2020 ◽  
Vol 21 (15) ◽  
pp. 5499
Author(s):  
Hannah L. Smith ◽  
Stephen A. Beers ◽  
Juliet C. Gray ◽  
Janos M. Kanczler
Keyword(s):  
Three Dimensional ◽  
Chorioallantoic Membrane ◽  
3D Models ◽  
In Ovo ◽  
Future Directions ◽  
3 Dimensional ◽  
In Vivo Animal Models

Treatment for osteosarcoma (OS) has been largely unchanged for several decades, with typical therapies being a mixture of chemotherapy and surgery. Although therapeutic targets and products against cancer are being continually developed, only a limited number have proved therapeutically active in OS. Thus, the understanding of the OS microenvironment and its interactions are becoming more important in developing new therapies. Three-dimensional (3D) models are important tools in increasing our understanding of complex mechanisms and interactions, such as in OS. In this review, in vivo animal models, in vitro 3D models and in ovo chorioallantoic membrane (CAM) models, are evaluated and discussed as to their contribution in understanding the progressive nature of OS, and cancer research. We aim to provide insight and prospective future directions into the potential translation of 3D models in OS.

Insights into bread melanoidins: fate in the upper digestive tract and impact on the gut microbiota using in vitro systems

2015 ◽  
Vol 6 (12) ◽  
pp. 3737-3745 ◽  
Author(s):  
Cynthia Helou ◽  
Sylvain Denis ◽  
Madeleine Spatz ◽  
David Marier ◽  
Véronique Rame ◽  
...  
Keyword(s):  
Small Intestine ◽  
Gut Microbiota ◽  
Digestive Tract ◽  
Batch Fermentation ◽  
Dramatic Decrease ◽  
Upper Digestive Tract ◽  
In Vitro Systems

Bread melanoidins are partially degraded in the small intestine and induce a dramatic decrease of enterobacteria during batch fermentation.

scholarly journals Three-dimensional migration of macrophages requires Hck for podosome organization and extracellular matrix proteolysis

Blood ◽  
2010 ◽  
Vol 115 (7) ◽  
pp. 1444-1452 ◽  
Author(s):  
Céline Cougoule ◽  
Véronique Le Cabec ◽  
Renaud Poincloux ◽  
Talal Al Saati ◽  
Jean-Louis Mège ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Spatial Organization ◽  
Ectopic Expression ◽  
Three Dimensional ◽  
Matrix Metalloproteases ◽  
3 Dimensional ◽  
Normal Expression ◽  
Migration Of Macrophages

Abstract Tissue infiltration of phagocytes exacerbates several human pathologies including chronic inflammations or cancers. However, the mechanisms involved in macrophage migration through interstitial tissues are poorly understood. We investigated the role of Hck, a Src-family kinase involved in the organization of matrix adhesion and degradation structures called podosomes. In Hck−/− mice submitted to peritonitis, we found that macrophages accumulated in interstitial tissues and barely reached the peritoneal cavity. In vitro, 3-dimensional (3D) migration and matrix degradation abilities, 2 protease-dependent properties of bone marrow–derived macrophages (BMDMs), were affected in Hck−/− BMDMs. These macrophages formed few and undersized podosome rosettes and, consequently, had reduced matrix proteolysis operating underneath despite normal expression and activity of matrix metalloproteases. Finally, in fibroblasts unable to infiltrate matrix, ectopic expression of Hck provided the gain–of–3D migration function, which correlated positively with formation of podosome rosettes. In conclusion, spatial organization of podosomes as large rosettes, proteolytic degradation of extracellular matrix, and 3D migration appeared to be functionally linked and regulated by Hck in macrophages. Hck, as the first protein combining a phagocyte-limited expression with a role in 3D migration, could be a target for new anti-inflammatory and antitumor molecules.

scholarly journals In vitro modelling of the mucosa of the oesophagus and upper digestive tract

2021 ◽  
Vol 0 ◽  
pp. 0-0
Author(s):  
Kyle Stanforth ◽  
Peter Chater ◽  
Iain Brownlee ◽  
Matthew Wilcox ◽  
Chris Ward ◽  
...  
Keyword(s):  
Digestive Tract ◽  
Upper Digestive Tract ◽  
In Vitro Modelling

Imitating Hypoxia and Tumor Microenvironment with Immune Evasion by Employing Three Dimensional in vitro Cellular Models: Impressive Tool in Drug Discovery

2021 ◽  
Vol 16 ◽  
Author(s):  
Suman Kumar Ray ◽  
Sukhes Mukherjee
Keyword(s):  
Cell Culture ◽  
Tumor Microenvironment ◽  
Immune Evasion ◽  
Cancer Biology ◽  
Three Dimensional ◽  
3D Cell Culture ◽  
Cell Culture Techniques ◽  
Culture Techniques ◽  
Hypoxic Tumor

: The heterogeneous tumor microenvironment is exceptionally perplexing and not wholly comprehended. Different multifaceted alignments lead to the generation of oxygen destitute situations within the tumor niche that modulate numerous intrinsic tumor microenvironments. Disentangling these communications is vital for scheming practical therapeutic approaches that can successfully decrease tumor allied chemotherapy resistance by utilizing the innate capability of the immune system. Several research groups have concerned with a protruding role for oxygen metabolism along with hypoxia in the immunity of healthy tissue. Hypoxia in addition to hypoxia-inducible factors (HIFs) in the tumor microenvironment plays an important part in tumor progression and endurance. Although numerous hypoxia-focused therapies have shown promising outcomes both in vitro and in vivo these outcomes have not effectively translated into clinical preliminaries. Distinctive cell culture techniques have utilized as an in vitro model for tumor niche along with tumor microenvironment and proficient in more precisely recreating tumor genomic profiles as well as envisaging therapeutic response. To study the dynamics of tumor immune evasion, three-dimensional (3D) cell cultures are more physiologically important to the hypoxic tumor microenvironment. Recent research has revealed new information and insights into our fundamental understanding of immune systems, as well as novel results that have been established as potential therapeutic targets. There are a lot of patented 3D cell culture techniques which will be highlighted in this review. At present notable 3D cell culture procedures in the hypoxic tumor microenvironment, discourse open doors to accommodate both drug repurposing, advancement, and divulgence of new medications and will deliberate the 3D cell culture methods into standard prescription disclosure especially in the field of cancer biology which will be discussing here.

scholarly journals Human Adipose Derived Cells in Two- and Three-Dimensional Cultures: Functional Validation of an In Vitro Fat Construct

2020 ◽  
Vol 2020 ◽  
pp. 1-14
Author(s):  
Robert Bender ◽  
Michelle McCarthy ◽  
Theodore Brown ◽  
Joanna Bukowska ◽  
Stanley Smith ◽  
...  
Keyword(s):  
Adipogenic Differentiation ◽  
Three Dimensional ◽  
3D Culture ◽  
Stromal Vascular Fraction ◽  
The United States ◽  
Nonalcoholic Fatty Liver ◽  
3 Dimensional ◽  
Treatment Of Obesity ◽  
On Chip

Obesity, defined as a body mass index of 30 kg/m2 or above, has increased considerably in incidence and frequency within the United States and globally. Associated comorbidities including cardiovascular disease, type 2 diabetes mellitus, metabolic syndrome, and nonalcoholic fatty liver disease have led to a focus on the mechanisms promoting the prevention and treatment of obesity. Commonly utilized in vitro models employ human or mouse preadipocyte cell lines in a 2-dimensional (2D) format. Due to the structural, biochemical, and biological limitations of these models, increased attention has been placed on “organ on a chip” technologies for a 3-dimensional (3D) culture. Herein, we describe a method employing cryopreserved primary human stromal vascular fraction (SVF) cells and a human blood product-derived biological scaffold to create a 3D adipose depot in vitro. The “fat-on-chip” 3D cultures have been validated relative to 2D cultures based on proliferation, flow cytometry, adipogenic differentiation, confocal microscopy/immunofluorescence, and functional assays (adipokine secretion, glucose uptake, and lipolysis). Thus, the in vitro culture system demonstrates the critical characteristics required for a humanized 3D white adipose tissue (WAT) model.

scholarly journals In situ measurement of ECM rheology and microheterogeneity in embedded and overlaid 3D pancreatic tumor stroma co-cultures via passive particle tracking

2017 ◽  
Vol 10 (06) ◽  
pp. 1742003 ◽  
Author(s):  
Dustin P. Jones ◽  
William Hanna ◽  
Gwendolyn M. Cramer ◽  
Jonathan P. Celli
Keyword(s):  
Mechanical Properties ◽  
Tumor Growth ◽  
Particle Tracking ◽  
Cancer Biology ◽  
Three Dimensional ◽  
Tumor Stroma ◽  
Physical Force ◽  
Culture Models

Tumor growth is regulated by a diverse set of extracellular influences, including paracrine crosstalk with stromal partners, and biophysical interactions with surrounding cells and tissues.Studies elucidating the role of physical force and the mechanical properties of the extracellular matrix (ECM) itself as regulators of tumor growth and invasion have been greatly catalyzed by the use of in vitro three-dimensional (3D) tumor models. These systems provide the ability to systematically isolate, manipulate, and evaluate impact of stromal components and extracellular mechanics in a platform that is both conducive to imaging and biologically relevant. However, recognizing that mechanoregulatory crosstalk is bi-directional and fully utilizing these models requires complementary methods for in situ measurements of the local mechanical environment. Here, in 3D tumor/fibroblast co-culture models of pancreatic cancer, a disease characterized by its prominent stromal involvement, we evaluate the use of particle-tracking microrheology to probe dynamic mechanical changes. Using videos of fluorescently labeled polystyrene microspheres embedded in collagen I ECM, we measure spatiotemporal changes in the Brownian motion of probes to report local ECM shear modulus and microheterogeneity. This approach reveals stiffening of collagen in fibroblast co-cultures relative to cultures with cancer cells only, which exhibit degraded ECM with heterogeneous microstructure. We further show that these effects are dependent on culture geometry with contrasting behavior for embedded and overlay cultures. In addition to potential application to screening stroma-targeted therapeutics, this work also provides insight into how the composition and plating geometry impact the mechanical properties of 3D cell cultures that are increasingly widely used in cancer biology.

scholarly journals Electrochemical mapping of oxygenation in the three-dimensional multicellular tumour hemi-spheroid

2019 ◽  
Vol 475 (2225) ◽  
pp. 20180647 ◽  
Author(s):  
Disha B. Sheth ◽  
Miklόs Gratzl
Keyword(s):  
Cancer Biology ◽  
Normal Tissue ◽  
Microelectrode Array ◽  
Three Dimensional ◽  
Blood Capillaries ◽  
Drug Molecules ◽  
Tumour Spheroid ◽  
Lethal Level ◽  
Drug Concentrations

Blood capillaries deliver oxygen and nutrients to surrounding micro-regions of tissue and carry away metabolic waste. In normal tissue, capillaries are close enough to keep all the cells viable. In solid tumours, the capillary system is chaotic and typical inter-capillary distances are larger than in normal tissue. Therefore, hypoxic regions develop. Drug molecules may not reach these areas at concentrations above the lethal level. The combined effect of low drug concentrations and local hypoxia, often exacerbated by acidity, leads to therapy failure. To better understand the interplay between hypoxia and poor drug penetration, oxygenation needs to be assessed in different areas of inter-capillary tissue. The multicellular tumour spheroid is a well-established three-dimensional (3D) in vitro model of the capillary microenvironment. It is used to mimic nascent tumours and micro-metastases as well. In this work, we demonstrate for the first time that dynamic intra-spheroidal oxygen maps can be obtained at the 3D multicellular tumour hemi-spheroid (MCH) using a non-invasive microelectrode array. The same oxygen distributions exist inside the equivalent but less accessible full spheroid. The MCH makes high throughput—high content analysis of spheroids feasible and thus can assist studies on basic cancer biology, drug development and personalized medicine.

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