scholarly journals Self-Assembling Peptide Hydrogels - PeptiGels® as a Platform for Hepatic Organoid Culture

2021 ◽  
Author(s):  
Adedamola Olayanju ◽  
Aline F Miller ◽  
Tahera Ansari ◽  
Christopher E. Goldring
Keyword(s):  
Ex Vivo ◽  
3D Culture ◽  
Molecular Techniques ◽  
3 Dimensional ◽  
Culture Systems ◽  
Self Assembling ◽  
Technological Platform ◽  
Peptide Hydrogels

AbstractA major challenge in advancing preclinical studies is the lack of robust in vitro culture systems that fully recapitulate the in vivo scenario together with limited clinical translational to humans. Organoids, as 3-dimensional (3D) self-replicating structures are increasingly being shown as powerful models for ex vivo experimentation in the field of regenerative medicine and drug discovery. Organoid formation requires the use of extracellular matrix (ECM) components to provide a 3D platform. However, the most commonly used ECM, essential for maintaining organoid growth is Matrigel and is derived from a tumorigenic source which limits its translational ability. PeptiGels® which are self-assembling peptide hydrogels present as alternatives to traditional ECM for use in 3D culture systems. Synthetic PeptiGels® are non-toxic, biocompatible, biodegradable and can be tuneable to simulate different tissue microenvironments. In this study, we validated the use of different types of PeptiGels® for porcine hepatic organoid growth. Hepatic organoids were assessed morphologically and using molecular techniques to determine the optimum PeptiGel® formulation. The outcome clearly demonstrated the ability of PeptiGel® to support organoid growth and offer themselves as a technological platform for 3D cultured physiologically and clinically relevant data.

scholarly journals From Clones to Buds and Branches: The Use of Lung Organoids to Model Branching Morphogenesis Ex Vivo

2021 ◽  
Vol 9 ◽  
Author(s):  
Ana Ivonne Vazquez-Armendariz ◽  
Susanne Herold
Keyword(s):  
Cell Fate ◽  
Molecular Mechanisms ◽  
Ex Vivo ◽  
3D Culture ◽  
Cell Types ◽  
Branching Morphogenesis ◽  
Cell Fate Decisions ◽  
Culture Systems

Three-dimensional (3D) organoid culture systems have rapidly emerged as powerful tools to study organ development and disease. The lung is a complex and highly specialized organ that comprises more than 40 cell types that offer several region-specific roles. During organogenesis, the lung goes through sequential and morphologically distinctive stages to assume its mature form, both structurally and functionally. As branching takes place, multipotent epithelial progenitors at the distal tips of the growing/bifurcating epithelial tubes progressively become lineage-restricted, giving rise to more differentiated and specialized cell types. Although many cellular and molecular mechanisms leading to branching morphogenesis have been explored, deeper understanding of biological processes governing cell-fate decisions and lung patterning is still needed. Given that these distinct processes cannot be easily analyzedin vivo, 3D culture systems have become a valuable platform to study organogenesisin vitro. This minireview focuses on the current lung organoid systems that recapitulate developmental events occurring before and during branching morphogenesis. In addition, we also discuss their limitations and future directions.

scholarly journals To Better Generate Organoids, What Can We Learn From Teratomas?

2021 ◽  
Vol 9 ◽  
Author(s):  
Hongyu Li ◽  
Lixiong Gao ◽  
Jinlin Du ◽  
Tianju Ma ◽  
Zi Ye ◽  
...  
Keyword(s):  
Animal Models ◽  
Disease Modeling ◽  
3D Culture ◽  
Cellular Heterogeneity ◽  
Developmental Studies ◽  
Genomic Profile ◽  
Organization Studies ◽  
Culture Systems

The genomic profile of animal models is not completely matched with the genomic profile of humans, and 2D cultures do not represent the cellular heterogeneity and tissue architecture found in tissues of their origin. Derived from 3D culture systems, organoids establish a crucial bridge between 2D cell cultures and in vivo animal models. Organoids have wide and promising applications in developmental research, disease modeling, drug screening, precision therapy, and regenerative medicine. However, current organoids represent only single or partial components of a tissue, which lack blood vessels, native microenvironment, communication with near tissues, and a continuous dorsal-ventral axis within 3D culture systems. Although efforts have been made to solve these problems, unfortunately, there is no ideal method. Teratoma, which has been frequently studied in pathological conditions, was recently discovered as a new in vivo model for developmental studies. In contrast to organoids, teratomas have vascularized 3D structures and regions of complex tissue-like organization. Studies have demonstrated that teratomas can be used to mimic multilineage human development, enrich specific somatic progenitor/stem cells, and even generate brain organoids. These results provide unique opportunities to promote our understanding of the vascularization and maturation of organoids. In this review, we first summarize the basic characteristics, applications, and limitations of both organoids and teratomas and further discuss the possibility that in vivo teratoma systems can be used to promote the vascularization and maturation of organoids within an in vitro 3D culture system.

Abstract 1717: Circulating tumor cells from in vitro 3D culture systems have tumor-initiating capacity in vivo

2016 ◽  
Author(s):  
Marina C. Cabrera ◽  
Elaine Hurt ◽  
Xiaoru Chen ◽  
Shi Xiaoqing ◽  
Haifeng Bao
Keyword(s):  
Tumor Cells ◽  
Circulating Tumor Cells ◽  
3D Culture ◽  
Culture Systems

scholarly journals Current Advances in 3D Tissue and Organ Reconstruction

2021 ◽  
Vol 22 (2) ◽  
pp. 830
Author(s):  
Georgia Pennarossa ◽  
Sharon Arcuri ◽  
Teresina De Iorio ◽  
Fulvio Gandolfi ◽  
Tiziana A. L. Brevini
Keyword(s):  
Cell Biology ◽  
Drug Testing ◽  
Three Dimensional ◽  
3D Culture ◽  
In Vitro Models ◽  
The Body ◽  
Cellular Functions ◽  
Culture Systems

Bi-dimensional culture systems have represented the most used method to study cell biology outside the body for over a century. Although they convey useful information, such systems may lose tissue-specific architecture, biomechanical effectors, and biochemical cues deriving from the native extracellular matrix, with significant alterations in several cellular functions and processes. Notably, the introduction of three-dimensional (3D) platforms that are able to re-create in vitro the structures of the native tissue, have overcome some of these issues, since they better mimic the in vivo milieu and reduce the gap between the cell culture ambient and the tissue environment. 3D culture systems are currently used in a broad range of studies, from cancer and stem cell biology, to drug testing and discovery. Here, we describe the mechanisms used by cells to perceive and respond to biomechanical cues and the main signaling pathways involved. We provide an overall perspective of the most recent 3D technologies. Given the breadth of the subject, we concentrate on the use of hydrogels, bioreactors, 3D printing and bioprinting, nanofiber-based scaffolds, and preparation of a decellularized bio-matrix. In addition, we report the possibility to combine the use of 3D cultures with functionalized nanoparticles to obtain highly predictive in vitro models for use in the nanomedicine field.

Self-assembling Peptides: From Bio-inspired Materials to Bone Regeneration

2008 ◽  
Vol 87 (7) ◽  
pp. 606-616 ◽  
Author(s):  
C. E. Semino
Keyword(s):  
Bone Regeneration ◽  
Nerve Repair ◽  
Three Dimensional ◽  
3D Culture ◽  
Functional Differentiation ◽  
Therapeutic Drug ◽  
Functional Requirements ◽  
Self Assembling

In recent years, the development of new biomaterials with specifications for tissue and organ functional requirements—such as proper biological, structural, and biomechanical properties as well as designed control for biodegradation and therapeutic drug-release capacity—is the main aim of many academic and industrial programs. Hence, the concept of molecular self-assembly is the driving force for the development of new biomaterials that support the growth and functional differentiation of cells and tissues in a controlled manner. The discovery, properties, and development of self-assembling peptides to be used as three-dimensional (3D) scaffolds based on their similarity (in structure and mechanical features) to extracellular matrices are described. Self-assembling peptides can be used for in vitro applications for cell 3D culture as well as in vivo for tissue regeneration such as bone and optical nerve repair, as well as for drug delivery of mediators to improve therapy, as in the case of myocardial infarction. Finally, the use of self-assembling materials in combination with a bioengineering platform is proposed to assist functional bone regeneration in cases of larger bone defects, including exposed fractures due to trauma and spinal disorders dealing with high loadings, as well as replacement of big bone structures due to tumors.

scholarly journals Bioengineered 3D Glial Cell Culture Systems and Applications for Neurodegeneration and Neuroinflammation

2017 ◽  
Vol 22 (5) ◽  
pp. 583-601 ◽  
Author(s):  
P. Marc D. Watson ◽  
Edel Kavanagh ◽  
Gary Allenby ◽  
Matthew Vassey
Keyword(s):  
Cell Culture ◽  
Drug Discovery ◽  
Glial Cell ◽  
Critical Role ◽  
3D Culture ◽  
Cell Types ◽  
Culture Systems ◽  
Cellular Environment

Neurodegeneration and neuroinflammation are key features in a range of chronic central nervous system (CNS) diseases such as Alzheimer’s and Parkinson’s disease, as well as acute conditions like stroke and traumatic brain injury, for which there remains significant unmet clinical need. It is now well recognized that current cell culture methodologies are limited in their ability to recapitulate the cellular environment that is present in vivo, and there is a growing body of evidence to show that three-dimensional (3D) culture systems represent a more physiologically accurate model than traditional two-dimensional (2D) cultures. Given the complexity of the environment from which cells originate, and their various cell–cell and cell–matrix interactions, it is important to develop models that can be controlled and reproducible for drug discovery. 3D cell models have now been developed for almost all CNS cell types, including neurons, astrocytes, microglia, and oligodendrocyte cells. This review will highlight a number of current and emerging techniques for the culture of astrocytes and microglia, glial cell types with a critical role in neurodegenerative and neuroinflammatory conditions. We describe recent advances in glial cell culture using electrospun polymers and hydrogel macromolecules, and highlight how these novel culture environments influence astrocyte and microglial phenotypes in vitro, as compared to traditional 2D systems. These models will be explored to illuminate current trends in the techniques used to create 3D environments for application in research and drug discovery focused on astrocytes and microglial cells.

A 3 Dimensional in Vitro Model to Test HL-60 Cell Line Sensitivity to Chemotherapy

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4882-4882
Author(s):  
Omar S. Aljitawi ◽  
Dandan Li ◽  
Da Zhang ◽  
Jonathan Mahnken ◽  
Suman Kambhampati ◽  
...  
Keyword(s):  
Cell Culture ◽  
Bone Marrow ◽  
3D Culture ◽  
Culture Conditions ◽  
Cytotoxic Chemotherapy ◽  
3 Dimensional ◽  
3D Microenvironment ◽  
3D Culture System

Abstract Abstract 4882 Introduction: Current in vitro drug testing models are based on 2-dimensional (2D) cell culture systems and therefore do not always predict in vivo responses. This lack of predictability of the 2D assays is believed to be related to the 3-dimensional (3D) microenvironment present in tissues or tumors. This 3D microenvironment, were cell-cell and cell-extracellular matrix (ECM) interactions occur, is fundamental for cell biologic activities. This is especially true for acute myeloid leukemia, were current 2-D cell culture models do not always predict clinical responses. This discrepancy in leukemia cell responses to chemotherapy in vivo, in comparison to in vitro, is at least partly related to leukemia cells interaction with the bone marrow microenvironment and their ability to establish niches. These niches offer partial protection from the effects of cytotoxic chemotherapy, otherwise termed cell adhesion-mediated drug resistance. In these experiments, we investigate the apoptotic effects of cytotoxic chemotherapy on HL-60 cell line cultured in a designed 3D AML cell culture model. In this 3D microenvironment, HL-60 cells were co-cultured with ex vivo expanded bone marrow mesenchyaml stem cells in a 3D synthetic scaffold. Aim: To examine the apoptotic effect of cytotoxic chemotherapy on HL-60 co-cultured with human bone marrow mesenchymal stem cells (huBM-MSCs) in 3D conditions. Methods: After several passages, expanded huBM-MSCs were seeded into PGA/PLLA 90/10 copolymer discs, 5-mm in diameter and 2-mm in thickness and allowed to attach to scaffold fibers and to expand over 2 weeks. Then, HL-60 were added and allowed to grow in the 3D culture system for another 10 days. HL-60 cells in 3D culture system were then exposed to doxorubicin given in two concentrations (25 and 50 μM) and incubated for 24 hours. HL-60 were then retrieved applying a combination of mechanical forces and using cell dissociation solution. FITC Annexin V Apoptosis Detection Kit was used to determine apoptosis. Apoptosis was confirmed by TUNEL assay. Proliferation of HL-60 cells in the 3D scaffold was assessed using Ki-67 stain of scaffold's cryosections. All tests were done in triplicates, and untreated HL-60 served as controls for treatment. Comparison was made with HL-60 cells alone and with HL-60 cells growing on a hu-BM-MSC monolayer. SAS version 9.2 (SAS Institute, Inc., 2002–2008) was used for statistical analysis Results: Virtually, all HL-60 cells treated with 25 or 50 μM underwent late apoptosis. Around.03% of HL-60 cells survived 25 μM concentration, none, however, survived 50 μM concentration. In 2D, most of HL-60 cells underwent necrosis, and to lesser extent late apoptosis. In sharp contrast, 17.8% of HL-60 cells survived 25μM concentration, nevertheless, only.27% of HL-60 cells treated with 50 μM concentration survived. The differences in apoptosis patterns between the three groups was statistically significant (P<.0001). Conclusion: compared to traditional cell culture conditions, the designed 3D culture conditions protected a higher percentage of HL-60 cells from undergoing apoptosis and necrosis. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Comparative Molecular Analysis of Cancer Behavior Cultured In Vitro, In Vivo, and Ex Vivo

Cancers ◽  
2020 ◽  
Vol 12 (3) ◽  
pp. 690 ◽  
Author(s):  
Nicholas R. Hum ◽  
Aimy Sebastian ◽  
Sean F. Gilmore ◽  
Wei He ◽  
Kelly A. Martin ◽  
...  
Keyword(s):  
Cancer Cell ◽  
Ex Vivo ◽  
Significant Proportion ◽  
3D Culture ◽  
Cell Phenotype ◽  
Primary Tumors ◽  
Altered Expression ◽  
Mesenchymal Transition

Current pre-clinical models of cancer fail to recapitulate the cancer cell behavior in primary tumors primarily because of the lack of a deeper understanding of the effects that the microenvironment has on cancer cell phenotype. Transcriptomic profiling of 4T1 murine mammary carcinoma cells from 2D and 3D cultures, subcutaneous or orthotopic allografts (from immunocompetent or immunodeficient mice), as well as ex vivo tumoroids, revealed differences in molecular signatures including altered expression of genes involved in cell cycle progression, cell signaling and extracellular matrix remodeling. The 3D culture platforms had more in vivo-like transcriptional profiles than 2D cultures. In vivo tumors had more cells undergoing epithelial-to-mesenchymal transition (EMT) while in vitro cultures had cells residing primarily in an epithelial or mesenchymal state. Ex vivo tumoroids incorporated aspects of in vivo and in vitro culturing, retaining higher abundance of cells undergoing EMT while shifting cancer cell fate towards a more mesenchymal state. Cellular heterogeneity surveyed by scRNA-seq revealed that ex vivo tumoroids, while rapidly expanding cancer and fibroblast populations, lose a significant proportion of immune components. This study emphasizes the need to improve in vitro culture systems and preserve syngeneic-like tumor composition by maintaining similar EMT heterogeneity as well as inclusion of stromal subpopulations.

scholarly journals Distinct niches within the extracellular matrix dictate fibroblast function in (cell free) 3D lung tissue cultures

2018 ◽  
Vol 314 (5) ◽  
pp. L708-L723 ◽  
Author(s):  
Gerald Burgstaller ◽  
Arunima Sengupta ◽  
Sarah Vierkotten ◽  
Gerhard Preissler ◽  
Michael Lindner ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Lung Tissue ◽  
Ex Vivo ◽  
3D Culture ◽  
Focal Adhesions ◽  
3D Cell Culture ◽  
Small Gtpases ◽  
Cellular Migration

Cues from the extracellular matrix (ECM) and their functional interplay with cells play pivotal roles for development, tissue repair, and disease. However, the precise nature of this interplay remains elusive. We used an innovative 3D cell culture ECM model by decellularizing 300-µm-thick ex vivo lung tissue scaffolds (d3D-LTCs) derived from diseased and healthy mouse lungs, which widely mimics the native (patho)physiological in vivo ECM microenvironment. We successfully repopulated all d3D-LTCs with primary human and murine fibroblasts, and moreover, we demonstrated that the cells also populated the innermost core regions of the d3D-LTCs in a real 3D fashion. The engrafted fibroblasts revealed a striking functional plasticity, depending on their localization in distinct ECM niches of the d3D-LTCs, affecting the cells’ tissue engraftment, cellular migration rates, cell morphologies, and protein expression and phosphorylation levels. Surprisingly, we also observed fibroblasts that were homing to the lung scaffold’s interstitium as well as fibroblasts that were invading fibrotic areas. To date, the functional nature and even the existence of 3D cell matrix adhesions in vivo as well as in 3D culture models is still unclear and controversial. Here, we show that attachment of fibroblasts to the d3D-LTCs evidently occurred via focal adhesions, thus advocating for a relevant functional role in vivo. Furthermore, we found that protein levels of talin, paxillin, and zyxin and phosphorylation levels of paxillin Y118, as well as the migration-relevant small GTPases RhoA, Rac, and CDC42, were significantly reduced compared with their attachment to 2D plastic dishes. In summary, our results strikingly indicate that inherent physical or compositional characteristics of the ECM act as instructive cues altering the functional behavior of engrafted cells. Thus, d3D-LTCs might aid to obtain more realistic data in vitro, with a high relevance for drug discovery and mechanistic studies alike.

scholarly journals Collagen hydrogel confinement of amyloid-β accelerates aggregation and reduces cytotoxic effects

2019 ◽  
Author(s):  
Laura W. Simpson ◽  
Gregory L. Szeto ◽  
Hacene Boukari ◽  
Theresa A. Good ◽  
Jennie B. Leach
Keyword(s):  
3D Culture ◽  
Amyloid Β ◽  
Correlation Spectroscopy ◽  
Great Promise ◽  
Culture Systems ◽  
Glass Substrates ◽  
Collagen Hydrogel ◽  
Β Sheet

AbstractAlzheimer’s disease (AD) is the most common form of dementia and is associated with the accumulation of amyloid-β (Aβ), a peptide whose aggregation has been associated with neurotoxicity. Drugs targeting Aβ have shown great promise in 2D in vitro models and mouse models, yet preclinical and clinical trials for AD have been highly disappointing. We propose that current in vitro culture systems for discovering and developing AD drugs have significant limitations; specifically, that Aβ aggregation is vastly different in these 2D cultures carried out on flat plastic or glass substrates vs. in a 3D environment, such as brain tissue, where Aβ confinement very likely alters aggregation kinetics and thermodynamics. In this work, we identified attenuation of Aβ cytotoxicity in 3D hydrogel culture compared to 2D cell culture. We investigated Aβ structure and aggregation in solution vs. hydrogel using Transmission Electron Microscopy (TEM), Fluorescence Correlation Spectroscopy (FCS), and Thioflavin T (ThT) assays. Our results reveal that the equilibrium is shifted to stable β-sheet aggregates in hydrogels and away from the relatively unstable/unstructured presumed toxic oligomeric Aβ species in solution. Volume exclusion imparted by hydrogel confinement stabilizes unfolded, presumably toxic species, promoting stable extended β-sheet fibrils. These results, taken together with the many recent reports that 3D hydrogel cell cultures enable cell morphologies and epigenetic changes that are more similar to cells in vivo compared to 2D cultures, strongly suggest that AD drugs should be tested in 3D culture systems as a step along the development pathway towards new, more effective therapeutics.

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