2018
2019
Journal Articles
Fish, the better model in human heart research? Zebrafish Heart aggregates as a 3D spontaneously cardiomyogenic in vitro model system
Patricia Hodgson 1, Jake Ireland 2, Bianka Grunow 3
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Abstract
The zebrafish (ZF) has become an essential model for biomedical, pharmacological and eco-toxicological heart research. Despite the anatomical differences between fish and human hearts, similarities in cellular structure and conservation of genes as well as pathways across vertebrates have led to an increase in the popularity of ZF as a model for human cardiac research. ZF research benefits from an entirely sequenced genome, which allows us to establish and study cardiovascular mutants to better understand cardiovascular diseases. In this review, we will discuss the importance of in vitro model systems for cardiac research and summarise results of in vitro 3D heart-like cell aggregates, consisting of myocardial tissue formed spontaneously from enzymatically digested whole embryonic ZF larvae (Zebrafish Heart Aggregate - ZFHA). We will give an overview of the similarities and differences of ZF versus human hearts and highlight why ZF complement established mammalian models (i.e. murine and large animal models) for cardiac research. At this stage, the ZFHA model system is being refined into a high-throughput (more ZFHA generated than larvae prepared) and stable in vitro test system to accomplish the same longevity of previously successful salmonid models. ZFHA have potential for the use of high-throughput-screenings of different factors like small molecules, nucleic acids, proteins and lipids which is difficult to achieve in the zebrafish in vivo screening models with lethal mutations as well as to explore ion channel disorders and to find appropriate drugs for safety screening.
Porous Chitosan Films Support Stem Cells and Facilitate Sutureless Tissue Repair
Herleen Ruprai, Sara Romanazzo, Jake Ireland, Kristopher Kilian, Damia Mawad, Laurel George, Richard Wuhrer, Jessica Houang, Daniel Ta, Simon Myers, and Antonio Lauto*
​Abstract
Photochemical tissue bonding with chitosan-based adhesive films is an experimental surgical technique that avoids the risk of thermal tissue injuries and the use of sutures to maintain strong tissue connection. This technique is advantageous over other tissue repair methods as it is minimally invasive and does not require mixing of multiple components before or during application. To expand the capability of the film to beyond just a tissue bonding device and promote tissue regeneration, in this study, we designed bioadhesive films that could also support stem cells. The films were modified with oligomeric chitosan to tune their erodibility and made porous through freeze-drying for better tissue integration. Of note, porous adhesive films (pore diameter ∼110 μm), with 10% of the chitosan being oligomeric, could retain similar tissue bonding strengths (13–15 kPa) to that of the nonporous chitosan-based adhesives used in previous studies when photoactivated. When tested in vitro, these films exhibited a mass loss of ∼20% after 7 days, swelling ratios of ∼270–300%, a percentage elongation of ∼90%, and both a tensile strength and Young’s modulus of ∼1 MPa. The physical properties of the films were suitable for maintaining the viability and multipotency of bone-marrow-derived human mesenchymal stem cells over the duration of culture. Thus, these biocompatible, photoactivated porous, and erodible adhesive films show promise for applications in controlled cell delivery and regenerative medicine.
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2021
Magnetic Nanocomposite Hydrogels for Directing Myofibroblast Activity in Adipose-Derived Stem Cells
Md Shariful Islam, Thomas G. Molley, Jake Ireland, Jamie J. Kruzic, Kristopher A. Kilian
​Abstract
Dynamic cell-culture materials that can change mechanical properties in response to extrinsic stimuli are emerging as promising tools for cell and tissue engineering research. However, most of these techniques involve a one-way stiffening or softening through changes in the materials chemistry, which does not allow reversibility. Here, the incorporation of superparamagnetic iron-oxide nanoparticles within poly(ethylene glycol) hydrogels as dynamic cell culture materials is demonstrated. Using simple permanent magnets and adipose-derived stem cells, a near twofold increase in cell spread area and an accompanying 20% enrichment in cells expressing alpha-smooth muscle actin is seen. This platform provides a means to study relationships between dynamic stiffening and cell behavior, using permanent magnets and clinically viable composite materials, with scope for use as a tool to enrich the myofibroblast population in stromal cells.
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Defined microenvironments trigger in vitro gastrulation in human pluripotent stem cells
Srivastava P, Romanazzo S, Ireland J, Nemec S, Molley TG, Jayathilaka P, Pandzic E, Yeola A, Chandrakanthan V, Pimanda J, Kilian K
​Abstract
Embryogenesis is orchestrated through local morphogen gradients and endometrial constraints that give rise to the three germ layers in a well-defined assembly. In vitro models of embryogenesis have been demonstrated by treating pluripotent stem cells in adherent or suspension culture with soluble morphogens and small molecules, which leads to tri-lineage differentiation. However, treatment with exogenous agents override the subtle spatiotemporal changes observed in vivo that ultimately underly the human body plan. Here we demonstrate how microconfinement of pluripotent stem cells on hydrogel substrates catalyses gastrulation-like events without the need for supplements. Within six hours of initial seeding, cells at the boundary show elevated cytoskeletal tension and yes-associated protein (YAP) activity, which leads to changes in cell and nuclear morphology, epithelial to mesenchymal transition, and emergence of defined patterns of primitive streak containing SRY-Box Transcription Factor 17 (SOX17) + T/BRACHYURY + cells. Immunofluorescence staining, transcript analysis, and the use of pharmacological modulators reveal a role for mechanotransduction-coupled non-canonical wingless-type (WNT) signalling in promoting epithelial to mesenchymal transition and multilayered organization within the colonies. These microscale gastruloids were removed from the substrate and encapsulated in 3D hydrogels, where biomaterials properties correspond to maintenance and spatial positioning of the primitive streak. Together, this approach demonstrates how materials alone can nurture embryonic gastrulation, thereby providing an in vitro model of early development.
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