Browsing by Author "Kinstlinger, Ian S."

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    A 3D printable perfused hydrogel vascular model to assay ultrasound-induced permeability
    (Royal Society of Chemistry, 2022) Royse, Madison K.; Means, A. Kristen; Calderon, Gisele A.; Kinstlinger, Ian S.; He, Yufang; Durante, Marc R.; Procopio, Adam T.; Veiseh, Omid; Xu, Jun
    The development of an in vitro model to study vascular permeability is vital for clinical applications such as the targeted delivery of therapeutics. This work demonstrates the use of a perfusion-based 3D printable hydrogel vascular model as an assessment for endothelial permeability and its barrier function. Aside from providing a platform that more closely mimics the dynamic vascular conditions in vivo, this model enables the real-time observation of changes in the endothelial monolayer during the application of ultrasound to investigate the downstream effect of ultrasound-induced permeability. We show an increase in the apparent permeability coefficient of a fluorescently labeled tracer molecule after ultrasound treatment via a custom MATLAB algorithm, which implemented advanced features such as edge detection and a dynamic region of interest, thus supporting the use of ultrasound as a non-invasive method to enhance vascular permeability for targeted drug therapies. Notably, live-cell imaging with VE-cadherin-GFP HUVECs provides some of the first real-time acquisitions of the dynamics of endothelial cell–cell junctions under the application of ultrasound in a 3D perfusable model. This model demonstrates potential as a new scalable platform to investigate ultrasound-assisted delivery of therapeutics across a cellular barrier that more accurately mimics the physiologic matrix and fluid dynamics.
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    Engineered tissues supported by convection and diffusion through dendritic vascular networks
    (2020-10-22) Kinstlinger, Ian S.; Miller, Jordan S
    Metabolic function in mammalian tissues is sustained by the delivery of oxygen and nutrients as well as the removal of waste through complex, three-dimensional (3D) networks of hierarchically organized blood vessels. However, fabrication of such 3D vascular networks within soft hydrogels remains one of the greatest challenges in tissue engineering. Sacrificial templates have proven useful for patterning perfusable vascular networks in engineered tissues, but such templates have been constrained in architectural complexity by limitations in the techniques which have been used to fabricate them. We hypothesized that these architectural limitations could be overcome by creating sacrificial vascular templates via selective laser sintering (SLS), an additive manufacturing process which uses a laser to fabricate solid structures from powdered raw materials. We developed an open-source SLS system and demonstrated its capacity to pattern biomimetic scale models of vascular topology. To adapt SLS fabrication for biocompatible and water-soluble materials which could be used sacrificially in the presence of cells, we identified carbohydrate powders formulations which are compatible with SLS and demonstrated laser sintering of carbohydrates into elaborate branched structures, including algorithmically-generated biomimetic branching networks which we term dendritic networks. Laser sintered carbohydrate templates were used to pattern perfusable vascular networks in a range of materials including natural and synthetically-derived biocompatible hydrogels, which can support cells in both the lumenal and parenchymal spaces. We leveraged this methodology to establish a complete pipeline encompassing generative vascular design, additive fabrication, perfusion culture, and volumetric spatial analysis of tissue performance. We identify heterogeneous zones of metabolic activity that emerge in perfused cell-laden hydrogels and we demonstrate that dendritic vascular networks can sustain cell metabolism deep within model tissues greater than 1 cm thick. We also seed endothelial cells, characterize convective transport through dendritic networks, and explore strategies to modulate the dynamics of changing cell densities within perfused gels. Finally, we demonstrate that perfusion culture through dendritic networks can support the survival and function of primary hepatocyte cultures. This approach for rapid design and biofabrication of engineered volumetric tissues offers an experimental strategy for interrogating the relationship between vascular network architecture, metabolite transport, and tissue function.
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    Open-Source Selective Laser Sintering (OpenSLS) of Nylon and Biocompatible Polycaprolactone
    (Public Library of Science, 2016) Kinstlinger, Ian S.; Bastian, Andreas; Paulsen, Samantha J.; Hwang, Daniel H.; Ta, Anderson H.; Yalacki, David R.; Schmidt, Tim; Miller, Jordan S.
    Selective Laser Sintering (SLS) is an additive manufacturing process that uses a laser to fuse powdered starting materials into solid 3D structures. Despite the potential for fabrication of complex, high-resolution structures with SLS using diverse starting materials (including biomaterials), prohibitive costs of commercial SLS systems have hindered the wide adoption of this technology in the scientific community. Here, we developed a low-cost, open-source SLS system (OpenSLS) and demonstrated its capacity to fabricate structures in nylon with sub-millimeter features and overhanging regions. Subsequently, we demonstrated fabrication of polycaprolactone (PCL) into macroporous structures such as a diamond lattice. Widespread interest in using PCL for bone tissue engineering suggests that PCL lattices are relevant model scaffold geometries for engineering bone. SLS of materials with large powder grain size (~500 μm) leads to part surfaces with high roughness, so we further introduced a simple vapor-smoothing technique to reduce the surface roughness of sintered PCL structures which further improves their elastic modulus and yield stress. Vapor-smoothed PCL can also be used for sacrificial templating of perfusable fluidic networks within orthogonal materials such as poly(dimethylsiloxane) silicone. Finally, we demonstrated that human mesenchymal stem cells were able to adhere, survive, and differentiate down an osteogenic lineage on sintered and smoothed PCL surfaces, suggesting that OpenSLS has the potential to produce PCL scaffolds useful for cell studies. OpenSLS provides the scientific community with an accessible platform for the study of laser sintering and the fabrication of complex geometries in diverse materials.