A novel organ-chip system emulates three-dimensional architecture of the human epithelia and the mechanical forces acting on it.

Successful translation of in vivo experimental data to human patients is an unmet need and a bottleneck in the development of effective therapeutics. Organ-on-Chip technology aims to address this need by leveraging recent significant advancements in microfabrication and biomaterials, which enable modeling of organs and their functionality. These microengineered chips offer researchers the possibility to recreate critical elements of native tissue architecture such as in vivo relevant tissue-tissue interface, air-liquid interface, and mechanical forces, including mechanical stretch and fluidic shear stress, which are crucial to recapitulate tissue level functions. Here, we present the development of a new, comprehensive 3D cell-culture system, where we combined our proprietary Organ-Chip technology with the advantages offered by three-dimensional organotypic culture. Leveraging microfabrication techniques, we engineered a flexible chip that consists of a chamber containing an organotypic epithelium, surrounded by two vacuum channels that can be actuated to stretch the hydrogel throughout its thickness. Furthermore, the ceiling of this chamber is a removable lid with a built-in microchannel that can be perfused with liquid or air and removed as needed for direct access to the tissue. The bottom part of this chamber is made from a porous flexible membrane which allows diffusive mass transport to and from the microfluidic channel positioned below the membrane. This additional microfluidic channel can be coated with endothelial cells to emulate a blood vessel and recapitulate endothelial interactions. Our results show that the Open-Top Chip design successfully addresses common challenges associated with the Organs-on-Chip technology, including the capability to incorporate a tissue-specific extracellular matrix gel seeded with primary stromal cells, to reproduce the architectural complexity of tissues by micropatterning the gel, and to extract the gel for H&E staining. We also provide proof-of-concept data on the feasibility of using the system with primary human skin and alveolar epithelial cells.

Robotic fluidic coupling and interrogation of multiple vascularized organ chips.

Organ chips can recapitulate organ-level (patho)physiology, yet pharmacokinetic and pharmacodynamic analyses require multi-organ systems linked by vascular perfusion. Here, we describe an 'interrogator' that employs liquid-handling robotics, custom software and an integrated mobile microscope for the automated culture, perfusion, medium addition, fluidic linking, sample collection and in situ microscopy imaging of up to ten organ chips inside a standard tissue-culture incubator. The robotic interrogator maintained the viability and organ-specific functions of eight vascularized, two-channel organ chips (intestine, liver, kidney, heart, lung, skin, blood-brain barrier and brain) for 3 weeks in culture when intermittently fluidically coupled via a common blood substitute through their reservoirs of medium and endothelium-lined vascular channels. We used the robotic interrogator and a physiological multicompartmental reduced-order model of the experimental system to quantitatively predict the distribution of an inulin tracer perfused through the multi-organ human-body-on-chips. The automated culture system enables the imaging of cells in the organ chips and the repeated sampling of both the vascular and interstitial compartments without compromising fluidic coupling.

Human iPSC-Derived Blood-Brain Barrier Chips Enable Disease Modeling and Personalized Medicine Applications.

The blood-brain barrier (BBB) tightly regulates the entry of solutes from blood into the brain and is disrupted in several neurological diseases. Using Organ-Chip technology, we created an entirely human BBB-Chip with induced pluripotent stem cell (iPSC)-derived brain microvascular endothelial-like cells (iBMECs), astrocytes, and neurons. The iBMECs formed a tight monolayer that expressed markers specific to brain vasculature. The BBB-Chip exhibited physiologically relevant transendothelial electrical resistance and accurately predicted blood-to-brain permeability of pharmacologics. Upon perfusing the vascular lumen with whole blood, the microengineered capillary wall protected neural cells from plasma-induced toxicity. Patient-derived iPSCs from individuals with neurological diseases predicted disease-specific lack of transporters and disruption of barrier integrity. By combining Organ-Chip technology and human iPSC-derived tissue, we have created a neurovascular unit that recapitulates complex BBB functions, provides a platform for modeling inheritable neurological disorders, and advances drug screening, as well as personalized medicine.

Bone-chip system to monitor osteogenic differentiation using optical imaging.

Human organoids and organ-on-chip systems to predict human responses to new therapies and for the understanding of disease mechanisms are being more commonly used in translational research. We have developed a bone-chip system to study osteogenic differentiation in vitro, coupled with optical imaging approach which provides the opportunity of monitoring cell survival, proliferation and differentiation in vitro without the need to terminate the culture. We used the mesenchymal stem cell (MSC) line over-expressing bone morphogenetic protein-2 (BMP-2), under Tet-Off system, and luciferase reporter gene under constitutive promoter. Cells were seeded on chips and supplemented with osteogenic medium. Flow of media was started 24 h later, while static cultures were performed using media reservoirs. Cells grown on the bone-chips under constant flow of media showed enhanced survival/proliferation, comparing to the cells grown in static conditions; luciferase reporter gene expression and activity, reflecting the cell survival and proliferation, was quantified using bioluminescence imaging and a significant advantage to the flow system was observed. In addition, the flow had positive effect on osteogenic differentiation, when compared with static cultures. Quantitative fluorescent imaging, performed using the osteogenic extra-cellular matrix-targeted probes, showed higher osteogenic differentiation of the cells under the flow conditions. Gene expression analysis of osteogenic markers confirmed the osteogenic differentiation of the MSC-BMP2 cells. Immunofluorescent staining performed against the Osteocalcin, Col1, and BSP markers illustrated robust osteogenic differentiation in the flow culture and lessened differentiation in the static culture. To sum, the bone-chip allows monitoring cell survival, proliferation, and osteogenic differentiation using optical imaging.

Human iPSC-Derived Endothelial Cells and Microengineered Organ-Chip Enhance Neuronal Development.

Human stem cell-derived models of development and neurodegenerative diseases are challenged by cellular immaturity in vitro. Microengineered organ-on-chip (or Organ-Chip) systems are designed to emulate microvolume cytoarchitecture and enable co-culture of distinct cell types. Brain microvascular endothelial cells (BMECs) share common signaling pathways with neurons early in development, but their contribution to human neuronal maturation is largely unknown. To study this interaction and influence of microculture, we derived both spinal motor neurons and BMECs from human induced pluripotent stem cells and observed increased calcium transient function and Chip-specific gene expression in Organ-Chips compared with 96-well plates. Seeding BMECs in the Organ-Chip led to vascular-neural interaction and specific gene activation that further enhanced neuronal function and in vivo-like signatures. The results show that the vascular system has specific maturation effects on spinal cord neural tissue, and the use of Organ-Chips can move stem cell models closer to an in vivo condition.