bims-orenst Biomed News
on Organs-on-chips and engineered stem cell models
Issue of 2021‒04‒25
seven papers selected by
Joram Mooiweer
University of Groningen

  1. Proc Natl Acad Sci U S A. 2021 Apr 27. pii: e2021569118. [Epub ahead of print]118(17):
      The de novo design of polar protein-protein interactions is challenging because of the thermodynamic cost of stripping water away from the polar groups. Here, we describe a general approach for designing proteins which complement exposed polar backbone groups at the edge of beta sheets with geometrically matched beta strands. We used this approach to computationally design small proteins that bind to an exposed beta sheet on the human transferrin receptor (hTfR), which shuttles interacting proteins across the blood-brain barrier (BBB), opening up avenues for drug delivery into the brain. We describe a design which binds hTfR with a 20 nM K d, is hyperstable, and crosses an in vitro microfluidic organ-on-a-chip model of the human BBB. Our design approach provides a general strategy for creating binders to protein targets with exposed surface beta edge strands.
    Keywords:  blood–brain barrier; computational protein design; drug delivery; neurological disease; transferrin receptor
  2. Lab Chip. 2021 Apr 20. 21(8): 1454-1474
      Drug development suffers from a lack of predictive and human-relevant in vitro models. Organ-on-chip (OOC) technology provides advanced culture capabilities to generate physiologically appropriate, human-based tissue in vitro, therefore providing a route to a predictive in vitro model. However, OOC technologies are often created at the expense of throughput, industry-standard form factors, and compatibility with state-of-the-art data collection tools. Here we present an OOC platform with advanced culture capabilities supporting a variety of human tissue models including liver, vascular, gastrointestinal, and kidney. The platform has 96 devices per industry standard plate and compatibility with contemporary high-throughput data collection tools. Specifically, we demonstrate programmable flow control over two physiologically relevant flow regimes: perfusion flow that enhances hepatic tissue function and high-shear stress flow that aligns endothelial monolayers. In addition, we integrate electrical sensors, demonstrating quantification of barrier function of primary gut colon tissue in real-time. We utilize optical access to the tissues to directly quantify renal active transport and oxygen consumption via integrated oxygen sensors. Finally, we leverage the compatibility and throughput of the platform to screen all 96 devices using high content screening (HCS) and evaluate gene expression using RNA sequencing (RNA-seq). By combining these capabilities in one platform, physiologically-relevant tissues can be generated and measured, accelerating optimization of an in vitro model, and ultimately increasing predictive accuracy of in vitro drug screening.
  3. Stem Cell Reports. 2021 Apr 16. pii: S2213-6711(21)00160-0. [Epub ahead of print]
      Neuromuscular junctions (NMJs) ensure communication between motor neurons (MNs) and muscle; however, in MN disorders, such as amyotrophic lateral sclerosis (ALS), NMJs degenerate resulting in muscle atrophy. The aim of this study was to establish a versatile and reproducible in vitro model of a human motor unit to investigate the effects of ALS-causing mutations. Therefore, we generated a co-culture of human induced pluripotent stem cell (iPSC)-derived MNs and human primary mesoangioblast-derived myotubes in microfluidic devices. A chemotactic and volumetric gradient facilitated the growth of MN neurites through microgrooves resulting in the interaction with myotubes and the formation of NMJs. We observed that ALS-causing FUS mutations resulted in reduced neurite outgrowth as well as an impaired neurite regrowth upon axotomy. NMJ numbers were likewise reduced in the FUS-ALS model. Interestingly, the selective HDAC6 inhibitor, Tubastatin A, improved the neurite outgrowth, regrowth, and NMJ morphology, prompting HDAC6 inhibition as a potential therapeutic strategy for ALS.
    Keywords:  FUS; HDAC6; Tubastatin A; amyotrophic lateral sclerosis; microfluidic device; neurite outgrowth; neurite regrowth; neuromuscular junction
  4. Front Bioeng Biotechnol. 2021 ;9 620128
      Angiogenesis is a complex, multicellular process that involves bidirectional interactions between extracellular matrix (ECM) and collectively invading endothelial cell (EC) sprouts that extend the microvasculature during development, wound healing, and disease processes. While many aspects of angiogenesis have been well studied, the relationship between endothelial sprout morphology and subsequent neovessel function remains relatively unknown. Here, we investigated how various soluble and physical matrix cues that regulate endothelial sprouting speed and proliferation correspond to changes in sprout morphology, namely, sprout stalk diameter. We found that sprout stalk cells utilize a combination of cytoskeletal forces and proteolysis to physically compact and degrade the surrounding matrix, thus creating sufficient space in three-dimensional (3D) ECM for lateral expansion. As increasing sprout diameter precedes lumenization to generate perfusable neovessels, this work highlights how dynamic endothelial stalk cell-ECM interactions promote the generation of functional neovessels during sprouting angiogenesis to provide insight into the design of vascularized, implantable biomaterials.
    Keywords:  angiogenesis; cell migration; cell proliferation; cytoskeletal forces; endothelial cell; extracellular matrix; microfluidic “lab-on-a-chip,”; proteolysis
  5. ACS Appl Mater Interfaces. 2021 Apr 20.
      Cancer is a multistep progressive disease that generally involves tumor growth, invasion, and metastasis. It is crucial to understand tumor progression for tumor diagnosis and therapy. However, tumor progression at an extremely early stage (EES) is barely demonstrated because EES tumors are too small to be detected by imaging. Herein, we, for the first time, replicated tumor progression at the EES on a microfluidic chip and uncovered the tumor behaviors affected by the tumor microenvironment. To mimic the progression of a single solid tumor at the EES, a HeLa cell spheroid was seeded and cultured on the chip, and a microvascular network was developed to integrate the microphysiological contexts around the tumor. We revealed not only the growth patterns and cell behaviors of tumor spheroids of different sizes under angiogenesis and fibroblast conditions but also the effect of tumor progression on peritumoral angiogenesis. We found that smaller tumors were more aggressive and that endotheliocytes and fibroblasts significantly accelerated both the proliferation and migration of tumor cells. In addition, we also first present the dynamic epithelial-mesenchymal transition process of tumor cells and the formation of vasculogenic mimicry at the EES. This work can provide insights for understanding tumor progression at the EES and offer new ideas for tumor therapy.
    Keywords:  angiogenesis; epithelial−mesenchymal transition; extremely early-stage tumor; microfluidic chip; vasculogenic mimicry
  6. Cell Mol Bioeng. 2021 Apr;14(2): 133-145
      Introduction: Neutrophils act as first responders during an infection, following signals from the pathogen as well as other host cells to migrate from blood vessels to the site of infection. This tightly regulated process is critical for pathogen clearance and, in many cases, eliminates the pathogen without the need for an additional immune response. It is, therefore, critical to understand what signals drive neutrophil migration to infection in a physiologically relevant environment.Methods: In this study, we used an infection-on-a-chip model to recapitulate many important aspects of the infectious microenvironment including an endothelial blood vessel, an extracellular matrix, and the environmental fungal pathogen Aspergillus fumigatus. We then used this model to visualize the innate immune response to fungal infection.
    Results: We found that A. fumigatus germination dynamics are influenced by the presence of an endothelial lumen. Furthermore, we demonstrated that neutrophils are recruited to and swarm around A. fumigatus hyphae and that the presence of monocytes significantly increases the neutrophil response to A. fumigatus. Using secreted protein analysis and blocking antibodies, we found that this increased migration is likely due to signaling by MIP-1 family proteins. Finally, we demonstrated that signal relay between neutrophils, mediated by LTB4 signaling, is also important for sustained neutrophil migration and swarming in response to A. fumigatus infection in our system.
    Conclusions: Taken together, these results suggest that paracrine signaling from both monocytes and neutrophils plays an important role in driving the neutrophil response to A. fumigatus.
    Keywords:  Endothelium; Fungal infection; Innate immunity; Microfluidics; Monocyte
  7. Sci Adv. 2021 Apr;pii: eabc1323. [Epub ahead of print]7(17):
      Existing three-dimensional (3D) culture techniques are limited by trade-offs between throughput, capacity for high-resolution imaging in living state, and geometric control. Here, we introduce a modular microscale hanging drop culture where simple design elements allow high replicates for drug screening, direct on-chip real-time or high-resolution confocal microscopy, and geometric control in 3D. Thousands of spheroids can be formed on our microchip in a single step and without any selective pressure from specific matrices. Microchip cultures from human LN229 glioblastoma and patient-derived mouse xenograft cells retained genomic alterations of originating tumors based on mate pair sequencing. We measured response to drugs over time with real-time microscopy on-chip. Last, by engineering droplets to form predetermined geometric shapes, we were able to manipulate the geometry of cultured cell masses. These outcomes can enable broad applications in advancing personalized medicine for cancer and drug discovery, tissue engineering, and stem cell research.