bims-orenst 2022-03-27 papers

bims-orenst

Biomed News

on Organs-on-chips and engineered stem cell models

Issue of 2022–03–27
eight papers selected by

Functional analysis of human brain endothelium using a microfluidic device integrating a cell culture insert.
Engineering neurovascular organoids with 3D printed microfluidic chips.
In-vitro engineered human cerebral tissues mimic pathological circuit disturbances in 3D.
Microfluidic platform for studying the anti-cancer effect of Ursolic acid on tumor spheroid.
Human Neurons Form Axon-Mediated Functional Connections with Human Cardiomyocytes in Compartmentalized Microfluidic Chip.
3D bioprinted, vascularized neuroblastoma tumor environment in fluidic chip devices for precision medicine drug testing.
Orthogonally induced differentiation of stem cells for the programmatic patterning of vascularized organoids and bioprinted tissues.
Kidney Decellularized Extracellular Matrix Enhanced the Vascularization and Maturation of Human Kidney Organoids.

APL Bioeng. 2022 Mar;6(1): 016103

Functional analysis of human brain endothelium using a microfluidic device integrating a cell culture insert.

Shigenori Miura, Yuya Morimoto, Tomomi Furihata, Shoji Takeuchi.

The blood-brain barrier (BBB) is a specialized brain endothelial barrier structure that regulates the highly selective transport of molecules under continuous blood flow. Recently, various types of BBB-on-chip models have been developed to mimic the microenvironmental cues that regulate the human BBB drug transport. However, technical difficulties in complex microfluidic systems limit their accessibility. Here, we propose a simple and easy-to-handle microfluidic device integrated with a cell culture insert to investigate the functional regulation of the human BBB endothelium in response to fluid shear stress (FSS). Using currently established immortalized human brain microvascular endothelial cells (HBMEC/ci18), we formed a BBB endothelial barrier without the substantial loss of barrier tightness under the relatively low range of FSS (0.1-1 dyn/cm2). Expression levels of key BBB transporters and receptors in the HBMEC/ci18 cells were dynamically changed in response to the FSS, and the effect of FSS reached a plateau around 1 dyn/cm2. Similar responses were observed in the primary HBMECs. Taking advantage of the detachable cell culture insert from the device, the drug efflux activity of P-glycoprotein (P-gp) was analyzed by the bidirectional permeability assay after the perfusion culture of cells. The data revealed that the FSS-stimulated BBB endothelium exhibited the 1.9-fold higher P-gp activity than that of the static culture control. Our microfluidic system coupling with the transwell model provides a functional human BBB endothelium with secured transporter activity, which is useful to investigate the bidirectional transport of drugs and its regulation by FSS.

DOI: https://doi.org/10.1063/5.0085564
Lab Chip. 2022 Mar 25.

Engineering neurovascular organoids with 3D printed microfluidic chips.

Idris Salmon, Sergei Grebenyuk, Abdel Rahman Abdel Fattah, Gregorius Rustandi, Thomas Pilkington, Catherine Verfaillie, Adrian Ranga.

The generation of tissue and organs requires close interaction with vasculature from the earliest moments of embryonic development. Tissue-specific organoids derived from pluripotent stem cells allow for the in vitro recapitulation of elements of embryonic development. However, they are not intrinsically vascularized, which poses a major challenge for their sustained growth, and for understanding the role of vasculature in fate specification and morphogenesis. Current organoid vascularization strategies do not recapitulate the temporal synchronization and spatial orientation needed to ensure in vivo-like early co-development. Here, we developed a human pluripotent stem cell (hPSC)-based approach to generate organoids which interact with vascular cells in a spatially determined manner. The spatial interaction between organoid and vasculature is enabled by the use of a custom designed 3D printed microfluidic chip which allows for a sequential and developmentally matched co-culture system. We show that on-chip hPSC-derived pericytes and endothelial cells sprout and self-assemble into organized vascular networks, and use cerebral organoids as a model system to explore interactions with this de novo generated vasculature. Upon co-development, vascular cells physically interact with the cerebral organoid and form an integrated neurovascular organoid on chip. This 3D printing-based platform is designed to be compatible with any organoid system and is an easy and highly cost-effective way to vascularize organoids. The use of this platform, readily performed in any lab, could open new avenues for understanding and manipulating the co-development of tissue-specific organoids with vasculature.

DOI: https://doi.org/10.1039/d1lc00535a
Commun Biol. 2022 Mar 23. 5(1): 254

In-vitro engineered human cerebral tissues mimic pathological circuit disturbances in 3D.

Aref Saberi, Albert P Aldenkamp, Nicholas A Kurniawan, Carlijn V C Bouten.

In-vitro modeling of brain network disorders such as epilepsy remains a major challenge. A critical step is to develop an experimental approach that enables recapitulation of in-vivo-like three-dimensional functional complexity while allowing local modulation of the neuronal networks. Here, by promoting matrix-supported active cell reaggregation, we engineered multiregional cerebral tissues with intact 3D neuronal networks and functional interconnectivity characteristic of brain networks. Furthermore, using a multi-chambered tissue-culture chip, we show that our separated but interconnected cerebral tissues can mimic neuropathological signatures such as the propagation of epileptiform discharges.

DOI: https://doi.org/10.1038/s42003-022-03203-4
Electrophoresis. 2022 Mar 22.

Microfluidic platform for studying the anti-cancer effect of Ursolic acid on tumor spheroid.

Shiqi Chang, Jing Wen, Yue Su, Huipeng Ma.

  At present, the probability that a new anti-tumor drug will eventually succeed in clinical trials is extremely low. In order to make up for this shortcoming, the use of a three-dimensional cell culture model for secondary screening is often necessary. Cell spheroid is the easiest three-dimensional model tool for drug screening. In this study, the microfluidic chip with microwell array was manufactured, which could allow forming tumor spheroids with uniform size and easily retrieving cell spheroids from the chip. Cell spheroids were successfully cultured for over 15 days and the survival rate was as high as 80%. Subsequently, cellular response to the Ursolic acid (UA) was observed on the chip. Compared to the monolayer culture cells in vitro, the tumor spheroids showed minor levels of epithelial-mesenchymal transition (EMT) fluctuation after drug treatment. The mechanism of cell spheroid resistance to UA was further verified by detecting the expression level of upstream pathway proteins. But the invasive ability of tumor spheroids was attenuated when the duration of action of UA extended. The anti-cancer effect of UA was innovatively evaluated on breast cancer by using the microfluidic device, which could provide a basis and direction for future pre-clinical research on UA. This article is protected by copyright. All rights reserved.

Keywords:  EMT; microfluidic chip; tumor spheroids; ursolic acid

DOI:  https://doi.org/10.1002/elps.202100382
Int J Mol Sci. 2022 Mar 15. pii: 3148. [Epub ahead of print]23(6):

Human Neurons Form Axon-Mediated Functional Connections with Human Cardiomyocytes in Compartmentalized Microfluidic Chip.

Martta Häkli, Satu Jäntti, Tiina Joki, Lassi Sukki, Kaisa Tornberg, Katriina Aalto-Setälä, Pasi Kallio, Mari Pekkanen-Mattila, Susanna Narkilahti.

  The cardiac autonomic nervous system (cANS) regulates cardiac function by innervating cardiac tissue with axons, and cardiomyocytes (CMs) and neurons undergo comaturation during the heart innervation in embryogenesis. As cANS is essential for cardiac function, its dysfunctions might be fatal; therefore, cardiac innervation models for studying embryogenesis, cardiac diseases, and drug screening are needed. However, previously reported neuron-cardiomyocyte (CM) coculture chips lack studies of functional neuron-CM interactions with completely human-based cell models. Here, we present a novel completely human cell-based and electrophysiologically functional cardiac innervation on a chip in which a compartmentalized microfluidic device, a 3D3C chip, was used to coculture human induced pluripotent stem cell (hiPSC)-derived neurons and CMs. The 3D3C chip enabled the coculture of both cell types with their respective culture media in their own compartments while allowing the neuronal axons to traverse between the compartments via microtunnels connecting the compartments. Furthermore, the 3D3C chip allowed the use of diverse analysis methods, including immunocytochemistry, RT-qPCR and video microscopy. This system resembled the in vivo axon-mediated neuron-CM interaction. In this study, the evaluation of the CM beating response during chemical stimulation of neurons showed that hiPSC-neurons and hiPSC-CMs formed electrophysiologically functional axon-mediated interactions.

Keywords:  axon-mediated; cardiomyocyte; coculture; functional interaction; human-induced pluripotent stem cell; microfluidic chip; neuron; organ-on-chip

DOI:  https://doi.org/10.3390/ijms23063148
Biofabrication. 2022 Mar 22.

3D bioprinted, vascularized neuroblastoma tumor environment in fluidic chip devices for precision medicine drug testing.

Daniel Nothdurfter, Christian Ploner, Débora C Coraça-Huber, Doris Wilflingseder, Thomas Müller, Martin Hermann, Judith Hagenbuchner, Michael J Ausserlechner.

  Neuroblastoma is an extracranial solid tumor which develops in early childhood and still has a poor prognosis. One strategy to increase cure rates is the identification of patient-specific drug responses in tissue models that mimic the interaction between patient cancer cells and tumor environment. We therefore developed a perfused and micro-vascularized tumor-environment model that is directly bioprinted into custom-manufactured fluidic chips. A gelatin-methacrylate/fibrin-based matrix containing multiple cell types mimics the tumor-microenvironment that promotes spontaneous micro-vessel formation by embedded endothelial cells. We demonstrate that both, adipocyte- and iPSC-derived mesenchymal stem cells can guide this process. Bioprinted channels are coated with endothelial cells post printing to form a dense vessel - tissue barrier. The tissue model thereby mimics structure and function of human soft tissue with endothelial cell-coated larger vessels for perfusion and micro-vessel networks within the hydrogel-matrix. Patient-derived neuroblastoma spheroids are added to the matrix during the printing process and grown for more than two weeks. We demonstrate that micro-vessels are attracted by and grow into tumor spheroids and that neuroblastoma cells invade the tumor-environment as soon as the spheroids disrupt. In summary, we describe the first bioprinted, micro-vascularized neuroblastoma - tumor-environment model directly printed into fluidic chips and a novel medium-throughput biofabrication platform suitable for studying tumor angiogenesis and metastasis in precision medicine approaches in future.

Keywords:  angiogenesis; fluidic device; induced pluripotent stem cell differentiation; laser manufacturing; mesenchymal stem cells; neuroblastoma; vascularization

DOI:  https://doi.org/10.1088/1758-5090/ac5fb7
Nat Biomed Eng. 2022 Mar 24.

Orthogonally induced differentiation of stem cells for the programmatic patterning of vascularized organoids and bioprinted tissues.

Mark A Skylar-Scott, Jeremy Y Huang, Aric Lu, Alex H M Ng, Tomoya Duenki, Songlei Liu, Lucy L Nam, Sarita Damaraju, George M Church, Jennifer A Lewis.

The generation of organoids and tissues with programmable cellular complexity, architecture and function would benefit from the simultaneous differentiation of human induced pluripotent stem cells (hiPSCs) into divergent cell types. Yet differentiation protocols for the overexpression of specific transcription factors typically produce a single cell type. Here we show that patterned organoids and bioprinted tissues with controlled composition and organization can be generated by simultaneously co-differentiating hiPSCs into distinct cell types via the forced overexpression of transcription factors, independently of culture-media composition. Specifically, we used such orthogonally induced differentiation to generate endothelial cells and neurons from hiPSCs in a one-pot system containing either neural or endothelial stem-cell-specifying media, and to produce vascularized and patterned cortical organoids within days by aggregating inducible-transcription-factor and wild-type hiPSCs into randomly pooled or multicore-shell embryoid bodies. Moreover, by leveraging multimaterial bioprinting of hiPSC inks without extracellular matrix, we generated patterned neural tissues with layered regions composed of neural stem cells, endothelium and neurons. Orthogonally induced differentiation of stem cells may facilitate the fabrication of engineered tissues for biomedical applications.

DOI: https://doi.org/10.1038/s41551-022-00856-8
Adv Sci (Weinh). 2022 Mar 24. e2103526

Kidney Decellularized Extracellular Matrix Enhanced the Vascularization and Maturation of Human Kidney Organoids.

Jin Won Kim, Sun Ah Nam, Jawoon Yi, Jae Yun Kim, Jong Young Lee, Seo-Yeon Park, Tugce Sen, Yoo-Mi Choi, Jae Yeon Lee, Hong Lim Kim, Hyung Wook Kim, Jiwhan Park, Dong-Woo Cho, Yong Kyun Kim.

  Kidney organoids derived from human pluripotent stem cells (hPSCs) have extensive potential for disease modelling and regenerative medicine. However, the limited vascularization and immaturity of kidney organoids have been still remained to overcome. Extracellular matrix (ECM) can provide mechanical support and a biochemical microenvironment for cell growth and differentiation. Here in vitro methods using a kidney decellularized extracellular matrix (dECM) hydrogel to culture hPSC-derived kidney organoids, which have extensive vascular network and their own endothelial cells, are reported. Single-cell transcriptomics reveal that the vascularized kidney organoids cultured using the kidney dECM have more mature patterns of glomerular development and higher similarity to human kidney than those cultured without the kidney dECM. Differentiation of α-galactosidase A (GLA)-knock-out hPSCs generated using CRISPR/Cas9 into kidney organoids by the culture method using kidney dECM efficiently recapitulate Fabry nephropathy with vasculopathy. Transplantation of kidney organoids with kidney dECM into kidney of mouse accelerates the recruitment of endothelial cells from the host mouse kidney and maintains vascular integrity with the more organized slit diaphragm-like structures than those without kidney dECM. The kidney dECM methodology for inducing extensive vascularization and maturation of kidney organoids can be applied to studies for kidney development, disease modeling, and regenerative medicine.

Keywords:  extracellular matrix; kidney; organoid; vascularization

DOI:  https://doi.org/10.1002/advs.202103526