bims-orenst Biomed News
on Organs-on-chips and engineered stem cell models
Issue of 2021‒03‒28
five papers selected by
Joram Mooiweer
University of Groningen

  1. Biotechnol Bioeng. 2021 Mar 25.
      In vitro models are becoming more advanced to truly present physiological systems while enabling high-throughput screening and analysis. Organ-on-a-chip devices provide remarkable results through the reconstruction of a three-dimensional (3D) cellular microenvironment although they need to be further developed in terms of multiple liquid patterning principle, material properties and scalability. Here we present a 3D anchor-based microfluidic injection-molded plastic array culture platform (Anchor-IMPACT) that enables selective, space-intensive patterning of hydrogels using anchor-island for high-throughput angiogenesis evaluation model. Anchor-IMPACT consists of a central channel and an anchor-island, integrating the array into an abbreviated 96-well plate format with a standard microscope slide size. The anchor-island enables selective 3D cell patterning without channel-to-channel contact or any hydrogel injection port using an anchor structure unlike conventional culture compartment. The hydrogel was patterned into defined regions by spontaneous capillary flow under hydrophilic conditions. We configured multiple cell patterning structures to investigate the angiogenic potency of colorectal cancer (CRC) cells in Anchor-IMPACT and the morphological properties of the angiogenesis induced by the paracrine effect were evaluated. In addition, the efficacy of anticancer drugs against angiogenic sprouts was verified by following dose-dependent responses. Our results indicate that Anchor-IMPACT offers not only a model of high-throughput experimentation but also an advanced 3D cell culture platform and can significantly improve current in vitro models while providing the basis for developing predictive preclinical models for biopharmaceutical applications. This article is protected by copyright. All rights reserved.
    Keywords:  Angiogenesis; Colorectal Cancer; High-throughput Screening; Microfluidics; Organ-on-a-Chip
  2. ACS Omega. 2021 Mar 16. 6(10): 6942-6952
      Human pluripotent stem cell (hPSC)-derived endothelial cells (ECs) are promising cell sources for drug discovery, tissue engineering, and studying or treating vascular diseases. However, hPSC-ECs derived from different culture methods display different phenotypes. Herein, we made a detailed comparative study of hPSC-ECs from three different culture systems (e.g., 2D, 3D PNIPAAm-PEG hydrogel, and 3D alginate hydrogel cultures) based on our previous reports. We expanded hPSCs and differentiated them into ECs in three culture systems. Both 3D hydrogel systems could mimic an in vivo physiologically relevant microenvironment to protect cells from shear force and prevent cell agglomeration, leading to a high culture efficiency and a high volumetric yield. We demonstrated that hPSC-ECs produced from both hydrogel systems had similar results as 2D-ECs. The transcriptome analysis showed that PEG-ECs and alginate-ECs displayed a functional phenotype due to their higher gene expressions in vasculature development, extracellular matrix, angiogenesis, and glycolysis, while 2D-ECs showed a proliferative phenotype due to their higher gene expressions in cell proliferation. Taken together, both PEG- and alginate-hydrogel systems will significantly advance the applications of hPSC-ECs in various biomedical fields.
  3. J Artif Organs. 2021 Mar 22.
      Understanding the active transport of substrates by the kidney in the renal proximal convoluted tubule is crucial for drug development and for studying kidney diseases. Currently, cell-based assays are applied for this this purpose, however, differences between assays and the body are common, indicating the importance of in vitro-in vivo discrepancies. Several studies have suggested that 3D cell cultures expose cells to a more physiological environments, thus, providing more accurate cell function results. To mimic the renal proximal tubule, we have developed a custom-made renal module (RM), containing a single polypropylene hollow fibre (Plasmaphan P1LX, 3M) that serves as a porous scaffold and compared to conventional Transwell cell-based bidirectional transport studies. In addition, a constant flow of media, exposed cells to a physiological shear stress of 0.2 dyne/cm2. MDCK-Mdr1a cells, overexpressing the rat Mdr1a (P-gp) transporter, were seeded onto the HF membrane surface coated with the basement membrane matrix Geltrex which facilitated cell adhesion and tight junction formation. Cells were then seeded into the HF lumen where attachment and tight junction formation were evaluated by fluorescence microscopy while epithelial barrier integrity under shear stress was shown to be achieved by day 7. qPCR results have shown significant changes in gene expression compared to cells grown on Transwells. Kidney injury marker such as KIM-1 and the hypoxia marker CA9 have been downregulated, while the CD133 (Prominin-1) microvilli marker has shown a fivefold upregulation. Furthermore, the renal transporter P-gp expression has been downregulated by 50%. Finally, bidirectional assays have shown that cells grown in the RM were able to reabsorb albumin with a higher efficiency compared to Transwell cell cultures while efflux of the P-gp-specific substrates Hoechst and Rhodamine 123 was decreased. These results further support the effect of the microenvironment and fluidic shear stress on cell function and gene expression. This can serve as the basis for the development of a microphysiological renal model for drug transport studies.
    Keywords:  3D cell culture; Drug transport; Fluidic shear stress; Hollow fibre; Renal function
  4. Front Bioeng Biotechnol. 2021 ;9 615639
      An islet-on-chip system in the form of a completely transparent microscope slide optically accessible from both sides was developed. It is made from laser-structured borosilicate glass and enables the parallel perifusion of five microchannels, each containing one islet precisely immobilized in a pyramidal well. The islets can be in inserted via separate loading windows above each pyramidal well. This design enables a gentle, fast and targeted insertion of the islets and a reliable retention in the well while at the same time permitting a sufficiently fast exchange of the media. In addition to the measurement of the hormone content in the fractionated efflux, parallel live cell imaging of the islet is possible. By programmable movement of the microscopic stage imaging of five wells can be performed. The current chip design ensures sufficient time resolution to characterize typical parameters of stimulus-secretion coupling. This was demonstrated by measuring the reaction of the islets to stimulation by glucose and potassium depolarization. After the perifusion experiment islets can be removed for further analysis. The live-dead assay of the removed islets confirmed that the process of insertion and removal was not detrimental to islet structure and viability. In conclusion, the present islet-on-chip design permits the practical implementation of parallel perifusion experiments on a single and easy to load glass slide. For each immobilized islet the correlation between secretion, signal transduction and morphology is possible. The slide concept allows the scale-up to even higher degrees of parallelization.
    Keywords:  NAD(P)H- and FAD-autofluorescence; borosilicate glass; calcium; femtosecond laser-structuring; insulin secretion; islet of langerhans; microfluidic perifusion system
  5. Front Microbiol. 2021 ;12 626370
      Trypanosoma cruzi (T. cruzi), the etiological agent of Chagas Disease (CD), is transmitted to humans by infected kissing bugs, blood transfusion, organ transplantation, and from mother-to-child. Congenital transmission is now considered an important route of CD spread in non-endemic countries where no routine testing of pregnant women for the disease is implemented. The main cellular mechanisms that lead to fetal infection by T. cruzi, despite the presence of a placental barrier, remain unclear. Mother-to-child transmission most likely occurs when bloodstream trypomastigotes reach the placental intervillous space and interact with the large cellular surface provided by the syncytioptrophoblasts. These highly specialized cells not only function as a physical obstacle between mother and fetus, but also modulate immune responses against pathogen infections. To overcome the limitations associated with the use of human fetal tissues, we employed a three-dimensional (3D) cell culture model to recreate the human placenta environment. In this system, the trophoblast-derived JEG-3 cell line is co-cultured with human brain microvascular endothelial cells attached to microcarrier beads in a rotating bioreactor. Here, we report that 3D culture of JEG-3/HBMEC spheroids promote JEG-3 cells differentiation revealed by the formation of syncytia and production of β human chorionic gonadotropin and human placental lactogen (hPL). Under these growth conditions, we demonstrate that 3D-grown JEG-3 cells have reduced susceptibility to T. cruzi infection compared to JEG-3 cells grown in conventional tissue culture flasks. We also show that 3D-cultured JEG-3 cells release paracrine factors in the supernatant that prevent T. cruzi infection of non-trophoblastic cell lines. Our in vitro model of T. cruzi vertical transmission may help better understand the molecular processes by which parasites bypass the human placental barrier and could be exploited to evaluate therapeutics to reduce congenital CD.
    Keywords:  3D culture system; Trypanosoma cruzi; chagas disease; congenital infection; human trophoblasts