bims-orenst 2022-06-05 papers

Issue of 2022–06–05
four papers selected by

Biotechnol J. 2022 Jun 02. e2100530

Compartmentalized microfluidic device for in vitro co-culture of retinal cells.

Devashree Jahagirdar, Shital Yadav, Manish Gore, Vikram Korpale, C S Mathpati, Subbulakshmi Chidambaram, Abhijit Majumder, Ratnesh Jain, Prajakta Dandekar.

The investigation is focused on the development of a compartmentalized microfluidic device for coculturing the cells of crucial retinal cellular layers and assessing cell-to-cell interactions. A perfusion-based microfluidic device was employed for coculturing the retinal epithelium (BPEI-1) and precursor (R28) cells. Computational validation was conducted for determining the pressure drop and fluid flow rate within the device microchannels. PDMS polymer used for fabrication ascertained gas permeability through the device and facilitated adherence of the cells over the glass surface, after coating with a cocktail of fibronectin and collagen. The precoated microfluidic device successfully supported cell proliferation and viability, under continuous perfusion of cell culture medium, at the flow rate of 1 μL/min. The barrier integrity of this coculture was confirmed by evaluating the permeability of fluorescently labeled molecules, wherein improved barrier functions were observed, as compared to a static model. The coculture expressed characteristic phenotypic protein markers like recoverin, PAX6, for retinal precursor cells, and RPE65 for retinal epithelial cells. The coculture also exhibited basal expression of TNF-α under normal conditions, as compared to cells treated with LPS, which was employed to induce an elevated inflammatory response. Additionally, the precursor cells expressed the photoreceptor marker, rhodopsin. Photoreceptor cells inherently possess sensitivity toward violet/blue light, which was validated in R28 cells by exposure to light having a wavelength of 405 nm, which significantly decreased cell viability via increased TNF-α production and reduced rhodopsin expression. This proof-of-concept investigation proved the functionality of the retinal coculture, which may be used as an appropriate perfusion-based, preclinical tool for the evaluation of novel retinal drugs and delivery systems. This article is protected by copyright. All rights reserved.

Keywords: co-culture; microfluidic; retina-on-a-chip; retinal epithelial cells; rod photoreceptor

DOI: https://doi.org/10.1002/biot.202100530

Proc Natl Acad Sci U S A. 2022 Jun 07. 119(23): e2118697119

A predictive microfluidic model of human glioblastoma to assess trafficking of blood-brain barrier-penetrant nanoparticles.

Joelle P Straehla, Cynthia Hajal, Hannah C Safford, Giovanni S Offeddu, Natalie Boehnke, Tamara G Dacoba, Jeffrey Wyckoff, Roger D Kamm, Paula T Hammond.

Significance The blood-brain barrier represents a major therapeutic challenge for the treatment of glioblastoma, and there is an unmet need for in vitro models that recapitulate human biology and are predictive of in vivo response. Here, we present a microfluidic model of vascularized glioblastoma featuring a tumor spheroid in direct contact with self-assembled vascular networks comprising human endothelial cells, astrocytes, and pericytes. This model was designed to accelerate the development of targeted nanotherapeutics and enabled rigorous assessment of a panel of surface-functionalized nanoparticles designed to exploit a receptor overexpressed in tumor-associated vasculature. Trafficking and efficacy data in the in vitro model compared favorably to parallel in vivo data, highlighting the utility of the vascularized glioblastoma model for therapeutic development.

Keywords: blood–brain barrier; drug delivery; glioblastoma; microfluidic; nanoparticle

DOI: https://doi.org/10.1073/pnas.2118697119

Adv Sci (Weinh). 2022 Jun 03. e2201785

Recapitulated Crosstalk between Cerebral Metastatic Lung Cancer Cells and Brain Perivascular Tumor Microenvironment in a Microfluidic Co-Culture Chip.

Hyunho Kim, Jason K Sa, Jaehoon Kim, Hee Jin Cho, Hyun Jeong Oh, Dong-Hee Choi, Seok-Hyeon Kang, Da Eun Jeong, Do-Hyun Nam, Hakho Lee, Hye Won Lee, Seok Chung.

Non-small cell lung carcinoma (NSCLC), which affects the brain, is fatal and resistant to anti-cancer therapies. Despite innate, distinct characteristics of the brain from other organs, the underlying delicate crosstalk between brain metastatic NSCLC (BM-NSCLC) cells and brain tumor microenvironment (bTME) associated with tumor evolution remains elusive. Here, a novel 3D microfluidic tri-culture platform is proposed for recapitulating positive feedback from BM-NSCLC and astrocytes and brain-specific endothelial cells, two major players in bTME. Advanced imaging and quantitative functional assessment of the 3D tri-culture model enable real-time live imaging of cell viability and separate analyses of genomic/molecular/secretome from each subset. Susceptibility of multiple patient-derived BM-NSCLCs to representative targeted agents is altered and secretion of serpin E1, interleukin-8, and secreted phosphoprotein 1, which are associated with tumor aggressiveness and poor clinical outcome, is increased in tri-culture. Notably, multiple signaling pathways involved in inflammatory responses, nuclear factor kappa-light-chain-enhancer of activated B cells, and cancer metastasis are activated in BM-NSCLC through interaction with two bTME cell types. This novel platform offers a tool to elucidate potential molecular targets and for effective anti-cancer therapy targeting the crosstalk between metastatic cancer cells and adjacent components of bTME.

Keywords: brain perivascular tumor microenvironment; cerebral metastatic lung cancer cells; microfluidic co-culture chip

DOI: https://doi.org/10.1002/advs.202201785

mBio. 2022 Jun 02. e0094422

Epithelial and Neutrophil Interactions and Coordinated Response to Shigella in a Human Intestinal Enteroid-Neutrophil Coculture Model.

Jose M Lemme-Dumit, Michele Doucet, Nicholas C Zachos, Marcela F Pasetti.

Polymorphonuclear neutrophils (PMN) are recruited to the gastrointestinal mucosa in response to inflammation, injury, and infection. Here, we report the development and the characterization of an ex vivo tissue coculture model consisting of human primary intestinal enteroid monolayers and PMN, and a mechanistic interrogation of PMN-epithelial cell interaction and response to Shigella, a primary cause of childhood dysentery. Cellular adaptation and tissue integration, barrier function, PMN phenotypic and functional attributes, and innate immune responses were examined. PMN within the enteroid monolayers acquired a distinct activated/migratory phenotype that was influenced by direct epithelial cell contact as well as by molecular signals. Seeded on the basal side of the intestinal monolayer, PMN were intercalated within the epithelial cells and moved paracellularly toward the apical side. Cocultured PMN also increased basal secretion of interleukin 8 (IL-8). Shigella added to the apical surface of the monolayers evoked additional PMN phenotypic adaptations, including increased expression of cell surface markers associated with chemotaxis and cell degranulation (CD47, CD66b, and CD88). Apical Shigella infection triggered rapid transmigration of PMN to the luminal side, neutrophil extracellular trap (NET) formation, and bacterial phagocytosis and killing. Shigella infection modulated cytokine production in the coculture; apical monocyte chemoattractant protein (MCP-1), tumor necrosis factor alpha (TNF-α), and basolateral IL-8 production were downregulated, while basolateral IL-6 secretion was increased. We demonstrated, for the first time, PMN phenotypic adaptation and mobilization and coordinated epithelial cell-PMN innate response upon Shigella infection in the human intestinal environment. The enteroid monolayer-PMN coculture represents a technical innovation for mechanistic interrogation of gastrointestinal physiology, host-microbe interaction, innate immunity, and evaluation of preventive/therapeutic tools. IMPORTANCE Studies of mucosal immunity and microbial host cell interaction have traditionally relied on animal models and in vitro tissue culture using immortalized cancer cell lines, which yield nonphysiological and often unreliable results. Herein, we report the development and characterization of an ex vivo enteroid-PMN coculture consisting of normal human intestinal epithelium and a mechanistic interrogation of PMN and epithelial cell interaction and function in the context of Shigella infection. We demonstrated tissue-driven phenotypic and functional adaptation of PMN and a coordinated epithelial cell and PMN response to Shigella, a primary cause of dysentery in young children in the developing world.

Keywords: Shigella; enteroid; epithelial cells; host-pathogen interactions; human; innate immunity; intestinal mucosa; neutrophils

DOI: https://doi.org/10.1128/mbio.00944-22