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On-chip perivascular niche supporting stemness of patient-derived glioma cells in a serum-free, flowable culture

Glioblastoma multiforme (GBM) is the most common and the most aggressive type of primary brain malignancy. Glioblastoma stem-like cells (GSCs) can migrate in vascular niches within or away from the tumour mass, increasing tumour resistance to treatments and contributing to relapses. To study individ...

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Autores principales: Gerigk, Magda, Bulstrode, Harry, Shi, HaoTian Harvey, Tönisen, Felix, Cerutti, Camilla, Morrison, Gillian, Rowitch, David, Huang, Yan Yan Shery
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The Royal Society of Chemistry 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8204159/
https://www.ncbi.nlm.nih.gov/pubmed/33969368
http://dx.doi.org/10.1039/d1lc00271f
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author Gerigk, Magda
Bulstrode, Harry
Shi, HaoTian Harvey
Tönisen, Felix
Cerutti, Camilla
Morrison, Gillian
Rowitch, David
Huang, Yan Yan Shery
author_facet Gerigk, Magda
Bulstrode, Harry
Shi, HaoTian Harvey
Tönisen, Felix
Cerutti, Camilla
Morrison, Gillian
Rowitch, David
Huang, Yan Yan Shery
author_sort Gerigk, Magda
collection PubMed
description Glioblastoma multiforme (GBM) is the most common and the most aggressive type of primary brain malignancy. Glioblastoma stem-like cells (GSCs) can migrate in vascular niches within or away from the tumour mass, increasing tumour resistance to treatments and contributing to relapses. To study individual GSC migration and their interactions with the perivasculature of the tumour microenvironment, there is a need to develop a human organotypic in vitro model. Herein, we demonstrated a perivascular niche-on-a-chip, in a serum-free condition with gravity-driven flow, that supported the stemness of patient-derived GSCs and foetal neural stem cells grown in a three-dimensional environment (3D). Endothelial cells from three organ origins, (i) human brain microvascular endothelial cells (hCMEC/D3), (ii) human umbilical vein endothelial cells (HUVECs) and, (iii) human lung microvascular endothelial cells (HMVEC-L) formed rounded microvessels within the extracellular-matrix integrated microfluidic chip. By optimising cell extraction protocols, systematic studies were performed to evaluate the effects of serum-free media, 3D cell cultures, and the application of gravity-driven flow on the characteristics of endothelial cells and their co-culture with GSCs. Our results showed the maintenance of adherent and tight junction markers of hCMEC/D3 in the serum-free culture and that gravity-driven flow was essential to support adequate viability of both the microvessel and the GSCs in co-culture (>80% viability at day 3). Endpoint biological assays showed upregulation of neovascularization-related genes (e.g., angiopoietins, vascular endothelial growth factor receptors) in endothelial cells co-cultured with GSCs in contrast to the neural stem cell reference that showed insignificant changes. The on-chip platform further permitted live-cell imaging of GSC – microvessel interaction, enabling quantitative analysis of GSC polarization and migration. Overall, our comparative genotypic (i.e. qPCR) and phenotypic (i.e. vessel permeability and GSC migration) studies showed that organotypic (brain cancer cells–brain endothelial microvessel) interactions differed from those within non-tissue specific vascular niches of human origin. The development and optimization of this on-chip perivascular niche, in a serum-free flowable culture, could provide the next level of complexity of an in vitro system to study the influence of glioma stem cells on brain endothelium.
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spelling pubmed-82041592021-06-29 On-chip perivascular niche supporting stemness of patient-derived glioma cells in a serum-free, flowable culture Gerigk, Magda Bulstrode, Harry Shi, HaoTian Harvey Tönisen, Felix Cerutti, Camilla Morrison, Gillian Rowitch, David Huang, Yan Yan Shery Lab Chip Chemistry Glioblastoma multiforme (GBM) is the most common and the most aggressive type of primary brain malignancy. Glioblastoma stem-like cells (GSCs) can migrate in vascular niches within or away from the tumour mass, increasing tumour resistance to treatments and contributing to relapses. To study individual GSC migration and their interactions with the perivasculature of the tumour microenvironment, there is a need to develop a human organotypic in vitro model. Herein, we demonstrated a perivascular niche-on-a-chip, in a serum-free condition with gravity-driven flow, that supported the stemness of patient-derived GSCs and foetal neural stem cells grown in a three-dimensional environment (3D). Endothelial cells from three organ origins, (i) human brain microvascular endothelial cells (hCMEC/D3), (ii) human umbilical vein endothelial cells (HUVECs) and, (iii) human lung microvascular endothelial cells (HMVEC-L) formed rounded microvessels within the extracellular-matrix integrated microfluidic chip. By optimising cell extraction protocols, systematic studies were performed to evaluate the effects of serum-free media, 3D cell cultures, and the application of gravity-driven flow on the characteristics of endothelial cells and their co-culture with GSCs. Our results showed the maintenance of adherent and tight junction markers of hCMEC/D3 in the serum-free culture and that gravity-driven flow was essential to support adequate viability of both the microvessel and the GSCs in co-culture (>80% viability at day 3). Endpoint biological assays showed upregulation of neovascularization-related genes (e.g., angiopoietins, vascular endothelial growth factor receptors) in endothelial cells co-cultured with GSCs in contrast to the neural stem cell reference that showed insignificant changes. The on-chip platform further permitted live-cell imaging of GSC – microvessel interaction, enabling quantitative analysis of GSC polarization and migration. Overall, our comparative genotypic (i.e. qPCR) and phenotypic (i.e. vessel permeability and GSC migration) studies showed that organotypic (brain cancer cells–brain endothelial microvessel) interactions differed from those within non-tissue specific vascular niches of human origin. The development and optimization of this on-chip perivascular niche, in a serum-free flowable culture, could provide the next level of complexity of an in vitro system to study the influence of glioma stem cells on brain endothelium. The Royal Society of Chemistry 2021-05-05 /pmc/articles/PMC8204159/ /pubmed/33969368 http://dx.doi.org/10.1039/d1lc00271f Text en This journal is © The Royal Society of Chemistry https://creativecommons.org/licenses/by-nc/3.0/
spellingShingle Chemistry
Gerigk, Magda
Bulstrode, Harry
Shi, HaoTian Harvey
Tönisen, Felix
Cerutti, Camilla
Morrison, Gillian
Rowitch, David
Huang, Yan Yan Shery
On-chip perivascular niche supporting stemness of patient-derived glioma cells in a serum-free, flowable culture
title On-chip perivascular niche supporting stemness of patient-derived glioma cells in a serum-free, flowable culture
title_full On-chip perivascular niche supporting stemness of patient-derived glioma cells in a serum-free, flowable culture
title_fullStr On-chip perivascular niche supporting stemness of patient-derived glioma cells in a serum-free, flowable culture
title_full_unstemmed On-chip perivascular niche supporting stemness of patient-derived glioma cells in a serum-free, flowable culture
title_short On-chip perivascular niche supporting stemness of patient-derived glioma cells in a serum-free, flowable culture
title_sort on-chip perivascular niche supporting stemness of patient-derived glioma cells in a serum-free, flowable culture
topic Chemistry
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8204159/
https://www.ncbi.nlm.nih.gov/pubmed/33969368
http://dx.doi.org/10.1039/d1lc00271f
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