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Quantitative characterization of the regulation of iron metabolism in glioblastoma stem-like cells using magnetophoresis.

ABSTRACT: This study focuses on different iron regulation mechanisms of glioblastoma (GBM) cancer stem-like cells (CSCs) and non-stem tumor cells (NSTCs) using multiple approaches: cell viability, density, and magnetophoresis. GBM CSCs and NSTCs were exposed to elevated iron concentration, and their magnetic susceptibility was measured using single cell magnetophoresis (SCM), which tracks the magnetic and settling velocities of thousands of individual cells passing through the magnetic field with a constant energy gradient. Our results consistently demonstrate that GBM NSTCs have higher magnetic susceptibility distribution at increased iron concentration compared with CSCs, and we speculate that it is because CSCs have the ability to store a high amount of iron in ferritin, whereas the free iron ions inside the NSTCs lead to higher magnetic susceptibility and reduced cell viability and growth. Further, their difference in magnetic susceptibility has led us to pursue a separate experiment using a quadrupole magnetic separator (QMS), a novel microfluidic device that uses a concentric channel and permanent magnets in a special configuration to separate samples based on their magnetic susceptibilities. GBM CSCs and NSTCs were exposed to elevated iron concentration, stained with two different trackers, mixed and introduced into QMS; subsequently, the separated fractions were analyzed by fluorescent microscopy. The separation results portray a successful label-less magnetic separation of the two populations.

SUBMITTER: Park KJ

PROVIDER: S-EPMC6693654 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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Quantitative characterization of the regulation of iron metabolism in glioblastoma stem-like cells using magnetophoresis.

Park Kyoung-Joo J KJ Kim James J Testoff Thomas T Adams Joseph J Poklar Miranda M Zborowski Maciej M Venere Monica M Chalmers Jeffrey J JJ

Biotechnology and bioengineering 20190424 7

This study focuses on different iron regulation mechanisms of glioblastoma (GBM) cancer stem-like cells (CSCs) and non-stem tumor cells (NSTCs) using multiple approaches: cell viability, density, and magnetophoresis. GBM CSCs and NSTCs were exposed to elevated iron concentration, and their magnetic susceptibility was measured using single cell magnetophoresis (SCM), which tracks the magnetic and settling velocities of thousands of individual cells passing through the magnetic field with a consta ...[more]

PMID: 30906984

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Project description:Glioblastoma (GBM) displays marked cellular and metabolic heterogeneity that varies among cellular microenvironments within a tumor. Metabolic targeting has long been advocated as a therapy against many tumors including GBM, but how lipid metabolism is altered to suit different microenvironmental conditions and whether cancer stem cells (CSCs) have altered lipid metabolism are outstanding questions in the field. We interrogated gene expression in separate microenvironments of GBM organoid models that mimic the transition between nutrient-rich and nutrient-poor pseudopalisading/perinecrotic tumor zones using spatial-capture RNA-sequencing. We revealed a striking difference in lipid processing gene expression and total lipid content between diverse cell populations from the same patient, with lipid enrichment in hypoxic organoid cores and also in perinecrotic and pseudopalisading regions of primary patient tumors. This was accompanied by regionally restricted upregulation of hypoxia-inducible lipid droplet-associated (HILPDA) gene expression in organoid cores and pseudopalisading regions of clinical GBM specimens, but not lower-grade brain tumors. CSCs have low lipid droplet accumulation compared to non-CSCs in organoid models and xenograft tumors, and prospectively sorted lipid-low GBM cells are functionally enriched for stem cell activity. Targeted lipidomic analysis of multiple patient-derived models revealed a significant shift in lipid metabolism between GBM CSCs and non-CSCs, suggesting that lipid levels may not be simply a product of the microenvironment but also may be a reflection of cellular state. CSCs had decreased levels of major classes of neutral lipids compared to non-CSCs, but had significantly increased polyunsaturated fatty acid production due to high fatty acid desaturase (FADS1/2) expression which was essential to maintain CSC viability and self-renewal. Our data demonstrate spatially and hierarchically distinct lipid metabolism phenotypes occur clinically in the majority of patients, can be recapitulated in laboratory models, and may represent therapeutic targets for GBM.

Project description:BackgroundIn this study, we investigated the feasibility of quantitative ultrashort echo time (qUTE) magnetic resonance (MR) imaging techniques in the detection and quantification of iron oxide nanoparticle (IONP)-labeled stem cells.MethodsA stem cell phantom containing multiple layers of unlabeled or labeled stem cells with different densities was prepared. The phantom was imaged with quantitative UTE (qUTE) MR techniques [i.e., UTE-T1 mapping, UTE-T2* mapping, and UTE-based quantitative susceptibility mapping (UTE-QSM)] as well as with a clinical T2 mapping sequence on a 3T clinical MR system. For T1 mapping, a variable flip angle (VFA) method based on actual flip angle imaging (AFI) technique was utilized. For T2* mapping and UTE-QSM, multiple images with variable, interleaved echo times including UTE images and gradient recalled echo (GRE) images were used. For UTE-QSM, the phase information from the multi-echo images was utilized and processed using a QSM framework based on the morphology-enabled dipole inversion (MEDI) algorithm. The qUTE techniques were also evaluated in an ex vivo experiment with a mouse injected with IONP-labeled stem cells.ResultsIn the phantom experiment, the parameters estimated with qUTE techniques showed high linearity with respect to the density of IONP-labeled stem cells (R2>0.99), while the clinical T2 parameter showed impaired linearity (R2=0.87). In the ex vivo mouse experiment, UTE-T2* mapping and UTE-QSM showed feasibility in the detection of injected stem cells with high contrast, whereas UTE-T1 and UTE-T2* showed limited detection. Overall, UTE-QSM demonstrated the best contrast of all, with other methods being subjected more to a confounding factor due to different magnetic susceptibilities of various types of neighboring tissues, which creates inhomogeneous contrast that behaves similar to IONP.ConclusionsIn this study, we evaluated the feasibility of a series of qUTE imaging techniques as well as conventional T2 mapping for the detection of IONP-labeled stem cells in vitro and ex vivo. UTE-QSM performed superior amongst other qUTE techniques as well as conventional T2 mapping in detecting stem cells with high contrast.

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Quantitative characterization of the regulation of iron metabolism in glioblastoma stem-like cells using magnetophoresis.

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Quantitative characterization of the regulation of iron metabolism in glioblastoma stem-like cells using magnetophoresis.

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