Differential exposure of tumor cells to microenvironmental cues greatly impacts cell phenotypes, raising a need for position based sorting of tumor cells amenable to multiple OMICs and functional analyses. One such key determinant of tumor heterogeneity in solid tumors is its vasculature. Proximity to blood vessels (BVs) profoundly affects tumor cell phenotypes due to differential availability of oxygen, gradient exposure to blood-borne substances and inputs by angiocrine factors. To unravel the whole spectrum of genes, pathways and phenotypes impacted by BVs and to determine spatial domains of vascular influences, we developed a methodology for sorting tumor cells according to their relative distance from BVs. The procedure exemplified here using glioblastoma (GBM) model is based on differential uptake of intra-venously injected, freely-diffusing fluorescent dye that allows separation of stroma-free tumor cells residing in different, successive microenvironments amenable for subsequent OMICs and functional analyses. This reliable, easy to use, cost effective strategy can be extended to all solid tumors to study the impact of vasculature or the lack of it.
Bibliographical noteFunding Information:
We thank members of the Eli Keshet’s laboratory for helpful suggestions during the standardization of this protocol. This work was supported by a DKFZ-MOST research cooperation grant (CA-178), a European Research Council Advance ERC Grant (322692-VASNICHE) and the Israel Cancer Research Fund Professorship Award to Eli Keshet. This protocol was adapted from previously published work (Kumar et al., 2019).
Copyright © 2020 The Authors; exclusive licensee Bio-protocol LLC.
- Blood vessels
- Spatial transcriptomics
- Tumor angiogenesis
- Tumor heterogeneity