High-resolution radioluminescence microscopy of FDG uptake in an engineered 3D tumor-stoma model

Abstract

The increased glucose metabolism of cancer cells is the basis for 18 F-uorodeoxyglucose positron emission tomography (FDG-PET). However, due to its coarse image resolution, PET is unable to resolve the metabolic role of cancer-associated stroma, which often in uences the metabolic reprogramming of a tumor. This study investigates the use of radioluminescence microscopy for imaging FDG uptake in engineered 3D tumor models with high resolution. METHODMulticellular tumor spheroids (A549 lung adenocarcinoma) were co-cultured with GFP-expressing human umbilical vein endothelial cells (HUVECs) within an arti cial extracellular matrix to mimic a tumor and its surrounding stroma. The tumor model was constructed as a 200 µm-thin 3D layer over a transparent CdWO 4 scintillator plate to allow high-resolution imaging of the cultured cells. After incubation with FDG, the radioluminescence signal was collected by a highly sensitive wide eld microscope. Fluorescence microscopy was performed using the same instrument to localize endothelial and tumor cells. RESULTSSimultaneous and co-localized bright eld, uorescence and radioluminescence imaging provided highresolution information on the distribution of FDG in the engineered tissue. The microvascular stromal compartment as a whole took up a large fraction of the FDG, comparable to the uptake of the tumor spheroids. In vitro gamma counting con rmed that A549 and HUVEC cells were both highly glycolytic with rapid FDG-uptake kinetics. Despite the relative thickness of the tissue constructs, an average spatial resolution of 64 ± 4 µm was achieved for imaging FDG. CONCLUSIONOur study demonstrates the feasibility of imaging the distribution of FDG uptake in engineered in vitro tumor models. With its high spatial resolution, the method can separately resolve tumor and stromal components. The approach could be extended to more advanced engineered cancer models but also to surgical tissue slices and tumor biopsies.

Modelling cancer metabolism in vitro: current improvements and future challenges

“…Glucose uptake measurements [61] 18F-FTHA Fatty acid Hepatic fatty acid uptake [63] high sensitivity and ability to identify various compounds in complex models, MS represents a powerful tool for metabolomics analysis [46]. In this respect, MS can be applied to virtually any 3D model.…”

“…The 18 Ffluorodeoxyglucose positron emission tomography (FDG-PET) is a glucose uptake-based methodology that has been extensively used for tumour imaging in vivo and which is also implemented in 3D models. For example, Khan et al [61] co-cultured lung adenocarcinoma with GFP-expressing human umbilical vein endothelial cells (HUVECs) in multicellular tumour spheroids within an artificial ECM. Using brightfield, fluorescence and radioluminescence imaging simultaneously, they provided high-resolution information about FDG distribution within 3D cultures, showing a large uptake of FDG in the microvascular stromal compartment.…”

FLASH Radiotherapy Using Single-Energy Proton PBS Transmission Beams for Hypofractionation Liver Cancer: Dose and Dose Rate Quantification

“…For proton PBS, currently, there is no widely accepted method to quantify the dose rate, despite multiple definitions proposed by different groups (19,21,22). There are several hypotheses, including oxygen depletion (26)(27)(28)(29), immune response (30), and peroxyl radical recombination (31), that have been proposed to explain the FLASH sparing effect. Another important aspect for practical PBS FLASH treatment planning comes from the existing machine parameters (19,32,33), such as beam current, magnet scanning speed, and minimum monitor unit (MU) of a spot.…”

Development of a Lensless Radiomicroscope for Cellular-Resolution Radionuclide Imaging

“…RLM has been used to study metabolism with 18 F-FDG (7) and cell proliferation with 18 F-fluorothymidine (1) in cell monolayers. The method can also be applied to image 3-dimensional (3D)-cultured cells including engineered tumor-stroma models (8) and patient-derived tumor organoids (9).…”