Research ArticleAPPLIED SCIENCES AND ENGINEERING

High-resolution tomographic analysis of in vitro 3D glioblastoma tumor model under long-term drug treatment

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Science Advances  06 Mar 2020:
Vol. 6, no. 10, eaay7513
DOI: 10.1126/sciadv.aay7513
  • Fig. 1 Conceptual figure for the workflow.

    (Top) Integration of the platform: 3D tumor tissue model and 2GMFMT. The platform enables long-term tissue culture and longitudinal assessment of tumor invasion. Bio-printing: 3D tissue containing a GBM spheroid and vascular channels were bioprinted within a plexiglass perfusion chamber and cultured with medium perfusion (with or without drug). Longitudinal imaging is conducted through a thick plexiglass for both tumor growth period and drug regimen. Noninvasive imaging was conducted through a transparent plexiglass chamber. Descanned configuration enabled dense sampling of the target. Each scan point (i.e., p, r, and s) represents a pixel in the raw data. As a representative, one detector raw datum was shown. A full frame was completed when raster scanning is finished (scan point number, s). Typically, a full field of view scanning is completed in ~20 s. (Bottom) Multimodal imaging and 3D reconstruction for all potential modalities. 2GMFMT imaging was performed on the model every 3 to 4 days with 2GMFMT. 2GMFMT presents superior data acquisition time for volumetric assessment of GBM brain tumor in comparison to its counterparts microscopic magnetic resonance imaging (μMRI) and laser scanning confocal microscopy (LSCM). WFFM, wide-field fluorescence microscopy. Photo credit: Vivian Lee, Northeastern University.

  • Fig. 2 Drug response of GBM cells cultured in different settings.

    Drug response of GBM cells cultured in different settings. (A to D) alamarBlue test results for testing metabolic activities of tumor cells and the overlay of phase contrast and fluorescent images of GBM cells cultured in 2D monolayer (A and B) and 3D suspended spheroid (C and D). (E) Invasive behavior of GBM cells before and after the treatment. SD02 cells from the embedded spheroid aggressively invaded into the surrounding matrix (day 26), regressed, and showed shrinkage of tumor core after drug treatment (14 days after drug treatment). However, some GBM cells survived the treatment and resumed its active invasion even with the continuing treatment (31 days after drug treatment). Top-view images (B, D, and E) were captured using WFFM. EC, endothelial cell.

  • Fig. 3 Imaging system rendered image and sensitivity characterization.

    (A) 2GMFMT system is depicted with plexiglass sample holder. The same optical pathway for excitation and emission light enabled coaligned raster scanning of the light and the detector array (i.e., camera) that entails descanned mode for data collection. EMCCD, electron-multiplying charge-coupled device. (B) Wide-field image of spheres before they were placed in the sample holder. Spheres were separated, d = 175 μm. Scale bars, 200 μm. (C and D) The x-y maximum intensity projection for LSCM and 2GMFMT, respectively. (E) The x-z cross section from μMRI volume. The known diameter and depth of spheres enabled to generate a ground truth (GT) image (F). μMRI (G), LSCM (H), and 2GMFMT (I) competed against each other. Performance criterion of volume error and sphericity showed that 2GMFMT overall delivered a better performance against its counterparts. (J) The table shows a detailed comparison for these volume error and sphericity metrics.

  • Fig. 4 Longitudinal intensity assessment of GBM brain tumor (tumor ID: 1).

    (A) WFFM that depicts the cellular invasion. (B) Epifluorescence equivalent raw data from 2GMFMT shows the diffused intensity signal from tumor cells. The 50% maximum intensity isoline (white) delineates the tumor core area, and the 10% maximum intensity isoline (black) shows the invasion area. Core area (C) and invasion area (D) values are shown for all data points, and representative data points are shown pictorially. In particular, the invasion area followed the same trend across modalities. (E) The segmented core areas from WFFM and MFMT 2D images for initial stage (day 26) and regressed stage (TMZ regimen at day 27). (F) The segmented invasion areas WFFM and MFMT 2D images for initial stage (day 26) and regressed stage (TMZ regimen at day 27). Scale bars, 500 μm.

  • Fig. 5 Longitudinal volumetric assessment of GBM brain tumor.

    Longitudinal volumetric assessment of GBM tumor 1 was demonstrated by comparing LSCM and 2GMFMT (A and B). μMRI provided collagen structure and was used as a coregistration landmark. The vascular construct, formed by endothellial cells, is shown in green, and the GBM is in red by both LSCM and 2GMFMT. (C) LSCM and 2GMFMT showed a similarity in volume variation trend with different rates of change. (D) Maximum intensity projections from LSCM and 2GMFMT were compared with LSCM images. Scale bar, 1 mm.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/6/10/eaay7513/DC1

    Fig. S1. 2D monolayer culture of GBM cells.

    Fig. S2. Suspension culture of 3D GBM spheroids.

    Fig. S3. Full EMCCD chip with 2 by 2 binning was used for most of the acquisition schemes.

    Fig. S4. Comparison of imaging modalities monitoring tumor growth on sample 2.

    Fig. S5. Comparison of imaging modalities monitoring tumor growth on sample 3.

    Fig. S6. Comparison of imaging modalities monitoring tumor growth on sample 4.

    Fig. S7. GBM spheroids cultured with glial cells and endothelial cells within soft ECM.

    Movie S1. Fluorescence sphere reconstruction (fluorescent beads with μMRI, LSCM, and MFMT).

    Movie S2. Tumor:1-2-3-4 MFMT-LSCM-MRI reconstruction.

    Movie S3. Sensitivity matrix generation.

  • Supplementary Materials

    The PDF file includes:

    • Fig. S1. 2D monolayer culture of GBM cells.
    • Fig. S2. Suspension culture of 3D GBM spheroids.
    • Fig. S3. Full EMCCD chip with 2 by 2 binning was used for most of the acquisition schemes.
    • Fig. S4. Comparison of imaging modalities monitoring tumor growth on sample 2.
    • Fig. S5. Comparison of imaging modalities monitoring tumor growth on sample 3.
    • Fig. S6. Comparison of imaging modalities monitoring tumor growth on sample 4.
    • Fig. S7. GBM spheroids cultured with glial cells and endothelial cells within soft ECM.
    • Legends for movies S1 to S3

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    Other Supplementary Material for this manuscript includes the following:

    • Movie S1 (.mp4 format). Fluorescence sphere reconstruction (fluorescent beads with μMRI, LSCM, and MFMT).
    • Movie S2 (.mp4 format). Tumor:1-2-3-4 MFMT-LSCM-MRI reconstruction.
    • Movie S3 (.mp4 format). Sensitivity matrix generation.

    Files in this Data Supplement:

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