Figure 1.
CRC-on-Chip tissue structure
(A) The organ-on-chip platform (schematic courtesy of Emulate, Inc.) consists of an epithelial channel (1) comprising epithelial and cancerous cells (3) and an endothelial channel (2) comprising HUVEC cells (4) separated by a porous membrane (5). To model cell-cell interactions in the TME, the CRC-on-Chip was modified to include layers of different cell types in the epithelial channel. CRC tumor cells were seeded on top of the epithelial cells. A stromal layer, comprised of CAFs, can be incorporated into the epithelial channel.
(B) Confocal fluorescence images of a chip cross-section spanning 106 μm from the top of the endothelial channel into the epithelial channel, highlighting the endothelial:epithelial tissue:tissue interface. HUVEC cells are labeled with anti-VE cadherin (red). Caco2 C2BBe1 cells labeled with anti-E-Cadherin (purple) form 3D-like structures in the top epithelial channel. HCT116 H2B-GFP cells grow in clusters on top of the Caco2 cells. Nuclei are labeled with DAPI (blue). Scale bar is 100 μm.
(C) Representative confocal immunofluorescent images of the epithelial (top) and endothelial (bottom) channels of an Intestine Chip (left) and CRC-on-Chip (right) stained for ZO-1 (gold) on day 6. DAPI (blue) labels the nuclei of the Caco2 C2BBe1 cells in the epithelial channel and HUVECs in the endothelial channel. White arrows designate HCT116 (green) in the epithelial channel of the CRC-on-Chip. Scale bars represent 200 μm. Images are maximum projections that span a 15 μm Z-height in the epithelial channel and a 10 μm Z-height in the endothelial channel with a 5 μm step size.
(D) The apparent permeability (Papp) of the intestinal epithelial cells in the top channel was not changed when HCT116 tumor cells were added to the CRC Chips. The concentration of inulin-FITC that diffused from the epithelial channel to the endothelial channel was used to calculate Papp (N = 3 Chips). Data are represented as mean ± SEM and analyzed using a two-way ANOVA; p > 0.05.