Investigation of Tumor Cell Behaviors on a Vascular Microenvironment-Mimicking Microfluidic Chip - PubMed (original) (raw)
Investigation of Tumor Cell Behaviors on a Vascular Microenvironment-Mimicking Microfluidic Chip
Rong Huang et al. Sci Rep. 2015.
Abstract
The extravasation of tumor cells is a key event in tumor metastasis. However, the mechanism underlying tumor cell extravasation remains unknown, mainly hindered by obstacles from the lack of complexity of biological tissues in conventional cell culture, and the costliness and ethical issues of in vivo experiments. Thus, a cheap, time and labor saving, and most of all, vascular microenvironment-mimicking research model is desirable. Herein, we report a microfluidic chip-based tumor extravasation research model which is capable of simultaneously simulating both mechanical and biochemical microenvironments of human vascular systems and analyzing their synergistic effects on the tumor extravasation. Under different mechanical conditions of the vascular system, the tumor cells (HeLa cells) had the highest viability and adhesion activity in the microenvironment of the capillary. The integrity of endothelial cells (ECs) monolayer was destroyed by tumor necrosis factor-α (TNF-α) in a hemodynamic background, which facilitated the tumor cell adhesion, this situation was recovered by the administration of platinum nanoparticles (Pt-NPs). This model bridges the gap between cell culture and animal experiments and is a promising platform for studying tumor behaviors in the vascular system.
Figures
Figure 1. Schematic illustration of the vascular model chip.
(a) Details of the components of the chip. (b) The structure of the integrated chip. (c) A photograph of an actual chip. The dark blue part delineates the microfluidic channel and the orange part indicates the stretch chamber. Scale bar: 1.5 cm.
Figure 2. Time-lapse images of HeLa cells under a physiological hemodynamic condition (FSS, 36.2 dynes cm−2, CS, 7.2 ± 1.1%, 1.17 Hz).
All the cells were stained with Calcein AM. (a) The schematic diagram illustrates flow direction of the fluid and positions of the tumor cells in the microchannel. (b)The red arrows indicate the travelling trace of a HeLa cell in the microchannel and its attachment on the substrate. (c) The process of the formation and breakdown of tumor cell clusters in the microchannel. The red lines track the same cells at different time points. (d) The intensity of the fluorescence of the cell highlighted by the red circle experienced an obvious decreasing process. Scale bar: 20 μm. The two-headed arrows indicate the direction of the CS, the one-headed arrows indicate the direction of the FSS.
Figure 3. Flow cytometry results indicate the effect of mechanical stimulation on apoptosis of the tumor cells.
The red dots in lower right quadrant of each flow cytometry diagram indicate apoptotic cells. (a) The apoptosis rate of tumor cells under physiological FSS and CS for 2 h. (b) The apoptosis rate of tumor cells under high level of FSS for 2 h. (c) The apoptosis rate of tumor cells under high level of FSS and physiological CS for 2 h. (d) The statistical data of the tumor apoptosis rates under different conditions for 2 h. (e) The apoptosis rate of tumor cells under physiological FSS and CS for 24 h. (f) The apoptosis rate of tumor cells under high level of FSS for 24 h. (g) The apoptosis rate of tumor cells under high level of FSS and physiological CS for 24 h. (h) The statistical data of the tumor apoptosis rates under different conditions for 24 h. **Conditions that were statistically different (P < 0.01). All results are shown plotted as mean ± SD with each group containing three experiments.
Figure 4. The single or combined impact of FSS and CS on the adhesion of the tumor cells on the chip.
(a) The effect of different levels of FSS on the adhesion of the tumor cells on the ECs monolayer. (b) The effect of different levels of CS on the adhesion of the tumor cells on the ECs monolayer. (c)The adhesion of the tumor cells (HeLa, green) on the ECs monolayer (HUVEC, orange) under different mechanical conditions simulating microenvironments of main artery, medium-sized artery, and capillary respectively for 2 h. Scale bar, 10 μm. (d) The statistical result of (c). **Conditions that were statistically different (P < 0.01).
Figure 5. Tumor cells-ECs monolayer interaction in TNF-α conditioned biochemical environment.
(a) The morphological change of the ECs monolayer in the absence or in the presence of TNF-α. (b) The adhesion of the tumor cells on the ECs monolayer and their morphological changes in the absence of TNF-α. (c) The adhesion of the tumor cells on the ECs monolayer and their morphological changes in the presence of TNF-α. In (b,c), the tumor cells were stained with Calcein Green AM. Scale bar, 20 μm. (d) The number of adhered tumor cells on the ECs monolayer in the absence or presence of TNF-α. (e) The spreading area of adhered tumor cells on the ECs monolayer in the absence or presence of TNF-α. (f) The aspect ratio of the adhered tumor cells in the absence or presence of TNF-α. (g) The result of flow cytometry indicates the apoptosis rate of the tumor cells in the absence of TNF-α at static condition for 24 h. (h) The result of flow cytometry indicates the apoptosis rate of the tumor cells in the presence of TNF-α at static condition for 24 h. (i) Statistical data of the tumor cell apoptosis in the absence or presence of TNF-α at static condition for 24 h. *Conditions that are statistically different (P < 0.05). **Conditions that were statistically different (P < 0.01). All results are shown plotted as mean ± SD with each group containing three experiments. Scale bar, 20 μm.
Figure 6. HeLa cells in whole blood on the chip.
(a) The adhesion of HeLa cells on the wall of the simulated major artery. (b) The adhesion of HeLa cells on the wall of the simulated medium-sized artery. (c) The adhesion of HeLa cells on the wall of the simulated capillary. (d) The statistic result of the numbers of adhered HeLa cells on the walls of different simulated blood vessels. **Conditions that are statistically different (P < 0.01). All results are shown plotted as mean ± SD with each group containing three experiments. Scale bar, 100 μm.
Figure 7. Pt-NPs treatment on the model.
(a) The morphological changes of the ECs monolayers under control conditions, treated with TNF-α, treated with TNF-α + Pt-NPs, respectively. (b) The areas of the tumor cells adhered on the ECs monolayer under three different conditions: control, treated with TNF-α, treated with TNF-α + Pt-NPs, respectively. (c) The number of adhered tumor cells on the ECs monolayer under three different conditions: control, treated with TNF-α, treated with TNF-α + Pt-NPs, respectively. *Conditions that are statistically different (P < 0.05). All results are shown plotted as mean ± SD with each group containing three experiments.
References
- Reymond N., d’Agua B. B. & Ridley A. J. Crossing the endothelial barrier during metastasis. Nat. Rev. Cancer 13, 858–870 (2013). - PubMed
- Chen Z. et al. Patterning mammalian cells for modeling three types of naturally occurring cell-cell interactions. Angew. Chem. Int. Ed. 48, 8303–8305 (2009). - PubMed
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