Screening method and application of tumor metastasis starting cells
A technology for initiating cells and tumor metastasis, applied in the field of cell screening, it can solve the problems of few tumor types, low enrichment rate, long time-consuming, etc., and achieve the effects of simple operation, high enrichment efficiency, and wide application.
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Embodiment 1
[0048] Breast cancer cells SKBR3 cells were suspended in cell culture medium and loaded into the circulatory system for 12h at 20dyne / cm 2 After blood flow shear stress treatment, its transfer ability and self-renewal ability were tested.
[0049] The detection step of metastasis ability is as follows: add the same number of cells in the control group and the treatment group to the upper layer of the 8 μm pore size Transwell chamber containing the culture medium containing low serum. Due to the attraction of the high serum in the lower layer, the tumor cells with strong migration ability can pass through the chamber to reach the transwell chamber. lower end.
[0050] The result is as image 3 shown, tumor cells treated with blood flow shear stress versus untreated cells (Control) and cells treated only in suspension (0 dyne / cm 2 ) more cells reached the bottom of the chamber, indicating a stronger migration ability.
[0051] The steps of cell self-renewal ability detection ...
Embodiment 2
[0054] The same number of tumor cells (20 dyne / cm 2 ), only suspension-treated cells (0 dyne / cm 2 ), untreated control cells (Control), and positive control cells (Tic) cultured by the tumor stem cell culture method were injected subcutaneously in nude mice, and tumor growth was measured regularly after 14-34 days of growth. .
[0055] The result is as Figure 5 shown, 20dyne / cm 2 Tumor cells treated with blood flow shear stress have stronger tumor-forming ability in nude mice.
Embodiment 3
[0057] The CXCR4 pathway plays an important role in the process of tumor metastasis. In this example, different concentrations of CXCR4 inhibitor-AMD3465 were selected to inhibit the activity of the CXCR4 pathway. Tumor cells were first treated with blood flow shear stress for 12 hours, then added with different concentrations of inhibitors and cultured for 24 hours, and then their metastatic ability was detected by Transwell chamber.
[0058] The result is as Image 6 As shown, with the increase of inhibitor concentration, the migration ability of the selected tumor-initiating cells also decreased gradually. It was shown that inhibiting the CXCR4 pathway can significantly inhibit the metastasis ability of the screened metastasis-initiating cells.
[0059] To sum up, the present invention mainly uses a peristaltic pump to generate blood flow shear stress that meets physiological conditions to screen metastasis-initiating cells that can effectively generate metastatic tumors. ...
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