Culture medium and culture method for esophageal cancer tumor organoid culture
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A culture method and technology for esophageal cancer, applied in the field of organoid culture, can solve the problems of slow growth of organoids, low culture success rate, long culture cycle, etc., to increase the number and survival rate, inhibit apoptosis, increase the number of production and The effect of viability
Active Publication Date: 2022-07-29
杭州艾名医学科技有限公司
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[0009] The technical problem to be solved by the present invention is to solve the problems of esophageal cancer organoids in the prior art that are usually prone to apoptosis during the culture process, the success rate of culture is low, the culture period of the original method is long, and the growth of organoids is slow. Provide an esophageal cancer organoid medium and culture method capable of improving the success rate of esophageal cancer organoid culture and the number of organoids formed, and shortening the culture cycle
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Embodiment 1
[0049] A culture medium for esophageal cancer organoids, the culture medium is composed of Advanced DMEM / F12 basal medium and specific addition factors 1×Glutamax (2mmol / L), 1×HEPES (10mmol / L), 1mmol / L N- Acetylcysteine, 10mmol / L Nicotinamide, 1×Penicillin-Streptomycin (0.1mg / mL), 1×B27, 10mmol / LSB202190, 500nmol / L A8301, 100μmol / L Gastrin, 100ng / mL R-spondin1, 100ng / mLNoggin, 50ng / mL EGF, as well as RKI-1447 and Zeaxanthin dipalmitate, where the concentration of RKI-144 was 100μmol / L and the concentration of Zeaxanthin dipalmitate was 5mmol / L.
[0050] A method for culturing esophageal cancer organoids, comprising the following steps:
[0051] 1) Obtain fresh esophageal cancer tissue (from Hangzhou Cancer Hospital, Ethics Committee: Hangzhou Cancer Hospital Ethics Committee, approval number: HZCH-2022), rinse repeatedly in pre-cooled PBS to remove obvious blood stains on the tissue.
[0052] 2) Cut it into pieces with scissors, transfer it to a 1.5mL Eppendorf tube, add it ...
Embodiment 2
[0060] Effects of different concentrations of RKI-1447 and Zeaxanthin dipalmitate on the culture of esophageal cancer organoids.
[0061] This Example 2 is basically the same as Example 1, except that the concentrations of RKI-1447 and Zeaxanthin dipalmitate in the esophageal cancer organoid culture medium are different; the details are shown in Table 1 below.
[0062] Table 1
[0063]
[0064] From Table 1 above, it can be seen that different concentrations of RKI-1447 and Zeaxanthin dipalmitate have a great effect on the number and viability of esophageal cancer organoids after culture. When 5 mmol / L was added, the number and viability of organoids could be significantly increased after 6 days of culture, and within this range, the higher the concentration of RKI-1447, the more obvious the effect. It shows that RKI-1447 can promote and inhibit the apoptosis of esophageal cancer organoids under the above addition amount. When the Zeaxanthin dipalmitate concentration was ...
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Abstract
The invention discloses a culture medium and a culture method for esophageal cancer tumor organoid culture. The culture medium for culturing the esophageal cancer tumor organoid comprises a basic culture medium and specific addition factors, and is characterized in that the specific addition factors comprise RKI-1447 with the use concentration of 10 to 100 [mu] mol/L and Zeaxanthin dipalmitate with the use concentration of 1 to 6 mmol/L. The invention further discloses a preparation method of the culture medium for culturing the esophageal cancer tumor organoid. The RKI-1447 is added into the esophageal cancer primary organoid culture solution, serves as a ROCK specific inhibitor and can inhibit apoptosis of separated esophageal cancer primary organoids, and the culture success rate and the growth rate of the esophageal cancer organoids are increased. The Zeaxanthin dipalmitate is added into the culture medium to serve as cell membrane surface fat connexin, so that information exchange between organoid cell membranes is promoted, the stability of a microenvironment in the culture process is maintained, cell nutrition metabolism is participated, and the number and motility rate of esophageal cancer organoid are increased. The RKI-1447 and the Zeaxanthin dipalmitate have a synergistic effect with each other, so that the generation quantity and the motility rate of the organoid can be remarkably improved.
Description
technical field [0001] The invention relates to the technical field of organoid culture, in particular to a culture medium and a culture method for esophageal cancer tumor organoid culture. Background technique [0002] Esophageal cancer is a highly malignant cancer, and the most commonly used models to study esophageal cancer are cell lines and patient-derived xenografts (PDX). Both model systems have considerable disadvantages. Tumor cell lines are mutated during culture and cannot mimic the original features of tumors well. Furthermore, cell cultures cannot mimic the interactions of tumor cells and other stromal cells in vivo because the cultured cells are single and lack the hierarchy of different cell types. PDX has a wide range of applications, however it does not adequately reflect the genetic characteristics and heterogeneity of human tumors. PDX is time-consuming, labor-intensive, long-term in culture, inefficient and difficult to work with high-throughput drug s...
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