Culture method and application of human colorectal cancer tissue organoid

Abstract

The invention discloses a culture method of human colorectal cancer tissue organoid, which comprises the following steps: washing human colorectal cancer tissues and putting them into a transfer culture medium, cleaning and shearing; further cleaning, centrifuging, carrying out enzyme digestion, filtering, and collecting cell clusters; and mixing the cell clusters with hydrogel, adding DMEM culture medium to culture after solidification to obtain the human colorectal cancer tissue organoids. According to the invention, gentamicin, amphotericin B, primocin and other antibiotics are added into the transfer culture medium, the digestion solution and the culture medium, so that microbial contamination can be reduced, and the method for obtaining the human colorectal cancer organoids through in-vitro culture is simple, convenient, rapid, high in success rate and good in organoid growth state. The invention further discloses application of the human colorectal cancer organoid in drug testing, and the co-culture model of human colorectal cancer tissue organoids and tumor-associated fibroblasts can be used as a patient individualized drug testing platform.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and in particular relates to a method for culturing human colorectal cancer tissue organoids and an application thereof. Background technique [0002] The human-derived tumor organoid (patient derived organoid, PDO) model is a new type of tumor research model that has emerged in recent years. Hans Clevers et al. proposed for the first time in 2009 the use of matrigel to culture mouse small intestinal crypt epithelial cells and Lgr5+ intestinal cells in vitro. Stem cells that can form microstructures similar to intestinal glands mimic the structure and function of the small intestine in vivo. Studies have shown that PDO and tumor tissue samples have highly similar genomic features and tumor molecular subtypes; at the same time, it reveals the importance of genotype-phenotype analysis at the single patient level. [0003] At present, almost all the establishment of organoids rely on animal-...

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Problems solved by technology

[0004] Brian et al. reported the successful construction of HA-based hydrogels for prostate cancer cell culture using aqueous solutions of thiolated HA and disulfide-containing polyethylene glycol diacrylate (PEGSSDA); the material The synthesis cost is low, the composition is precise and controllable, and it is transparent, stable and porous, and can support the growth of tumor cells and stromal cells at the same time; this study successfully used this HA-based hydrogel as a catalyst for prostate cancer tumor cells and stromal cells. Multilayer cultured biomaterials have constructed a high-throughput drug testing platform, but so far there have been no reports on the use of hydrogels to establish human colorectal cancer (CRC) organoids in vitro

[0005] The PDO model contains only epithelial cells, but does not contain mesenchymal cells in the original tumor tissue, which has become the most important defect of PDO, leading to its inaccurate clinical response in some applications, and seriously hindering its role in immune defense, Applications in tumor metastasis and invasion, microenvironment regulation, etc.

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