Culture medium for normal epithelial cell of human or mammal, culture methods, normal epithelial cell and application of normal epithelial cell

Abstract

The invention relates to a culture medium for a normal epithelial cell of a human or mammal, primary separation culture, subculture, 3D gas-liquid culture and 3D matrigel culture methods, the normal epithelial cell generated by using the culture medium and the culture methods and application of the normal epithelial cell to a toxicological evaluation system. The culture medium is prepared by mixing DMEM and Ham's F-12NUTRIENT MIX according to the volume ratio of 3:1 and also adding 4-6% of fetal calf serum, 1-3nM triiodothyronine, 0.4-0.65% of insulin-transferrin-selenium reagent, 4-6mu g/ml transferrin, 9-11ng/mL epidermal growth factors, 0.3-0.5mu g/mL hydrocortisone, 0.5-1.5nM cholera toxin, 0.4-0.6mu g/mL amphoterrible B, 35-45mu g/mL gentamicin, 45-55nM calpeptin, 35-45ng/ml recombinant human IL-1RA and 3mu g/ml recombinant human R-Spondin-1. The culture medium disclosed by the invention can be used for carrying out separation culture or subculture on the normal epithelial cell of the human or the mammal and any other various tissue source, rapidly proliferating the normal epithelial cell in vitro and establishing a cell line; and the normal epithelial cell is a normal diploid cell and is applied to the toxicological evaluation system of the human or the mammal.

Description

technical field [0001] The present invention relates to the field of cell biology and toxicology, in particular to culture medium, primary separation culture, subculture, air-liquid 3D culture and Matrigel 3D culture method of human or mammalian normal epithelial cells, as well as the culture medium, culture Methods Generated normal epithelial cells and their application in toxicology evaluation system. Background technique [0002] Environmental pollution has seriously endangered people's health, and respiratory diseases caused by it have also emerged continuously. Human epithelial cells are the first to be the main damage target. Organ-specifically differentiated epithelial cells perform important functions such as gas exchange in the lungs, filtration in the kidneys, detoxification and binding in the liver, insulin secretion by pancreatic islet cells, and skin resistance to harmful environments. Therefore, epithelial cell-based in vitro models have been used to investig...

AI Technical Summary

Problems solved by technology

Although many cell immortalization methods have been established, most of them introduce exogenous viruses and proto-oncogenes into target cells through virus introduction and gene transfection. In the internal physiological pathway, cells undergo unexpected changes. Compared with normal cells, their biological properties such as karyotype and serum-dependent type may change, and some even show the characteristics of tumor cells. The established immortalized cells cannot maintain its original normal phenotype

However, so far, it has not been possible to establish a normal cell line without gene transfer and immortalization as a research model in toxicology

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