The invention discloses a 
simulation three-dimensional 
incubator and a culture method for 
mouth mucosa epithelial cells and applications of the 
incubator and the culture method. The 
incubator comprises a single or a plurality of basic structural units, wherein each basic 
structural unit consists of an inner container and an outer container which are tightly nested with each other, and are prepared from an organic material for 
cell culture; and the bottom of the inner container consists of a permeable supporting film, and a certain gap is formed between the bottom of the inner container and the bottom of the outer container. The incubator takes the factors of 
nutrient supply of the 
mouth mucosa epithelial cells, blood vessels and 
blood vessel endothelial cells into account, and the growing environment of the 
mouth mucosa epithelial cells in the incubator is close to that in a 
human body, so that the incubator can be used for physiological, pharmacological, toxicological and micro-ecological studies of mouth mucosas, and can also be used for constructing human 
tissue engineering mouth mucosas and repairing and reconstructing mucosal 
coloboma. The new technology of the 
simulation three-dimensional incubator promotes an 
in vitro culture model of a mouth mucosa to a new height, so that related studies are closer to clinical practices.