Culture of human intestinal epithelial cell using the dissociating enzyme thermolysin and endothelin-3
Braz. j. med. biol. res
;
43(5): 451-459, May 2010. tab, ilus
Article
Dans Anglais
| LILACS
| ID: lil-546337
ABSTRACT
Epithelium, a highly dynamic system, plays a key role in the homeostasis of the intestine. However, thus far a human intestinal epithelial cell line has not been established in many countries. Fetal tissue was selected to generate viable cell cultures for its sterile condition, effective generation, and differentiated character. The purpose of the present study was to culture human intestinal epithelial cells by a relatively simple method. Thermolysin was added to improve the yield of epithelial cells, while endothelin-3 was added to stimulate their growth. By adding endothelin-3, the achievement ratio (viable cell cultures/total cultures) was enhanced to 60 percent of a total of 10 cultures (initiated from 8 distinct fetal small intestines), allowing the generation of viable epithelial cell cultures. Western blot, real-time PCR and immunofluorescent staining showed that cytokeratins 8, 18 and mouse intestinal mucosa-1/39 had high expression levels in human intestinal epithelial cells. Differentiated markers such as sucrase-isomaltase, aminopeptidase N and dipeptidylpeptidase IV also showed high expression levels in human intestinal epithelial cells. Differentiated human intestinal epithelial cells, with the expression of surface markers (cytokeratins 8, 18 and mouse intestinal mucosa-1/39) and secretion of cytokines (sucrase-isomaltase, aminopeptidase N and dipeptidylpeptidase IV), may be cultured by the thermolysin and endothelin-3 method and maintained for at least 20 passages. This is relatively simple, requiring no sophisticated techniques or instruments, and may have a number of varied applications.
Texte intégral:
Disponible
Indice:
LILAS (Amériques)
Sujet Principal:
Bacillus thermoproteolyticus neutral proteinase
/
Techniques de culture cellulaire
/
Cellules épithéliales
/
Muqueuse intestinale
/
Intestin grêle
Limites du sujet:
Humains
langue:
Anglais
Texte intégral:
Braz. j. med. biol. res
Thème du journal:
Biologie
/
Médicament
Année:
2010
Type:
Article
Pays d'affiliation:
Chine
Institution/Pays d'affiliation:
Shanghai Jiao Tong University Affiliated Sixth People's Hospital/CN
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