[Study on inducing differentiation of human amniotic epithelial cells into insulin secreting cells in vitro].
OBJECTIVE
To investigate the differentiation of human amniotic epithelial cells (hAECs) into insulin secreting cells (ISCs) in vitro.


METHODS
The hAECs were isolated from human amnion by trypsin digestion, and the phenotype of the isolated cells were identified by flow cytometry and immunocytochemical staining. The hAECs at passage 3 were treated with nicotinamide and N2 supplement to investigate their differentiation into ISCs. At different times after differentiation, the expression of insulin and beta2 microglobulin (beta2-MG) was determined by immunocytochemical staining, while the content of insulin in supernatant from cultured hAECs was detected by radioimmunoassay and the expressions of insulin, pancreatic and duodenal homeobox factor-1 (PDX-1) mRNA were detected by reverse transcriptase-polymerase chain reaction (RT-PCR).


RESULTS
(1) hAECs expressed high percent of CD29, CD73, CD166 and CK19. (2) At 7, 14 and 21 days, the percentages of insulin-positive cells in induced groups were 74.00% +/- 1.73%, 75.33% +/- 1.15% (see symbol) 75.67% +/- 0.58% respectively, which were negative in control groups. (3) At 7, 14 and 21 days, contents of insulin in supernatant from induced groups were (328.47 +/- 3.22) microIU/ml, (332.26 +/- 1.22) microIU/ml and (329.68 +/- 2.57) microIU/ml respectively, they were significantly higher than those in control groups (All P < 0.01). (4) PDX-1 mRNA and beta2-MG were expressed before and after the induction of hAECs, but insulin mRNA was expressed only in the induced groups.


CONCLUSIONS
hAECs can differentiate into ISCs, having the potential application for therapy of type I diabetes.
Key Laboratory of Cell Engineering of Guizhou Province, Affiliated Hospital of Zunyi Medical College, Zunyi 563003, China.
Yu-Jie
YJ
Zhao
Ning
N
Fang
Dai-Xiong
DX
Chen
Li-Mei
LM
Yu
Huang-Fei
HF
Yu
Chun-Hua
CH
Zhao
D000650:Amnion / Q000166:cytology*; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000502:physiology*; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000166:cytology*; D005434:Flow Cytometry; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000378:metabolism; D050417:Insulin-Secreting Cells / Q000166:cytology*; D012333:RNA, Messenger / Q000235:genetics; D015534:Trans-Activators / Q000378:metabolism; D001613:beta 2-Microglobulin / Q000378:metabolism
D018398:Homeodomain Proteins; D007328:Insulin; D012333:RNA, Messenger; D015534:Trans-Activators; D001613:beta 2-Microglobulin; C084010:pancreatic and duodenal homeobox 1 protein