In vitro pancreas duodenal homeobox-1 enhances the differentiation of pancreatic ductal epithelial cells into insulin-producing cells.
OBJECTIVE
To observe whether pancreatic and duodenal homeobox factor-1 enhances the differentiation of pancreatic ductal epithelial cells into insulin-producing cells in vitro.


METHODS
Rat pancreatic tissue was submitted to digestion by collegenase, ductal epithelial cells were separated by density gradient centrifugation and then cultured in RPMI1640 medium with 10% fetal bovine serum. After 3-5 passages, the cells were incubated in a six-well plate for 24 h before transfection of recombination plasmid XlHbox8VP16. Lightcycler quantitative real-time RT-PCR was used to detect the expression of PDX-1 and insulin mRNA in pancreatic epithelial cells. The expression of PDX-1 and insulin protein was analyzed by Western blotting. Insulin secretion was detected by radioimmunoassay. Insulin-producing cells were detected by dithizone-staining.


RESULTS
XlHbox8 mRNA was expressed in pancreatic ductal epithelial cells. PDX-1 and insulin mRNA as well as PDX-1 and insulin protein were significantly increased in the transfected group. The production and insulin secretion of insulin-producing cells differentiated from pancreatic ductal epithelial cells were higher than those of the untransfected cells in vitro with a significant difference (1.32 +/- 0.43 vs 3.48 +/- 0.81, P < 0.01 at 5.6 mmol/L; 4.86 +/- 1.15 vs 10.25 +/- 1.32, P < 0.01 at 16.7 mmol/L).


CONCLUSIONS
PDX-1 can differentiate rat pancreatic ductal epithelial cells into insulin-producing cells in vitro. In vitro PDX-1 transfection is a valuable strategy for increasing the source of insulin-producing cells.
Liu
Tao
T
Department of Pancreatic Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China.
Wang
Chun-You
CY
Yu
Feng
F
Gou
Shan-Miao
SM
Wu
He-Shui
HS
Xiong
Jiong-Xin
JX
Zhou
Feng
F
D000818:Animals; D002454:Cell Differentiation; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism*; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism*; D007328:Insulin / Q000378:metabolism*; D008297:Male; D010183:Pancreatic Ducts / Q000166:cytology / Q000378:metabolism*; D010957:Plasmids; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D018398:Homeodomain Proteins; D007328:Insulin; D012333:RNA, Messenger; D015534:Trans-Activators; C084010:pancreatic and duodenal homeobox 1 protein