Coculture of rat oral keratinocytes with inactive 3T3 fibroblasts
- VernacularTitle:兔口腔黏膜细胞与灭活的3T3成纤维细胞的体外共培养
- Author:
chao, LI
;
yue-min, XU
;
lu-jie, SONG
;
lei, CUI
;
shuo, YIN
- Publication Type:Journal Article
- Keywords:
oral keratinocytes;
3T3 cells;
urethra;
tissue engineering
- From:Journal of Shanghai Jiaotong University(Medical Science)
2006;0(11):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To explore the cultural method of oral keratinocytes and make preparations for further investigation in using oral keratinocytes as a new choice of seed cells for the reconstruction of tissue-engineered urethra. Methods Oral keratinocytes of rabbits were isolated in vitro and seeded onto a culture dish with a feeder layer of 3T3 mouse fibroblasts inhibited by mitomycin(i3T3) or a culture dish without i3T3 respectively. Cell morphology was observed and cell growth was detected at intervals.Meanwhile,oral keratinocytes obtained from in vitro culture were performed immunofluorescence staining with broad-spectrum keratin antibody(AE1/AE3) and keratin 19 antibody(K19).The percentage of positive cells of passage 2 reactive to AE1/AE3 was assessed by flow cytometry. ResultsOral keratinocytes seeded onto a feeder layer of i3T3 exhibited finer morphous,better amplification capability,and could be passed for 7 or 8 generations.However,those cultured without i3T3 took on various morphous and could only be subcultured 2 generations before ageing.It was indicated by immunofluorescence staining that oral keratinocytes obtained from in vitro culture were positive for AE1/AE3 staining and 40% were positive for K19 staining.The result of flow cytometry revealed that the amout of positive keratinocytes reactive to AE1/AE3 was more than 95% of total cellular score. Conclusion Oral keratinocytes of rabbits can be cocultured with i3T3 in vitro and magnitude quantity can be attained,laying a favourable foundation for oral keratinocytes as a new choice of seed cells for urethral reconstruction with tissue engineering.
