Inducing expression of β-nerve growth factor and its effect on limbal stem cells

doi:10.16098/j.issn.0529-1356.2017.04.009

Abstract

Abstract:

Objective To construct recombinant lentiviral vector carrying rat β-nerve growth factor (β-NGF) gene by inducing expression of the Tet-on 3G tetracycline induced expression system, and to observe its overexpression in HEK293FT cells and effects on limbal stem cells（LSCs）. Methods After transfection, β-NGF protein was detected to express on HEK293FT cells following the increase of doxycycline (Dox)dose. After primary cultured, LSCs were allocated to four groups with different doses of Dox: control group（non-transfection group）,1000, 100 and 1000μg/L of Dox inducing groups (A,B,C). The expressions of p63,NGF,p38,TrkA were observed. Results After transfection, β-NGF protein was detected to express on HEK293FT cells and LSCs following the increase of Dox dose. The experimental groups expressed different quantities of p63,p38, and TrkA, which was statistically significant. Conclusion The β-NGF gene could be expressed successfully on LSCs by Tet-on 3G induced expression system. These result highlight that NGF is benefit LSCs amplification in vitro,and maintain their stemness.

Key words:

β-nerve growth factor, Tet-on 3G tetracycline induced expression system;Doxycycline, Limbal stem cells, Western blotting, Immunofluorescence, Rat

GAO Jie DING Ling HU Rong HUANG Yue SU Min LI Hong. Inducing expression of β-nerve growth factor and its effect on limbal stem cells[J]. Acta Anatomica Sinica, 2017, 48(4): 421-427.

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