Effect of basic fibroblast growth factor gene transfection on bio-behavior of rat extraocular muscle satellite cells
- VernacularTitle:碱性成纤维细胞生长因子基因转染对鼠眼外肌成肌细胞生物学行为的影响
- Author:
Ligui ZHANG
;
Chuanfu WANG
;
Xiaohui HUANG
- Publication Type:Journal Article
- From:
Chinese Journal of Tissue Engineering Research
2007;11(33):6706-6711
- CountryChina
- Language:Chinese
-
Abstract:
BACKGROUND: Experiments have demonstrated that exogenous basic fibroblast growth factor (bFGF) promotes the growth and survival of skeletal muscle satellite cells, and endogenous bFGF also has obvious effect on muscular repair.But whether bFGF gene transfection has the same effect on extraocular muscle satellite cells is unclear.OBJECTIVE: The goal of this study was to investigate the effect of bFGF gene transfection on the bio-behavior of rat extraocular muscle satellite cells.DESIGN: Single sample observation.SETTING: Department of Ophthalmology, Qingdao University Medical College.MATERIALS: Primary rat extraocular muscle satellite cells were cultured (purity ＞ 90%). Human PEGFP-N3-bFGF eukaryotic expression vector was successfully constructed (reported in other papers).METHODS: This study was carried out in the Central Laboratory of Qingdao University Medical College between September 2005 and December 2006. Experimental intervention and grouping: During the experiment, 3 groups were divided: Experimental group, in which, recombinant PEGFP-N3-bFGF was used for transfection; Control group A, in which, empty-load PEGFP-N3 was used for transfection; Control group B: in which, only F 10 medium (0.1 volume fraction of fetal bovine serum)was used for transfection. Recombinant PEGFP-N3-bFGF plasmid was used to transfect rat extraocular muscle satellite cells cultured in vitro by liposome-mediated transgenic technology. Experimental evaluation: bFGF expression was observed by immubohistochemical method; Transfection efficiency was detected with fluorescence microscope; The protein secretion of bFGF of satellite cells in each group was detected by SBC-ELISA method on the 1st, 3rd, 5th, 9th, 11th, 13th, 15th, 21st and 28th days of culture; The proliferation of rat extraocular muscle satellite cells in each group was detected by methyl thiazolyl tetrazolium(MTT) method; Creatine kinase (CK) activity of cells in each group was determined according to the A value of some standard whose concentration was known.MAIN OUTCOME MEASURES: ① Transfection efficiency of bFGF gene to rat extraocular muscle satellite cells cultured in vitro and bFGF gene expression after transfection; ②bFGF protein secretion after transfection, and effect of bFGF gene transfection on the growth and proliferation of extraocular muscle satellite cells.RESULTS: ① Transfection efficiency of bFGF gene to rat extraocular muscle satellite cells cultured in vitrowas (42.8±1.2)%. ② Immunohistochemical detection showed positive reaction. ②Transfected cells could secrete bFGF protein,which was the highest on the 5th day (662.935 ng/L). ④ The proliferation activity of transfected cells was obviously enhanced, and its A value was significantly higher than that of control group at the same time point (P ＜ 0.05). ⑤CK value was higher than that of the control group from the 5th day after transfection to the end (P ＜ 0.01).CONCLUSION: bFGF gene transfering into rat extraocular muscle satellite cells can make extraocular muscle satellite cells to secrete bFGF, promote cell proliferation, survival and differentiation.
