Effect of carboxymethylated chitosan on expression of c-fos and c-jun and cell cycle in rat Schwann cells cultured in vitro

Abstract

Objective To investigate the effects of carboxymethylated chitosan (CMCS) on Schwann cells (SCs) proliferation and expression of c-fos,c-jun and Cyclin E.Methods Twenty 3-5 days old SPF SD rats were ures,weighing 25-30 g,take the sciatic nerves in vitro SCs,via S-100 immunofluorescence markers.In the logarithmic growth phase 2nd generation SCs,application methyl thiazol tet-razolium (MTT) and BrdU staining for different concentrations of CMCS SCs proliferation.The SCs are divided into four groups,adding a final concentration of CMCS adjustment 50 (B group),100 (C group),200 mg/L (D group) and control group (A) an equal volume phosphate buffer (PBS) ; reverse tran-scriptase PCR assay within SCs c-fos,c-jun and Cyclin E gene expression;Western blotting assay c-fos,c-jun and Cyclin E protein expression changes.Results S-100 immunofluorescence markers showing more than 90％ positive cells.MTT results showed that each group SCs absorbance (A) values were:control group (0.141 3 ±0.005 2),10 mg/L CMCS treatment group (0.152 5 ±0.006 1),50 mg/L CMCS treat-ment group (0.164 8 ±0.007 2),100 mg/L CMCS treatment group (0.172 6 ±0.006 2),200 mg/L CMCS treatment group (0.193 8 ±0.008 1),500 mg/L CMCS treatment group (0.194 7 ±0.006 2),with the concentration increased proliferation CMCS.CMCS BrdU staining results showed that in the concentration range of 50-200 mg/L can promote SCs DNA synthesis and increased with increasing concentrations of CMCS,200 mg/L CMCS most significant effect,compared with control group increased by 2 to 3 times.Reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting analysis showed,B,C,D group c-foc,c-jun and Cyclin E mRNA and protein expression were significantly higher in group A (P ＜0.05) ;among the most significant effect D group.Conclusion CMCS can promote the SCs proliferation and promotes cell cycle related gene c-fos,c-jun and Cyclin E expression. Key words: Carboxymethylated chitosan ; Schwann cells ; Proliferation ; Cyclin