Studies On The Purification, Culture And Proliferation Agents Of Rat Olfactory Ensheathing Cells

Objective:â‘ To establish an economic, simple, effective method for primary culture and purification of Olfactory ensheathing cells (OECs) in adult rats with modified differential adherence and serum-free conditioned medium supplemented with supernatant of OECs culture.â‘¡Fully explored the proliferation effect of conditioned medium consist of serum,Forskolin,basic fibroblast growth factor (bFGF) and a variety of combined proliferation agent on OECs, to solve the quantity and source problem of OECs in the basic and clinical research.Methods:â‘ OECs was sterilely collected from Olfactory bulb and digested for resuspendion. The diluted monoplast suspension was equally assigned into 3 samples, and separately used for simple modified differential adherence, simple serum-free conditioned medium, and modified differential adherence + serum-free conditioned medium. After intervention by 12 h + 24 h differential adherence purification and serum-free conditioned medium for 36-48 hours, the OECs growth condition and morphological features of each group were observed using inverted microscope. OECs were subjected to immunofluorescence, and the purity of OECs was calculated.â‘¡With the principle of randomization, the purified OECs were divided into six groups: blank control group, conditioned medium consist of serum group,Forskolin group, bFGF group, Forskolin + bFGF group and the combined proliferation agent group. After intervention, the proliferation effect of each group was detected by 5-bromodeoxyuridine incorporation and MTT activity assay.Results:â‘ Following modified differential adherence + serum-free conditioned medium, the purity of OECs was 88%~92%, which was obviously superior to another control groups.â‘¡The conditioned medium consist of serum and Forskolin could promote the vitality and number of OECs compared with the control group (P<0.05), and their effect were moderate. However,the bFGF could significantly increase the vitality and number of OECs compared with the control group (P<0.001), and its effect was relatively intense. In addition, Forskolin+bFGF group and the combined proliferation agents group could significantly improve the number of OECs (P<0.001). It was clear that combined application of these proliferation agents had a synergy effect.Conclusion :â‘ This experiment establishes a set of the novelty culture and purification method for OECs isolated from adult rats.â‘¡The conditioned medium consist of serum, Forskolin and bFGF have the proliferation effect on the OECs. Combined three proliferation agents could significantly promote the vitality and number of OECs, then combined application may be used as a means to amplify primary OECs...