The Research On NGF Promoting Schwann Cell Proliferation And Up-regulated Expression Of NF-KB

Objective: It was found that NGF can promote the proliferation ofSchwann cells. Meanwhile, expression of NF-KB can be observed in Schwanncells that are proliferating. And it has been clear that NF-KB has a role ofanti-apoptosis and promoting proliferation of cells. This study was designed toexplore the relationships between NGF and SCs proliferation and up-regulatedexpression of NF-KB, aiming at supporting data for further research onproliferation mechanism of Schwann cells.Methods: Ten healthy rabbits were included in this experiment. The sciaticnerve was removed under aseptic condition after rabbits received e anesthesia.Schwann cells were isolated and cultured. The S-1000immunocytochemistrywas used to identify cell purity. The Schwann cells were plated at1×105/ml inthree Petri dishes with rat tail collagen: experimental group (adding40ng/mlof NGF); blockage group (adding of NGF+anti-NGFR); control group (addingan equal volume PBS). Then cell cultured dishes were placed in a37°C with5%CO2for5days. The S-100fluorescence labeled cells were observed under afluorescence microscope and counted. The RNA and protein were extracted for related tests. Each experiment was repeated in three times.Result: After the first24h, the adherent SCs with clear form can beobserved under the inverted contrast microscope. There were two kinds of cells:Schwann cells and fibroblasts. Schwann cells have elongated or round nucleus,dimensional sense of strong and very full of spindle-shaped body. And bothends of the elongated protrusions around the cell body were long brightband. The other kind of cells is fibroblasts with shallow nucleus and2to3nucleoli. S-100immunohistochemical staining showed that the purity ofSchawann cells cultured in vitro was about91%。The OD values results showed:In NGF group, at the24~72h, proliferation rate of Schwann cells increasedobviously. It reached the peak at5thdays and then slowed down gradually.There were obvious difference at each time point between NGF group and thecontrol. CLSM imaging displayed: NF-KB green fluorescent could be observedmostly in cytoplasmic domain in both the experimental group and the NGFgroup; there are significant difference between the experimental and the normalgroup, but not between the experimental and the control. The results showed: theexperimental group and NGF group had significant differences (p <0.01), whilethe control group and the experimental group, there was no significantdifference (p>0.05). The real-time PCR results showed that: compared withControl group, blocked group, there was significant increase of expression ofNF-KB; but no difference can be detected between the control and the blockedgroup. Conclusions:1. SCs can be separated and proliferated effectively by tissueanchored combined with differential attachment, low oncentrations of trypsindigestion and G-418.2. Adding exogenous NGF (40ng/L) can significantlyimprove the appreciation rate of the SC, with obvious up-regulated expression ofNF–KB.3. Up-regulated expression of NF-KB can be observed in the SCs that areproliferating, and the expression of NF-KB in fastest-growing SCs is the highest.And it has been clear that NF-KB has a role of anti-apoptosis and promotingproliferation of cells. It is concluded that NF-KB can promote SCs proliferation,but further research should be carried out to confirm its direct relationship onSCs proliferation.