| Objective: Schwann cells were the peripheral nervous system primary glial cell . It had important function in repairing nerve injury. Though the recovery of nerve injury did not ideality, the nerve tissue engineering would bring goodly foreground. Due to the core of peripheral nerve tissue engineering was Schwann cells, acquired a lot of Schwann cells was the basic require of peripheral nerve tissue engineering by culture cell in vitro. In order to meet requirement, we must be solve the problem. To study the mechanism of nerve growth factor (NGF)in promotion of proliferation of Schwann cells of fetus rabbit has significant. Furthermore, the tissue of embryo did not possess integrity characteristic of biology and the system of immunity dose not grow mature. It was in harmony with the environment of autoeciousness and reaction of exclude doesn't remarkably occur. The nervous system of fetus had grown mature. The Schwann cells be cultured with fetus rabbit sciatic nerve was feasible. When the cells were transplanted, the reaction of immunity hadn't happened.Methods: Prepared before experimentation, included confecting culture medium and collagenase of tail's rat, prepared utensil used culture. Adopted pregnancy New Zealand white rabbit, it was executed and paunched and taken out fetus rabbit from the abdomen rapidly and it was dip in 75% alcohol 5 minute, then flushed out the alcohol with D-hank's. Under operation microscope, the fetus rabbit sciatic was fetched, removed the ectoblastic of sciatic nerve and cut it into piece about 0.1mm3.We adopted the method of transplant piece to culture primary Schwann cells. The Schwann cells from which primary cultured were separated and purified by the way of speed different adhesion after 14 days. The separated and purified Schwann cells from fetus rabbit sciatic nerve were divided into 2 groups; the NGF group and control group. After 24 hours the second Schwann cells cultured, the Schwann cells were observed under phase contrast microscope and counted every day. Proliferation index(PI)was index and processed statistic analysis.Results: On the sixth day, the cell was seen around fetus rabbit sciatic piece in the utensil. The cell had two type. one was form shuttle, owned 2～3 slightness prominence and a majority of cells had double pole. The ellipse nuclear of cell evident. There was a light circle in the vital cell. That is growth faint. Occasionally, another cell was visible in the utensil. Its body greatness, flat, abnormity morphology, short and many prominence and the shallow nuclear of cell has 2～3 nucleolus. Along with time process, cells forth extend and density gradually enhance. Finally, the cell had creped fully on the bottom of utensil. The cell arrayed trim in district of cell bloom. It grew flower around transplant piece after 14 days. If the ectoblast of sciatic nerve was removed very clear, the Schwann cells had highly purity. It is alone Schwann cells. Occasionally, fibroblast was visible in the utensil. After the second cultured 24 hours, the Schwann cell was observed under phase contrast microscope. There was a large of cell adhering to bottom of utensil in the experiment group and control group. A minority of cell suspended and dies. The vital cell presented form of shuttle, owned double pole and a minority of cells presented triangle. After 48 and 72 hours, the cell polymerized, presented end and end, side by side, their array of vortex-shaped or fence-shaped, ellipse karyon. The Schwann cells were confirmed by rabbit anti-S100 protein antibody immunocytochemical staining. In the 60 visual fields of 20 petri dishes, the Schwann cells were counted. Two groups mean plus/minus standard deviation of proliferation index were compared with. The result of two groups mean plus/minus standard deviation of proliferation index t-test:the third day, t value 12.62,P value <0.001；the fourth day,t value 10.29,P value <0.001；the fifth day,t value 11.55,P value <0.001；the sixth day,t value 24.88,P value <0.001；the seventh day,t value... |
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