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| Experimental study on culturing Schwann cells of neonatal rats in vitro |
| ZHOU Min HU Ming HE Songming HUANG Guotao CHEN Qian HONG Li |
| Department of Gynecology and Obstetrics, People's Hospital of Wuhan University, Hubei Province, Wuhan 430060, China |
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Abstract Objective To explore a culture of SCs method with higher cell purity and shorter time by comparing the two methods including the double-enzyme digestion and single-enzyme digestion of primary culture of Schwann cells (SCs) using the neonatal rats. Methods Bilaterally sciatic nerves were dissected from 4 to 5-day-old neonatal SD rats and double-enzyme digestion method and single-enzyme digestion method which both combined with hemi-explants-culture method, differential digestion method, differential adhesion method, and cytosine arabinoside were used to culture and purify SCs, respectively. The morphology of the SCs was observed under the inverted microscope at 48 hours and 6 days of culture. Then the SCs were digested and passaged after 7 days in culture and counted under the cell counter. The purity of SCs was identified by S-100 immunofluorescence staining. CCK-8 was used to measure the P3 cells growth curve, respectively. Results With the same number of neonatal rats, double-enzyme digestion method acquired SCs at least twice than single-enzyme digestion method. (95.29±1.71)% of S-100 positive SCs cultured were shown by immunofluorescence staining in the double-enzyme digestion method group, and (72.41±2.16)% in the single-enzyme digestion method group (P < 0.05). There was no difference between the two cultures in the growth curve of P3 cells (P > 0.05). Conclusion The double-enzyme digestion combined with hemi-explants-culture method, differential digestion method, differential adhesion method, and cytosine arabinoside may obtain sufficient amount of high-purity SCs in a short time and be applied in the further cytological experimental research on peripheral nerve regeneration.
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