Viability of cultured nerve grafts: An assessment of proliferation of Schwann cells and fibroblasts

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Abstract
Previous studies demonstrated that the viability of nerve grafts had a positive effect on nerve regeneration, while the cold storage of nerve grafts obtained few viable cells at the later stage. The purpose of this study was to examine the cellular activities of Schwann cells and fibroblasts in cultured nerve grafts prior to transplantation. 2.5-cm long sciatic nerve grafts were harvested from 75 male Lewis rats. Two different media were utilized to culture the nerve grafts up to 3 weeks: Dulbecco's modified eagle medium (DMEM) only or DMEM supplemented with 2 μM forskolin and 10 μg/ml pituitary exact (mitogen medium for Schwann cells). In vivo predegenerated and normal nerve grafts were used as positive and negative controls, respectively. We employed a 5-bromo-2′-deoxyuridine (BrdU) incorporation method to evaluate the proliferating cells in the cultured nerve grafts. S-100 and vimentin immunostaining were used to estimate the presence of Schwann cells and fibroblasts in all nerve grafts at different intervals. The results showed that the proliferating cells increased progressively under culture conditions. The proliferating cells distributed evenly in small fascicles (average diameter 251 ± 71.5 μm), whereas they appeared mainly in the margin of large fascicles (average diameter 624 ± 87.3 μm). The mitogen medium stimulated Schwann cell multiplication more significantly in comparison with DMEM after 3 days of culture (P < 0.01), however, there were fewer fibroblasts present in the mitogen medium than in DMEM after 2 days of culture (P < 0.01). It is suggested that the viability of nerve grafts can be preserved under culture conditions. Furthermore, the cellular activity of the Schwann cells and fibroblasts in nerve grafts can be manipulated in in vitro Wallerian degeneration. © 1999 Wiley-Liss, Inc. MICROSURGERY 19:356–363 1999