Objective: The functional roles of protein kinase C (PKC) in the neurite outgrowth and nerve regeneration remain controversial. The present study was aimed to investigate the role of PKC in neurite outgrowth, by studying their regulatory effects on neurite elongation in spinal cord neurons in vitro.
Methods: The anterior-horn neurons of spinal cord from embryonic day 14 (E14) Sprague-Dawley (SD) rats were dissociated, purified and cultured in the serum-containing medium. The ratio of membrane-PKC (mPKC) activity to cytoplasm-PKC (cPKC) activity (m/c-PKC) was studied at different time points during culture.
Results: Between 3-11 d of culture, the change of m/c-PKC activity ratio and PKC-betaII expression in the neurite were both significantly correlated with neurite outgrowth (r=0.95, P< 0.01; r=0.73, P< 0.01, respectively). Moreover, PMA, an activator of PKC, induced a dramatic elevation in the m/c-PKC activity ratio, accompanied with the increase in neurite length (r=0.99, P< 0.01). In contrast, GF 109203X, an inhibitor of PKC, significantly inhibited neurite elongation, which could not be reversed by PMA.
Conclusion: PKC activity may be important in regulating neurite outgrowth in spinal cord neurons, and betaII isoform of PKC probably plays a major role in this process.
目的: 关于蛋白激酶C (PKC)在神经元突起生长和神经再生中的作用, 目前仍存有争议。 本研究主要观察PKC对离体培养的脊髓神经元生长的调节作用, 旨在阐明PKC对突起生长的调节作用。
方法: 分离纯化胎龄14天(E14)的SD胎鼠的脊髓前角神经元, 进行原代培养, 并检测不同时相点膜/浆PKC活性(m/c-PKC activity)的比值。
结果: 神经元培养3–11 d 期间, 神经元内m/c-PKC比值以及PKC-βII在突起中的表达水平均与突起生长呈显著相关关系(r = 0.95, P < 0.01; r = 0.73, P < 0.01)。 此外, PKC激动剂PMA能显著提高m/c-PKC比值, 且与神经突起的生长一致(r = 0.99, P < 0.01)。 而PKC抑制剂GF 109203X 则能显著抑制突起生长, 且不被PMA作用所逆转。
结论: PKC的活性在脊髓神经元突起生长调节中具有重要作用, 其中βII亚型可能扮演重要角色。