粒腺體功能異常抑制神經生長因子所調控之PC12細胞神經軸突的生長及細胞凋亡現象

Abstract

粒腺體在細胞中扮演著重要的角色，它除了是細胞內提供能量的主要來源，也調節著細胞內鈣離子的濃度，更是調控細胞凋亡的途徑中不可或缺的胞器。因此，當粒腺體功能異常就會直接影響到細胞內生理功能。在神經系統中，神經細胞的生長與分化是需要粒腺體提供正常功能才得以維持的生理行為，過去的研究顯示，粒腺體功能的缺失異常被認為是造成神經功能異常現象或是神經退化性疾病的主要原因之一。在本研究中，我們利用老鼠嗜鉻細胞瘤（rat pheochromocytoma cells，PC12 cells），經神經生長因子（nerve growth factor，NGF）誘導分化後具有類神經細胞功能的特性，並採用五胺基酮戊酸光動力方式（5- aminolevulinic acid mediated photodynamic treatment，ALA- PDT）反覆多次處理，選殖出穩定具有粒腺體功能缺失的PC12變異株細胞，進而觀察粒腺體功能缺失對PC12細胞的影響為何？ 研究結果顯示：（1）利用五胺基酮戊酸光動力方式（ALA-PDT）反覆多次處理確實造成PC12細胞粒腺體功能缺失，也觀察到細胞內鈣離子有下降現象，並且使得細胞體積上有變大的現象。（2）當存在有神經生長因子（nerve growth factor）的狀況下，粒腺體功能缺失的PC12變異株細胞分化後的外觀發生變化，並且神經突觸（Neurite）的分化延長能力上受到阻礙。（3）缺乏神經生長因子，在無血清（serum- free）或低血清（low- serum）的培養狀況下，粒腺體功能缺失的PC12變異株細胞發生細胞凋亡（apoptosis）現象下降。（4）粒腺體功能缺失的PC12變異株細胞，細胞內ERK分子活化現象降低；而JNK分子活性則無明顯變化。（5）經由生物晶片（microarray）分析，PC12變異株細胞內與鈣離子、神經等等相關的基因表現量有下降現象。

In cells, mitochondria are the major ATP source produced by oxidative phosphorylation. In addition, mitochondria are critical for other aspects of cell function, such as modulating calcium levels. Consequently, mitochondrial dysfunction contributes to a wide range of human diseases, including neurodegenerative diseases. Neurons are highly dependent on oxidative energy metabolism, therefore normal mitochondrial function is the key pathogenic factor in a number of neurodegenerative disorders. In our study, using 5- aminolevulinic acid mediated photodynamic treatment（ALA-PDT）, PC12 variants with dysfunctional mitochondria were established. The purpose of this study is to address the post-PDT responses in PC12 cells survived from mitochondrial photodamage induced by ALA-PDT. We found：（1）ALA-PDT could be used to establish enlarged PC12 variants with dysfunctional mitochondria.（2）In the presence of NGF, morphology was changed and neurite elongation was suppressed in differentiated PC12 variants with dysfunctional mitochondria.（3）In the absence of NGF, apoptotic cell death was suppressed in PC12 variants with dysfunctional mitochondria in serum-free or low-serum medium.（4）Activated ERK decreased in PC12 variants with dysfunctional mitochondria, but JNK not.（5）In mRNA microarray analysis, expression of calcium- and neuro- related gene decreased in PC12 variants with dysfunctional mitochondria.