利用TF-1細胞探討細胞自戕和核醣核酸水解酵素的關係

Abstract

在細胞死亡的研究中，apoptosis（細胞自戕）逐漸成?眾多研究者感興趣的焦點。僅管真正與apoptosis進行的過程有關的現象和機制到目前?止還沒有徹底發現，但由於很多研究者的參與，使我們能對apoptosis有愈來愈深的瞭解。 進行apoptosis的細胞在型態上的變化有核濃縮及形成碎片，細胞萎縮，細胞內的胞器重組，及表面形成氣泡而使細胞逐漸碎裂成片。在大部分的例子中，進行apoptosis這種死亡的細胞內同時也有因活化DNase I (去氧核醣核酸水解酵素 I)而導至chromosomal DNA（染色體去氧核醣核酸）碎裂成片的現象。近來更發現當細胞進行apoptosis時，其內的28S rRNA（核醣體核醣核酸）會被RNase（核醣核酸水解酵素）在特定位置切斷的現象。 本論文中主要目的乃在探討TF-1細胞進行apoptosis時是否有特定之RNase被活化。實驗採用兩種方法讓TF-1細胞進行apoptosis，一?去除其生長因數，GM-CSF (granulocye-macrophage colony stimulating-factor，顆粒球細胞-巨噬細胞株落刺激因數），一?加入TPA。結果發現 TF-1細胞因去除GM-CSF而引起的apoptosis時有一些特殊的RNase可被活化，然而這些RNase在其他原因所造成的死亡現象（例如，由熱處理或DMSO引起的細胞壞死，或是由TPA引起的apoptosis）中並無發現可被活化。另外，在D2細胞（不依賴GM-CSF即可生長的TF-1突變細胞株）中發現在TF-1細胞因去除GM-CSF導致的apoptosis中所引起的、可活化的RNase也存在，某些則無。這些RNase在TPA處理過的D2細胞中亦均消失。但在TPA所引起的apoptosis卻有另外其他群的RNase被活化。 因此，由上述所發現之實驗證據可推測，在細胞產生apoptosis時有些RNase會被活化，但這些RNase的種類會因細胞種類不同或引起apoptosis的因素不同而有異。

Apoptosis, a suicide- like process, is a major focus for a wide variety of biological investigators, recently.The detail mechanism of apoptosis has not been identified thoroughly, but its morphological phenomena and the basic machinery is more and more clearly. The morphological changes of cellular apoptosis include nuclear condensation and fragmentation followed by a loss of cell viability, cell shrinkage accompanied by organelle rearrangements, and surface blebbing leading to cell fragmentation. In most cases, apoptotic cell death is also accompanying by fragmentation of chromosomal DNA resulting from the activation of DNase I. Most recently, it is even identified that 28S rRNA sites specifically cleaved in cells undergoing apoptosis. One major aim of the present study was to find those specific RNase activated in apoptotic systems. For these purpose, the RNase activity was studied in the GM-CSF depression of TF-1 cells induced apoptosis. We found that some RNase was present in activated form when GM-CSF depression no matter in TF-1 cell or D2 cell, TF-1 mutant and GM-CSF independent, but absent in other cellular death, e.g. necrosis induced by heat-shock or DMSO treatment, or apoptosis induced by TPA. But, some RNase indeed present only the GM-CSF induced apoptotic TF-1 cells. Furthermore, we also found there has RNase activity of some other groups increased by TPA induction. Therefore, we suggest that there has some specific RNase activity could be induced in cellular apoptotic death, and which kinds of RNase would be activated depends on cell types of the inducer of apoptotic system.