星狀膠質細胞( astrocytes )是中樞神經系統中主要的膠質細胞，負責中樞神經系統的恆定、神經細胞生成、以及突觸形成等作用。在中樞神經系統發育時期，星狀膠質細胞的形狀會變成擁有複雜觸角(elaborated process)的放射狀形態。而在細胞培養過程中，當星狀膠質細胞受到刺激而進行分化時也可以觀察到星狀細胞的出現。然而，目前對於放射狀形態產生的分子機制尚未明瞭。已有文獻指出，Mps one binder 2 (Mob2)會和nuclear Ser/Thr kinase Dbf2-related protein (NDR1)共同作用來參與調控細胞週期，但Mob2在中樞神經系統中的研究甚少。本研究發現，星狀膠質細胞在沒有血清的培養環境中，由纖維母細胞狀轉變為有多觸角的細胞形態，並且細胞內Mob2的基因表現有顯著的上升。星狀膠質細胞以cAMP類似物-dbcAMP處理6小時後，觀察到Mob2基因表現量增加，並且在24小時的處理下，觀察到星狀膠質細胞轉變為放射狀形態。促胰液素/高血糖素/VIP成員之一PACAP，可以促進神經新生和星狀膠質細胞的分化。我們發現PACAP處理星狀膠質細胞6小時後，可以增加Mob2基因表現，並且此分子處理星狀膠質細胞24小時後，細胞形態轉變為放射狀形態。而進一步的結果發現，利用cAMP/PKA的抑制劑KT5720和CaMKII抑制劑KN93，可以抑制由dbcAMP所增加的Mob2基因表現。KT5720和細胞內鈣離子嵌合物BAPTA-AM，可以抑制由PACAP所增加的Mob2的基因表現。以上結果顯示cAMP/Ca2+ 會刺激星狀膠質細胞中Mob2的基因表現。進一步利用pcDNA-Mob2 (pMob2)及pEGFP-N2(N2)共同轉殖的方法，並將轉殖後的星狀膠質細胞培養在含有血清的培養液中，結果發現帶有GFP+的細胞形態為放射狀的型態。另一方面，利用siRNA-Mob2及N2共同轉殖的方式，在dbcAMP處理下星狀膠質細胞的觸角呈現萎縮的現象。以上的結果推測，Mob2參與調控星狀膠質細胞放射狀形態的產生。在轉殖pMob2/N2的細胞中，利用Texas Red-X phalloidin進行F-actin螢光染色，可以發現到會有短的絲狀足(filopodia)分散在延伸的觸角上。然而，利用Q-PCR分析發現，在dbcAMP的處理下並不會影響細胞中actin及GFAP的mRNA表現量。由以上的結果推測，中樞神經系統發育過程中，Mob2或許會在cAMP刺激之下，透過影響actin的重新排列而導致星狀膠質細胞轉變成為放射狀的形態。

Astrocytes which are a major population of CNS neural cells play an important role in neuronal homeostasis, neurogenesis, and synaptogenesis. During CNS development, astrocytes become stellate cells with elaborated processes. In culture, astrocytic stellation is observed when these cells undergo differentiation in response to stimuli. Nevertheless, the molecular mechanism for astrocytic stellation is poorly understood. Mps one binder (Mob) proteins that form stable complex with a nuclear Ser/Thr kinase Dbf2-related protein (NDR1), has been implicated to regulate cell cycle progression. Yet, little is known about their functional role in CNS neural cells. In this study, we found that Mob2 mRNA upregulation accompanied with astrocytic stellation after serum deprivation. Furthermore, the addition of the membrane permeable cAMP analogue dbcAMP not only caused an increase in Mob2 mRNA expression at 6 h, but also significantly induced astrocytic stellation at 24 h. Similarly, PACAP that is a member of the vasoactive intestinal peptide (VIP) /glucagon peptide family and stimulates neurogenesis and gliogenesis, induced an increase in Mob2 mRNA expression at 6 h, followed by astrocytic stellation at 24 h. The cAMP/PKA inhibitor KT5720 and the CaMKII inhibitor KN93 blocked dbcAMP-induced upregulation of Mob2 mRNA expression in astrocytes. KT5720 and intracellular calcium chelator BAPTA-AM also blocked PACAP-induced Mob2 mRNA upregulation. These results suggest that cAMP/Ca2+ signaling stimulates Mob2 transcription in astrocytes. Molecular approach using cotransfection of pcDNA-Mob2 (pMob2) with pEGFP-N2 (N2) in serum-treated astrocytes indicated that there was an evident stellate morphological change of GFP+ cells. In addition, interference of Mob2 expression using siRNA-Mob2 induced process withdrawal in dbcAMP-treated astrocytes. These results point to the role of Mob2 in the regulation of astrocytic stellation. In F-actin staining using Texas Red-X phalloidin, the short filopodia scattered in the elongated process of GFP+ astrocytes was observed after cotransfection of pMob2 with N2. However, Q-PCR analysis on actin and GFAP mRNA indicated that treatment with dbcAMP had no effect on GFAP and actin production. Based on our findings, we suggest that Mob2 may mediate astrocytic stellation by cAMP-dependent pathway through actin rearrangement in developing CNS.

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