dbcAMP 誘導神經前驅細胞分化中 鈣訊號和 Akt 去磷酸化之影響

Involvement of calcium signaling and dephosphorylation of Akt in dbcAMP-induced neuronal differentiation of neural progenitor cells

林哲志(Jhe-Jhih Lin)

指導教授 ： 金亭佑

中原大學/工學院/生物科技研究所/碩士(2009年)

https://doi.org/10.6840/CYCU.2009.00858

摘要

目前來說，對於調控神經幹/前驅細胞分化為神經細胞的胞內訊號因子了解有限。利用初級培養的老鼠神經前驅細胞為模組，我們發現 dbcAMP 具有誘導神經分化作用的特性，因此藉此去了解其中分子機轉。在老鼠神經前驅細胞中，dbcAMP 可誘導胞內鈣離子的濃度快速上升。且於型態上，也伴隨著神經突長以及神經特有細胞骨架蛋白 Tuj1 和 MAP2 的表現。前處理鈣離子螯合劑 BAPTA/AM，明顯的阻斷 dbcAMP 所誘導的神經分化。除此之外，在 dbcAMP 加藥處理後，發現 Akt 有早期去磷酸化反應。藉由抑制劑 BAPTA/AM、FK506 等去探索 dbcAMP 誘導 Akt 去磷酸化訊號路徑，結果發現，鈣離子內流對於 Akt 早期去磷酸化現象是必要的，然而其去磷酸化作用並非經由 calcineurin 催化。更進一步，前處理 Akt-PI3K 抑制劑 LY294002，並無法直接造成神經突長現象。綜合以上證據顯示，胞內鈣離子濃度上升和早期 Akt 去磷酸化反應可能與神經分化早期相關。

關鍵字

神經前驅細胞 ； 鈣離子 ； dbcAMP ； Akt

並列摘要

The intracellular signaling events controlling neural stem/progenitor cells differentiation into neurons are poorly understood. Using primary culture of rat neural progenitor cells (rNPCs) as the model system, we characterized dbcAMP-induced neuronal differentiation which lead me to understand the underlying mechanisms. In rNPCs, dbcAMP induced a rapid increase of the cytosolic Ca2+ concentration. On the morphology, it also accompanied neurite outgrowth and expression of neuronal marker Tuj1 and MAP2. Pretreatment of NPCs with BAPTA/AM, a permeable intracellular calcium chelator, significantly blocked dbcAMP-induced neuronal differentiation. Besides, early dephosphorylation of Akt was observed after dbcAMP treatment. To explore the signal pathway of dbcAMP-induced Akt dephosphorylation, we pretreat rNPCs with inhibitors, such as BAPTA/AM, calcineurin inhibitor FK-506 and so on. These data showed that Ca2+ influx is essential for early dephosphorylation of Akt, but it doesn’t catalysis by calcineurin. Moreover, pretreatment of NPCs with LY294002, an Akt-PI3K inhibitor, can’t direct induced neurite outgrowth. Taken together, these results provide initial evidence that intracellular calcium elevation and Akt dephosphorylation could potentially influence early aspects of neuronal differentiation, such as neuronal fate specification.

並列關鍵字

neural progenitor cells ； calcium ； Akt ； dbcAMP

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