Ishizuka Toshiaki, Goshima Hazuki, Ozawa Ayako, Watanabe Yasuhiro
Department of Pharmacology, National Defense Medical College, Tokorozawa, Saitama, Japan.
Clin Exp Pharmacol Physiol. 2014 May;41(5):345-50. doi: 10.1111/1440-1681.12224.
Activation of serotonin (5-hydroxytryptamine; 5-HT) receptors plays a role in adult neurogenesis and differentiation of neural progenitor cells (NPC). Herein, we examined the involvement of 5-HT receptors in the differentiation of mouse induced pluripotent stem (iPS) cells into NPC. To induce embryoid body (EB) formation, mouse iPS cells were cultured on ultralow-attachment dishes. All-trans retinoic acid (ATRA; 1 μmol/L) and/or 5-HT (0.03 or 0.1 μmol/L) was added to the EB cultures for 4 days and then EB plated on gelatin-coated plates were cultured for 7 or 14 days. Immunofluorescence staining revealed that mouse iPS cells expressed both 5-HT2A and 5-HT4 receptors and, to a lesser extent, 5-HT1A receptors. Treatment with 5-HT significantly enhanced the ATRA-induced expression of nestin, a specific marker for NPC, and phosphorylation of cAMP response element-binding protein (CREB). Pretreatment of EB cultures with either 1 μmol/L GR113808 (a selective 5-HT4 receptor antagonist) or 1 μmol/L H89 (a protein kinase (PKA) inhibitor) significantly inhibited these effects of 5-HT. These findings suggest that stimulation of 5-HT4 receptors may enhance ATRA-induced neural differentiation of mouse iPS cells through activation of PKA and CREB.
血清素（5-羟色胺；5-HT）受体的激活在成体神经发生和神经祖细胞（NPC）的分化中起作用。在此，我们研究了5-HT受体在小鼠诱导多能干细胞（iPS）向NPC分化过程中的作用。为诱导胚状体（EB）形成，将小鼠iPS细胞接种于超低吸附培养皿中培养。向EB培养物中加入全反式维甲酸（ATRA；1 μmol/L）和/或5-HT（0.03或0.1 μmol/L），培养4天，然后将接种于明胶包被培养皿上的EB再培养7或14天。免疫荧光染色显示，小鼠iPS细胞表达5-HT2A和5-HT4受体，5-HT1A受体表达较少。5-HT处理显著增强了ATRA诱导的NPC特异性标志物巢蛋白的表达以及环磷酸腺苷反应元件结合蛋白（CREB）的磷酸化。用1 μmol/L GR113808（一种选择性5-HT4受体拮抗剂）或1 μmol/L H89（一种蛋白激酶（PKA）抑制剂）预处理EB培养物可显著抑制5-HT的这些作用。这些发现表明，刺激5-HT4受体可能通过激活PKA和CREB增强ATRA诱导的小鼠iPS细胞的神经分化。
