Expression and function of the non-neuronal cholinergic system in murine embryonic stem cells

小鼠胚胎干细胞非神经元胆碱能系统的表达和功能

• 批准号: 164641675
• 负责人: Professor Charles James Kirkpatrick, Ph.D.
• 金额: --
• 依托单位: Institut für allgemeine Pathologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2010
• 资助国家: 德国
• 起止时间: 2009-12-31 至 2015-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/164641675?language=en
• 关键词: Expression; function; non; neuronal; cholinergic

The non-neuronal cholinergic system, which has been described at first by the applicants together with a research group from the States and Japan, is meanwhile accepted in the scientific community worldwide. Acetylcholine is synthesized by primitive organisms (bacteria, protozoa, plants) as well as by more or less all so called non-neuronal cells including the human organism (for example epithelial, endothelial, mesenchymal and immune cells). Non-neuronal acetylcholine is involved via auto- and paracrine mechanisms stimulating nicotinic and muscarinic receptors in the regulation of cellular proliferation and differentiation. Embryonic stem are characterized by their potency to differentiate into the somatic cell types indicated above. In the last 2 years the applicants have demonstrated the synthesis of acetylcholine and a remarkable high fractional release rate (2% per min) of acetylcholine in the murine stem cell line CGR8. The release is mediated by organic cation transporters in addition to so far unknown pathways. The synthesizing enzyme choline acetyltransferase is localized within the cytosol and the cell nucleus. Upon removal of leukemia inhibitory factor (LIF) CGR8 cells lose their pluripotency and show early differentiation. At this time the synthesis, cell content and release of acetylcholine is substantially up-regulated, which may indicate its functional role in early differentiation. In the present application the cellular mechanism of the upregulation will be investigated. Therefore, the interaction with intracellular signal transduction pathways known to interfer with the state of pluripotency will be examined (STAT3, PI3PK, MAPK, AC, PKA and PKC). Moreover it will be tested, whether comparable mechanisms are active in induced stem cells (iPS) and in embryonic stem cell of non-human primates.In context with iPS it has to be tested, whether the cholinergic system differs between the original differentiated somtic cell line and the induced pluripotent cell line and possible key signals modifying the cholinergic system should be identified. Moreover it will be tested whether the expression of nicotinic and muscarinic receptors will be changed during differentiation. In conclusion, the present "follow-application" should allow, to illuminate regulation processes of differentiation and de-differentiation within the cutting edge of pluripotency by investigating ther cholinergic system.

非神经元胆碱能系统首先由申请人与来自美国和日本的研究小组一起描述，同时被全世界科学界所接受。乙酰胆碱由原始生物体（细菌、原生动物、植物）以及或多或少所有所谓的非神经元细胞（包括人类有机体）（例如上皮细胞、内皮细胞、间充质细胞和免疫细胞）合成。非神经元乙酰胆碱通过自分泌和旁分泌机制刺激烟碱和毒蕈碱受体参与细胞增殖和分化的调节。胚胎干的特征在于其分化成上述体细胞类型的能力。 在过去的两年中，申请人已经证明了在小鼠干细胞系CGR8中乙酰胆碱的合成和乙酰胆碱显着高的部分释放率（每分钟2%）。除了迄今为止未知的途径之外，该释放还由有机阳离子转运蛋白介导。合成酶胆碱乙酰转移酶位于细胞质和细胞核内。去除白血病抑制因子 (LIF) 后，CGR8 细胞会失去多能性并显示出早期分化。此时乙酰胆碱的合成、细胞含量和释放显着上调，这可能表明其在早期分化中的功能作用。在本申请中，将研究上调的细胞机制。因此，将检查与已知干扰多能性状态的细胞内信号转导途径的相互作用（STAT3、PI3PK、MAPK、AC、PKA 和 PKC）。此外，还将测试类似的机制在诱导干细胞（iPS）和非人类灵长类动物的胚胎干细胞中是否活跃。在 iPS 的背景下，必须测试原始分化体细胞之间的胆碱能系统是否不同应鉴定细胞系和诱导多能细胞系以及可能改变胆碱能系统的关键信号。此外，将测试烟碱和毒蕈碱受体的表达在分化过程中是否会改变。总之，目前的“后续应用”应该允许通过研究胆碱能系统来阐明多能性前沿内分化和去分化的调节过程。