CNS stem and progenitor cell differentiation into functional neuronal circuits in three-dimensional collagen gels
CNS stem and progenitor cell differentiation into functional neuronal circuits in three-dimensional collagen gels
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DOI：
10.1016/j.expneurol.2003.10.016
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发表时间：
2004-12-01
影响因子：
5.3
通讯作者：
Barker, JL
中科院分区：
文献类型：
--
作者：
Ma, W;Fitzgerald, W;Barker, JL
哺乳动物中枢神经系统（CNS）损伤后自我修复的能力很低，神经元不能增殖。因此，结合神经干细胞和祖细胞以及生物衍生的聚合物支架的神经组织工程可能会彻底改变治疗受损CNS组织的医学方法。神经干细胞和神经祖细胞从胚胎大鼠皮质或皮质下神经上皮细胞内分散的I型胶原蛋白，和细胞胶原蛋白的结构培养在无血清培养基中含有碱性成纤维细胞生长因子。胶原包埋的干细胞和祖细胞活跃地扩增并有效地产生神经元，其产生神经元极性、神经递质、离子通道/受体和兴奋性。Ca ~（2+）显像显示BrdU（+）/TuJ_1（-）细胞分化为BrdU（-）/TuJ_1（+）细胞时，伴随着神经递质功能受体表达的变化，从胆碱能和嘌呤能转变为主要的GABA能和谷氨酸能。自发性突触后电流记录从前体细胞衍生的神经元膜片钳和这些电流部分阻断10 μ M荷包牡丹碱，并完全阻断另外10 μ M的红藻氨酸受体拮抗剂CNQX，表明GABA能和谷氨酸能突触活动的外观。用内吞标记物FM 1 -43染色证明在胶原包埋的神经元中发生活性突触囊泡再循环。这些结果表明，在3D胶原凝胶中培养的神经干细胞和祖细胞重现了CNS干细胞的发育;这是CNS干细胞和祖细胞衍生的功能性突触和神经元网络在3D基质中形成的首次证明。神经祖细胞在3D胶原凝胶中的增殖能力和神经元分化潜力表明其在体内促进神经元再生的尝试中的潜在用途。(C)2004爱思唯尔公司All rights reserved.
The mammalian central nervous system (CNS) has little capacity for self-repair after injury, and neurons are not capable of proliferating. Therefore, neural tissue engineering that combines neural stem and progenitor cells and biologically derived polymer scaffolds may revolutionize the medical approach to the treatment of damaged CNS tissues. Neural stem and progenitor cells isolated from embryonic rat cortical or subcortical neuroepithelium were dispersed within type I collagen, and the cell-collagen constructs were cultured in serum-free medium containing basic fibroblast growth factor. The collagen-entrapped stem and progenitors actively expanded and efficiently generated neurons, which developed neuronal polarity, neurotransmitters, ion channels/receptors, and excitability. Ca2+ imaging showed that differentiation from BrdU(+)/TuJ1(-) to BrdU(-)/TuJ1(+) cells was accompanied by a shift in expression of functional receptors for neurotransmitters from cholinergic and purinergic to predominantly GABAergic and glutamatergic. Spontaneous postsynaptic currents were recorded by patch-clamping from precursor cell-derived neurons and these currents were partially blocked by 10-muM bicuculline, and completely blocked by additional 10 muM of the kainate receptor antagonist CNQX, indicating an appearance of both GABAergic and glutamatergic synaptic activities. Staining with endocytotic marker FM1-43 demonstrated active synaptic vesicle recycling occurring among collagen-entrapped neurons. These results show that neural stem and progenitor cells cultured in 3D collagen gels recapitulate CNS stem cell development; this is the first demonstration of CNS stem and progenitor cell-derived functional synapse and neuronal network formation in a 3D matrix. The proliferative capacity and neuronal differentiating potential of neural progenitors in 3D collagen gels suggest their potential use in attempts to promote neuronal regeneration in vivo. (C) 2004 Elsevier Inc. All rights reserved.