Combined extrinsic and intrinsic manipulations exert complementary neuronal enrichment in embryonic rat neural precursor cultures: an in vitro and in vivo analysis.
Combined extrinsic and intrinsic manipulations exert complementary neuronal enrichment in embryonic rat neural precursor cultures: an in vitro and in vivo analysis.
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DOI：
10.1002/cne.22027
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发表时间：
2009-07-01
期刊：
影响因子：
--
通讯作者：
Sagen J
中科院分区：
文献类型：
--
作者：
Furmanski O;Gajavelli S;Lee JW;Collado ME;Jergova S;Sagen J
许多CNS病症在γ-氨基丁酸抑制性信号传导的失调中具有共同的病理学。在去抑制部位移植GABA释放细胞有望缓解疾病症状，且副作用比传统药物疗法少。我们操纵FGF-2剥夺和MASH 1转录因子水平，试图扩增培养的大鼠胚胎神经前体细胞（NPC）的默认GABA能神经元命运，用于移植研究。幼稚和MASH 1慢病毒转导的NPC在FGF-2中维持或剥夺FGF-2不同的时间长度。免疫染色和定量分析表明，GABA-和β-III-微管蛋白免疫反应细胞通常通过连续传代减少，表明在体外扩增的大鼠神经球中神经原性潜能的丧失。然而，FGF-2剥夺导致来自传代的神经球的GABA能细胞的数量小，但显著增加。与幼稚和GFP慢病毒转导的克隆相反，MASH 1转导导致BrdU掺入和克隆集落大小增加。Western blotting显示MASH 1过表达和FGF-2剥夺可增加β-III-微管蛋白表达和降低CNY表达，而FGF-2剥夺单独减弱GFAP表达。这些结果表明，低FGF-2信号传导和MASH 1活性可以协同作用以富集NPC培养物的GABA神经元表型。当移植到成年大鼠脊髓中时，这种组合也产生了GABA能神经元。这些发现表明，即使成功利用默认的GABA能神经元前体命运，外源性和内源性操作的组合也可能是实现NSC移植物恢复功能的全部潜力所必需的。
Numerous CNS disorders share a common pathology in dysregulation of γ-amino butyric acid inhibitory signaling. Transplantation of GABA-releasing cells at the site of disinhibition holds promise for alleviating disease symptoms with fewer side effects than traditional drug therapies. We manipulated FGF-2 deprivation and MASH1 transcription factor levels in an attempt to amplify the default GABAergic neuronal fate in cultured rat embryonic neural precursor cells (NPCs) for use in transplantation studies. Naïve and MASH1 lentivirus-transduced NPCs were maintained in FGF-2 or deprived of FGF-2 for varying lengths of time. Immunostaining and quantitative analysis showed that GABA- and β-III-tubulin-immunoreactive cells generally decreased through successive passages, suggesting a loss of neurogenic potential in rat neurospheres expanded in vitro. However, FGF-2 deprivation resulted in a small, but significantly increased population of GABAergic cells derived from passaged neurospheres. In contrast to naïve and GFP lentivirus-transduced clones, MASH1 transduction resulted in increased BrdU incorporation and clonal colony size. Western blotting showed that MASH1 overexpression and FGF-2 deprivation additively increased β-III-tubulin and decreased CNPase expression, while FGF-2 deprivation alone attenuated GFAP expression. These results suggest that low FGF-2 signaling and MASH1 activity can operate in concert to enrich NPC cultures for a GABA neuronal phenotype. When transplanted into the adult rat spinal cord, this combination also yielded GABAergic neurons. These findings indicate that, even for successful utilization of the default GABAergic neuronal precursor fate, a combination of both extrinsic and intrinsic manipulations will likely be necessary to realize the full potential of NSC grafts in restoring function.
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影响因子：
5.3
作者：
Gernert, M;Thompson, KW;Tobin, AJ
通讯作者：
Tobin, AJ
影响因子：
4.4
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作者：
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