Department of Anatomy, Dongguk University Graduate School of Medicine, Gyeongju 38066, Republic of Korea.
Department of Anatomy, Dongguk University Graduate School of Medicine, Gyeongju 38066, Republic of Korea.
Phytomedicine. 2018 Jul 15;46:164-175. doi: 10.1016/j.phymed.2018.04.012. Epub 2018 Apr 6.
The hippocampus is a vulnerable brain region that is implicated in learning and memory impairment by two pathophysiological features, that is, neurite regression and synaptic dysfunction, and stigmasterol (ST), a cholesterol-equivalent phytosterol, is known to facilitate neuromodulatory effects.
To investigate the neuromodulatory effects of ST on the development of central nervous system neurons and the molecular bases of these effects in primary hippocampal neurons.
Rat embryonic (E18-19) brain neurons were cultured in the absence or presence of ST (75 µM). Neuritogenic activities of ST were evident by increases in various morphometric parameters. To identify underlying affected genes, total RNA was isolated on day in vitro 12 (DIV 12) and mRNA high throughput sequencing (mRNA-Seq) was performed. Affected key genes for neuronal development were identified using bioinformatics tools and their upregulations were confirmed by immunocytochemistry.
Among the differentially expressed 17,337 RefSeq genes, 445 genes (up/down 293/157) passed the p-value < 0.05 criterion, 52 genes (up/down; 37/13) had a p-value < 0.05 and a false discovery rate (FDR) q-value of < 0.2, and 24 genes (up/down; 20/4) passed the more stringent criterion of both p < 0.05 and q < 0.05. After applying a stringent FDR q-value cutoff of < 0.2, it was found ST induced many immediate early genes (IEGs), and that a major proportion of upregulated genes were related to central nervous system (CNS) development (neurite outgrowth or synaptic transmission). In a Venn diagram for CNS development Gene Ontologies (GOs) (i.e., axon development, dendrite development, modulation of synaptic transmission), Reln emerged as a central player in these processes, and highly interconnected 'hub' genes, including Dcx, Egr1, Ntrk2, and Slc24a2, were revealed by gene co-expression networks. Finally, transcriptomic data was confirmed by immunocytochemistry of primary hippocampal neurons.
The study indicates that ST upregulates genes for neuritogenesis and synaptogenesis, and suggests ST be viewed as a potential resource for improving brain functions.
海马体是一个易受损伤的大脑区域，它涉及学习和记忆障碍的两个病理生理特征，即神经突回缩和突触功能障碍，而豆甾醇（ST）是一种胆固醇等效植物甾醇，已知它具有促进神经调节作用。
研究 ST 对中枢神经系统神经元发育的神经调节作用及其在原代海马神经元中的作用机制。
在无 ST（75μM）或有 ST 的情况下培养大鼠胚胎期（E18-19）脑神经元。ST 的神经发生活性通过各种形态计量参数的增加来证明。为了鉴定潜在的受影响基因，在体外培养第 12 天（DIV12）分离总 RNA，并进行 mRNA 高通量测序（mRNA-Seq）。使用生物信息学工具鉴定影响神经元发育的关键基因，并通过免疫细胞化学确认其上调。
在差异表达的 17337 个 RefSeq 基因中，445 个基因（上调/下调 293/157）通过了 p 值<0.05 的标准，52 个基因（上调/下调；37/13）通过了 p 值<0.05 和假发现率（FDR）q 值<0.2 的标准，24 个基因（上调/下调；20/4）通过了更严格的 p<0.05 和 q<0.05 的标准。在应用严格的 FDR q 值截止值<0.2 后，发现 ST 诱导了许多即时早期基因（IEGs），并且上调基因的主要比例与中枢神经系统（CNS）发育（神经突生长或突触传递）有关。在与中枢神经系统（CNS）发育相关的基因本体论（GO）的 Venn 图（即轴突发育、树突发育、突触传递的调节）中，Reln 作为这些过程的核心参与者出现，并且通过基因共表达网络揭示了高度互联的“枢纽”基因，包括 Dcx、Egr1、Ntrk2 和 Slc24a2。最后，通过对原代海马神经元进行免疫细胞化学证实了转录组数据。
该研究表明 ST 上调了神经发生和突触发生的基因，并表明 ST 可被视为改善大脑功能的潜在资源。
