Department of Anatomical Sciences and Molecular Biology, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran.
Department of Genetic, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran.
J Mol Neurosci. 2019 Dec;69(4):597-607. doi: 10.1007/s12031-019-01388-8. Epub 2019 Jul 30.
Retinal degenerative diseases lead to blindness due to poorly regenerative potential of the retina. Recently, cell therapy is more considered for degenerative diseases. Autologous mesenchymal stem cells derived from adipose tissue are a suitable source for this purpose. Therefore, we conducted a stepwise efficient method to differentiate human adipose-derived stem cells (hADSCs) into retinal precursor-like cells in vitro. We compared two differentiation protocols, monolayer and hanging drop cultures. Through the defined medium and 3D hanging drop culture method, we could achieve up to 75% retinal precursor gene expression profile (PAX6, RAX, CHX10, and CRX) from hADSCs. By imitation of in vivo development, for direct conversion of stem cells into retinal cells, the suppression of the BMP, Nodal, and Wnt signaling pathways was carried out by using three small molecules. The hADSCs were primarily differentiated into anterior neuroectodermal cells by expression of OTX2, SIX3, and Β-TUB III and then the differentiated cells were propelled into the retinal cells. According to our data from real-time PCR, RT-PCR, immunocytochemistry, and functional assay, it seems that the hanging drop method improved retinal precursor differentiation yield which these precursor-like cells respond to glutamate neurotransmitter. Regarding the easy accessibility and immunosuppressive properties of hADSCs and more efficient hanging drop method, this study may be useful for future autologous cell therapy of retinal degenerative disorders.
视网膜退行性疾病由于视网膜再生潜力差而导致失明。最近，细胞疗法更多地被用于退行性疾病。源自脂肪组织的自体间充质干细胞是这种方法的合适来源。因此，我们采用逐步有效的方法将人脂肪源性干细胞（hADSCs）在体外分化为视网膜前体细胞样细胞。我们比较了两种分化方案，即单层和悬滴培养。通过定义的培养基和 3D 悬滴培养方法，我们可以从 hADSCs 中达到高达 75%的视网膜前体细胞基因表达谱（PAX6、RAX、CHX10 和 CRX）。通过模仿体内发育，为了将干细胞直接转化为视网膜细胞，通过使用三种小分子抑制 BMP、Nodal 和 Wnt 信号通路。hADSCs 通过表达 OTX2、SIX3 和 Β-TUB III 初步分化为前神经外胚层细胞，然后将分化的细胞推向视网膜细胞。根据我们的实时 PCR、RT-PCR、免疫细胞化学和功能测定数据，悬滴法似乎提高了视网膜前体细胞的分化产量，这些前体细胞样细胞对谷氨酸神经递质有反应。鉴于 hADSCs 易于获得且具有免疫抑制特性，以及更有效的悬滴法，这项研究可能对未来视网膜退行性疾病的自体细胞治疗有用。
