National Foundation for Liver Research, Cell Laboratory, Dr. Rela Institute and Medical Centre, Chennai 600 044, India.
Department of Biomedical Science, Bharathidasan University, Tiruchirappalli 620024, India.
ACS Chem Neurosci. 2021 Feb 17;12(4):704-718. doi: 10.1021/acschemneuro.0c00728. Epub 2021 Jan 29.
Hypoxia is considered a key factor in cellular differentiation and proliferation, particularly during embryonic development; the process of early neurogenesis also occurs under hypoxic conditions. Apart from these developmental processes, hypoxia preconditioning or mild hypoxic sensitization develops resistance against ischemic stroke in deteriorating tissues. We therefore hypothesized that neurons resulting from hypoxia-regulated neuronal differentiation could be the best choice for treating brain ischemia, which contributes to neurodegeneration. In this study, infrapatellar fat pad (IFP), an adipose tissue present beneath the knee joint, was used as the stem cell source. IFP-derived stem cells (IFPSCs) are totally adherent and are mesenchymal stem cells. The transdifferentiation protocol involved hypoxia preconditioning, the use of hypoxic-conditioned medium, and maintenance in maturation medium with α-lipoic acid. The differentiated cells were characterized using microscopy, reverse transcription PCR, real time PCR, and immunocytochemistry. To evaluate the epigenetic reprogramming of IFPSCs to become neuron-like cells, methylation microarrays were performed. Hypoxia preconditioning stabilized and allowed for the translocation of hypoxia inducible factor 1α into the nucleus and induced achaete-scute homologue 1 and doublecortin expression. Following induction, the resultant cells expressed neuronal markers neuron-specific enolase, neurofilament-light chain, growth associated protein 43, synaptosome associated protein 25, and β-III tubulin. The differentiated neural-lineage cells had functional gene expression pertaining to neurotransmitters, their release, and their receptors. The molecular signaling mechanisms regulated developmental neurogenesis. Furthermore, the physiological condition regulated neurotransmitter respecification or switching during IFPSC differentiation to neurons. Thus, differentiated neurons were fabricated against the ischemic region to treat neurodegenerative diseases.
缺氧被认为是细胞分化和增殖的关键因素，特别是在胚胎发育过程中；早期神经发生也在缺氧条件下发生。除了这些发育过程，缺氧预处理或轻度低氧致敏会在恶化组织中产生对缺血性中风的抗性。因此，我们假设缺氧调节的神经元分化产生的神经元可能是治疗脑缺血的最佳选择，这有助于神经退行性变。在这项研究中，髌下脂肪垫（IFP）被用作干细胞来源。IFP 衍生的干细胞（IFPSCs）是完全贴壁的间充质干细胞。转分化方案涉及缺氧预处理、使用缺氧条件培养基以及在含α-硫辛酸的成熟培养基中维持。使用显微镜、逆转录 PCR、实时 PCR 和免疫细胞化学对分化细胞进行了特征描述。为了评估 IFPSC 向神经元样细胞的表观遗传重编程，进行了甲基化微阵列分析。缺氧预处理稳定并允许缺氧诱导因子 1α易位到细胞核中，并诱导 achaete-scute 同源物 1 和双皮质素表达。诱导后，所得细胞表达神经元标志物神经元特异性烯醇化酶、神经丝轻链、生长相关蛋白 43、突触体相关蛋白 25 和 β-III 微管蛋白。分化的神经谱系细胞具有与神经递质及其释放和受体相关的功能性基因表达。分子信号机制调节发育神经发生。此外，生理条件调节 IFPSC 分化为神经元时神经递质的重新指定或转换。因此，针对缺血区域制造分化神经元以治疗神经退行性疾病。
