Single-Cell Gene Expression Analysis: Implications for Neurodegenerative and Neuropsychiatric Disorders

Technical and experimental advances in microaspiration techniques, RNA amplification, quantitative real-time polymerase chain reaction (qPCR), and cDNA microarray analysis have led to an increase in the number of studies of single-cell gene expression. In particular, the central nervous system (CNS) is an ideal structure to apply single-cell gene expression paradigms. Unlike an organ that is composed of one principal cell type, the brain contains a constellation of neuronal and noneuronal populations of cells. A goal is to sample gene expression from similar cell types within a defined region without potential contamination by expression profiles of adjacent neuronal subpopulations and noneuronal cells. The unprecedented resolution afforded by single-cell RNA analysis in combination with cDNA microarrays and qPCR-based analyses allows for relative gene expression level comparisons across cell types under different experimental conditions and disease states. The ability to analyze single cells is an important distinction from global and regional assessments of mRNA expression and can be applied to optimally prepared tissues from animal models as well as postmortem human brain tissues. This focused review illustrates the potential power of single-cell gene expression studies within the CNS in relation to neurodegenerative and neuropsychiatric disorders such as Alzheimer's disease (AD) and schizophrenia, respectively.     

转录因子EB和自噬在神经退行性疾病发病机制中的作用

自噬广泛存在于真核细胞中,与机体生理和病理过程的发生发展密切联系.自噬主要参与长寿蛋白质的降解,以清除受损或多余的蛋白质和细胞器,是细胞自我降解的过程之一.自噬通常被分为三类:大自噬、分子伴侣介导的自噬和小自噬.自噬溶酶体途径(ALP)功能障碍导致蛋白质聚集,从而产生异常蛋白质和无效细胞器的积累,这些特征是阿尔茨海默病(Alzheimer disease,AD)、帕金森病(Parkinson disease,PD)和亨廷顿病等神经退行性疾病(Huntington disease,HD)的标志.自噬的过程受一系列复杂的信号分子的调控,其中一个主要调节因子是转录因子EB(TFEB),是转录因子MiT家族的成员之一.研究表明,TFEB可通过积极调节自噬体形成和自噬体-溶酶体融合参与自噬,此外它还通过溶酶体胞吐作用提高细胞内的清除作用.因此作为自噬溶酶体生物发生的主要调节因子,TFEB已被广泛证明激活后可以从病理方面改善这些疾病.我们回顾分析ALP和TFEB的调节及其对神经退行性疾病的影响,同时展望ALP和TFEB在疾病病理中的复杂作用及其治疗意义.     

MicroRNA，lncRNA与神经退行性疾病

综述了microRNA和lncRNA在一些神经退行性疾病病理生理中的作用机制．随着社会生产的发展,人类文明的进步,人口日益老年化,神经退行性疾病正在全球范围内流行,严重地危害着人类的健康．尽管长期的研究使人们对神经退行性疾病有了比较全面和深入的了解,但是其背后隐藏的发病机制仍然是个谜．人类基因组约98%的转录产物为非编码RNA(ncRNA),在生命活动中有着许多鲜为人知的广泛而多样性的生物功能．小分子RNA(microRNA)是研究得相对比较深入的一类小ncRNA,最近2～3年,长非编码RNA(lncRNA)受到人们的重视,已积累了一些相关研究成果．     

萝卜硫素的成药性及其在肿瘤和神经退行性疾病中的作用

萝卜硫素是一种来源于十字花科类蔬菜的天然活性物质,具有强大的抗氧化和抗癌能力。研究表明,萝卜硫素可通过调节Nrf2、NF-κB、HSF1-HSP信号通路和调控表观遗传,从而作用于II相解毒酶、HDAC和DNMT,影响癌症、神经退行性疾病等疾病的发生、发展。此外,由于萝卜硫素具有易吸收易代谢等特点,用纳米材料包裹萝卜硫素可提高其生物利用度和稳定性,更好发挥其疗效。近年来已有多项I/II期临床实验显示,萝卜硫素具有良好的成药性,这表明萝卜硫素在治疗癌症和神经退行性疾病方面有极高的药用潜力。本文主要针对萝卜硫素的药代动力学、作用靶点、安全性及其在肿瘤和神经退行性疾病中的研究进展进行综述,为萝卜硫素未来应用于肿瘤和神经退行性疾病的治疗提供参考。     

Protease Pathways in Peptide Neurotransmission and Neurodegenerative Diseases

1. Recent research demonstrates the critical importance of neuroproteases for the production of peptide neurotransmitters, and for the production of toxic peptides in major neurodegenerative diseases that include Alzheimer's (AD) and Huntington's diseases. This review describes the strategies utilized to identify the appropriate proteases responsible for producing active peptides for neurotransmission, with application of such approaches for defining protease mechanisms in neurodegenerative diseases.2. Integration of multidisciplinary approaches in neurobiology, biochemistry, chemistry, proteomics, molecular biology, and genetics has been utilized for neuroprotease studies. These investigations have identified secretory vesicle cathepsin L for the production of the enkephalin opioid peptide neurotransmitter and other neuropeptides. Furthermore, new results using these strategies have identified secretory vesicle cathepsin B for the production of β-amyloid (Aβ) in the major regulated secretory pathway that provides activity-dependent secretion of Aβ peptides, which accumulate in AD.3. CNS neuroproteases that participate in peptide neurotransmission and in neurodegenerative diseases represent new candidate drug targets that may be explored in future research for the development of novel therapeutic agents for neurological conditions.     

The Triggering Receptor Expressed on Myeloid Cells 2: A Molecular Link of Neuroinflammation and Neurodegenerative Diseases

The triggering receptor expressed on myeloid cells (TREM) 2 is a member of the immunoglobulin superfamily of receptors and mediates signaling in immune cells via engagement of its co-receptor DNAX-activating protein of 12 kDa (DAP12). Homozygous mutations in TREM2 or DAP12 cause Nasu-Hakola disease, which is characterized by bone abnormalities and dementia. Recently, a variant of TREM2 has also been associated with an increased risk for Alzheimer disease. The selective expression of TREM2 on immune cells and its association with different forms of dementia indicate a contribution of this receptor in common pathways of neurodegeneration.     

Caspase-3：治疗神经退行性疾病的新靶点

Caspase-3是caspases家族（一类天冬氨酸特异性酶切半胱氨酸蛋白酶）中的成员,是哺乳动物细胞凋亡的关键蛋白酶.随着研究的深入,发现caspase-3在神经退行性疾病的病理过程中起着很重要的角色.Caspase-3在这些疾病的病理过程中,不仅仅是起着凋亡的效应器作用,还能直接与老年性痴呆症、帕金森氏症、亨廷顿舞蹈病、脊椎小脑失调等疾病的致病蛋白质分子相互作用,参与致病机制.因此,caspase-3是治疗神经退行性疾病的新靶点,寻找caspase-3高效高选择性的抑制剂将为治疗神经退行性疾病提供新的途径.     

线粒体蛋白CHCHD10与神经退行性疾病

CHCHD10是核基因编码的线粒体蛋白，主要位于线粒体膜间隙，在维持线粒体结构的完整性和线粒体功能方面起关键作用。CHCHD10基因突变或功能缺失与额颞叶痴呆、肌萎缩侧索硬化症、帕金森病、阿尔茨海默病等多种神经退行性疾病的发生、发展密切相关。线粒体损伤是多种神经退行性疾病的一个共同特点，CHCHD10基因突变或功能缺失同样会导致线粒体结构和功能的异常。本文从CHCHD10结构及其线粒体功能角度总结近年来所发表的研究进展，讨论CHCHD10基因突变或功能缺失引起线粒体损伤的机制。研究CHCHD10在维持线粒体结构和功能中的作用机制，将有助于理解神经退行性疾病的致病机理，并为探究这些疾病的干预策略奠定基础。     

线粒体融合、分裂与神经变性疾病

线粒体是一种处于高度运动状态的频繁地进行融合与分裂的细胞器.在生理状态下,线粒体的融合与分裂处于一种平衡的状态,这种平衡受线粒体融合蛋白1/2（Mfn1/2）、视神经萎缩蛋白1（OPA1）和动力相关蛋白1（Drp1）的调节. Mfn1/2介导线粒体外膜的融合,而OPA1则参与线粒体内膜的融合,这些蛋白受泛素化和蛋白水解的调控. Drp1参与线粒体的分裂过程,受多种翻译后修饰的调节,如磷酸化、泛素化、SUMO化和S 硝基化.对于神经元来说,线粒体融合分裂的动态平衡对保证神经元末梢长距离运输和能量平均分布是非常重要的.因此,线粒体融合分裂异常可能是许多神经变性疾病的致病因素之一.对线粒体融合而言,Mfn2错义突变将导致遗传性运动感觉神经病2型（CMT2A）;OPA1错义突变将引起显性遗传性视神经萎缩(ADOA),而就线粒体分裂而言,Drp1突变与多系统功能障碍的新生儿致死性相关.     

Coupled Single-Cell CRISPR Screening and Epigenomic Profiling Reveals Causal Gene Regulatory Networks

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DNA Damage Profiling in Motor Neurons: A Single-Cell Analysis by Comet Assay

We developed a method to measure DNA damage in single motor neurons (MN). A cell fraction enriched in viable -motor neurons was isolated from adult rat spinal cord. This cell preparation was used to measure the vulnerability of the MN genome to different reactive oxygen species (ROS). MN were exposed in vitro to hydrogen peroxide, nitric oxide and peroxynitrite. Specific types of DNA lesions (e.g., abasic sites, single-strand breaks, and double-strand breaks) were measured using single-cell gel electrophoresis (comet assay). The MN genome was very susceptible to attack by ROS. Different ROS induced different DNA damage profiles in MN. MN were also isolated from adult rats with sciatic nerve avulsions to show that DNA damage emerges early during their degeneration in vivo. This study demonstrates that the comet assay is a feasible method for profiling DNA lesions in the genome of single MN. Viable mature MN can be isolated and used for in vitro models of MN genotoxicity and can be isolated from in vivo models of MN degeneration for profiling DNA damage on a single-cell basis.     

组织转谷氨酰胺酶与神经退行性疾病

组织转谷氨酰胺酶(tissue transglutaminase,tTG)广泛分布于各种组织及细胞中,是一个多功能蛋白质。tTG能催化Ca^2 依赖的蛋白质交联反应,并在多种生物学过程中起到了重要作用,如细胞生长与分化、受体介导的胞吞作用、细胞黏附、细胞形态的维持以及细胞凋亡等。已有研究表明,tTG可能在多种神经退行性疾病的病理生理过程中起到了重要作用。现就近年来有关tTG与神经退行性疾病研究的一些进展做一介绍。     

Molecular Mechanisms of Cellular Senescence in Neurodegenerative Diseases

Neurodegenerative diseases, such as Alzheimer’s and Parkinson’s, are characterized by several pathological features, including selective neuronal loss, aggregation of specific proteins, and chronic inflammation. Aging is the most critical risk factor of these disorders. However, the mechanism by which aging contributes to the pathogenesis of neurodegenerative diseases is not clearly understood. Cellular senescence is a cell state or fate in response to stimuli. It is typically associated with a series of changes in cellular phenotypes such as abnormal cellular metabolism and proteostasis, reactive oxygen species (ROS) production, and increased secretion of certain molecules via senescence-associated secretory phenotype (SASP). In this review, we discuss how cellular senescence contributes to brain aging and neurodegenerative diseases, and the relationship between protein aggregation and cellular senescence. Finally, we discuss the potential of senescence modifiers and senolytics in the treatment of neurodegenerative diseases.     

诱导多能干细胞及其在神经退行性疾病研究中的应用

用干细胞转录因子OCT4、SOX2、c-MYC和KLF4进行体细胞重编程产生具有胚胎干细胞特性的诱导多能干细胞(iPS细胞)是干细胞研究领域的突破性进展。近年来,iPS细胞的研究从产生方法、重编程机理及实际应用方面不断取得进展。由于iPS细胞的产生可取自体细胞,因而克服了胚胎干细胞应用的伦理学和免疫排斥等缺陷,为iPS细胞的临床应用开辟了广阔的前景。该文将对iPS细胞的产生方法、重编程机理及其在神经性退行性疾病的研究与应用进行文献综述,反映近几年iPS细胞最新研究成果,并阐述了用病人iPS细胞模型探讨帕金森氏病、老年性痴呆症、脊髓侧索硬化症、脊髓肌肉萎缩症及舞蹈症等5种常见神经性退行性疾病发病机理的研究现状。     

Subcellular Pathology of Human Neurodegenerative Disorders: Alzheimer-Type Dementia and Huntington''s Disease

Activities of enzyme markers of subcellular organelles have been measured in brain tissue from subjects with Alzheimer-type dementia (ATD) and Huntington's disease (HD). Significant increases in the activity of the lysosomal enzyme beta-glucuronidase were observed in both ATD temporal cortex and HD putamen. It is suggested that beta-glucuronidase activity may be a useful biochemical indicator of cellular damage in the CNS. A significant reduction in neutral alpha-glucosidase activity was observed in ATD temporal cortex and HD putamen. This change may reflect an alteration in glycoconjugate processing and may relate to the susceptibility of neurones to the degenerative processes of ATD and HD.