Derivation and dopaminergic differentiation of human drug addict-specific iPS cel

人吸毒者特异性 iPS 细胞的衍生和多巴胺能分化

• 批准号: 8247948
• 负责人: Yu Luo
• 金额: $ 19.63万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8247948
• 关键词: Addictive Behavior; Adult; Alcohol or Other Drugs use; Alcohols; Biological Assay; Cell Line; Cell physiology; Cells; Characteristics; Complex; DNA; DNA Fingerprinting; Derivation procedure; Development; Discipline; Disease; Drug Addiction; Drug Controls; Drug user; Ethnography; Family; Fibroblasts; Functional disorder; Gene Expression; Genes; Genetic; Genetic Polymorphism; Genomics; Goals; Health; Human; Human Development; Illicit Drugs; Individual; Karyotype determination procedure; Lead; Legal; Link; Midbrain structure; Minisatellite Repeats; Mixed Function Oxygenases; Molecular; Molecular Target; Neurons; Pathway interactions; Patients; Pharmaceutical Preparations; Play; Pluripotent Stem Cells; Population; Protocols documentation; Research; Resources; Rewards; Role; Sampling; Skin; Societies; Somatic Cell; Source; Substance abuse problem; Substance of Abuse; System; Testing; Therapeutic; Tobacco; Validation; World Health Organization; addiction; adverse outcome; base; brain cell; calbindin; cell type; cohort; cost; dopamine transporter; dopaminergic differentiation; dopaminergic neuron; drug addict; drug of abuse; genetic variant; human disease; induced pluripotent stem cell; insight; monoamine; neurotransmission; novel; pluripotency; promoter; psychostimulant; relating to nervous system; response; skills; social; stem; stem cell technology; vesicular monoamine transporter

DESCRIPTION (provided by applicant): Addiction can be defined as a loss of control of substance use despite adverse consequences. Addiction to legal and illegal substances destroys the lives of both addicted subjects and their families, exerting an enormous cost and burden on society. The molecular- and cellular-based mechanisms that contribute to the initiation and development of addiction remain to be elucidated. Estimates have suggested that 40-60% of the vulnerability to addiction may be attributable to genetic aberrations. Multiple chromosomal regions have been linked to addiction including those containing the dopamine transporter (DAT) and vesicular monoamine transporter (VMAT2) genes. Current efforts to understand how polymorphisms in these monoamine transporters contribute to the molecular mechanisms of addiction are severely hindered by the inability to directly interrogate neural cell types from the patients. Patient-specific sources of cells carrying specific genetic variants that are capable of robust and reproducible differentiation into specific neural lineages do not exist. We propose to develop a cell-based system whereby neural cells from afflicted individuals can be functionally assayed to interrogate the molecular mechanisms underlying addiction. To achieve this goal we have developed a cutting-edge proposal that that incorporates the skill and expertise of multiple disciplines. In Aim 1 we will derive and characterize patient-specific, induced pluripotent stem (iPS) cells from addiction patients and controls that carry monoamine transporter polymorphisms. Since midbrain dopaminergic system play a prominent role in natural and drug related reward pathways and represent a common substrate for drugs of abuse, in Aim 2 we will differentiate patient-specific iPS cells line into dopaminergic neurons and carry out a detailed and functional characterization of these cells to identify their molecular characteristics (i.e. A9, A10, mesolimbic or mesocortical dopaminergic neurons). In Aim 3, we will characterize, compare, and functionally assay these patient-specific, iPS cell-derived dopaminergic neurons from control and addiction patients that carry polymorphisms for hDAT1 and hVMAT2 gene. There is great potential for patient-specific iPS cell technology to profoundly impact our understanding of human development and disease by providing genetically distinct, functional sources of human cells. By completing the aims set forth in this proposal we expect to provide a detailed characterization of dopaminergic neurotransmission function in patients afflicted with addiction and provide insight into the pathophysiology of this complex disease as well as the contribution of genetic variants in monoamine transporter genes to addiction. We have established an interdisciplinary team that combines strengths in ethnographic study of drug addicts, neural differentiation and dopaminergic function analysis, as well as pluripotency and iPS cells to interrogate novel questions about the cellular and molecular dysfunction that contributes to addiction. We expect that results from our studies will have immediate relevance to the understanding and treatment of this human disease. PUBLIC HEALTH RELEVANCE: Addiction can be defined as a loss of control of substance use despite adverse consequences. The World Health Organization estimates that there are 2 billion alcohol users, 1.3 billion tobacco users and 185 million illicit drug users worldwide. People who are addicted cannot control their need for the substance of abuse, even in the face of negative health, social or legal consequences. As a result, addiction to legal and illegal substances destroys the lives of both patients and their families, exerting tremendous cost and burden on the society. Unfortunately, the cause of addiction is currently unknown. The molecular- and cellular-based mechanisms that contribute to the initiation and development of addition remain to be elucidated. Results of our studies will provide a detailed characterization of brain cell function in patients afflicted with addiction and will offer insight into the mechanisms that contribute to this complex, devastating disease.

描述（由申请人提供）：成瘾可以定义为尽管有不良后果，但对物质使用失去控制。对法律的和非法物质的成瘾破坏了成瘾者及其家庭的生活，给社会带来了巨大的成本和负担。分子和细胞为基础的机制，有助于成瘾的启动和发展仍有待阐明。据估计，40-60%的成瘾可能归因于遗传畸变。多个染色体区域与成瘾有关，包括含有多巴胺转运蛋白（DAT）和囊泡单胺转运蛋白（VMAT 2）基因的区域。目前的努力，以了解这些单胺转运蛋白的多态性如何有助于成瘾的分子机制是严重阻碍了无法直接询问神经细胞类型的患者。不存在携带能够稳健且可再现地分化成特定神经谱系的特定遗传变体的细胞的患者特异性来源。我们建议开发一个基于细胞的系统，使神经细胞从受影响的个人可以进行功能分析，以询问成瘾的分子机制。为了实现这一目标，我们开发了一个尖端的提案，其中包含了多个学科的技能和专业知识。在目标1中，我们将从携带单胺转运蛋白多态性的成瘾患者和对照组中获得并表征患者特异性诱导多能干细胞（iPS）。由于中脑多巴胺能系统在天然和药物相关的奖赏途径中发挥重要作用，并且代表滥用药物的常见底物，在目的2中，我们将使患者特异性iPS细胞系分化为多巴胺能神经元，并对这些细胞进行详细的功能表征以鉴定其分子特征（即A9、A10、中脑边缘或中皮层多巴胺能神经元）。在目标3中，我们将表征，比较和功能分析这些患者特异性，iPS细胞衍生的多巴胺能神经元从控制和成瘾患者携带hDAT 1和hVMAT 2基因多态性。患者特异性iPS细胞技术具有巨大的潜力，通过提供遗传上不同的人类细胞功能来源，深刻影响我们对人类发育和疾病的理解。通过完成本提案中提出的目标，我们期望提供成瘾患者多巴胺能神经传递功能的详细表征，并深入了解这种复杂疾病的病理生理学以及单胺转运蛋白基因中遗传变异对成瘾的贡献。我们已经建立了一个跨学科的团队，结合了药物成瘾者的人种学研究，神经分化和多巴胺能功能分析，以及多能性和iPS细胞的优势，以询问有关成瘾的细胞和分子功能障碍的新问题。我们希望我们的研究结果将与理解和治疗这种人类疾病直接相关。 公共卫生相关性：成瘾可以定义为尽管有不良后果但对物质使用失去控制。世界卫生组织估计，全世界有20亿酗酒者、13亿吸烟者和1.85亿非法药物使用者。上瘾的人无法控制他们对滥用物质的需求，即使面对负面的健康、社会或法律的后果。结果，对法律的和非法物质的成瘾破坏了患者及其家庭的生活，给社会带来了巨大的成本和负担。不幸的是，成瘾的原因目前尚不清楚。分子和细胞为基础的机制，有助于启动和发展的此外仍有待阐明。我们的研究结果将提供成瘾患者脑细胞功能的详细特征，并将提供对导致这种复杂，毁灭性疾病的机制的深入了解。