The Brain and Maternal Microchimerism

大脑和母体微嵌合现象

• 批准号: 10216869
• 负责人: J. Lee Nelson
• 金额: $ 16.48万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10216869
• 关键词: Abbreviations; Address; Adult; Affect; Age; Alleles; Area; Autoimmune; Autopsy; Biological Assay; Biology; Blood; Brain; Brain Neoplasms; Cardiac Myocytes; Cell Nucleus; Cells; Characteristics; Child; Childhood; Clinical; Custom; DNA; Development; Disease; Epilepsy; Excision; Female; Fetus; Fluorescent in Situ Hybridization; Foundations; Gene Expression; Gene Expression Profile; Genetic Polymorphism; Genotype; Glioma; Grouping; HLA Antigens; Health; Heart; Hepatocyte; Human; Immune; Immunofluorescence Immunologic; Individual; Intractable Epilepsy; Islet Cell; Kidney; Knowledge; Liver; Methodology; Microchimerism; Mothers; Nuclear RNA; Operative Surgical Procedures; Organ; Pancreas; Patients; Persons; Pharmaceutical Preparations; Phenotype; Population; Pregnancy; Prevalence; Resected; Role; Sampling; Signal Transduction; Small Nuclear RNA; Specimen; Spleen; Swab; Testing; Tissue Sample; Tissues; Work; X Chromosome; Y Chromosome; brain tissue; cell type; experimental study; fetal; genotyped patients; male; neoplastic; nervous system disorder; offspring; pediatric patients; transcriptome; transcriptome sequencing

PROJECT SUMMARY/ABSTRACT Microchimerism (Mc) refers to harboring a small amount of cells or DNA from a genetically distinct individual. Many years after the physical union of mother and child ends maternal Mc (MMc) is found in her offspring, and Mc of fetal origin in the mother. MMc has been found in children and adults in blood and organs, including heart, liver, spleen, kidney and pancreas. In experimental and human studies Mc appears to be differentiated, creating for example MMc as cardiac myocytes in the heart, islet  cells in the pancreas and hepatocytes in the liver. Despite the importance of the brain to human health and function a fundamental gap of knowledge exists for MMc in the brain. The overall purpose of this proposal is to generate foundational knowledge about MMc cell types, quantities and the MMc transcriptome in human brain. To establish the origin of Mc as specific to the mother requires maternal participation which is generally not available for childhood autopsies. Aim 1 has two parts. Part 1 of Aim 1 will investigate MMc in pediatric patients who have surgical excision for medication refractory epilepsy, for whom mothers are available to participate. HLA and other polymorphism genotyping is done from maternal DNA extracted from a buccal swab sample. After genotyping patients and mothers, each mother-child pair is reviewed to identify a non-transmitted, non-shared polymorphism i.e. unique to the mother. A maternal-specific assay is next selected from a panel of HLA- and other polymorphism-specific quantitative PCR (qPCR) assays we have developed for this purpose. DNA extracted from excised brain tissue is then interrogated for MMc using the selected custom assay for each mother-child pair. A similar approach will be employed to study brain resected from age comparable patients undergoing surgery for gliomas for which maternal participation can be included. Part 2 of the Aim 1 approach will select brain tissues from males to study by fluorescence in situ hybridization (FISH) with X- and Y-chromosome specific probes; this aspect of the Aim 1 approach will permit including brain tissue from children without neurologic disease from autopsy from whom maternal participation is not required. Female cells with two X-chromosome signals, presumed maternal, will be counted with XY male cells enumerated in the same area. Immunofluorescence (IF) will be added to evaluate cell phenotypes. In Aim 2 we will conduct single nuclei RNA sequencing (snRNA-seq) analysis on brain tissue samples. The snRNA-seq studies will comprehensively evaluate what type of cells in the brain are derived from MMc and will assess the MMc transcriptome. The ways in which MMc may affect the brain are multiple and diverse to the potential benefit and/or detriment of a child. In addition to informativeness for epilepsy, if naturally acquired MMc is a basic aspect of biology as we hypothesize, the proposed work will have created a foundation from which diverse disorders of the human brain can be investigated including conditions that are developmental, autoimmune, degenerative, or neoplastic, underscoring significance.

项目摘要/摘要 微chimerismism（MC）是指遗传上不同个体的少量细胞或DNA。 在她的后代中发现母亲和儿童结束后的Mater MC（MMC）多年，并且 母亲的胎儿来源。在包括心脏在内的血液和器官的儿童和成人中发现了MMC 肝，脾脏，肾脏和胰腺。在实验和人类研究中，MC似乎是分化的，创造了 例如，MMC作为心脏的心肌细胞，胰腺中的胰岛细胞和肝脏中的肝细胞。 尽管大脑对人类健康和功能的重要性，但仍存在基本知识差距 大脑中的MMC。该提案的总体目的是生成有关MMC单元的基础知识 人脑中的类型，数量和MMC转录组。确定MC的起源是特定于 母亲需要孕产妇参与，这通常不适合儿童尸检。 AIM 1有两个 部分。 AIM 1的第1部分将调查在药物外科手术惊喜的儿科患者中的MMC 难治性癫痫，母亲可以参加。 HLA和其他多态性基因分型是 从从颊拭子样品中提取的母体DNA完成。在基因分型患者和母亲之后，每个 审查母子对，以识别一种非传递，非共享的多态性，即母亲独有的。 接下来是从HLA-和其他多态性特异性定量的小组中选择的特定于母亲的测定 我们为此目的开发的PCR（QPCR）测定法。从极好的脑组织中提取的DNA就是 使用每个母子对的选定自定义测定法对MMC进行询问。类似的方法将是 用于研究从年龄可比患者接受手术的神经胶质瘤的大脑 可以包括孕产妇的参与。 AIM 1方法的第2部分将从男性中选择脑组织 通过荧光原位杂交（FISH）与X-和Y-Cromosoms特异性问题；目标的这一方面1 方法将允许包括来自没有神经疾病的儿童的脑组织 不需要孕产妇参与。具有两个X染色体信号的女性细胞，假定的Mater将是 用在同一区域列举的XY雄性细胞计数。免疫荧光（如果）将添加以评估 细胞表型。在AIM 2中，我们将对脑组织进行单核RNA测序（SNRNA-SEQ）分析 样品。 SNRNA-Seq研究将全面评估大脑中哪种类型的细胞。 MMC并将评估MMC转录组。 MMC可能影响大脑的方式是多重的，并且 潜水员对儿童的潜在利益和/或损害。除了癫痫的信息性之外 获得的MMC是我们假设的生物学的基本方面，拟议的工作将创造一个基础 可以从其中研究人脑的发散疾病，包括发育状况， 自身免疫性，退化性或肿瘤，强调意义。