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.

项目总结/摘要 微嵌合体（Microchimerism，Mc）是指从遗传上不同的个体中携带少量细胞或DNA。 在母亲和孩子的身体结合结束多年后，母亲的Mc（MMc）在她的后代中被发现， 母体中胎儿起源的MC。在儿童和成人的血液和器官中发现了MMc，包括心脏， 肝脏脾脏肾脏和胰腺在实验和人体研究中，Mc似乎是分化的， 例如MMc，如心脏中的心肌细胞、胰腺中的胰岛β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， 大脑中的MMc本提案的总体目的是生成有关MMc细胞的基础知识 类型、数量和人脑中的MMc转录组。为了确定Mc的来源， 母亲需要母亲的参与，这通常不适用于儿童尸检。目标1有两个 零件.目标1的第1部分将研究因药物治疗而接受手术切除的儿科患者中的MMc 难治性癫痫，母亲可以参加。HLA和其他多态性基因分型是 从母亲的口腔拭子样本中提取DNA。在对患者和母亲进行基因分型后，每个人 检查母子对以鉴定非传播的、非共享的多态性，即母亲独有的多态性。 接下来，从HLA和其他多态性特异性定量分析组中选择母体特异性分析。 我们为此开发了PCR（qPCR）检测。然后从切除的脑组织中提取DNA， 使用针对每个母子对选择的定制测定询问MMc。类似的方法将是 用于研究从接受神经胶质瘤手术的年龄相当的患者中切除的大脑， 可以包括产妇参与。Aim 1方法的第2部分将选择男性的脑组织进行研究 通过X和Y染色体特异性探针的荧光原位杂交（FISH）; Aim 1的这一方面 这种方法将允许包括来自尸检的没有神经系统疾病的儿童的脑组织， 不需要产妇参与。具有两个X染色体信号的女性细胞，假定为母体细胞，将被 用相同区域中计数的XY雄性细胞计数。将增加免疫荧光（IF）以评价 细胞表型在目标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的细胞来源于哪种类型 MMc并将评估MMc转录组。MMc可能影响大脑的方式是多种多样的， 对儿童的潜在利益和/或损害。除了癫痫的信息量，如果自然 正如我们假设的那样，获得性MMc是生物学的一个基本方面，所提出的工作将为我们创造一个基础。 从中可以研究人类大脑的各种疾病，包括发育性疾病， 自身免疫性、退行性或肿瘤性，强调了重要性。