The Brain and Maternal Microchimerism

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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.