Description

Background: Microglia are the resident innate immune cells of the central nervous system (CNS) and play an integral role in maintaining brain homeostasis and in the progression of many neurological disorders. Some of the roles of microglia include synapse maintenance, phagocytosis of apoptotic cells and protein aggregates, and surveillance of the CNS. In diseased microglia, these roles can become dysregulated, leading to a multitude of neurodegenerative diseases. In diseases like Alzheimer’s Disease, microglia fail to take up and clear amyloid-β (Aβ) and tau, resulting in plaque formation. In multiple sclerosis, microglia are hyperactive and demyelination of the subcortical and cortical brain matter occur. One disease where microglia are thought to be the main pathogenic cell type is in a rare pediatric histiocytic disorder called Langerhans Cell Histiocytosis (LCH). LCH is caused by somatic MAPK pathway mutations in myeloid progenitor cells. Patients with LCH often develop a progressive neurodegenerative disorder. Since microglia are derived from yolk sac myeloid progenitors, we hypothesize that microglia with a MAPK mutation are driving the neurodegeneration seen in patients with LCH. The goal for my part of the project is to use techniques developed in the Coufal Lab to understand the role of microglia in LCH and broadly their dysregulation in neurodegenerative disorders. Methods: Using patient-reprogrammed iPSC lines, I learned how iPSCs are developed into iHPCs, and then differentiated into mature microglia. Several microglial morphology assays were learned; ramification quantification of microglia to observe what morphological phenotypes are present, and phagocytosis assays being some. Taking iPSCs that were previously reprogrammed from a patient with LCH, I will CRISPR in the LCH mutation, subclone them, run an Alu1 restriction digest, and send in my samples for sequencing to see if I got the mutation of interest. Upon getting the mutation of interest, I will then start the microglia maturation process and run numerous assays on them. Results/Discussion: I learned of various techniques used when modeling pediatric neurodegenerative disease such as LCH. Such rare neuroinflammatory pediatric diseases can be reprogrammed to iPSCs to make microglia and cerebral organoids to understand their diseases better and to test therapies.