John Cintron

John Cintron

Incoming sophomore (Class of 2027) at Johns Hopkins University, Chemical & Biomolecular Engineering major. GPCR drug discovery project

Microglia, one of the most essential glial cells found abundantly in the brain, have essential homeostatic functions but are also known to enter neuroinflammatory states (DAMs) during disease. This program has been truly life-changing. The amazing BSRP staff and my excellent mentor all have contributed to my love for research. It’s allowed me to develop soft and technical skills that are beneficial in this modern world, and I feel that it’s helped me develop a path for my professional and personal future.A myeloid and microglia-specific G-protein coupled receptor (GPCR) of interest, has been reported to be necessary for the maintenance of normal homeostatic microglial function and the inhibition of proinflammatory response. The activation of this GPCR has therapeutic potential in promoting homeostatic function and inhibiting the proinflammatory activity of microglia, thereby reducing neuronal damage in neurodegenerative and neuroinflammatory disorders like Alzheimer’s Disease. By comparing mice with a knockout of our GPCR of interest and C57BL/6J (wild-type) littermates at 18 days of age utilizing immunofluorescence staining and fluorescence microscopy imaging, we were able to scan slides of brain tissue to evaluate the spatial localization of proteins of interest, relative to microglia. Machine learning image analysis was performed in a “double-blind” manner, based on genotype independent high and low extreme scenario training images (8wk old outbred mice, and 6 month old 5xFAD mice). Results have shown that knockout mice have significant increases in the expression of SPP1, APOC1, Annexin A5, particularly in non-microglia cells. During early postnatal development, these proteins are often found in association with neuronal support cells which typically undergo apoptosis prior to p18.  We then isolated regions of interest that showed elevated levels of Spp1, ApoC1, or Annexin A5 and found that cellular regions demonstrating these markers showed a lower association with microglia proximity and a general reduction of microglia coverage in knockout mice where as wild-type mice show an accumulation of microglia in contrast. Thus, we propose that our GPCR of interest in microglia is essential for the removal of neuronal support cells that arise as a result of normal brain development and that the knockout of this gene resulted in a buildup of these cells by day 18.

 

Project: Effects of Microglia-Associated GPCR Knockout on Brain Development in Mice

Mentor: Diana Bohannon, Stanley Center Therapeutics