Lab Experience at the Stroke and Vascular Dysfunctions Laboratory – Mario Negri Institute – Nicholas Giulio Raccagni

Laboratory: Stroke and Vascular Dysfunctions Laboratory
Principal Investigator: Dr. Stefano Fumagalli
Tutor: Dr. Aurora Bianchi
Duration: September 9, 2024 – October 4, 2024

The Stroke and Vascular Dysfunctions Laboratory at the Mario Negri Institute in Milan investigates the mechanisms underlying stroke and vascular-related neurological disorders, with a particular focus on neuroinflammation in ischemic brain injuries. Under the guidance of Dr. Fumagalli and Dr. Bianchi, I was engaged in the lab’s dynamic research environment, learning from two key projects.

The first project delved into the role of mannose-binding lectin (MBL) in ischemic brain injury. Recognized as a crucial initiator of the lectin complement pathway, MBL’s activation has been linked to exacerbating brain damage post-ischemia. Through this project, I gained insights into how MBL contributes to local microvascular thrombosis and inflammation, and the potential of pharmacological interventions to mitigate its harmful effects.

The second project centered on alpha-synuclein, a protein predominantly associated with neurodegenerative diseases such as Parkinson’s. Recent studies suggest that alpha-synuclein may also influence ischemic stroke pathophysiology. My involvement in this research allowed me to observe the expression and aggregation patterns of alpha-synuclein in post-ischemic brain tissues, enhancing my understanding of its impact on neuronal survival and recovery.

Throughout this rotation, I developed proficiency in several laboratory techniques essential for neuroscience research. I learned to operate a cryostat for preparing thin mouse brain tissue sections, facilitating detailed histological and immunofluorescence analyses in in-vivo models. Instead, culturing immortalized human brain microvascular endothelial cells (iHBMECs) and subjecting them to oxygen and glucose deprivation (OGD) conditions provided a hands-on understanding of in vitro ischemic models. This process encompassed routine tasks such as medium changes, cell passaging, and seeding cells in multi-well plates for subsequent assays.

Post-OGD, I assisted at RNA and protein extractions from cell samples, enabling quantitative real-time PCR (qRT-PCR) and Western blotting to assess gene and protein expression changes. Additionally, I employed spectrophotometry to measure RNA concentrations, ensuring the quality and accuracy of samples for molecular experiments.

This immersive experience not only enhanced my technical skills but also deepened my understanding of the complex interplay between vascular pathology and neurodegeneration. The mentorship of Dr. Fumagalli and Dr. Bianchi was invaluable, providing me with insights into experimental design, data interpretation, and the broader implications of our research findings.

I am grateful for this opportunity, which has solidified my interest in pursuing a career at the intersection of neurology and vascular biology.