PhD Scientific Days 2025

Poster Session I. - F: Pharmaceutical Sciences and Health Technologies

Microglia-Induced Neurodegeneration in SH-SY5Y Cells: An In Vitro Model of Oxidative Stress and Insulin Resistance in Alzheimer’s Disease

Name of the presenter

Barcza-Paszternák Alexandra

Institute/workplace of the presenter

Department of Pharmacodynamics, Semmelweis University

Authors

Alexandra Barcza-Paszternák¹, Kamilla Varga¹, Tamás Tábi¹

1: Department of Pharmacodynamics, Semmelweis University

Text of the abstract

Introduction:
Neuroinflammation plays a key role in the pathology of Alzheimer’s disease (AD), promoting neuronal dysfunction and degeneration. Although numerous studies have investigated the role of individual inflammatory mediators, there is a need for a focused yet biologically relevant in vitro model capable of replicating immune cell-induced neuronal damage.
Aims:
Our aim was to establish a simplified model of inflammation-induced neurodegeneration by treating retinoic acid-differentiated SH-SY5Y neuroblastoma cells with conditioned medium (CM) derived from LPS-activated BV2 microglial cells.
Methods:
To evaluate the impact of BV2-derived inflammatory mediators on neuronal function, SH-SY5Y cells were treated with conditioned medium from LPS-activated BV2 cells. Oxidative stress was measured using DCFDA and HE staining; mitochondrial function was assessed via JC-1 and MitoTracker dyes. Glucose uptake was quantified using the 2-NBDG assay, and insulin sensitivity was evaluated by determining IC₅₀ values from insulin dose–response curves. Autophagic activity was analyzed by Acridine Orange staining, insulin signaling was assessed by measuring phosphorylated GSK-3β levels using ELISA, and protein aggregation was detected with Thioflavin S staining.
Results:
After short-term exposure (3 h), BV2-conditioned medium significantly increased ROS and superoxide levels, accompanied by a marked reduction in mitochondrial membrane potential. Mitochondrial mass remained unchanged, while glucose uptake was significantly impaired. Notably, insulin treatment failed to restore glucose uptake, and the insulin IC₅₀ increased nearly sixfold (from 35 pM to 198 pM), indicating pronounced insulin resistance. After prolonged treatment (72 h), oxidative stress partially normalized, whereas insulin responsiveness continued to decline, confirming the chronic impact of microglia-derived inflammation on neuronal metabolism. Acridine Orange staining revealed reduced autophagic activity and accumulation of acidic vesicles, indicating autophagy inhibition. Phospho-GSK-3β levels decreased following BV2+LPS treatment and were restored only at higher insulin concentrations, while liraglutide treatment normalized phosphorylation, suggesting that inflammation-induced insulin resistance is reversible. Thioflavin S staining indicated the accumulation of protein aggregates.
Conclusion:
Our microglia-based in vitro model reproduces key Alzheimer’s-related features such as oxidative stress, mitochondrial dysfunction, and insulin resistance. It provides a simple, scalable tool to study inflammation-driven neuronal damage and to screen potential neuroprotective compounds.