Functional deficits in immortalized microglial cell lines limit their utility for modeling neuroinflammation

Microglia, the resident immune cells of the central nervous system (CNS), play a pivotal role in brain development and disease. Although primary microglia serve as the gold standard for in vitro investigations, their limited proliferative capacity has driven the widespread use of immortalized cell lines, with the BV-2 cell line being the most prominent. However, accumulating evidence demonstrates that BV-2 cells exhibit profound functional discrepancies relative to primary microglia.

This study aimed to generate optimized lentivirus-mediated human telomerase reverse transcriptase (hTERT)-immortalized microglial cell lines, and to compare their functional similarity to primary microglia and the BV-2 cells.

We established two murine microglial cell lines (CM-A1, CM-A3) via lentiviral hTERT transduction. Cellular characteristics including CD11b expression, cell morphology, phagocytic capacity, and transcriptional responses of inflammatory genes to lipopolysaccharide (LPS) were comprehensively evaluated.

CM-A1 and CM-A3 exhibited higher surface CD11b expression and basal phagocytic capacity relative to BV-2 cells. Nevertheless, all immortalized cell lines presented prominent functional defects, including reduced surface CD11b abundance, impaired morphological transformation following LPS stimulation, loss of LPS-triggered phagocytic upregulation, and suppressed induction of pro-inflammatory genes.

These findings reveal that both hTERT-immortalized microglial lines and BV-2 cells fail to recapitulate the critical dynamic functional responses observed in primary microglia, highlighting the importance of rigorous model selection in neuroimmunological research.