[Construction and mechanisms of D-galactose-induced blood-brain barrier organoid aging model].

OBJECTIVETo establish a D-galactose-induced aging model of the blood-brain barrier(BBB) with organoid technology and explore the pathophysiological mechanisms of BBB aging.

METHODSA BBB model was constructed on an organ-on-a-chip using human cerebral microvascular endothelial cells(hCMEC/D3), followed by D-galactose exposure to induce aging. The model was evaluated by measuring transendothelial electrical resistance(TEER), apparent permeability coefficient(Papp) of fluorescein, β-galactosidase staining, live/dead cell staining, lactate dehydrogenase(LDH) activity, and reactive oxygen species(ROS) levels. The expression of tight junction proteins(ZO-1, Occludin), senescence-associated gene(p53), and anti-inflammatory genes(HO-1, Nrf2) was assessed using RT-qPCR to characterize aging features.

RESULTSThe optimal concentration for inducing BBB organoid aging was 50 mg/mL D-galactose. The model group significantly reduced TEER((19.46±0.98)Ω·cm~2 vs. (22.07±0.21)Ω·cm~2, P<0.05), increased Papp of fluorescein((1.08±0.02)×10~(-4 )cm/s vs. (4.39±0.01)×10~(-5 )cm/s, P<0.05), elevated β-galactosidase-positive cells, decreased live cells, increased dead cells, and increased LDH release and ROS levels. RT-qPCR analysis showed downregulation of ZO-1 and Occludin, along with upregulation of p53, HO-1, and Nrf2.

CONCLUSIOND-galactose induced aging-associated damage in the BBB organoid model, providing a reliable in vitro biomimetic platform for studying BBB aging mechanisms and potential therapeutic interventions for neurodegenerative diseases. This model contributes to the understanding of the pathophysiological processes underlying neurological disorders.