Total flavones of Rhododendron protect against BBB injury following cerebral ischemia/reperfusion via promoting astrocytic polarization towards A2 subtype.

This study investigates the role and underlying mechanisms of total flavones of Rhododendron (TFR) in blood-brain barrier (BBB) disruption following transient bilateral common carotid artery occlusion-induced cerebral ischemia/reperfusion (I/R), focusing on whether A2 astrocyte polarization mediates the protective effects of TFR. RNA sequencing analysis was performed to elucidate the molecular mechanisms underlying the differential effects of A1 and A2 astrocytes on endothelial function. TFR treatment significantly reduced Evans blue (EB) extravasation and attenuated the downregulation of claudin-5, ZO-1, occludin, and CD31 in mouse hippocampal tissues. Furthermore, TFR enhanced the production and release of cystathionine-β-synthase (CBS)-derived H2S from astrocytes. Both TFR and TFR-induced A2 astrocytes promoted the expression of tight junction proteins in endothelial cells (ECs) following oxygen-glucose deprivation/reoxygenation (OGD/R). RNA sequencing revealed upregulation of Caspase-11 in A1 astrocytes and downregulation in A2 astrocytes. Moreover, A1 astrocytes exhibited increased expression of matrix metalloproteinase (MMP)-9 and TNF-α, whereas A2 astrocytes showed elevated levels of IL-10. Collectively, these findings indicate that TFR mitigates BBB disruption after cerebral I/R through dual mechanisms: directly upregulating tight junction proteins in brain microvascular endothelial cells and indirectly promoting astrocyte polarization toward the A2 subtype, which is associated with reduced Caspase-11 expression, enhanced IL-10 secretion, and increased CBS-derived H2S production.