Tanshione IIA reduces astrocyte swelling after oxygen-glucose deprivation and reoxygenation by inhibition of the HMGB1/RAGE/NF-κB/IL-6 pro-inflammatory axis and AQP4 over-clustering in the plasma membrane

Abstract Background: Tanshione IIA (TSO IIA) has emerged as a powerful anti-inflammatory compound with multiple therapeutic properties. Here, we investigated the role of TSO IIA in astrocytic swelling caused by ischemia and reperfusion-like injury in an in vitro model and the molecular mechanisms underlying this effect. Methods: Primary brain astrocytes were cultured in vitro in conditions of glucose and oxygen deprivation and reoxygenation (OGD/R). We investigated the effects of TSO IIA treatment on cell swelling and injury, and the expression levels of AQP4 protein in the plasma membrane. We then studied the involvement of the HMGB1/RAGE/NF-κB/IL-6 pro-inflammatory axis in TSO IIA-mediated protection. Results: Treatment with TSO IIA alleviated OGD/R-induced astrocytic swelling, changes in cell morphology and ultrastructure, the release of LDH, and the over-clustering of AQP4 protein in the plasma membrane. In addition, TSO-IIA significantly reduced over-expression of HMGB1 protein in the cytoplasm and surrounding medium, the high levels of NF-κB protein in the nucleus, and of IL-6 protein in cytoplasm and extracellular media induced by OGD/R. Combination of TSO IIA and recombinant HMGB1 (rHMGB1) reversed these effects. Inhibition of RAGE, the receptor of HMGB1, via administration of FPS-ZM1 (a RAGE antagonist) induced similar results to those of TSO IIA. The combination of HMGB1 and FPS-ZM1 did not alter the effects mediated by FPS-ZM1 alone. In addition, exogenous IL-6 reversed TSO IIA-mediated effect on AQP4 protein clustering in the plasma membrane, cell swelling and LDH activity.Conclusions: TSO IIA significantly reduced astrocyte swelling after OGD/R injury in vitro, via blocking activation of the HMGB1/RAGE/NF-κB/IL-6 pro-inflammatory axis and thereby decreasing the expression of AQP4 in plasma membrane.