The role of connexin43 in hemorrhagic transformation after thrombolysis in vivo and in vitro.

Thrombolysis with recombinant tissue plasminogen activator (rtPA) is the most effective drug treatment for acute ischemic stroke within 4.5h after symptom onset. However, the use of rtPA may increase the risk of hemorrhagic transformation (HT), particularly when it is administered after the first 4.5h. However, no effective treatments are available to reduce the HT risk. Disruption of the blood-brain barrier (BBB) is central to the genesis of HT. Connexin43 (Cx43)-mediated gap junction intercellular communication (GJIC) has been demonstrated to regulate the integrity of the BBB in ischemia. We investigated the effect of Cx43 on BBB permeability during rtPA-induced HT. Spontaneously hypertensive rats (SHRs) underwent a 1.5-h middle cerebral artery occlusion and were treated with rtPA at 4.5h. The rats were sacrificed at 24h, and their brains were evaluated for BBB permeability and the expression of tight junction (TJ) proteins and Cx43. We examined whether the effects were Cx43 dependent using multiple Cx43 inhibitors. Phosphorylated Cx43 (p-Cx43) but not total Cx43 protein expression was increased after rtPA treatment. Delayed rtPA administration induced significant HT and BBB disruption. These effects were attenuated by inhibitors that blocked GJIC and Cx43 phosphorylation and expression but not Cx43 redistribution. Additionally, rtPA administration upregulated p-Cx43 expression in hypoxia/reoxygenation (H/R)-exposed brain endothelial cells. These effects were suppressed by the phosphatidylinositol 3'-kinase (PI3K) inhibitor LY294002 and the extracellular signal-regulated kinase 1/2 (ERK1/2) inhibitor U0126. We suggest that rtPA-associated hemorrhage due to an alteration in the integrity of the BBB is highly associated with an increase in p-Cx43 resulting from the activation of the PI3K and ERK pathways.