The Regional Specific Alterations in BBB Permeability are Relevant to the Differential Responses of 67-kDa LR Expression in Endothelial Cells and Astrocytes Following Status Epilepticus

Abstract

Status epilepticus (a prolonged seizure activity, SE) differently affects vasogenic edema formation and dystrophin-aquaporin 4 (AQP4) expressions between the rat hippocampus and the piriform cortex (PC). In the present study, we explored whether the 67-kDa laminin receptor (LR) expression was relevant to the regional specific susceptibility of vasogenic edema at 3 days after SE. In spite of no difference in expression levels of 67-kDa LR, dystrophin, and AQP4 under physiological conditions, SE-induced serum extravasation was more severe in the PC than the hippocampus. Western blots demonstrated that SE reduced expression levels of 67-kDa LR, dystrophin, and AQP4 in the PC, but not in the hippocampus proper. Immunofluorescent studies revealed that SE increased 67-kDa LR expression in reactive CA1 astrocyte, but reduced it in the PC and the molecular layer of the dentate gyrus due to massive astroglial loss. Furthermore, SE decreased expressions of endothelial 67-kDa LR and SMI-71 (endothelial brain barrier antigen) in these regions. The 67-kDa LR neutralization evoked serum extravasation in these regions of normal animals without astroglial loss. Similar to SE, 67-kDa LR neutralization also reduced dystrophin-AQP4 expressions in the PC more than the total hippocampus. Furthermore, 67-kDa LR IgG infusion increased phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2), but not c-Jun N-terminal kinase, independent of phosphoprotein enriched in astrocytes of 15 kDa (PEA15) activity. Co-treatment of U0126 (an ERK1/2 inhibitor) alleviated vasogenic edema formation and the reduced dystrophin-AQP4 expressions induced by 67-kDa LR neutralization. The 67-kDa LR IgG infusion also increased the susceptibility to SE induction. Therefore, our findings suggested that the cellular specific alterations in 67-kDa LR expression might be involved in the severity of SE-induced vasogenic edema formation in regional specific manners, which might affect the susceptibility to SE induction.

Blockade of 67-kDa Laminin Receptor Facilitates AQP4 Down-Regulation and BBB Disruption via ERK1/2-and p38 MAPK-Mediated PI3K/AKT Activations

“…Furthermore, 67LR neutralization activates p38 MAPK-mediated VEGF expression, which also increases vascular permeability [17]. Indeed, SB202190 alleviates serum extravasation induced by SE and 67LR IgG infusion, while it does not affect AQP4 expression [17,18]. Consistent with these reports, the present study reveals that 67LR neutralization and SE increased p38 MAPK-mediated PI3K/AKT phosphorylations.…”

“…Alzet 1007D osmotic pump contained (1) vehicle, (2) SB202190 (0.3 mg/mL), (3) wortmannin (0.1 nmol), (4) 3CAI (25 μM), and (5) U0126 (25 μM). In our previous studies [14,[16][17][18]30,31], each treatment did not show behavioral and neurological defects and could not change the seizure susceptibility and seizure severity in response to pilocarpine. Three days after surgery, this group was induced with SE by lithium chloride (LiCl)-pilocarpine.…”

“…Recently, we have reported that blockade of 67LR functionality is involved in serum extravasation through p38 MAPK-VEGF signaling pathway. Furthermore, 67LR neutralization decreases AQP4 expression and enhanced ERK1/2 phosphorylation, which are reversed by U0126 [17,18]. As AQP4 deletion or its inhibition worsens vasogenic edema [10][11][12], we have speculated that 67LR neutralization-induced ERK1/2 activation would down-regulate AQP4 expression.…”

“…Serum extravasation was measured, as previously described [14,17,18,31]. Aforementioned, sections were incubated in biotinylated rat IgG and ABC complex (Vector, #PK-6100, Burlingame, CA, USA, diluted 1:200).…”

“…Wortmannin, 3CAI, and U0126 increase the AQP4 expression level without affecting p38 MAPK phosphorylation. Since 67LR is identified in astrocytes and vascular endothelial cells [17,18,33,34], 67LR IgG infusion may likely affect p38 MAPK, AKT, and ERK1/2 phosphorylations in these cell populations. To identify the cellular profiles of the altered p38 MAPK, AKT, and ERK1/2 phosphorylations induced by 67LR neutralization, we performed the double immunofluorescent study.…”