Inhibition of prostaglandin E₂ receptor EP3 mitigates thrombin-induced brain injury

Abstract

Prostaglandin E 2 EP3 receptor is the only prostaglandin E 2 receptor that couples to multiple G-proteins, but its role in thrombin-induced brain injury is unclear. In the present study, we exposed mouse hippocampal slice cultures to thrombin in vitro and injected mice with intrastriatal thrombin in vivo to investigate the role of EP3 receptor in thrombin-induced brain injury and explore its underlying cellular and molecular mechanisms. In vitro, EP3 receptor inhibition reduced thrombin-induced hippocampal CA1 cell death. In vivo, EP3 receptor was expressed in astrocytes and microglia in the perilesional region. EP3 receptor inhibition reduced lesion volume, neurologic deficit, cell death, matrix metalloproteinase-9 activity, neutrophil infiltration, and the number of CD68 þ microglia, but increased the number of Ym-1 þ M2 microglia. RhoA-Rho kinase levels were increased after thrombin injection and were decreased by EP3 receptor inhibition. In mice that received an intrastriatal injection of autologous arterial blood, inhibition of thrombin activity with hirudin decreased RhoA expression compared with that in vehicle-treated mice. However, EP3 receptor activation reversed this effect of hirudin. These findings show that prostaglandin E 2 EP3 receptor contributes to thrombin-induced brain damage via Rho-Rho kinase-mediated cytotoxicity and proinflammatory responses.

Cerebroprotection of flavanol (-)-epicatechin after traumatic brain injury via Nrf2-dependent and -independent pathways

“…An accelerating rotarod was used to measure motor function and balance [36]. The rotarod treadmill (Accuscan, Inc., Columbus, OH, USA) provided a motor balance and coordination assessment.…”

“…Brains were removed and stored in 4% paraformaldehyde overnight and then transferred to 30% sucrose in PBS for another 48 h. Frozen serial coronal brain sections were sliced at 30 μm on a cryostat. Sections were used for Luxol fast blue staining (day 28, n=5/group), Cresyl violet staining (days 3 and 28, n = 8/group), Fluoro-Jade B (FJB) staining (day 3, n=6/group), and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL; day 3, n=6/group) [32, 36]. Mice assigned to propidium iodide (PI) staining (day 3, n=6/group) and hydroethidine analysis (day 3, n=6/group) were perfused with PBS only.…”

Cerebroprotection by salvianolic acid B after experimental subarachnoid hemorrhage occurs via Nrf2- and SIRT1-dependent pathways

“…In brief, brain sections were stained with H&E or Cresyl violet solution according to standard procedures [33,44,48–50], and then mounted onto microscope slides with Permount (Thermo Fisher Scientific). Staining was visualized under a light microscope.…”

“…Immunofluorescence staining was performed according to our previously published protocols [44,50]. Briefly, brain sections (6 μm) or cultured cells were incubated with primary antibodies against 8-hydroxyguanosine (8-OHdG, Abcam), Nrf2 (1:100, Abcam), SIRT1 (1:50, Santa Cruz), caspase-3 (1:100, Cell Signaling), and NeuN (1:200, cat# MAB377, EMD Millipore, USA) overnight at 4 °C.…”

Cerebroprotection by the neuronal PGE2 receptor EP2 after intracerebral hemorrhage in middle-aged mice

“…We and others have shown that EP1 and EP3 receptor signaling has toxic effects in in vitro and in vivo models of ICH 6,7 and in thrombin-induced brain injury. 8 In contrast, EP2 receptor signaling protects cultured neurons under various conditions through the cAMP/PKA or cAMP/Epac pathway [9][10][11][12] and regulates microglial activation and function in vitro. [13][14][15][16] These findings reflect cell-specific differences in EP2 signaling.…”