Ferulic acid regulates the AKT/GSK-3β/CRMP-2 signaling pathway in a middle cerebral artery occlusion animal model

Abstract

Ferulic acid, a component of the plants Angelica sinensis (Oliv.) Diels and Ligusticum chuanxiong Hort, exerts a neuroprotective effect by regulating various signaling pathways. This study showed that ferulic acid treatment prevents the injury-induced increase of collapsin response mediator protein 2 (CRMP-2) in focal cerebral ischemia. Glycogen synthase kinase-3β (GSK-3β) regulates CRMP-2 function through phosphorylation of CRMP-2. Moreover, the pro-apoptotic activity of GSK-3β is inactivated by phosphorylation by Akt. This study investigated whether ferulic acid modulates the expression of CRMP-2 and its upstream targets, Akt and GSK-3β, in focal cerebral ischemia. Male rats were treated immediately with ferulic acid (100 mg/kg, i.v.) or vehicle after middle cerebral artery occlusion (MCAO), and then cerebral cortices were collected 24 hr after MCAO. MCAO resulted in decreased levels of phospho-Akt and phospho-GSK-3β, while ferulic acid treatment prevented the decrease in the levels of these proteins. Moreover, phospho-CRMP-2 and CRMP-2 levels increased during MCAO, whereas ferulic acid attenuated these injury-induced increases. These results demonstrate that ferulic acid regulates the Akt/GSK-3β/CRMP-2 signaling pathway in focal cerebral ischemic injury, thereby protecting against brain injury.

Keywords: Akt; CRMP-2; Ferulic acid; GSK-3β; neuroprotection.

Figure 1

Collapsin response mediator protein 2 (CRMP-2) protein spots identified by MALDI-TOF in the cerebral cortex from vehicle+middle cerebral artery occlusion (MCAO), ferulic acid+MCAO, vehicle+sham, ferulic acid+sham animals. Circles indicate the CRMP-2 protein spots. The intensity of spots was measured using PDQuest software. The ratio of intensity is described as spots intensity of these animals to spots intensity of sham+vehicle animals. Data (n=5) are shown as mean±SEM. ^*P<0.05. Mw and IP indicate molecular weight and isoelectrical point, respectively.

Figure 2

Western blot analysis of phospho-Akt and Akt in the cerebral cortex from vehicle+middle cerebral artery occlusion (MCAO), ferulic acid+MCAO, vehicle+sham, ferulic acid+sham animals. Each lane represents an individual experimental animal. Densitometric analysis is represented as these proteins intensity to β-actin intensity. Molecular weight (kDa) is depicted at right. Data (n=5) are represented as mean±SEM. ^*P<0.05.

Figure 3

Western blot analysis of phospho-GSK-3β and GSK-3β in the cerebral cortex from vehicle + middle cerebral artery occlusion (MCAO), ferulic acid+MCAO, vehicle+sham, ferulic acid+sham animals. Each lane represents an individual experimental animal. Densitometric analysis is represented as these proteins intensity to β-actin intensity. Molecular weight (kDa) is depicted at right. Data (n=5) are represented as mean±SEM. ^*P<0.05.

Figure 4

Western blot analysis of phospho-CRMP-2 and CRMP-2 in the cerebral cortex from vehicle+middle cerebral artery occlusion (MCAO), ferulic acid+MCAO, vehicle+sham, ferulic acid+sham animals. Each lane represents an individual experimental animal. Densitometric analysis is represented as these proteins intensity to β-actin intensity. Molecular weight (kDa) is depicted at right. Data (n=5) are represented as mean±SEM. ^*P<0.05.