Resveratrol Alleviates Cognitive Impairment in Chronic Cerebral Hypoperfusion by Targeting Lingo-1, NgR1, p75, and RhoA/ROCK-2 Pathways

Abstract

Background: Chronic cerebral hypoperfusion (CCH), a pathophysiological state linked to vascular dementia and cognitive impairment, involves the NgR1/Lingo-1/p75 signaling complex implicated in neurodegenerative processes. Resveratrol (RES), a neuroprotective compound, was investigated for its potential to mitigate CCH-induced cognitive deficits by targeting this pathway.

Objectives: This study examined RES's ability to improve cognitive impairment in CCH by suppressing the NgR1/Lingo-1/p75 complex and downstream RhoA-ROCK2 signaling.

Methods: Rats were divided into five groups: Control, CCH + Ethanol (ETH), CCH, CCH + resveratrol (RES), and RES. Chronic cerebral hypoperfusion was induced via permanent bilateral carotid artery occlusion (2VO). Cognitive function was assessed using the Morris Water Maze (MWM). Hippocampal morphology in CA1, CA3, and dentate gyrus (DG) regions was analyzed via H&E staining. The expression levels of Lingo-1, NgR1, P75, RhoA, and ROCK2 signaling pathway were detected by western blot and quantitative real-time PCR (qRT-PCR).

Results: Chronic cerebral hypoperfusion rats showed elevated protein expression of Lingo-1, p75, RhoA, and ROCK2, though NgR1 remained unchanged. The RES treatment significantly reduced these protein levels. Similarly, mRNA levels of all five targets increased in CCH, but RES notably lowered Lingo-1 and NgR1 expression. The MWM tests revealed RES improved spatial learning and memory deficits in 2VO rats. H&E staining demonstrated RES's neuroprotective effects, preserving hippocampal neuron integrity.

Conclusions: Resveratrol alleviates CCH-induced cognitive impairment by downregulating the Lingo-1/NgR1/p75 signaling axis and inhibiting RhoA-ROCK2 pathways. These findings highlight RES's potential as a therapeutic agent for vascular cognitive impairment associated with chronic hypoperfusion.

Keywords: Chronic Cerebral Hypoperfusion; Lingo-1; NgR1; Resveratrol; p75.

Figure 1.. Experimental design–chronic cerebral hypoperfusion (CCH) model was induced by bilateral carotid artery occlusion (2VO). Rats received resveratrol (RES; 5 mg/kg, i.p.) or ethanol daily for 35 days following surgery. Day 0 refers to the day of the surgery.

Figure 2.. The effects of resveratrol (RES) treatment on spatial learning and memory in rats following chronic cerebral hypoperfusion (CCH) in the Morris Water Maze (MWM) test – A, the distance traveled during the training days; B, the distance traveled in the target quadrant without a platform; C, the time spent during the training days; D, the time spent in the target quadrant without a platform; and E, representative pathways on the last day of training trials. Data are expressed as mean ± SEM (n = 12/group). Differences between groups were determined by ANOVA followed by Tukey's test; * P < 0.05, ** P < 0.01, and *** P < 0.001 vs. Control (Ctl); & P < 0.05 vs. CCH + Ethanol (CCH+ETH); # P < 0.05 and ## P < 0.01 vs. CCH.

Figure 3.. Lingo-1 expression – the protein (A) and mRNA (B) expression levels of Lingo-1 in the hippocampus were assessed in the following groups: Control (Ctl), chronic cerebral hypoperfusion (CCH), CCH + Ethanol (ETH), CCH + Resveratrol (RES) and RES. Results were normalized to β-actin expression. Data are expressed as mean ± SEM (n = 6); * P < 0.05 vs. Ctl, # P < 0.05 and ## P < 0.01 vs. CCH group.

Figure 4.. NgR1 expression – the protein (A) and mRNA (B) expression levels of NgR1 in the hippocampus were assessed in the following groups: Control (Ctl), chronic cerebral hypoperfusion (CCH), CCH + Ethanol (CCH + ETH), CCH+Resveratrol (CCH + RES) and resveratrol (RES). Results were normalized to β-actin expression. Data are expressed as mean ± SEM (n = 6); * P < 0.05 vs. Ctl, & P < 0.05 vs. CCH+ETH, # P < 0.05 and ## P < 0.01 vs. CCH group.

Figure 5.. P75 expression – the protein (A) and mRNA (B) expression levels of P75 in the hippocampus were assessed in the following groups: Control (Ctl), chronic cerebral hypoperfusion (CCH), CCH + ethanol (ETH), CCH + resveratrol (RES) and RES. Results were normalized to β-actin expression. Data are expressed as mean ± SEM (n = 6); * P < 0.05 and ** P < 0.01 vs. Ctl, # P < 0.05 and ## P < 0.01 vs. CCH group.

Figure 6.. RhoA expression – the protein (A) and mRNA (B) expression levels of RhoA in the hippocampus were assessed in the following groups: Control (Ctl), chronic cerebral hypoperfusion (CCH), CCH + ethanol (ETH), CCH + resveratrol (RES) and RES. Results were normalized to β-actin expression. Data are expressed as mean ± SEM (n = 6); * P < 0.05, ** P < 0.01 and *** P < 0.001 vs. Ctl, # P < 0.05, ## P < 0.01 and ### P < 0.001 vs. CCH.

Figure 7.. Rock2 expression – the protein (A) and mRNA (B) expression levels of ROCK2 in the hippocampus were assessed in the following groups: Control (Ctl), chronic cerebral hypoperfusion (CCH), CCH + ethanol (ETH), CCH + resveratrol (RES) and RES. Results were normalized to β-actin expression. Data are expressed as mean ± SEM (n = 6); * P < 0.05 and ** P < 0.01 vs. Ctl, # P < 0.05 and ## P < 0.01 vs. CCH.

Figure 8.. H&E staining in the CA1, CA3 and dentate gyrus (DG) of the rat hippocampus – neuronal death in the hippocampus was assessed in the following groups: Control (Ctl), chronic cerebral hypoperfusion (CCH), CCH + ethanol (ETH), CCH + resveratrol (RES) and RES groups. In the Ctl group, the cytoplasm is stained, and the nuclei are not fragmented in CA1, CA3 and DG. However, neurons are degenerated, and the nuclei are dense in CA1, CA3 and DG in the CCH and CCH + ETH groups. Resveratrol prevented CCH-induced neuronal death in the hippocampus. (Magnification 40X, 400X, scale bar = 200 µm or 50 µm).