Reduced Apoptotic Injury by Phenothiazine in Ischemic Stroke through the NOX-Akt/PKC Pathway

Abstract

Phenothiazine treatment has been shown to reduce post-stroke ischemic injury, though the underlying mechanism remains unclear. This study sought to confirm the neuroprotective effects of phenothiazines and to explore the role of the NOX (nicotinamide adenine dinucleotide phosphate oxidase)/Akt/PKC (protein kinase C) pathway in cerebral apoptosis. Sprague-Dawley rats underwent middle cerebral artery occlusion (MCAO) for 2 h and were randomly divided into 3 different cohorts: (1) saline, (2) 8 mg/kg chlorpromazine and promethazine (C+P), and (3) 8 mg/kg C+P as well as apocynin (NOX inhibitor). Brain infarct volumes were examined, and cell death/NOX activity was determined by assays. Western blotting was used to assess protein expression of kinase C-δ (PKC-δ), phosphorylated Akt (p-Akt), Bax, Bcl-XL, and uncleaved/cleaved caspase-3. Both C+P and C+P/NOX inhibitor administration yielded a significant reduction in infarct volumes and cell death, while the C+P/NOX inhibitor did not confer further reduction. In both treatment groups, anti-apoptotic Bcl-XL protein expression generally increased, while pro-apoptotic Bax and caspase-3 proteins generally decreased. PKC protein expression was decreased in both treatment groups, demonstrating a further decrease by C+P/NOX inhibitor at 6 and 24 h of reperfusion. The present study confirms C+P-mediated neuroprotection and suggests that the NOX/Akt/PKC pathway is a potential target for efficacious therapy following ischemic stroke.

Rapid Intervention of Chlorpromazine and Promethazine for Hibernation-Like Effect in Stroke: Rationale, Design, and Protocol for a Prospective Randomized Controlled Trial

“…Our studies in rat ischemic stroke models demonstrated a neuroprotective effect of C+P. This protection was due to the reduction in the reactive oxygen species (ROS)-mediated oxidative injury ( 15 ), reduction in brain inflammation determined by the expression of TNF-α, IL-1 β, ICAM-1, VCAM-1, NF-κB ( 16 ), and reduction in apoptotic signal cascade ( 17 ), as well as improved integrity of blood-brain barrier ( 18 ). Importantly, the neuroprotection was induced in both hypothermic or normothermic conditions after ischemic stroke.…”

“…In addition, our previous studies have demonstrated a neuroprotective effect of C+P in rat ischemic stroke models ( 15 ). The neuroprotective effect was due to the reduction in reactive oxygen species (ROS)-mediated oxidative injury ( 15 ), brain inflammation ( 16 ), apoptosis ( 17 ), and reduction in blood-brain barrier (BBB) dysfunction after ischemic stroke ( 18 ). Importantly, this neuroprotection was not completely dependent upon the reduction of body temperature.…”

Melatonin attenuates reactive astrogliosis and glial scar formation following cerebral ischemia and reperfusion injury mediated by GSK‐3β and RIP1K

“…Previous studies have demonstrated that apoptosis plays an important role in the pathogenesis of cerebral I/R and leads to cerebral infarction (Sun et al, 2015; Tong et al, 2019; Uzdensky, 2019). Our results demonstrated that melatonin reduced the infarct volume and the number of TUNEL‐positive cells in the ischemic brain.…”

Phenothiazine Inhibits Neuroinflammation and Inflammasome Activation Independent of Hypothermia After Ischemic Stroke

“…Chlorpromazine, a phenothiazine, was shown to suppress the production of proin ammatory cytokines including TNF-α, IL-1β, and IL-6 [31][32][33]. Moreover, the neuroprotective role of C + P in ischemic stroke seemed to be dependent on its amelioration of hyperglycolysis [8], blood-brain barrier disruption [28], in ammation [11], in ammasome activation [12], and apoptosis [34]. C + P can induce hypothermia, but its neuroprotective effect post-ischemic stroke was partially independent of its hypothermic effect [6,8,11,12].…”