Elsevier

European Journal of Pharmacology

Volume 724, 5 February 2014, Pages 43-50
European Journal of Pharmacology

Neuropharmacology and analgesia
Candesartan and glycyrrhizin ameliorate ischemic brain damage through downregulation of the TLR signaling cascade

https://doi.org/10.1016/j.ejphar.2013.12.032Get rights and content

Abstract

Stroke is the second leading cause of death in industrialized countries and the most frequent cause of permanent disability in adults worldwide. The final outcome of stroke is determined not only by the volume of the ischemic core, but also by the extent of secondary brain damage inflicted to penumbral tissues by brain swelling, impaired microcirculation, and inflammation. The only drug approved for the treatment ischemic stroke is recombinant tissue plasminogen activator (rt-PA). The current study was designed to investigate the protective effects of candesartan (0.15 mg/kg, orally) and glycyrrhizin (30 mg/kg, orally) experimentally-induced ischemic brain damage in C57BL/6 mice (middle cerebral artery occlusion, MCAO) in comparison to the effects of a standard neuroprotective drug (cerebrolysin, 7.5 mg/kg, IP). All drugs were administered 30 min before and 24 h after MCAO. Both candesartan and glycyrrhizin ameliorated the deleterious effects of MCAO as indicated by the improvement in the performance of the animals in behaviour tests, reduction in brain infarction, neuronal degeneration, and leukocyte infiltration. In addition, MCAO induced a significant upregulation in the different elements of the TLR pathway including TLR-2 and TLR-4, Myd88, TRIF and IRF-3 and the downstream effectors TNF-α, IL-1β, IL-6 and NF-kB. All these changes were significantly ameliorated by treatment with candesartan and glycyrrhizin. The results of the current study represent a new indication for both candesartan and glycyrrhizin in the management of ischemic stroke with effects comparable to those of the standard neuroprotective drug cerebrolysin.

Introduction

Stroke is the second leading cause of death in industrialized countries and the most frequent cause of permanent disability in adults worldwide. Despite advances in understanding the pathophysiology of cerebral ischemia, therapeutic options for acute ischemic stroke remain very limited (Woodruff et al., 2011).

Toll like receptors (TLRs) are one group of pattern recognition receptors (PRRs) that detect damage-associated molecular pattern molecules (DAMPs) released from dead/dying cells (Carty and Bowie, 2011, Takeda and Akira, 2005). In the brain, TLRs are expressed in microglia, astrocytes, oligodendrocytes and neurons (Ahmad et al., 2013). Activation of microglial cells in response to cerebral ischemia is associated with a significant induction of TLR-2 and TLR-4, which are considered to mediate the inflammatory reaction involved in the pathophysiological processes of ischemic injury (Kong and Le, 2011).

Several lines of clinical and experimental evidence suggest an important role of the brain rennin-angiotensin (RAS) in ischemic brain injury (Li et al., 2005). Endogenous levels of angiotensin II (Ang II) (Mecca et al., 2009) and angiotensin II type-1 (AT1) receptor (Edvinsson, 2008) are increased following stroke and angiotensin receptor blockers (ARBs) have been shown to attenuate the deleterious effects in animal stroke models (Mecca et al., 2009).

Angiotensin II, acting via AT1 receptor, promotes vasoconstriction, oxidative stress, inflammation and atherosclerosis (Dasu et al., 2009). Angiotensin II also induces an increase in reactive oxygen spices and a proinflammatory phenotype via AT1 receptor (Dasu et al., 2009).

Stimulation of AT2 receptors promotes cell differentiation and regeneration in neuronal tissue (Li et al., 2005). Also angiotensin II, acting through the AT2 receptor, represents a neurotrophic factor for neurons in the central nervous system (Li et al., 2005).

Glycyrrhizin (GL) has diverse biological activities, including anti-inflammatory, anti-viral, anti-liver cancer, antioxidative, immunomodulatory, cardioprotective and hepatoprotective activities (Kim et al., 2012, Schrofelbauer et al., 2009). GL attenuated ischemia-induced learning and memory deficits and protected against cell damage in the brain (Hou et al., 2010).

The current study was designed to investigate the protective effects of candesartan (ARB at a sub-hypotensive dose) and glycyrrhizin (a natural compound with anti-inflammatory activity) against experimentally-induced ischemic brain damage (middle cerebral artery occlusion, MCAO) in C57BL/6 mice as indicated by alterations in behaviour, infarct volume, neuronal degeneration and leukocyte infiltration. Also the current investigation explored the effects of ischemia and the tested compounds on the expression/activity of the different components on the toll-like receptor (TLR) pathway including TLR-2 and TLR-4, MYD88, TRIF and IRF-3 and the downstream effectors TNF-α, IL-1β, IL-6 and NF-κB. The effects of the tested compounds were compared to the effects of the standard neuroprotective drug; Cerebrolysin.

Section snippets

Animals

C57BL/6 mice (18–24 g) purchased from Theodor Bilharzias Center, Cairo, Egypt were used in all experiments. The mice were kept under standard environmental and nutritional conditions throughout the investigation. This study was performed in accordance with the guidelines of the Ethical Committee for animal handling at Zagazig University (ECAHZU). All experiments, surgery, neurological scores and infarct volume determination were performed in a randomized manner.

Mice were randomly distributed

Results

Daily administration of candesartan (0.15 mg/kg) or glycyrrhizin (30 mg/kg) did not cause any significant change in blood pressure (data not shown).

Fig. 1 shows the change in behaviour induced by the different treatments. Treatment with candesartan, glycyrrhizin and cerebrolysin, caused significant reduction in neurological scores compared with ischemic group reaching 2.2, 1.5 and 2.2 vs 3.2, respectively (Fig. 1A).

The chimney test pointed out early deficits of motor coordination in MCAO mice.

Discussion

Few neurological conditions are as complex and devastating as stroke (Moskowitz et al., 2010). Currently, thrombolysis with tissue plasminogen activator (t-PA) is the only effective therapy, but due to its narrow therapeutic window and safety concern; fewer than 5% of stroke patients receive this treatment (Woodruff et al., 2011). It is crucial to expand the narrow therapeutic opportunities for this devastating condition.

Cerebral ischemia evokes a strong inflammatory response characterized by

Conclusion

In conclusion, the present study provides a new evidence for a protective role of candesartan and GL against ischemic brain damage which was mediated through downregulation of the TLR signaling pathway with subsequent reduction in brain inflammation. The activities of both drugs were comparable to that of a commonly used neuroprotective agent (CL) which might indicate a beneficial co treatment. In the current study, candesartan and GL were used 30 min before and 24 h after MCAO and it is expected

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