Elsevier

Journal of Ethnopharmacology

Volume 197, 2 February 2017, Pages 147-156
Journal of Ethnopharmacology

Sameerpannag Ras Mixture (SRM) improved neurobehavioral deficits following acute ischemic stroke by attenuating neuroinflammatory response

https://doi.org/10.1016/j.jep.2016.07.059Get rights and content

Abstract

Ethnopharmacological relevance

Cerebral ischemic stroke is one of the leading causes of death and long-term disability worldwide. Unfortunately, due to the failure of most of drugs in clinical trials recently, attentions have moved towards the traditional system of medicines including Ayurveda. In Ayurveda, Sameerpannag Ras (SR) is a mineral and metallic origin based formulation which has been used for the treatment of arthritis and chronic systematic inflammatory disorder. The current study was designed to investigate the neuroprotective effects of Sameerpannag Ras Mixture (SRM), on the neurobehavioral dysfunction and associated neuroinflammation, induced by transient Internal Carotid Artery Occlusion (ICAO) in mice model.

Materials and methods

In the present study, mice were treated with Sameerpannag Ras Mixture (SRM) at the dose of 40 mg/kg body weight by oral gavages for 3 and 7days respectively, twice a day, after the induction of ICAO for 90 min followed by reperfusion. The efficacy of SRM was examined by scoring neurological behavioral deficit using the standard neurological deficit score (NDS), grip strength and rotarod performance tests at different time intervals of post-ICAO.

Results

Post-ischemic treatment with Sameerpannag Ras Mixture (SRM) at 40 mg/kg significantly reduced Neurological Deficit Score and improved the motor coordination at different time intervals post-ICAO. The analysis of RT-qPCR data showed that transient cerebral ischemia could induce the inflammatory response genes in the affected striatal region of ICAO group, as compared to sham group, on day3 and day7 post-ICAO. Interestingly, SRM treatment showed marked improvement in the ischemia-induced neurobehavioral deficits by attenuating ischemia-induced neuroinflammatory response at both gene and protein level.

Conclusion

The present study suggests that Sameerpannag Ras Mixture (SRM) treatment ameliorates behavioral outcomes after mild ischemia through the suppression of a number of inflammatory response genes involved in neuronal damage. This is the first report of the molecular mechanism underlying the significant neuroprotective action of the Ayurvedic drug, Sameerpannag Ras Mixture (SRM), using a mild stroke in mice model.

Introduction

The most common cause of cerebral stroke is sudden occlusion of blood vessel, resulting in hypoxia or ischemia driving a complex series of events that includes excitotoxicity, oxidative stress, blood brain barrier dysfunction and post ischemic inflammation, that eventually leads to cell death (del Zoppo et al., 2000, Gartshore et al., 1997, Liu et al., 2006, Moskowitz et al., 2010, Tuttolomondo et al., 2014). Several mechanisms are involved in the pathophysiology of cerebral stroke; however, increasing evidence shows that inflammation process plays an important role in the progression of stroke pathogenesis (Barone and Feuerstein, 1999, Godinez-Rubi et al., 2013, Iadecola and Anrather, 2011, Lakhan et al., 2009, Muir et al., 2007). In the acute phase of stroke, several pro-inflammatory cytokines such as tumor necrosis factor-α (TNFα), interleukins (ILs) like IL-1α, IL-1β, IL-6, IL-20 as well as other potential cytotoxic molecules including nitric oxide (NO) and reactive oxygen species (ROS)are released in and around the affected brain region/s in response to stroke (Clausen et al., 2008, Jin et al., 2010, Lambertsen et al., 2012, Lucas et al., 2006). These post-ischemic inflammatory changes contribute to the dysfunction of blood brain barrier (BBB) and promote the brain edema formation, which eventually lead to neuronal cell death (Candelario-Jalil et al., 2009, Tuttolomondo et al., 2014, Yang and Rosenberg, 2011). Therefore, therapeutic targeting of the inflammatory pathway has become an important area of research in ischemic stroke.

Today in modern medicine era, to identify a drug that is safe, effective and affordable, is major challenge. Most of the clinical trials with the potential novel chemical entities failed in recent time (Stein, 2015). For this reason, the investigations have turned to the traditional medicine knowledge such as Ayurveda, which is used from the ancient time (Chopra and Doiphode, 2002, Patwardhan and Mashelkar, 2009). In India, Ayurveda, a traditional healing system, originated approximately six thousand years ago (Garodia et al., 2007, Sharma et al., 2007). According to the reports, it is well documented that Ayurveda medicines has been evaluated for the treatment of rheumatoid arthritis, bronchial asthma, depression, anxiety, learning, memory and focal cerebral ischemia (Chaudhary et al., 2003, Chowdhuri et al., 2002, Gupta et al., 2004, Khanna et al., 2007, Limpeanchob et al., 2008). Global acceptance of Ayurveda approach is not expensive and the compounds or drug isolated from the medicinal plants will be safe; however, additional clinical trials are needed to examine this potential traditional therapeutics.

This study aims to elucidate the possible mechanism of finding the neuroprotective effect of Sameerpannag Ras, an Ayurveda formulation, in acute cerebral ischemia mouse model. SR in Ayurveda is well known for mineral and metallic origin (rasushdhi) based medicine being in use for the treatment of rheumatoid arthritis, a chronic systematic inflammatory disorder and also been recommended in brain stroke as well (Ayurvedic formulary of India Kupipakva rasayan [Ayurveda Aushadhgunadharmashastra 4part]). It is also recommended in sannipat roga (unconsciousness, situation in which all three humors such as vata, pitta and kapha are vitiated), Kaphonmad (mania due to predominant kapha), Kaphaja sandhibandh (stiffness of joints due to kapha dosha), jvar (fever), swasa (asthmatic problem) and kasa (cough). Other constituents such as Giloy satva and Prawal pishti of sameerpannag ras mixture (SRM) possess the anti-oxidant properties (Meshram et al., 2013, Onkar et al., 2012, Shah and Goyal, 2010). Additionally, Ayurveda has novel approach in inflammation via its anti-inflammatory agent to treat many chronic diseases(Aggarwal et al., 2011). So here, the objective of the study is to investigate the effect of Sameerpannag Ras Mixture (SRM), if any, on the neurobehavioral deficit due to the striatal damage and whether its action is though attenuating the neuroinflammatory molecular responses in our internal carotid artery occlusion (ICAO) mouse model of acute ischemic stroke (AIS).

Section snippets

Animals and housing

All animal procedures were approved by the Institutional Animal Ethics Committee (IAEC/29/2013-14) at CCMB, Hyderabad, India. Adult male CD1 mice of 8-10 weeks old and weighing 30-35gm were used for the experiments. Animals were acclimatized to the experimental room environment maintained at ~24 °C and 12 h/12 h: light/dark schedule. Food and water were available ad libitum to the animals. After their habituation period, animals were randomly grouped according to experimental conditions.

Internal Carotid Artery Occlusion (ICAO) model

The

Effect of Sameerpannag Ras Mixture (SRM) on cerebral ischemia induced behavioral deficit

We assessed the effect of Sameerpannag Ras Mixture (SRM) treatment on the recovery of behavioral functions using rotarod performance test and grip strength with various doses (SI. Fig. 1a and b) and by evaluating its efficacy at behavioral level we fixed the dose as 40 mg/kg/dose, twice daily. Sameerpannag Ras Mixture (SRM) at the dose of 40 mg/kg attenuated the cerebral ischemia-induced behavioral deficits. Neurobehavioral assessment was performed on day1, day3, day5, and day7 post-ICAO. On NDS

Discussion

Cerebral stroke is one of the leading causes of mortality and morbidity in the world (Grady et al., 2015, Wardlaw et al., 2012). But ironically no new drug has come up in spite of intense research in the field. So, attention has moved towards uncovering the molecular mechanisms underlying the effectiveness of alternative medicine and in this direction our study demonstrates for the first time the molecular mechanisms involved in the efficacy of Sameerpannag Ras Mixture (SRM), an Ayurvedic

Acknowledgments

This research was supported by the Council of Scientific and Industrial Research (CSIR), India Network Projects [(BSC0103-UNDO to SC) and (BSC0115-miND to AK)]. ABW was supported by senior research fellowship of the University Grants Commission (UGC) (Sr.No. 2121130812 Ref. No:18-12/2011(II)EU-V). Director, NIPER, Mohali, India was greatly acknowledged for providing all kind of support for this work. In addition, the authors would like to specially acknowledge Mrs. B. Jyothilakshmi for the

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