Elsevier

Neuropharmacology

Volume 61, Issue 4, September 2011, Pages 622-631
Neuropharmacology

MLC901, a Traditional Chinese Medicine protects the brain against global ischemia

https://doi.org/10.1016/j.neuropharm.2011.05.003Get rights and content

Abstract

Global ischemia leads to damage in the hippocampal CA1 region and is associated with behavioral deficits. NeuroAid (MLC601 and MLC901), a Traditional Chinese Medicine is used in China for patients after stroke. We have investigated here the effects of MLC901 on brain injury and deficits after global ischemia in the rat. Global ischemia induced by four-vessel occlusion resulted in degeneration of CA1 neurons. MLC901 (0.074 mg/ml) prevented both necrosis and apoptosis of neurons up to 3 h after ischemia. These positive MLC901 effects were associated with a decrease in Bax expression and in levels of the lipid peroxidation product malondialdehyde. Using the PI3-kinase inhibitor LY294002 we also demonstrated the critical role of the Akt pathway in MLC901-mediated neuroprotection. MLC901 enhanced neurogenesis. Furthermore, MLC901 improved functional recovery of rats after global ischemia as assessed by the Morris water maze. In this test MLC901 reduced the increase in escape latency and in swim distance induced by ischemia. MLC901 also improved post-ischemic grip strength. If observations made with rats can be extended to humans, then MLC901 will represent a novel therapeutic strategy after cardiac arrest with a clinically interesting time window of protection.

Highlights

► We studied effects of MLC901 on rat brain injury and deficits after global ischemia. ► MLC901 prevented necrosis and apoptosis of neurons up to 3 h after ischemia. ► MLC901 enhanced neurogenesis. ► We showed the critical role of the Akt pathway in MLC901-mediated neuroprotection. ► MLC901 decreased functional deficits induced by ischemia.

Introduction

Global cerebral ischemia is a clinical outcome occurring as a consequence of cardiac arrest, reversible severe hypotension or other situations that deprive the brain of oxygen and glucose. Each year, estimated 350,000 people undergo sudden cardiac arrest in Europe (Herlitz et al., 1999). Recovery without residual neurologic damage after global ischemia is rare. Besides the dramatic clinical aspects of the disease, subsequent neurological injuries represent a considerable financial burden in medical and rehabilitation expenses and lost productivity. Hypothermia has been described as the only therapy which can improve outcome after cardiac arrest (Nolan et al., 2003, Sterz et al., 2006, Zhao et al., 2007). The need for new therapeutic strategies is imperative (Ginsberg, 2008).

The pyramidal neurons of the CA1 region in the hippocampus are among the cells most vulnerable to loss of blood supply to the brain in humans and rodents (Pulsinelli and Brierley, 1979). Cell death occurs days after the ischemic insult, a phenomenon termed delayed neuronal death, which is associated with severe behavioral impairments. Global ischemia when it is not lethal is often followed by some types of recovery. A large part of the recovery after ischemia is associated with the capacity of the brain to induce spontaneous adult neurogenesis, which generates functional neurons in subventricular and subgranular zones of the hippocampus (Alvarez-Buylla and Lim, 2004, Eriksson et al., 1998, Sharp et al., 2002, Di Filippo et al., 2008, Zhang et al., 2008). However, brain self-repair by neuronal replacement from endogenous precursors is insufficient and functional recovery remains incomplete. Amplification of this self-repair mechanism could be a promising strategy for developing restorative therapies for global ischemia.

Interestingly, NeuroAid (MLC601) is a Traditional Chinese Medicine (TCM), which was first registered by the Sino Food and Drug Administration in 2001 after being evaluated in clinical trials in China as a drug to facilitate recovery after stroke (Chen et al., 2009). It combines 9 herbal and 5 animal components. In a previous study, we demonstrated in a rodent model of focal ischemia that MLC601 and also MLC901, which is a simplified version with only the nine herbal components, improved survival, protected the brain from ischemic injury and drastically decreased functional deficits (Heurteaux et al., 2010). MLC601/MLC901 also prevented neuronal death in an in vitro model of excitotoxicity using cultures of cortical neurons exposed to glutamate. In addition, MLC601/MLC901 treatments have been shown to induce neurogenesis, promote cell proliferation and stimulate the development of a dense axonal and dendritic network. In the present work we investigated the therapeutic effectiveness of MLC901 on brain injury and motor deficits induced by global ischemia (four-vessel occlusion model) in rats. We also deciphered some of mechanisms associated with the effects of this TCM by analyzing MLC901 actions on neuronal cell death (necrosis and apoptosis), lipid peroxidation, phosphinositide-3-kinase/Akt pathway and neurogenesis.

Section snippets

Animals

All experiments were performed on male Wistar rats (250 g) from Charles River Laboratories (France) and used according to policies on the care and use of laboratory animals of European Communities Council Directive (86/609/EEC). The local Ethics Committee approved the experiments (protocol numbers NCA/2006/10-1 and NCA/2006/10-2). All efforts were made to minimize animal suffering and reduce the number of animals used. The animals were housed under controlled laboratory conditions with a 12-h

Neuroprotective effect of MLC901 on necrotic and apoptotic cell death induced by global ischemia

To assay the potential neuroprotective effects of MLC901 against global ischemia, we carried out a dose-effect response in the rat 4 four-vessel occlusion model. Animals received an intraperitoneal MLC901 injection 30 min post-ischemia followed by one injection per day for seven days after reperfusion at the concentrations/ml of 74, 7.4, 0.74 or 0.074 mg/ml (n = 6 per experimental group). The two lower doses of MLC901 injected (0.74 and 0.074 mg/ml) induced a 100% survival rate of rats as

Discussion

The classical development of drugs for neuroprotection and/or repair in ischemic situations normally goes from molecular targets to animal models and to clinical trials in humans. Things are different with Traditional Chinese Medicine, that has long been administered in humans but that often needs a scientific rationale for its putative mode of action. NeuroAid (MLC601) has been first tested in clinical trials conducted in Asia. These trials as well as clinical reports have demonstrated a high

Disclosure/conflict of interest

This work is supported by a CNRS/MOLEAC contract. ML is vice-president for research of Moleac.

Acknowledgments

The authors are very grateful to D. Picard (Moleac Singapore) for helpful and inspiring discussions concerning the program and particularly the follow-up of the clinical effects of MLC601 and for providing MLC901 capsules. We thank Dr A. Patel for careful reading of the paper. This work is supported by the Centre National de la Recherche Scientifique (CNRS). HQ is very grateful for the financial support of CNRS/CHU Nice.

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