An in vitro study of neuroprotective properties of traditional Chinese herbal medicines thought to promote healthy ageing and longevity

Abstract

Background: Age is the leading risk factor for acute and chronic neurodegenerative diseases. The Shen Nong Ben Cao Jing, the oldest known compendium of Chinese materia media, lists herbal medicines that were believed to exert neither fast acting pharmacological effects nor discernible toxicity, but to promote general health and longevity. In modern terms, these herbal medicines could be considered as complementary health care products for prevention rather than treatment of diseases. In the present study, we examined whether a selection of 13 such herbal medicines exhibited neuroprotective activity.

Methods: The antioxidant capacity of the herbal extracts was determined using three non-cellular assays measuring the total phenol content (FCR assay), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging capacity and oxygen radical absorbance capacity (ORAC). Cytotoxic effects of the herbal extracts were assayed in cultured mouse cortical neurons and their neuroprotective activities were studied using staurosporine-induced apoptosis of the cultured neurons.

Results: Most of the herbal extracts showed negligible toxic effects at 100 μg/ml. However, Polygonum multiflorum and Rhodiola rosea exhibited some neurotoxicity at this concentration. Extracts of Ganoderma lucidum, Glycyrrhiza glabra, Schizandra chinensis, and Polygonum cuspidatum inhibited staurosporine-induced apoptosis by 30 - 50% in a dose-dependent manner. The neuroprotective effects of Polygonum cuspidatum were predominantly due to its major ingredient, resveratrol. The effective herbal extracts showed various levels of reactive oxygen species (ROS) scavenging capacity, which was significantly correlated with their neuro- protective activity. However, P. multiflorum and R. rosea extracts proved to be the exception as they exhibited a high level of antioxidant capacity, but did not exhibit neuroprotective effects in cell-based assay.

Conclusions: This in vitro study provides evidence for neuroprotective activity of some Chinese herbal medicines traditionally used to promote healthy ageing and longevity. Our results provide a justification for further study of these herbal extracts in neurodegenerative animal models to assess their safety and effectiveness as a basis for subsequent clinical trials. These herbal medicines might potentially offer a novel preemptive neuroprotective approach in neurodegenerative diseases and might be developed for use in persons at risk.

Figure 1

Cytotoxic effects of herbal extracts in cultured primary mouse cortical neurons. The neuronal cells were exposed to 100 μg/ml of the herbal extracts for 24 h. Culture media (neurobasal medium supplemented with 2% B-27) containing vehicle only were used as a negative control (defined as 0%) and media containing staurosporine were used as positive control (defined as 100%).

Figure 2

Neuroprotective effects of the herbal extracts. The relative cytotoxicity of neuronal cells was measured for the cell cultures (n = 3) co-treated with 100 μg/ml of the herbal extracts and 500 nM staurosporine for 24 h. * p value < 0.05; ** p value < 0.01, and *** p value < 0.001 compared with the positive control.

Figure 3

Dose-dependence of neuroprotective effects of selected herbal extracts and purified compounds. The herbal extracts, Glycyrrhiza glabra(A, left panel), Schizandra chinensis is (B, left panel) and Polygonum cuspidatum(C, left panel), were used in the concentration range of 3 – 100 μg/ml. The pure compounds, glabridin (A, right panel), schisandrin (B, right panel) and resveratrol (C, right panel) were used in the range of 1 – 65 μg/ml. * p value < 0.05; ** p value < 0.01, and *** p value < 0.001 compared with the positive control.

Figure 4

Correlation analysis between the neuroprotective activity and antioxidant capacity of the herbal extracts. The neuroprotective activity of the herbal extracts was normalized to the cytotoxicity of staurosporine (100%). The antioxidant activities were determined by non-cellular DPPH, ORAC and FCR assays. Gallic acid equivalence (GAE) was used for relative quantification of the antioxidant activity. The correlation curves were plotted and the correlation level was analyzed for all 13 herbal extracts (A) and for selected bioactive herbal extracts (B), G. lucidum (4), G. glabra (5), S. chinensis (12), and P. cuspidatum (13), using GraphPad Prism software.