Neuroprotective activity of compounds of Clematis mandshurica against glutamate-induced cell death in HT22 cells

Clematis mandshurica (C. mandshurica) is a traditional crude drug for treatment of urethritis, carbuncles and carcinoma [1 – 3]. We revealed that C. mandshurica have neuroprotective activity against glutamate-induced cell death in HT22 cells. We isolated and identified 24 compounds from C. mandshurica extract. The neuroprotective effect of 1 – 24 compounds was evaluated against glutamate-induced cell death in hippocampal HT22 cell line As a result, methyl 5-(4-methylcyclohexa-1,4-dienyl)hexanoate showed higher neuroprotective effect (relative protection of 28.74 ± 4.32% at 100µM) than that of other compounds. Methyl 5-(4-methylcyclohexa-1,4-dienyl)hexanoate also reduced ROS levels (84.28 ± 7.5% of 100µM) and intracellular Ca²⁺ accumulation (102.58 ± 5.04% at 100µM). Methyl 5-(4-methylcyclohexa-1,4-dienyl)hexanoate significantly recovered the mitochondrial membrane potential to 92.98 ± 2.13% of control at 100µM. Methyl 5-(4-methylcyclohexa-1,4-dienyl)hexanoate also significantly recovered glutamate-induced depletion of GSH (86.60 ± 1.99 at 100µM), GR (87.85 ± 2.46% at 100µM) and GPx (90.01 ± 5.13% at 100µM). These results suggest that Methyl 5-(4-methylcyclohexa-1,4-dienyl)hexanoate exerted neuroprotective effect by antioxidant effect. Neuroprotective effect of methyl 5-(4-methylcyclohexa-1,4-dienyl)hexanoate may be associated with anti-oxidantive effect.

No.	Compound
1	(2S,3S,4R,5S)-2-(3,4-dihydroxy-3,4-dihydro-2 H-pyran-2-yloxy)-6-(hydroxymethyl)tetrahydro-2 H-pyran-3,4,5-triol
2	6-((2S,3S,4R,5S)-3,4,5-trihydroxy-6-(hydroxymethyl) tetrahydro-2 H-pyran-2-yloxy)-5,6-dihydro-2 H-pyran-2,4,5-triol
3	methyl nonanoate
4	methyl ester heptanoic acid
5	(E)-methyl dodec-9-enoate
6	linolenic acid
7	linoleic acid
8	5-[(β-D-glucopyranosyloxy)methyl]-, (5S)- 2(5 H)-furanone
9	hexahydro-3a,7a-dimethyl-, (3aR,7aS)-rel-4(1 H)-isobenzofuranone
10	3a,7,7-trimethylhexahydro-2 H-furo[2,3-c]pyran-2-one
11	γ,4-dimethyl-1,4-cyclohexadiene-1-butanoic acid
12	methyl 5-(4-methylcyclohexa-1,4-dienyl)hexanoate
13	Stigmasterol
14	stigmasterol glucoside
15	β-sitosterol
16	pyrocincholic acid methyl ester
17	Hederagenin
18	(3S,3'S,4R,4'R,5S,5'S,6S,6'S)-6,6'-(2-(hydroxymethyl)-3-(3-methylbut-2-enyl)-1,4-phenylene)bis(oxy)bis(2(hydroxymethyl)tetrahy -dro-2 H-pyran-3,4,5-triol)
19	ferulic acid
20	2-ethynyl-1,4-dimethoxybenzene
21	6-ethoxy-3-oxochroman-8-carbaldehyde
22	2,4-dihydroxy-6-(hydroxymethyl) benzaldehyde
23	2-[(8-acetyl-6-methoxy-4-methyl-2-oxo-2 H-1-benzopyran-7-yl)oxy]-acetic acid
24	2,3-dihydro-5-methyl-7-benzofurancarboxaldehyde

Acknowledgements: This research was supported by Kangwon National University.

Keywords: Clematis mandshurica, neuroprotective effect, oxidative stress, isolation of compounds.

References:

[1] The Korea Food and Drug Administration. The Korean herbal pharmacopoeia: notification; 2014: 2014 – 2074.

[2] Jung YB, Roh KJ, Jung JA, Jung K, Yoo H, Cho YB, Kwak WJ, Kim DK, Kim KH, Han CK. Effect of SKI 306X, a new herbal anti-arthritic agent, in patients with osteoarthritis of the knee: a double-blind placebo controlled study. Am J Chin Med 2001; 29: 485 – 491.

[3] Lee SW, Chung WT, Choi SM, Kim KT, Yoo KS, Yoo YH. Clematis mandshurica protected to apoptosis of rat chondrocytes. J Ethnopharmacol 2005; 101: 294 – 298.