Synergistic neuroprotective effect of microglial‑conditioned media treated with geniposide and ginsenoside Rg1 on hypoxia injured neurons

Wang,Jun; Hou,Jincai; Lei,Hongtao; Fu,Jianhua; Pan,Yanshu; Liu,Jianxun

doi:10.3892/mmr.2015.4094

Synergistic neuroprotective effect of microglial‑conditioned media treated with geniposide and ginsenoside Rg1 on hypoxia injured neurons

Authors:
- Jun Wang
- Jincai Hou
- Hongtao Lei
- Jianhua Fu
- Yanshu Pan
- Jianxun Liu
View Affiliations

Affiliations: Experiment Center, Institute of Basic Theory, Academy of Chinese Medical Sciences, Beijing 100700, P.R. China, Institute of Basic Medical Sciences of Xiyuan Hospital, Beijing Key Laboratory of Pharmacology of Chinese Materia Medica, Beijing 100091, P.R. China, Morphology Laboratory, The Experimental Research Center, Academy of Chinese Medical Sciences, Beijing 100700, P.R. China, Graduate School, Beijing University of Chinese Medicine, Beijing 100029, P.R. China
Published online on: July 20, 2015 https://doi.org/10.3892/mmr.2015.4094
Pages: 5328-5334

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

The synergistic mechanism underlying the effects of multi‑component combined drug use for complex diseases remains to be fully elucidated. Microglial activation following ischemia can either affect neural survival or cause neuronal injury. The aim of the present study was to determine the synergistic effect of geniposide and ginsenoside Rg1, based on microglial‑neuronal communication. N2a neuronal cells were divided into the following seven groups: Control group; normal cultured microglial cells in conditioned medium (N‑MG‑CM) group; oxygen‑glucose deprivation (OGD) model group; OGD‑injured MG‑CM (I‑MG‑CM) group; geniposide‑treated MG‑CM (G‑MG‑CM) group; ginsenoside Rg1‑treated MG‑CM (R‑MG‑CM) group; and combination‑treated MG‑CM (C‑MG‑CM) group. A series of assays were used to detect the effects of the different MG‑CM on neurons in terms of: (i) cell viability, determined using a Cell Counting Kit‑8; (ii) lactate dehydrogenase (LDH) leakage rate; (iii) expression of NMDAR1 and activated caspase‑3, detected using western blotting; (iv) mitochondrial transmembrane potential, determined by JC‑1; and (v) mitochondrial ultrastructural features, determined using electron microscopy. The experimental results demonstrated that MG‑CM including the integrated use of geniposide and ginsenoside Rg1 significantly protected neuronal cell viability and inhibited LDH leakage, suppressed the expression of N‑methyl‑D‑aspartate receptor subunit 1 and activated caspase‑3, increased the mitochondrial transmembrane potential and improved the mitochondrial ultrastructure. MG‑CM from separately used geniposide or ginsenoside Rg1 demonstrated differential neuroprotection at different levels. These findings revealed that the synergistic drug combination of geniposide and ginsenoside Rg1 in the treatment of stroke is a feasible approach for use.

View Figures

View References

1	Feigin VL, Lawes CM, Bennett DA, Barker-Collo SL and Parag V: Worldwide stroke incidence and early case fatality reported in 56 population-based studies: A systematic review. Lancet Neurol. 8:355–369. 2009. View Article : Google Scholar : PubMed/NCBI
2	Faure S, Oudart N, Javellaud J, Fournier A, Warnock DG and Achard JM: Synergistic protective effects of erythropoietin and olmesartan on ischemic stroke survival and post-stroke memory dysfunctions in the gerbil. J Hypertens. 24:2255–2261. 2006. View Article : Google Scholar : PubMed/NCBI
3	Smadja D, Chausson N, Joux J, Saint-Vil M, Signaté A, Edimonana M, Jeannin S, Bartoli B, Aveillan M, Cabre P and Olindo S: A new therapeutic strategy for acute ischemic stroke: Sequential combined intravenous tpa-tenecteplase for proximal middle cerebral artery occlusion based on first results in 13 consecutive patients. Stroke. 42:1644–1647. 2011. View Article : Google Scholar : PubMed/NCBI
4	Zhang J, Li P and Wang Y, Liu J, Zhang Z, Cheng W and Wang Y: Ameliorative effects of a combination of baicalin, jasminoidin and cholic acid on ibotenic acid-induced dementia model in rats. PLoS One. 8:e566582013. View Article : Google Scholar : PubMed/NCBI
5	Liu J, Zhou CX, Zhang ZJ, Wang LY, Jing ZW and Wang Z: Synergistic mechanism of gene expression and pathways between jasminoidin and ursodeoxycholic acid in treating focal cerebral ischemia-reperfusion injury. CNS Neurosci Ther. 18:674–682. 2012. View Article : Google Scholar : PubMed/NCBI
6	Lai AY and Todd KG: Microglia in cerebral ischemia: Molecular actions and interactions. Can J Physiol Pharmacol. 84:49–59. 2006. View Article : Google Scholar : PubMed/NCBI
7	Wake H, Moorhouse AJ, Jinno S, Kohsaka S and Nabekura J: Resting microglia directly monitor the functional state of synapses in vivo and determine the fate of ischemic terminals. J Neurosci. 29:3974–3980. 2009. View Article : Google Scholar : PubMed/NCBI
8	Ransohoff RM and Perry VH: Microglial physiology: Unique stimuli, specialized responses. Annu Rev Immunol. 27:119–145. 2009. View Article : Google Scholar : PubMed/NCBI
9	Hanisch UK and Kettenmann H: Microglia: Active sensor and versatile effector cells in the normal and pathologic brain. Nature Neurosci. 10:1387–1394. 2007. View Article : Google Scholar : PubMed/NCBI
10	Barclay AN, Wright GJ, Brooke G and Brown MH: CD200 and membrane protein interactions in the control of myeloid cells. Trends Immunol. 23:285–290. 2002. View Article : Google Scholar : PubMed/NCBI
11	Moon JB, Lee CH, Park CW, Cho JH, Hwang IK, Yoo KY, Choi JH, Shin HC and Won MH: Neuronal degeneration and microglial activation in the ischemic dentate gyrus of the gerbil. J Vet Med Sci. 71:1381–1386. 2009. View Article : Google Scholar : PubMed/NCBI
12	Taylor DL, Pirianov G, Holland S, McGinnity CJ, Norman AL, Reali C, Diemel LT, Gveric D, Yeung D and Mehmet H: Attenuation of proliferation in oligodendrocyte precursor cells by activated microglia. J Neurosci Res. 88:1632–1644. 2010.PubMed/NCBI
13	Hur J, Lee P, Kim MJ, Kim Y and Cho YW: Ischemia-activated microglia induces neuronal injury via activation of gp91phox NADPH oxidase. Biochem Biophys Res Commun. 391:1526–1530. 2010. View Article : Google Scholar
14	Liu Y, Hua Q, Lei H and Li P: Effect of Tong Luo Jiu Nao on Aβ-degrading enzymes in AD rat brains. J Ethnopharmacol. 137:1035–1046. 2011. View Article : Google Scholar : PubMed/NCBI
15	Li C, Pan Y, Jia X, Li K, Liu C, Wang X and Li P: Effects of Huanqin (dried root of Scutellaria baicalensis) and Zhizi (dried fruit of Gardenia jasminoides) used in combination on ischemic cascade reaction in the rat model of focal cerebral ischemia and reperfusion. Beijing Zhong Yi Yao Da Xue Xue Bao. 25:31–33. 2002.In Chinese.
16	Li K: Systematic review of Sanqi agents for acute ischaemic stroke. Lin Chuang Hui Cui. 22:1–5. 2007.In Chinese.
17	Chen J, Sun M, Wang X, Lu J, Wei Y, Tan Y, Liu Y, Götz J, He R and Hua Q: The herbal compound geniposide rescues formaldehyde-induced apoptosis in N2a neuroblastoma cells. Sci China Life Sci. 57:412–421. 2014. View Article : Google Scholar : PubMed/NCBI
18	Xie CL, Wang WW, Xue XD, Zhang SF, Gan J and Liu ZG: A systematic review and meta-analysis of Ginsenoside-Rg1 (G-Rg1) in experimental ischemic stroke. Sci Rep. 5:77902015. View Article : Google Scholar : PubMed/NCBI
19	Wang J, Li PT, Du H, Hou JC, Li WH, Pan YS and Chen HC: Tong Luo Jiu Nao injection, a traditional Chinese medicinal preparation, inhibits MIP-1β expression in brain microvascular endothelial cells injured by oxygen-glucose deprivation. J Ethnopharmacol. 141:151–157. 2012. View Article : Google Scholar : PubMed/NCBI
20	Wang J, Hou J, Zhang P, Li D, Zhang C and Liu J: Geniposide reduces inflammatory responses of oxygen-glucose deprived rat microglial cells via inhibition of the TLR4 signaling pathway. Neurochem Res. 37:2235–2248. 2012. View Article : Google Scholar : PubMed/NCBI
21	Wang J, Li PT, Du H, Hou JC, Li WH, Pan YS, Hua Q and Chen HC: Impact of paracrine signals from brain microvascular endothelial cells on microglial proliferation and migration. Brain Res Bull. 86:53–59. 2011. View Article : Google Scholar : PubMed/NCBI
22	Carey KM, Comee MR, Donovan JL and Kanaan AO: A polypill for all? Critical review of the polypill literature for primary prevention of cardiovascular disease and stroke. Ann Pharmacother. 46:688–695. 2012. View Article : Google Scholar : PubMed/NCBI
23	Zimmermann GR, Lehár J and Keith CT: Multi-target therapeutics: When the whole is greater than the sum of the parts. Drug Discov Today. 12:34–42. 2007. View Article : Google Scholar : PubMed/NCBI
24	Li H, Xu J, Wang X and Yuan G: Protective effect of ginsenoside Rg1 on lidocaine-induced apoptosis. Mol Med Rep. 9:395–400. 2014.
25	Umebayashi D, Natsume A, Takeuchi H, Hara M, Nishimura Y, Fukuyama R, Sumiyoshi N and Wakabayashi T: Blockade of gap junction hemichannel protects secondary spinal cord injury from activated microglia-mediated glutamate exitoneurotoxicity. J Neurotrauma. 31:1967–1974. 2014. View Article : Google Scholar : PubMed/NCBI
26	Kawasaki Y, Kohno T, Zhuang ZY, Brenner GJ, Wang H, Van Der Meer C, Befort K, Woolf CJ and Ji RR: Ionotropic and metabotropic receptors, protein kinase A, protein kinase C, and Src contribute to C-fiber-induced ERK activation and cAMP response element-binding protein phosphorylation in dorsal horn neurons, leading to central sensitization. J Neurosci. 24:8310–8321. 2004. View Article : Google Scholar : PubMed/NCBI
27	Novitskaya YA, Dravolina OA, Zvartau EE, Danysz W and Bespalov AY: Interaction of blockers of ionotropic NMDA receptors and metabotropic glutamate receptors in a working memory test in rats. Neurosci Behav Phsyiol. 40:807–811. 2010. View Article : Google Scholar
28	Xu GY, Liu S, Hughes MG and McAdoo DJ: Glutamate-induced losses of oligodendrocytes and neurons and activation of caspase-3 in the rat spinal cord. Neuroscience. 153:1034–1047. 2008. View Article : Google Scholar : PubMed/NCBI
29	Yin J, Li H, Feng C and Zuo Z: Inhibition of brain ischemia-caused notch activation in microglia may contribute to isoflurane postconditioning-induced neuroprotection in male rats. CNS Neurol Disord Drug Targets. 13:718–732. 2014. View Article : Google Scholar : PubMed/NCBI
30	Yang X and Huang N: Berberine induces selective apoptosis through the AMPK-mediated mitochondrial/caspase pathway in hepatocellular carcinoma. Mol Med Rep. 8:505–510. 2013.PubMed/NCBI
31	Wang L, Chen M, Yuan L, Xiang Y, Zheng R and Zhu S: 14,15-EET promotes mitochondrial biogenesis and protects cortical neurons against oxygen/glucose deprivation-induced apoptosis. Biochem Biophys Res Commun. 450:604–609. 2014. View Article : Google Scholar : PubMed/NCBI
32	Wang J, Wang P, Li S, Wang S, Li Y, Liang N and Wang M: Mdivi-1 prevents apoptosis induced by ischemia-reperfusion injury in primary hippocampal cells via inhibition of reactive oxygen species-activated mitochondrial pathway. J Stroke Cerebrovasc Dis. 23:1491–1499. 2014. View Article : Google Scholar : PubMed/NCBI
33	Lee WK and Thévenod F: A role for mitochondrial aquaporins in cellular life-and-death decisions? Am J Physiol Cell Physiol. 291:C195–C202. 2006. View Article : Google Scholar : PubMed/NCBI
34	Wang J, Hou JC, Xiang LH, Zhang J and Ju DH: Compatibility of geniposide and ginsenoside rg1: Their regulating roles in secretion of anoxia induction injured microglia inflammatory cytokines. Zhongguo Zhong Xi Yi Jie He Za Zhi. 34:91–95. 2014.In Chinese. PubMed/NCBI