Ginkgolide B enhances the differentiation of preosteoblastic MC3T3-E1 cells through VEGF: Involvement of the p38 MAPK signaling pathway

Luo,Jiaquan; Zhong,Yu; Huang,Sheng; Li,Liangping; Zhang,Chi; Zou,Xuenong

doi:10.3892/mmr.2016.5829

Ginkgolide B enhances the differentiation of preosteoblastic MC3T3-E1 cells through VEGF: Involvement of the p38 MAPK signaling pathway

Authors:
- Jiaquan Luo
- Yu Zhong
- Sheng Huang
- Liangping Li
- Chi Zhang
- Xuenong Zou
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Affiliations: Guangdong Provincial Key Laboratory of Orthopedics and Traumatology, The First Affiliated Hospital of Sun Yat‑Sen University, Guangzhou, Guangdong 510080, P.R. China, Department of Pharmacology, Peking University International Hospital, Beijing 102206, P.R. China
Published online on: October 12, 2016 https://doi.org/10.3892/mmr.2016.5829
Pages: 4787-4794

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Abstract

Ginkgolide B (GB) is one of the ginkgolides isolated from the leaves of the Ginkgo biloba tree. Our previous study indicated that GB promotes the proliferation, migration and adhesion of endothelial progenitor cells, and the induction of angiogenesis through vascular endothelial factor (VEGF). In the present study, the effects of GB on the differentiation of MC3T3‑E1 cells and the signaling pathway involved were investigated in vitro. The MC3T3‑E1 cell viability activities were assessed using an MTS assay. Measurements of alkaline phosphatase activity and Alizarin Red staining were used to identify osteoblastic differentiation of the MC3T3‑E1 cells. The mRNA and secretion levels of VEGF were detected using reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) analysis and enzyme-linked immunosorbent assays, respectively. The protein expression levels of phosphorylation‑associated markers were detected using western blot analysis and associated gene expression was determined using RT‑qPCR analysis. It was found that GB significantly promoted alkaline phosphatase activity and osteoblastic mineralization in the MC3T3‑E1 cells. In addition, the mRNA expression and secretion levels of VEGF in the MC3T3‑E1 cells were significantly increased in MC3T3‑E1 cells treated with GB. SB203580, a specific inhibitor of p38 mitogen‑activated protein (MAP) kinase, markedly suppressed the GB‑induced p38 kinase phosphorylation and GB‑induced synthesis of VEGF. PD98059, an inhibitor of the upstream kinase, which activates p44/p42 MAP kinase, had minimal effect on the GB‑induced phosphorylation of p44/p42 MAP kinase or the GB‑induced synthesis of VEGF. Taken together, these results indicated that GB promoted osteoblastic differentiation of the MC3T3‑E1 cells through VEGF, and that the p38, but not the p44/p42 MAP kinase signaling pathway, was involved in the GB‑induced synthesis of VEGF.

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1	Liang L: The history of cultivation techniques of Ginkgo biloba in China. Agricultural Archaeology. 1:259–261. 2002.
2	Dias MA, Sampaio AL, Venosa AR, Meneses EA and Oliveira CA: The chemopreventive effect of Ginkgo biloba extract 761 against cisplatin ototoxicity: A pilot study. Int Tinnitus J. 19:12–19. 2015. View Article : Google Scholar : PubMed/NCBI
3	Shen J, Wang J, Zhao B, Hou J, Gao T and Xin W: Effects of EGb 761 on nitric oxide and oxygen free radicals, myocardial damage and arrhythmia in ischemia-reperfusion injury in vivo. Biochim Biophys Acta. 1406:228–236. 1998. View Article : Google Scholar : PubMed/NCBI
4	Zhang WR, Hayashi T, Kitagawa H, Sasaki C, Sakai K, Warita H, Wang JM, Shiro Y, Uchida M and Abe K: Protective effect of ginkgo extract on rat brain with transient middle cerebral artery occlusion. Neurol Res. 22:517–521. 2000. View Article : Google Scholar : PubMed/NCBI
5	Ahlemeyer B, Mowes A and Krieglstein J: Inhibition of serum deprivation- and staurosporine-induced neuronal apoptosis by Ginkgo biloba extract and some of its constituents. Eur J Pharmacol. 367:423–430. 1999. View Article : Google Scholar : PubMed/NCBI
6	Abdel-Wahab BA and Abd El-Aziz SM: Ginkgo biloba protects against intermittent hypoxia-induced memory deficits and hippocampal DNA damage in rats. Phytomedicine. 19:444–450. 2012. View Article : Google Scholar : PubMed/NCBI
7	Nash KM and Shah ZA: Current perspectives on the beneficial role of Ginkgo biloba in neurological and cerebrovascular disorders. Integr Med Insights. 10:1–9. 2015. View Article : Google Scholar : PubMed/NCBI
8	Nabavi SM, Habtemariam S, Daglia M, Braidy N, Loizzo MR, Tundis R and Navabi SF: Neuroprotective effects of Ginkgolide B against ischemic stroke: A review of current literature. Curr Top Med Chem. 15:2222–2232. 2015. View Article : Google Scholar : PubMed/NCBI
9	Li R, Chen B, Wu W, Bao L, Li J and Qi R: Ginkgolide B suppresses intercellular adhesion molecule-1 expression via blocking nuclear factor-kappaB activation in human vascular endothelial cells stimulated by oxidized low-density lipoprotein. J Pharmacol Sci. 110:362–369. 2009. View Article : Google Scholar : PubMed/NCBI
10	Huang M, Qian Y, Guan T, Huang L, Tang X and Li Y: Different neuroprotective responses of Ginkgolide B and bilobalide, The Two Ginkgo Components Hyperglycemia. Eur J Pharmacol. 677:71–76. 2012. View Article : Google Scholar : PubMed/NCBI
11	Huang JYSJMS: Protective effects of ginkgolide B on cerebral ischemia reperfusion injury in rats. Chin Pharmacol Bull. 269–272. 2008.
12	Botao Y, Ma J, Xiao W, Xiang Q, Fan K, Hou J, Wu J and Jing W: Protective effect of ginkgolide B on high altitude cerebral edema of rats. High Alt Med Biol. 14:61–64. 2013. View Article : Google Scholar : PubMed/NCBI
13	Tang Y, Huang B, Sun L, Peng X, Chen X and Zou X: Ginkgolide B promotes proliferation and functional activities of bone marrow-derived endothelial progenitor cells: Involvement of Akt/eNOS and MAPK/p38 signaling pathways. Eur Cell Mater. 21:459–469. 2011.PubMed/NCBI
14	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) Method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
15	Yang Y, Chin A, Zhang L, Lu J and Wong RW: The role of traditional Chinese medicines in osteogenesis and angiogenesis. Phytother Res. 28:1–8. 2014. View Article : Google Scholar : PubMed/NCBI
16	Li N, Jiang Y, Wooley PH, Xu Z and Yang SY: Naringin promotes osteoblast differentiation and effectively reverses ovariectomy-associated osteoporosis. J Orthop Sci. 18:478–485. 2013. View Article : Google Scholar : PubMed/NCBI
17	Huh JE, Yang HR, Park DS, Choi DY, Baek YH, Cho EM, Cho YJ, Kang-Il K, Kim DY and Lee JD: Puerariae radix promotes differentiation and mineralization in human osteoblast-like SaOS-2 cells. J Ethnopharmacol. 104:345–350. 2006. View Article : Google Scholar : PubMed/NCBI
18	Lamant V, Mauco G, Braquet P, Chap H and Douste-Blazy L: Inhibition of the metabolism of platelet activating factor (PAF-acether) by three specific antagonists from Ginkgo biloba. Biochem Pharmacol. 36:2749–2752. 1987. View Article : Google Scholar : PubMed/NCBI
19	Chan PC, Xia Q and Fu PP: Ginkgo biloba leave extract: Biological, medicinal and toxicological effects. J Environ Sci Health C Environ Carcinog Ecotoxicol Rev. 25:211–244. 2007. View Article : Google Scholar : PubMed/NCBI
20	Xie H, Cui Z, Wang L, Xia Z, Hu Y, Xian L, Li C, Xie L, Crane J, Wan M, et al: PDGF-BB secreted by preosteoclasts induces angiogenesis during coupling with osteogenesis. Nat Med. 20:1270–1278. 2014. View Article : Google Scholar : PubMed/NCBI
21	Street J, Winter D, Wang JH, Wakai A, McGuinness A and Redmond HP: Is human fracture hematoma inherently angiogenic? Clin Orthop Relat Res. 224–237. 2000. View Article : Google Scholar : PubMed/NCBI
22	Geiger F, Bertram H, Berger I, Lorenz H, Wall O, Eckhardt C, Simank HG and Richter W: Vascular endothelial growth factor gene-activated matrix (VEGF165-GAM) enhances osteogenesis and angiogenesis in large segmental bone defects. J Bone Miner Res. 20:2028–2035. 2005. View Article : Google Scholar : PubMed/NCBI
23	D'Alimonte I, Nargi E, Mastrangelo F, Falco G, Lanuti P, Marchisio M, Miscia S, Robuffo I, Capogreco M, Buccella S, et al: Vascular endothelial growth factor enhances in vitro proliferation and osteogenic differentiation of human dental pulp stem cells. J Biol Regul Homeost Agents. 25:57–69. 2011.PubMed/NCBI
24	Hah YS, Jun JS, Lee SG, Park BW, Kim DR, Kim UK, Kim JR and Byun JH: Vascular endothelial growth factor stimulates osteoblastic differentiation of cultured human periosteal-derived cells expressing vascular endothelial growth factor receptors. Mol Biol Rep. 38:1443–1450. 2011. View Article : Google Scholar : PubMed/NCBI
25	Tan YY, Yang YQ, Chai L, Wong RW and Rabie AB: Effects of vascular endothelial growth factor (VEGF) on MC3T3-E1. Orthod Craniofac Res. 13:223–228. 2010. View Article : Google Scholar : PubMed/NCBI
26	Deckers MM, Karperien M, van der Bent C, Yamashita T, Papapoulos SE and Löwik CW: Expression of vascular endothelial growth factors and their receptors during osteoblast differentiation. Endocrinology. 141:1667–1674. 2000. View Article : Google Scholar : PubMed/NCBI
27	Tokuda H, Kozawa O, Miwa M and Uematsu T: p38 mitogen-activated protein (MAP) kinase but not p44 p42 MAP kinase is involved in prostaglandin E1-induced vascular endothelial growth factor synthesis in osteoblasts. J Endocrinol. 170:629–638. 2001. View Article : Google Scholar : PubMed/NCBI
28	Kozawa O, Kawamura H, Hatakeyama D, Matsuno H and Uematsu T: Endothelin-1 induces vascular endothelial growth factor synthesis in osteoblasts involvement of p38 mitogen-activated protein kinase. Cellular Signal. 12:375–380. 2000. View Article : Google Scholar
29	Tokuda H, Hatakeyama D, Akamatsu S, Tanabe K, Yoshida M, Shibata T and Kozawa O: Involvement of MAP kinases in TGF-beta-stimulated vascular endothelial growth factor synthesis in osteoblasts. Arch Biochem Biophys. 415:117–125. 2003. View Article : Google Scholar : PubMed/NCBI
30	Chen M, Chen PM, Dong QR, Huang Q, She C and Xu W: p38 signaling in titanium particle-induced MMP-2 secretion and activation in differentiating MC3T3-E1 cells. J Biomed Mater Res A. 102:2824–2832. 2014. View Article : Google Scholar : PubMed/NCBI
31	Greenblatt MB, Shim JH, Zou W, Sitara D, Schweitzer M, Hu D, Lotinun S, Sano Y, Baron R, Park JM, et al: The p38 MAPK pathway is essential for skeletogenesis and bone homeostasis in mice. J Clin Invest. 120:2457–2473. 2010. View Article : Google Scholar : PubMed/NCBI
32	Hu Y, Chan E, Wang SX and Li B: Activation of p38 mitogen-activated protein kinase is required for osteoblast differentiation. Endocrinology. 144:2068–2074. 2003. View Article : Google Scholar : PubMed/NCBI
33	Thouverey C and Caverzasio J: The p38α MAPK positively regulates osteoblast function and postnatal bone acquisition. Cell Mol Life Sci. 69:3115–3125. 2012. View Article : Google Scholar : PubMed/NCBI