Atherosclerosis is one common chronic inflammatory disease in which angiogenesis is involved. Here we established an in vitro cell model of angiogenesis made by human dermal microvascular endothelial cells (HMEC-1) and work to investigate the role of triptolide (TPL) in this model. To induce angiogenesis, HMEC-1 cells were cultured in Matrigel-conditioned medium. The ratio of tubes to nucleus was detected. To evaluate angiogenesis, Western blot assay was carried out to detect endothelial nitric oxide synthase (eNOS), vascular endothelial growth factor receptor-2 (VEGFR2) and VEGF. Cell counting kit-8 was utilized to estimate the viability of HMEC-1 cells. microRNA (miR)-92a was analyzed by qRT-PCR. The targeting relationship between integrin subunit alpha 5 (ITGA5) and miR-92a was verified through luciferase activity assay. The effects of ITGA5 on signaling transducers (ERK, PI3K, and AKT) in a phosphorylated form were valued using Western blot method. After stimulated by TPL, LY294002 and PD98059, the alteration in phosphorylation of the signaling transducers was evaluated by Western blot assay. The ratio of tubes to nucleus and angiogenesis related factors were increased with the delaying of culture time. TPL decreased the expression of angiogenesis factors. Furthermore, miR-92a was upregulated by TPL and miR-92a silence upregulated angiogenesis factors. In addition, TPL decreased ITGA5 which was proved as a target of miR-92a. ITGA5 overexpression resulted in the abundance of angiogenesis factors while ITGA5 silence led to the opposite results. Meanwhile, ITGA5 overexpression increased phosphorylation of ERK, PI3K and AKT while ITGA5 silence reversed the trend. TPL (as an anti-angiogenesis agent) suppressed angiogenesis by upregulating miR-92a, and miR-92a-mediated down-regulation of ITGA5 blocked the signaling transduction of ERK and PI3K/AKT pathways.
This is a preview of subscription content, access via your institution.
Buy single article
Instant access to the full article PDF.
Tax calculation will be finalised during checkout.
Subscribe to journal
Immediate online access to all issues from 2019. Subscription will auto renew annually.
Tax calculation will be finalised during checkout.
The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.
Andreou I, Sun X, Stone PH, Edelman ER, Feinberg MW (2015) miRNAs in atherosclerotic plaque initiation, progression, and rupture. Trends Mol Med 21:307–318
Bonauer A, Carmona G, Iwasaki M, Mione M, Koyanagi M, Fischer A, Burchfield J, Fox H, Doebele C, Ohtani K, Chavakis E, Potente M, Tjwa M, Urbich C, Zeiher AM, Dimmeler S (2009) MicroRNA-92a controls angiogenesis and functional recovery of ischemic tissues in mice. Science 324:1710–1713
Cai J, Jiang WG, Ahmed A, Boulton M (2006) Vascular endothelial growth factor-induced endothelial cell proliferation is regulated by interaction between VEGFR-2, SH-PTP1 and eNOS. Microvasc Res 71:20–31
Chen J, Qiao Y, Tang B, Chen G, Liu X, Yang B, Wei J, Zhang X, Cheng X, Du P, Jiang W, Hu Q, Hua ZC (2017) Modulation of Salmonella tumor-colonization and Intratumoral anti-angiogenesis by Triptolide and its mechanism. Theranostics 7:2250–2260
Dai J, Peng L, Fan K, Wang H, Wei R, Ji G, Cai J, Lu B, Li B, Zhang D, Kang Y, Tan M, Qian W, Guo Y (2009) Osteopontin induces angiogenesis through activation of PI3K/AKT and ERK1/2 in endothelial cells. Oncogene 28:3412–3422
Efimenko A, Sagaradze G, Akopyan Z, Lopatina T, Kalinina N (2016) Data supporting that miR-92a suppresses angiogenic activity of adipose-derived mesenchymal stromal cells by down-regulating hepatocyte growth factor. Data Brief 6:295–310
Gallant-Behm CL, Piper J, Dickinson BA, Dalby CM, Pestano LA, Jackson AL (2018) A synthetic microRNA-92a inhibitor (MRG-110) accelerates angiogenesis and wound healing in diabetic and nondiabetic wounds. Wound Repair Regen 26:311–323
Gimbrone MA Jr, Garcia-Cardena G (2016) Endothelial cell dysfunction and the pathobiology of atherosclerosis. Circ Res 118:620–636
Giral H, Kratzer A, Landmesser U (2016) MicroRNAs in lipid metabolism and atherosclerosis. Best Pract Res Clin Endocrinol Metab 30:665–676
Hu H, Luo L, Liu F, Zou D, Zhu S, Tan B, Chen T (2016) Anti-cancer and Sensibilisation effect of Triptolide on human epithelial ovarian Cancer. J Cancer 7:2093–2099
Huang M, Qiu Q, Xiao Y, Zeng S, Zhan M, Shi M, Zou Y, Ye Y, Liang L, Yang X, Xu H (2016) BET Bromodomain suppression inhibits VEGF-induced angiogenesis and vascular permeability by blocking VEGFR2-mediated activation of PAK1 and eNOS. Sci Rep 6:23770
Huang Y, Tang S, Ji-Yan C, Huang C, Li J, Cai AP, Feng YQ (2017) Circulating miR-92a expression level in patients with essential hypertension: a potential marker of atherosclerosis. J Hum Hypertens 31:200–205
Insull W Jr (2009) The pathology of atherosclerosis: plaque development and plaque responses to medical treatment. Am J Med 122:S3–s14
Kalinina N, Klink G, Glukhanyuk E, Lopatina T, Efimenko A, Akopyan Z, Tkachuk V (2015) miR-92a regulates angiogenic activity of adipose-derived mesenchymal stromal cells. Exp Cell Res 339:61–66
Lee SJ, Namkoong S, Kim YM, Kim CK, Lee H, Ha KS, Chung HT, Kwon YG, Kim YM (2006) Fractalkine stimulates angiogenesis by activating the Raf-1/MEK/ERK- and PI3K/Akt/eNOS-dependent signal pathways. Am J Physiol Heart Circ Physiol 291:H2836–H2846
Loyer X, Potteaux S, Vion AC, Guerin CL, Boulkroun S, Rautou PE, Ramkhelawon B, Esposito B, Dalloz M, Paul JL, Julia P, Maccario J, Boulanger CM, Mallat Z, Tedgui A (2014) Inhibition of microRNA-92a prevents endothelial dysfunction and atherosclerosis in mice. Circ Res 114:434–443
Lu F, Zhao LY, Zhang ZM, Zou Q, Yu XL, Wei CY (2018) The intervention of enalapril maleate and folic acid tablet on the expressions of the GRP78 and CHOP and vascular remodeling in the vascular smooth muscle cells of H-hypertensive rats with homocysteine. Eur Rev Med Pharmacol Sci 22:2160–2168
Luo L, Yang T (2016) Triptolide inhibits the progression of atherosclerosis in apolipoprotein E(−)/(−) mice. Exp Ther Med 12:2307–2313
Michel JB, Martin-Ventura JL, Nicoletti A, Ho-Tin-Noe B (2014) Pathology of human plaque vulnerability: mechanisms and consequences of intraplaque haemorrhages. Atherosclerosis 234:311–319
Munoz-Vega M, Masso F, Paez A, Carreon-Torres E, Cabrera-Fuentes HA, Fragoso JM, Perez-Hernandez N, Martinez LO, Najib S, Vargas-Alarcon G, Perez-Mendez O (2018) Characterization of immortalized human dermal microvascular endothelial cells (HMEC-1) for the study of HDL functionality. Lipids Health Dis 17:44
Murata K, Ito H, Yoshitomi H, Yamamoto K, Fukuda A, Yoshikawa J, Furu M, Ishikawa M, Shibuya H, Matsuda S (2014) Inhibition of miR-92a enhances fracture healing via promoting angiogenesis in a model of stabilized fracture in young mice. J Bone Miner Res 29:316–326
Natarajan R, Fisher BJ, Fowler AA 3rd (2003) Regulation of hypoxia inducible factor-1 by nitric oxide in contrast to hypoxia in microvascular endothelium. FEBS Lett 549:99–104
Nazari-Jahantigh M, Wei Y, Noels H, Akhtar S, Zhou Z, Koenen RR, Heyll K, Gremse F, Kiessling F, Grommes J, Weber C, Schober A (2012) MicroRNA-155 promotes atherosclerosis by repressing Bcl6 in macrophages. J Clin Invest 122:4190–4202
Nguyen LTH, Muktabar A, Tang J, Dravid VP, Thaxton CS, Venkatraman S, Ng KW (2017) Engineered nanoparticles for the detection, treatment and prevention of atherosclerosis: how close are we? Drug Discov Today 22:1438–1446
Ohyagi-Hara C, Sawada K, Kamiura S, Tomita Y, Isobe A, Hashimoto K, Kinose Y, Mabuchi S, Hisamatsu T, Takahashi T, Kumasawa K, Nagata S, Morishige K, Lengyel E, Kurachi H, Kimura T (2013) miR-92a inhibits peritoneal dissemination of ovarian cancer cells by inhibiting integrin alpha5 expression. Am J Pathol 182:1876–1889
Otsuka F, Yasuda S, Noguchi T, Ishibashi-Ueda H (2016) Pathology of coronary atherosclerosis and thrombosis. Cardiovasc Diagn Ther 6:396–408
Ridker PM, Everett BM, Thuren T, MacFadyen JG, Chang WH, Ballantyne C, Fonseca F, Nicolau J, Koenig W, Anker SD, Kastelein JJP, Cornel JH, Pais P, Pella D, Genest J, Cifkova R, Lorenzatti A, Forster T, Kobalava Z, Vida-Simiti L, Flather M, Shimokawa H, Ogawa H, Dellborg M, Rossi PRF, Troquay RPT, Libby P, Glynn RJ (2017) Antiinflammatory therapy with Canakinumab for atherosclerotic disease. N Engl J Med 377:1119–1131
Santulli G (2015) microRNAs distinctively regulate vascular smooth muscle and endothelial cells: functional implications in angiogenesis, atherosclerosis, and in-stent restenosis. Adv Exp Med Biol 887:53–77
Song JM, Molla K, Anandharaj A, Cornax I, MG OS, Kirtane AR, Panyam J, Kassie F (2017) Triptolide suppresses the in vitro and in vivo growth of lung cancer cells by targeting hyaluronan-CD44/RHAMM signaling. Oncotarget 8:26927–26940
Wei Y, Nazari-Jahantigh M, Chan L, Zhu M, Heyll K, Corbalan-Campos J, Hartmann P, Thiemann A, Weber C, Schober A (2013) The microRNA-342-5p fosters inflammatory macrophage activation through an Akt1- and microRNA-155-dependent pathway during atherosclerosis. Circulation 127:1609–1619
WHO. 2014. Global staus report on noncommunicable diseases 2014.Editor^editors: World Health Organization
Yin YL, Zhu ML, Wan J, Zhang C, Pan GP, Lu JX, Ping S, Chen Y, Zhao FR, Yu HY, Guo T, Jian X, Liu LY, Zhang JN, Wan GR, Wang SX, Li P (2017) Traditional Chinese medicine xin-mai-jia recouples endothelial nitric oxide synthase to prevent atherosclerosis in vivo. Sci Rep 7:43508
Zhang Q, Kandic I, Kutryk MJ (2011) Dysregulation of angiogenesis-related microRNAs in endothelial progenitor cells from patients with coronary artery disease. Biochem Biophys Res Commun 405:42–46
Zhou HF, Liu XY, Niu DB, Li FQ, He QH, Wang XM (2005) Triptolide protects dopaminergic neurons from inflammation-mediated damage induced by lipopolysaccharide intranigral injection. Neurobiol Dis 18:441–449
Ziaei S, Halaby R (2016) Immunosuppressive, anti-inflammatory and anti-cancer properties of triptolide: a mini review. Avicenna J Phytomed 6:149–164
This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.
Conflict of interest
The authors declare that there are no conflicts of interest.
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
1. TPL inhibits HMEC-1 cell angiogenesis;
2. TPL upregulates the expression of miR-92a;
3. TPL inhibits cell angiogenesis via upregulating miR-92a expression;
4 ITGA5 is a target of miR-92a;
5. Overexpression of ITGA5 promotes angiogenesis while ITGA5 silence inhibits angiogenesis.
About this article
Cite this article
Xu, X., Tian, L. & Zhang, Z. Triptolide inhibits angiogenesis in microvascular endothelial cells through regulation of miR-92a. J Physiol Biochem 75, 573–583 (2019). https://doi.org/10.1007/s13105-019-00707-2