Microglia-mediated neuroinflammation plays an important role in focal ischemic stroke, a disorder with no effective therapeutic agents. Since microglial polarization to the M2 phenotype and reduction of oxidative stress are mediated through AMP-activated protein kinase (AMPK) and nuclear factor erythroid 2-related factor 2 (Nrf2) activation, we assessed the dual therapeutic effect of AMPK and Nrf2 activation by a novel neuroprotectant HP-1c in the treatment of ischemic stroke.
Results:
We developed a novel class of hybrids (HP-1a–HP-1f) of telmisartan and 2-(1-hydroxypentyl)-benzoate (HPBA) as a ring-opening derivative of NBP. The most promising hybrid, HP-1c, exhibited more potent anti-inflammatory and neuroprotective effects in vitro and reduced brain infarct volume and improved neurological deficits in a rat model of transient focal cerebral ischemia when compared with telmisartan alone, NBP alone, or a combination of telmisartan and NBP. HP-1c had a therapeutic window of up to 24 h, ameliorated ischemic cerebral injury in permanent focal cerebral ischemia, and improved motor function. The beneficial effects of HP-1c in ischemic stroke were associated with microglial polarization to the M2 phenotype and reduced oxidative stress. HP-1c also shifted the M1/M2 polarization in a mouse neuroinflammatory model. The anti-inflammatory and anti-oxidative effects of HP-1c were associated with AMPK-Nrf2 pathway activation for neuroprotection. We showed that HP-1c penetrates the brain, has a plasma half-life of around 3.93 h, and has no toxicity in mice.
Innovation and Conclusion:
Our study results suggest that HP-1c, with dual AMPK- and Nrf2-activating properties, may have potential in further studies as a novel therapy for ischemic stroke. Antioxid. Redox Signal. 28, 141–163.
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References
1.
AlfieriA, SrivastavaS, SiowRC, ModoM, FraserPA, and MannGE. Targeting the Nrf2-Keap1 antioxidant defence pathway for neurovascular protection in stroke. J Physiol, 589: 4125–4136, 2011.
2.
Barry-LanePA, PattersonC, van der MerweM, HuZ, HollandSM, YehET, and RungeMS. p47phox is required for atherosclerotic lesion progression in ApoE(-/-) mice. J Clin Invest, 108: 1513–1522, 2001.
BruneB, DehneN, GrossmannN, JungM, NamgaladzeD, SchmidT, von KnethenA, and WeigertA. Redox control of inflammation in macrophages. Antioxid Redox Signal, 19: 595–637, 2013.
5.
ChangS, RuanWC, XuYZ, WangYJ, PangJ, ZhangLY, LiaoH, and PangT. The natural product 4,10-aromadendranediol induces neuritogenesis in neuronal cells in vitro through activation of the ERK pathway. Acta Pharmacol Sin, 38: 29–40, 2017.
6.
ChenS, WenX, ZhangW, WangC, LiuJ, and LiuC. Hypolipidemic effect of oleanolic acid is mediated by the miR-98-5p/PGC-1beta axis in high-fat diet-induced hyperlipidemic mice. FASEB J, 31: 1085–1096, 2017.
7.
de SouzaLF, BarretoF, da SilvaEG, AndradesME, GuimaraesEL, BehrGA, MoreiraJC, and BernardEA. Regulation of LPS stimulated ROS production in peritoneal macrophages from alloxan-induced diabetic rats: involvement of high glucose and PPARgamma. Life Sci, 81: 153–159, 2007.
8.
ElkahlounAG, HafkoR, and SaavedraJM. An integrative genome-wide transcriptome reveals that candesartan is neuroprotective and a candidate therapeutic for Alzheimer's disease. Alzheimers Res Ther, 8: 5, 2016.
9.
FengX, PengY, LiuM, and CuiL. DL-3-n-butylphthalide extends survival by attenuating glial activation in a mouse model of amyotrophic lateral sclerosis. Neuropharmacology, 62: 1004–1010, 2012.
10.
FerminiB and FossaAA. The impact of drug-induced QT interval prolongation on drug discovery and development. Nat Rev Drug Discov, 2: 439–447, 2003.
11.
GaoY, XuX, ChangS, WangY, XuY, RanS, HuangZ, LiP, LiJ, ZhangL, SaavedraJM, LiaoH, and PangT. Totarol prevents neuronal injury in vitro and ameliorates brain ischemic stroke: potential roles of Akt activation and HO-1 induction. Toxicol Appl Pharmacol, 289: 142–154, 2015.
12.
GiriS, NathN, SmithB, ViolletB, SinghAK, and SinghI. 5-aminoimidazole-4-carboxamide-1-beta-4-ribofuranoside inhibits proinflammatory response in glial cells: a possible role of AMP-activated protein kinase. J Neurosci, 24: 479–487, 2004.
13.
GreenMF, AndersonKA, and MeansAR. Characterization of the CaMKKbeta-AMPK signaling complex. Cell Signal, 23: 2005–2012, 2011.
14.
HennA, LundS, HedtjarnM, SchrattenholzA, PorzgenP, and LeistM. The suitability of BV2 cells as alternative model system for primary microglia cultures or for animal experiments examining brain inflammation. ALTEX, 26: 83–94, 2009.
15.
HuX, LeakRK, ShiY, SuenagaJ, GaoY, ZhengP, and ChenJ. Microglial and macrophage polarization-new prospects for brain repair. Nat Rev Neurol, 11: 56–64, 2015.
16.
HuX, LiP, GuoY, WangH, LeakRK, ChenS, GaoY, and ChenJ. Microglia/macrophage polarization dynamics reveal novel mechanism of injury expansion after focal cerebral ischemia. Stroke, 43: 3063–3070, 2012.
17.
HuaK, ShengX, LiTT, WangLN, ZhangYH, HuangZJ, and JiH. The edaravone and 3-n-butylphthalide ring-opening derivative 10b effectively attenuates cerebral ischemia injury in rats. Acta Pharmacol Sin, 36: 917–927, 2015.
18.
ItoT, YamakawaH, BregonzioC, TerronJA, Falcon-NeriA, and SaavedraJM. Protection against ischemia and improvement of cerebral blood flow in genetically hypertensive rats by chronic pretreatment with an angiotensin II AT1 antagonist. Stroke, 33: 2297–2303, 2002.
19.
JiangT, YuJT, ZhuXC, WangHF, TanMS, CaoL, ZhangQQ, GaoL, ShiJQ, ZhangYD, and TanL. Acute metformin preconditioning confers neuroprotection against focal cerebral ischaemia by pre-activation of AMPK-dependent autophagy. Br J Pharmacol, 171: 3146–3157, 2014.
20.
JinQ, ChengJ, LiuY, WuJ, WangX, WeiS, ZhouX, QinZ, JiaJ, and ZhenX. Improvement of functional recovery by chronic metformin treatment is associated with enhanced alternative activation of microglia/macrophages and increased angiogenesis and neurogenesis following experimental stroke. Brain Behav Immun, 40: 131–142, 2014.
21.
JinR, YangG, and LiG. Inflammatory mechanisms in ischemic stroke: role of inflammatory cells. J Leukoc Biol, 87: 779–789, 2010.
22.
JinW, WangH, YanW, XuL, WangX, ZhaoX, YangX, ChenG, and JiY. Disruption of Nrf2 enhances upregulation of nuclear factor-kappaB activity, proinflammatory cytokines, and intercellular adhesion molecule-1 in the brain after traumatic brain injury. Mediators Inflamm, 2008: 725174, 2008.
23.
JooMS, KimWD, LeeKY, KimJH, KooJH, and KimSG. AMPK facilitates nuclear accumulation of Nrf2 by phosphorylating at Serine 550. Mol Cell Biol, 36: 1931–1942, 2016.
24.
KangKW, LeeSJ, and KimSG. Molecular mechanism of nrf2 activation by oxidative stress. Antioxid Redox Signal, 7: 1664–1673, 2005.
25.
Keleku-LukweteN, SuzukiM, OtsukiA, TsuchidaK, KatayamaS, HayashiM, NaganumaE, MoriguchiT, TanabeO, EngelJD, ImaizumiM, and YamamotoM. Amelioration of inflammation and tissue damage in sickle cell model mice by Nrf2 activation. Proc Natl Acad Sci U S A, 112: 12169–12174, 2015.
26.
KurlandDB, GerzanichV, KarimyJK, WooSK, VennekensR, FreichelM, NiliusB, BryanJ, and SimardJM. The Sur1-Trpm4 channel regulates NOS2 transcription in TLR4-activated microglia. J Neuroinflammation, 13: 130, 2016.
27.
LeiteHR, Oliveira-LimaOC, Pereira LdeM, OliveiraVE, PradoVF, PradoMA, PereiraGS, and MassensiniAR. Vesicular acetylcholine transporter knock down-mice are more susceptible to inflammation, c-Fos expression and sickness behavior induced by lipopolysaccharide. Brain Behav Immun, 57: 282–292, 2016.
28.
LiC, WangJ, FangY, LiuY, ChenT, SunH, ZhouXF, and LiaoH. Nafamostat mesilate improves function recovery after stroke by inhibiting neuroinflammation in rats. Brain Behav Immun, 56: 230–245, 2016.
29.
LiL, ZhangB, TaoY, WangY, WeiH, ZhaoJ, HuangR, and PeiZ. DL-3-n-butylphthalide protects endothelial cells against oxidative/nitrosative stress, mitochondrial damage and subsequent cell death after oxygen glucose deprivation in vitro. Brain Res, 1290: 91–101, 2009.
30.
LinHY, HuangBR, YehWL, LeeCH, HuangSS, LaiCH, LinH, and LuDY. Antineuroinflammatory effects of lycopene via activation of adenosine monophosphate-activated protein kinase-alpha1/heme oxygenase-1 pathways. Neurobiol Aging, 35: 191–202, 2014.
31.
LiuCL, LiaoSJ, ZengJS, LinJW, LiCX, XieLC, ShiXG, and HuangRX. dl-3n-butylphthalide prevents stroke via improvement of cerebral microvessels in RHRSP. J Neurol Sci, 260: 106–113, 2007.
32.
LiuX, LiuJ, ZhaoS, ZhangH, CaiW, CaiM, JiX, LeakRK, GaoY, ChenJ, and HuX. Interleukin-4 is essential for microglia/macrophage M2 polarization and long-term recovery after cerebral ischemia. Stroke, 47: 498–504, 2016.
33.
LiuXM, PeytonKJ, ShebibAR, WangH, KorthuisRJ, and DuranteW. Activation of AMPK stimulates heme oxygenase-1 gene expression and human endothelial cell survival. Am J Physiol Heart Circ Physiol, 300: H84–H93, 2011.
34.
LongaEZ, WeinsteinPR, CarlsonS, and CumminsR. Reversible middle cerebral artery occlusion without craniectomy in rats. Stroke, 20: 84–91, 1989.
35.
LuDY, TangCH, ChenYH, and WeiIH. Berberine suppresses neuroinflammatory responses through AMP-activated protein kinase activation in BV-2 microglia. J Cell Biochem, 110: 697–705, 2010.
36.
MacleodMR, FisherM, O'CollinsV, SenaES, DirnaglU, BathPM, BuchanA, van der WorpHB, TraystmanRJ, MinematsuK, DonnanGA, and HowellsDW. Reprint: good laboratory practice: preventing introduction of bias at the bench. Int J Stroke, 4: 3–5, 2009.
37.
MakarTK, GerzanichV, NimmagaddaVK, JainR, LamK, MubarizF, TrislerD, IvanovaS, WooSK, KwonMS, BryanJ, BeverCT, and SimardJM. Silencing of Abcc8 or inhibition of newly upregulated Sur1-Trpm4 reduce inflammation and disease progression in experimental autoimmune encephalomyelitis. J Neuroinflammation, 12: 210, 2015.
38.
MoC, WangL, ZhangJ, NumazawaS, TangH, TangX, HanX, LiJ, YangM, WangZ, WeiD, and XiaoH. The crosstalk between Nrf2 and AMPK signal pathways is important for the anti-inflammatory effect of berberine in LPS-stimulated macrophages and endotoxin-shocked mice. Antioxid Redox Signal, 20: 574–588, 2014.
39.
O'NeillLA and HardieDG. Metabolism of inflammation limited by AMPK and pseudo-starvation. Nature, 493: 346–355, 2013.
40.
PangT, BenickyJ, WangJ, OrecnaM, Sanchez-LemusE, and SaavedraJM. Telmisartan ameliorates lipopolysaccharide-induced innate immune response through peroxisome proliferator-activated receptor-gamma activation in human monocytes. J Hypertens, 30: 87–96, 2012.
41.
PangT, WangYJ, GaoYX, XuY, LiQ, ZhouYB, XuL, HuangZJ, LiaoH, ZhangLY, GaoJR, YeQ, and LiJ. A novel GSK-3beta inhibitor YQ138 prevents neuronal injury induced by glutamate and brain ischemia through activation of the Nrf2 signaling pathway. Acta Pharmacol Sin, 37: 741–752, 2016.
42.
RemuzziA, SangalliF, MacconiD, TomasoniS, CattaneoI, RizzoP, BonandriniB, BrescianiE, LongarettiL, GagliardiniE, ContiS, BenigniA, and RemuzziG. Regression of renal disease by angiotensin II antagonism is caused by regeneration of kidney vasculature. J Am Soc Nephrol, 27: 699–705, 2016.
43.
SaavedraJM. Angiotensin II AT(1) receptor blockers as treatments for inflammatory brain disorders. Clin Sci (Lond), 123: 567–590, 2012.
44.
SaavedraJM. Evidence to consider angiotensin II receptor blockers for the treatment of early Alzheimer's disease. Cell Mol Neurobiol, 36: 259–279, 2016.
45.
SagD, CarlingD, StoutRD, and SuttlesJ. Adenosine 5'-monophosphate-activated protein kinase promotes macrophage polarization to an anti-inflammatory functional phenotype. J Immunol, 181: 8633–8641, 2008.
46.
SandbergM, PatilJ, D'AngeloB, WeberSG, and MallardC. NRF2-regulation in brain health and disease: implication of cerebral inflammation. Neuropharmacology, 79: 298–306, 2014.
47.
SatohT, McKercherSR, and LiptonSA. Nrf2/ARE-mediated antioxidant actions of pro-electrophilic drugs. Free Radic Biol Med, 65: 645–657, 2013.
48.
SmirnovaNA, Haskew-LaytonRE, BassoM, HushpulianDM, PayappillyJB, SpeerRE, AhnYH, RakhmanI, ColePA, PintoJT, RatanRR, and GazaryanIG. Development of Neh2-luciferase reporter and its application for high throughput screening and real-time monitoring of Nrf2 activators. Chem Biol, 18: 752–765, 2011.
49.
SonTG, CamandolaS, ArumugamTV, CutlerRG, TelljohannRS, MughalMR, MooreTA, LuoW, YuQS, JohnsonDA, JohnsonJA, GreigNH, and MattsonMP. Plumbagin, a novel Nrf2/ARE activator, protects against cerebral ischemia. J Neurochem, 112: 1316–1326, 2010.
50.
SurhYJ. NF-kappa B and Nrf2 as potential chemopreventive targets of some anti-inflammatory and antioxidative phytonutrients with anti-inflammatory and antioxidative activities. Asia Pac J Clin Nutr, 17Suppl 1: 269–272, 2008.
WangCY, WangZY, XieJW, WangT, WangX, XuY, and CaiJH. Dl-3-n-butylphthalide-induced upregulation of antioxidant defense is involved in the enhancement of cross talk between CREB and Nrf2 in an Alzheimer's disease mouse model. Neurobiol Aging, 38: 32–46, 2016.
53.
WangF, MaJ, HanF, GuoX, MengL, SunY, JinC, DuanH, LiH, and PengY. DL-3-n-butylphthalide delays the onset and progression of diabetic cataract by inhibiting oxidative stress in rat diabetic model. Sci Rep, 6: 19396, 2016.
54.
WangG, ShiY, JiangX, LeakRK, HuX, WuY, PuH, LiWW, TangB, WangY, GaoY, ZhengP, BennettMV, and ChenJ. HDAC inhibition prevents white matter injury by modulating microglia/macrophage polarization through the GSK3beta/PTEN/Akt axis. Proc Natl Acad Sci U S A, 112: 2853–2858, 2015.
55.
WangH, KhorTO, SawCL, LinW, WuT, HuangY, and KongAN. Role of Nrf2 in suppressing LPS-induced inflammation in mouse peritoneal macrophages by polyunsaturated fatty acids docosahexaenoic acid and eicosapentaenoic acid. Mol Pharm, 7: 2185–2193, 2010.
56.
WangX, LiY, ZhaoQ, MinZ, ZhangC, LaiY, JiH, PengS, and ZhangY. Design, synthesis and evaluation of nitric oxide releasing derivatives of 3-n-butylphthalide as antiplatelet and antithrombotic agents. Org Biomol Chem, 9: 5670–5681, 2011.
57.
WangX, WangL, LiT, HuangZ, LaiY, JiH, WanX, XuJ, TianJ, and ZhangY. Novel hybrids of optically active ring-opened 3-n-butylphthalide derivative and isosorbide as potential anti-ischemic stroke agents. J Med Chem, 56: 3078–3089, 2013.
58.
WangX, ZhaoQ, WangX, LiT, LaiY, PengS, JiH, XuJ, and ZhangY. Studies on the enantiomers of ZJM-289: synthesis and biological evaluation of antiplatelet, antithrombotic and neuroprotective activities. Org Biomol Chem, 10: 9030–9040, 2012.
59.
XiongXY, LiuL, and YangQW. Functions and mechanisms of microglia/macrophages in neuroinflammation and neurogenesis after stroke. Prog Neurobiol, 142: 23–44, 2016.
60.
XuM, YangL, HongLZ, ZhaoXY, and ZhangHL. Direct protection of neurons and astrocytes by matrine via inhibition of the NF-kappaB signaling pathway contributes to neuroprotection against focal cerebral ischemia. Brain Res, 1454: 48–64, 2012.
61.
XuY, WangY, XuY, LiJ, LiaoH, ZhangL, and PangT. Development of a novel phosphorylated AMPK protection assay for high-throughput screening using TR-FRET assay. J Biomol Screen, 20: 906–912, 2015.
XueLX, ZhangT, ZhaoYW, GengZ, ChenJJ, and ChenH. Efficacy and safety comparison of DL-3-n-butylphthalide and Cerebrolysin: effects on neurological and behavioral outcomes in acute ischemic stroke. Exp Ther Med, 11: 2015–2020, 2016.
64.
YagiK, KitazatoKT, UnoM, TadaY, KinouchiT, ShimadaK, and NagahiroS. Edaravone, a free radical scavenger, inhibits MMP-9-related brain hemorrhage in rats treated with tissue plasminogen activator. Stroke, 40: 626–631, 2009.
65.
YoshidaH, YanaiH, NamikiY, Fukatsu-SasakiK, FurutaniN, and TadaN. Neuroprotective effects of edaravone: a novel free radical scavenger in cerebrovascular injury. CNS Drug Rev, 12: 9–20, 2006.
66.
ZangY, YuLF, PangT, FangLP, FengX, WenTQ, NanFJ, FengLY, and LiJ. AICAR induces astroglial differentiation of neural stem cells via activating the JAK/STAT3 pathway independently of AMP-activated protein kinase. J Biol Chem, 283: 6201–6208, 2008.
67.
ZhangBB, ZhouG, and LiC. AMPK: an emerging drug target for diabetes and the metabolic syndrome. Cell Metab, 9: 407–416, 2009.
68.
ZhangR, XuM, WangY, XieF, ZhangG, and QinX. Nrf2-a promising therapeutic target for defensing against oxidative stress in stroke. Mol Neurobiol, 2016.
69.
ZhouX, CaoY, AoG, HuL, LiuH, WuJ, WangX, JinM, ZhengS, ZhenX, AlkayedNJ, JiaJ, and ChengJ. CaMKKbeta-dependent activation of AMP-activated protein kinase is critical to suppressive effects of hydrogen sulfide on neuroinflammation. Antioxid Redox Signal, 21: 1741–1758, 2014.
70.
ZhuYP, BrownJR, SagD, ZhangL, and SuttlesJ. Adenosine 5'-monophosphate-activated protein kinase regulates IL-10-mediated anti-inflammatory signaling pathways in macrophages. J Immunol, 194: 584–594, 2015.
71.
ZimmermannK, BaldingerJ, MayerhoferB, AtanasovAG, DirschVM, and HeissEH. Activated AMPK boosts the Nrf2/HO-1 signaling axis—a role for the unfolded protein response. Free Radic Biol Med, 88: 417–426, 2015.
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