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Cellular and Molecular Neurobiology

, Volume 39, Issue 2, pp 301–319 | Cite as

Myosin IIA Regulated Tight Junction in Oxygen Glucose-Deprived Brain Endothelial Cells Via Activation of TLR4/PI3K/Akt/JNK1/2/14-3-3ε/NF-κB/MMP9 Signal Transduction Pathway

  • Yanni LvEmail author
  • Wen Liu
  • Zhaohui Ruan
  • Zixuan Xu
  • Longsheng Fu
Original Research
  • 55 Downloads

Abstract

Non-muscle myosin heavy chain IIA (NMMHC IIA), a member of Myosin II family, plays a critical role in various cellular physiological processes. Our previous research had suggested that NMMHC IIA could participate in regulating tight junction morphological changes induced by ischemia stroke. Thus, in the current study, we attempted to uncover the regulation pattern of NMMHC IIA on tight junction dysfunction in oxygen glucose-deprived (OGD) mouse brain bEND.3 endothelial cells. The regulation of NMMHC IIA on tight junction in OGD-stimulated bEND.3 cells was evaluated by western blotting assay. Morphologic change of occludin, claudin-5, and ZO-1 tight junction proteins was compared with pretreatment with NMMHC II inhibitor blebbistatin via immunohistochemical staining. Detection of activation of NMMHC IIA on OGD-mediated tight junction transduction pathway was investigated via Koch’s postulate using corresponding protein inhibitor. Our results showed that NMMHC IIA was activated in OGD-stimulated bEND.3 endothelial cells. The inhibition of NMMHC IIA could attenuate the morphologic change of occludin, claudin-5, and ZO-1 tight junction proteins. NMMHC IIA participated in regulating downstream transduction pathway TLR4, phosphatidylinositol 3-kinase (PI3K), Akt, JNK1/2, 14-3-3ε, nuclear factor kappa B (NF-кB) and matrix metalloprotein 9 (MMP9). Blocking of these pathways using indicated inhibitors demonstrated that NMMHC IIA destroyed the connection of tight junction via the activation of TLR4/PI3K/Akt/JNK1/2/14-3-3ε/NF-κB/MMP9 pathway. Our study described the key role of NMMHC IIA in OGD-stimulated mouse brain bEND.3 endothelial cells, while also exhibited the molecule effect on tight junction dysfunction via TLR4/PI3K/Akt/JNK1/2/14-3-3ε/NF-κB/MMP9 signal transduction pathway.

Keywords

Non-muscle myosin heavy chain IIA Mouse brain bEND.3 endothelial cells Tight junction dysfunction TLR4/PI3K/Akt/JNK1/2/14-3-3ε/NF-κ/BMMP9 

Notes

Acknowledgements

This work was supported by Foundation Project: National Natural Science Foundation of China (81760094, 31602111, 81860020), China; Jiangxi Provincial Education Department Science and Technology Research Youth Project (No. GJJ160238); The Foundation of Jiangxi Provincial Department of Science and Technology Youth Project (No. 20171BAB215021). The funder has no role in the study design, data collection and analysis, decision to publish or preparation of the manuscript.

Author Contributions

All authors contributed to the research. YL conceived the experiments. YL, WL, ZR, ZX, LF conducted the experiments and analyzed the results. All authors contributed to the writing and editing of the manuscript.

Compliance with Ethical Standards

Conflict of interest

None of the authors have any conflicts of interest, including financial, personal or other relationships, with other individuals or organizations.

Ethical Approval

All animal care and experimental procedures were conducted according to standard ethical guidelines (National Institutes of Health Guide to the use of Laboratory Animals) and approved by Institutional Animal Care and use Committee of Nanchang University. All efforts were made to minimize the number of mice used and their suffering.

References

  1. Butt T, Mufti T, Humayun A, Rosenthal PB, Khan S, Khan S et al (2010) Myosin motors drive long range alignment of actin filaments. J Biol Chem 285:4964–4974CrossRefGoogle Scholar
  2. Butt AQ, Ahmed S, Maratha A, Miggin SM (2014) 14-3-3ε and 14-3-3σ inhibit Toll-like receptor (TLR)-mediated proinflammatory cytokine induction. J Biol Chem 287:38665–38679CrossRefGoogle Scholar
  3. Cai M, Yu Z, Wang L, Song X, Zhang J, Zhang Z et al (2016) Tongxinluo reduces brain edema and inhibits post-ischemic inflammation after middle cerebral artery occlusion in rats. J Ethnopharmacol 181:136–145CrossRefGoogle Scholar
  4. Campos Y, Qiu X, Gomero E, Wakefield R, Horner L, Brutkowski W et al (2016) Alix-mediated assembly of the actomyosin-tight junction polarity complex preserves epithelial polarity and epithelial barrier. Nat Commun 7:11876CrossRefGoogle Scholar
  5. Choi C, Kwon J, Lim S, Helfman DM (2016) Integrin β1, myosin light chain kinase and myosin IIA are required for activation of PI3K-AKT signaling following MEK inhibition in metastatic triple negative breast cancer. Oncotarget 7:63466–63487Google Scholar
  6. Clark K, Middelbeek J, Dorovkov MV, Figdor CG, Ryazanov AG, Lasonder E et al (2008) The alpha-kinases TRPM6 and TRPM7, but not eEF-2 kinase, phosphorylate the assembly domain of myosin IIA, IIB and IIC. FEBS Lett 582:2993–2997CrossRefGoogle Scholar
  7. Daquinag AC, Zhang Y, Amaya-Manzanares F, Simmons PJ, Kolonin MG (2011) An isoform of decorin is a resistin receptor on the surface of adipose progenitor cells. Cell Stem Cell 9:2011Google Scholar
  8. Funami K, Matsumoto M, Obuse C, Seya T (2016) 14-3-3-zeta participates in TLR3-mediated TICAM-1 signal-platform formation. Mol Immunol 73:60–68CrossRefGoogle Scholar
  9. Garbuzova-Davis S, Mirtyl S, Sallot SA, Hernandez-Ontiveros DG, Haller E, Sanberg PR (2013) Blood-brain barrier impairment in MPS III patients. BMC Neurol 13:174CrossRefGoogle Scholar
  10. Greene C, Campbell M (2016) Tight junction modulation of the blood brain barrier: CNS delivery of small molecules. Tissue Barriers 4:e1138017CrossRefGoogle Scholar
  11. Ivanov AI, Bachar M, Babbin BA, Adelstein RS, Nusrat A, Parkos CA (2007) A unique role for nonmuscle myosin heavy chain IIA in regulation of epithelial apical junctions. PLoS ONE 2:e658CrossRefGoogle Scholar
  12. Kim T (2015) Determinants of contractile forces generated in disorganized actomyosin bundles. Biomech Model Mechanobiol 14:345–355CrossRefGoogle Scholar
  13. Kolega J (2006) The role of myosin II motor activity in distributing myosin asymmetrically and coupling protrusive activity to cell translocation. Mol Biol Cell 17:4435–4445CrossRefGoogle Scholar
  14. Kovács M, Tóth J, Hetényi C, Málnási-Csizmadia A, Sellers JR (2004) Mechanism of blebbistatin inhibition of myosin II. J Biol Chem 279:35557–35563CrossRefGoogle Scholar
  15. Liu W, Hendren J, Qin X, Shen J, Liu KJ (2009) Normobaric hyperoxia attenuates early blood-brain barrier disruption by inhibiting MMP-9-mediated occludin degradation in focal cerebral ischemia. J Neurochem 108:811–820CrossRefGoogle Scholar
  16. Liu WY, Wang ZB, Zhang LC, Wei X, Li L (2012) Tight junction in blood-brain barrier: an overview of structure, regulation, and regulator substances. CNS Neurosci Ther 18(8):609–615CrossRefGoogle Scholar
  17. Liu T, Ye Y, Zhang X, Zhu A, Yang Z, Fu Y et al (2016) Downregulation of non-muscle myosin IIA expression inhibits migration and invasion of gastric cancer cells via the c-Jun N-terminal kinase signaling pathway. Mol Med Rep 13:1639–1644CrossRefGoogle Scholar
  18. Lu P, Chen J, Yan L, Yang L, Zhang L, Dai J, Hao Z, Bai T, Xi Y, Li Y, Kang Z, Xv J, Sun G, Yang T (2018) RasGRF2 promotes migration and invasion of colorectal cancer cells by modulating expression of MMP9 through Src/Akt/NF-κB pathway. Cancer Biol Ther 10:1–9CrossRefGoogle Scholar
  19. Luissint AC, Artus C, Glacial F, Ganeshamoorthy K, Couraud PO (2012) Tight junctions at the blood brain barrier: physiological architecture and disease-associated dysregulation. Fluids Barriers CNS 9:23CrossRefGoogle Scholar
  20. Lv Y, Fu L (2018) The potential mechanism for Hydroxysafflor yellow A attenuating blood-brain barrier dysfunction via tight junction signaling pathways excavated by an integrated serial affinity chromatography and shotgun proteomics analysis approach. Neurochem Int 112:38–48CrossRefGoogle Scholar
  21. Lv Y, Qian Y, Fu L, Chen X, Zhong H, Wei X (2015) Hydroxysafflor yellow A exerts neuroprotective effects in cerebral ischemia reperfusion-injured mice by suppressing the innate immune TLR4-inducing pathway. Eur J Pharmacol 769:324–332CrossRefGoogle Scholar
  22. Mashukova A, Wald FA, Salas PJ (2011) Tumor necrosis factor alpha and inflammation disrupt the polarity complex in intestinal epithelial cells by a posttranslational mechanism. Mol Cell Biol 31:756–765CrossRefGoogle Scholar
  23. Naydenov NG, Feygin A, Wang D, Kuemmerle JF, Harris G, Conti MA et al (2016) Nonmuscle Myosin IIA Regulates Intestinal Epithelial Barrier in vivo and Plays a Protective Role During Experimental Colitis. Sci Rep 6:24161CrossRefGoogle Scholar
  24. Neal CL, Xu J, Li P, Mori S, Yang J, Neal NN et al (2012) Overexpression of 14-3-3ζ in cancer cells activates PI3K via binding the p85 regulatory subunit. Oncogene 31:897–906CrossRefGoogle Scholar
  25. Nomura M, Shimizu S, Sugiyama T, Narita M, Ito T, Matsuda H et al (2003) 14-3-3 Interacts directly with and negatively regulates pro-apoptotic Bax. J Biol Chem 278:2058–2065CrossRefGoogle Scholar
  26. Pozuelo-Rubio M (2012) 14-3-3 Proteins are Regulators of Autophagy. Cells 1:754–773CrossRefGoogle Scholar
  27. Schuster TB, Costina V, Findeisen P, Neumaier M, Ahmad-Nejad P (2011) Identification and functional characterization of 14-3-3 in TLR2 signaling. J Proteome Res 10:4661–4670CrossRefGoogle Scholar
  28. Stamatovic SM, Johnson AM, Keep RF, Andjelkovic AV (2016) Junctional proteins of the blood-brain barrier: new insights into function and dysfunction. Tissue Barriers 4:e1154641CrossRefGoogle Scholar
  29. Turner JR (2006) Molecular basis of epithelial barrier regulation: from basic mechanisms to clinical application. Am J Pathol 169:1901–1909CrossRefGoogle Scholar
  30. Utech M, Ivanov AI, Samarin SN, Bruewer M, Turner JR, Mrsny RJ et al (2005) Mechanism of IFN-gamma-induced endocytosis of tight junction proteins: myosin II-dependent vacuolarization of the apical plasma membrane. Mol Biol Cell 16:5040–5052CrossRefGoogle Scholar
  31. Wang XT, Pei DS, Xu J, Guan QH, Sun YF, Liu XM et al (2007) Opposing effects of Bad phosphorylation at two distinct sites by Akt1 and JNK1/2 on ischemic brain injury. Cell Signal 19:1844–1856CrossRefGoogle Scholar
  32. Wang Y, Liu Q, Xu Y, Zhang Y, Lv Y, Tan Y et al (2016a) Ginsenoside Rg1 Protects against Oxidative Stress-induced Neuronal Apoptosis through Myosin IIA-actin Related Cytoskeletal Reorganization. Int J Biol Sci 12:1341–1356CrossRefGoogle Scholar
  33. Wang ZG, Cheng Y, Yu XC, Ye LB, Xia QH, Johnson NR et al (2016b) bFGF Protects Against Blood-Brain Barrier Damage Through Junction Protein Regulation via PI3K-Akt-Rac1 Pathway Following Traumatic Brain Injury. Mol Neurobiol 53:7298–7311CrossRefGoogle Scholar
  34. Wen H, Watry DD, Marcondes MC, Fox HS (2004) Selective decrease in paracellular conductance of tight junctions: role of the first extracellular domain of claudin-5. Mol Cell Biol 24:8408–8417CrossRefGoogle Scholar
  35. Wickman GR, Julian L, Mardilovich K, Schumacher S, Munro J, Rath N et al (2013) Blebs produced by actin-myosin contraction during apoptosis release damage-associated molecular pattern proteins before secondary necrosis occurs. Cell Death Differ 20:1293–1305CrossRefGoogle Scholar
  36. Wu JS, Cheung WM, Tsai YS, Chen YT, Fong WH, Tsai HD et al (2009) Ligand-activated peroxisome proliferator-activated receptor-gamma protects against ischemic cerebral infarction and neuronal apoptosis by 14-3-3 epsilon upregulation. Circulation 119:1124–1134CrossRefGoogle Scholar
  37. Zhai K, Tang Y, Zhang Y, Li F, Wang Y, Cao Z et al (2015) NMMHC IIA inhibition impedes tissue factor expression and venous thrombosis via Akt/GSK3β-NF-κB signalling pathways in the endothelium. Thromb Haemost 114:173CrossRefGoogle Scholar
  38. Zhaocheng J, Jinfeng L, Luchang Y, Yequan S, Feng L, Kai W (2016) Ginkgolide A inhibits lipopolysaccharide-induced inflammatory response in human coronary artery endothelial cells via downregulation of TLR4-NF-κB signaling through PI3K/Akt pathway. Pharmazie 71(10):588–591Google Scholar
  39. Zhou C, Wang Y, Peng J, Li C, Liu P, Shen X (2017) SNX10 Plays a Critical Role in MMP9 Secretion via JNK-p38-ERK Signaling Pathway. J Cell Biochem 118:4664–4671CrossRefGoogle Scholar
  40. Zhu Y, Bu Q, Liu X, Hu W, Wang Y (2014) Neuroprotective effect of TAT-14-3-3e fusion protein against cerebral ischemia/reperfusion injury in rats. PLoS ONE 9:e93334CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Pharmacy departmentThe First Affiliated Hospital of Nanchang UniversityNanchangChina
  2. 2.Jiangxi Provincial Institute of Traditional Chinese MedicineNanchangChina

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