Abstract
Summary
Activated acid-sensing ion channel 1a (ASIC1a) is involved in acid-induced osteoclastogenesis by regulating activation of the transcription factor NFATc1. These results indicated that ASIC1a activation by extracellular acid may cause osteoclast migration and adhesion through Ca2+-dependent integrin/Pyk2/Src signaling pathway.
Introduction
Osteoclast adhesion and migration are responsible for osteoporotic bone loss. Acidic conditions promote osteoclastogenesis. ASIC1a in osteoclasts is associated with acid-induced osteoclastogenesis through modulating transcription factor NFATc1 activation. However, the influence and the detailed mechanism of ASIC1a in regulating osteoclast adhesion and migration, in response to extracellular acid, are not well characterized.
Methods
In this study, knockdown of ASIC1a was achieved in bone marrow macrophage cells using small interfering RNA (siRNA). The adhesion and migration abilities of osteoclast precursors and osteoclasts were determined by adhesion and migration assays, in vitro. Bone resorption was performed to measure osteoclast function. Cytoskeletal changes were assessed by F-actin ring formation. αvβ3 integrin expression in osteoclasts was measured by flow cytometry. Western blotting and co-immunoprecipitation were performed to measure alterations in integrin/Pyk2/Src signaling pathway.
Results
Our results showed that blockade of ASIC1a using ASIC1a-siRNA inhibited acid-induced osteoclast precursor migration and adhesion, as well as osteoclast adhesion and bone resorption; we also demonstrated that inhibition of ASIC1a decreased the cell surface αvβ3 integrin and β3 protein expression. Moreover, blocking of ASIC1a inhibited acidosis-induced actin ring formation and reduced Pyk2 and Src phosphorylation in osteoclasts and also inhibited the acid-induced association of the αvβ3 integrin/Src/Pyk2.
Conclusion
Together, these results highlight a key functional role of ASIC1a/αvβ3 integrin/Pyk2/Src signaling pathway in migration and adhesion of osteoclasts.
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References
Cosman F, de Beur SJ, LeBoff MS, Lewiecki EM, Tanner B, Randall S, Lindsay R (2014) Clinician’s guide to prevention and treatment of osteoporosis. Osteoporos Int 25:2359–2381
Cooper C, Campion G, Melton LJ 3rd (1992) Hip fractures in the elderly: a world-wide projection. Osteoporos Int 2:285–289
Udagawa N, Takahashi N, Jimi E et al (1999) Osteoblasts/stromal cells stimulate osteoclast activation through expression of osteoclast differentiation factor/RANKL but not macrophage colony-stimulating factor: receptor activator of NF-kappa B ligand. Bone 25:517–523
Brazier H, Pawlak G, Vives V, Blangy A (2009) The Rho GTPase Wrch1 regulates osteoclast precursor adhesion and migration. Int J Biochem Cell Biol 41:1391–1401
Yuan FL, Xu MH, Li X, Xinlong H, Fang W, Dong J (2016) The roles of acidosis in osteoclast biology. Front Physiol 7:222
Kikuta J, Kawamura S, Okiji F, Shirazaki M, Sakai S, Saito H, Ishii M (2013) Sphingosine-1-phosphate-mediated osteoclast precursor monocyte migration is a critical point of control in antibone-resorptive action of active vitamin D. Proc Natl Acad Sci U S A 110:7009–7013
Arnett TR (2010) Acidosis, hypoxia and bone. Arch Biochem Biophys 503:103–109
Komarova SV, Pereverzev A, Shum JW, Sims SM, Dixon SJ (2005) Convergent signaling by acidosis and receptor activator of NF-kappaB ligand (RANKL) on the calcium/calcineurin/NFAT pathway in osteoclasts. Proc Natl Acad Sci U S A 102:2643–2648
Waldmann R, Champigny G, Bassilana F, Heurteaux C, Lazdunski M (1997) A proton-gated cation channel involved in acid-sensing. Nature 386:173–177
Xiong ZG, Zhu XM, Chu XP et al (2004) Neuroprotection in ischemia: blocking calcium-permeable acid-sensing ion channels. Cell 118:687–698
Xiong ZG, Chu XP, Simon RP (2006) Ca2+-permeable acid-sensing ion channels and ischemic brain injury. J Membr Biol 209:59–68
Hey JG, Chu XP, Seeds J, Simon RP, Xiong ZG (2007) Extracellular zinc protects against acidosis-induced injury of cells expressing Ca2+-permeable acid-sensing ion channels. Stroke 38:670–673
Duan B, Wu LJ, Yu YQ, Ding Y, Jing L, Xu L, Chen J, Xu TL (2007) Upregulation of acid-sensing ion channel ASIC1a in spinal dorsal horn neurons contributes to inflammatory pain hypersensitivity. J Neurosci 27:11139–11148
Weng XC, Zheng JQ, Li J, Xiao WB (2007) Underlying mechanism of ASIC1a involved in acidosis-induced cytotoxicity in rat C6 glioma cells. Acta Pharmacol Sin 28:1731–1736
Yuan FL, Zhao MD, Jiang DL, Jin C, Liu HF, Xu MH, Hu W, Li X (2016) Involvement of acid-sensing ion channel 1a in matrix metabolism of endplate chondrocytes under extracellular acidic conditions through NF-kappaB transcriptional activity. Cell Stress Chaperones 21:97–104
Li X, Wu FR, Xu RS, Hu W, Jiang DL, Ji C, Chen FH, Yuan FL (2014) Acid-sensing ion channel 1a-mediated calcium influx regulates apoptosis of endplate chondrocytes in intervertebral discs. Expert Opin Ther Targets 18:1–14
Li X, Xu RS, Jiang DL, He XL, Jin C, Lu WG, Su Q, Yuan FL (2013) Acid-sensing ion channel 1a is involved in acid-induced osteoclastogenesis by regulating activation of the transcription factor NFATc1. FEBS Lett 587:3236–3242
He Y, Staser K, Rhodes SD et al (2011) Erk1 positively regulates osteoclast differentiation and bone resorptive activity. PLoS One 6:e24780
Jansen ID, Vermeer JA, Bloemen V, Stap J, Everts V (2012) Osteoclast fusion and fission. Calcif Tissue Int 90:515–522
Ahn H, Kim JM, Lee K, Kim H, Jeong D (2012) Extracellular acidosis accelerates bone resorption by enhancing osteoclast survival, adhesion, and migration. Biochem Biophys Res Commun 418:144–148
Nakamura I, Duong LT, Rodan SB, Rodan GA (2007) Involvement of alpha(v)beta3 integrins in osteoclast function. J Bone Miner Metab 25:337–344
Faccio R, Teitelbaum SL, Fujikawa K, Chappel J, Zallone A, Tybulewicz VL, Ross FP, Swat W (2005) Vav3 regulates osteoclast function and bone mass. Nat Med 11:284–290
Novack DV, Faccio R (2011) Osteoclast motility: putting the brakes on bone resorption. Ageing Res Rev 10:54–61
Tomomura M, Hasegawa H, Suda N, Sakagami H, Tomomura A (2012) Serum calcium-decreasing factor, caldecrin, inhibits receptor activator of NF-kappaB ligand (RANKL)-mediated Ca2+ signaling and actin ring formation in mature osteoclasts via suppression of Src signaling pathway. J Biol Chem 287:17963–17974
Duong LT, Lakkakorpi P, Nakamura I, Rodan GA (2000) Integrins and signaling in osteoclast function. Matrix Biol 19:97–105
Arnett TR (2008) Extracellular pH regulates bone cell function. J Nutr 138:415S–418S
Pereverzev A, Komarova SV, Korcok J, Armstrong S, Tremblay GB, Dixon SJ, Sims SM (2008) Extracellular acidification enhances osteoclast survival through an NFAT-independent, protein kinase C-dependent pathway. Bone 42:150–161
Takeuchi S, Hirukawa K, Togari A (2013) Acidosis inhibits mineralization in human osteoblasts. Calcif Tissue Int 93:233–240
Hu W, Chen FH, Yuan FL, Zhang TY, Wu FR, Rong C, Jiang S, Tang J, Zhang CC, Lin MY (2012) Blockade of acid-sensing ion channels protects articular chondrocytes from acid-induced apoptotic injury. Inflamm Res 61:327–335
Rong C, Chen FH, Jiang S, Hu W, Wu FR, Chen TY, Yuan FL (2012) Inhibition of acid-sensing ion channels by amiloride protects rat articular chondrocytes from acid-induced apoptosis via a mitochondrial-mediated pathway. Cell Biol Int 36:635–641
Yuan FL, Chen FH, Lu WG, Li X, Wu FR, Li JP, Li CW, Wang Y, Zhang TY, Hu W (2010) Acid-sensing ion channel 1a mediates acid-induced increases in intracellular calcium in rat articular chondrocytes. Mol Cell Biochem 340:153–159
Teitelbaum SL (2011) The osteoclast and its unique cytoskeleton. Ann N Y Acad Sci 1240:14–17
Zhao H, Liu X, Zou H et al (2016) Osteoprotegerin disrupts peripheral adhesive structures of osteoclasts by modulating Pyk2 and Src activities. Cell Adhes Migr 10:299–309
Jurdic P, Saltel F, Chabadel A, Destaing O (2006) Podosome and sealing zone: specificity of the osteoclast model. Eur J Cell Biol 85:195–202
McHugh KP, Hodivala-Dilke K, Zheng MH, Namba N, Lam J, Novack D, Feng X, Ross FP, Hynes RO, Teitelbaum SL (2000) Mice lacking beta3 integrins are osteosclerotic because of dysfunctional osteoclasts. J Clin Invest 105:433–440
Hassanpour S, Jiang H, Wang Y, Kuiper JW, Glogauer M (2014) The actin binding protein adseverin regulates osteoclastogenesis. PLoS One 9:e109078
Jiang H, Wang Y, Viniegra A, Sima C, McCulloch CA, Glogauer M (2015) Adseverin plays a role in osteoclast differentiation and periodontal disease-mediated bone loss. FASEB J 29:2281–2291
Lev S, Moreno H, Martinez R, Canoll P, Peles E, Musacchio JM, Plowman GD, Rudy B, Schlessinger J (1995) Protein tyrosine kinase PYK2 involved in Ca(2+)-induced regulation of ion channel and MAP kinase functions. Nature 376:737–745
Duong LT, Lakkakorpi PT, Nakamura I, Machwate M, Nagy RM, Rodan GA (1998) PYK2 in osteoclasts is an adhesion kinase, localized in the sealing zone, activated by ligation of alpha(v)beta3 integrin, and phosphorylated by src kinase. J Clin Invest 102:881–892
Sanjay A, Houghton A, Neff L et al (2001) Cbl associates with Pyk2 and Src to regulate Src kinase activity, alpha(v)beta(3) integrin-mediated signaling, cell adhesion, and osteoclast motility. J Cell Biol 152:181–195
Acknowledgments
This study was supported by the Natural Science Foundation of China (81270011; 81472125) and the Natural Science Foundation of Jiangsu Province (Grant BK20151114) and Foundation of Traditional Chinese Medicine of Jiangsu Province (YB201578).
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Experimental protocols of this study were approved by the Ethics Committee of Medical College of Nantong University (20141101-001).
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X. Li and J. X. Ye are co-first authors.
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Li, X., Ye, JX., Xu, MH. et al. Evidence that activation of ASIC1a by acidosis increases osteoclast migration and adhesion by modulating integrin/Pyk2/Src signaling pathway. Osteoporos Int 28, 2221–2231 (2017). https://doi.org/10.1007/s00198-017-4017-0
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DOI: https://doi.org/10.1007/s00198-017-4017-0