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Arthritis Research & Therapy
Published in: Arthritis Research & Therapy 3/2012

Open Access 01-06-2012 | Research article

The effects of oxygen tension and antiaging factor Klothoon Wnt signaling in nucleus pulposus cells

Authors: Akihiko Hiyama, Fumiyuki Arai, Daisuke Sakai, Katsuya Yokoyama, Joji Mochida

Published in: Arthritis Research & Therapy | Issue 3/2012

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Abstract

Introduction

The goals of this study were to examine the oxemic regulation of Wnt signaling to explore whether Wnt signaling accelerates the age-related degeneration of nucleus pulposus cells, and if so, to define the mechanism underlying this effect. We investigated the expression of Klotho, a newly identified antiaging gene, and whether its regulation is attributable to the suppression of Wnt signaling.

Methods

Rat nucleus pulposus cells were cultured under normoxic (21% O2) or hypoxic (2% O2) conditions, and the expression and promoter activity of Wnt signaling and Klotho were evaluated. The effect of Klotho protein was examined with transfection experiments, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, senescence-associated β-galactosidase staining, and cell-cycle analysis. To determine the methylation status of the Klotho promoter region, bisulfite genomic sequencing analysis was performed. Its relation with the activation of Wnt signaling was assessed. We also examined whether the expression of Klotho could block the effects of pathological Wnt expression in nucleus pulposus cells.

Results

Nucleus pulposus cells exhibited increased β-catenin mRNA and protein under the hypoxic condition. Klotho protein was expressed in vivo, and protein and messenger RNA expression decreased under the hypoxic condition. Klotho treatment decreased cell proliferation and induced the quiescence of nucleus pulposus cells. In addition, Klotho treatment inhibited expression of β-catenin gene and protein compared with untreated control cells.

Conclusions

These data indicate that Wnt signaling and Klotho form a negative-feedback loop in nucleus pulposus cells. These results suggest that the expression of Klotho is regulated by the balance between upregulation and downregulation of Wnt signaling.
Appendix
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Literature
1.
Hiyama A, Mochida J, Iwashina T, Omi H, Watanabe T, Serigano K, Iwabuchi S, Sakai D: Synergistic effect of low-intensity pulsed ultrasound on growth factor stimulation of nucleus pulposus cells. J Orthop Res. 2007, 25: 1574-15781. 10.1002/jor.20460.CrossRefPubMed
2.
Hiyama A, Mochida J, Iwashina T, Omi H, Watanabe T, Serigano K, Tamura F, Sakai D: Transplantation of mesenchymal stem cells in a canine disc degeneration model. J Orthop Res. 2008, 26: 589-600. 10.1002/jor.20584.CrossRefPubMed
3.
Watanabe T, Sakai D, Yamamoto Y, Iwashina T, Serigano K, Tamura F, Mochida J: Human nucleus pulposus cells significantly enhanced biological properties in a coculture system with direct cell-to-cell contact with autologous mesenchymal stem cells. J Orthop Res. 2010, 28: 623-630.PubMed
4.
Weng LH, Wang CJ, Ko JY, Sun YC, Wang FS: Control of Dkk-1 ameliorates chondrocyte apoptosis, cartilage destruction, and subchondral bone deterioration in osteoarthritic knees. Arthritis Rheum. 2010, 62: 1393-1402. 10.1002/art.27357.CrossRefPubMed
5.
Rudnicki JA, Brown AM: Inhibition of chondrogenesis by Wnt gene expression in vivo and in vitro. Dev Biol. 1997, 185: 104-118. 10.1006/dbio.1997.8536.CrossRefPubMed
6.
Zhu M, Chen M, Zuscik M, Wu Q, Wang YJ, Rosier RN, O'Keefe RJ, Chen D: Inhibition of β-catenin signaling in articular chondrocytes results in the development of osteoarthritis. Arthritis Rheum. 2008, 58: 2053-2064. 10.1002/art.23614.PubMedCentralCrossRefPubMed
7.
Behrens J, Jerchow BA, Würtele M, Grimm J, Asbrand C, Wirtz R, Kühl M, Wedlich D, Birchmeier W: Functional interaction of an axin homolog, conductin, with β-catenin, APC, and GSK3β. Science. 1998, 280: 596-599. 10.1126/science.280.5363.596.CrossRefPubMed
8.
Behrens J, von Kries JP, Kühl M, Bruhn L, Wedlich D, Grosschedl R, Birchmeier W: Functional interaction of β-catenin with the transcription factor LEF-1. Nature. 1996, 382: 638-642. 10.1038/382638a0.CrossRefPubMed
9.
Hartmann C, Tabin CJ: Dual roles of Wnt signaling during chondrogenesis in the chicken limb. Development. 2000, 127: 3141-3159.PubMed
10.
Hiyama A, Sakai D, Risbud MV, Tanaka M, Arai F, Abe K, Mochida J: Enhancement of intervertebral disc cell senescence by WNT/β-catenin signaling-induced matrix metalloproteinase expression. Arthritis Rheum. 2010, 62: 3036-3047. 10.1002/art.27599.PubMedCentralCrossRefPubMed
11.
Hiyama A, Sakai D, Tanaka M, Arai F, Nakajima D, Abe K, Mochida J: The relationship between the Wnt/β-catenin and TGF-β/BMP signals in the intervertebral disc cell. J Cell Physiol. 2011, 226: 1139-1148. 10.1002/jcp.22438.CrossRefPubMed
12.
Urban JP, Smith S, Fairbank JC: Nutrition of the intervertebral disc. Spine. 2004, 29: 2700-2709. 10.1097/01.brs.0000146499.97948.52.CrossRefPubMed
13.
Agrawal A, Guttapalli A, Narayan S, Albert TJ, Shapiro IM, Risbud MV: Normoxic stabilization of HIF-1α drives glycolytic metabolism and regulates aggrecan gene expression in nucleus pulposus cells of the rat intervertebral disk. Am J Physiol Cell Physiol. 2007, 293: C621-C631. 10.1152/ajpcell.00538.2006.CrossRefPubMed
14.
Walmsley R: The development and growth of the intervertebral disc. Edinb Med J. 1953, 60: 341-364.PubMed
15.
Erwin WM, Inman RD: Notochord cells regulate intervertebral disc chondrocyte proteoglycan production and cell proliferation. Spine. 2006, 31: 1094-1099. 10.1097/01.brs.0000216593.97157.dd.CrossRefPubMed
16.
Butler WE: Comparative anatomy and development of the mammalian disc. The Biology of the Intervertebral Disc. Edited by: Gosh P. 1989, Boca Raton, FL: CRC Press, 84-108.
17.
Hunter CJ, Matyas JR, Duncan NA: The notochordal cell in the nucleus pulposus: a review in the context of tissue engineering. Tissue Eng. 2003, 9: 667-677. 10.1089/107632703768247368.CrossRefPubMed
18.
Ben-Dov IZ, Galitzer H, Lavi-Moshayoff V, Goetz R, Kuro-o M, Mohammadi M, Sirkis R, Naveh-Many T, Silver J: The parathyroid is a target organ for FGF23 in rats. J Clin Invest. 2007, 117: 4003-4008.PubMedCentralPubMed
19.
Hu MC, Shi M, Zhang J, Pastor J, Nakatani T, Lanske B, Razzaque MS, Rosenblatt KP, Baum MG, Kuro-o M, Moe OW: Klotho: a novel phosphaturic substance acting as an autocrine enzyme in the renal proximal tubule. FASEB J. 2010, 9: 3438-3450.CrossRef
20.
Li SA, Watanabe M, Yamada H, Nagai A, Kinuta M, Takei K: Immunohistochemical localization of Klotho protein in brain, kidney, and reproductive organs of mice. Cell Struct Funct. 2004, 29: 91-99. 10.1247/csf.29.91.CrossRefPubMed
21.
Kuro-o M, Matsumura Y, Aizawa H, Kawaguchi H, Suga T, Utsugi T, Ohyama Y, Kurabayashi M, Kaname T, Kume E, Iwasaki H, Iida A, Shiraki-Iida T, Nishikawa S, Nagai R, Nabeshima YI: Mutation of the mouse klotho gene leads to a syndrome resembling ageing. Nature. 1997, 390: 45-51. 10.1038/36285.CrossRefPubMed
22.
Kurosu H, Yamamoto M, Clark JD, Pastor JV, Nandi A, Gurnani P, McGuinness OP, Chikuda H, Yamaguchi M, Kawaguchi H, Shimomura I, Takayama Y, Herz J, Kahn CR, Rosenblatt KP, Kuro-o M: Suppression of aging in mice by the hormone Klotho. Science. 2005, 309: 1829-1833. 10.1126/science.1112766.PubMedCentralCrossRefPubMed
23.
Nabeshima Y: Toward a better understanding of Klotho. Sci Aging Knowledge Environ. 2006, 8: pe11-
24.
Kawano K, Ogata N, Chiano M, Molloy H, Kleyn P, Spector TD, Uchida M, Hosoi T, Suzuki T, Orimo H, Inoue S, Nabeshima Y, Nakamura K, Kuro-o M, Kawaguchi H: Klotho gene polymorphisms associated with bone density of aged postmenopausal women. J Bone Miner Res. 2002, 17: 1744-1751. 10.1359/jbmr.2002.17.10.1744.CrossRefPubMed
25.
Ogata N, Matsumura Y, Shiraki M, Kawano K, Koshizuka Y, Hosoi T, Nakamura K, Kuro-O M, Kawaguchi H: Association of Klotho gene polymorphism with bone density and spondylosis of lumber spine in postmenopausal women. Bone. 2002, 31: 37-42. 10.1016/S8756-3282(02)00786-X.CrossRefPubMed
26.
Mullin BH, Wilson SG, Islam FM, Calautti M, Dick IM, Devine A, Prince RL: Klotho gene polymorphisms are associated with osteocalcin levels but not bone density of aged postmenopausal women. Calcif Tissue Int. 2005, 77: 145-151. 10.1007/s00223-004-0291-x.CrossRefPubMed
27.
Yamada Y, Ando F, Niino N, Shimokata H: Association of polymorphisms of the androgen receptor and klotho genes with bone mineral density in Japanese women. J Mol Med (Berl). 2005, 83: 50-57. 10.1007/s00109-004-0578-4.CrossRef
28.
Liu H, Fergusson MM, Castilho RM, Liu J, Cao L, Chen J, Malide D, Rovira II, Schimel D, Kuo CJ, Gutkind JS, Hwang PM, Finkel T: Augmented Wnt signaling in a mammalian model of accelerated aging. Science. 2007, 317: 803-806. 10.1126/science.1143578.CrossRefPubMed
29.
Choi BH, Kim CG, Lim Y, Lee YH, Shin SY: Transcriptional activation of the human Klotho gene by epidermal growth factor in HEK293 cells; role of Egr-1. Gene. 2010, 450: 121-127. 10.1016/j.gene.2009.11.004.CrossRefPubMed
30.
Reinhold MI, Kapadia RM, Liao Z, Naski MC: The Wnt-inducible transcription factor Twist1 inhibits chondrogenesis. J Biol Chem. 2006, 281: 1381-1388.CrossRefPubMed
31.
Yamada Y, Miyashita T, Savagner P, Horton W, Brown KS, Abramczuk J, Xie HX, Kohno K, Bolander M, Bruggeman L: Regulation of the collagen II gene in vitro and in transgenic mice. Ann N Y Acad Sci. 1990, 580: 81-87. 10.1111/j.1749-6632.1990.tb17920.x.CrossRefPubMed
32.
Sato N, Meijer L, Skaltsounis L, Greengard P, Brivanlou AH: Maintenance of pluripotency in human and mouse embryonic stem cells through activation of Wnt signaling by a pharmacological GSK-3-specific inhibitor. Nat Med. 2004, 10: 55-63. 10.1038/nm979.CrossRefPubMed
33.
Risbud MV, Fertala J, Vresilovic EJ, Albert TJ, Shapiro IM: Nucleus pulposus cells upregulate PI3K/Akt and MEK/ERK signaling pathways under hypoxic conditions and resist apoptosis induced by serum withdrawal. Spine. 2005, 30: 882-889. 10.1097/01.brs.0000159096.11248.6d.CrossRefPubMed
34.
Camilli TC, Xu M, O'Connell MP, Chien B, Frank BP, Subaran S, Indig FE, Morin PJ, Hewitt SM, Weeraratna AT: Loss of Klotho during melanoma progression leads to increased filamin cleavage, increased Wnt5A expression, and enhanced melanoma cell motility. Pigment Cell Melanoma Res. 2011, 24: 175-186. 10.1111/j.1755-148X.2010.00792.x.PubMedCentralCrossRefPubMed
35.
Ishihara H, Urban JP: Effects of low oxygen concentrations and metabolic inhibitors on proteoglycan and protein synthesis rates in the intervertebral disc. J Orthop Res. 1999, 17: 829-835. 10.1002/jor.1100170607.CrossRefPubMed
36.
Risbud MV, Guttapalli A, Albert TJ, Shapiro IM: Hypoxia activates MAPK activity in rat nucleus pulposus cells: regulation of integrin expression and cell survival. Spine. 2005, 30: 2503-2509. 10.1097/01.brs.0000186326.82747.13.CrossRefPubMed
37.
Mwale F, Ciobanu I, Giannitsios D, Roughley P, Steffen T, Antoniou J: Effect of oxygen levels on proteoglycan synthesis by intervertebral disc cells. Spine. 2011, 36: E131-E138. 10.1097/BRS.0b013e3181d52b9e.CrossRefPubMed
38.
Kaidi A, Williams AC, Paraskeva C: Interaction between β-catenin and HIF-1 promotes cellular adaptation to hypoxia. Nat Cell Biol. 2007, 9: 210-217. 10.1038/ncb1534.CrossRefPubMed
39.
Agrawal A, Gajghate S, Smith H, Anderson DG, Albert TJ, Shapiro IM, Risbud MV: Cited2 modulates hypoxia-inducible factor-dependent expression of vascular endothelial growth factor in nucleus pulposus cells of the rat intervertebral disc. Arthritis Rheum. 2008, 58: 3798-3808. 10.1002/art.24073.CrossRefPubMed
40.
41.
Kurosu H, Ogawa Y, Miyoshi M, Yamamoto M, Nandi A, Rosenblatt KP, Baum MG, Schiavi S, Hu MC, Moe OW, Kuro-o M: Regulation of fibroblast growth factor-23 signaling by klotho. J Biol Chem. 2006, 281: 6120-6123. 10.1074/jbc.C500457200.PubMedCentralCrossRefPubMed
42.
Kuro-o M: Klotho as a regulator of fibroblast growth factor signaling and phosphate/calcium metabolism. Curr Opin Nephrol Hypertens. 2006, 15: 437-441. 10.1097/01.mnh.0000232885.81142.83.CrossRefPubMed
43.
Doi S, Zou Y, Togao O, Pastor JV, John GB, Wang L, Shiizaki K, Gotschall R, Schiavi S, Yorioka N, Takahashi M, Boothman DA, Kuro-o M: Klotho inhibits transforming growth factor-b1 (TGF-b1) signaling and suppresses renal fibrosis and cancer metastasis in mice. J Biol Chem. 2011, 286: 8655-8665. 10.1074/jbc.M110.174037.PubMedCentralCrossRefPubMed
44.
Kuro-o M: Endocrine FGFs and Klothos: emerging concepts. Trends Endocrinol Metab. 2008, 19: 239-245. 10.1016/j.tem.2008.06.002.CrossRefPubMed
45.
Maldonado BA, Oegema TR: Initial characterization of the metabolism of intervertebral disc cells encapsulated in microspheres. J Orthop Res. 1992, 22: 2885-2893.
46.
Monaghan AP, Kioschis P, Wu W, Zuniga A, Bock D, Poustka A, Delius H, Niehrs C: Dickkopf genes are co-ordinately expressed in mesodermal lineages. Mech Dev. 1999, 87: 45-56. 10.1016/S0925-4773(99)00138-0.CrossRefPubMed
47.
Semënov MV, Tamai K, Brott BK, Kühl M, Sokol S, He X: Head inducer Dickkopf-1 is a ligand for Wnt coreceptor LRP6. Curr Biol. 2001, 11: 951-961. 10.1016/S0960-9822(01)00290-1.CrossRefPubMed
Metadata
Title
The effects of oxygen tension and antiaging factor Klothoon Wnt signaling in nucleus pulposus cells
Authors
Akihiko Hiyama
Fumiyuki Arai
Daisuke Sakai
Katsuya Yokoyama
Joji Mochida
Publication date
01-06-2012
Publisher
BioMed Central
Published in
Arthritis Research & Therapy / Issue 3/2012
Electronic ISSN: 1478-6362
DOI
https://doi.org/10.1186/ar3830

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