Skip to main content

Advertisement

SpringerLink
Log in

Involvement of endoplasmic reticulum stress in homocysteine-induced apoptosis of osteoblastic cells

  • Original Article
  • Published:
Journal of Bone and Mineral Metabolism Aims and scope Submit manuscript

Abstract

Hyperhomocysteinemia has been shown to increase the incidence of osteoporosis and osteoporotic fractures. Endoplasmic reticulum (ER) stress was recently shown to be associated with apoptosis in several types of cells. In this study, we determined the effect of homocysteine (Hcy) on the apoptosis of osteoblastic cells and investigated whether ER stress participates in Hcy-induced osteoblast apoptosis. Human osteoblastic cells were incubated with Hcy. Hcy dose-dependently decreased cell viability and increased apoptosis in osteoblastic cells. Osteoblastic cells are more susceptible to Hcy-mediated cell death than other cell types. Expression of cleaved caspase-3 was significantly increased by Hcy, and pretreatment with caspase-3 inhibitor rescued the cell viability by Hcy. Hcy treatment led to an increase in release of mitochondrial cytochrome c. It also triggered ER stress by increased expression of glucose-regulated protein 78, inositol-requiring transmembrane kinase and endonuclease 1α (IRE-1α), spliced X-box binding protein, activating transcription factor 4, and C/EBP homologous protein. Silencing IRE-1α expression by small interfering RNA effectively suppressed Hcy-induced apoptosis of osteoblastic cells. Our results suggest that hyperhomocysteinemia induces apoptotic cell death in osteoblasts via ER stress.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

Abbreviations

IRE-1α:

Inositol-requiring transmembrane kinase and endonuclease 1α

sXBP-1:

Spliced X-box binding protein

ATF4:

Activating transcription factor 4

Bax:

Bcl-2-associated X protein

Bcl-2:

B-cell lymphoma 2

CHOP:

C/EBP homologous protein

ER:

Endoplasmic reticulum

FLS:

Fibroblast-like synoviocytes

GRP78:

Glucose-regulated protein 78

Hcy:

Homocysteine

HUVECs:

Human umbilical vein endothelial cells

PERK:

PKR-like ER-associated kinase

UPR:

Unfolded protein response

References

  1. Kanis JA, McCloskey EV, Johansson H, Oden A (2009) Approaches to the targeting of treatment for osteoporosis. Nat Rev Rheumatol 5:425–431

    PubMed  CAS  Google Scholar 

  2. Seeman E (2002) Pathogenesis of bone fragility in women and men. Lancet 359:1841–1850

    PubMed  Google Scholar 

  3. Manolagas SC (1998) Cellular and molecular mechanisms of osteoporosis. Aging (Milano) 10:182–190

    CAS  Google Scholar 

  4. Weinstein RS, Manolagas SC (2000) Apoptosis and osteoporosis. Am J Med 108:153–164

    PubMed  CAS  Google Scholar 

  5. Harvey N, Dennison E, Cooper C (2010) Osteoporosis: impact on health and economics. Nat Rev Rheumatol 6:99–105

    PubMed  Google Scholar 

  6. Levasseur R (2009) Bone tissue and hyperhomocysteinemia. Jt Bone Spine 76:234–240

    CAS  Google Scholar 

  7. Yap S, Naughten E (1998) Homocystinuria due to cystathionine beta-synthase deficiency in Ireland: 25 years’ experience of a newborn screened and treated population with reference to clinical outcome and biochemical control. J Inherit Metab Dis 21:738–747

    PubMed  CAS  Google Scholar 

  8. Testai FD, Gorelick PB (2010) Inherited metabolic disorders and stroke part 2: homocystinuria, organic acidurias, and urea cycle disorders. Arch Neurol 67:148–153

    PubMed  Google Scholar 

  9. van Meurs JB, Dhonukshe-Rutten RA, Pluijm SM et al (2004) Homocysteine levels and the risk of osteoporotic fracture. N Engl J Med 350:2033–2041

    PubMed  Google Scholar 

  10. McLean RR, Jacques PF, Selhub J, Tucker KL, Samelson EJ, Broe KE, Hannan MT, Cupples LA, Kiel DP (2004) Homocysteine as a predictive factor for hip fracture in older persons. N Engl J Med 350:2042–2049

    PubMed  CAS  Google Scholar 

  11. Gjesdal CG, Vollset SE, Ueland PM, Refsum H, Meyer HE, Tell GS (2007) Plasma homocysteine, folate, and vitamin B 12 and the risk of hip fracture: the hordaland homocysteine study. J Bone Miner Res 22:747–756

    PubMed  CAS  Google Scholar 

  12. Herrmann M, Wildemann B, Claes L, Klohs S, Ohnmacht M, Taban-Shomal O, Hubner U, Pexa A, Umanskaya N, Herrmann W (2007) Experimental hyperhomocysteinemia reduces bone quality in rats. Clin Chem 53:1455–1461

    PubMed  CAS  Google Scholar 

  13. Gupta S, Kuhnisch J, Mustafa A, Lhotak S, Schlachterman A, Slifker MJ, Klein-Szanto A, High KA, Austin RC, Kruger WD (2009) Mouse models of cystathionine beta-synthase deficiency reveal significant threshold effects of hyperhomocysteinemia. FASEB J 23:883–893

    PubMed  CAS  Google Scholar 

  14. Anagnostis P, Karagiannis A, Kakafika AI, Tziomalos K, Athyros VG, Mikhailidis DP (2009) Atherosclerosis and osteoporosis: age-dependent degenerative processes or related entities? Osteoporos Int 20:197–207

    PubMed  CAS  Google Scholar 

  15. Zhang K, Kaufman RJ (2008) From endoplasmic-reticulum stress to the inflammatory response. Nature 454:455–462

    PubMed  CAS  Google Scholar 

  16. Ji C, Kaplowitz N (2003) Betaine decreases hyperhomocysteinemia, endoplasmic reticulum stress, and liver injury in alcohol-fed mice. Gastroenterology 124:1488–1499

    PubMed  CAS  Google Scholar 

  17. Kim HJ, Cho HK, Kwon YH (2008) Synergistic induction of ER stress by homocysteine and beta-amyloid in SH-SY5Y cells. J Nutr Biochem 19:754–761

    PubMed  CAS  Google Scholar 

  18. Chigurupati S, Wei Z, Belal C, Vandermey M, Kyriazis GA, Arumugam TV, Chan SL (2009) The homocysteine-inducible endoplasmic reticulum stress protein counteracts calcium store depletion and induction of CCAAT enhancer-binding protein homologous protein in a neurotoxin model of Parkinson disease. J Biol Chem 284:18323–18333

    PubMed  CAS  Google Scholar 

  19. Zhang C, Cai Y, Adachi MT, Oshiro S, Aso T, Kaufman RJ, Kitajima S (2001) Homocysteine induces programmed cell death in human vascular endothelial cells through activation of the unfolded protein response. J Biol Chem 276:35867–35874

    PubMed  CAS  Google Scholar 

  20. Shirakawa K, Maeda S, Gotoh T et al (2006) CCAAT/enhancer-binding protein homologous protein (CHOP) regulates osteoblast differentiation. Mol Cell Biol 26:6105–6116

    PubMed  CAS  Google Scholar 

  21. Pereira RC, Stadmeyer LE, Smith DL, Rydziel S, Canalis E (2007) CCAAT/enhancer-binding protein homologous protein (CHOP) decreases bone formation and causes osteopenia. Bone 40:619–626

    PubMed  CAS  Google Scholar 

  22. Yoo SA, Park BH, Park GS, Koh HS, Lee MS, Ryu SH, Miyazawa K, Park SH, Cho CS, Kim WU (2006) Calcineurin is expressed and plays a critical role in inflammatory arthritis. J Immunol 177:2681–2690

    PubMed  CAS  Google Scholar 

  23. Lee SJ, Kim KM, Namkoong S et al (2005) Nitric oxide inhibition of homocysteine-induced human endothelial cell apoptosis by down-regulation of p53-dependent Noxa expression through the formation of S-nitrosohomocysteine. J Biol Chem 280:5781–5788

    PubMed  CAS  Google Scholar 

  24. Yuan Q, Jiang DJ, Chen QQ, Wang S, Xin HY, Deng HW, Li YJ (2007) Role of asymmetric dimethylarginine in homocysteine-induced apoptosis of vascular smooth muscle cells. Biochem Biophys Res Commun 356:880–885

    PubMed  CAS  Google Scholar 

  25. Kim DJ, Koh JM, Lee O, Kim NJ, Lee YS, Kim YS, Park JY, Lee KU, Kim GS (2006) Homocysteine enhances apoptosis in human bone marrow stromal cells. Bone 39:582–590

    PubMed  CAS  Google Scholar 

  26. Orrenius S, Zhivotovsky B, Nicotera P (2003) Regulation of cell death: the calcium-apoptosis link. Nat Rev Mol Cell Biol 4:552–565

    PubMed  CAS  Google Scholar 

  27. Degli Esposti M (2004) Mitochondria in apoptosis: past, present and future. Biochem Soc Trans 32:493–495

    PubMed  CAS  Google Scholar 

  28. Cotter TG (2009) Apoptosis and cancer: the genesis of a research field. Nat Rev Cancer 9:501–507

    PubMed  CAS  Google Scholar 

  29. Hengartner MO (2000) The biochemistry of apoptosis. Nature 407:770–776

    PubMed  CAS  Google Scholar 

  30. Althausen S, Paschen W (2000) Homocysteine-induced changes in mRNA levels of genes coding for cytoplasmic- and endoplasmic reticulum-resident stress proteins in neuronal cell cultures. Brain Res Mol Brain Res 84:32–40

    PubMed  CAS  Google Scholar 

  31. Shen X, Ellis RE, Lee K et al (2001) Complementary signaling pathways regulate the unfolded protein response and are required for C. elegans development. Cell 107:893–903

    PubMed  CAS  Google Scholar 

  32. Cox JS, Shamu CE, Walter P (1993) Transcriptional induction of genes encoding endoplasmic reticulum resident proteins requires a transmembrane protein kinase. Cell 73:1197–1206

    PubMed  CAS  Google Scholar 

  33. Bellido T, Ali AA, Plotkin LI, Fu Q, Gubrij I, Roberson PK, Weinstein RS, O’Brien CA, Manolagas SC, Jilka RL (2003) Proteasomal degradation of Runx2 shortens parathyroid hormone-induced anti-apoptotic signaling in osteoblasts. A putative explanation for why intermittent administration is needed for bone anabolism. J Biol Chem 278:50259–50272

    PubMed  CAS  Google Scholar 

  34. Tsuboi M, Kawakami A, Nakashima T et al (1999) Tumor necrosis factor-alpha and interleukin-1beta increase the Fas-mediated apoptosis of human osteoblasts. J Lab Clin Med 134:222–231

    PubMed  CAS  Google Scholar 

  35. Suhara T, Fukuo K, Yasuda O, Tsubakimoto M, Takemura Y, Kawamoto H, Yokoi T, Mogi M, Kaimoto T, Ogihara T (2004) Homocysteine enhances endothelial apoptosis via upregulation of Fas-mediated pathways. Hypertension 43:1208–1213

    PubMed  CAS  Google Scholar 

  36. Szegezdi E, Logue SE, Gorman AM, Samali A (2006) Mediators of endoplasmic reticulum stress-induced apoptosis. EMBO Rep 7:880–885

    PubMed  CAS  Google Scholar 

  37. Zhang K, Kaufman RJ (2006) The unfolded protein response: a stress signaling pathway critical for health and disease. Neurology 66:S102–S109

    PubMed  CAS  Google Scholar 

  38. Morishima N, Nakanishi K, Takenouchi H, Shibata T, Yasuhiko Y (2002) An endoplasmic reticulum stress-specific caspase cascade in apoptosis. Cytochrome c-independent activation of caspase-9 by caspase-12. J Biol Chem 277:34287–34294

    PubMed  CAS  Google Scholar 

  39. Cai Y, Zhang C, Nawa T, Aso T, Tanaka M, Oshiro S, Ichijo H, Kitajima S (2000) Homocysteine-responsive ATF3 gene expression in human vascular endothelial cells: activation of c-Jun NH(2)-terminal kinase and promoter response element. Blood 96:2140–2148

    PubMed  CAS  Google Scholar 

  40. Zhang C, Kawauchi J, Adachi MT, Hashimoto Y, Oshiro S, Aso T, Kitajima S (2001) Activation of JNK and transcriptional repressor ATF3/LRF1 through the IRE1/TRAF2 pathway is implicated in human vascular endothelial cell death by homocysteine. Biochem Biophys Res Commun 289:718–724

    PubMed  CAS  Google Scholar 

  41. Chen YR, Meyer CF, Tan TH (1996) Persistent activation of c-Jun N-terminal kinase 1 (JNK1) in gamma radiation-induced apoptosis. J Biol Chem 271:631–634

    PubMed  CAS  Google Scholar 

  42. Ron D, Walter P (2007) Signal integration in the endoplasmic reticulum unfolded protein response. Nat Rev Mol Cell Biol 8:519–529

    PubMed  CAS  Google Scholar 

  43. Zinszner H, Kuroda M, Wang X, Batchvarova N, Lightfoot RT, Remotti H, Stevens JL, Ron D (1998) CHOP is implicated in programmed cell death in response to impaired function of the endoplasmic reticulum. Genes Dev 12:982–995

    PubMed  CAS  Google Scholar 

  44. Yang X, Matsuda K, Bialek P et al (2004) ATF4 is a substrate of RSK2 and an essential regulator of osteoblast biology; implication for Coffin–Lowry syndrome. Cell 117:387–398

    PubMed  CAS  Google Scholar 

  45. Zhang P, McGrath B, Li S, Frank A, Zambito F, Reinert J, Gannon M, Ma K, McNaughton K, Cavener DR (2002) The PERK eukaryotic initiation factor 2 alpha kinase is required for the development of the skeletal system, postnatal growth, and the function and viability of the pancreas. Mol Cell Biol 22:3864–3874

    PubMed  CAS  Google Scholar 

  46. Lisse TS, Thiele F, Fuchs H et al (2008) ER stress-mediated apoptosis in a new mouse model of osteogenesis imperfecta. PLoS Genet 4:e7

    PubMed  Google Scholar 

  47. Berglund S, Sodergren A, Wallberg Jonsson S, Rantapaa Dahlqvist S (2009) Atherothrombotic events in rheumatoid arthritis are predicted by homocysteine—a six-year follow-up study. Clin Exp Rheumatol 27:822–825

    PubMed  CAS  Google Scholar 

  48. Lazzerini PE, Selvi E, Lorenzini S, Capecchi PL, Ghittoni R, Bisogno S, Catenaccio M, Marcolongo R, Galeazzi M, Laghi-Pasini F (2006) Homocysteine enhances cytokine production in cultured synoviocytes from rheumatoid arthritis patients. Clin Exp Rheumatol 24:387–393

    PubMed  CAS  Google Scholar 

  49. Kokame K, Kato H, Miyata T (1996) Homocysteine-respondent genes in vascular endothelial cells identified by differential display analysis. GRP78/BiP and novel genes. J Biol Chem 271:29659–29665

    PubMed  CAS  Google Scholar 

  50. Blass S, Union A, Raymackers J, Schumann F, Ungethum U, Muller-Steinbach S, De Keyser F, Engel JM, Burmester GR (2001) The stress protein BiP is overexpressed and is a major B and T cell target in rheumatoid arthritis. Arthritis Rheum 44:761–771

    PubMed  CAS  Google Scholar 

  51. Hampton RY (2002) ER-associated degradation in protein quality control and cellular regulation. Curr Opin Cell Biol 14:476–482

    PubMed  CAS  Google Scholar 

  52. Amano T, Yamasaki S, Yagishita N et al (2003) Synoviolin/Hrd1, an E3 ubiquitin ligase, as a novel pathogenic factor for arthropathy. Genes Dev 17:2436–2449

    PubMed  CAS  Google Scholar 

Download references

Acknowlegment

This work was supported by grants from the Korea Health 21R&D Project, Ministry of Health & Welfare, Republic of Korea (No. A040018), and the Korea Healthcare Technology R&D Project, Ministry for Health, Welfare and Family Affairs (No. A092258 and A084948), and National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (No. 314-2008-1-E00113).

Conflict of interest

None.

Author information

Authors and Affiliations

  1. Division of Rheumatology, Department of Internal Medicine, College of Medicine, Yeouido St. Mary’s Hospital, The Catholic University of Korea, #62 Yeouido-dong, Yeongdeungpo-ku, Seoul, South Korea

    Su-Jung Park, Ki-Jo Kim, Wan-Uk Kim, Il-Hoan Oh & Chul-Soo Cho

Authors
  1. Su-Jung Park
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ki-Jo Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Wan-Uk Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Il-Hoan Oh
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Chul-Soo Cho
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Chul-Soo Cho.

Additional information

S.-J. Park and K.-J. Kim contributed equally to this work.

About this article

Cite this article

Park, SJ., Kim, KJ., Kim, WU. et al. Involvement of endoplasmic reticulum stress in homocysteine-induced apoptosis of osteoblastic cells. J Bone Miner Metab 30, 474–484 (2012). https://doi.org/10.1007/s00774-011-0346-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00774-011-0346-9

Keywords

Search

Navigation