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Growth Plate Zonal Microarray Analysis Shows Upregulation of Extracellular Matrix Genes and Downregulation of Metalloproteinases and Cathepsins following Irradiation

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Abstract

Although the growth plate matrix area fraction increases after irradiation, extracellular matrix (ECM) gene expression in this context has not been studied. The hypothesis was that normally expressed ECM genes would be upregulated after irradiation. The right limbs of six Sprague-Dawley 5-week-old rats were irradiated with the left limbs as controls. Half of the animals were harvested after 1 week and half after 2. Microarray was conducted from normal and irradiated tibial growth plate proliferative zone (PZ) and hypertrophic zone (HZ) chondrocytes separated by laser microdissection at each time point. In situ hybridization (ISH) and real-time polymerase chain reaction (PCR) were used to confirm expression of selected genes. At 1 and 2 weeks after irradiation, both normally expressed ECM genes and others not highly expressed in the normal growth plate showed upregulation. Metalloproteinases and cathepsins were downregulated. PZ gene expression after irradiation exhibited features of the normal HZ, suggesting premature terminal differentiation. ECM genes not highly expressed in the normal growth plate included several members of the small leucine-rich proteins and the ezrin–radixin–moesin family. The effects of irradiation on cathepsin K (Ctsk), integrin binding sialoprotein (Ibsp), and procollagen II alpha 1 (Col2a1), as determined by ISH and real-time PCR, were highly correlated with the microarray results. Accumulation of matrix following radiation injury to the growth plate correlated well with changes in gene expression. Upregulation of genes not normally highly expressed in the noninjured growth plate suggests their importance in the injury and repair response.

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References

  1. Goldwein JW (1991) Effects of radiation therapy on skeletal growth in childhood. Clin Orthop 262:101–107

    PubMed  Google Scholar 

  2. Damron TA, Mathur S, Horton JA, Strauss J, Margulies B, Grant W, Farnum CE, Spadaro JA (2004) Temporal changes in PTHrP, Bcl-2, Bax, caspase, TGF-beta, and FGF-2 expression following growth plate irradiation with or without radioprotectant. J Histochem Cytochem 52:157–167

    PubMed  CAS  Google Scholar 

  3. Sandell LJ, Sugai JV, Trippel SB (1994) Expression of collagens I, II, X, and XI and aggrecan mRNAs by bovine growth plate chondrocytes in situ. J Orthop Res 12:1–14

    Article  PubMed  CAS  Google Scholar 

  4. Fukunaga T, Yamashiro T, Oya S, Takeshita N, Takigawa M, Takano-Yamamoto T (2003) Connective tissue growth factor mRNA expression pattern in cartilages is associated with their type I collagen expression. Bone 33:911–918

    Article  PubMed  CAS  Google Scholar 

  5. Unsold C, Pappano WN, Imamura Y, Steiglitz BM, Greenspan DS (2002) Biosynthetic processing of the pro-alpha 1(V)2pro-alpha 2(V) collagen heterotrimer by bone morphogenetic protein-1 and furin-like proprotein convertases. J Biol Chem 277:5596–5602

    Article  PubMed  CAS  Google Scholar 

  6. Wang Y, Middleton F, Horton JA, Reichel L, Farnum CE, Damron TA (2004) Microarray analysis of proliferative and hypertrophic growth plate zones identifies differentiation markers and signal pathways. Bone 35:1273–1293

    Article  PubMed  CAS  Google Scholar 

  7. Middleton FA, Mirnics K, Pierri JN, Lewis DA, Levitt P (2002) Gene expression profiling reveals alterations of specific metabolic pathways in schizophrenia. J Neurosci 22:2718–2729

    PubMed  CAS  Google Scholar 

  8. Weinreb M, Shinar D, Rodan GA (1990) Different pattern of alkaline phosphatase, osteopontin, and osteocalcin expression in developing rat bone visualized by in situ hybridization. J Bone Miner Res 5:831–842

    Article  PubMed  CAS  Google Scholar 

  9. DeRisi JL, Iyer VR, Brown PO (1997) Exploring the metabolic and genetic control of gene expression on a genomic scale. Science 278:680–686

    Article  PubMed  CAS  Google Scholar 

  10. Abdul M, Hoosein N (2000) Changes in beta-2 microglobulin expression in prostate cancer 5:168–172

  11. Zhang H, Liew CC, Marshall KW (2002) Microarray analysis reveals the involvement of beta-2 microglobulin (B2M) in human osteoarthritis. Osteoarthritis Cartilage 10:950–960

    Article  PubMed  CAS  Google Scholar 

  12. Matsuoka Y, Li X, Bennett V (2000) Adducin: structure, function and regulation. Cell Mol Life Sci 57:884–895

    Article  PubMed  CAS  Google Scholar 

  13. Fukata Y, Oshiro N, Kinoshita N, Kawano Y, Matsuoka Y, Bennett V, Matsuura Y, Kaibuchi K (1999) Phosphorylation of adducin by Rho-kinase plays a crucial role in cell motility. J Cell Biol 145:347–361

    Article  PubMed  CAS  Google Scholar 

  14. Singh A, Hitchcock-DeGregori SE (2003) Local destabilization of the tropomyosin coiled coil gives the molecular flexibility required for actin binding. Biochemistry 42:14114–14121

    Article  PubMed  CAS  Google Scholar 

  15. Xie L, Law BK, Aakre ME, Edgerton M, Shyr Y, Bhowmick NA, Moses HL (2003) Transforming growth factor beta-regulated gene expression in a mouse mammary gland epithelial cell line. Breast Cancer Res 5:R187–R198

    Article  PubMed  CAS  Google Scholar 

  16. Gustafsson E, Aszodi A, Ortega N, Hunziker EB, Denker HW, Werb Z, Fassler R (2003) Role of collagen type II and perlecan in skeletal development. Ann N Y Acad Sci 995:140–150

    Article  PubMed  CAS  Google Scholar 

  17. Brekken RA, Sage EH (2001) SPARC, a matricellular protein: at the crossroads of cell-matrix communication. Matrix Biol 19:816–827

    Article  PubMed  CAS  Google Scholar 

  18. Bengtsson E, Morgelin M, Sasaki T, Timpl R, Heinegard D, Aspberg A (2002) The leucine-rich repeat protein PRELP binds perlecan and collagens and may function as a basement membrane anchor. J Biol Chem 277:15061–15068

    Article  PubMed  CAS  Google Scholar 

  19. Marshansky V, Wang X, Bertrand R, Luo H, Duguid W, Chinnadurai G, Kanaan N, Vu MD, Wu J (2001) Proteasomes modulate balance among proapoptotic and antiapoptotic Bcl-2 family members and compromise functioning of the electron transport chain in leukemic cells. J Immunol 166:3130–3142

    PubMed  CAS  Google Scholar 

  20. Loveridge N, Farquharson C, Hesketh JE, Jakowlew SB, Whitehead CC, Thorp BH (1993) The control of chondrocyte differentiation during endochondral bone growth in vivo: changes in TGF-beta and the proto-oncogene c-myc. J Cell Sci 105(pt 4):949–956

    PubMed  CAS  Google Scholar 

  21. Shen Z, Gantcheva S, Mansson B, Heinegard D, Sommarin Y (1998) Chondroadherin expression changes in skeletal development. Biochem J 330(pt 1):549–557

    PubMed  CAS  Google Scholar 

  22. Neame PJ, Tapp H, Azizan A (1999) Noncollagenous, nonproteoglycan macromolecules of cartilage. Cell Mol Life Sci 55:1327–1340

    Article  PubMed  CAS  Google Scholar 

  23. Worapamorn W, Xiao Y, Li H, Young WG, Bartold PM (2002) Differential expression and distribution of syndecan-1 and -2 in periodontal wound healing of the rat. J Periodontal Res 37:293–299

    Article  PubMed  CAS  Google Scholar 

  24. Knudson CB, Knudson W (2001) Cartilage proteoglycans. Semin Cell Dev Biol 12:69–78

    Article  PubMed  CAS  Google Scholar 

  25. Gregory KE, Keene DR, Tufa SF, Lunstrum GP, Morris NP (2001) Developmental distribution of collagen type XII in cartilage: association with articular cartilage and the growth plate. J Bone Miner Res 16:2005–2016

    Article  PubMed  CAS  Google Scholar 

  26. Martin TA, Harrison G, Mansel RE, Jiang WG (2003) The role of the CD44/ezrin complex in cancer metastasis. Crit Rev Oncol Hematol 46:165–186

    Article  PubMed  Google Scholar 

  27. Bretscher A, Edwards K, Fehon RG (2002) ERM proteins and merlin: integrators at the cell cortex. Nat Rev Mol Cell Biol 3:586–599

    Article  PubMed  CAS  Google Scholar 

  28. Hiscox S, Jiang WG (1999) Ezrin regulates cell-cell and cell-matrix adhesion, a possible role with E-cadherin/beta-catenin. J Cell Sci 112(pt 18):3081–3090

    PubMed  CAS  Google Scholar 

  29. Masumoto J, Sagara J, Hayama M, Hidaka E, Katsuyama T, Taniguchi S (1998) Differential expression of moesin in cells of hematopoietic lineage and lymphatic systems. Histochem Cell Biol 110:33–41

    Article  PubMed  CAS  Google Scholar 

  30. Iozzo RV (1999) The biology of the small leucine-rich proteoglycans. Functional network of interactive proteins. J Biol Chem 274:18843–18846

    CAS  Google Scholar 

  31. Wilda M, Bachner D, Just W, Geerkens C, Kraus P, Vogel W, Hameister H (2000) A comparison of the expression pattern of five genes of the family of small leucine-rich proteoglycans during mouse development. J Bone Miner Res 15:2187–2196

    Article  PubMed  CAS  Google Scholar 

  32. Raouf A, Ganss B, McMahon C, Vary C, Roughley PJ, Seth A (2002) Lumican is a major proteoglycan component of the bone matrix. Matrix Biol 21:361–367

    Article  PubMed  CAS  Google Scholar 

  33. Santra M, Mann DM, Mercer EW, Skorski T, Calabretta B, Iozzo RV (1997) Ectopic expression of decorin protein core causes a generalized growth suppression in neoplastic cells of various histogenetic origin and requires endogenous p21, an inhibitor of cyclin-dependent kinases. J Clin Invest 100:149–157

    Article  PubMed  CAS  Google Scholar 

  34. Petersson U, Hultenby K, Wendel M (2003) Identification, distribution and expression of osteoadherin during tooth formation. Eur J Oral Sci 111:128–136

    Article  PubMed  CAS  Google Scholar 

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Acknowledgement

The authors acknowledge Karen Gentile, Joseph Spadaro, Judy Strauss, and Bryan Margulies for their assistance with this project. Funding was provided by National Institutes of Health (National Cancer Institute) grant R01-CA83892 and the David G. Murray Endowed Professorship.

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Authors and Affiliations

  1. Musculoskeletal Sciences Research Laboratory, Department of Orthopedic Surgery, State University of New York Upstate Medical University, Suite 130, 550 Harrison Street, Syracuse, NY, 13210, USA

    Mingliang Zhang, Yan Wang, Jason A. Horton & Timothy A. Damron

  2. Microarray Core Facility, Department of Neuroscience and Physiology, State University of New York Upstate Medical University, Syracuse, NY, 13210, USA

    Frank A. Middleton

  3. Department of Biomedical Sciences, Cornell University, T4018 Veterinary Research Tower, Ithaca, NY, 14853, USA

    Cornelia E. Farnum

Authors
  1. Mingliang Zhang
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  2. Yan Wang
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  3. Frank A. Middleton
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  4. Jason A. Horton
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  5. Cornelia E. Farnum
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  6. Timothy A. Damron
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Correspondence to Timothy A. Damron.

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Zhang, M., Wang, Y., Middleton, F.A. et al. Growth Plate Zonal Microarray Analysis Shows Upregulation of Extracellular Matrix Genes and Downregulation of Metalloproteinases and Cathepsins following Irradiation. Calcif Tissue Int 81, 26–38 (2007). https://doi.org/10.1007/s00223-007-9025-1

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  • DOI: https://doi.org/10.1007/s00223-007-9025-1

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