Immunologic Regulation of Bone Development

  • Mark C. Horowitz
  • Joseph A. Lorenzo
Part of the Advances in Experimental Medicine and Biology book series (volume 602)

A regulatory network comprised of transcription factors PU.1, Ikaros, E2A, EBF, and Pax5 control B cell fate specification and differentiation. Early B Cell Factor-1 (EBF-1) is essential for B cell fate specification while Pax5 is required for B cell development. Mice deficient in Pax5 or EBF-1 have a developmental arrest of B cell differentiation at the pro-B cell stage, which results in the absence of mature B cells. We analyzed the bone phenotype of Pax5 and EBF-1 wild-type (+/+) and homozygous mutant (–/–) mice to determine if the loss of these transcription factors regulated bone cell development.

Bones from Pax5–/– mice were strikingly osteopenic 15 days after birth, with increased numbers of osteoclasts, and decreased trabecular number. The number of osteoblasts in Pax5–/– bones and their function in vitro were not different from controls. In addition, Pax5 was not expressed by wild-type osteoblasts. To investigate the origin of the in vivo increase in osteoclasts, Pax5–/– or +/+ spleen cells were cultured with M-CSF and RANKL and multinucleated, TRAP+ cells counted. Cells from Pax5–/– spleen produced 5-10 times more osteoclasts than did controls.

Tibia from EBF-1–/– mice had a striking increase in osteoblasts lining bone surfaces. Consistent with this was an increase in osteoid thickness and in the bone formation rate. This correlated with a 2-fold increase in serum osteocalcin. However, in vitro proliferation and ALP of mutant osteoblasts did not differ from control. In contrast, osteoclast number was similar in 4 week-old +/+ and –/– mice; however, at 12 weeks the number of osteoclasts was more than twice that of controls. These data correlated with an increase in bone volume at 12 weeks of age. The most striking aspect of the EBF-1–/– bones was the presence of adipocytes, which filled the marrow space. The adipocytes in the marrow were present at both 4 and 12 weeks of age. Increased fat was also seen in the liver of mutant mice. However, subcutaneous fat was almost absent in EBF-1–/– mice. Importantly, EBF-1 mRNA was expressed in wild-type osteoblasts and in adipocytes. Loss of EBF-1 and Pax5 causes distinct, non-overlapping bone phenotypes. It is important to understand why this network of transcription factors, which are so important for B cell development, have such striking effects on bone cell growth and development.


Osteoclast Precursor Paired Domain Pax5 Expression Cell Fate Specification Bone Phenotype 


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  1. Adams, B., P. Dorfler, A. Aguzzi, Z. Kozmik, P. Urbanek, I. Maurer-Fogy, and M. Busslinger. 1992. Pax5 encodes the transcription factor BSAP and is expressed in B-lymphocytes, the developing CNS, and adult testis. Genes Dev 6: 1589–1607.CrossRefPubMedGoogle Scholar
  2. Akerblad, P., U. Lind, D. Liberg, K. Bamberg, and M. Sigvardsson. 2002. Early B-cell factor (O/E-1) is a promoter of adipogenesis and involved in control of genes important for terminal adipocyte differentiation. Mol Cell Biol 22: 8015–8025.CrossRefPubMedGoogle Scholar
  3. Baldwin, C.T., C.F. Hoth, J.A. Amos, E.O. da-Silva, and A. Milunsky. 1992. An exonic mutation in the HuP2 paired domain gene causes Waardenburg’s syndrome. Nature 355: 637–638.CrossRefPubMedGoogle Scholar
  4. Balling, R., U. Deutsch, and P. Gruss. 1988. Undulated, a mutation affecting the development of the mouse skeleton, has a point mutation in the paired box of Pax1. Cell 55: 531–535.CrossRefPubMedGoogle Scholar
  5. Baumgartner, M., D. Bopp, M. Burri, and L.M. Nol. 1987. Structure of two genes at the gooseberry locus related to the paired gene and their spatial expression during Drosophila embryogenesis. Genes Dev 1: 1247–1267.CrossRefPubMedGoogle Scholar
  6. Bopp, D., M. Burri, S. Baumgartner, G. Frigerio, and M. Noll. 1986. Conservation of a large protein domain in the segmentation gene paried and in functionally related genes of Drosphila. Cell 47: 1033–1040.CrossRefPubMedGoogle Scholar
  7. Burri, M., Y. Tromvoulis, D. Bopp, G. Frigerio, and L.M. Nol. 1989. Conservation of the paired domain in metazoans and its structure in three isolated human genes. EMBO J 8: 1183–1190.Google Scholar
  8. Campos-Ortega, J.A. 1998. The genetics of the Drosophila achaete-scute gene complex: a historical appraisal. Int J Dev Biol 42: 291–297.PubMedGoogle Scholar
  9. Crozatier, M., D. Valle, L. Dubois, S. Ibnsouda, and A. Vincent. 1996. Collier, a novel regulator of Drosophila head development, is expressed in a single mitotic domain. Curr. Biol 6: 707–718.CrossRefPubMedGoogle Scholar
  10. Dambly-Chaudiere, C., and M. Vervoort. 1998. The bHLH genes in neural development. Int J Dev Biol. 42: 269–273.PubMedGoogle Scholar
  11. Dowell, P., and D.W. Cooke. 2002. Olf-1/early B cell factor is a regulator of gult4 gene expression in 3T3–L1 adipocytes. J Biol Chem 277: 1712–1718.CrossRefPubMedGoogle Scholar
  12. Epstein, D.J., M. Vekemans, and P. Gros. 1991. Splotch (Sp2H), a mutation affecting development of the mouse neural tube, shows a deletion within the paired homeodomain of Pax3. Cell 57: 767–774.CrossRefGoogle Scholar
  13. Garel, S., F. Marin, M.G. Mattei, C. Vesque, A. Vincent, and P. Charnay. 1997. Family of Ebf/Olf-1-related genes potentially involved in neuronal differentiation and regional specification in the central nervous system. Dev Dyn 210: 191–205.CrossRefPubMedGoogle Scholar
  14. Georgopoulos, K., M. Bigby, J-H. Want, A. Molnar, P. Wu, S. Winandy, and A. Sharpe. 1994. The Ikaros gene is required for the development of all lymphoid lineages. Cell 79: 143–156.CrossRefPubMedGoogle Scholar
  15. Hagman, J., C. Belanger, A. Travis, C.W. Turck, and R. Grosschedl. 1993. Cloning and functional characterization of early B-cell factor, a regulator of lymphocyte-specific gene expression. Genes Dev 7: 760–773.CrossRefPubMedGoogle Scholar
  16. Hardy, R.R., C.E. Carmack, S.A. Shinton, J.D. Kemp, and K. Hayakawa. 1991. Resolution, and characterization of pro-B and Pre-pro-B cell stages in normal mouse bone marrow. J Exp Med 173: 1213–1225.CrossRefPubMedGoogle Scholar
  17. Hata, A., J. Seoane, G. Lagna, E. Montalvo, A. Hemmati-Brivanlou, and J. Massague. 2000. OAZ uses distinct DNA- and protein-binding zinc fingers in separate BMP-Smad and Olf signaling pathways. Cell 100: 229–240.CrossRefPubMedGoogle Scholar
  18. Hill, E.R., J. Favor, B.L.M. Hogan, C.C. Ton, G.F Saunders, I.M. Hanson, J. Prosser, T. Jordan, N.D. Hastie, and V. van Heyningen. 1991. Mouse Small eye results from mutations in a paired-like homeobox-containing gene. Nature 354: 522–525.CrossRefPubMedGoogle Scholar
  19. Hofbauer, L.C., and A.E. Heufelder. 2000. The role of receptor activator of nuclear factor-HB ligand and osteoprotegerin in the pathogenesis and treatment of metabolic bone diseases. J Clin Endocrinol Metab 85(7): 2355–2363.CrossRefPubMedGoogle Scholar
  20. Horowitz, M.C., Y. Xi, D.L. Pflugh, D.G.T. Hesslein, D.G. Schatz, J.A. Lorenzo, and A.L.M. Bothwell. 2004. Pax5 deficient mice exhibit early onset osteopenia with increased osteoclast progenitors. J Immunol 173: 6583–6591.PubMedGoogle Scholar
  21. Horowitz, M.C., A.L.M. Bothwell, D.G.T. Hesslein, D.L. Pflugh, and D.G. Schatz. 2005. B cells and osteoblast and osteoclast development. Immunol Rev 208: 141–153.CrossRefPubMedGoogle Scholar
  22. Lin, H., and R. Grosschedl. 1995. Failure of B-cell differentiation in mice lacking the transcription factor EBF. Nature 376: 263–267.CrossRefPubMedGoogle Scholar
  23. O’Riordan, M., and R. Grosschedl. 1999. Coordinate regulation of B cell differentiation by the transcription factors EBF and E2A. Immunity 11: 21–31.CrossRefPubMedGoogle Scholar
  24. Ton, C.C., H. Hirvonen, H. Miwa, M.M. Weil, P. Monaghan, T. Jordan, V. van Heyningen, N.D. Hastie, H. Meijers-Heijboer, M. Drechsler, B. Royer-Pokora, F. Collins, A. Swaroop, L.C. Strong, and G.F. Saunders. 1991. Positional cloning and characterization of a paired-box-and homeobox-containing gene from the anirida region. Cell 57: 1059–1074.CrossRefGoogle Scholar
  25. Tondravi, M.M., S.R. McKercher, K. Anderson, J.M. Erdmann, M. Quiroz, R. Maki, and S.L. Teitelbaum. 1997. Osteopetrosis in mice lacking haematopoietic transcription factor PU.1. Nature 386: 81–84.CrossRefPubMedGoogle Scholar
  26. Urbánek, P., Z-Q. Wang, I. Fetka, E.F. Wagner, and M. Busslinger. 1994. Complete block of early B cell differentiation and altered patterning of the posterior midbrain in mice lacking Pax5/BSAP. Cell 79: 901–912.CrossRefPubMedGoogle Scholar
  27. Walther, C., J.I. Guenet, D. Simon, U. Deutsch, B. Jostes, M.D. Goulding, D. Plachov, R. Balling, P. Gruss. 1991. Pax: A murine multigene family of paired box-containing genes. Genomics 11: 424–434.CrossRefPubMedGoogle Scholar
  28. Wang, M.M., and R.R. Reed. 1993. Molecular cloning of the olfactory neuronal transcription factor Olf-1 by genetic selection in yeast. Nature 364: 121–126.CrossRefPubMedGoogle Scholar
  29. Wang, S.S., R.Y.L. Tsai, and R.R. Reed. 1997. The characterization of the Olf-1/EBF-like HLH transcription factor family: implications in olfactory gene regulation and neuronal development. J Neurosci 17: 4149–4158.PubMedGoogle Scholar
  30. Wang, S.S., A.G. Betz, and R.R. Reed. 2002. Cloning of a novel Olf-1/EBF-like gene, O/E-4, by degenerate oligo-based direct selection. Mol Cell Neurosci 20: 404–414.CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2007

Authors and Affiliations

  • Mark C. Horowitz
    • 1
  • Joseph A. Lorenzo
    • 2
  1. 1.Department of Orthopaedics and RehabilitationYale University School of MedicineNew HavenUSA
  2. 2.Department of MedicineUniversity of Connecticut Health CenterNew HavenUSA

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