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Ectopic mineralized cartilage formation in human undifferentiated pancreatic adenocarcinoma explants grown in nude mice

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Abstract

Mineralized as well as nonmineralized cartilage-like structures enclosing cells resembling chondrocytes were found in human-derived undifferentiated but not in poorly differentiated pancreatic adenocarcinoma explants grown in nude mice. The structures reacted with anti-mouse IgG but not with antibodies against human cytokeratin 19, indicating that the newly formed tissue was of mouse origin. High activity of alkaline phosphatase was found in cell layers surrounding the structures and in cells embedded in the matrix. The extracellular matrix was strongly positive after Sirius red staining, reacted with anti-collagen type II antibodies, and the presence of proteoglycans was demonstrated with Alcian blue staining and by metachromasia after Giemsa staining. Electron microscopic inspection revealed the presence of bundles of both thick collagenous fibrils with low levels of fine filamentous material and thin collagenous fibrils with high concentrations of filamentous components. The majority of both types of matrices was found to be partially or completely calcified. The mean area density of the cartilage-like structures in the undifferentiated tumors was 0.31%. The frequent formation of the cartilage-like structures in the rapidly growing undifferentiated explants and its absence in the slowly growing, more differentiated explants suggest that low oxygen tensions in combination with altered levels of growth factors, such as members of the transforming growth factor β superfamily, create conditions that induce differentiation of fibroblasts to chondrocytes. It is concluded that these human tumors grown in nude mice can be used as an in vivo model to study ectopic formation of mineralized cartilage.

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References

  1. Russell RGG, Kanis JA (1984) Ectopic calcification and ossification. In: Nordin BEC (ed) Metabolic bone and stone disease, 2nd ed. Churchill Livingstone, Edinburgh, p 344

    Google Scholar 

  2. Russell RGG, Caswell AM, Hearn PR, Sharrard RM (1986) Calcium in mineralized tissues and pathological calcification. Br Med Bull 42:435–446

    Google Scholar 

  3. Puzas JE, Miller MD, Rosier RN (1989) Pathologic bone formation. Clin Orthop Rel Res 245:269–281

    Google Scholar 

  4. Nollen AJG (1986) Effects of ethylhydroxydiphosphonate (EHDP) on heterotopic ossification. Acta Orthop Scand 57:358–361

    Google Scholar 

  5. Lotke PA (1970) Ossification of the achilles tendon: report of 7 cases. J Bone Joint Surg 52A:157

    Google Scholar 

  6. Finerman GAM, Stover SL (1981) Heterotopic ossification following hip replacement or spinal cord injury. Two clinical studies with EHDP. Metab Bone Dis Rel Res 4, 5:337–342

    Google Scholar 

  7. Smith R, Triffitt JT (1986) Bones in muscles: the problems of soft tissue ossification. Quart J Med 235:985–990

    Google Scholar 

  8. Conner JM (1983) Soft tissue ossification. Springer-Verlag, New York

    Google Scholar 

  9. Galasko CSB (1986) Skeletal metastases. Butterworths, London, p 25

    Google Scholar 

  10. Vogels IMC, Van Noorden CJF, Hoeben KA, Korper W, Jonges GN, Everts V (1993) Use of frozen biological material for combined light and electron microscopy. Ultrastruct Pathol 17:537–546

    Google Scholar 

  11. Kloeppel G, Otto V, Baisch H, Von Bülow M (1988) Growth, stability and experimental treatment of human tumors after transplantation on thymus aplastic nude mice. Verh Dtsch Ges Path 72:1–16.

    Google Scholar 

  12. Coen H, Kint H, Kuyper CF, Jonges GN, Dumarey N, Pauwels M, Kloeppel G, Roels F (1992) Growth behavior of human pancreatic carcinoma xenograft correlates with nuclear features. Cytometry 13:775–781

    Google Scholar 

  13. Van Noorden CJF, Frederiks WM (1992) Enzyme histochemistry: a laboratory manual of current methods. Oxford University Pres Oxford

    Google Scholar 

  14. Frederiks WM, Marx F, Myagkaya GL (1989) The ‘nothing dehydrogenase’ reaction and the detection of ischaemic damage. Histochem J 21:565–573

    Google Scholar 

  15. Beertsen W, Van den Bos T (1992) Alkaline phosphatase induces the mineralization of sheets of collagen implanted subcutaneously in the rat. J Clin Invest 89:1974–1980

    Google Scholar 

  16. Marks SC, Grolman M (1987) Tartrate-resistant acid phosphatase in mononuclear and multinuclear cells during the bone resorption of tooth eruption. J Histochem Cytochem 35:1227–1230

    Google Scholar 

  17. Bianco P, Ballanti P, Bonucci E (1988) Tartrate-resistant acid phosphatase activity in rat osteoblasts and osteocytes. Calcif Tissue Int 43:167–171

    Google Scholar 

  18. James J, Bosch KS, Zuyderhoudt FMJ, Houtkooper JM, Van Gool J (1986) Histophotometric estimation of volume density of collagen as an indication of fibrosis in rat liver. Histochemistry 85:129–133

    Google Scholar 

  19. Shoobridge MPK (1983) A new principle in polychrome staining: a system of automated staining, complementary to hematoxylin and eosin, and usable as a research tool. Stain Technol 58:245–258

    Google Scholar 

  20. Bancroft JD, Cook HC (1984) Manual of histological techniques. Churchill Liyingstone, Edinburgh

    Google Scholar 

  21. Weibel ER (1969) Stereological principles for morphometry in electron microscopic cytology. Int Rev Cytol 26:235–302

    Google Scholar 

  22. Urist MR (1965) Bone formation by autoinduction. Science 150: 893

    Google Scholar 

  23. Huggins CB, Urist MR (1970) Dentin matrix transformations: rapid induction of alkaline phosphatase and cartilage. Science 167:896–898

    Google Scholar 

  24. Reddi AH, Anderson WA (1976) Collagenous bone matrix-induced endochondral ossification and hemopoiesis. J Cell Biol 69:557–572

    Google Scholar 

  25. Reddi AH, Huggins CB (1972) Biochemical sequences in the transformation of normal fibroblasts in adolescent rat. Proc Natl Acad Sci USA 69:1601–1605

    Google Scholar 

  26. Seyedin SM, Thompson AY, Rosen DM, Piez KA (1983) In vitro induction of cartilage-specific macromolecules by a bone extract. J Cell Biol 97:1950–1953

    Google Scholar 

  27. Hunter GK, Holmyard DP, Pritzker KPH (1993) Calcification of chick vertebral chondrocytes grown in agarose gels: a biochemical and ultrastructural study. J Cell Sci 104:1031–1038

    Google Scholar 

  28. Friedenstein AJ, Chailakhjan RK, Lalykina KS (1970) The development of fibroblast colonies in monolayer cultures of guinea pig bone marrow and spleen cells. Cell Tissue Kinet 3:393–403.

    Google Scholar 

  29. Friedenstein AJ (1976) Precursor cells of melanocytes. Int Rev Cytol 47:327–355

    Google Scholar 

  30. Ashton BA, Allen TD, Howlett CR, Eagleson CC, Hattori A, Owen M (1980) Formation of bone and cartilage by marrow stromal cells in diffusion chambers in vivo. Clin Orthop Rel Res 151:294–307

    Google Scholar 

  31. Patt HM, Maloney M, Flannery ML (1982) Hematopoietic microenvironment transfer by stromal fibroblasts derived from bone marrow varying in cellularity. Exp Hematol 10:738–742

    Google Scholar 

  32. Bab I, Ashton BA, Gazit D, Marx G, Williamson MC, Owen ME (1986) Kinetics and differentiation of marrow stromal cells in diffusion chambers in vivo. J Cell Sci 84:139–151

    Google Scholar 

  33. Benayahu D, Kletter Y, Zipori D, Wientroub S (1989) Bone marrow-derived stromal cell line expressing osteoblastic phenotype in vitro and osteogenic capacity in vivo. J Cell Physiol 140:1–7

    Google Scholar 

  34. Bogomoletz WV (1991) Cystic tumours of the exocrine pancreas. In: Anthony PP MacSween RNM (eds) Recent advances in histopathology, vol 15. Churchill Livingstone, Edinburgh, p 141

    Google Scholar 

  35. Charpin C, Bonnier P, Habib MC, Andrac L, Vacheret H, Lavaut MN, Ansaldi JL, Allasia C, Piana L (1992) X-raying of sliced surgical specimens during surgery: an improvement of the histological diagnosis of impalpable breast lesions with microcalcifications. Anticancer Res 12:1737–1746

    Google Scholar 

  36. Wolman M (1991) Minerals pigments and inclusion bodies. In: Filipe MI, Lake BD (eds) Histochemistry in pathology, 2nd ed. Churchill Livingstone, Edinburgh, p 419

    Google Scholar 

  37. Ahmed A (1975) Calcification in human breast carcinomas: ultrastructural observations. J Pathol 117:247–251

    Google Scholar 

  38. Sakuma Y, Mori M (1992) Experimental calcification in rat submandibular gland. Cell Mol Biol 38:413–427

    Google Scholar 

  39. Triatanfallou A, Harrison JD, Garrett JR (1993) Analytical ultrastructural investigation of microliths in salivary glands of cat. Histochem J 25:183–190

    Google Scholar 

  40. Iozzo RV, Cohen I (1993) Altered proteoglycan gene expression and the tumor stroma. Experientia 49:447–455

    Google Scholar 

  41. Bassett CAL, Herrmann I (1961) Influence of oxygen concentration and mechanical factors on differentiation of connective tissues in vitro. Nature 190:460–461

    Google Scholar 

  42. Shaw JL, Bassett CAL (1967) The effects of varying oxygen concentrations on osteogenesis and embryonic cartilage in vitro. J Bone Joint Surg 49A:73–80

    Google Scholar 

  43. Pawelek JM (1969) Effects of thyroxine and low oxygen tension on chondrogenic expression in cell culture. Dev Biol 19:52–72

    Google Scholar 

  44. Allen RG, Balin AK (1989) Oxidative influence in development and differentiation: an overview of a free radical theory of development. Free Rad Biol Med 6:631–661

    Google Scholar 

  45. Yabu M, Takaoka K, Hashimoto J, Fujita H (1992) Immunohis-tochemical, autoradiographic and electron microscopic studies on the transformation of fibroblasts into chondrocytes in the mouse subfascia induced by bone morphogenetic protein. Histochemistry 97:463–468

    Google Scholar 

  46. Murray SS, Murray EJB, Glackin CA, Urist MR (1993) Bone morphogenetic protein inhibits differentiation and affects expression of Helix-Loop-Helix regulatory molecules in myoblastic cells. J Cell Biochem 53:51–60

    Google Scholar 

  47. Centrella M, McCarthy TL, Canalis E (1987) Transforming growth factor β is a bifunctional regulator of replication and collagen synthesis in osteoblast-enriched cell cultures from fetal rat bone. J Biol Chem 262:2869–2874

    Google Scholar 

  48. Seyedin S, Thompson AY, Bentz H, Rosen DM, McPherson JM, Conti A, Siegel NR, Gallupi JR, Piez KA (1986) Cartilage-inducing factor-A. Apparent identity to transforming growth factor-β. J Biol Chem 261:5693–5695

    Google Scholar 

  49. Wozney JM, Rosen V, Celeste AJ, Mitsock LM, Whitters MJ, Kriz RW, Hewick RM, Wang EA (1988) Novel regulators of bone formation: molecular clones and activities. Science 242: 1528–1534

    Google Scholar 

  50. Celeste AJ, Iannazzi JA, Taylor BC, Hewick RM, Rosen V, Wang EA, Wozney JM (1990) Identification of transforming growth factor β family members present in bone-inductive protein purified from bovine bone. Proc Natl Acad Sci USA 87: 9843–9847

    Google Scholar 

  51. Hunter GK, Douglas P, Holmyard DP, Pritzker KPH (1993) Calcification of chick vertebral chondrocytes grown in agarose gels: a biochemical and ultrastructural study. J Cell Sci 104: 1031–1038

    Google Scholar 

  52. Deren JA, Kaplan FS, Brighton CT (1990) Alkaline phosphatase production by periosteal cells at various oxygen tensions in vitro. J Bone Joint Surg 49A:73–80

    Google Scholar 

  53. Brighton CT, Ray R, Soble L, Kuettner K (1969) In vitro epiphyseal plate growth in various oxygen tensions. J Bone Joint Surg 51A:1383

    Google Scholar 

  54. Brighton CT, Krebs A (1972) Oxygen tension of healing fractures in rabbits. J Bone Joint Surg 54A:323

    Google Scholar 

  55. Vaupel P, Kallinowski F, Okunieff P (1989) Blood flow, oxygen and nutrient supply, and metabolic microenvironment of human tumors: a review. Cancer Res 49:6449–6465

    Google Scholar 

  56. Leith JT, Padfield G, Faulkner L, Michelson S (1991) Hypoxic fractions in xenografted human colon tumors. Cancer Res 51: 5139–5143

    Google Scholar 

  57. Mueller-Klieser W, Walenta S (1993) Geographical mapping of metabolites in biological tissue with quantitative bioluminescence and single photon imaging. Histochem J 25:407–420

    Google Scholar 

  58. Massagué J (1958) Transforming growth factors. Isolation, characterization, and interaction with cellular receptors. Prog Med Virol 32:142–158

    Google Scholar 

  59. Centrella M, Massagué J, Canalis E (1986) Human plateletderived transforming growth factor stimulates parameters of bone growth in fetal rat calvariae. Endocrinology 119:2306–2312

    Google Scholar 

  60. Luyten FP, Yu YM, Yanagishita M, Vukicevic S, Glenn Hammonds R, Reddi AH (1992) Natural bovine osteogenin and recombinant human bone morphogenetic protein-2B are equipotent in the maintenance of proteoglycans in bovine articular cartilage explant cultures. J Biol Chem 267:3691–3695

    Google Scholar 

  61. Amitani K, Nakata Y, Stevens J (1974) Bone induction by lyophilized osteosarcoma in mice. Calcif Tissue Res 16:305–313

    Google Scholar 

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Authors and Affiliations

  1. Laboratory of Cell Biology and Histology, Academic Medical Center, University of Amsterdam, Meibergdreef 15, 1105 AZ, Amsterdam, The Netherlands

    C. J. F. Van Noorden, G. N. Jonges, I. M. C. Vogels, K. A. Hoeben & V. Everts

  2. Experimental Oral Biology Group, Department of Periodontology, Academic Medical Center, University of Amsterdam, P.O. Box 22700, 1100 DE, Amsterdam, The Netherlands

    K. A. Hoeben & V. Everts

  3. Animal House Amsterdam, Academic Medical Center, University of Amsterdam, P.O. Box 22700, 1100 DE, Amsterdam, The Netherlands

    B. Van Urk

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  1. C. J. F. Van Noorden
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  3. I. M. C. Vogels
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  4. K. A. Hoeben
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  6. V. Everts
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Van Noorden, C.J.F., Jonges, G.N., Vogels, I.M.C. et al. Ectopic mineralized cartilage formation in human undifferentiated pancreatic adenocarcinoma explants grown in nude mice. Calcif Tissue Int 56, 145–153 (1995). https://doi.org/10.1007/BF00296347

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  • DOI: https://doi.org/10.1007/BF00296347

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