Cell and Tissue Research

, Volume 256, Issue 2, pp 365–372 | Cite as

Specific adhesion between pheochromocytoma (PC12) cells and adrenal medullary endothelial cells in co-culture

  • Yaffa Mizrachi
  • Peter I. Lelkes
  • Richard L. Ornberg
  • Gertrude Goping
  • Harvey B. Pollard


Chromaffin cells in the adrenal medulla are found in close proximity to capillary endothelial cells, thereby forming the classical endocrine complex. To examine the possible chemical basis of their interaction in more detail, we have grown bovine adrenal medullary endothelial (BAME) cells in monolayer cultures and added to them pheochromocytoma (PC12) cells, a chromaffin tumor cell line of rats. The PC12 cells were chosen because of the similarities they share with adrenal medullary chromaffin cells. PC12 cells rapidly attached to BAME cells cultures, their rate of adhesion being significantly enhanced over binding of PC12 cells to either uncoated plates or to monolayers of unrelated cell cultures. Consistent with this observation, we noted that the extracellular matrix (ECM) derived from the BAME cells did not enhance PC12 cell adhesion and did not promote neurite sprouting as previously described for ECM derived from corneal endothelial cells. The specific adhesion between PC12 and BAME cells could be abolished by cell surface extracts derived from these two cells but not by extracts derived from unrelated cell types. This activity was heat-labile, sensitive to trypsin and, to a lesser extent, to neuraminidase. We therefore conclude that PC12 cells may interact with BAME cells by specific proteinaceous adhesive factors associated with their plasma membranes. These interactions might represent the formative role of cell-cell contacts in the organization of the developing adrenal gland.

Key words

Adrenal medullary endothelial cells Pheochromocytoma (PC12) cells Co-culture Cell surface extracts Adhesion Cell-cell interactions Bovine Rat 



bovine adrenal medullary endothelial cells


Dulbecco's modified essential medium


extracellular matrix


Eagle's modified essential medium


fetal calf serum


phosphate-buffered saline


rat pheochromocytoma cells


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Akeson R, Warren SL (1986) PC12 adhesion and neurite formation on selected substrates are inhibited by some glycosaminoglycans and a fibronectin-derived tetrapeptide. Exp Cell Res 162:347–362Google Scholar
  2. Alitalo K, Kuismanen E, Myllylia R, Kiistala U, Asko-Seljavaara S, Vaheri A (1982) Extracellular matrix proteins of human epidermal keratinocytes and feeder 3T3 cells. J Cell Biol 94:497–505Google Scholar
  3. Banerjee KD, Ornberg RL, Youdim MBH, Heldman E, Pollard HB (1985) Endothelial cells from bovine adrenal medulla develop capillary-like growth patterns in culture. Proc Natl Acad Sci USA 82:4702–4706Google Scholar
  4. Bethea CL, Kozak SL (1984) Effect of extracellular matrix on PC12 cell shape and dopamine processing. Mol Cell Endocrinol 37:391–429Google Scholar
  5. Bischoff R (1986) Rapid adhesion of nerve cell to muscle fibers from adult rats is mediated by sialic acid-binding receptor. J Cell Biol 102:2273–2280Google Scholar
  6. Bozyczko D, Horwitz AF (1986) The participation of a putative cell surface receptor for laminin and fibronectin in peripheral neurite extension. J Neurosci 6:1241–1251Google Scholar
  7. Carmichael SW, Spangroli DB, Frederickson R, Krause WJ, Gulberson JL (1987) The opossum adrenal medulla; I. Postnatal development and normal anatomy. Am J Anat 179:211–219Google Scholar
  8. Castellot JJ, Rosenberg RD, Karnovsky MJ (1984) Endothelium, heparin and regulation of vascular smooth muscle growth. Jaffe EA (ed) Biology of Endothelial Cells. Martinus Nijhoff Publishers, pp 118–128Google Scholar
  9. Cole GJ, Loewy A, Glaser L (1986) Neuronal cell adhesion depends on interactions of N-CAM with heparin-like molecules. Nature 320:445–449Google Scholar
  10. Collins F (1980) Neurite outgrowth induced by the substrate associated material from non-neuronal cells. Dev Biol 79:247–252Google Scholar
  11. Darnule AT, Stozky G, Darnule TV, Turino GM, Mandl I (1983) Identification of surface antigens of endothelial cells. Immunol Commun 12:351–362Google Scholar
  12. Doupe AJ, Landis SC, Patterson PH (1985) Environmental influences in the development of neural crest derivatives: Glucocorticoids, growth factors and chromaffin cell plasticity. J Neurosci 5:2119–2142Google Scholar
  13. Dubois C, Magnani JL, Grunwald GB, Spitalnik SL, Trisler GD, Nirenberg M, Ginzburg V (1986) Monoclonal antibody 18B8, which detects synapse-associated antigens, binds to ganglioside GT3 (II3[NeuAC]3 Laccer). J Biol Chem 261:3826–3830Google Scholar
  14. Edelman GM (1983) Cell adhesion molecules. Science 219:450–457Google Scholar
  15. Forsberg EJ, Feuerstein G, Shohami E, Pollard HB (1987) Adenosine triphosphate stimulates inositol phospholipid metabolism and prostacyclin formation in adrenal medullary endothelial cells via P2-purinergic receptors. Proc Natl Acad Sci USA 84:5630–5634Google Scholar
  16. Gerritsen ME (1987) Functional heterogeneity of vascular endothelial cells. Pharmacology 36:2701–2711Google Scholar
  17. Greene LA, Tischler AS (1976) Establishment of noradrenergic clonal line of rat adrenal pheochromocytoma cells which respond to nerve growth factor. Proc Natl Acad Sci USA 73:2424–2428Google Scholar
  18. Grumet M, Hoffman S, Grossin KL, Edelman GM (1985) Cytotactin, an extracellular matrix protein of neural and non-neural tissue that mediate glianeuron interactions. Proc Natl Acad Sci USA 82:8075–8079Google Scholar
  19. Heimark RL, Schwartz SM (1985) The role of membrane-membrane interactions in the regulation of endothelial cell growth. J Cell Biol 100:1934–1940Google Scholar
  20. Janzer RC, Raff MC (1987) Astrocytes induce blood-brain barrierproperties in endothelial cells in vitro. Nature 325:253–257Google Scholar
  21. Kelner KL, Burns L, Pollard HB, MacRae R, Bakry LA, Srivastiva M (1987) Steroid control of neurotransmitter synthesis: Glucocorticoid regulation of phenylethanolamine-N-methyl transferase. In: Stolk F, Richard DC, Fuxe A (eds) Neuronal Epinephrine (In press)Google Scholar
  22. Kishi M, Nakajo S, Shibayama T, Nakaya TK, Nakamura Y (1980) Cell surface proteins extracted with urea from Ah-66 hepatoma ascites cells. J Biochem 87:135–141Google Scholar
  23. Lander AD, Fujii DK, Reichardt LF (1985) Purification of a factor that promotes neurite outgrowth: isolation of laminin and associated molecules. J Cell Biol 101:898–913Google Scholar
  24. Landreth GE, Williams LK, McCutchen C (1985) Wheat germ agglutinin blocks the biological effects of nerve growth factor. J Cell Biol 101:1690–1694Google Scholar
  25. Langley OK, Aletsee MC, Gratzl M (1987) Endocrine cells share expression of N-CAM with neurons. FEBS Lett 220:108–112Google Scholar
  26. Lelkes PI, Mizrachi Y, Ornberg R, Naranjo J, Pollard HB (1987) Specific interaction between PC12 cells and adrenal medullary endothelial cells in coculture: An in vitro model for early stages of developmental differentiation in neurosecretion. Neuroscience 22a: 818Google Scholar
  27. Manthorpe M, Engvall E, Ruoslahti E, Longo FM, Davis GE, Varon S (1983) Laminin promotes neuritic regeneration from cultured peripheral and central neurons. J Cell Biol 97:1882–1890Google Scholar
  28. Mizrachi Y, Lelkes PI, Pollard HB (1986) PC12 cells and adrenal medullary endothelial cells form complexes in co-culture analogous to chromaffin cell endothelial interactions seen in mature adrenal medulla tissue. Soc Neurosci [Abstr] 320:10Google Scholar
  29. Pappas GD, Sagan J (1986) Fine structure of PC12 cell implants in the rat spinal cord. Neurosci Lett 70:59–64Google Scholar
  30. Patterson PH (1978) Environmental determination of autonomic neurotransmitter function. Ann Rev Neurosci 1:1–17Google Scholar
  31. Pollard HB, Ornberg R, Levine M, Kelner K, Morita K, Levine R, Forsberg E, Brocklehurst KW, Doung L, Lelkes PI, Heldman E, Youdim M (1985) Hormone secretion by exocytosis with emphasis on information from the chromaffin cell system. Vitam Horm 42:109–195Google Scholar
  32. Ramachandran CK, Sanders K, Melnykovich G (1986) Enhancement in the adhesion of tumor cells to endothelial cells by decreased cholesterol synthesis. Cancer Res 46:2520–2525Google Scholar
  33. Risau W, Hallmann R, Albrecht U, Henke-Fahle S (1986) Brain induces expression of an early cell surface marker for blood-brain barrier-specific endothelium. EMBO J 5:3179–3183Google Scholar
  34. Siegelman M, Bond MW, Gallatin WM, St John T, Smith HT, Fried VA, Weissman IL (1986) Cell surface molecule associated with lymphocytes homing is a ubiquitinated branched chain glycoprotein. Science 231:823–825Google Scholar
  35. Tanford C (1980) The hydrophobic effect: Formation of micelles and biological membranes. John Wiley and Sons, New YorkGoogle Scholar
  36. Thiery JF, Duband JL, Tucker GC (1985) Cell migration in the vertebrate embryo. Role of cell adhesion and tissue environment in pattern formation. Ann Rev Cell Biol 1:91–113Google Scholar
  37. Tischler AS, Greene LA (1980) Phenotypic plasticity of pheochromocytoma and normal adrenal medullary cells. Adv Biochem Psychopharmacol 25:61–68Google Scholar
  38. Trisler D, Benekstein J, Danies MP (1986) Antibody to a molecular marker of cell position inhibits synapse formation in the retina. Proc Natl Acad Sci USA 83:4194–4198Google Scholar
  39. Vlodavsky I, Levi A, Lax I, Fuxs Z, Schlessinger J (1982) Induction of cell attachment and morphological differentiation in pheochromocytoma cell line and embryonal sensory cells by extracellular matrix. Dev Biol 93:285–300Google Scholar
  40. Waymire JC, Bennett WF, Boehme R, Hawkins I, Gilmer-Waymire K, Haycock JW (1983) Bovine adrenal chromaffin cell high yield purification and viability in suspension culture. JNeurosci Methods 7:323–351Google Scholar
  41. Whittenberger B, Raben D, Lieberman MA, Glaser L (1978) Inhibition of growth of 3T3 cells by extract of surface membranes. Proc Natl Acad Sci USA 75:5457–5461Google Scholar
  42. Youdim MB, Banerjee DK, Pollard HB (1984) Isolated chromaffin cells from adrenal medulla contain primarily monamine oxidase B. Science 224:619–621Google Scholar

Copyright information

© Springer-Verlag 1989

Authors and Affiliations

  • Yaffa Mizrachi
    • 1
  • Peter I. Lelkes
    • 1
  • Richard L. Ornberg
    • 1
  • Gertrude Goping
    • 1
  • Harvey B. Pollard
    • 1
  1. 1.Laboratory of Cell Biology and Genetics, NIDDK, National Institutes of HealthBethesdaUSA

Personalised recommendations