Summary
The human carcinoma line RPMI 2650 produces autocrine factors; they are detected by the ability of RPMI 2650 conditioned medium (CM) to stimulate growth in soft agar of RPMI 2650 cells plated at low density. The autocrine activity in crude CM can be fractionated by ultrafiltration into a lower molecular weight (MW) fraction (R1–30), which concentrates molecules in the 1000–30 000 Da range; and a higher MW fraction (R30) with molecules greater than 30 000 Da in a more concentrated form. R1–30 is labile to acid, base, and heat treatment, whereas R30 is stable to (and sometimes activated by) these treatments. Boiling of R30, however, renders it labile to acid, base, and trypsin treatments. CM can be separated into a weakly heparin-binding fraction (with stability properties similar, but not identical, to R1–30), and a non-heparin binding fraction (with stability properties similar to R30). RPMI 2650 cells secrete transforming growth factor (TGF)α- and TGFβ-like molecules, but the R1–30 fraction can be distinguished from these TGFs, and from most other known growth factors, by its unusual combination of acid lability and weak affinity for heparin. Since the R30/non-heparin binding fraction is rendered labile by boiling or acid treatment, it may represent a bound or conformationally stable form of a growth factor.
Similar content being viewed by others
References
Arai, K.; Lee, F.; Miyajima, A., et al. Cytokines: co-ordinators of immune and inflammatory responses. Ann. Rev. Biochem. 59:783–836; 1990.
Bazill, G. W.; Haynes, M.; Garland, J., et al. Characterization and partial purification of a haemopoietic cell growth factor in WEH1-3 cell conditioned medium. Biochemistry J. 210:747–759; 1983.
Blat, C.; Villaudy, J.; Delbe, J., et al. Purification from transformed mouse fiber of a cell growth inhibitor which is an IGF-binding protein. Growth Factors 6:65–75; 1992.
Browder, T. M.; Dunbar, C.; Nierhuis, A. Private and public autocrine loops in neoplastic cells. Cancer Cells 1:9–17; 1989.
Brown, C. A.; Halper, J. Mitogenic effects of transforming growth factor type e on epithelial and fibroblastic cells—comparison with other growth factors. Exp. Cell Res. 190:233–242. 1990.
Clark-Lewis, I.; Kent, S. B. H.; Schrader, J. W. Purification to apparent homogeneity of a factor stimulating the growth of multiple lineages of hemopoietic cells. Biol. Chem. 259:7488–7494; 1983.
Clynes, M.; Dooley, M.; Carey, B., et al.In vitro production of growth regulators for human epithelial cells. In: Spier, R. E.; Griffiths, J. B.; Meignier, B., eds. Production of biologicals from animal cells in culture. London: Butterworth-Heineman; 1991:932–734.
DeLarco, J. E.; Todaro, G. J. Growth factors from murine sarcoma virus transformed cells. Proc. Natl. Acad. Sci. USA 75:4001–4005; 1978.
Dennis, P. A.; Sabsela, O.; Harper, P., et al.α 2-Macroglobulin is a binding protein for basic fibroblast growth factor. J. Biol. Chem. 264:7210–7216; 1989.
Derynck, R.; Balentien, E.; Han, J. M., et al. Recombinant expression, biochemical characterisation and biological activities of the human MGSA/gro protein. Biochemistry 29:10225–10233; 1990.
Deuel, T. F. Polypeptide growth factors: roles in normal and abnormal cell growth. Ann. Rev. Cell Biol. 3:443–492; 1987.
Dunnington, D. J.; Scott, R. G.; Anzano, M. A., et al. Characterisation and partial purification of human epithelial transforming growth factor. J. Cell. Biochem. 44:229–239; 1990.
Finch, P. W.; Rubin, J. S.; Miki, T., et al. Human KGF is FGF-related with properties of a paracrine effector of epithelial cell growth. Science 245:752–755; 1989.
Glovine, F. S.; Duft, G. W. Molecular and functional aspects of human PRFs. In: Peptide regulatory factors. London: Lancet Series, Edward Arnold; 1989:125–155.
Heldin, C. H.; Westermark, B. Growth factors as transforming proteins. Eur. J. Biochem. 184:487–496; 1989.
Higashiyama, S.; Abraham, J. A.; Miller, J., et al. A heparin-binding growth factor secreted by macrophage-like cells that is related to EGF. Science 251:936–938; 1991.
Hilton, D. J.; Nicola, N. A.; Metcalf, D. Purification of a murine leukemia inhibitory factor from Krebs ascites cells. Anal. Biochem. 173:359–367; 1988.
Hirai, R.; Yamanoaka, K.; Mitsui, H. Isolation and partial purification of a new class of transforming growth factor from an avian sarcoma virus-transformed rat cell line. Cancer Res. 43:5742–5746; 1983.
Hirano, T.; Taga, T.; Nakano, N., et al. Purification to homogeneity and characterization of human B-cell differentiation factor (BCDF or BSFp-2). Proc. Natl. Acad. Sci. USA 82:5490–5494; 1985.
Huang, S.; O’Grady, P.; Huang, J. Human transforming growth factor-β α 2-macroglobulin complex is a latent form of transforming growth factorβ. J. Biol. Chem. 263:1535–1541; 1988.
Huang, J. S.; Huang, S. S.; Deuel, T. F. Specific covalent binding of platelet-derived growth factor to human plasmaα 2-macroglobulin. Proc. Natl. Acad. Sci. USA 81:342–346; 1984.
Jaye, M.; Burgess, W. H.; Shaw, A. B., et al. Biological equivalence of natural bovine and recombinant humanα-endothelial cell growth factors. J. Biol. Chem. 262:16612–16617; 1987.
Jubinsky, P. T.; Stanley, E. R. Purification of hemopoietin 1: A multilineage hemopoietic growth factor. Proc. Natl. Acad. Sci. USA 82:2764–2768; 1985.
Jullien, P.; Berg, T. M.; Lawrence, D. A. Acidic cellular environments: activation of latent TGFβ and sensitisation of cellular responses to TGFβ and EGF. Int. J. Cancer 43:886–891; 1989.
Li, Y.; Milner, P. G.; Chauhan, A. K., et al. Cloning and expression of a developmentally regulated protein that induces mitogenic and neurite outgrowth activity. Science 250:1690–1694; 1990.
Lobb, R. R.; Harper, J. W.; Fett, J. W. Purification of heparin-binding growth factors. Anal. Biochem. 154:1–14; 1986.
Lopez-Casillas, F.; Cheifetz, S.; Doody, J., et al. Structure and expression of the membrane proteoglycan betaglycan, a component of the TGF-β receptor system. Cell 67:785–795; 1991.
Lyons, R. M.; Moses, H. L. Transforming growth factors and the regulation of cell proliferation. Eur. J. Biochem. 187:467–473; 1990.
Merenmies, J.; Rauvala, H. Molecular cloning of an 18 kD growth-associated protein of developing brain. J. Biol. Chem. 265:16721–16724; 1990.
Miyazono, K.; Hellman, V.; Wernstedt, C., et al. Latent high molecular weight complex of transforming growth factor-β1. J. Biol. Chem. 263:6407–6415; 1988.
Moore, G.; Sandberg, A. Studies of a human tumor cell line, with a diploid karyotype. Cancer 117:170–173; 1963.
Moses, H. L.; Yong, E. Y.; Pietenpol, J. TGF-β stimulation and inhibition of cell proliferation: new mechanistic insights. Cell 63:245–247; 1990.
Myoken, Y.; Kayada, Y.; Okamoto, T., et al. Vascular endothelial cell growth factor (VEGF) produced by A431 human epidermoid carcinoma cells and identification of VEGF membrane binding sites. Proc. Natl. Acad. Sci. USA 88:5819–5823; 1991.
Namen, A. E.; Schmierer, A. E.; March, C. J., et al. B-cell precursor growth-promoting activity. Purification and characterization of a growth factor active on lymphocyte precursors. J. Exp. Med. 167:988–1002; 1988.
Nicola, N. A.; Metcalf, D.; Matsumoto, M., et al. Purification of a factor inducing differentiation in murine myelomonocytic leukemia cells. Biol. Chem. 258:9017–9023; 1983.
O’Connor-McCourt, M. D.; Wakefield, L. M. Latant transforming growth factor-β in serum. J. Biol. Chem. 262:14090–14099; 1987.
Ohara, J.; Lahet, S.; Inman, J., et al. Partial purification of murine B cell stimulatory factor (BSF)-1. Immunology 135:2518–2523; 1985.
Peveri, P.; Walz, A.; Dewald, B., et al. A novel neutrophil-activating factor produced by human mononuclear phagocytes. J. Exp. Med. 167:1547–1559; 1988.
Rizzino, A.; Ruff, E. Fibroblast growth factor induces the soft agar growth of non-transformed cell lines. In Vitro Cell. Dev. Biol. 22:749–755; 1986.
Roberts, R.; Gallagher, J.; Spooner, E., et al. Heparin sulphate-bound growth factors, a mechanism for cell mediated haemopoiesis. Nature 332:376–378; 1988.
Rowe, J. M.; Kasper, S.; Shiu, R. P. C., et al. Purification and characterisation of a human tumor-derived growth factor. Cancer Res. 46:1408–1412; 1986.
Rubin, J. S.; Osada, H.; Finch, P. W., et al. Purification and characterisation of a newly identified growth factor for epithelial cells. Proc. Natl. Acad. Sci. USA 86:802–806; 1989.
Sasoki, R.; Yanagawa, S. I.; Chuba, H. Isolation of human erythropoietin with monoclonal antibodies. Methods Enzymol. 147:328–340; 1987.
Sauder, D. N.; Stanulis-Praeger, B. M.; Gilchrest, B. A. Autocrine growth stimulation of human keratinocytes by epidermal cell-derived thymocyte-activating factor: implications for skin aging. Arch. Dermatol. Res. 280:71–76; 1988.
Schroder, J. M.; Mrowietz, U.; Christophers, E. Identification of different charged species of a human monocyte derived neutrophil activating peptide (MONAP). Biochem. Biophys. Res. Commun. 152:277–284; 1988.
Sero, M.; Sasada, R.; Kurokawa, T., et al. Carboxyl-terminal structure of basic fibroblast growth factor significantly contributes to its affinity for heparin. Eur. J. Biochem. 188:239–245; 1990.
Shearer, M.; Taylor-Popadimitrou, J. Regulation of cell growth by interferon. Cancer Metastasis Rev. 6:199–221; 1987.
Sherry, B.; Cerami, A. Cachectin/tumor necrosis factor exerts endocrine, paracrine and autocrine control of inflammatory responses. J. Cell. Biol. 107:1270–1277; 1988.
Slack, J. M. W.; Peptide regulatory factors in embryonic development. In: Peptide regulatory factors. London: Lancet Series, Edward Arnold; 1989 :91–101.
Sporn, M. B.; Roberts, A. B. Peptide growth factors are multifunctional. Nature 332:217–219; 1988.
Stern, A. S.; Pan, Y. E.; Urdal, D. L., et al. Purification to homogeneity and partial characterization of interleukin 2 from a human T-cell leukemia. Proc. Natl. Acad. Sci. USA 81:871–875; 1984.
Takatsu, K.; Kikuchi, Y.; Takahashi, T., et al. Interleukin 5, a T-cell-derived B-cell differentiation factor also induces cytotoxic T lymphocytes. Proc. Natl. Acad. Sci. USA 84:4234–4238; 1987.
Van Zoelan, E. Phenotypic transformation of normal rat kidney cells: a model for studying cellular alterations in oncogenesis. Crit. Rev. Oncogenesis 2:311–333; 1991.
Watson, J. D.; Crossier, P. S.; March, C. J., et al. Purification to homogeneity of a human hematopoietic growth factor that stimulates the growth of a murine interleukin 3-dependent cell line. J. Immunol. 137:854–857; 1986.
Wu, D.; Kan, M.; Sato, G. M., et al. Characterization and molecular cloning of a putative binding protein for heparin-binding growth factors. J. Biol. Chem. 266:16778–16785; 1991.
Zarling, J. M.; Shoyab, M.; Marguert, H., et al. A growth regulator produced by differentiated histocytic lymphoma cells. Proc. Natl. Acad. Sci. USA 83:9739–9743; 1986.
Zarnegar, R.; Michalopoulos, G. Purification and biological characterisation of human hepatopoietin A, a polypeptide growth factor for hepatocytes. Cancer Res. 49:3314–3320; 1989.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Carey, B.M., Dooley, M., Weedle, R. et al. Production of autostimulatory growth factors by the human carcinoma line, RPMI 2650. In Vitro Cell Dev Biol - Animal 29, 153–160 (1993). https://doi.org/10.1007/BF02630947
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02630947