Summary
We have compared PC12 cell lines derived from different laboratories and the newly developed mouse pheochromocytoma (MPC) cell line. Morphologically, there were distinct differences in size, shape, adherence, and clumping behavior, which varied in response to different culture media, growth substrates, and nerve growth factor. Quantitative messenger ribonucleic acid (mRNA) analysis showed significant variability in the expression of the catecholaminergic biosynthetic enzymes tyrosine hydroxylase (TH) phenylethanolamine N-methyltransferase (PNMT), the noradrenaline transporter (NAT), and neuron-specific enolase (NSE) between all lines examined. Of most significance were the increased levels of PNMT mRNA in the MPC cells, which were to 15-fold greater than in the PC12 cell lines grown under the same conditions in Dulbecco modified Eagle medium (P<0.05). Growth of MPC cells in Roswell Park Memorial Institute media induced a further significant increase in PNMT gene expression (P≤0.05). Immunohistochemistry for TH, PNMT. and NAT was generally consistent with mRNA analysis, with the MPC cells demonstrating strong immunoreactivity, for PNMT. The MPC cells showed the highest levels of desipramine-sensitive [3H] noradrenaline uptake activity (threefold > than PC12 American Type Culture Center line, P≤0.05), despite relatively low levels of NAT mRNA. These results indicate that PC12 cell lines should be carefully chosen for optimal utility in the study of chromaffin cell or sympathetic neuron biology and that cell features will be influenced by type of media and substrate chosen. Furthermore, they confirm that the new MPC cell line is likely a useful model for the study of adrenergic mechanisms or studies involving NAT.
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References
Acheson, A. L.; Naujoks, K.; Thoenen, H., Nerve growth factor-mediated enzyme induction in primary cultures of bovine adrenal chromaffin cells: specificity and level of regulation. J. Neurosci. 4:1771–1780; 1984.
Akeson, R.; Warren, S. L. PC12 adhesion and neurite formation on selected substrates are inhibited by some glycosaminoglycans and a fibronectin-derived tetrapeptide. Exp. Cell Res. 162:347–362; 1986.
Andreassi, J. L.; Eggleston, W. B., Fu, G.; Stewart, J. K.. Phenylethanolamine N-methyltransferase mRNA in rat hypothalamus and cerebellum. Brain Res. 779:289–291; 1998.
Bar-Sagi, D.; Feramisco, J. R. Microinjection of the ras oncogene protein into PC12 cells induces morphological differentiation. Cell 42:841–848; 1985.
Beaujean, D.; Rosenbaum, C.; Muller, H. W.; Willemsen, J. J.; Lenders, J.; Bornstein, S. R. Combinatorial code of growth factors and neuropeptides define neuroendocrine differentiation in PC12 cells. Exp. Neurol. 184:348–358; 2003.
Bitler, C. M.; Zhang, M. B.; Howard, B. D. PC12 variants deficient in catecholamine transport. J. Neurochem. 47:1286–1293; 1986.
Bothwell, M. A.; Schechter, A. L.; Vaughn, K. M.. Clonal variants of PC12 pheochromocytoma cells with altered response to nerve growth factor. Cell 21:857–866; 1980.
Bruss, M.; Porzgen, P.; Bryan-Lluka, L. J.; Bonisch, H. The rat norepinephrine transporter. molecular cloning from PC12 cells and functional expression. Mol. Brain Res. 52:257–262; 1997.
Burton, L. D.; Kippenberger, A. G.; Lingen, B., Bruss, M.; Bonisch, H.; Christie, D. L. A variant of the bovine noradrenaline transporter reveals the importance of the C-terminal region for correct targeting to the membrane and functional expression. Biochem. J. 330:909–914; 1998.
Cahill, A. L.; Eertmoed, A. L.; Mangoura, D.; Perlman, R. L., Differential regulation of phenylethanolamine N-methyltransferase expression in two distinct subpopulations of bovine chromaffin cells. J. Neurochem. 67:1217–1224; 1996.
Comer, A. M.; Qi, J. G.; Christie, D. L.; Gibbons, H. M.; Lipski, J. Noradrenaline transporter expression in the pons and medulla oblongata of the rat—localisation to noradrenergic and some C1 adrenegic neurones. Mol. Brain Res. 62:65–76; 1998.
Comer, A. M.; Yip, S.; Lipski, J. Detection of weakly expressed genes in the rostral ventrolateral medulla of the rat using micropunch and RT-PCR techniques. Clin. Exp. Hypertens. 24:755–759; 1997.
Corey, J. M.; Feldman, E. E. Substrate patterning: an emerging technology for the study of neuronal behavior. Exp. Neurol. 184 (Suppl. 1): S89-S96; 2003.
Dragunow, M.; Xu, R.; Walton, M., et al. c-Jun promotes neurite outgrowth and survival in PC12 cells. Mol. Brain Res. 83:20–33; 2000.
Eaton, M. J.; Duplan, H. Useful cell lines derived from the adrenal medulla. Mol. Cell. Endocrinol. 228:39–52; 2004.
Edwards, M. A.; Loxley, R. A.; Powers-Martin, K.; Lipski, J.; McKitrick, D. J.; Arnolda, L. F.; Phillips, J. K. Unique levels of expression of N-Methyl-D-Aspartate receptor subunits and neuronal nitric oxide synthase in the rostral ventrolateral medulla of the spontaneously hypertensive rat. Mol. Brain Res. 129:33–43; 2004.
Evinger, M. J.; Cikos, S.; Nwafor-Anene, V.; Powers, J. F.; Tischler, A. S. Hypoxia activates multiple transcriptional pathways in mouse pheochromocytoma cells. Ann. N.Y. Acad. Sci. 971:61–65; 2002.
Fujii, D. K.; Massoglia, S. L.; Savion, N.; Gospodarowicz, D.. neurite outgrowth and protein synthesis by PC12 cells as a function of substratum and nerve growth factor. J. Neurosci. 2:1157–1175; 1982.
Fukuda, M.; Gotoh, Y.; Tachibana, T.; Dell, K.; Hattori, S.; Yoneda, Y.; Nishida, E.. Induction of neurite outgrowth by MAP kinase in PC12 cells. Oncogene 11:239–244; 1995.
Genda, T.; Sakamoto, M.; Ichida, T.; Asakura, H.; Hiohashi, S. Loss of cellcell contact is induced by integrin-mediated cell substratum adhesion in highly-motile and highly-metastatic hepatocellular carcinoma cells. Lab. Invest 80:387–394; 2000.
Greene, L. A.; Rein, G. Release, storage and uptake of catecholamines by a clonal cell line of nerve growth factor (NGF) responsive pheo-chromocytoma cells. Brain Res, 129:247–263; 1977.
Greene, L. A.; Tischler, A. S. Estalishment of a noradrenergic clonal line of rat adrenal pheochromocytoma cells which respond to nerve growth factor. Proc. Natl. Acad. Sci. USA 73:2424–2428; 1976.
Harder, R.; Bonisch, H. Effects of monovalent ions on the transport of noradrenaline across the plasma membrane of neuronal cells (PC-12 cells). J. Neurochem. 45:1154–1162; 1985.
Kippenberger, A. G.; Palmer, D. J.; Comer, A. M.; Lipski, J.; Burton, L. D.; Christie, D. L. Localization of the noradrenaline transporter in rat adrenal medulla and PC12 cells: evidence for its association with secretory granules in PC12 cells. J. Neurochem. 73:1024–1032; 1999.
Leoni, C.; Menegon, A.; Benfenati, F.; Toniolo, D.; Pennuto, M.; Valtorta, F. neurite extension occurs in the absence of regulated exocytosis in PC12 subclones. Mol. Biol. Cell 10:2919–2931; 1999.
Leppa, S.; Saffrich, R.; Ansorge, W.; Bohmann, D. Differential regulation of c-Jun by ERK and JNK during PC12 cell differentiation. EMBO J. 17:4404–4413; 1998.
Livak, K. J.; Schmittgen, T. D. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T) method. Methods 25:402–408;2001.
Marangos, P. J.; Zis A. P.; Clark, R. L.; Goodwin, F. K. Neuronal, nonneuronal and hybrid forms of enolase in brain: structural, immunological and functional comparisons. Brain Res. 150:117–133; 1978.
Naranjo-Suarez, S.; Castellanos, M. C.; Alvarez-Tejado, M.; Vara, A.; Landazuri, M. O.; del Peso, L.: Down-regulation of hypoxia-inducible factor-2 in PC12 cells by nerve growth factor stimulation. J. Biol. Chem. 278:31895–31901; 2003.
Phillips, J. K.; Dubey, R.; Sesiashvili, E.; Takeda, M.; Christie, D. L.; Lipski, J. Differential expression of the noradrenaline transporter in adrenergic chromaffin cells, ganglion cells and nerve fibres of the rat adrenal medulla. J. Chem. Neuroanat. 21:95–104; 2001.
Powers, J. F.; Evinger, M. J.; Tsokas, P.; Bedri, S. Alroy, J.; Shahsavari, M. Tischler, A. S. Pheochromocytoma cell lines from heterozygous neurofibromatosis knockout mice. Cell Tissue Res. 302:309–320; 2000.
Powers, J. F.; Schelling, K. H.; Brachold, J. M.; Tischler, A. S.. Plasticity of pheochromocytoma cell lines from neurofibromatosis knockout mice. Ann. N.Y. Acad. Sci. 971:371–378; 2002.
Ramachandran, B.; Houben, K.; Rozenberg, Y. Y.; Haigh, J. R.; Varpetian, A.; Howard, B. D. Differential expression of transporters for norepinephrine and glutamate in wild type, variant, and WNT1-expressing PC12 cells. J. Biol. Chem. 268:23891–23897; 1993.
Rao, R. R.; Kisaalita, W. S. Biochemical and electrophysiological differentiation profile of a human neuroblastoma (IMR-32) cell line. In Vitro Cell Dev. Biol. 38A:450–456; 2002.
Tischler, A. S. Chromaffin cells as models of endocrine cells and neurons. Ann. N. Y. Acad. Sci. 971:366–370; 2002.
Tischler, A. S.; Greene, L. A. Nerve growth factor-induced process formation by cultured rat pheochromocytoma cells. Nature 258:341–342; 1975.
Tischler, A. S.; Perlman, R. L.; Morse, G. M.; Sheard, B. E. Glucocorticoids increase catecholamine synthesis and storage in PC12 pheochromocytoma cell cultures. J. Neurochem. 40:364–370; 1983.
Tischler, A. S.; Powers, J. F.; Alroy, J. Animal models of pheochromocytoma. Histol. Histopathol. 19:883–895; 2004.
Turner, D. C.; Flier, L. A.; Carbonetto, S. Magnesium-dependent attachment and neurite outgrowth by PC12 cells on collagen and laminin substrata. Dev. Biol. 121:510–525; 1987.
Uchida, J.; Kiuchi, Y.; Ohno M.; Yura, A.; Oguchi, K. Ca(2+)-dependent enhancement of [3H]noradrenaline uptake in PC12 cells through calmodulin-dependent kinases. Brain Res. 809:155–164; 1998.
Unsworth, B. R.; Hayman, G. T.; Carroll A.; Lelkes, P. I. Tissue-specific alternative mRNA splicing of phenylethanolamine N-methyltransferase (PNMT) during development by intron retention. Int. J. Dev. Neurosci. 17:45–55; 1999.
Xiao, J.; Zhou, Q.; Liu, Y. Variant PC12 cell line that spontaneously differentiates and extends neuritic processes. J. Neurosci. Res. 69:104–109; 2002.
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Dixon, D.N., Loxley, R.A., Barron, A. et al. Comparative studies of PC12 and mouse pheochromocytoma-derived rodent cell lines as models for the study of neuroendocrine systems. In Vitro Cell.Dev.Biol.-Animal 41, 197–206 (2005). https://doi.org/10.1290/0411077.1
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DOI: https://doi.org/10.1290/0411077.1