Regulation of Bcl-2 oncoprotein levels with differentiation of human neuroblastoma cells Journal Article
| Authors: | Hanada, M.; Krajewski, S.; Tanaka, S.; Cazals-Hatem, D.; Spengler, B. A.; Ross, R. A.; Biedler, J. L.; Reed, J. C. |
| Article Title: | Regulation of Bcl-2 oncoprotein levels with differentiation of human neuroblastoma cells |
| Abstract: | When established in culture, human neuroblastoma cell lines typically are comprised of heterogeneous cellular subpopulations, including neuroblastic (N-type), substrate-adherent (S-type), and intermediate (I-type) cells that can be distinguished by their characteristic morphologies and expression of differentiation-associated antigens. Here we examined the relative levels of the Bcl-2 oncoprotein in 15 clones derived from four different neuroblastoma cell lines. Among six clones isolated from the SK-N-SH line, levels of p26-Bcl-2 correlated with morphology and differentiation markers with the hierarchy of bcl-2 expression being: N-type cells > N/I-type > I-type > S-type. Furthermore, stimulation of one of the N-type clones, SH-SY5Y, with the phorbol ester, 12-O-tetradecanoylphor-bol-13-acetate, induced differentiation toward a more neuronal-like phenotype and resulted in a 5- to 10-fold elevation in the relative levels of Bcl-2 protein. High relative amounts of p26-Bcl-2 protein were also found in an N-type done derived from the SMS-KCN line. In two N-type clones derived from the LA-N-1 line, however, levels of Bcl-2 protein were only moderately elevated, and in one N-type done from the SK-N-BE(2) line the levels of Bcl-2 protein were low. Thus, high relative levels of Bcl-2 oncoprotein are not a universal feature of N-type cells (three of six clones tested). In contrast, all 5 of the S-type clones evaluated contained relatively low levels of Bcl-2 protein, suggesting that these cells (which may represent embryonic precursors of Schwann, glial, and melanocytic cells) do not typically express the bcl-2 gene at high levels. Consistent with this inverse correlation between Bcl-2 protein levels and S-type characteristics, stimulation of an I-type done derived from the SK-N-BE(2) line with 5-bromodeoxyuridine was accompanied by an accumulation of S-type cells in these cultures, decreased Bcl-2 protein, diminutions in the neuronal markers neurofilament-M and neuron-specific enclase, and an increase in the relative levels of the S-type marker proteins vimentin and β-2-mcroglobulin. Conversely, stimulation of this I-type done with retinoic acid resulted in an accumulation of N-type cells (which are thought to represent embryonic precursors of sympathetic neurons), decreased vimentin and β-2-mcroglobulin, increased neurofilament-M, and a marked elevation in p26-Bcl-2. lb begin to explore the functional significance of variations in the levels of Bcl-2 protein among neuroblastomas, a cell line with I-type characteristics and relatively low levels of p26-Bcl-2 (IMR-5) was stably infected with either a control or Bcl-2-encoding retrovirus, thus producing the lines IMR-5-BCL-2 and IMR-5-NEO. IMR-5-BCL-2 cells contained 5-fold higher levels of Bcl-2 protein than IMR-NEO and were markedly more resistant to killing by several chemotherapeutic drugs as measured in short-term cytotoxicity assays. Taken together, these findings indicate that Bcl-2 protein levels are regulated during the differentiation of at least some neuroblastoma cell clones in culture, thus raising the possibility of pharmacologically altering bcl-2 gene expression in neuroblastomas in vivo and thereby modulating sensitivity to chemotherapeutic drugs. © 1993, American Association for Cancer Research. All rights reserved. |
| Keywords: | cancer chemotherapy; controlled study; oncoprotein; human cell; proto-oncogene proteins; doxorubicin; dose response; antineoplastic agents; methotrexate; antigen expression; cell viability; cell survival; cell division; etoposide; cell line; vincristine; cell differentiation; drug resistance; tumor cells, cultured; transfection; time factors; gene transfer; oncogenes; monoclonal antibody; gene expression regulation, neoplastic; dimethyl sulfoxide; kinetics; neuroblastoma; immunocytochemistry; immunoblotting; beta 2 microglobulin; drug cytotoxicity; cell clone; nerve cell differentiation; proto-oncogene proteins c-bcl-2; retinoic acid; clone cells; retrovirus; phorbol esters; polyacrylamide gel electrophoresis; broxuridine; 3 (4,5 dimethyl 2 thiazolyl) 2,5 diphenyltetrazolium bromide; neuron specific enolase; vimentin; phorbol 13 acetate 12 myristate; tetradecanoylphorbol acetate; carcinogens; tretinoin; neurofilament protein; polyclonal antibody; protein content; human; priority journal; article; support, non-u.s. gov't; support, u.s. gov't, p.h.s.; triton x 100; 4alpha phorbol didecanoate |
| Journal Title: | Cancer Research |
| Volume: | 53 |
| Issue: | 20 |
| ISSN: | 0008-5472 |
| Publisher: | American Association for Cancer Research |
| Date Published: | 1993-10-15 |
| Start Page: | 4978 |
| End Page: | 4986 |
| Language: | English |
| PUBMED: | 8402688 |
| PROVIDER: | scopus |
| DOI/URL: | |
| Notes: | Source: Scopus |
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