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American Head & Neck Society
Annual Meeting, April 10-11, 2013
JW Marriott Grande Lakes
Orlando, Florida

During the
Combined Otolaryngology Spring Meeting
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MALIGNANT TRANSFORMATION OF MURINE 3T3 CELLS BY CO-CULTURING WITH NASOPHARYNGEAL CARCINOMA CELLS

Presentation: P007
Topic: Basic Science - Molecular / Cellular Biology
Type: Poster
Date: Wednesday - Thursday, April 10 - 11, 2013
Session: Designated Poster viewing times
Authors: Wen-son Hsieh, Dr, Fredrik Petersson, Associate, professor, De -Yun Wang, Associate, professor, Kwok Seng Loh, Associate, professor, Fenggang Yu, PhD
Institution(s): National University of Singapore

Mouse 3T3 fibroblasts have been widely used as a feeder layer to promote human epithelial cell cultures. In this study, we attempted to culture nasopharyngeal carcinoma (NPC) cells on 3T3 cells feeders, since the establishment of NPC primary cell culture is notoriously difficult due to early growth arrest.


Unexpectedly, a sub-population of 3T3 feeder cells were transformed by the NPC tumor cells. The transformed 3T3 cells acquired an accelerated growth rate, lost contact inhibition, and displayed anchorage-independent multilayer growth in vitro. These cells were highly malignant in vivo exemplified by very robust tumor formation in immunodeficient nude mice. At 1.5X106 cells, tumors were formed with 100% of incidence and the diameter reached 2.5 cm within 2 weeks. Histopathologic examination of the tumors revealed obviously malignant features, including increased nuclear to cytoplasmic ratio, tumor giant cells, large nucleoli, abundant mitotic figures with also readily identified atypical forms. Most strikingly, the immunohistochemical study of the tumors revealed that a significant proportion (>80%) expressed cytokeratin (AE1/AE3). Instead of forming sarcomatous tumors, they developed into tumor cells that showed a carcinomatous phenotype somewhat resembling the original NPC, albeit the detection of Epstein-Barr virus (EBV) turned out negative by EBV-encoded RNA (EBER) in-situ hybridization. Tumors were formed with the same efficacy even after 50 in vitro passages of the cells, indicating the transformation was sustained rather than transient.

Furthermore, the transformations were reproducible by using NPC cells from different patients with or without direct cell-cell contact, suggesting some soluble factors released by NPC tumor cells mediated the event. However, NPC cells alone were not sufficient to confer the transformed characteristics of cancer cells onto normal human fibroblasts and nasopharyngeal epithelial cells.


These findings highlight potential roles for soluble factors (e.g. exsomes, microvesicles) in the induction of horizontal cellular transformation and contribution to the onset and development of human NPC disease.
 

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