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Quiros RM , Valianou M , Kwon Y , Brown KM , Godwin AK , Cukierman E
Ovarian normal and tumor-associated fibroblasts retain in vivo stromal characteristics in a 3-D matrix-dependent manner
Gynecol Oncol. 2008 Jul;110(1) :99-109
PMID: 18448156    PMCID: PMC2612536   
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OBJECTIVE: Due to a lack of experimental systems, little is known about ovarian stroma. Here, we introduce an in vivo-like 3-D system of mesenchymal stromal progression during ovarian tumorigenesis to support the study of stroma permissiveness in human ovarian neoplasias. METHODS: To sort 3-D cultures into 'normal,' 'primed' and 'activated' stromagenic stages, 29 fibroblastic cell lines from 5 ovarian tumor samples (tumor ovarian fibroblasts, TOFs) and 14 cell lines from normal prophylactic oophorectomy samples (normal ovarian fibroblasts, NOFs) were harvested and characterized for their morphological, biochemical and 3-D culture features. RESULTS: Under 2-D conditions, cells displayed three distinct morphologies: spread, spindle, and intermediate. We found that spread and spindle cells have similar levels of alpha-SMA, a desmoplastic marker, and consistent ratios of pFAKY(397)/totalFAK. In 3-D intermediate cultures, alpha-SMA levels were virtually undetectable while pFAKY(397)/totalFAK ratios were low. In addition, we used confocal microscopy to assess in vivo-like extracellular matrix topography, nuclei morphology and alpha-SMA features in the 3-D cultures. We found that all NOFs presented 'normal' characteristics, while TOFs presented both 'primed' and 'activated' features. Moreover, immunohistochemistry analyses confirmed that the 3-D matrix-dependent characteristics are reminiscent of those observed in in vivo stromal counterparts. CONCLUSIONS: We conclude that primary human ovarian fibroblasts maintain in vivo-like (staged) stromal characteristics in a 3-D matrix-dependent manner. Therefore, our stromal 3-D system offers a tool that can enhance the understanding of both stromal progression and stroma-induced ovarian tumorigenesis. In the future, this system could also be used to develop ovarian stroma-targeted therapies.
Quiros, Roderick M Valianou, Matthildi Kwon, Youngjoo Brown, Kimberly M Godwin, Andrew K Cukierman, Edna CA-06927/CA/NCI NIH HHS/United States P50 CA083638/CA/NCI NIH HHS/United States R01-CA113451/CA/NCI NIH HHS/United States Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't United States Gynecologic oncology Gynecol Oncol. 2008 Jul;110(1):99-109. Epub 2008 May 2.