Abstract
Ovarian cancer, a highly metastatic disease, is the fifth leading cause of cancer-related deaths in women. Chickens are widely used as a model for human ovarian cancer as they spontaneously develop epithelial ovarian tumors similar to humans. The cellular and molecular biology of chicken ovarian cancer (COVCAR) cells, however, have not been studied. Our objectives were to culture COVCAR cells and to characterize their invasiveness and expression of genes and proteins associated with ovarian cancer. COVCAR cell lines (n = 13) were successfully maintained in culture for up to19 passages, cryopreserved and found to be viable upon thawing and replating. E-cadherin, cytokeratin and α-smooth muscle actin were localized in COVCAR cells by immunostaining. COVCAR cells were found to be invasive in extracellular matrix and exhibited anchorage-independent growth forming colonies, acini and tube-like structures in soft agar. Using RT-PCR, COVCAR cells were found to express E-cadherin, N-cadherin, cytokeratin, vimentin, mesothelin, EpCAM, steroidogenic enzymes/proteins, inhibin subunits-α, βA, βB, anti-müllerian hormone, estrogen receptor [ER]-α, ER-β, progesterone receptor, androgen receptor, and activin receptors. Quantitative PCR analysis revealed greater N-cadherin, vimentin, and VEGF mRNA levels and lesser cytokeratin mRNA levels in COVCAR cells as compared with normal ovarian surface epithelial (NOSE) cells, which was suggestive of epithelial-mesenchymal transformation. Western blotting analyses revealed significantly greater E-cadherin levels in COVCAR cell lines compared with NOSE cells. Furthermore, cancerous ovaries and COVCAR cell lines expressed higher levels of an E-cadherin cleavage product when compared to normal ovaries and NOSE cells, respectively. Cancerous ovaries were found to express significantly higher ovalbumin levels whereas COVCAR cell lines did not express ovalbumin thus suggesting that the latter did not originate from oviduct. Taken together, COVCAR cell lines are likely to improve our understanding of the cellular and molecular biology of ovarian tumors and its metastasis.
Highlights
Ovarian cancer is the fifth leading cause of cancer-related deaths in women [1,2,3]
Described for the first time is a simple method for harvesting, long-term culturing, and cryopreservation of chicken ovarian tumor cells from ascites
The procedure is based on a previously reported method for culturing primary ovarian tumor cells obtained from ascites in human subjects with advanced stages of ovarian cancer [22]
Summary
Ovarian cancer is the fifth leading cause of cancer-related deaths in women [1,2,3]. According to an estimate by the National Cancer Institute, the number of new cases of ovarian cancer diagnosed in women will be 22,880 and deaths caused by ovarian cancer will be 15,500 by the end of 2012 [4]. Animal models that develop epithelial ovarian cancer spontaneously are important for cancer prevention studies and to understand early events in ovarian tumorigenesis. In this regard, the domestic chicken (Gallus domesticus) is the most appropriate animal model for human epithelial ovarian cancer [7,8,9,10]. Laying hens develop epithelial ovarian tumors spontaneously with similar etiology, undergo similar disease progression [11], and exhibit a high rate of ovarian cancer incidence (25–40% between 2 and 4 years of age). Hens ovulate almost daily and have 450 ovulations or more within 2 years of age Such incessant ovulation is likely to contribute to the high rate of ovarian cancer incidence by possibly causing oxidative DNA damages in ovarian surface epithelial cells (OSE; [12])
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