Role of hCG in vasculogenic mimicry in OVCAR-3 ovarian cancer cell line.
OBJECTIVE
Vasculogenic mimicry (VM) is induced by hypoxia in 3-dimensional culture of ovarian cancer cells. By using this 3D model system, we explored the expression of human chorionic gonadotropin (hCG) and its effects on VM formation in ovarian cancer cell line OVCAR-3 both under normoxic and hypoxic conditions.


METHODS
Vasculogenic mimicry was identified by morphological observation and detection of vascular cell marker expressed by OVCAR-3. Potential formation of tumor channels was observed by light microscopy and scanning electron microscopy. Expression of vascular cell marker CD31, vascular endothelial growth factor, and Factor VIII were detected by flow cytometry, immunochemistry, and Western blot. Expression of hCG was investigated by enzyme-linked immunosorbent assay, chemiluminescence immunoassay, real-time polymerase chain reaction (PCR), and Western blot. Expression of hypoxia-inducible factor-1α (HIF-1α) was detected by real-time PCR, Western blot, and blocked by small interference RNA. Incubation of OVCAR-3 with recombinant hCG was used to evaluate its effect on VM formation. The specificity of the effect of hCG was assessed by inhibition with the neutralizing anti-hCG antibody.


RESULTS
OVCAR-3 cells formed vessel-like network structures and expressed vascular marker significantly under hypoxia in 3D. The expression level of hCG under hypoxia was significantly higher than that under normoxia. Attenuating hypoxia-inducible factor (HIF)-1α expression via small interference RNA resulted in a significantly decreased hCG expression in OVCAR-3, which indicated that the effect of hypoxia on hCG expression was mediated through HIF-1α. Treatment of OVCAR-3 with 5000 mU/mL hCG resulted in the presence of tumor cell-lined vasculature and significant elevation in vascular marker expression, even under normoxia. Expression level of vascular marker and HIF-1α in OVCAR-3 increased in response to hCG treatment in a dose-dependent manner. The effect of hCG was inhibited by the neutralizing anti-hCG antibody.


CONCLUSIONS
Human chorionic gonadotropin has the potential to induce VM in OVCAR-3. Human chorionic gonadotropin might have synergistic hypoxia-induced effect on vascular marker and HIF-1α expression.
Department of Obstetrics and Gynecology, The Affiliated Hospital of Nantong University, People's Republic of China. sumin_nt@126.com
Min
M
Su
Weiwei
W
Wei
Xiangxiang
X
Xu
Xiaoying
X
Wang
Caoyi
C
Chen
Li
L
Su
Yuquan
Y
Zhang
D045744:Cell Line, Tumor; D048430:Cell Shape; D006063:Chorionic Gonadotropin / Q000502:physiology*; D005260:Female; D006801:Humans; D000860:Hypoxia; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D009389:Neovascularization, Pathologic; D010051:Ovarian Neoplasms / Q000098:blood supply*; D034741:RNA, Small Interfering
D006063:Chorionic Gonadotropin; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D034741:RNA, Small Interfering