Yu Shuangxing, Murph Mandi M, Lu Yiling, Liu Shuying, Hall Hassan S, Liu Jinsong, Stephens Clifton, Fang Xianjun, Mills Gordon B
Department of Systems Biology, Division of Cancer Medicine, MD Anderson Cancer Center, Houston, TX 77030, USA.
J Natl Cancer Inst. 2008 Nov 19;100(22):1630-42. doi: 10.1093/jnci/djn378. Epub 2008 Nov 11.
Lysophosphatidic acid (LPA) acts through the cell surface G protein-coupled receptors, LPA1, LPA2, or LPA3, to elicit a wide range of cellular responses. It is present at high levels in intraperitoneal effusions of human ovarian cancer increasing cell survival, proliferation, and motility as well as stimulating production of neovascularizing factors. LPA2 and LPA3 and enzymes regulating the production and degradation of LPA are aberrantly expressed by ovarian cancer cells, but the consequences of these expression changes in ovarian cancer cells were unknown.
Expression of LPA1, LPA2, or LPA3 was inhibited or increased in ovarian cancer cells using small interfering RNAs (siRNAs) and lentivirus constructs, respectively. We measured the effects of changes in LPA receptor expression on cell proliferation (by crystal violet staining), cell motility and invasion (using Boyden chambers), and cytokines (interleukin 6 [IL-6], interleukin 8 [IL-8], and vascular endothelial growth factor [VEGF]) production by enzyme-linked immunosorbent assay. The role of LPA receptors in tumor growth, ascites formation, and cytokine production was assessed in a mouse xenograft model. All statistical tests were two-sided.
SKOV-3 cells with increased expression of LPA receptors showed increased invasiveness, whereas siRNA knockdown inhibited both migration (P < .001, Student t test) and invasion. Knockdown of the LPA2 or LPA3 receptors inhibited the production of IL-6, IL-8, and VEGF in SKOV-3 and OVCAR-3 cells. SKOV-3 xenografts expressing LPA receptors formed primary tumors of increased size and increased ascites volume. Invasive tumors in the peritoneal cavity occurred in 75% (n = 4) of mice injected with LPA1 expressing SKOV-3 and 80% (n = 5) of mice injected with LPA2 or LPA3 expressing SKOV-3 cells. Metastatic tumors expressing LPA1, LPA2, and LPA3 were identified in the liver, kidney, and pancreas; tumors expressing LPA2 and LPA3 were detected in skeletal muscle; and tumors expressing LPA2 were also found in the cervical lymph node and heart. The percent survival of mice with tumors expressing LPA2 or LPA3 was reduced in comparison with animals with tumors expressing beta-galactosidase.
Expression of LPA2 or LPA3 during ovarian carcinogenesis contributes to ovarian cancer aggressiveness, suggesting that the targeting of LPA production and action may have potential for the treatment of ovarian cancer.
溶血磷脂酸（LPA）通过细胞表面G蛋白偶联受体LPA1、LPA2或LPA3发挥作用，引发广泛的细胞反应。它在人卵巢癌腹腔积液中含量很高，可提高细胞存活率、增殖能力和运动能力，并刺激新生血管形成因子的产生。LPA2和LPA3以及调节LPA产生和降解的酶在卵巢癌细胞中异常表达，但这些表达变化在卵巢癌细胞中的后果尚不清楚。
分别使用小干扰RNA（siRNA）和慢病毒构建体在卵巢癌细胞中抑制或增加LPA1、LPA2或LPA3的表达。我们通过结晶紫染色测量LPA受体表达变化对细胞增殖的影响，使用博伊登小室测量细胞运动和侵袭能力，并通过酶联免疫吸附测定法测量细胞因子（白细胞介素6 [IL-6]、白细胞介素8 [IL-8]和血管内皮生长因子[VEGF]）的产生。在小鼠异种移植模型中评估LPA受体在肿瘤生长、腹水形成和细胞因子产生中的作用。所有统计检验均为双侧检验。
LPA受体表达增加的SKOV-3细胞侵袭性增强，而siRNA敲低则抑制迁移（P <.001，学生t检验）和侵袭。敲低LPA2或LPA3受体可抑制SKOV-3和OVCAR-3细胞中IL-6、IL-8和VEGF的产生。表达LPA受体的SKOV-3异种移植形成了更大的原发性肿瘤和更多的腹水。在注射表达LPA1的SKOV-3细胞的小鼠中以及注射表达LPA2或LPA3的SKOV-3细胞的小鼠中，分别有75%（n = 4）和80%（n = 5）在腹腔内发生侵袭性肿瘤。在肝脏、肾脏和胰腺中发现了表达LPA1、LPA2和LPA3的转移性肿瘤；在骨骼肌中检测到表达LPA2和LPA3的肿瘤；在颈部淋巴结和心脏中也发现了表达LPA2的肿瘤。与表达β-半乳糖苷酶的肿瘤小鼠相比，表达LPA2或LPA3的肿瘤小鼠的存活率降低。
卵巢癌发生过程中LPA2或LPA3的表达促进了卵巢癌的侵袭性，这表明针对LPA产生和作用的靶向治疗可能具有治疗卵巢癌的潜力。
