卵巢癌G蛋白偶联受体1（OGR1）最初从人类卵巢癌细胞系HEY中克隆得到，它可被生物活性脂分子鞘鞍醇磷脂酰胆碱（SPC）和质子（H+）激活，同时具有不依赖于配体的组成型活性。在前列腺癌细胞中，OGR1被发现可发挥抑制细胞迁移的功能。然而，OGR1在其他细胞中的作用机制还缺少深入的研究。我们实验室前期工作显示，OGR1可以通过Gα12/13-PI3K/Akt-Rac1信号通路明显抑制乳腺癌细胞的迁移。本论文首先通过转染获得稳定高表达OGR1的人乳腺癌细胞系MCF-7细胞克隆（MCF-7-OGR1），发现MCF-7-OGR1条件培养基可有效地抑制MCF-7细胞的迁移。因此，我们推测MCF-7-OGR1细胞可向其条件培养基中分泌特定因子，该因子可以有效抑制乳腺癌细胞的迁移。进一步的实验表明，条件培养基抑制细胞迁移的活性不会受到微波加热（高热，10min）的破坏，但可以通过亲脂的活性炭的处理而使其消除，由此分析活性因子很可能是脂类分子。采用质谱技术分析条件培养基中的脂类成分，发现溶血磷脂酸（LPA）含量在MCF-7-OGR1条件培养基中明显上升。通过用单甘油酯酶（MG-lipase）处理条件培养基、以及单独加入不同浓度的LPA测试其对MCF-7迁移的影响等实验，进一步确定LPA是条件培养基中抑制细胞迁移的活性成分。本论文工作还发现，在高表达OGR1的MCF-7细胞中，自家趋化素（Autotaxin, ATX）的表达较普通MCF-7细胞显著性上升，从而导致LPA的合成增加，且ATX的表达受到Gα12/13-PI3K/Akt信号通路的调控；另外，LPA受体LPA4也受到OGR1的调控而表达上升。虽然在之前的报道中，LPA大多呈现出刺激迁移的作用，但我们发现MCF-7细胞对不同浓度的LPA诱导产生不同的应答：高浓度的LPA可促进MCF-7的迁移，而低浓度的LPA则抑制MCF-7细胞的迁移。通过siRNA实验分别干扰LPA的不同受体LPA2和LPA4发现，当LPA4的表达被干扰后，低浓度的LPA不能再有效抑制MCF-7细胞迁移；而干扰LPA2的表达后，高浓度的LPA不再显著地促进MCF-7细胞迁移。另外，在MCF-7、MDA-MB-231和MDA-MB-468细胞中高表达LPA4均可显著降低细胞的迁移能力，这些都有力的证明了在乳腺癌MCF-7细胞中，LPA4与LPA2具有拮抗功能：在低浓度的LPA刺激下，LPA4发挥着主要的抑制迁移作用；而在高浓度LPA的刺激下，LPA2发挥着主要的促迁移作用。 因此，我们首次发现OGR1可通过上调ATX和LPA4受体表达来抑制MCF-7细胞的迁移，并对LPA2和LPA4受体在MCF-7细胞迁移中的不同功能进行了初步探究。这不仅有助于进一步了解OGR1抑制肿瘤迁移的相关下游机制，并且拓宽了对ATX-LPA信号通路在肿瘤迁移中作用的认识，为以ATX-LPA为靶点治疗肿瘤提供了新的策略。

Ovarian cancer G-protein coupled receptor 1 (OGR1), was first cloned from a human ovarian cancer cell line HEY and has been shown to be proton-sensing and activated by sphingosylphosphorylcholine. In addition, it is likely to have a constitutive activity (ligand-independent). OGR1 has been shown to be a tumor metastasis suppressor in prostate cancer. However, the role and signaling properties of OGR1 in other cancer cells has not been reported (or has only been minimally studied). We have recently found that overexpression of OGR1 could inhibit breast cancer cell migration through a Gα12/13-PI3K/Akt-Rac1 pathway. In this work, we characterized that OGR1-induced secretion of a soluble factor in the conditioned medium (CM) responsible for the migratory inhibition activity in breast cancer cells. The CM from OGR1 stable-overexpressed MCF-7 breast cancer cells (MCF-7-OGR1), but not those from the parental or vector-transfected MCF-7 cells, effectively suppressed the migration of parental MCF-7 cells. This activity was heat-insensitive and removed by lipophilic dextran-charcoal, suggesting lipid factors are likely to be involved. Mass spectrometry analyses of lipid profile from the CM revealed that lysophosphatidic acid (LPA) was one of the major mediators of the inhibitory effect, which was confirmed by using MG-lipase digestion and testing LPA’s effect with the corresponding concentrations on cell migration directly. We showed, for the first time, that up-regulation of autotaxin, the major LPA production enzyme, and one of the six LPA receptors, LPA4, were responsible for the increased LPA production in OGR1-expressing cells. Through RNAi experiments of LPA2 and LPA4, we provide new insights into the crosstalk between LPA receptors. We demonstrated that LPA2 plays a dominated migration-promoting role under 5μM LPA, while LPA4, in contrast to other stimulatory LPA receptors, mediated a major migration inhibitory role in response to low concentrations of LPA.We show here the first evidence that OGR1 up-regulates ATX and LPA4, leading to increase LPA in CM and inhibited cell migration in MCF-7 cell line. While the detailed mechanisms by which OGR1 regulate ATX and LPA4 remain to be further investigated, the results in this study will not only expand our understanding of LPA signaling, but also provide very critical information in designing and developing new strategies targeting the ATX-LPA axis in cancer treatment.