Purpose

Paracrine interactions with the stromal environment, including fibroblasts, may be important in the pathogenesis of ovarian cancer. Here, we evaluated the effect of conditioned media derived from ovarian fibroblasts (fibroblast-CMs) and their major cytokines on the growth of ovarian cancer cells, as well as the involvement of mitogen-activated protein kinases (MAPKs) and AKT in mediating this effect.

Methods

Ovarian cancer cells were cultured in serum-free media (SF), or conditioned media of fibroblasts derived from normal ovary (CM1) and ovarian tumor tissue (CM2). Cell proliferation was measured by MTT assay. Phosphorylation of MAPKs and AKT was evaluated by Western blotting. Specific inhibitors of MAPKs and AKT were used to evaluate their respective involvement in mediating increased cell growth. Cytokine levels in fibroblast-CMs were measured using Luminex assays. Immunohistochemical staining was conducted for CXCL1, CXCR2 and phosphorylated p38 in primary ovarian tumors.

Results

CM1 and CM2 significantly increased the growth of ovarian cancer cells relative to SF. In OVCAR3 and OVCAR4 cells, p38 phosphorylation was strongly induced by fibroblast-CMs, and pre-treatment with a p38 inhibitor prevented the growth increase induced by fibroblast-CMs. Fibroblasts secreted high levels of IL-6, IL-8, MCP1 and CXCL1. Treatment with only CXCL1 (1 μg/ml) increased cell growth and p38 phosphorylation. Treatment with a CXCR2 inhibitor effectively prevented p38 activation and cell growth induced by fibroblast-CMs. High expression of both CXCL1 and CXCR2 correlated with high expression of phosphorylated p38 in primary ovarian tumors.

Conclusions

From our data, we conclude that CXCL1 is a key factor derived from ovarian fibroblasts that is responsible for increased ovarian cancer cell growth in part through p38 activation. Phosphorylated p38 can be used as a biomarker to predict CXCL1-CXCR2 interaction in vivo.

中文翻译：

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通过 CXCL1-CXCR2 相互作用的上皮-间质通讯通过 p38 激活刺激卵巢癌细胞的生长。

目的

与基质环境（包括成纤维细胞）的旁分泌相互作用可能在卵巢癌的发病机制中很重要。在这里，我们评估了源自卵巢成纤维细胞 (fibroblast-CMs) 的条件培养基及其主要细胞因子对卵巢癌细胞生长的影响，以及丝裂原活化蛋白激酶 (MAPK) 和 AKT 在介导这种影响中的作用.

方法

卵巢癌细胞在无血清培养基 (SF) 或源自正常卵巢 (CM1) 和卵巢肿瘤组织 (CM2) 的成纤维细胞的条件培养基中培养。通过MTT测定法测量细胞增殖。通过蛋白质印迹评估 MAPK 和 AKT 的磷酸化。MAPK 和 AKT 的特异性抑制剂用于评估它们各自参与介导细胞生长增加的情况。使用 Luminex 测定法测量成纤维细胞 CM 中的细胞因子水平。对原发性卵巢肿瘤中的 CXCL1、CXCR2 和磷酸化 p38 进行免疫组织化学染色。

结果

相对于 SF，CM1 和 CM2 显着增加了卵巢癌细胞的生长。在 OVCAR3 和 OVCAR4 细胞中，成纤维细胞 CM 强烈诱导 p38 磷酸化，用 p38 抑制剂预处理可防止成纤维细胞 CM 诱导的生长增加。成纤维细胞分泌高水平的 IL-6、IL-8、MCP1 和 CXCL1。仅用 CXCL1 (1 μg/ml) 处理会增加细胞生长和 p38 磷酸化。用 CXCR2 抑制剂治疗可有效阻止 p38 激活和成纤维细胞 CM 诱导的细胞生长。CXCL1 和 CXCR2 的高表达与原发性卵巢肿瘤中磷酸化 p38 的高表达相关。

结论

根据我们的数据，我们得出结论，CXCL1 是源自卵巢成纤维细胞的关键因子，它部分通过 p38 激活负责增加卵巢癌细胞的生长。磷酸化的 p38 可用作预测体内CXCL1-CXCR2 相互作用的生物标志物。