Background/Objective: Clear cell renal cell carcinoma (ccRCC) is characterized by a high degree of functional intratumoral heterogeneity (ITH). This is highlighted by the finding that tumor cell proliferation and intracellular signaling occur preferentially in the tumor periphery. The driving forces for such a spatial organization are largely unknown. Herein, we investigate the role of the tumor microenvironment in the control of tumor cell proliferation and functional ITH. Methods: Conditioned media (CM) derived from nonmalignant peritumoral kidney tissue were used to stimulate RCC cells in vitro. A neutralization assay was used to characterize the role of FGF-2 in the CM. The molecular mechanisms underlying the action of CM on RCC cells were investigated using immunoblotting, flow cytometry and immunofluorescence microscopy. Lastly, a series of ccRCCs were stained for Ki-67 and p27Kip1, and expression was analyzed in both tumor periphery and center. Results: We show that CM derived from nonmalignant kidney cells adjacent to an RCC can downregulate the expression of the CDK inhibitor p27Kip1 through enhanced protein degradation in an FGF-2-dependent fashion. FGF-2 functions mainly through the PI3K/AKT pathway downstream of its receptors, and RCC cells with constitutively high AKT activity show not only an enhanced degradation of p27Kip1 through the Emi1-Skp2 axis, but also a subcellular mislocalization of p27Kip1 to the cytoplasmic compartment. Such a mislocalization was also detected in the tumor periphery in vivo suggesting that p27Kip1 plays an important role in shaping this spatial niche. Conclusions: Our results suggest that the tumor microenvironment is involved in shaping the tumor peripheral niche by stimulating the enhanced proliferation that is characteristic for this zone.

中文翻译：

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微环境衍生的 FGF-2 通过调节 p27 ^Kip1刺激肾细胞癌细胞增殖：对空间小生境形成和功能性肿瘤内异质性的影响

背景/目的：透明细胞肾细胞癌 (ccRCC) 的特征在于高度的功能性瘤内异质性 (ITH)。肿瘤细胞增殖和细胞内信号传导优先发生在肿瘤外围的发现强调了这一点。这种空间组织的驱动力在很大程度上是未知的。在此，我们研究了肿瘤微环境在控制肿瘤细胞增殖和功能性 ITH 中的作用。方法：使用来源于非恶性瘤周肾组织的条件培养基 (CM) 在体外刺激 RCC 细胞。中和试验用于表征 FGF-2 在 CM 中的作用。使用免疫印迹、流式细胞术和免疫荧光显微镜研究了 CM 对 RCC 细胞作用的分子机制。最后，对一系列 ccRCC 进行 Ki-67 和 p27Kip1 染色，并分析肿瘤外围和中心的表达。结果：我们表明，来源于与 RCC 相邻的非恶性肾细胞的 CM 可以通过 FGF-2 依赖性方式增强蛋白质降解来下调 CDK 抑制剂 p27Kip1 的表达。FGF-2 主要通过其受体下游的 PI3K/AKT 通路发挥作用，具有高 AKT 活性的 RCC 细胞不仅显示出通过 Emi1-Skp2 轴增强的 p27Kip1 降解，而且显示出 p27Kip1 亚细胞错误定位到细胞质区室. 在体内肿瘤外围也检测到这种错误定位，表明 p27Kip1 在塑造这个空间生态位方面起着重要作用。结论：