基于肥大细胞的募集探讨前列腺癌微环境中PKD3-uPA信号轴促肿瘤血管新生的作用和机制

The recruitment of mast cells have been found to play a critical role in the tumor microenvironment remodelling and tumor angiogenesis. Our current data show that constitutive active PKD3 expression in prostate cancer cells not only significantly leads to increase the migration of human mast cells via urokinase-tpye plasminogen activator(uPA) secretion by Boden chamber chemotactic assay, but also enhances the expression of angiogenic factor FGF-2 in the human mast cells. However, how constitutive active PKD3 in prostate cancer cells regulates the recruitment of mast cells and tumor-promoting angiogenesis in the prostate tumor microenvironment remains poorly understood. The objective of the present proposal is to explore the the hypothesis for the first time that PKD3-uPA axis in prostate cancer cells promotes the recruitment of mast cells to the peritumoral compartment and tumor-promoting angiognesis in prostate cancer microenvironment. To test this hypothesis,the proposed study will utilize the following AIMs: AIM 1: verify the corelations between constitutive active PKD3 and recruitment and number of mast cells in clinical tumor samples. AIM 2 : determine PKD3-uPA axis contributes to bone marrow-derived mast cells migration by in vitro chemotactic assay. AIM 3 : investigate the role of PKD3-uPA axis in the regulation of mast cells recruitment and tumor angiogenesis in the prostate tumor microenvironment using the orthotropic implantation of mouse tumor cells. AIM 4 : explore the expression and mechanism underlying PKD3-uPA induces angiogenic factors in the mast cells. This proposal will provide a new insight into the mechanism of PKD3-uPA axis in the promotions of mast cells tumor angiogenesis in the prostate cancer microenvironment.

肥大细胞的募集在肿瘤微环境重塑及肿瘤血管新生中起着重要的作用。我们最近的研究显示，前列腺癌细胞中组成性激活和表达的PKD3促进uPA的分泌而诱导肥大细胞的迁移及血管新生因子FGF-2的表达。在前列腺癌肿瘤微环境中组成性激活的PKD3如何调控肥大细胞的募集并促进肿瘤的血管新生尚不清楚。本研究首次探讨PKD3-uPA信号轴诱导的肥大细胞募集并促进肿瘤血管新生与调控机理，内容包括：（1）明确前列腺癌临床肿瘤标本中PKD3的表达和激活与肥大细胞的募集的相关性；（2）体外细胞趋化迁移证实PKD3-uPA信号轴促进骨髓源肥大细胞的迁移；（3）肿瘤原位移植模型确证PKD3-uPA信号轴调控肥大细胞募集及促肿瘤的血管新生；（4）探讨PKD3-uPA信号轴诱导肥大细胞血管新生因子的表达和调控机理。本研究从PKD3-uPA信号轴这个新视点为揭示前列腺癌微环境中肥大细胞促肿瘤血管新生机制奠定基础。

肥大细胞在多种肿瘤微环境以及肿瘤微血管形成中扮演着重要的作用，但是肥大细胞在前列腺癌肿瘤微环境以及肿瘤微血管形成的作用的机理尚不完全清楚。本研究通过一系列细胞和分子生物学研究和策略探讨了PKD2/3如何通过肥大细胞的募集迁移，促进肿瘤的血管新生，从而促进前列腺癌的生长及进展。结果显示，（1）通过对临床前列腺癌组织芯片分析，对比正常的前列腺组织，在前列腺癌样本中PKD磷酸化水平、CD31、C-KIT的表达量显著增加。而且PKD磷酸化水平与微血管密度呈现显著的相关性。（2）PKD活性的抑制及敲低前列腺癌细胞DU145和PC-3M细胞中PKD2或PKD3的表达抑制p815肥大细胞及小鼠骨髓源肥大细胞（BMMCs）的趋化迁移；（3）PKD2和PKD3通过NF-kB和ERK1/2信号通路调节SCF,CCL5和CCL11的表达，从而募集肥大细胞至肿瘤周围，促进肥大细胞的血管生成因子的表达，并最终调节肿瘤的血管形成；（4）PKD2和PKD3调节NF-kB p65 或AP1与SCF、CCL5和CCL11基因的启动子区域结合；（5）抑制PKD的活性降低异位移植的小鼠肿瘤的生长、肥大细胞的募集及肿瘤的血管生成；（6）前列腺癌临床资料显示PKD磷酸化与微血管密度及肥大细胞募集有关。上述结果提示PKD2和PKD3可能在前列腺癌的肿瘤的血管生成中起着极为重要的作用，本研究为进一步揭示前列腺癌的血管生成机理以及临床治疗晚期前列腺癌潜在靶标的筛选提供了新的思路。