血小板介导肿瘤血管形成和稳定性的机制分析-mechanism analysis of platelet-mediated tumor angiogenesis and stability.docxVIP

影响肿瘤血管形成，我们使用 anti-PECAM-1 和 anti-α–SMA 双染法检查了肿瘤组织血管密度和覆盖率。血小板减少组的 PECAM-1-positive 毛细血管密度明显低于对照组。肿瘤内的毛细血管覆盖率（α–SMA-positive 的壁细胞）亦明显少于对照组。5.减少血小板降低 肿瘤缺血和 HIF-1α 表达：为了更好地探寻血小板减少导致肿瘤转移 的分子机制，我们首先通过检查肿瘤组织中哌莫硝唑复合物的生成评 价了缺氧水平，结果显示血小板减少组的缺氧明显低于对照组。此外， 缺氧诱导的转录因子 HIF-1α 表达明显低于对照组。6.血小板与肿瘤细 胞共培养促进肿瘤生长和肿瘤转移：血小板与肿瘤细胞 B16/F10 以 1000:1 的比例混悬共培养（37℃,5%CO2）30min 后接种于小鼠背部，同时设未共培养的对照组。待肿瘤生长 24 天后，共培养组的肿瘤大 小(3968 ± 296 mm3 Vs. 2956 ± 180 mm3, P0.05)和肿瘤重量(5.529 ±0.35 g Vs. 3.943 ± 0.74 g, P0.05)明显大于对照组，且肿瘤的肺转移明 显增加。通过检查肿瘤组织中哌莫硝唑复合物的生成评价了缺氧水 平，结果显示共培养组的缺氧明显高于对照组。7.血小板改变肿瘤内 的血管生成因子水平：在处死小鼠的前一天对肿瘤组织进行微透析， 透析液以 VEGF-Elisa 试剂盒进行定量分析，结果显示血小板减少组 的 VEGF 明显低于对照组(P0.05)。结论：1.血小板促进肿瘤细胞的 迁移，即增强肿瘤细胞的侵袭性。2.血液循环中的血小板在肿瘤的生 长和侵袭中扮演了重要角色,不改变肿瘤的大小，但是促进肿瘤转移。 3.肿瘤微环境中血小板与肿瘤细胞的粘附增加肿瘤的生长和转移。4. 血小板减少改变肿瘤微环境中血管的数量和成熟度。5.血小板可能参与了肿瘤缺血，血小板数量的减少使转录因子 HIF-1α 表达下调。6.血小板可能参与了肿瘤微环境血管生成因子的分泌。 关键词： 血小板；肿瘤；血管生成；血管稳定性；转移Role of platelet in the regulation of tumor angiogenesis and vessel stabilityAbstract:Objective:Platelets play a fundamental role in maintaining hemostasis and have been shown to participate in hematogenous dissemination of tumor cells. Abundant platelets were detected in the tumor microenvironment outside of the blood vessel. However, it is unknown whether a platelet-tumor cell interaction outside of the bloodstream plays a role in regulating primary tumor growth, angiogenesis, and metastasis initiation. The current study was designed to test the hypothesis that platelets influence metastasis by mediating the migration of tumor cells to blood vessels in the tumor microenvironment, giving rise to increased intravasation.Method:1.in vitro studies: We modifiedthree-dimensional Matrigel as described previously. B16/F10 cells (1×103cells/mL) were suspended with PLTs(1×106 cells/mL) in 250μL matrigel and pipetted into the upper chambers of 24-well Transwell inserts. Insertswere placed into lower chamber wells that were filled with DMEM (600 μL) containing 2.5% FBS.After incubating in standard cell cu