| 刘志豪,谭万龙,蓝创歆,朱俊逸,吴海渤.欧前胡素调节Gas6/Axl轴对膀胱癌荷瘤小鼠免疫功能的影响[J].现代生物医学进展英文版,2023,(20):3809-3814. |
| 欧前胡素调节Gas6/Axl轴对膀胱癌荷瘤小鼠免疫功能的影响 |
| Effect of Imperatorin Regulating Gas6/Axl Axis on Immune Function of Bladder Cancer Bearing Mice |
| Received:May 18, 2023 Revised:June 13, 2023 |
| DOI：10.13241/j.cnki.pmb.2023.20.002 |
| 中文关键词: 欧前胡素 Gas6/Axl信号通路 膀胱癌 免疫功能 |
| 英文关键词: Imperatorin Gas6/Axl signaling pathway Bladder cancer Immune function |
| 基金项目:广东省自然科学基金项目（2017A030307037） |
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| 中文摘要: |
| 摘要 目的：探讨欧前胡素（IMP）调节生长停滞特异蛋白6（Gas6）/Axl轴对膀胱癌荷瘤小鼠免疫功能的影响。方法：从60只SPF级雄性C57BL/6小鼠中随机选取12只小鼠作为对照组（CON组），其余小鼠平均分为Model组、IMP组、Gas6组、IMP+Gas6组，每组12只，往其左侧前肢皮下注射0.2 mL浓度为1×107个/mL的鼠源性膀胱癌肿瘤细胞系T739细胞溶液（Model组和IMP组注射对照载体的T739稳转株细胞，Gas6组和IMP+Gas6组注射过表达Gas6的T739稳转细胞株）造模。造模成功后，IMP组及IMP+Gas6组通过腹腔注射5 mg/kg的IMP；CON组、Model组及Gas6组通过腹腔注射等量的生理盐水，2天1次，持续5次。测量小鼠肿瘤体积和重量；测定胸腺指数和脾脏指数；酶联免疫吸附测定（ELISA）法检测血清白介素2（IL-2）、γ干扰素（IFN-γ）、血管内皮生长因子（VEGF）水平；苏木精-伊红（HE）染色检查肿瘤组织病理变化；流式细胞术检测T细胞亚群；TUNEL染色检测细胞凋亡；蛋白质印迹法（Western blot）检测Gas6/Axl通路相关蛋白表达。结果：Model组肿瘤细胞整体上排列松散，出现不同程度的核固缩、核碎裂现象；IMP则可以逆转该现象，使肿瘤细胞排列重新变得紧密，细胞核固缩程度提升，核碎裂现象降低；Model组较CON组肿瘤体积、肿瘤质量、CD8+细胞百分比、Gas6、Axl蛋白水平、VEGF水平显著增加（P<0.05），胸腺指数和脾脏指数、IL-2、IFN-γ、CD4+细胞百分比、CD4+/CD8+、肿瘤细胞凋亡率显著下降（P<0.05）；与Model组相比，IMP组胸腺指数和脾脏指数、IL-2、IFN-γ、CD4+细胞百分比、CD4+/CD8+、肿瘤细胞凋亡率显著升高（P<0.05），肿瘤体积、肿瘤质量、CD8+细胞百分比、Gas6、Axl蛋白水平、VEGF水平显著下降（P<0.05）；而Gas6组结果与IMP组趋势相反；而IMP+Gas6组的结果显示：IMP能逆转Gas6过表达后对膀胱癌荷瘤小鼠的促瘤作用。结论：IMP可能通过下调Gas6/Axl信号通路改善膀胱癌荷瘤小鼠免疫功能，从而产生抑瘤效果。 |
| 英文摘要: |
| ABSTRACT Objective: To investigate the effect of imperatorin (IMP) on the immune function of bladder cancer bearing mice by regulating the growth arrest specific protein 6 (Gas6)/Axl axis. Methods: 12 mice were randomly selected from 60 SPF grade male C57BL/6 mice as the control group (CON group). The remaining mice were evenly divided into Model group, IMP group, Gas6 group, and IMP+Gas6 group, with 12 mice in each group. A concentration of 0.2 mL was injected subcutaneously into their left forelimb at a concentration of 1×107/mL murine bladder cancer cell line T739 cell solution (model group and IMP group were injected with T739 stable transfection cell line of control vector, Gas6 group and IMP+Gas6 group were injected with T739 stable transfection cell line expressing Gas6) for modeling. After successful modeling, the IMP group and IMP+Gas6 group received intraperitoneal injection of 5 mg/kg of IMP. The CON group, Model group, and Gas6 group were injected with an equal amount of physiological saline via intraperitoneal injection, once every 2 days, for 5 consecutive times. Measure the volume and weight of mouse tumors. Measure thymus index and spleen index. Serum interleukin-2 (IL-2), γ Interferon (IFN-γ), vascular endothelial growth factor (VEGF) levels were detected by enzyme linked immunosorbent assay (ELISA). Hematoxylin eosin (HE) staining was used to examine the pathological changes of tumor tissue. T cell subpopulations was detected by flow cytometry. TUNEL staining was used to detect cell apoptosis. Western blot (Western blot) was used to detect the expression of Gas6/Axl pathway related proteins. Results: The tumor cells in Model group were loosely arranged, with pyknosis and Karyorrhexis in different degrees. IMP can reverse this phenomenon, making the tumor cell arrangement become compact again, the degree of nuclear pyknosis increases, and the Karyorrhexis phenomenon decreases. The model group showed a significant increase in tumor volume, tumor mass, CD8+ cell percentage, Gas6, Axl protein levels, and VEGF levels compared to the CON group (P<0.05), as well as thymus and spleen indices, IL-2, and IFN-γ, the percentage of CD4+cells, CD4+/CD8+, and tumor cell apoptosis rate significantly decreased (P<0.05). Compared with the Model group, the IMP group had thymic index, spleen index, IL-2, IFN-γ, the percentage of CD4+cells, CD4+/CD8+ cells, and tumor cell apoptosis rate significantly increased (P<0.05), while tumor volume, tumor mass, percentage of CD8+ cells, Gas6, Axl protein levels, and VEGF levels significantly decreased (P<0.05). The results of the Gas6 group showed opposite trends to those of the IMP group. The results of IMP+Gas6 group showed that IMP could reverse the tumor promoting effect of overexpression of Gas6 on bladder cancer bearing mice. Conclusion: IMP may improve the immune function of bladder cancer bearing mice by down-regulation of Gas6/Axl signaling pathway, thus producing anti-tumor effect. |
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