文章摘要
徐叶峰,刘云霞,姚勇伟,等.骨肉瘤肺转移大鼠模型建立及清肺固金汤的干预研究[J].浙江中医药大学学报,2021,(1):8-15, 33.
骨肉瘤肺转移大鼠模型建立及清肺固金汤的干预研究
Establishment of Rat Model of Osteosarcoma with Pulmonary Metastasis and Intervention Study of Qingfei Gujin Decoction
投稿时间:2020-04-20  
DOI:10.16466/j.issn1005-5509.2021.01.002
中文关键词: 骨肉瘤  肺转移  肺虚痰阻证  清肺固金汤  炎性因子  趋化因子受体
英文关键词: osteosarcoma  pulmonary metastasis  pulmonary deficiency phleqm obstruction syndrome  Qingfei Gujin Decoction  inflammatory factor  chemokines receptor
基金项目:浙江省自然科学基金青年项目(LQ18H290001);浙江省中医药科技计划项目(2015ZB081、2018ZB107)
作者单位
徐叶峰 杭州市第三人民医院 杭州 310009 
刘云霞 杭州市第三人民医院 杭州 310009 
姚勇伟 杭州市第三人民医院 杭州 310009 
王翌庆 杭州市第三人民医院 杭州 310009 
杨洁文 杭州市第三人民医院 杭州 310009 
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中文摘要:
      [目的]建立“肺虚痰阻证”骨肉瘤肺转移病证结合大鼠模型,探讨清肺固金汤对骨肉瘤肺转移大鼠的干预作用。[方法]将80只SD大鼠随机分为4组,空白对照组、骨肉瘤模型组、骨肉瘤肺转移模型组和清肺固金组,选用骨肉瘤UMR-106细胞株建立SD大鼠骨肉瘤肺转移模型。骨肉瘤肺转移模型建立成功后,采用冷风刺激加烟熏法建立“肺虚痰阻证”模型。骨肉瘤模型组、骨肉瘤肺转移模型组给予0.9%氯化钠溶液灌胃,清肺固金组给予清肺固金汤灌胃,连续给药8周。观察分析清肺固金汤对各组大鼠体质量、原发灶肿瘤生长及肺转移情况的干预作用。采用酶联免疫吸附法(enzyme-linked immunosorbent assay,ELISA)检测各组大鼠血清及瘤体内白细胞介素-10(interleukin-10,IL-10)、转化生长因子-β1(transforming growth factor-β1,TGF-β1)含量。以Western blot法检测大鼠右胫骨组织趋化因子基元受体4(CXC motif receptor 4,CXCR4)蛋白表达。[结果]除空白对照组外,其余各组接种大鼠均出现肉眼可见的右膝关节肿块,X线检查提示右胫骨干骺端骨质破坏,呈团片状密度增高影;病理检查右胫骨可见核大深染的骨肉瘤细胞,肺组织可见转移性骨肉瘤结节。与骨肉瘤模型组比较,骨肉瘤肺转移模型组体质量减轻,清肺固金组体质量增加,差异有统计学意义(P<0.05)。与骨肉瘤模型组、骨肉瘤肺转移模型组比较,清肺固金组瘤体质量、体积均明显减少,差异有统计学意义(P<0.05,P<0.01)。骨肉瘤肺转移模型组的肺转移灶数目最多,其中Ⅲ+Ⅳ级结节最多,而清肺固金组肺转移灶数目及Ⅲ+Ⅳ级结节数目均为最低。与空白对照组比较,骨肉瘤模型组、骨肉瘤肺转移模型组的血清IL-10、TGF-β1含量以及右胫骨CXCR4蛋白表达均增加,差异有统计学意义(P<0.05,P<0.01)。与骨肉瘤模型组比较,骨肉瘤肺转移模型组血清及瘤体内IL-10、TGF-β1含量,右胫骨CXCR4蛋白表达均增加,清肺固金组IL-10、TGF-β1含量及CXCR4蛋白表达均明显下降,差异有统计学意义(P<0.05,P<0.01)。[结论]“肺虚痰阻证”骨肉瘤肺转移病证结合大鼠模型临床契合度好,成功率高,稳定性佳。清肺固金汤具有较好的抗骨肉瘤肺转移作用,其机制可能与下调IL-10、TGF-β1及CXCR4表达有关。
英文摘要:
      [Objective] To establish the “pulmonary deficiency phlegm obstruction syndrome” osteosarcoma pulmonary metastasis rat model of combination of disease and syndrome, and explore the effect of Qingfei Gujin Decoction on osteosarcoma pulmonary metastasis rats.[Methods] Eighty SD rats were divided into 4 groups, blank control group, osteosarcoma model group, osteosarcoma lung metastasis model group, Qingfei Gujin group, and the osteosarcoma UMR-106 cell line was selected to establish SD rat osteosarcoma pulmonary metastasis model. After the SD rat osteosarcoma pulmonary metastasis model was established, the model of “pulmonary deficiency phlegm obstruction syndrome” was established through cold wind stimulation plus fumigation. Rats in osteosarcoma model group and osteosarcoma pulmonary metastasis model group were given 0.9% sodium chloride solution by gavage, and Qingfei Gujin group was given Qingfei Gujin Decoction by gavage for 8 weeks. The interventional effect of Qingfei Gujin Decoction on the body weight, primary tumor growth and pulmonary metastasis of each group was observed and analyzed. The contents of interleukin-10(IL-10) and transforming growth factor-β1(TGF-β1) in serum and tumors were detected by enzyme-linked immunosorbent assay(ELISA). Western blot was used to detect the expression of CXC motif receptor 4(CXCR4) in the right tibia of rats. [Results] Except blank control group, the rats in other three groups had visible right knee joint mass, X-ray examination showed destruction of right tibial metaphyseal bone and patchy density increase shadow; pathological examination showed large and deeply stained osteosarcoma cells in right tibia, and metastatic osteosarcoma nodules in lung tissue. Compared with osteosarcoma model group, the body weight of osteosarcoma pulmonary metastasis model group was lighter, while that of Qingfei Gujin group was heavier(P<0.05). Compared with osteosarcoma model group and osteosarcoma pulmonary metastasis model group, the mass and volume of tumor in Qingfei Gujin group were significantly reduced(P<0.05, P<0.01). The number of pulmonary metastases was the most in osteosarcoma lung metastasis model group, and the number of grade Ⅲ+Ⅳ nodules was the highest, while the number of Qingfei Gujin group were the lowest. Compared with blank control group, the contents of serum IL-10, TGF-β1 and the expression of CXCR4 protein in right tibia of osteosarcoma model group and osteosarcoma pulmonary metastasis model group were increased, and the differences were statistically significant(P<0.05, P<0.01). Compared with osteosarcoma model group, the contents of IL-10, TGF-β1 and CXCR4 protein expression in the right tibia of osteosarcoma pulmonary metastasis model group increased, while the contents of IL-10, TGF-β1 and CXCR4 protein expression in the Qingfei Gujin group decreased significantly(P<0.05, P<0.01). [Conclusion] The“pulmonary deficiency phlegm obstruction syndrome” osteosarcoma pulmonary metastasis rat model of combination of disease and syndrome has a good clinical fit, high success rate and good stability. Qingfei Gujin Decoction has good anti-osteosarcoma pulmonary metastasis effect, and its mechanism may be related to the down-regulation of IL-10, TGF-β1 and CXCR4 expression.
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