温下方乙酸乙酯提取部位调控CAFs介导的Hh-Gli1通路抑制A549移植瘤生长的作用机制  

Mechanism of ethyl acetate extract from Wenxia Formula on regulating Hedgehog-Gli1 signaling pathway mediated by cancer associated fibroblasts for inhibiting anti-A549 transplanted tumor

作　　者：王盟 毕倩宇[1] 季旭明 WANG Meng;BI Qian-yu;JI Xu-ming(School of Basic Medical Science,Zhejiang Chinese Medical University,Hangzhou 310053,China;Department of Medicine,Jining No.1 People's Hospital,Jining 272000,China)

机构地区：[1]浙江中医药大学基础医学院,杭州310053 [2]济宁市第一人民医院医务部,济宁272000 [3]浙江中医药大学基础医学院方剂教研室,杭州310053

出　　处：《中华中医药杂志》2021年第9期5499-5504,共6页China Journal of Traditional Chinese Medicine and Pharmacy

基　　金：国家自然科学基金项目(No.81573871,No.81774198)。

摘　　要：目的:观察温下方乙酸乙酯提取部位(WFEA)对肿瘤相关成纤维细胞(CAFs)介导的Hh-Gli1通路的影响,探讨其抑制A549移植瘤生长的作用机制。方法:建立肺癌A549裸鼠皮下移植瘤模型,随机分为A549组、模型组、顺铂组,WFEA高、中、低剂量组和环巴胺组,药物干预5周。采用超高压液相色谱-飞行时间质谱仪(UPLC-Q-TOF-MS)分析WFEA的主要化学成分;采用小动物活体成像、HE染色、免疫组化等方法检测WFEA在A549移植瘤裸鼠体内的抗肿瘤活性;采用Western blot法和实时PCR法检测CAFs介导的Hh-Gli1信号通路水平。结果:采用UPLC-Q-TOF-MS方法共定性出WFEA的15种化学成分;与模型组比较,WFEA各给药组裸鼠肿瘤的光子强度均有不同程度的降低(P<0.05,P<0.01)。模型组HE染色可见细胞排列紧密,瘤细胞形态不规则,未见明显的坏死,WFEA各给药组肿瘤细胞均有不同程度坏死,胞浆破裂,细胞膜不完整;与模型组比较,WFEA各给药组FN蛋白表达以及PTCH1、SMO和Gli1的蛋白和mRNA表达水平均有不同程度的降低(P<0.01,P<0.05)。结论:WFEA对A549移植瘤有一定的抑制作用,其作用机制可能与抑制CAFs介导的Hh-Gli1信号通路有关。Objective: To observe the effects of ethyl acetate extract(WFEA) of wenxia Formula on cancer-associated fibroblasts(CAFs)-mediated Hh-Gli1 pathway, and to explore its inhibitory effect on the growth of A549 transplanted tumor and mechanism. Methods: The A549 nude mice subcutaneous transplanted tumor model of lung cancer was established and randomly divided into A549 group, model group, cisplatin group, WFEA high-, medium-, low-dose group and cyclopamine group.Drug intervention lasted for 5 weeks. UPLC-Q-TOF-MS was used to analyze the main chemical components of the extracted part of WFEA;tumor volume, tumor weight, small animal imaging, HE staining, immunohistochemistry were used to detect the antitumor activity of WFEA in A549 nude mice;Western blot and real-time PCR were used to detect the level of CAFs mediated Hh-Gli1 signal pathway, and the possible mechanism of its inhibition of tumor was revealed. Results: Chemical constituents of WFEA were identified by UPLC-Q-TOF-MS;Compared with the model group, the photon intensity of tumor in nude mice in each administration group of wfea decreased in varying degrees(P<0.05, P<0.01). In the model group, he staining showed that the cells were closely arranged, the tumor cells were irregular in shape, and there was no obvious necrosis. The tumor cells in each administration group of wfea had different degrees of necrosis, cytoplasmic rupture and incomplete cell membrane;Compared with the model group, the expression of FN protein and the expression levels of protein and mRNA of PTCH1, SMO and Gli1 in wfea groups decreased in varying degrees(P<0.01, P<0.05). Conclusion: The WFEA under the temperature has a certain inhibitory effect on A549, and its mechanism may be related to the intervention of CAFs mediated Hh-Gli1 signal pathway.

关 键 词：温下方 肿瘤相关成纤维细胞(CAFs) 超高压液相色谱-飞行时间质谱仪(UPLC-Q-TOF-MS) Hh-Gli1 乙酸乙酯提取部位 A549移植瘤 肺癌 

分 类 号：R285.5[医药卫生—中药学]