香砂六君子汤合除痰解毒中药对Lewis肺癌小鼠免疫逃逸的影响  

Effect of Xiangsha Liu Junzitang Combined with Phlegm-removing and Detoxifying Traditional Chinese Medicine on Immune Escape in Lewis Lung Cancer Mice

作　　者：杨倩 王淑美 冯诗函 詹雪 夏琳涛 吴明星 YANG Qian;WANG Shumei;FENG Shihan;ZHAN Xue;XIA Lintao;WU Mingxing(Chongqing Key Laboratory of Traditional Chinese Medicine(TCM)for Prevention and Cure of Metabolic Diseases,Chongqing 400016,China;College of TCM,Chongqing Medical University,Chongqing 400016,China)

机构地区：[1]中医药防治代谢性疾病重庆市重点实验室,重庆400016 [2]重庆医科大学中医药学院,重庆400016

出　　处：《中国实验方剂学杂志》2024年第7期78-86,共9页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家中医临床研究基地业务建设科研专项(JDZX2015073);重庆市自然科学基金面上项目(CSTB2022NSCQ-MSX1564)。

摘　　要：目的:观察香砂六君子汤合除痰解毒中药对Lewis肺癌小鼠免疫逃逸的作用及机制。方法:取60只SPF级C57BL/6J雄鼠,于右侧腋中线皮下注射Lewis细胞悬液0.2 m L(含细胞数2×10^(6)个/m L),接种7 d后,将造模成功的小鼠随机分为模型组、顺铂组、香砂六君子汤低、中、高剂量组、联合组,每组10只。香砂六君子汤低、中、高剂量组分别灌胃17.88、35.75、71.50 g·kg^(-1)香砂六君子汤加减溶液,1次/d,顺铂腹腔注射小鼠用量换算为5 mg·kg^(-1),2次/周,各组均连续干预14 d,每2 d测量肿瘤体积。治疗结束后称取各组小鼠肿瘤质量并计算抑瘤率;苏木素-伊红(HE)染色观察各组肿瘤的形态学特征;实时荧光定量聚合酶链式反应(Real-time PCR)检测各组肿瘤组织中自然杀伤(NK)细胞杀伤受体2(NKG2D)、核糖核酸输出因子-1(RAE-1)、γ干扰素(IFN-γ)mRNA含量;免疫组化法(IHC)、蛋白免疫印迹法(Western blot)检测各组肿瘤RAE-1、NKG2D、IFN-γ表达;Western blot检测各组小鼠肿瘤中IL-6、JAK2、p-JAK2、STAT3、p-STAT3及脾脏组织中NKG2D、RAE-1蛋白含量。结果:与模型组比较,香砂六君子汤各剂量组肿瘤质量均下降,差异无统计学意义;肿瘤体积均缩小(P<0.05,P<0.01);病理学形态改善;香砂六君子汤中剂量组NKG2D、RAE-1、IFN-γ mRNA含量均升高(P<0.05,P<0.01),肿瘤组织中NKG2D、RAE-1、IFN-γ蛋白表达均升高(P<0.05,P<0.01),p-JAK2、p-STAT3蛋白表达下降(P<0.05,P<0.01);脾脏组织中,香砂六君子汤各剂量组NKG2D、RAE-1蛋白表达均显著升高(P<0.01)。与顺铂组比较,香砂六君子汤中剂量组NKG2D、RAE-1、IFN-γ mRNA含量升高,差异无统计学意义;IHC显示联合组NKG2D、IFN-γ蛋白表达显著升高(P<0.01),Western blot结果表明RAE-1、NKG2D、IFN-γ蛋白表达升高(P<0.05,P<0.01);联合组p-JAK2、p-STAT3蛋白表达下降(P<0.05,P<0.01);脾脏组织中,香砂六君子汤中剂量组、联合组NKG2D、RAE-1蛋白表达显著升高(P<0.01)�Objective:To study the effect and mechanism of Xiangsha Liu Junzitang combined with phlegm-removing and detoxifying traditional Chinese medicine on immune escape in Lewis lung cancer mice.Method:A total of 60 specific-pathogen-free (SPF)-grade C57BL/6J male mice were injected subcutaneously with 0.2 m L of Lewis cell suspension (containing 2×10~6 cells·m L^(-1)) in the right mid-axillary line.After 7 days,the mice that had been successfully modeled were randomly divided into six groups:the model group,the cisplatin group,the Xiangsha Liu Junzitang low-,medium-,and high-dose groups,and the combined group,with 10 mice in each group.The Xiangsha Liu Junzitang low-,medium-and high-dose groups were gavaged with 17.88,35.75,71.50 g·kg^(-1) Xiangsha Liu Junzitang solution once a day,respectively,and the dosage of cisplatin intraperitoneally injected into the mice was converted to 5 mg·kg^(-1) twice a week,and the tumour volumes of each group were measured every two days.The intervention lasted for 14 consecutive days.At the end of treatment,the tumour mass of mice in each group was weighed and the tumour inhibition rate was calculated.The morphological characteristics of tumours in each group were observed by hematoxylin-eosin(HE) staining.Fluorescent quantitative real-time polymerase chain reaction (Real-time PCR) assay was used to detect messenger ribonucleic acid (mRNA) contents of the natural killer group 2 member D (NKG2D) receptor,ribonucleic acid export-1 (RAE-1),and γ interferon (IFN-γ) in the tumour tissues of each group.NKG2D,RAE-1,and IFN-γ mRNA in tumour tissues of each group.Immunohistochemistry (IHC) and Western blot were applied to detect the expressions of RAE-1,NKG2D,and IFN-γ in tumour tissues of each group,and Western blot was used to detect the expressions of interleukin-6 (IL-6),Janus kinase 2 (JAK2),p-JAK2,signal transducer and activator of transcription 3 (STAT3),and p-STAT3 in tumour tissues of each group,as well as the protein levels of NKG2D,and RAE-1 in spleen tissues of each group.Res

关 键 词：香砂六君子汤 LEWIS肺癌 免疫逃逸 NK细胞杀伤受体2(NKG2D) 核糖核酸输出因子-1(RAE-1) 

分 类 号：R2-0[医药卫生—中医学] R22R242R285.5