23-羟基白桦酸调控免疫细胞干预NNK联合LPS致小鼠肺部炎癌转化的作用  

23-Hydroxybetulinic Acid Inhibits NNK and LPS-induced Lung Inflammation-related Carcinogenesis by Regulating Immune Cells in Mice

作　　者：刘彭浩邦 段文彬 陈亚娟 陈兰英[1,2] LIU Penghaobang;DUAN Wenbin;CHEN Yajuan;CHEN Lanying(Jiangxi University of Chinese Medicine,Nanchang 330004,China;National Engineering Research Center for Manufacturing Technology of Traditional Chinese Medicine Solid Preparation,Nanchang 330006,China)

机构地区：[1]江西中医药大学,南昌330004 [2]中药固体制剂制造技术国家工程研究中心,南昌330006

出　　处：《中国实验方剂学杂志》2025年第6期98-106,共9页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(82060732,81860720)。

摘　　要：目的:观察白头翁主要成分23-羟基白桦酸(23-HBA)抑制4-甲基亚硝胺基-1-3-吡啶基-1-丁酮(NNK)联合脂多糖(LPS)致小鼠肺部炎癌转化作用及对免疫细胞的影响,并初步探索该作用机制。方法:建立NNK联合LPS诱导的小鼠炎癌转化模型,随机分为空白组,模型组,阿司匹林组(10 mg·kg^(-1)),23-HBA低、中、高剂量组(3.75、7.5、15 mg·kg^(-1)),持续给药26周,实验结束后取脾脏、肺脏及外周血。计算肺脏、脾脏指数;苏木素-伊红(HE)染色观察小鼠肺组织病理变化;免疫组化(IHC)检测肺组织中甲状腺转录因子-1(TTF-1)、神经元特异性烯醇化酶(NSE)和增殖细胞核抗原(Ki-67)的表达水平;高通量蛋白芯片检测小鼠血清中白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)和肿瘤坏死因子-α(TNF-α)含量;流式细胞术(FACS)检测肺组织和脾组织中巨噬细胞、髓源性抑制细胞(MDSC)、耗竭性T淋巴细胞表达情况。运用口袋分子对接的方法预测23-HBA对Janus激酶2(JAK2)、含Src同源2结构域蛋白酪氨酸磷酸酶(SHP2)蛋白与细胞因子信号抑制物3(SOCS3)的对接能。蛋白免疫印迹法(Western blot)检测23-HBA对M1活化巨噬细胞与肺腺癌细胞A549中磷酸化信号传导及转录激活蛋白3(p-STAT3)及其下游蛋白p53与SHP2蛋白的表达水平。结果:与正常组比较,模型组小鼠肺脏、脾脏指数均有不同程度地升高(P<0.05,P<0.01),TTF-1、NSE与Ki-67蛋白表达明显上升(P<0.05,P<0.01),血清TNF-α、IL-1β、IL-6含量均显著升高(P<0.01),巨噬细胞的数量显著降低(P<0.01),耗竭型T细胞和MDSCs的数量明显升高(P<0.05,P<0.01)。与模型组比较,23-HBA各剂量组小鼠脾脏和胸腺指数明显下降(P<0.05),23-HBA中剂量组肺指数明显下降(P<0.05),23-HBA高、中剂量组可改善模型组NNK联合LPS致小鼠肺部炎性浸润灶与恶性病变灶的发生,23-HBA中、高剂量组TTF-1较模型组表达明显下降(P<0.05,P<0.01),23-HBA各剂量组小鼠较模型组NSE�Objective:To investigate the suppressive effect of 23-hydroxybetulinic acid(23-HBA),a key constituent of Pulsatillae Radix,on the pulmonary inflammation-related carcinogenesis induced by the combined exposure of 4(-methylnitrosamino)-1(-3-pyridyl)-1-butanone(NNK)and lipopolysaccharide(LPS)in mice,alongside exploring its influence on immune cells and delving into the underlying mechanisms.Methods:A murine model of pulmonary inflammation-related carcinogenesis induced by NNK combined with LPS was established.Mice were randomly assigned into blank control,model,aspirin(10 mg·kg^(-1)),and low-,medium-,and high-dose(3.75,7.5,15 mg·kg^(-1),respectively)23-HBA groups.The treatment lasted for 26 weeks,after which the spleen,lung,and peripheral blood samples were collected.Lung and spleen indices were calculated.Histopathological changes in the lung tissue were observed by hematoxylin-eosin staining.Immunohistochemistry was employed to assess the expression levels of thyroid transcription factor-1(TTF-1),neuron-specific enolase(NSE),and proliferating cell nuclear antigen(Ki-67)in the lung tissue.High-throughput protein microarray was employed to measure the levels of interleukin-1β(IL-1β),interleukin-6(IL-6),and tumor necrosis factor-α(TNF-α)in the mouse serum.Flow cytometry was employed to evaluate the expression of macrophages,myeloid-derived suppressor cells(MDSCs),and exhausted T lymphocytes in the lung and spleen tissue.Molecular docking was performed to predict the binding affinity of 23-HBA to Janus kinase 2(JAK2),Src homology 2 domain-containing phosphatase 2(SHP2),and suppressor of cytokine signaling 3(SOCS3).Western blot was performed to assess the protein levels of phosphorylated-signal transducer and activator of transcription 3(p-STAT3),p53,and SHP2 in the M1-activated macrophages and A549 lung adenocarcinoma cells treated with 23-HBA.Results:Compared with the normal group,the lung and spleen indexes of the model group were increased to varying degrees(P<0.05,P<0.01),the expression of TTF-1,NSE and Ki-6

关 键 词：23-羟基白桦酸 4-甲基亚硝胺基-1-3-吡啶基-1-丁酮(NNK)联合脂多糖(LPS) 肺部组织 炎癌转化 免疫细胞 

分 类 号：R242[医药卫生—中医临床基础] R285[医药卫生—中医学] R734