补肾活血汤对乳腺癌骨转移模型裸鼠Bax/Bcl-2及cleaved Caspase-3蛋白表达的影响  

Effect of Bushen Huoxuetang on Expression of Bax/Bcl-2 and Cleaved Caspase-3 in Nude Mice with Bone Metastasis of Breast Cancer

作　　者：朱蓉 郭雪斌 杨满英 陈雪珍 胡玉蝶 杨争[3] 袁博[3] 胡金辉[3] ZHU Rong;GUO Xuebin;YANG Manying;CHEN Xuezhen;HU Yudie;YANG Zheng;YUAN Bo;HU Jinhui(Yiyang First Hospital of Tradition Chinese Medicine,Yiyang 413000,China;Hunan University of Chinese Medicine,Changsha 410208,China;The First Hospital of Hunan University of Chinese Medicine,Changsha 410021,China)

机构地区：[1]益阳市第一中医医院,湖南益阳413000 [2]湖南中医药大学,长沙410208 [3]湖南中医药大学第一附属医院,长沙410021

出　　处：《中国实验方剂学杂志》2023年第8期133-141,共9页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：湖南省教育厅基金项目(20A373);湖南中医药大学校级基金项目(2019XJJJ039);湖南省科技厅科普专项(2021ZK4108);湖南省中医药管理局重点基金项目(C2022020)。

摘　　要：目的:观察补肾活血汤对乳腺癌骨转移模型裸鼠细胞凋亡及B细胞淋巴瘤-2(Bcl-2)相关X蛋白(Bax)/Bcl-2及切割型胱天蛋白酶-3(cleaved Caspase-3)蛋白表达的影响,探讨其抑制骨质破坏的作用机制。方法:将30只BALB/c雌性裸鼠随机分成空白组6只、造模组24只。将乳腺癌细胞株4T1经裸鼠右下肢胫骨注射构建乳腺癌骨转移模型,将成膜裸鼠随机分为模型组、补肾活血汤组、唑来膦酸组、联合用药组,每组6只。补肾活血汤组予以补肾活血汤中药灌胃36.67 g·kg^(-1),每日1次,唑来膦酸组予以皮下注射100μg·kg^(-1),每周2次,联合用药组给予相同剂量补肾活血汤组灌胃联合唑来膦酸皮下注射,正常组和模型组予以等体积蒸馏水灌胃,连续14 d,次日脊柱脱臼法处死,观察裸鼠一般情况及体质量变化,截取每组裸鼠右下肢,称重,X射线扫描及苏木素-伊红(HE)染色观察裸鼠骨组织形态学变化、原位末端标记法(TUNEL)检测裸鼠骨组织凋亡情况,蛋白免疫印迹法(Western blot)检测裸鼠骨组织中Bax/Bcl-2及cleaved Caspase-3蛋白表达情况。结果:与空白组比较,模型组第5~15天裸鼠体质量显著减轻(P<0.01),右下肢组织质量显著增加(P<0.01),与模型组比较,补肾活血汤组、唑来膦酸组、联合用药组第10~15天裸鼠体质量显著增加(P<0.01),右下肢组织质量显著降低(P<0.01)。HE染色显示与空白组比较,模型组、补肾活血汤组、唑来膦酸组、联合用药组裸鼠肿瘤细胞异型性明显,核染色深,骨转移明确。X射线扫描示与空白组比较,模型组裸鼠右下肢出现明显溶骨性破坏,胫骨近段、膝关节完整结构基本丧失,与模型组比较,补肾活血汤组、唑来膦酸组、联合用药组右下肢溶骨性病变明显减少,部分结构完整,对骨质破坏具有抑制作用。TUNEL染色显示与空白组比较,模型组骨转移瘤细胞凋亡率降低,与模型组比较,补肾活血汤组、唑来膦酸组、�Objective:To study the effect of Bushen Huoxuetang on the apoptosis and the expression of B-cell lymphoma(Bcl-2)-associated X protein(Bax)/Bcl-2 and cleaved cysteine-containing aspartate proteolytic enzyme-3(cleaved Caspase-3)in the nude mouse model of bone metastasis of breast cancer,and explore the mechanism of Bushen Huoxuetang in inhibiting bone destruction.Method:Thirty BALB/c female nude mice were randomly assigned into blank group(n=6)and model group(n=24).The suspension of 4T1breast cancer cells was injected into the tibia of mouse right lower limb to establish model of bone metastasis of breast cancer.The successfully modeled nude mice were randomly assigned into model group,Bushen Huoxuetang group,zoledronic acid group,and combined drug group,with 6 mice in each group.Bushen Huoxuetang was administrated at a dose of 36.67 g·kg^(-1),once a day,and zoledronic acid was administrated by subcutaneous injection at a dose of 100μg·kg^(-1),twice a week.The combined drug group was administrated with the same doses of Bushen Huoxuetang group by gavage and zoledronic acid by subcutaneous injection.The mice in the blank group and the model group were administrated with the same volume of distilled water by gavage for 14 days.On the next day at the end of drug administration,the mice were sacrificed by cervical dislocation.The general situation and weight changes of the mice were examined.The right lower limb was collected,and X-ray scanning and hematoxylin-eosin(HE)staining methods were used for observation of pathological changes in the bone.The terminal deoxynucleotidyl transferase d UTP nick-end labeling(TUNEL)was employed to detect the apoptosis of bone tissue in nude mice,and Western blot to determine the expression of Bax/Bcl-2 and cleaved Caspase-3 in the bone tissue.Result:Compared with the blank group,the modeling reduced the body weight(P<0.01)and increased the right lower limb weight of the nude mice(P<0.01).Compared with the model group,Bushen Huoxuetang,zoledronic acid,and their combination increase

关 键 词：补肾活血汤 乳腺癌骨转移 凋亡 B细胞淋巴瘤-2(Bcl-2) Bcl-2相关X蛋白(Bax) 切割型胱天蛋白酶-3(cleaved Caspase-3) 

分 类 号：R22[医药卫生—中医基础理论] R242[医药卫生—中医学] R2-031R285.5