附子-白术药对对乳腺癌骨转移裸鼠TGF-β/Smads/Gli2/PTHrP信号通路的影响

Effect of Aconiti Lateralis Radix Praeparata-Atractylodis Macrocephalae Rhizoma on TGF-β1/Smads/Gli2/PTHrP Signaling Pathway in Nude Mice with Bone Metastasis of Breast Cancer

目的:研究附子-白术药对对乳腺癌骨转移裸鼠的保护作用及其作用机制。方法:裸鼠随机分为正常组、模型组、唑来膦酸组、附子-白术药对组,采用左心室注射乳腺癌细胞MDA-MB-231BO法建立骨转移模型,并进行相应干预。观察裸鼠骨转移程度,微计算机断层扫描(Micro-CT)检测裸鼠胫骨矿物质密度(BMD),骨体积(BV),选取感兴趣区域(ROI)体积(TV),计算骨体积分数BV/TV,抗酒石酸酸性磷酸酶染色法(TRAP)观察骨转移灶中破骨细胞表达,蛋白免疫印迹法(Western blot)法检测转化生长因子-β_1(TGF-β1),Smad3,Smad4,胶质瘤相关癌基因同源蛋白2(Gli2)和甲状旁腺激素相关蛋白(PTHr P)的表达。结果:与模型组比较,附子-白术药对组骨转移程度显著降低(P<0.01),裸鼠胫骨BMD,BV/TV显著升高(P<0.01),TRAP(+)细胞数量显著减少(P<0.01),骨转移灶中TGF-β_1,Smad3,Gli2和PHTr P表达均显著降低(P<0.01),Smad4表达均明显升高(P<0.05)。结论:附子-白术药对可减轻乳腺癌骨转移裸鼠溶骨性损伤,其机制可能与其上调Smad4蛋白表达,下调TGF-β1,Smad3,Gli2和PTHr P蛋白表达,调控TGF-β1/Smads/Gli2/PTHr P信号通路有关。

Objective： To investigate the therapeutic efficacy of Aconiti Lateralis Radix PraeparataAtractylodis Macrocephalae Rhizoma on nude mice with bone metastasis of breast cancer and its mechanism.Method： Nude mice were randomly divided into normal group,model group,zoledronate group and Aconiti Lateralis Radix Praeparata-Atractylodis Macrocephalae Rhizoma group. Except for normal group,MDA-MB-231 BO cells were injected into the left ventricle to create the model of bone metastasis of breast cancer. Aconiti Lateralis Radix Praeparata-Atractylodis Macrocephalae Rhizoma group nude mice were orally administered with Aconiti Lateralis Radix Praeparata-Atractylodis Macrocephalae Rhizoma. Normal group,model group and zoledronate group nude mice were orally administered with physiological saline,and zoledronate was injected into the abdominal cavity in zoledronate group. The degree of bone metastases was evaluated. Bone mineral density（ BMD）,bone volume（ BV）,range of interests（ ROI） and total volume（ TV） were evaluated by micro-CT. Osteoclasts in bone metastasis were evaluated by tartrate resistant acid phosphatase dye（ TRAP）. Transforming growth factor-β1（ TGF-β1）,drosophila mothers against decapentaplegic protein 3（ Smad3）, Smad4, Glioma-associated oncogene homologue 2（ Gli2） and Parathyroid hormone-related protein（ PTHr P） in bone metastasis tissues were evaluated by Western blot. Result： The degree of bone metastases,BMD and BV/TV in Aconiti Lateralis Radix PraeparataAtractylodis Macrocephalae Rhizoma group were higher than those of model group（ P〈0. 01）. The number of TRAP（ ＋） cells in Aconiti Lateralis Radix Praeparata-Atractylodis Macrocephalae Rhizoma group was lower than that of model group（ P〈0. 01）. The expression of TGF-β1,Smad3,Gli2 and PHTr P protein expressions of bone metastases tissues in Aconiti Lateralis Radix Praeparata-Atractylodis Macrocephalae Rhizoma group were lower than those of model group（ P〈0. 01）,and the expression of Smad4 in Aconiti Lateralis Radix Praeparata-Atractylodis Macrocephalae Rhizoma group was higher than that of model group（ P〈0. 05）. Conclusion： Aconiti Lateralis Radix Praeparata-Atractylodis Macrocephalae Rhizoma has a therapeutic effect on soluble osseous injury in nude mice with bone metastasis of breast cancer,and the mechanisms of action may be that Aconiti Lateralis Radix Praeparata-Atractylodis Macrocephalae Rhizoma could up-regulate Smad4,down-regulate TGF-β1,Smad3,Gli2 and PTHr P,and regulate TGF-β1/Smads/Gli2/PTHrP signaling pathway.