阳和化岩汤通过ER/PI3K/Akt/mTOR通路逆转乳腺癌他莫昔芬耐药

目的:通过观察阳和化岩汤对他莫昔芬(TAM)耐药型乳腺癌移植瘤的作用,及对雌激素受体(ER)/磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)/哺乳动物雷帕霉素靶蛋白(mTOR)交互通路的影响,探讨阳和化岩汤逆转乳腺癌TAM耐药的可能机制。方法:50只小鼠随机分为5组,空白组、模型组、阳和化岩汤组、依维莫司组以及阳和化岩汤+依维莫司组。通过切除双侧卵巢建立肾虚模型,其中空白组行假手术处理。造模后恢复3 d,5组小鼠均通过皮下肿瘤接种乳腺癌TAM耐药细胞(MCF-7/TAM-)建立乳腺癌TAM耐药型移植瘤模型。建模成功后,阳和化岩汤组给予阳和化岩汤灌胃(给药剂量为中药制剂20 mL·kg^-1),依维莫司组给予依维莫司腹腔注射(10 mg·kg^-1),阳和化岩汤+依维莫司组给予阳和化岩汤灌胃+依维莫司腹腔注射,空白组、模型组给予磷酸盐缓冲液(PBS)灌胃+腹腔注射;均连续给药28 d,1次/d。给药结束后分离肿瘤组织、称质量,计算抑瘤率;苏木素-伊红(HE)染色观察肿瘤组织病理学改变;免疫荧光和实时荧光定量聚合酶链式反应(Real-time PCR)分别检测肿瘤组织PI3K,Akt,mTOR,ER蛋白和mRNA表达情况。结果:与模型组比较,阳和化岩汤组d12,d20,d28肿瘤体积、瘤质量显著降低(P<0.01),抑瘤率显著增加(P<0.01);阳和化岩汤组能明显降低肿瘤细胞密度,引起肿瘤细胞坏死;与模型组比较,阳和化岩汤组、依维莫司组和阳和化岩汤+依维莫司组均能抑制PI3K,Akt,mTOR蛋白及mRNA的表达(P<0.05,P<0.01);与空白组比较,阳和化岩汤组、依维莫司组和阳和化岩汤+依维莫司组均能抑制ER蛋白及mRNA的表达(P<0.01);与模型组比较,阳和化岩汤+依维莫司组ER mRNA表达量显著减少(P<0.01)。结论:阳和化岩汤可抑制TAM耐药型乳腺癌移植瘤生长,其机制可能是通过下调ER/PI3K/Akt/mTOR交互信号通路关键分子的表达,从而达到逆转乳腺癌TAM耐药的作用。

Yanghe Huayantang Reverses Tamoxifen Resistance in Breast Cancer Through ER/PI3K/Akt/mTOR Pathway

Objective:To explore the possible mechanism of Yanghe Huayantang in reversing the drug resistance of breast cancer by observing the effect of Yanghe Huayantang on the transplant tumor of tamoxifen(TAM)-resistant breast cancer and its influences on the interaction pathway of estrogen receptor(ER)/phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)/mammalian rapamycin target protein(mTOR).Method: Fifty mice were randomly divided into 5 groups:blank group,model group,Yanghe Huayantang group,everolimus group,and Yanghe Huayantang+everolimus group. The model of kidney deficiency was established by bilateral ovariectomy,and the blank group was treated with sham operation. Three days after the establishment of the model,all the five groups of mice were inoculated with breast cancer TAM drug-resistant cells(MCF-7/TAM-)to establish breast cancer TAM-resistant transplanted tumor model. After successful modeling,Yanghe Huayantang group received intragastric administration of Yanghe Huayantang(traditional Chinese medicine preparation 20 mL·kg^-1),everolimus group received intraperitoneal injection of everolimus(10 mg·kg^-1). Yanghe Huayantang + everolimus group received Yanghe Huayantang by intragastric administration and everolimus by intraperitoneal injection. The blank group and model group received intragastric administration and intraperitoneal injection of phosphate buffer(PBS). Drug administration was lasted for 28 days in all groups,once a day. After administration,the tumor tissue was separated and weighed,and the tumor inhibition rate was calculated. Hematoxylin-eosin(HE) staining was used to observe the pathological changes of tumor tissue. Immunofluorescence and Real-time fluorescence quantitative polymerase chain reaction(Real-time PCR)were used to detect the expression of PI3K,Akt,mTOR,ER protein and mRNA in tumor tissue. Result: Compared with the model group,the tumor volume and tumor weight of Yanghe Huayantang group decreased significantly on the 12 th,20 th and 28 th days(P<0.01),and the tumor inhibition rate increased significantly(P<0.01). Yanghe Huayantang group significantly reduced the density of tumor cells and caused tumor cell necrosis. Compared with the model group,Yanghe Huayantang group,everolimus group and Yanghe Huayantang+everolimus group inhibited the expression of PI3K,Akt,mTOR protein and mRNA(P<0.05,P<0.01). Compared with the blank group,Yanghe Huayantang group,everolimus group and Yanghe Huayantang+everolimus group all inhibited the protein and mRNA expression of ER,and mRNA expression of ER in Yanghe Huayantang+everolimus group was significantly lower than that in the model group(P<0.01). Conclusion: Yanghe Huayantang can inhibit the growth of TAM-resistant breast cancer. The mechanism may be that Yanghe Huayantang can reverse the TAM resistance of breast cancer by down-regulating the expression of key molecules of ER/PI3K/Akt/mTOR cross-signal pathway.