基于MEKK1/SEK1/JNK1/AP-1通路探讨三七总皂苷对4T1乳腺癌荷瘤小鼠肿瘤模型的影响

Effect of Panax Notoginseng Saponins on Breast Cancer Cell Line 4T1 in Tumor-bearing Mice Through MEKK1/SEK1/JNK1/AP-1 Pathway

目的:通过研究三七总皂苷对4T1乳腺癌荷瘤小鼠促分裂原激活的蛋白激酶的激酶1(MEKK1)/应激激活的蛋白激酶/细胞外信号调节的蛋白激酶(SAPK/Erk激酶,SEK1)/c-Jun氨基末端激酶1(JNK1)/激活蛋白-1(AP-1)通路的影响,探讨三七总皂苷抑制肿瘤的可能机制。方法:建立4T1乳腺癌荷瘤小鼠肿瘤模型,将48只造模成功的小鼠随机分为三七总皂苷低、中、高剂量组(10,20,40 mg·kg^-1)和模型组,每组12只,三七总皂苷各剂量组腹腔注射剂量为10 mL·kg^-1,模型组给予相同剂量的生理盐水,连续给药28 d。给药结束后分离肿瘤组织、称质量、切片、匀浆等,采用脱氧核苷酸末端转移酶(TdT)介导的dUTP缺口末端标记法(TUNEL)染色检测肿瘤细胞凋亡;实时荧光定量聚合酶链式反应(Real-time PCR)检测肿瘤组织MEKK1,SEK1,JNK1,AP-1 mRNA表达;蛋白免疫印迹法(Western blot)检测4T1乳腺癌荷瘤小鼠肿瘤组织MEKK1,SEK1,JNK1,AP-1蛋白表达。结果:与模型组比较,三七总皂苷中、高剂量组的肿瘤质量明显下降(P<0.05);肿瘤细胞凋亡数量随三七总皂苷剂量的升高明显升高(P<0.05);三七总皂苷中、高剂量组肿瘤组织中MEKK1,SEK1,JNK1,AP-1 mRNA和蛋白表达明显升高(P<0.05)。结论:三七总皂苷对4T1乳腺癌荷瘤小鼠肿瘤模型具有抑制作用,其作用机制可能与激活MEKK1/SEK1/JNK1/AP-1信号通路有关。

Objective:To explore the potential mechanisms of Panax Notoginseng Saponins(PNS)on growth inhibition of breast cancer cell line 4T1 in tumor-bearing mice by investigating the mitogen-activated protein kinase kinase kinase 1(MEKK1)/stress activated protein kinase(SAPK)/extracellular regulated protein kinases(Erk)Kinase(SEK1)/c-Jun N-terminal kinase 1(JNK1)/activator protein-1(AP-1)signaling pathways.Method:The 4T1 breast cancer mice model was established.Forty-eight mice with successful modeled and randomly divided into the low,medium and high-dose PNS groups(10,20,40 mg·kg^-1)and the model control group(12 mice in each group).The PNS groups received intraperitoneal injection with dosage of 10 mL·kg^-1,while the controlled group was given the same dosage of saline.After administration with PNS for 28 days,tumor tissues were isolated,weighed,sliced and homogenized.Tumor cell apoptosis was detected by TdT mediated-dUTP nick end labeling(TUNEL)staining.The mRNA expressions of MEKK1,SEK1,JNK1 and AP-1 in tumor tissue were detected by Real-time polymerase chain reaction(Real-time PCR).The protein expressions of MEKK1,SEK1,JNK1 and AP-1 in tumor tissue were detected by immunofluorescence staining and Western blot.Result:Compared with model group,the tumor weights of medium-dose and high-dose PNS groups were decreased significantly(P<0.05).TUNEL staining showed that the number of apoptotic tumor cells increased with the rise of dosage of PNS(P<0.05).The medium-dose and high-dose PNS groups showed a significant increase in the mRNA expressions of MEKK1,SEK1,JNK1 and AP-1 as well as the protein expressions of MEKK1,SEK1,JNK1 and AP-1 in tumor tissues(P<0.05),with statistically significant differences(P<0.05).Conclusion:PNS could inhibit the tumor growth of breast cancer cell line 4T1 in tumorbearing mice,which may be related to the activation of MEKK1/SEK1/JNK1/AP-1 signaling pathways.