摘要
目的 以乳腺癌原位移植模型为基础,通过慢性不可预知性应激联合孤养法构建肝气郁滞型三阴性乳腺癌伴肝转移小鼠模型.方法 采用随机数字表法将6周龄BALB/c雌性小鼠40只分为4组,每组10只:对照组,正常饲养2周后予以乳腺垫接种4T1-luc乳腺癌细胞株,后期不给于刺激处理;模型组,慢性不可预知性应激(CUS)联合孤养;6-羟基多巴胺组(6-OHDA组),CUS联合孤养+6-OHDA;解郁消癥颗粒组,CUS联合孤养+解郁消癥颗粒.观察对比各组小鼠一般状况、旷场试验、蔗糖水偏嗜实验、促肾上腺皮质激素释放激素(CRH)、皮质酮(CORT)及去甲肾上腺素(NE)水平、肝脏HE染色和肝转移细胞凋亡水平,并行小鼠活体成像检测感兴趣区域(ROI)肿瘤荧光强度,磁共振成像检测肝脏转移灶数量.结果 对照组、模型组、解郁消癥颗粒组和6-OHDA组糖水偏好、旷场试验中心区域的活动距离和穿梭次数得分,组间比较差异均有统计学意义(F=0.34,P＜0.01;F=363.90,P＜0.01;F=0.046,P＜0.001).模型组糖水偏好低于对照组,活动距离低于解郁消癥颗粒组,穿梭次数低于解郁消癥颗粒组,均P＜0.01.对照组、模型组、解郁消癥颗粒组和6-OHDA组中血清CRH、CORT和NE表达量组间比较差异均有统计学意义(F=2 501.00,P＜0.01;F=155.30,P＜0.001;F=149.80,P＜0.01).模型组 CRH、CORT 和 NE 表达量(87.71±5.33,198.90±5.64,371.00± 16.08)均高于解郁消癥颗粒组(48.87±3.33,156.00±5.60,262.90±7.98),均P＜0.01.解郁消癥颗粒组与模型组比较,ROI区域减少,肝转移数量减少,乳腺癌肝转移细胞凋亡指数增加,均P＜0.01.结论 以乳腺癌原位移植小鼠模型为基础,应用CUS联合孤养法能够良好模拟三阴性乳腺癌伴肝转移肝气郁滞证,可为探索肝气郁滞在三阴性乳腺癌伴肝转移中的证候实质及相关治疗药物的药理机制提供实验载体.
Abstract
Objective To establish a mouse model of liver qi depression triple negative breast cancer with liver metastasis by using chronic unpredictable stress combined with separation.Method In this experiment,6-week-old BALB/c female mice were selected.In the model group,4Tl-luc breast cancer cell line was inoculated in the breast pad for 2 weeks by chronic unpredictable stress(CUS)combined with solitary rearing method and continued to stimulate for 4 weeks.In the same way,6-hydroxydopamine group(6-OHDA group)and Jie Yu Xiao Zheng Granule group were established.Observe and compare the general condition of mice in each group,open field test,sucrose water preference test,serum HP A axis related hormones(CRH,CORT,NE)level,liver HE staining,liver metastasis cell apoptosis level,and the tumor fluo-rescence intensity and the number of liver metastases detected by mouse living imaging.Result There was a statistically significant difference between the control group,model group,Jie Yu Xiao Zheng Granule group,and 6-OHDA group in sugar water preference,activity distance and shuttle frequency scores in the central area of the open field experiment(F=0.34,P＜0.01;F=363.90,P＜0.01;F=0.046,P＜0.001).The preference for sugar water in the model group was significantly lower than the control group,the difference was statistically significant(P＜0.01).The activity distance of the model group was significantly lower than the Jie Yu Xiao zheng Granule group,the difference was statistically signifi-cant(P＜0.01).The shuttle frequency of the model group was significantly lower than the Jie Yu Xiao zheng Granule group,the difference was statistically significant(P＜0.01).The average expression levels of serum CRH,CORT,and NE in the control group,model group,Jie Yu Xiao zheng Granule group,and 6-OHDA group were statistically signifi-cant(F=2 501.00,P＜0.01;F=155.30,P＜0.001;F=149.80,P＜0.01).The expression level of CRH in the mod-el group was 87.71±5.33,which was significantly higher than the Jie Yu Xiao Zheng Granule group,which was 48.87± 3.33,with a statistically significant difference(P＜0.01).The expression level of CORT in the model group was 198.90± 5.64,which was significantly higher than that in the Jie Yu Xiao Zheng Granule group,which was 156.00±5.60,with a statistically significant difference(P＜0.01).The expression level of NE in the model group was 371.00±16.08,which was significantly higher than that in the Jie Yu Xiao Zheng Granule group,which was 262.90±7.98,with a statistically significant difference(P＜0.01).Compared with the Jie Yu Xiao Zheng Granule group,the model group showed a de-crease in the ROI area,a decrease in the number of liver metastases(both P＜0.01).Compared with the model group and the Jie Yu Xiao Zheng Granule group,the apoptosis index of liver metastasis cells of breast cancer in the Jie Yu Xiao Zheng Granule group increased,and the difference was statistically significant(P＜0.01).Conclusion Based on the xeno-graft mouse model of breast cancer,the application of chronic unpredictable stress combined with separation can well sim-ulate triple negative breast cancer with liver metastasis of liver qi depression type,which can provide an experimental car-rier for exploring the essence of stagnation of liver qi depression in triple negative breast cancer with liver metastasis and the pharmacological mechanism of related therapeutic drugs.
NETL
NSTL
万方数据