培元抗癌汤对人乳腺癌MDA-MB-231细胞裸鼠移植瘤生长及细胞凋亡的影响

Effect of Peiyuan Anticancer Decoction on growth and apoptosis of human breast cancer MDA-MB-231 cell xenograft in nude mice

目的探讨培元抗癌汤对人乳腺癌MDA-MB-231细胞裸鼠移植瘤生长及细胞凋亡的影响。方法选取30只BALB/c-nu雌性裸鼠,通过皮下注射MDA-MB-231细胞建立人乳腺移植瘤模型。将建模达标的载瘤鼠随机分为对照组、阳性对照组以及低、中、高剂量组,每组6只。低、中、高剂量组分别给予200、300、400 mg/kg培元抗癌汤灌胃,阳性对照组给予皮下注射乌司他丁(3 mg/kg),对照组不做任何干预治疗。给药期间观察各组载瘤鼠的一般情况。连续给药21 d后,处死各组载瘤鼠,取出乳腺肿瘤称重并计算肿瘤的抑制率,苏木精-伊红染色观察肿瘤组织形态学,检测肿瘤组织B细胞淋巴瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)、含半胱氨酸的天冬氨酸蛋白水解酶(Caspase)-9和Caspase-7蛋白表达水平。取人乳腺癌MDA-MB-231细胞,设立细胞空白组、3μmol/L乌司他丁组以及200μmol/L、300μmol/L、400μmol/L培元抗癌汤组;除细胞空白组外其他组加入相应剂量的药物,干预48 h后检测细胞凋亡率。结果对照组载瘤鼠体重减轻、活动减少、反应迟钝;与对照组比较,各给药组载瘤鼠体重增加、反应较为灵敏。对照组、低剂量组、高剂量组、阳性对照组、中剂量组的肿瘤抑制率依次升高(均P<0.05)。HE染色结果显示,对照组和低剂量组可见明显异形细胞,细胞核增大,核异性明显;高中剂量组和阳性对照组肿瘤细胞出现了不同程度退变,可见细胞核碎片和凋亡细胞,中剂量组肿瘤细胞明显退变。与对照组和低剂量组比较,其他3组的促凋亡蛋白(Bax、Caspase-9、Caspase-7)相对表达量升高,抗凋亡蛋白Bcl-2相对表达量降低(均P<0.05),中剂量组各促凋亡蛋白相对表达量最高、抗凋亡蛋白相对表达量最低(均P<0.05)。其他3组细胞的凋亡率高于细胞空白组和200μmol/L培元抗癌汤组,300μmol/L培元抗癌汤组细胞凋亡率高于400μmol/L培元抗癌汤组和3μmol/L乌司他丁组(均P<0.05)。结论培元抗癌汤能有效抑制乳腺癌MDA-MB-231细胞裸鼠移植肿瘤生长,并调控凋亡相关蛋白的表达,从而诱导肿瘤细胞发生凋亡。

Objective To investigate the effect of Peiyuan Anticancer Decoction on the growth and apoptosis of human breast cancer MDA-MB-231 cell xenograft in nude mice.Methods Thirty BALB/c-nu female nude mice were selected,and human breast cancer xenograft model was established by hypodermic injection of MDA-MB-231 cells.The tumor-bearing mice up to the standard were randomized to control group,positive control group,low-dose group,medium-dose group or high-dose group,with 6 mice in each group.The low-,medium-and high-dose groups were gavaged with Peiyuan Anticancer Decoction with concentrations of 200 mg/kg,300 mg/kg and 400 mg/kg,respectively,the positive control group was hypodermically injected with ulinastatin(3 mg/kg),and the control group did not receive any intervention.The general condition of the tumor-bearing mice was observed during medication in all groups.After 21 days of continuous medication,the tumor-bearing mice in all groups were killed,then the tumor inhibitory rate was calculated after removal and weighing of breast tumor,the histocytomorphology of the tumor was observed using hematoxylin-eosin(HE)staining;besides,expression levels of B-cell lymphoma-2(Bcl-2),Bcl-2 associated X protein(Bax),cysteinyl aspartate specific proteinase(Caspase)-9 and Caspase-7 proteins in tumor tissue were determined.Human breast cancer MDA-MB-231 cells were collected,and the cell-blank group,the 3μmol/L ulinastatin group,and the 200μmol/L Peiyuan Anticancer Decoction group,the 300μmol/L Peiyuan Anticancer Decoction group and the 400μmol/L Peiyuan Anticancer Decoction group were established;except the cell-blank group the other groups were added with the drugs of corresponding concentrations,and the cells apoptosis rate was detected after a 48-hour intervention.Results Weigh loss,inactivity and unresponsiveness were observed in the tumor-bearing mice of the control group;and compared with the control group,mice in all medication groups gained more weight and were more responsive.The tumor inhibitory rate increased in the order of the control group,the low-dose group,the high-dose group,the positive control group,and the medium-dose group(all P<0.05).HE staining results showed that notable heterocysts with enlarged nucleus and obvious nuclear atypia were found in the control group and the low-dose group;cell degeneration in various degrees with nuclear fragments and apoptotic cells was observed in the medium-and high-dose groups and the positive control group,and the tumor cells in the medium-dose group exhibited obvious degeneration.Compared with the control group and the low-dose group,the remaining three groups obtained increased relative expression of pro-apoptotic proteins(Bax,Caspase-9 and Caspase-7)and decreased relative expression of anti-apoptotic protein Bcl-2(all P<0.05),and the highest relative expression of all pro-apoptotic proteins and the lowest relative expression of anti-apoptotic protein were found in the medium-dose group(all P<0.05).The other three groups yielded a higher cell apoptosis rate as compared with the cell-blank group and the 200μmol/L Peiyuan Anticancer Decoction group,and the rate was higher in the 300μmol/L Peiyuan Anticancer Decoction group than in the 400μmol/L Peiyuan nticancer Decoction group or the 3μmol/L ulinastatin group(all P<0.05).Conclusion Peiyuan Anticancer Decoction can effectively inhibit the growth of human breast cancer MDA-MB-231 cell xenograft in nude mice,and regulate the apoptosis-related proteins expression,thus inducing tumor cell apoptosis.