山椒子烯酮对急性T淋巴细胞白血病的抗肿瘤药效及安全性评价

Anti-tumor Efficacy and Safety of Zeylenone on Acute T Lymphoblastic Leukemia

目的:探究山椒子烯酮对急性T淋巴细胞白血病的抗肿瘤药效、机制及安全性。方法:体外培养Molt-4细胞,不同浓度山椒子烯酮(0.2,0.4,0.8,1.6,3.2μmol·L^-1)给药48 h,通过细胞增殖-毒性检测法(CCK-8)检测Molt-4细胞的活性。非肥胖糖尿病/重症联合免疫缺陷(NOD/SCID)小鼠随机分为正常组,模型组,长春新碱组(1 mg·kg^-1),山椒子烯酮低、中、高剂量组(12.5,25,50 mg·kg^-1)。除正常组外,小鼠预先照射60Co,皮下接种Molt-4细胞建立Molt-4细胞移植瘤小鼠模型。第13天,小鼠脱臼处死并解剖。计算抑瘤率、相对肿瘤增殖率、肝指数及脾指数;苏木素-伊红(HE)染色观察肝组织、脾组织的病理变化;蛋白免疫印迹法(Western blot)检测肿瘤组织中磷酸化信号转导与转录激活因子-3(p-STAT3),B细胞淋巴瘤-2(Bcl-2),Bcl-2相关X蛋白(Bax),半胱氨酸天冬氨酸蛋白酶-3(Caspase-3)蛋白的表达。结果:体外实验结果表明,与空白组比较,山椒子烯酮对Molt-4细胞有明显的抑制作用,半数抑制浓度(IC50)为1.49μmol·L^-1。体内实验结果表明,与模型组比较,山椒子烯酮中剂量组和高剂量组抑瘤效果显著,抑制率分别为50.24%和60.75%(P<0.01)。病理结果表明,与长春新碱组比较,山椒子烯酮对肝组织、脾组织损伤较小,安全性更好。蛋白印迹结果表明,与模型组比较,山椒子烯酮中剂量组和高剂量组可以显著下调p-STAT3,Caspase-3,Bcl-2蛋白的表达(P<0.05,P<0.01),上调Bax蛋白的表达(P<0.05,P<0.01)。结论:山椒子烯酮有明显抑制Molt-4细胞增殖,诱导Molt-4细胞凋亡的作用,其作用机制可能与下调p-STAT3,Caspase-3,Bcl-2蛋白的表达,上调Bax蛋白的表达有关。同时,山椒子烯酮对肝组织、脾组织的损伤较小,安全性更好。

Objective: To investigate the anti-tumor efficacy,mechanism and safety of zeylenone on acute T lymphocytic leukemia. Method: In vitro,Molt-4 cells were treated with various concentrations of zeylenone(0. 2,0. 4,0. 8,1. 6,3. 2 μmol·L^-1)for 48 h,and the cell viability was measured with cell counting kit-8(CCK-8) assay. nonobese diabetic-severce combined immunodeficient mice(NOD/SCID) mice were randomly divided into six groups:normal group,model group,vincristine group(1 mg·kg^-1),low-dose zeylenone group(12.5 mg·kg^-1), m edium-dose zeylenone group(25 m g·kg^-1), high-dose zeylenone group(50 m g·kg^-1). With the exception of normal group,mice were pre-irradiated with60 Co and inoculated subcutaneously with Molt-4 cells to establish the Molt-4 xenograft model. Then NOD/SCID mice were sacrificed after 13 days of administration. The tumor inhibition rates,relative tumor growth rates and organ indexes were calculated. Hematoxylin and eosin(HE)staining was used to observe the pathological changes of liver and spleen tissues in mice. The expressions of phosphorylation signal transducer and activator of transcription(p-STAT3),B-cell lymphoma-2(Bcl-2),Bcl-2-associated X protein(Bax)and cysteine aspartatespecific protease-3(Caspase-3)were detected in tumor tissues by Western blot. Result: In vitro,zeylenone had an obvious inhibitory effect on Molt-4 cells. IC50 values of zeylenone was 1. 49 μmol·L^-1. In vivo,compared with the model group,medium and high-dose zeylenone groups had significant tumor inhibition effects,with the inhibition rates of 50. 24% and 60. 75%,respectively(P<0. 01). Additionally,liver and spleen injuries were slight in the above mentioned two groups compared with the vincristine group,indicating that zeylenone was safe. Western blot analysis showed that medium and high-dose zeylenone groups showed significant declines in proteins p-STAT3,Caspase-3 and Bcl-2,and marked increases in pro-apoptotic protein Bax compared with the model group(P<0. 05,P<0. 01). Conclusion: zeylenone could obviously inhibit the proliferation and induce the apoptosis of Molt-4 cells;and its mechanism may be related to the down-regulation of p-STAT3,Caspase-3,Bcl-2 and the up-regulation of Bax expressions. In addition,zeylenone had less damage to liver and spleen,and was safer than vincristine.