摘要
目的探讨毛蕊花糖苷(ACT)联合人工合成的PI3K/AKT信号通路抑制剂-2-吗啉代-8-苯基色酮(Ly294002)对人肝癌细胞系HepG2裸鼠皮下移植瘤生长的抑制作用。方法建立HepG2裸鼠移植瘤,分组设置空白组、模型组、Ly294002(20mg/kg)组、ACT(80mg/kg)组、ACT+Ly294002(80mg/kg+20mg/kg)组,共5组,每组6只。给药期间,对肿瘤体积进行监测;给药结束后,称取肿瘤质量,计算肿瘤抑制率和生长率;采用HE染色、透射电镜和免疫荧光法分别对肿瘤组织的病理形态、超微结构和Ki-67及PCNA表达水平进行观察或统计分析,酶联免疫吸附试验(ELISA)检测裸鼠血清中白介素-6(IL-6)、超敏C反应蛋白(hs-CRP)、甲胎蛋白(AFP)和肿瘤坏死因子(TNF-α)活性水平。结果模型组、ACT组、Ly294002组和ACT+Ly294002组肿瘤质量分别为(0.90±0.24)、(0.63±0.18)、(0.66±0.05)和(0.45±0.04)g,F=8.533,P<0.01。HE染色结果显示,模型组有大量存活肿瘤细胞,Ly294002组和ACT组存活细胞相对减少,而ACT+Ly294002组肿瘤存活区域明显减少,坏死区面积增大。免疫荧光结果显示,模型组、ACT组、Ly294002组、ACT+Ly294002组Ki-67表达水平分别为(100.00±0.00)%、(179.92±3.96)%、(58.91±1.04)%和(1.21±0.04)%,F=4039.224,P<0.01。模型组、ACT组、Ly294002组、ACT+Ly294002组PCNA表达水平分别为(100.00±0.00)%、(31.39±0.87)%、(6.56±1.01)%和(1.81±0.42)%,F=12591.668,P<0.01。超微结构观察结果显示,模型组细胞核异型,核膜局部凹陷,形成核切迹、多核,肝肿瘤细胞细胞器不丰富;Ly294002组肿瘤细胞的细胞核受到损伤,异染色质团集;ACT组核膜溶解,絮状样改变,线粒体脊溶解;ACT+Ly294002组肿瘤细胞损伤最为严重,出现空泡化,细胞质和细胞器广泛溶解,异染色质边际化。与空白组相比,模型组荷瘤裸鼠血清中IL-6、hs-CRP、AFP和TNF-α活性水平均升高,均P<0.01;与模型组相比,ACT组、Ly294002组和ACT+Ly294002组IL-6、hs-CRP、AFP和TNF-α活性水平均有不同程度下降,其中Ly294002组IL-6和TNF-α的下降具有统计学意义,P<0.05;ACT+Ly294002组IL-6、hs-CRP和TNF-α的下降具有统计学意义,P<0.01;与Ly294002组相比,ACT+Ly294002组hs-CRP活性下降,P<0.05。结论ACT和Ly294002对HepG2裸鼠移植瘤的生长均有抑制作用,且联合用药具有协同增效的作用。
Objective To investigate the inhibitory effect of acteoside(ACT)combined with 2-(4-Morpholinyl)-8-phenyl-4H-1-benzopyran-4-one(Ly294002),which is a synthetic inhibitor of PI3K/AKT signaling pathway,on the growth of carcinoma xenograft(human hepatoma cells HepG2)in nude mice.Methods HepG2 nude mice xenograft was established and set groups as control group,model group,Ly294002(20mg/kg)group,ACT(80mg/kg)group,ACT+Ly294002(80mg/kg+20mg/kg)group,with 6 in each group.During administration,the tumor volume was monitored.After administration,the wet weight of the tumor was measured,and the tumor inhibition rate and growth rate were calculated.The pathological morphology,ultrastructure and Ki-67 and PCNA expression levels of tumor tissues were observed or statistically analyzed by HE staining,transmission electron microscopy and immunofluorescence,respectively.The activity of interleukin-6(IL-6),hypersensitive C-reactive protein(hs-CRP),alpha-fetoprotein(AFP)and activity of tumor necrosis factor(TNF-α)in serum of nude mice were detected by enzyme-linked immunosorbent assay(ELISA).Results The wet weight of tumor in model group,ACT group,Ly294002 group,and ACT+Ly294002 group were(0.90±0.24)g,(0.63±0.18)g,(0.66±0.05)g and(0.45±0.04)g,F=8.533,P<0.01.The HE staining results showed that there were a large number of living tumor cells in the model group,and the number of living cells in the Ly294002 group and the ACT group was relatively reduced,while the number of tumor survival areas in the ACT+Ly294002 group was significantly reduced,and the area of necrotic areas was increased.The immunofluorescence results showed that the expression levels of Ki-67 in model group,ACT group,Ly294002 group and ACT+Ly294002 group were(100.00±0.00)%,(179.92±3.96)%,(58.91±1.04)%and(1.21±0.04)%,respectively,F=4039.224,P<0.01.The expression levels of PCNA in model group,ACT group,Ly294002 group and ACT+Ly294002 group were(100.00±0.00)%,(31.39±0.87)%,(6.56±1.01)%and(1.81±0.42)%respectively,F=12591.668,P<0.01.The ultrastructural observation showed that in the model group,the nuclei were heterotypic,the nuclear membrane was locally depressed,and the nuclear notch and multinuclear were formed,and the organelles of liver tumor cells were not abundant;In Ly294002 group,the nuclei of tumor cells were damaged and heterochromatin clumped;In ACT group,nuclear membrane was dissolved,flocculent changes were observed,and mitochondrial ridge was dissolved;In ACT+Ly294002 group,the tumor cells were most severely damaged,with vacuolation,widespread dissolution of cytoplasm and organelles,and marginal heterochromatin.Compared with the control group,the IL-6,hs-CRP,AFP and TNF-α activity levels in the serum of nude mice bearing tumor in the model group were all increased(P<0.01);Compared with the model group,the IL-6,hs-CRP,AFP and TNF-α activity levels in ACT group,Ly294002 group and ACT+Ly294002 group were all decreased to different degrees,including the decrease of IL-6 and TNF-α in Ly294002 group with statistically significant(P<0.05),and the decrease of IL-6,hs-CRP and TNF-α in ACT+Ly294002 group with significant difference(P<0.01).Compared with Ly294002 group,the activity of hs-CRP in ACT+Ly294002 group decreased(P<0.05).Conclusion ACT and Ly294002 can inhibit the growth of HepG2 nude mice xenograft,and the combination of ACT and Ly294002 has a synergistic effect.
作者
文丽梅
张嘉伟
陈樟
组皮艳木·马拜提
姑丽米热·艾力
阿依谢姆古丽·马木提
王泽坤
杨建华
胡君萍
WEN Limei;ZHANG Jiawei;CHEN Zhang;ZUPIYANMU Mabaiti;GULIMIRE Aili;AYIXIEMUGULI Mamuti;WANG Zekun;YANG Jianhua;HU Junping(Pharmaceutical Department,First Affiliated Hospital of Xinjiang Medical University,Urumqi 830011,China;College of Pharmacy,Xinjiang Medical University,Urumqi 830017,China)
出处
《中华肿瘤防治杂志》
CAS
北大核心
2023年第15期904-911,共8页
Chinese Journal of Cancer Prevention and Treatment
基金
新疆维吾尔自治区自然科学基金(2021D01C347)
新疆医科大学2021年16期大学生创新创业训练计划(202110760009)
