摘要
目的:探讨褪黑素(melatonin,MLT)对小鼠肿瘤细胞自噬信号通路的影响,从而阐述褪黑素发挥抗肿瘤效果的机制。方法:构建H22肝癌模型小鼠30只,每组10只,分别为生理盐水组、10 mg/kg褪黑素组、20 mg/kg褪黑素组。将小鼠肿瘤组织制备成切片后于光学显微镜和电镜下观察切片。利用荧光免疫组化法检测每个视野中的阳性细胞数平均百分比,以该切片阳性细胞百分比为基础进行计分。通过蛋白印迹试验对蛋白进行定性及定量测定,最后利用荧光显微镜下观察细胞内的绿色荧光,记录图像,分析计数LC3阳性细胞与总细胞数的比例,反映细胞发生自噬的程度。结果:MLT处理的动物其自噬形态较生理盐水组明显增多。随机成像后MLT处理组自噬液泡总数明显高于生理盐水组(P <0. 05)。经MLT处理的H22荷瘤小鼠的肿瘤组织中Beclin-1、LC3-Ⅱ表达水平明显高于生理盐水组(P <0. 05)。MLT处理的H22荷瘤小鼠肿瘤组织中,LC3亮点有所增强。MLT处理的H22荷瘤小鼠肿瘤组织中Akt和mT OR磷酸化水平较生理盐水组明显降低(P <0. 05)。结论:褪黑素在H22荷瘤小鼠中诱导自噬,褪黑素能够诱导自噬过程标志蛋白Beclin-1和LC3的表达,并且能够抑制H22荷瘤小鼠中的Akt/mT OR信号通路。
Objective:To discuss the effect of melatonin(MLT)on autophagy signal pathway in mice tumor cells and the mechanism of melatonin to play an antitumor effect.Methods:30 models of H22 tumor BALB/c mice were established with 10 rats in each group,including the normal saline group,the 10 mg/kg and the 20 mg/kg melatonin group.The tumor tissue of mice was sliced and sectioned by optical microscope and electron microscope.The average percentage of the number of positive cells in each field of vision was detected by the method of immunofluorescence,which was based on the percentage of the positive cells of the slice.The protein was qualitatively and quantitatively determined by Western blotting.At last,the green fluorescence in cells was observed under fluorescence microscope,and the images were recorded.The ratio of LC3 positive cells to total cells was counted and analyzed,which reflects the degree of autophagy.Results:The autophagic morphology of the MLT treated animals was significantly higher than that in the normal saline group.The total number of autophagic vacuoles in the MLT treatment group was significantly higher than that of the normal saline group after random imaging(P<0.05).In the tumor tissue of H22 bearing mice treated with melatonin,expression levels of Beclin-1 and LC3-Ⅱwere significantly higher than normal saline group(P<0.05)and the bright spot of LC3 was enhanced.The level of phosphorylation of Akt and mTOR in the tumor tissues of melatonin treated H22 tumor mice was significantly lower than that in the normal saline group(P<0.05).Conclusion:Melatonin induces autophagy in H22 tumor mice and induces the expression of autophagy Beclin-1 and LC-3.MLT can also inhibit the Akt/mTOR signaling pathway in H22 tumor bearing mice.
作者
刘畅
张宗恺
贾佳
陈点点
阮新建
刘彦芳
张侠
Liu Chang;Zhang Zongkai;Jia Jia;Chen Diandian;Ruan Xinjian;Liu Yanfang;Zhang Xia(Department of Oncology,the Seventh Medical Center of PLA General Hospital,Beijing 100700,China)
出处
《现代肿瘤医学》
CAS
2019年第5期733-738,共6页
Journal of Modern Oncology
基金
国家自然科学基金(编号:81073087)
