摘要
目的应用血管内皮抑素(ES)干预肺腺癌A549细胞荷瘤鼠,研究其对A549荷瘤鼠的调节作用及其作用机制。方法取40只雌性BALB/c裸鼠4周龄,体重15~20 g。通过肺腺癌细胞株A549建立荷瘤鼠模型,利用重组ES作用于小鼠移植性肿瘤模型,将成瘤后的30只裸鼠按随机数字表法分为PCMV-Script-ES组、PCMV-Script组和对照组,每组各10只。每组均通过瘤体内注射给药。PCMV-Script-ES组注射含有endostatin的重组质粒0.2 mL,PCMV-Script组注射无endostatin的空载体0.2 mL,对照组注射等体积的0.9%氯化钠溶液,连续注射14 d。测量并记录3组裸鼠体重并计算抑瘤率,采用TUNEL法检测瘤体组织中细胞凋亡指数。分别使用不同浓度(0、10、20、40μg/mL)PCMV-Script-ES作用于A549细胞,于处理12、24和48 h后,利用MTT法检测其对A549细胞增殖的抑制作用;选择抑制率最高的时间点,利用链亲和素过氧化酶法(SP法)检测A549细胞中Bcl-2和Bax蛋白的表达情况。结果(1)与治疗前相比,治疗后14 d,3组小鼠肿瘤体积均显著增加,PCMV-Script-ES组小鼠肿瘤体积均小于PCMV-Script组和对照组,差异有统计学意义(P<0.05)。与治疗前相比,治疗后14 d,PCMV-Script-ES组小鼠体重增加,PCMV-Script组和对照组小鼠体重均显著下降(P<0.05)。治疗后14 d,PCMV-Script-ES组小鼠体重显著小于PCMV-Script组和对照组,差异均有统计学意义(P<0.05)。(2)PCMV-Script-ES组抑瘤率显著高于PCMV-Script组和对照组,差异均有统计学意义(P<0.05)。(3)PCMV-Script-ES组凋亡细胞数量较对照组和PCMV-Script组显著增多,差异有统计学意义(P<0.05)。(4)随着PCMV-Script-ES的浓度增加,其抑制率逐渐增加;在药物浓度20μg/mL时,作用48 h后,抑制作用明显增强;在药物浓度为40μg/mL时,抑制率随着作用时间的增加而增强。(5)PCMV-Script-ES处理A549细胞时,随着作用剂量增加,肺腺癌细胞中Bcl-2蛋白表达阳性率显著降低,而Bax蛋白表达阳性率则显著增加,差异均有统计学意义(P<0.05)。结论ES可通过抑制A549肺腺癌细胞增殖和促进细胞凋亡从而对肺腺癌荷瘤鼠发挥显著抑瘤作用,且其作用效果随浓度和作用时间增加而增强。
Objective To apply endostatin to lung cancer A549 cell tumor-bearing mice,and to study its regulatory effect on A549 tumor-bearing mice and its mechanism.Methods Forty female BALB/c nude mice were taken at 4 weeks of age,weighing 15-20 g.The lung adenocarcinoma cell line A549 was used to establish a tumor-bearing mouse model,and recombinant ES was used to act on the transplanted tumor model in mice.The nude mice after tumor formation were randomly divided into PCMV-Script-ES group,PCMV-Script group and control group,with 10 mice in each group.Each group was administered by intratumoral injection.The PCMV-Script-ES group was injected with 0.2 mL of the recombinant plasmid containing endostatin,the PCMV-Script group was injected with 0.2 mL of the empty vector without endostatin,and the control group was injected with an equal volume of normal saline for 14 consecutive days.The body weight of the three groups of nude mice was measured and recorded,and the tumor inhibition rate was calculated.The TUNEL method was used to detect the apoptosis index in the tumor tissue.Different concentrations(0,10,20,40μg/mL)of PCMV-Script-ES were used to act on A549 cells.After 12,24 and 48 h treatment,the MTT method was used to detect its inhibitory effect on the proliferation of A549 cells;The time point with the highest inhibition rate was selected,and the expression of Bcl-2 and Bax protein in A549 cells was detected by the streptavidin peroxidase method(SP method).Results(1)Compared with before treatment,14 days after treatment,the tumor volume of the three groups of mice increased significantly,and the tumor volume of the mice in the PCMV-Script-ES group was less than that in the PCMV-Script group and the control group,the difference was statistically significant(P<0.05).Compared with before treatment,mice in PCMV-Script-ES group gained weight after treatment,and mice in PCMV-Script group and control group decreased significantly(P<0.05).After treatment,the weight of the mice in the PCMV-Script-ES group was significantly less than that of the PCMV-Script group and the control group,the difference was statistically significant(P<0.05).(2)The tumor inhibition rate of PCMV-Script-ES group was higher than that of PCMV-Script group and control group,the difference was statistically significant(P<0.05).(3)The number of apoptotic cells in the PCMV-Script-ES group was significantly higher than that of the control groupand PCMV-Script group,the difference was statistically significant(P<0.05).(4)As the concentration of PCMV-Script-ES increases,its inhibition rate gradually increases;when the drug concentration is 20μg/mL,and after 48 hours of action,the inhibitory effect was significantly enhanced;when the drug concentration was 40μg/mL,the inhibitory rate increased with the increase of the action time.(5)When PCMV-Script-ES treats A549 cells,as the dose increases,the positive rate of Bcl-2 protein expression in lung adenocarcinoma cells was significantly reduced,while the positive rate of Bax protein expression was significantly increased,the differences were statistically significant(P<0.05).Conclusion ES can inhibit the proliferation of A549 lung adenocarcinoma cells and promote cell apoptosis,thereby exerting a significant anti-tumor effect on lung adenocarcinoma tumor-bearing mice,and its effect is enhanced with the increase of concentration and time of action.
作者
肖珊珊
张卓尔
杨巧红
XIAO Shan-shan;ZHANG Zhuo-er;YANG Qiao-hong(Department of Pathology,First Affiliated Hospital of Guangzhou University of Traditional Chinese Medicine,Guangzhou Guangdong 510405,China;Department of Clinical Medicine,Southern Medical University,Guangzhou Guangdong 518056,China;Department of Pathology,School of Basic Medicine,Guangzhou University of Chinese Medicine,Guangzhou Guangdong 510006,China.)
出处
《临床和实验医学杂志》
2021年第18期1912-1916,共5页
Journal of Clinical and Experimental Medicine
基金
广东省中医药局科研项目(编号:20211122)。
