黄芩素联合吉西他滨诱导人胰腺癌细胞凋亡的研究

发布时间：2018-04-05 02:28

本文选题：胰腺癌　切入点：黄芩素　出处：《南京中医药大学》2017年博士论文

【摘要】：胰腺癌是恶性程度高、死亡率高的消化系统肿瘤,由于胰腺癌的诊断困难,发病早期症状不明显,目前对胰腺癌的治疗仍不理想,生存率不足5%。目前吉西他滨是胰腺癌辅助化疗的一线药物,但对胰腺癌患者生存率的提高仍然不足。近年来,多个临床试验将吉西他滨和其他药物联合治疗胰腺癌,但是效果仍不理想。因此,寻找更有效的治疗胰腺癌的药物,或者能联合吉西他滨达到更好治疗效果的药物十分重要。中药黄芩的主要有效成分是黄芩素,黄芩素也是黄芩主要发挥药理作用的成分。研究发现,黄芩素具有多种药理学作用,特别对多种肿瘤具有显著的抑制效果。小剂量短时间经过黄芩素的处理,肿瘤细胞能得到有效抑制,而当大剂量长时间黄芩素处理后,能诱导肿瘤细胞凋亡。因此,我们考虑通过小剂量的黄芩素联合吉西他滨是否能有效的抑制胰腺癌细胞的生长,成为潜在的治疗手段。研究方法(1)体外研究黄芩素联合吉西他滨对胰腺癌细胞的抑制作用及机制1、MTT检测黄芩素、吉西他滨和两种药物联用对胰腺癌细胞系CFPAC-1增殖的抑制作用;2、Hoechst33258染色,荧光显微镜下观察黄芩素及黄芩素联合吉西他滨对胰腺癌细胞的影响;3、Annexin-V APC/7-AAD双染法检测黄芩素、吉西他滨和两种药物联用对胰腺癌细胞凋亡的影响;4、黄芩素、吉西他滨和两种药物联用在胰腺癌细胞中对凋亡相关蛋白表达的影响;(2)药物治疗胰腺癌裸鼠移植模型将人胰腺癌细胞系CFPAC-1注射到裸鼠背部建立裸鼠移植瘤模型,待皮下肿瘤生长到2-4 mm开始给药,分别给予黄芩素、吉西他滨和两种药物联用,记录裸鼠体重及皮下移植瘤体积,共观察28天。然后处死裸鼠,取肿瘤进行称重和免疫组化染色。实验结果1、黄芩素或吉西他滨均能显著抑制人胰腺癌细胞系CFPAC-1细胞增殖,并且具有药物浓度依耐性,并且两种药物联用对细胞增殖抑制效果更为显著;2、黄芩素与吉西他滨联用能诱导人胰腺癌细胞系CFPAC-1细胞凋亡;3、黄芩素与吉西他滨联用诱导胰腺癌细胞凋亡是通过调控凋亡蛋白表达改变起作用的;4、黄芩素与吉西他滨联用能显著抑制人胰腺癌细胞裸鼠移植瘤的生长,并且作用效果明显优于单药的作用。结论黄芩素可以抑制胰腺癌细胞的增殖,诱导其分化。体内外实验研究表明,联合黄芩素和吉西他滨可以明显抑制人胰腺癌细胞的增殖,效果优于单药处理。黄芩素和吉西他滨联用将是成为胰腺癌治疗的新手段。
[Abstract]:Pancreatic cancer is a kind of digestive system tumor with high malignancy and high mortality. Because of the difficulty in diagnosis of pancreatic cancer, the early symptoms of pancreatic cancer are not obvious, so the treatment of pancreatic cancer is still not ideal, and the survival rate is less than 5%.Gemcitabine is the first-line adjuvant chemotherapy for pancreatic cancer, but the improvement of survival rate is still insufficient.In recent years, several clinical trials have combined gemcitabine with other drugs to treat pancreatic cancer, but the results are still unsatisfactory.Therefore, it is important to find more effective drugs for pancreatic cancer, or combination of gemcitabine for better treatment.The main active component of Scutellaria baicalensis is baicalin, and baicalin is the main pharmacological component of Scutellaria baicalensis.It was found that baicalin has many pharmacological effects, especially on many tumors.After the treatment of baicalin for a short time, the tumor cells could be effectively inhibited, but when the large dose of baicalin was treated for a long time, the apoptosis of tumor cells could be induced.Therefore, we consider whether small doses of baicalin combined with gemcitabine can effectively inhibit the growth of pancreatic cancer cells as a potential treatment.Methods 1) to study the inhibitory effect of baicalin combined with gemcitabine on pancreatic cancer cells in vitro and its mechanism. The inhibitory effects of baicalin, gemcitabine and two drugs on the proliferation of pancreatic cancer cell line CFPAC-1 were detected by MTT assay.The effects of baicalin and combination of baicalin and gemcitabine on pancreatic cancer cells were observed under fluorescence microscope. The effects of baicalin, gemcitabine and two drugs on apoptosis of pancreatic cancer cells were detected by Annexin-V APC/7-AAD double staining.Effects of gemcitabine and two drugs on the expression of apoptosis-related protein in pancreatic cancer cellsBaicalin, gemcitabine and two drugs were used to record the body weight of nude mice and the volume of subcutaneous transplanted tumor, and the tumor volume was observed for 28 days after the subcutaneous tumor grew to 2-4 mm and was given baicalin, gemcitabine and two kinds of drugs respectively.Then the nude mice were killed and the tumor was weighed and stained by immunohistochemistry.Results 1. Baicalin or gemcitabine could significantly inhibit the proliferation of human pancreatic cancer cell line CFPAC-1, and had concentration-dependent drug tolerance.The combination of baicalin and gemcitabine can induce apoptosis of human pancreatic cancer cell line CFPAC-1, baicalin and gemcitabine can induce apoptosis of pancreatic cancer cell line through the combination of baicalin and gemcitabine.Regulating the expression of apoptotic protein and regulating the expression of apoptotic protein, baicalin combined with gemcitabine could significantly inhibit the growth of human pancreatic cancer cell xenografts in nude mice.And the effect was obviously better than that of single drug.Conclusion baicalin can inhibit the proliferation and induce the differentiation of pancreatic cancer cells.In vitro and in vivo experiments showed that the combination of baicalin and gemcitabine could significantly inhibit the proliferation of human pancreatic cancer cells.Baicalin combined with gemcitabine will be a new treatment for pancreatic cancer.
【学位授予单位】：南京中医药大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R735.9

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