藥學(xué)論文復(fù)方腸泰抑制大腸癌的作用及機理初探

1、復(fù)方腸泰抑制大腸癌的作用及機理初探 【畢業(yè)論文標(biāo)題】 復(fù)方腸泰抑制大腸癌的作用及機理初探【英文標(biāo)題】 primary study of fu-fang-chang-tai on the inhibition to colon cancer【中文摘要】 大腸癌(colorectal cancer,crc)是世界范圍內(nèi)最常見的惡性腫瘤之一。本文通過整體動物和體外細(xì)胞生物學(xué)實驗,從整體動物、組織器官、細(xì)胞和分子生物學(xué)的不同水平,觀察復(fù)方腸泰(ffct)對人結(jié)腸癌細(xì)胞sw480裸鼠種植瘤的抑制作用;對人結(jié)腸癌細(xì)胞sw480細(xì)胞增殖的抑制作用,對其細(xì)胞周期、凋亡以及凋亡作用途徑進(jìn)行初步探索;并探索了對h

2、22荷瘤小鼠免疫功能的影響。本文的實驗方法如下:通過ffct對人結(jié)腸癌細(xì)胞sw480體外生長的影響以及裸鼠接種sw480瘤株后的體內(nèi)抑瘤作用,進(jìn)行體內(nèi)外的抗腫瘤藥效作用研究,并對ffct的臨床處方進(jìn)行篩選;采用噻唑藍(lán)(mtt)比色法測定不同濃度ffct在不同作用時間內(nèi)對體外培養(yǎng)的sw480細(xì)胞增殖的影響,同時應(yīng)用流式細(xì)胞術(shù)檢測其對細(xì)胞增殖周期及凋亡的影響;檢測經(jīng)不同濃度的ffct作用后sw480結(jié)腸癌細(xì)胞內(nèi)caspase-3的活性,比較經(jīng)ffct作用以及同時經(jīng)ffct和caspase-3抑制劑ac-devd-cho作用后,體外培養(yǎng)的人結(jié)腸癌sw480細(xì)胞凋亡的影響;檢測ffct對h22荷瘤小鼠

3、瘤重、免疫器官濕重以及溶血素抗體生成的影響。實驗結(jié)果表明:“五號處方”的ffct能夠顯著抑制人結(jié)腸癌細(xì)胞sw480的增殖;抑制sw480裸鼠種植瘤的瘤重,且裸鼠的體重均未見明顯減輕,未見化學(xué)藥常見的毒性作用。ffct“處方五”的體內(nèi)外抑瘤作用優(yōu)于其它4個處方。fftc可在g0/g1期以及g2/m期阻滯人結(jié)腸癌細(xì)胞sw480的增殖,使s期的活細(xì)胞比率降低;在一定范圍內(nèi),ffct的濃度越高、作用時間越長,對腫瘤細(xì)胞生長的抑制作用越強,凋亡率也越高。經(jīng)ffct培養(yǎng)液分別作用不同時間后的caspase-3酶活性,與對照組相比均有顯著性變化,呈現(xiàn)出先增高活性,隨后逐漸降低的趨勢;加入caspase-3抑

4、制劑ac-devd-cho的ffct各濃度、時間組結(jié)腸癌細(xì)胞sw480的早期凋亡率均有所增加,細(xì)胞的壞死率與晚期凋亡率的總和也有所增加,與空白組比較且呈一定的量-效、時-效關(guān)系,但是與沒有加入ac-devd-cho時相比,各濃度、時間組的sw480早期凋亡率均有所下降,細(xì)胞的壞死率與晚期凋亡率的總和沒有顯著性變化。ffct與抗腫瘤藥物環(huán)磷酰胺(cy)有一定的協(xié)同作用,抑制瘤重增長;能明顯改善cy化療后h22荷瘤小鼠的脾臟、胸腺等臟器的萎縮,提高胸腺和脾臟的臟器系數(shù),使因化療引起的機體免疫功能低下有一定的恢復(fù),并且能夠升高h(yuǎn)22荷瘤小鼠血清溶血素水平,使cy所致的血清溶血素含量下降有明顯恢復(fù),從

5、而可提高h(yuǎn)22荷瘤小鼠的體液免疫能力。本文得出如下的結(jié)論:ffct可抑制大腸癌荷瘤裸鼠的瘤重,并可恢復(fù)和增強荷瘤小鼠的免疫力;ffct可抑制體外培養(yǎng)的人結(jié)腸癌細(xì)胞sw480的增殖,其作用機理可能與阻止g0/g1期以及g2/m周期的細(xì)胞生長、促進(jìn)sw480細(xì)胞凋亡相關(guān)聯(lián)。實驗結(jié)果還提示caspase-3途徑可能是ffct促進(jìn)人結(jié)腸癌細(xì)胞sw480凋亡的重要途徑之一,但可能還存在其它作用途徑,有待今后作進(jìn)一步的深入研究?！居⑽恼?colorectal cancer is one of the most common cancers in both men and women.to observ

6、e the effects of fu-fang-chang-tai(ffct) on the growth of sw480 human colon cancer xenografted in athymic mice,and on cell growth,cell generation cycle and apoptosis of human colon carcinoma cell sw480,and on the immune function of mice with carcinoma through in vivo animal experiments and in vitro

7、cell experiments.the mechanisms were explored at levels from whole animal and histological to cytobiological and molecule-biological.the methods were used as follows:the best one was sieved from several prescriptions of ffct through the experimental effects on the growth of human colon cancer cells

8、sw480 and sw480 xenografted in athymic mice.the effects of different saturation of ffct in different action time on cell multiplication of sw480 in vitro culture were determined with methyl thiazolyl tetrazolium(mtt) chromatometry.at the same time the changes of cell generation cycle and apoptosis r

9、ate were detected with flow cytometry.caspase-3 enzyme activities in ffct-treated sw480 cells were determined. the comparison was made between apoptosis rates in ffct-treated cells and ac-devd-cho with ffct-treated cells.the effects of ffct acting on the tumors, immune organ weight and the formation

10、 of hemolysin antibody of the mice with hepatocarcinoma h22 were studied.the results came out as follows:the growth of human colon carcinoma cell sw480 and sw480 xenografted in athymic mice could be significantly inhibited by 5th ffct, while the body weights of the mice treated by 5th ffct were not

11、light which compared with the control group.the effectiveness of 5th ffct was better than other prescriptions. the generation of colon carcinoma cell sw480 in the stage of g0/g1 and g2/m could be interrupted by ffct.to some extent,the more saturation and time of ffct acted,the more of inhibitory act

12、ion on tumor cell growth and apoptosis rate showed.caspase-3 enzyme activity in ffct-treated sw480 cells increased at first and then lysised with time went compared with the control group.both apoptosis rates and death rates in ac-devd-cho with ffct-treated cells increased significantly compared wit

13、h the control group.apoptosis rates in ac-devd-cho-with ffct-treated groups were lower than in corresponding ffct-treated groups while death rates were similar with them. there were some synergistic actions in cyclophosphamide united with ffct to inhibit tumor growth.it could improve the situation o

14、f organ atrophy and low formation of hemolysin antibody of the tumor-beating mice caused by chemotherapy in that organism immunity.the humoral immunity could be improved.the conclusion indicated that:colon carcinoma could be cured by ffct through the growth of tumor weight inhibition and the immune

15、function improvement.the growth cycle of tumer cells could be interrupted and cell apoptosis could be induced to inhibit the growth of human colon carcinoma cell sw480 on the effect of ffct.caspase-3 way was an important but not the only way in the process of sw480 apoptosis.【中文關(guān)鍵詞】 復(fù)方腸泰; 裸鼠種植瘤; 人結(jié)腸

16、癌細(xì)胞sw480; 細(xì)胞周期; 細(xì)胞凋亡; caspase-3; 荷瘤小鼠; 免疫 【英文關(guān)鍵詞】 fu-fang-chang-tai; xenografted human colon carcinoma cell; sw480; apoptosis ; caspase-3 ; tumor-bearing mice ; immune function 【論文目錄】摘要 5-7 abstract 7-8 第一章 緒論 14-32 1.1 大腸癌的機理研究 14-19 1.1.1 大腸癌的現(xiàn)代醫(yī)學(xué)發(fā)病機理 14-16 1.1.2 中醫(yī)對大腸癌病因病機的認(rèn)識 16-19 1.2 中西醫(yī)藥對大腸癌治療

17、的研究進(jìn)展 19-23 1.2.1 現(xiàn)代醫(yī)學(xué)研究進(jìn)展 19-21 1.2.2 中醫(yī)藥研究進(jìn)展 21-23 1.3 存在的問題與展望 23-24 1.4 本實驗的相關(guān)工作 24-28 參考文獻(xiàn) 28-32 第二章 ffct處方篩選實驗(第一批) 32-41 2.1 主要實驗材料 32-34 2.1.1 實驗細(xì)胞與動物 32 2.1.2 藥物與試劑 32-33 2.1.3 主要儀器 33-34 2.2 實驗方法 34-36 2.2.1 不同處方ffct對sw480細(xì)胞生長的影響(mtt法) 34-35 2.2.2 ffct各處方對人結(jié)腸癌細(xì)胞sw480裸鼠種植瘤的影響 35 2.2.3 數(shù)據(jù)分析

18、35-36 2.3 實驗結(jié)果 36-39 2.3.1 不同處方ffct稀釋液對sw480細(xì)胞生長的影響 36-38 2.3.2 ffct各處方對人結(jié)腸癌細(xì)胞sw480裸鼠種植瘤的影響 38-39 2.4 小結(jié)與討論 39-40 參考文獻(xiàn) 40-41 第三章 ffct處方(第二批)篩選實驗 41-48 3.1 主要實驗材料 41-42 3.1.1 實驗細(xì)胞與動物 41 3.1.2 藥物與試劑 41-42 3.1.3 主要儀器 42 3.2 實驗方法 42-43 3.2.1 不同處方ffct稀釋液對sw480細(xì)胞生長的影響(mtt法) 42 3.2.2 ffct各處方對人結(jié)腸癌細(xì)胞sw480裸鼠種

19、植瘤的影響 42-43 3.2.3 數(shù)據(jù)分析 43 3.3 實驗結(jié)果 43-46 3.3.1 不同處方ffct稀釋液對sw480細(xì)胞生長的影響 43-45 3.3.2 ffct各處方對人結(jié)腸癌細(xì)胞sw480裸鼠種植瘤的影響 45-46 3.4 小結(jié)與討論 46-48 第四章 ffct抑制人結(jié)腸癌sw480細(xì)胞增殖的實驗研究 48-64 4.1 主要實驗材料 48-49 4.1.1 實驗細(xì)胞 48 4.1.2 藥物與試劑 48-49 4.1.3 主要儀器 49 4.2 實驗方法 49-53 4.2.1 ffct及陽性藥的加藥前處理 49-50 4.2.2 ffct對結(jié)腸癌細(xì)胞的細(xì)胞增殖抑制作用(

20、mtt法) 50-51 4.2.3 ffct對結(jié)腸癌細(xì)胞周期的影響 51 4.2.4 ffct誘導(dǎo)結(jié)腸癌細(xì)胞凋亡的研究 51-52 4.2.5 數(shù)據(jù)分析 52-53 4.3 實驗結(jié)果 53-61 4.3.1 ffct對結(jié)腸癌細(xì)胞的細(xì)胞增殖抑制作用:采用mtt比值法 53-56 4.3.2 ffct對結(jié)腸癌細(xì)胞周期的影響 56-59 4.3.3 ffct誘導(dǎo)結(jié)腸癌細(xì)胞凋亡的研究 59-61 4.4 小結(jié)與討論 61-63 參考文獻(xiàn) 63-64 第五章 ffct通過caspase-3途徑促進(jìn)人結(jié)腸癌細(xì)胞sw480凋亡機制的探討 64-76 5.1 主要實驗材料 64-65 5.1.1 實驗細(xì)胞 64 5.1.2 藥物與試劑 64-65 5.1.3 主要儀器 65 5.2 實驗方法 65-69 5.2.1 ffct及陽性藥的加藥前處理 65 5.2.2 annexin v/pi法檢測經(jīng)ffct作用后sw480結(jié)腸癌細(xì)胞凋亡 65-66 5.2.3 檢測經(jīng)ffct作用后sw480結(jié)腸癌細(xì)胞內(nèi)caspase-3的活性 66-67 5.2.4 檢測經(jīng)ffct和caspase-3抑制劑ac-devd-cho共同作用后sw480結(jié)腸癌細(xì)胞內(nèi)caspase-3的活性 67-68 5.2.5 annexin v/pi法檢測經(jīng)ffct和caspase-3抑制劑ac-devd-c