本文選題：重樓 + 胃癌　； 參考：《南京中醫(yī)藥大學(xué)》2017年博士論文

【摘要】：[目的]研究重樓乙醇提取物(Parispolyphlla ethanol extract,PPE)對(duì)ICR小鼠的急性毒性劑量,探討重樓在腫瘤綜合治療中的應(yīng)用,并考察其體外的抗腫瘤作用及機(jī)制;探討胃癌患者的術(shù)前血清腫瘤標(biāo)記物水平和手術(shù)后脈管內(nèi)癌栓之間的關(guān)系,初步探討胃癌臨床應(yīng)用中藥聯(lián)合氟尿嘧啶類化療的療效及其對(duì)療效相關(guān)分子調(diào)節(jié)作用。[方法]1、用ICR小鼠進(jìn)行急性毒性實(shí)驗(yàn),以生存狀態(tài)、血液學(xué)指標(biāo)變化、腹腔臟器變化為觀察指標(biāo),并計(jì)算PPE的半數(shù)致死量(median leath death,LD50)劑量。2、用MTT法及細(xì)胞克隆形成方法檢測(cè)PPE,以及PPE聯(lián)合放療對(duì)人胃癌細(xì)胞MKN-45的生長(zhǎng)抑制;用PI染色法檢測(cè)PPE,以及PPE聯(lián)合放療對(duì)人胃癌細(xì)胞株MKN-45細(xì)胞周期的影響;Western-blot檢測(cè)PPE對(duì)凋亡相關(guān)蛋白水平的影響;運(yùn)用RT-PCR法檢測(cè)PPE對(duì)氟尿嘧啶類藥物敏感基因胸苷酸合成酶(thymidylatesynthase,TS)的mRNA水平的變化。3、通過對(duì)我院普通外科進(jìn)行胃癌根治性手術(shù)的1005例患者,檢測(cè)手術(shù)前一周內(nèi)的外周血腫瘤標(biāo)志物CEA、CA19-9、CA242、CA72-4、CA125的水平,并對(duì)術(shù)后標(biāo)本進(jìn)行包括脈管癌栓在內(nèi)的多項(xiàng)病理學(xué)檢查。入組患者基線水平數(shù)據(jù)按照各外周血腫瘤標(biāo)志物的狀況被分組,分析每個(gè)亞組標(biāo)志物水平對(duì)脈管癌栓狀態(tài)的影響。評(píng)價(jià)各亞組腫瘤標(biāo)志物水平預(yù)測(cè)脈管癌栓的敏感性、特異性、陽性預(yù)測(cè)價(jià)值、陰性預(yù)測(cè)價(jià)值;追蹤胃癌根治術(shù)Ⅲ期的輔助化療患者共270名,化療前后患者均進(jìn)行外周血TS mRNA檢測(cè)。將入組患者分成中藥結(jié)合化療和單純化療組。入組患者每月檢測(cè)外周血腫瘤標(biāo)志物CEA、CA19-9、CA242、CA72-4、CA125的水平,每三月復(fù)查影像學(xué)評(píng)價(jià)病情進(jìn)展。比較兩組患者的外周血腫瘤學(xué)標(biāo)志物變化情況,1年內(nèi)無復(fù)發(fā)生存率,以及化療前后外周血TS mRNA水平變化。[結(jié)果]1、單次給藥后獲得PPE對(duì)ICR小鼠的LD50值約為:12mg/ml,給藥劑量越接近LD50值的小鼠,其白細(xì)胞較正常小鼠組明顯下降,紅細(xì)胞及血小板無明顯變化,藥物對(duì)小鼠臟器未見明顯毒性作用。2、PPE對(duì)人胃癌細(xì)胞MKN-45的抑制率呈時(shí)間和劑量依賴性;重樓聯(lián)合放療可以降低腫瘤細(xì)胞的生存率;重樓醇提物單獨(dú)作用將MKN-45細(xì)胞阻滯在S期,而與放療聯(lián)合細(xì)胞周期阻滯在G2/M期;凋亡分析顯示PPE可以誘導(dǎo)腫瘤細(xì)胞凋亡,同時(shí)PPE與放療以協(xié)同作用方式誘導(dǎo)腫瘤細(xì)胞凋亡。PPE可下調(diào)Bcl-2蛋白的表達(dá)及上調(diào)Bax蛋白的表達(dá)。PPE可下調(diào)TS mRNA水平,在MKN-45細(xì)胞系中下調(diào)至0.59 ±0.07倍。3、入組的1005名患者中共有540名(53.73%)患者發(fā)現(xiàn)術(shù)后存在血管內(nèi)癌栓。對(duì)于至少有一項(xiàng)外周血腫瘤標(biāo)志物升高的患者來說,其脈管癌栓發(fā)生危險(xiǎn)性是沒有腫瘤標(biāo)志物升高患者的2.9倍。對(duì)于至少有四項(xiàng)外周血腫瘤標(biāo)志物升高的患者,對(duì)比沒有腫瘤標(biāo)志物升高的患者,其脈管癌栓發(fā)生風(fēng)險(xiǎn)是其16.4倍;輔助化療聯(lián)合中藥治療組與單純輔助化療組比較,6周期輔助化療后外周血腫瘤標(biāo)志物較化療前上升超過20%者分別有27名(19.3%)及34名(26.2%,P=0.178);1年內(nèi)病情復(fù)發(fā)的患者分別有31名(22.1%)及43名(33.1%,P=0.044);兩組間化療前外周血TS mRNA表達(dá)水平無顯著差異(聯(lián)合組:1.31,化療組:1.28,P=0.93),6周期化療后單純化療組患者外周血TS mRNA表達(dá)平均水平較化療前上升(1.28,1.42,P=0.43),化療聯(lián)合中藥治療組患者外周血TS mRNA表達(dá)平均水平較化療前下降(1.31,0.98,P=0.12),但均未達(dá)到統(tǒng)計(jì)學(xué)顯著性差異。[結(jié)論]1、PPE具有毒性,可能是通過血液白細(xì)胞的變化所致,對(duì)于腹腔臟器沒有明顯影響,臨床應(yīng)用需關(guān)注血液學(xué)檢測(cè)指標(biāo)。2、PPE具有顯著的體外抗腫瘤作用,其抗腫瘤作用的機(jī)理可能與影響細(xì)胞周期,下調(diào)Bcl-2蛋白的表達(dá)及上調(diào)Bax蛋白的表達(dá)來誘導(dǎo)腫瘤細(xì)胞凋亡,下調(diào)TS基因表達(dá)水平提高化療藥物敏感性等作用有關(guān);PPE與放療結(jié)合可以阻滯細(xì)胞周期,誘導(dǎo)腫瘤細(xì)胞凋亡,其機(jī)制可能與釋放活性氧相關(guān)。重樓是一味在腫瘤綜合治療中非常有前景的中藥。3、術(shù)前外周血腫瘤標(biāo)記物可以用于預(yù)測(cè)術(shù)后靜脈癌栓狀態(tài),對(duì)于術(shù)前血漿腫瘤標(biāo)志物升高的患者我們需要考慮更加積極的術(shù)前治療。4、胃癌Ⅲ期根治術(shù)后輔助化療患者中,聯(lián)合中藥治療較單純化療能顯著減少腫瘤復(fù)發(fā)風(fēng)險(xiǎn),其機(jī)制可能與中藥能夠降低患者TS mRNA表達(dá)從而提高氟尿嘧啶類化療藥物療效相關(guān)。
[Abstract]:[Objective] to study the acute toxic dose of Parispolyphlla ethanol extract (PPE) on ICR mice, to explore the application of heavy building in the comprehensive treatment of tumor, and to investigate the anti-tumor effect and mechanism in vitro, and to explore the relationship between the level of serum tumor markers before operation and the relationship between the tumor thrombus in the pulse tube after operation. The effect of the combination of traditional Chinese medicine and fluorouracil in the clinical application of gastric cancer and the regulating effect on the therapeutic effect of fluorouracil. [methods]1, ICR mice were used for acute toxicity test, the survival state, the change of Hematology Index, the change of abdominal organ as the observation index, and the dosage of median leath death, LD50), and the dosage of.2, and MTT. The method and cell clone formation method were used to detect the growth inhibition of PPE and PPE combined with radiotherapy for human gastric cancer cell MKN-45; the effect of PPE and PPE combined with radiotherapy on the MKN-45 cell cycle of human gastric cancer cell line were detected by PI; Western-blot was used to detect the effect of PPE on the level of apoptosis related proteins; PPE on fluorouracil was detected by RT-PCR method. Changes in the mRNA level of the thymidylatesynthase (TS) of the substance sensitive gene.3. The level of peripheral blood tumor markers, CEA, CA19-9, CA242, CA72-4, and CA125, were detected in 1005 patients undergoing radical gastrectomy for gastric cancer in general surgery in our hospital, and many of the postoperative specimens were included in the postoperative specimens including the vascular tumor thrombus. The baseline level data of the patients were grouped according to the status of the peripheral blood tumor markers, and the effects of each subgroup marker level on the vascular tumor thrombus status were analyzed. The sensitivity, specificity, positive predictive value and negative predictive value of the tumor markers in each subgroup were evaluated. 270 patients were treated with adjuvant chemotherapy in stage III, and peripheral blood TS mRNA detection was performed before and after chemotherapy. The patients were divided into groups of Chinese medicine combined with chemotherapy and chemotherapy alone. The patients enrolled in the group were detected the level of peripheral blood tumor markers CEA, CA19-9, CA242, CA72-4, CA125 every month, and evaluated the progress of the disease every March. Compare the two groups. There was no recurrence survival rate within 1 years, and the change of TS mRNA in peripheral blood before and after chemotherapy. [results]1, the LD50 value of PPE to ICR mice after a single dose of drug was about 12mg/ml, the more the dosage was closer to the LD50 value, the leukocytes were significantly lower than those of the normal mice, and the red blood cells and platelets had no obvious changes. The drug has no obvious toxic effect on mouse organs.2, and the inhibitory rate of PPE on human gastric cancer cell MKN-45 is time and dose dependent; heavy building combined with radiotherapy can reduce the survival rate of tumor cells; heavy building alcohol extracts separate MKN-45 cells in S phase alone, and the cell cycle block in G2/M phase with radiotherapy; apoptosis analysis shows PPE. The apoptosis of tumor cells could be induced, and the apoptosis of tumor cells was induced by PPE and radiotherapy in a synergistic manner..PPE could down regulate the expression of Bcl-2 protein and up regulate the expression of Bax protein,.PPE can down regulate the level of TS mRNA, down to 0.59 + 0.07 times.3 in MKN-45 cell line, and 540 (53.73%) patients in the 1005 patients of the group were found to exist after the operation. Intravascular thrombus. For patients with at least one increase in peripheral blood tumor markers, the risk of the vascular tumor thrombus was 2.9 times as high as those without a tumor marker. For patients with at least four peripheral blood tumor markers, the risk of the vascular tumor thrombus was 16. compared to those who had no higher tumor markers. 4 times, compared with the adjuvant chemotherapy group and the simple adjuvant chemotherapy group, there were 27 (19.3%) and 34 (26.2%, P=0.178) of peripheral blood tumor markers in the 6 cycle adjuvant chemotherapy than before chemotherapy, and 31 (22.1%) and 43 (33.1%, P=0.044) in 1 years, respectively, and TS mRNA of peripheral blood before chemotherapy before chemotherapy, and TS mRNA before chemotherapy. There was no significant difference in expression level (1.31, 1.31, chemotherapy group: 1.28, P=0.93). The average level of TS mRNA expression in peripheral blood of patients with chemotherapy after 6 cycles was higher than that before chemotherapy (1.28,1.42, P=0.43). The level of TS mRNA expression in peripheral blood of patients with chemotherapy combined with Chinese medicine treatment group was lower than that before chemotherapy (1.31,0.98, P=0.12), but no statistics were reached. [conclusion]1, PPE is toxic and may be caused by changes in blood leucocyte, it has no obvious influence on abdominal organs. Clinical application should pay attention to hematology detection index.2, PPE has significant antitumor effect in vitro, its anti-tumor mechanism can affect cell cycle, down regulation of Bcl-2 protein expression and The expression of Bax protein can induce apoptosis of tumor cells and reduce the level of TS gene expression to improve the chemosensitivity. The combination of PPE and radiotherapy can block cell cycle and induce apoptosis of tumor cells. The mechanism may be related to the release of reactive oxygen species. The anterior peripheral blood tumor markers can be used to predict the postoperative venous tumor thrombus status. For patients with elevated plasma tumor markers, we need to consider more active preoperative treatment of.4. In patients with adjuvant chemotherapy after stage III radical gastrectomy, the combination of traditional Chinese medicine and chemotherapy can significantly reduce the risk of tumor recurrence. The mechanism may be related to the possible mechanism. Chinese medicine can reduce the expression of TS mRNA in patients, thereby enhancing the efficacy of fluorouracil chemotherapy drugs.
【學(xué)位授予單位】：南京中醫(yī)藥大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：R735.2

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