食管腺癌细胞雌激素受体表达与化疗敏感性的研究

发布时间：2018-04-23 00:42

本文选题：食管腺癌 + Barrett’s食管　；参考：《天津医科大学》2015年博士论文

【摘要】：目的①探讨雌激素受体ERα、ERβ及其亚型在Barrett’s食管细胞系及食管腺癌细胞系细胞中的表达,并比较两种细胞系的表达差异。②探讨不同雌激素受体ERα和ERβ表达的食管腺癌细胞系对化疗药5-FU、顺铂、他莫昔芬的敏感性,并对两组数据进行相关分析,以期发现化疗敏感性与雌激素受体表达的关系。③探讨5-FU、顺铂联合他莫昔芬对不同雌激素受体表达细胞系细胞生长的影响。方法①对Barrett’s食管细胞系BAR-T、Ch TERT、Gi TERT、Qh TERT及食管腺癌细胞系OE19、OE33、SKGT4、OACP4C、FLO-1、OACM5.1进行培养,培养至细胞融合度约70%时收割细胞、提取蛋白,用EZQ工具包测蛋白浓度。采用Western blotting方法测定雌激素受体ERα、ERβ及其亚型在不同食管细胞系细胞中的表达水平。②利用MTS法测定最佳的MTS观察时间;建立各细胞系96小时生长曲线;根据细胞生长曲线决定96孔板每孔种植细胞的数量;对种植于96孔板OE19、OE33、SKGT4、OACP4C、FLO-1、OACM5.1等细胞系细胞分别予不同浓度5-FU、顺铂、他莫昔芬化学治疗72小时,比较各细胞系对药物的不同敏感性并将其结果与雌激素受体表达结果进行Spearman相关分析。③根据其雌激素受体表达的不同,我们选取雌激素受体ERβ高表达、ERα低表达的OE19及雌激素受体ERβ低表达、ERα高表达OACM5.1作为联合化疗对象。对选取的细胞系进行5-FU联合他莫昔芬及顺铂联合他莫昔芬联合化学治疗72小时,对其生长情况进行MTS检测。结果①通过Western blotting检测,Barrett’s食管细胞系细胞与食管腺癌细胞系细胞均不同程度地表达ERα或者ERβ,ERα表达以61k Da、90k Da亚型为主,ERβ以59k Da、51.5k Da及44.9k Da亚型为主。然而,与Brrett’s食管细胞系相比,食管腺癌细胞系细胞雌激素受体无论ERα或者ERβ表达水平均较高,而且在亚型ERβ44.9k Da表达上,两组相比差异具有统计学意义(P0.05)。②最佳的MTS观察时间为加MTS溶液后4小时内,以2小时或3小时观察为最佳,而24小时则不适宜观察。根据各细胞系96小时生长曲线,其每孔的适宜种植细胞数OE19、OE33、SKGT4应为2000个每孔,OACP4C、FLO-1、OACM5.1应为3000个每孔。各细胞系对5-FU、顺铂、他莫昔芬药物的化疗敏感性不同,对其不同的敏感性与雌激素受体表达行Spearman相关分析得知,在各细胞系中,5-FU、顺铂、他莫昔芬的敏感性与雌激素受体ERα61k Da、ERα90k Da、ERβ59k Da、ERβ51.5k Da、ERβ44.9k Da亚型表达无相关性,而雌激素受体亚型ERβ51.5k Da与他莫昔芬的敏感性呈负相关。③在雌激素受体ERβ高表达、ERα低表达细胞系OE19中,他莫昔芬能显著增加顺铂对细胞的杀伤作用,而他莫昔芬联合5-FU则无此作用。在雌激素受体ERβ低表达、ERα高表达细胞系OACM5.1中,他莫昔芬联合顺铂或5-FU均不能增加其杀伤作用。结论①Barrett’s食管细胞系细胞与食管腺癌细胞系细胞均不同程度地表达ERα、ERβ及其亚型,然而食管腺癌细胞系细胞雌激素受体ERα、ERβ表达水平均较高,尤其是ERβ44.9k Da的表达具有统计学意义,提示我们可以将此亚型作为鉴别Barrett’s食管和食管腺癌的指标之一。同时,我们在食管腺癌细胞系细胞观察到ERα61k Da亚型的表达。②不同的食管腺癌细胞系细胞对5-FU、顺铂、他莫昔芬的化疗敏感性是不同的。细胞对他莫昔芬的化疗敏感性与雌激素受体ERβ51.5k Da的表达呈负相关,提示雌激素受体某些亚型的表达与他莫昔芬化疗敏感性密切相关。③在高雌激素受体β、低α表达的食管腺癌细胞系中,他莫昔芬与顺铂具有协同杀伤作用。在高雌激素受体α、低β表达的食管腺癌细胞系则无此作用。这提示我们,对于高雌激素受体β、低α表达食管腺癌患者可以尝试他莫昔芬与顺铂联合治疗,以提高临床疗效。
[Abstract]:Objective: To investigate the expression of estrogen receptor ER alpha, ER beta and its subtypes in Barrett 's esophagus cell lines and esophageal adenocarcinoma cell lines, and to compare the differences in the expression of two cell lines. 2. The sensitivity of esophageal adenocarcinoma cell lines with different estrogen receptor ER alpha and ER beta expression to chemotherapy drug 5-FU, cisplatin and tamoxifen, and the number of two groups. To explore the relationship between chemosensitivity and the expression of estrogen receptor. (3) to explore the effect of 5-FU, cisplatin and tamoxifen on the growth of cell lines in different estrogen receptors. (1) Barrett 's esophageal cell line BAR-T, Ch TERT, Gi TERT, Qh TERT, and esophageal adenocarcinoma cell lines are OE19. OE33 -1, OACM5.1 were cultured to reaping cells, extracting protein and measuring protein concentration by EZQ toolkit. Western blotting method was used to determine the expression level of estrogen receptor ER alpha, ER beta and its subtypes in different esophageal cell lines. (2) the optimal MTS observation time was determined by MTS method, and each cell line 96 was established by MTS method. Hourly growth curve; determine the number of cells per hole of the 96 orifice according to the cell growth curve; the cell lines grown on the 96 orifice OE19, OE33, SKGT4, OACP4C, FLO-1, OACM5.1, and other cell lines were treated with different concentrations of 5-FU, cisplatin, and tamoxifen for 72 hours, and compared the different sensitivity of each cell line to the drug and the result and estrogen Spearman correlation analysis of the receptor expression results. (3) we select the high expression of estrogen receptor ER beta, low expression of ER alpha, low expression of ER alpha and low expression of ER beta in estrogen receptor, and ER alpha high expression OACM5.1 as a combined chemotherapy object. The growth situation of Finn was detected by MTS for 72 hours. Results (1) the expression of Barrett 's esophageal cell line cells and esophageal adenocarcinoma cell lines expressed ER alpha or ER beta in varying degrees by Western blotting. The expression of ER a was dominated by 61k Da, 90K Da subtype. Compared with the rrett 's esophageal cell line, the levels of both ER A and ER beta in the esophageal adenocarcinoma cell line were higher, and the difference between the two groups was statistically significant (P0.05) in the expression of the subtype ER beta 44.9k Da. The best time for MTS observation was to be best observed in 2 hours or 3 hours after the addition of MTS solution, and 24 small. According to the 96 hour growth curve of each cell line, the number of suitable planting cells per pore OE19, OE33, SKGT4 should be 2000 per pore, OACP4C, FLO-1, OACM5.1 should be 3000 per pore. The chemosensitivity of each cell line to 5-FU, cisplatin and tamoxifen is different, and its sensitivity to the expression of estrogen receptor is Spearman. The sensitivity of 5-FU, cisplatin and tamoxifen to the estrogen receptor ER alpha 61k Da, the ER alpha 90K Da, the ER beta 59K Da, the ER beta 51.5k, and the estrogen receptor subtype are negatively correlated with the sensitivity of tamoxifen. In cell line OE19, tamoxifen can significantly increase cisplatin's killing effect on cells, while tamoxifen combined with 5-FU without this effect. In the low expression of estrogen receptor ER beta, in the ER alpha high expression cell line OACM5.1, tamoxifen combined with cisplatin or 5-FU can not increase its killing effect. The cell line cells express ER alpha, ER beta and its subtypes in varying degrees. However, the expression level of estrogen receptor ER A and ER beta in the cell lines of the esophageal adenocarcinoma cell lines is higher, especially the expression of ER beta 44.9k Da, which suggests that we can use this subtype as one of the indicators to identify Barrett 's esophagus and esophageal adenocarcinoma. The expression of ER alpha 61k Da subtype was observed in the cells of the esophageal adenocarcinoma cell line. 2. The chemotherapeutic sensitivity of different esophageal adenocarcinoma cell lines to 5-FU, cisplatin and tamoxifen was different. The chemosensitivity of cells to tamoxifen was negatively correlated with the expression of the estrogen receptor ER beta 51.5k Da, suggesting the expression of some of the estrogen receptor subtypes and the expression of some of the estrogen receptor subtypes. Mooxifen is closely related to chemotherapy sensitivity. Tamoxifen has a synergistic killing effect with cisplatin in the high estrogen receptor beta and low alpha expression of esophageal adenocarcinoma cell lines. There is no effect in the high estrogen receptor alpha and low beta expression of the esophageal adenocarcinoma cell lines. This suggests that we have high estrogen receptor beta, low alpha expression of esophageal adenocarcinoma patients. We try to combine tamoxifen with cisplatin to improve the clinical efficacy.

【学位授予单位】：天津医科大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R735.1

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