基于SIRT1介导的P53信号通路复方蜥蜴散从毒瘀交阻论治胃癌机制研究

Promoting cell apoptosis,inhibiting proliferation and differentiation is the important link of blocking gastric cancer development.Our topic group research shows,toxin and blood stasis cross resistance syndrome is main syndrome of precancerous lesion of gastric cancer and gastric cancer.Based on the early stage of the different particle combination agent to cure precancerous lesions of gastric cancer with the ningxia eremias multiocellata,found that the prescriptions could effectively control P53,promote cell apoptosis,decrease CEA,block the canceration. P53 is an important role substrate of SIRT1,an important factor related with blood stasis syndrome;SIRT1 can regulate tumor cell apoptosis in ischemia,hypoxia environment,namely,toxin and blood stasis cross resistance,mucosal dystrophy environment mechanism of activating the same.Therefore,we propose the compound lizards powder may be mediated through SIRT1,and control the P53 expression is the main blocking gastric cancer mechanism.This topic applicate TUNEL,immunohistochemistry, MTT,Western-blot method and tissue microarray technique,to observate of SIRT1 regulation of human gastric cancer cell SGC-7901 in vitro and vaccination in Kunming nude mice gastric carcinoma model in P53 expression intervention with the compound lizards powder,the MTT and TUNEL in gastric mucosal cell apoptosis,the flow cytometry in cell cycle,to verificate anticancer hypothesis of the compound lizards powder by SIRT1 mediated regulation P53 expression,explain anticancer mechanism.

促进细胞凋亡、抑制增殖分化是阻断胃癌发生重要环节。本课题组研究表明，胃癌前病变及胃癌证候以毒瘀交阻为主，前期开展以宁夏密点麻蜥为主的复方蜥蜴散治疗胃癌前病变研究，发现该方可有效调控P53,促进细胞凋亡，降低CEA,阻断癌变。P53是SIRT1重要作用底物之一，是血瘀证重要相关因子；SIRT1可调控肿瘤细胞凋亡，在缺血缺氧环境下，即毒瘀交阻，黏膜失养环境下激活机制相同。因此,我们提出复方蜥蜴散可能通过介导SIRT1，进而调控p53表达是阻断胃癌主要机制。本课题拟以免疫组化、Western-blot、Envision等方法及组织芯片技术，观察复方蜥蜴散基于SIRT1调控对人SGC-7901胃癌细胞株体外培养及接种昆明种裸鼠胃癌模型P53表达干预，MTT、TUNEL测细胞凋亡,流式细胞仪测细胞周期，验证复方蜥蜴散通过介导SIRT1调控P53表达抗癌假说，阐释该方抗癌机制。

促进细胞凋亡、抑制增殖分化是阻断胃癌发生重要环节。本项目基于胃癌证候以毒瘀交阻为主，肿瘤细胞在缺血缺氧即毒瘀交阻，黏膜失养环境下激活机制相一致的病机，通过研究复方蜥蜴散基于SIRT1调控对人SGC-7901胃癌细胞株体外培养及体内接种昆明种裸鼠胃癌模型P53表达干预，MTT、TUNEL测细胞凋亡,流式细胞仪测细胞周期，阐释该方抗癌机制。.体外实验研究中，通过制备复方蜥蜴散不同微粒组合剂含药SD大鼠血清，加于人胃癌细胞SGC-7901培养24h后，检测表明含药大鼠血清干预组细胞总凋亡率大于生理盐水阴性对照组及华蟾素片阳性对照组、5-Fu阳性对照组，复方蜥蜴散高剂量组G1期细胞比例大于其他中药组，P<0.05。各含药血清组SIRT1 、P53蛋白表达均低于生理盐水阴性对照组；高剂量组表达低于其他药物组，P<0.05。复方蜥蜴散拆方含药SD大鼠血清组间比较，以补气组作用较佳，其次为解毒通络组；各干预组P53、bax表达水平与生理盐水对照组比较，呈显著升高趋势，以复方蜥蜴散组为著。.此外，宁夏密点麻蜥不同部位含药SD大鼠血清各组可明显抑制SGC-7901胃癌细胞增殖，诱导凋亡，宁夏密点麻蜥尾部组细胞总凋亡率大于其他各组，宁夏密点麻蜥不同部位含药大鼠血清干预组Sirt1和P53蛋白表达低于生理盐水对照组，P<0.05；尾部组与其他各组比较，Sirt1和P53蛋白表达低明显，P<0.05。.宁夏密点麻蜥含药大鼠血清各干预组OD均值小于生理盐水对照组，P53、Bax以及E-cad蛋白表达水平与生理盐水组比显著升高，MMP-9蛋白表达水平与生理盐水组比降低。随着血清药物浓度的增高，MMP-9的表达呈依赖性降低，P53、Bax以及E-cad的表达呈依赖性增强。.体内抗癌研究中，建立裸鼠原位移植SGC7901胃癌模型，随机分为生理盐水对照组、华蟾素对照组、复方蜥蜴散高、中、低计量组，药物连续干预21天后处理所有小鼠；将肿瘤组织进行免疫组织化学染色，TUNEL法检测凋亡细胞；Western－Blot检测胃癌细胞SIRT1 、P53蛋白、HIF-1α、PI3K、AKT蛋白表达水平的变化。复方蜥蜴散不同微粒组合剂可能通过降低SIRT1 、P53蛋白表达，阻断HIF-1α介导PI3K/AKT，抑制EMT而抗癌。