IL-6/mTOR参与NCP转变为PIN过程中的作用机制研究

It has demonstrated that non-controlled prostate inflammation (NCP) can change into prostatic intraepithelial neoplasia(PIN), however, little is known about the its key regulatory molecules mechanisms and regulation rules.We previously study show that mTOR in PIN markedly higher than NCP, this suggestion that the abnormal expression of mTOR relate with the development of PIN.it is also showed that IL-6 and autophagy are play an important role in the process of NCP change into PIN;and IL-6 through mTOR pathway regulate the development of inflammation, this suggestion that the mechanisms of NCP change into PIN result from molecular regulatory by IL-6/mTOR.So, We will intend to study breakthrough point of IL-6/mTOR,apply the new animal models of PIN,aim to the the process of PIN, constuct the animal models of NCP change into PIN; revealing the role of IL-6 、mTOR and autophagy in the NCP change into PIN;revealing its mechanisms of the molecular regulatory;constuct the key regulatorysites and signaling pathways in NCP change into PIN, to provide the new strategies and targets with the PIN.

研究表明前列腺非可控性炎症（NCP）可转变为前列腺上皮内瘤变（PIN），但目前对此过程关键分子调控机制和规律仍然不清。我们前期研究显示mTOR在PIN表达明显比NCP高，提示mTOR的异常表达可能与PIN的发生发展相关。已有研究显示IL-6和自噬在NCP转变为PIN的过程中起重要作用；且IL-6通过mTOR信号通路调控炎症发生发展过程，提示NCP转变为PIN的机制可能是由IL-6/mTOR组成的分子网络调控的结果。因此，本项目以IL-6/mTOR为切入点，拟采用国际最新的PIN动物模型，针对PIN发生发展过程，运用分子生物学及动物模型等方法，构建NCP转变为PIN的动物模型；阐明IL-6 、mTOR和自噬在此过程中的作用；揭示IL-6在NCP转变为PIN过程中分子调控机制；构建其关键调控位点和信号通路，为PIN的防治提供新的策略与作用靶点。

本研究从分子、细胞、动物、临床等多层次探索了非可控性前列腺炎（NCP）与前列腺上皮内瘤变（PIN）发生之间的内在关系。首先，课题组在慢性前列腺炎、前列腺癌患者血清中发现了IL-6水平的差异，IL-6水平在前列腺癌患者中升高。并进一步在前列腺组织标本中发现了mTOR通路中相关蛋白的表达差异，前列腺癌组织中Akt、mTOR、Raptor和Rictor表达水平较前列腺炎组织高（P＜0.05）。成功构建NCP大鼠模型，分离获取NCP大鼠前列腺上皮细胞，并进一步转染Akt/ERG过表达的慢病毒，筛选表达绿色荧光蛋白的细胞，qPCR检测证实Akt及ERG基因mRNA表达水平明显较对照组和空白组升高（P＜0.05）。成功获取mTOR通路过度激活的NCP大鼠前列腺上皮细胞，进一步进行Western Blot检测mTOR相关蛋白，发现实验组中p-Akt、p-mTOR、Raptor及Rictor蛋白表达明显升高（P<0.05）。在成瘤实验中，对非肥胖糖尿病/重症联合免疫缺陷(NOD-SCID)裸鼠皮下注射Akt/ERG过表达前列腺上皮细胞，30天后皮下肿瘤平均体积达602.3±46.2 mm3。使用不同浓度mTOR抑制剂Rapamycin瘤内注射时，肿瘤的体积增长受到抑制，且随着Rapamycin注射的剂量增加，生长抑制受到抑制更明显（P＜0.05）。进一步检测各组中自噬通路相关蛋白，随着Rapamycin注射的剂量增加，LC3 II/LC3 I比值逐渐升高，Baclin 1蛋白逐渐升高，p62蛋白逐渐降低，提示自噬过程增强。本研究初步阐明，IL-6在前列腺癌患者中有较高表达，mTOR通路在PIN发生时过度激活，而mTOR抑制剂Rapamycin可以通过增强自噬过程来阻止PIN的发展，为前列腺癌的治疗提供了新的策略与作用靶点。