microRNA调控乳腺癌围手术期T淋巴细胞免疫抑制的机制研究

The progression and metastasis of breast cancer is associated with mechanisms that inhibit the anti-tumor activity of T cells. Surgical excision is the mainstay of treatment for potentially curable breast cancer. It is increasingly recognized that anaesthetic technique and other perioperative factors have the potential to effect the anti-tumor function of immune system. However the mechanism is still unknown. MicroRNA (miRNA) is an important regulator of gene expression and cell differentiation. Based on our preliminary results, we intend to explore the regulation of miRNA on differentiation and immune function of T cells in peripheral blood of breast cancer patients on the different time points during perioperation. High-throughput microRNA microarray analysis, quantitative RT-PCR, and bioinformatics method will be used to detect the changes of miRNA expression and its target genes expression in different subtypes of lymphocytes in breast cancer patients perioperatively. Lentivirus vectors will be used to increase or decrease the expression of microRNA to observe its effect on T cell immunologic function. This would provide new clues to explain the biological mechanisms of immunosuppression in patients with breast cancer during perioperative period and also will provide new potential targets of RNA interference on the regulation of immune function in breast cancer patients.

乳腺癌的发展和转移与宿主的免疫功能有密切关系。围手术期的麻醉及手术等因素会导致机体抗肿瘤免疫功能降低，但机制仍旧不清楚。由于微小分子RNA（miRNA）是调控基因表达和细胞分化的重要因子，我们基于前期的研究基础和预实验数据，将进一步探索调控围手术期乳腺癌患者外周血T细胞分化命运的miRNA及其功能。通过采用高通量和定性定量的RNA组学检测技术，分析围手术期乳腺癌患者T淋巴细胞miRNA表达水平的变化，筛选不同亚型T细胞中差异表达的特定miRNA及其目标基因的表达，进一步探索干预特异的miRNA水平对T淋巴细胞分化方向的调控作用，从而观察它们对T细胞免疫功能的影响，并在此基础上研究控制T细胞分化方向的特异miRNA调控的分子通路。为阐明调控围手术期乳腺癌患者免疫抑制的生物学机制提供新线索，并为研制RNA干扰药物调控乳腺癌患者免疫功能提供新靶点。

乳腺癌的发展和转移与宿主的免疫功能有密切关系，围手术期的麻醉药物可能会使T淋巴细胞亚群比例发生变化，导致机体抗肿瘤免疫功能降低。本项目使用流式细胞术检测乳腺浸润性导管癌患者组与乳腺纤维瘤患者组围术期外周血中T淋巴细胞亚群比例，发现乳腺癌患者在术后72小时内CD4+、Thl细胞比例均较术前显著降低，Treg细胞比例显著升高。说明乳腺浸润性导管癌患者术后72小时内T细胞抗肿瘤免疫受抑制。关联性分析发现乳腺浸润性导管癌患者围术期T细胞亚群的变化与患者肿瘤负荷以及免疫组化标志HER2,ER相关，从而说明T细胞亚群变化与术后复发转移密切相关。.在麻醉药物对T淋巴细胞亚群的影响的研究中，我们发现右美托咪啶逆转T淋巴细胞亚群的改变，减轻术后细胞免疫功能的抑制。而吸入麻醉药七氟醚通过阻断原始T细胞向Th1细胞方向分化从而导致细胞免疫抑制。.乳腺癌患者术后肿瘤复发转移与术后肿瘤细胞休眠打破密切相关。我们使用裸鼠乳腺癌转移模型发现吗啡组裸鼠肺部转移灶数目较多，复发肿瘤增殖活跃、凋亡减少，肿瘤微血管密度增加；吗啡组复发肿瘤p-Akt、c-Myc、TSP-1蛋白表达升高。表明吗啡促进裸鼠乳腺癌术后复发肿瘤血管新生，促进癌细胞增殖、抑制凋亡，可能通过激活PI3K—c-Myc信号通路促使休眠的乳腺癌细胞脱离休眠状态发生肺转移。而我们使用非甾体抗炎药酮洛酸可抑制通过抑制cox-2受体进而抑制akt－c-myc、p-nfkb通路达到抑癌作用。上述实验发现阿片类镇痛药、非甾体抗炎药在乳腺癌术后细胞休眠复发过程中起到了重要作用，探究了术后镇痛药物的选择对乳腺癌转移、复发的影响及可能机制，为乳腺癌患者术后镇痛药物的选择提供依据。