塞来昔布对结肠腺瘤干/前体细胞β-catenin/Lgr5分子的调控研究

Prevention is very important in tumor. Colonic adenoma stem cell is a new field for the prevention of colon cancer, and now, colonic adenoma stem cell has been separated and cultured sucessfully. NSAIDs can prevent of colon cancer effectively, but due to its complications, its clinical applications are limited. Some literature and our previous study found that aspirin inhibit the expressions of “stemness” makers in colon cancer cell, and can regulate the β-catenin (a key molecular of wnt pathway) expression in colon cancer. But the specific mechanism of NSAIDs regulating colon adenoma stem cells is unclear. In our research, the human colon adenoma stem / progenitor cells will be separated and cultured. And celecoxib will be used to intervent these cells, and β-catenin will be interfered and overexpressed, wnt signa will be activated too. Then the phosphorylated and non-phosphorylated β-catenin, downstream “stemness” (such as lgr5) and differentiation related makers will be detected, and biological behaviors (such as tumorsphere formation) and tumorigenicity in nude will be also observed. Eventually, the theory of celecoxib regulating wnt /β-catenin signal pathway, then regulating the "stemness" of colonic adenoma stem / progenitor cells, then affecting the self-renewal and differentiation of colonic adenoma stem / progenitor cells. And we may find fundamental targets for NSAIDs preventing and treating of colon cancer.

肿瘤重在预防，结肠腺瘤干细胞的提出，为结肠癌的预防提出新的理念，目前可成功分离、培养之。非甾体类抗炎药(NSAIDs)预防结肠癌有效，能明显减少甚至消退结肠腺瘤，但并发症多，限制临床应用。文献报道及我们前期研究发现，NSAIDs抑制结肠癌“干性”标志物表达，并在结肠癌参与调节Wnt通路关键分子β-catenin。但NSAIDs如何调控结肠腺瘤干细胞不清楚。本课题拟原代培养人结肠腺瘤干/前体细胞，采用塞来昔布干预，通过对β-catenin干扰和过表达，以及对Wnt信号激动等，检测磷酸化和非磷酸化β-catenin、下游Lgr5等干性靶标和肠分化标志物表达，观察细胞成球等生物学行为以及裸鼠成瘤能力变化，最终确立塞来昔布调节Wnt/β-catenin信号通路——抑制结肠腺瘤干/前体细胞“干性”——影响结肠腺瘤干/前体细胞自我更新、分化等，为寻找NSAIDs预防结肠癌根本作用靶点提供理论依据。

肿瘤干细胞是肿瘤的“种子”和“源泉”，该理论的提出，为结肠癌的防治提出新的理念。非甾体类抗炎药(NSAIDs)防治结肠癌有效，但如何调控结肠肿瘤干细胞“干性”、分化和上皮间充质转变(EMT)这些根本机制目前不清楚。本课题通过流式双标分选培养结肠肿瘤干/前体细胞，以及动物实验，分别采用NSAIDs塞来昔布干预，通过检测磷酸化和非磷酸化β-catenin表达变化，以及下游Lgr5、 CD133等干性靶标、MUC2等肠分化标志物、E-cadherin、Vimentin等EMT相关标志物表达，观察对细胞增殖、成球、迁移和侵袭等生物学行为影响情况，以及结合人体标本实验，最终确立塞来昔布通过调节Wnt/β-catenin信号通路抑制结肠肿瘤干/前体细胞“干性”，影响结肠肿瘤干/前体细胞自我更新、分化以及EMT转变，最终影响结肠肿瘤的发生、进展，并且通过对5-脂氧合酶(5-LOX)抑制剂和miRNA-302b对结肠癌“干性”的调控机制研究，从肿瘤干细胞这一根本理论出发更深入地探讨了NSAIDs塞来昔布等防治结肠肿瘤可能存在的根本分子机制，为寻找NSAIDs防治结肠癌的根本作用靶点提供理论依据。