Prostaglandin E(2) Activates YAP and a Positive-Signaling Loop to Promote Colon Regeneration After Colitis but Also Carcinogenesis in Mice.
Prostaglandin E(2) Activates YAP and a Positive-Signaling Loop to Promote Colon Regeneration After Colitis but Also Carcinogenesis in Mice.
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DOI：
10.1053/j.gastro.2016.11.005
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发表时间：
2017-02
期刊：
影响因子：
29.4
通讯作者：
Myung SJ
中科院分区：
文献类型：
--
作者：
Kim HB;Kim M;Park YS;Park I;Kim T;Yang SY;Cho CJ;Hwang D;Jung JH;Markowitz SD;Hwang SW;Yang SK;Lim DS;Myung SJ
前列腺素E2(PGE2)是调节组织再生的炎症介质，但其持续激活与肿瘤的发生有关。对PGE2信号通路中促成肿瘤形成的因素知之甚少。我们研究了河马信号通路中的转录共激活因子YAP1是否介导了PGE2的功能。将表达转基因或小发夹RNA的载体分别导入DLD-1和SW480结肠癌细胞株，并与重组PGE2共同孵育，分别用免疫印迹、免疫荧光、定量逆转录聚合酶链式反应、转录报告和细胞增殖实验进行分析。用葡聚糖硫酸钠(DSS)诱导C57/BL6小鼠(对照组)、羟基前列腺素脱氢酶15基因缺失小鼠(15-PGDH基因敲除小鼠)、YAP1基因敲除小鼠(YAP基因敲除小鼠)和双基因敲除小鼠结肠炎模型。一些小鼠也被给予吲哚美辛以阻止PGE2的合成。15-PGDH基因敲除小鼠与肠道特异性破坏了萨尔瓦多家族包含1个基因(Sav1)的WW结构域的小鼠杂交，Sav1编码河马信号的激活剂。我们对26例结肠炎相关癌患者和51例年龄和性别匹配的结直肠癌(非结肠炎)患者的结肠活检标本进行了免疫组织化学分析。结肠癌细胞株与前列腺素E_2共同孵育后，cAMP反应元件结合蛋白1(CREB_1)磷酸化，YAP_1mRNA和蛋白水平及YAP_1‘S转录活性增加。这导致前列腺素-过氧化物合成酶2基因(Ptgs2或COX2)和前列腺素E受体4基因(PTGER4或EP4)转录增加。与PGE2孵育可促进结肠癌细胞系的增殖，但不能促进YAP1基因敲除的细胞的增殖。对照组小鼠在注射DSS后出现结肠炎，但注射PGE2导致这些小鼠的结肠再生。然而，YAP基因敲除小鼠不能再生结肠组织，并在注射DSS后不久死亡。在停用DSS后，15-PGDH基因敲除小鼠比对照小鼠更快地再生结肠组织，体重、结肠长度和结肠炎组织学评分恢复得更快。这些作用可被注射消炎痛逆转。SAV1基因敲除或15-PDGH基因敲除小鼠在结肠炎诱导后没有发生自发性肿瘤，但SAV1/15-PDGH双基因敲除小鼠出现息肉，最终发展为原位癌。给这些小鼠注射吲哚美辛可防止自发肿瘤的形成。在人类散发性结直肠癌和结肠炎相关肿瘤中，PGE2水平与YAP水平相关。PGE2信号增强YAP1的表达和转录活性，导致COX2和EP4的表达增加，从而激活一个正向信号循环。该途径促进结肠癌细胞系的增殖和结肠炎小鼠的结肠组织再生。该通路的结构性激活导致了小鼠息肉和结肠癌的形成。
Prostaglandin E2 (PGE2) is mediator of inflammation that regulates tissue regeneration, but its continual activation has been associated with carcinogenesis. Little is known about factors in the PGE2 signaling pathway that contribute to tumor formation. We investigated whether yes associated protein 1 (YAP1), a transcriptional co-activator in the Hippo signaling pathway, mediates PGE2 function. DLD-1 and SW480 colon cancer cell lines were transfected with vectors expressing transgenes or small hairpin RNAs and incubated with recombinant PGE2, with or without pharmacologic inhibitors of signaling proteins, and analyzed by immunoblot, immunofluorescence, quantitative reverse transcription PCR, transcriptional reporter, and proliferation assays. Dextran sodium sulfate (DSS) was given to induce colitis in C57/BL6 (control) mice, as well as in mice with disruption of the hydroxyprostaglandin dehydrogenase 15 gene (15-PGDH-knockout mice), Yap1 gene (YAP-knockout mice), and double knockout mice. Some mice were also given indomethacin to block PGE2 synthesis. 15-PGDH knockout mice were crossed with mice with intestine-specific disruption of the Salvador family WW domain containing 1 gene (Sav1), which encodes an activator of Hippo signaling. We performed immunohistochemical analyses of colon biopsy samples from 26 patients with colitis-associated cancer and 51 age- and sex-matched patients with colorectal cancer (without colitis). Incubation of colon cancer cell lines with PGE2 led to phosphorylation of cAMP responsive element binding protein 1 (CREB1) and increased levels of YAP1 mRNA and protein and YAP1’s transcriptional activity. This led to increased transcription of the prostaglandin-endoperoxide synthase 2 gene (PTGS2 or COX2) and prostaglandin E receptor 4 gene (PTGER4 or EP4). Incubation with PGE2 promoted proliferation of colon cancer cell lines, but not cells with knockdown of YAP1. Control mice developed colitis after administration of DSS, but injection of PGE2 led to colon regeneration in these mice. However, YAP-knockout mice did not regenerate colon tissues and died soon after administration of DSS. 15-PGDH-knockout mice regenerated colon tissues more rapidly than control mice after withdrawal of DSS, and had faster recovery of body weight, colon length, and colitis histology scores. These effects were reversed by injection of indomethacin. SAV1 -knockout or 15-PDGH-knockout mice did not develop spontaneous tumors following colitis induction, but SAV1/15-PDGH double knockout mice developed polyps that eventually progressed to carcinoma in situ. Administration of indomethacin to these mice prevented spontaneous tumor formation. Levels of PGE2 correlated with those of YAP levels in human sporadic colorectal tumors and colitis-associated tumors. PGE2 signaling increases expression and transcriptional activities of YAP1, leading to increased expression of COX2 and EP4 to activate a positive signaling loop. This pathway promotes proliferation of colon cancer cell lines and colon tissue regeneration in mice with colitis. Constitutive activation of this pathway led to formation of polyps and colon tumors in mice.