Fusobacterium nucleatum Increases Proliferation of Colorectal Cancer Cells and Tumor Development in Mice by Activating Toll-Like Receptor 4 Signaling to Nuclear Factor-κB, and Up-regulating Expression of MicroRNA-21.
Fusobacterium nucleatum Increases Proliferation of Colorectal Cancer Cells and Tumor Development in Mice by Activating Toll-Like Receptor 4 Signaling to Nuclear Factor-κB, and Up-regulating Expression of MicroRNA-21.
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具核梭杆菌通过激活核因子 kappa B 的 Toll 样受体 4 信号传导以及上调 MicroRNA-21 的表达来增加小鼠结直肠癌细胞的增殖和肿瘤的发展
DOI：
10.1053/j.gastro.2016.11.018
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发表时间：
2017-03
期刊：
影响因子：
29.4
通讯作者：
Ma Y
中科院分区：
文献类型：
--
作者：
Yang Y;Weng W;Peng J;Hong L;Yang L;Toiyama Y;Gao R;Liu M;Yin M;Pan C;Li H;Guo B;Zhu Q;Wei Q;Moyer MP;Wang P;Cai S;Goel A;Qin H;Ma Y
全球近20%的癌症负担可能与感染因子有关。具核梭杆菌通过不清楚的机制促进上皮细胞的肿瘤形成。我们的目的是鉴定F nucleatum诱导的microRNA（miRNAs），并评估其促进小鼠结直肠癌发生的能力。将结肠直肠癌（CRC）细胞系与F nucleatum或对照试剂一起孵育，并在增殖和愈合测定中进行分析。将HCT 116、HT 29、LoVo和SW 480 CRC细胞系与F nucleatum或磷酸盐缓冲盐水（PBS对照）一起孵育，并在染色质免疫沉淀测定中分析miRNA表达模式。将细胞与miRNA模拟物、对照序列或小干扰（si）RNA孵育;在荧光素酶测定中测量报告构建体的表达。将CRC细胞与F nucleatum或PBS孵育，并注射到BALB/C裸鼠中;测量异种移植肿瘤的生长。C57 BL APCmin/+、C57 BL miR 21 a −/−和具有全长miR 21 a的C57 BL小鼠（对照）通过管饲法给予F nucleatum;一些小鼠给予氧化偶氮甲烷（AOM）和葡聚糖硫酸钠（DSS）以诱导结肠炎和结肠肿瘤。收集肠组织并计数肿瘤。通过ELISA分析来自小鼠的血清样品的细胞因子水平。我们进行了原位杂交分析，以检测结直肠癌细胞中F核的富集。对来自中国的90例肿瘤和匹配的非肿瘤组织中的F核DNA进行了探索，以进行表达相关性分析;将来自日本的125例肿瘤组织中的F核DNA水平与其生存时间进行了比较。与对照细胞相比，F nucleatum增加了CRC细胞系的增殖和侵袭活性。大肠癌细胞系感染F nucleatum形成更大的肿瘤，更迅速，在裸鼠比未感染的细胞。APCmin/+小鼠灌胃F nucleatum发展显着更多的结肠直肠肿瘤比小鼠给予PBS和较短的生存时间。我们发现，给予F核的小鼠血清中的几种炎症因子（白细胞介素17 F [IL 17 F]、IL 21、IL 22和MIP 3A）显著增加。我们发现，与PBS相比，在用F nucleatum孵育的CRC中，50种miRNA显著上调，52种miRNA显著下调; miR 21的水平增加最多（超过4倍）。miR 21的抑制剂阻止了培养物中F nucleatum诱导细胞增殖和侵袭。与对照小鼠相比，miR 21 a −/−小鼠在给予AOM和DSS后出现便血和腹泻的时间较晚，并且存活时间较长。miR 21 a −/−小鼠的结肠直肠肿瘤较少，尺寸较小，并且miR 21 a −/−小鼠比对照小鼠存活时间更长。我们发现RASA 1，它编码一个RAS GT3，是一个目标基因一致下调的细胞中过表达miR 21和上调暴露于miR 21抑制剂的细胞。用具核F感染细胞通过激活TLR 4信号传导至MYD 88增加miR 21的表达，导致核因子NFκB的激活。与来自患者的非肿瘤结肠组织相比，肿瘤组织中F核DNA和miR 21的水平增加（并且在晚期肿瘤组织中甚至更多）。具有高量F核DNA和miR 21的肿瘤患者的生存时间比具有较低量的肿瘤患者的生存时间短。我们发现用具核F感染CRC细胞可增加其增殖、侵袭活性和在小鼠中形成异种移植肿瘤的能力。F nucleatum激活TLR 4信号传导至MYD 88，导致核因子NFκB的激活和miR 21表达的增加;该miRNA降低RAS GTP酶RASA 1的水平。具有高量组织F核DNA和miR 21的患者表现出不良结局的较高风险。
Nearly 20% of the global cancer burden can be linked to infectious agents. Fusobacterium nucleatum promotes tumor formation by epithelial cells via unclear mechanisms. We aimed to identify microRNAs (miRNAs) induced by F nucleatum and evaluate their ability to promote colorectal carcinogenesis in mice. Colorectal cancer (CRC) cell lines were incubated with F nucleatum or control reagents and analyzed in proliferation and would healing assays. HCT116, HT29, LoVo, and SW480 CRC cell lines were incubated with F nucleatum or phosphate buffer saline (PBS control) and analyzed for miRNA expression patterns and in chromatin immunoprecipitation assays. Cells were incubated with miRNAs mimics, control sequences, or small interfering (si) RNAs; expression of reporter constructs was measured in luciferase assays. CRC cells were incubated with F nucleatum or PBS and injected into BALB/C nude mice; growth of xenograft tumors was measured. C57BL APCmin/+, C57BL miR21a−/−, and C57BL mice with full-length miR21a (controls) were given F nucleatum by gavage; some mice were given azoxymethane (AOM) and dextran sodium sulfate (DSS) to induce colitis and colon tumors. Intestinal tissues were collected and tumors were counted. Serum samples from mice were analyzed for cytokine levels by ELISAs. We performed in situ hybridization analyses to detect enrichment of F nucleatum in CRC cells. F nucleatum DNA in 90 tumor and matched non-tumor tissues from patients in China were explored for the expression correlation analysis; levels in 125 tumor tissues from patients in Japan were compared with their survival times. F nucleatum increased proliferation and invasive activities of CRC cell lines, compared with control cells. CRC cell lines infected with F nucleatum formed larger tumors, more rapidly, in nude mice than uninfected cells. APCmin/+ mice gavaged with F nucleatum developed significantly more colorectal tumors than mice given PBS and had shorter survival times. We found several inflammatory factors to be significantly increased in serum from mice given F nucleatum (interleukin 17F [IL17F], IL21, IL22, and MIP3A). We found 50 miRNAs to be significantly upregulated and 52 miRNAs to be significantly downregulated in CRCs incubated with F nucleatum vs PBS; levels of miR21 increased by the greatest amount (more than 4-fold). Inhibitors of miR21 prevented F nucleatum from inducing cell proliferation and invasion in culture. miR21a−/− mice had a later appearance of fecal blood and diarrhea after administration of AOM and DSS, and had longer survival times, compared with control mice. The colorectum of miR21a−/− mice had fewer tumors, of smaller size, and the miR21a−/− mice survived longer than control mice. We found RASA1, which encodes a RAS GTPase, to be one of the target genes consistently downregulated in cells that overexpressed miR21 and upregulated in cells exposed to miR21 inhibitors. Infection of cells with F nucleatum increased expression of miR21 by activating TLR4 signaling to MYD88, leading to activation of the nuclear factor NFκB. Levels of F nucleatum DNA and miR21 were increased in tumor tissues (and even more so in advanced tumor tissues), compared with non-tumor colon tissues from patients. Patients whose tumors had high amounts of F nucleatum DNA and miR21 had shorter survival times than patients whose tumors had lower amounts. We found infection of CRC cells with F nucleatum to increase their proliferation, invasive activity, and ability to form xenograft tumors in mice. F nucleatum activates TLR4 signaling to MYD88, leading to activation of the nuclear factor NFκB and increased expression of miR21; this miRNA reduces levels of the RAS GTPase RASA1. Patients with both high amount of tissue F nucleatum DNA and miR21 demonstrated a higher risk for poor outcomes.
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影响因子：
5.4
作者：
Han YW
通讯作者：
Han YW
影响因子：
24.5
作者：
Mima K;Nishihara R;Qian ZR;Cao Y;Sukawa Y;Nowak JA;Yang J;Dou R;Masugi Y;Song M;Kostic AD;Giannakis M;Bullman S;Milner DA;Baba H;Giovannucci EL;Garraway LA;Freeman GJ;Dranoff G;Garrett WS;Huttenhower C;Meyerson M;Meyerhardt JA;Chan AT;Fuchs CS;Ogino S
通讯作者：
Ogino S
影响因子：
30.3
作者：
Liu S;da Cunha AP;Rezende RM;Cialic R;Wei Z;Bry L;Comstock LE;Gandhi R;Weiner HL
通讯作者：
Weiner HL
影响因子：
16.6
作者：
Cantini L;Isella C;Petti C;Picco G;Chiola S;Ficarra E;Caselle M;Medico E
通讯作者：
Medico E
影响因子：
4.4
作者：
Liu, Hongqi;Redline, Raymond W.;Han, Yiping W.
通讯作者：
Han, Yiping W.