Wang Hong, Liu Anna, Kuo Yingyi, Chi Eric, Yang Xu, Zhang Lanjing, Yang Chung S
Susan L. Cullman Laboratory for Cancer Research, Department of Chemical Biology and Center for Cancer Prevention Research, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, 164 Frelinghuysen Road, Piscataway, NJ 08854, USA.
Department of Pathology, University Medical Center of Princeton, Plainsboro, NJ 08536, USA and.
Carcinogenesis. 2016 Jul;37(7):723-730. doi: 10.1093/carcin/bgw054. Epub 2016 May 2.
Obesity is associated with an increased risk of cancer. To study the promotion of dietary carcinogen-induced gastrointestinal cancer by obesity, we employed 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) to induce intestinal tumorigenesis in CYP1A-humanized (hCYP1A) mice, in which mouse Cyp1a1/1a2 was replaced with human CYP1A1/1A2 Obesity was introduced in hCYP1A mice by breeding with Lepr(db/+) mice to establish the genetically induced obese hCYP1A-Lepr(db/db) mice or by feeding hCYP1A mice a high-fat diet. PhIP induced the formation of small intestinal tumors at the ages of weeks 28-40 in obese hCYP1A mice, but not in lean hCYP1A mice. No tumors were found in colon and other gastrointestinal organs in the lean or obese mice. Using immunohistochemistry (IHC), we found strong positive staining of NF-κB p65, pSTAT3 and COX2 as well as elevated levels of nuclear β-catenin (Ctnnb1) in small intestinal tumors, but not in normal tissues. By sequencing Apc and Ctnnb1 genes, we found that most PhIP-induced small intestinal tumors in obese mice carried only a single heterozygous mutation in Apc By bisulfite-sequencing of CpG islands of Apc, we found DNA hypermethylation in a CpG cluster located in its transcription initiation site, which most likely caused the inactivation of the wild-type Apc allele. Our findings demonstrate that PhIP-induced small intestinal carcinogenesis in hCYP1A-db/db mice is promoted by obesity and involves Apc mutation and inactivation by DNA hypermethylation. This experimental result is consistent with the association of obesity and the increased incidence of small intestinal cancer in humans in recent decades.
肥胖与癌症风险增加相关。为研究肥胖对饮食致癌物诱导的胃肠道癌症的促进作用，我们使用2-氨基-1-甲基-6-苯基咪唑并[4,5-b]吡啶（PhIP）在CYP1A人源化（hCYP1A）小鼠中诱导肠道肿瘤发生，其中小鼠Cyp1a1/1a2被人CYP1A1/1A2取代。通过与Lepr(db/+)小鼠杂交，在hCYP1A小鼠中引入肥胖，以建立基因诱导肥胖的hCYP1A-Lepr(db/db)小鼠，或通过给hCYP1A小鼠喂食高脂饮食来引入肥胖。PhIP在28-40周龄时诱导肥胖hCYP1A小鼠发生小肠肿瘤，但在瘦的hCYP1A小鼠中未诱导出肿瘤。在瘦小鼠或肥胖小鼠的结肠和其他胃肠道器官中未发现肿瘤。通过免疫组织化学（IHC），我们发现小肠肿瘤中NF-κB p65、pSTAT3和COX2呈强阳性染色，以及核β-连环蛋白（Ctnnb1）水平升高，但在正常组织中未发现。通过对Apc和Ctnnb1基因进行测序，我们发现肥胖小鼠中大多数PhIP诱导的小肠肿瘤仅在Apc中携带一个杂合突变。通过对Apc的CpG岛进行亚硫酸氢盐测序，我们发现在其转录起始位点的一个CpG簇中存在DNA高甲基化，这很可能导致野生型Apc等位基因失活。我们的研究结果表明，肥胖促进了hCYP1A-db/db小鼠中PhIP诱导的小肠致癌作用，并且涉及Apc突变以及DNA高甲基化导致的失活。这一实验结果与近几十年来肥胖与人类小肠癌发病率增加之间的关联一致。
