Hamada K, Umemoto A, Kajikawa A, Seraj M J, Monden Y
Second Department of Surgery, School of Medicine, University of Tokushima, Japan.
Carcinogenesis. 1994 Sep;15(9):1911-5. doi: 10.1093/carcin/15.9.1911.
The in vitro experiment was carried out following 32P-postlabeling analysis to determine the DNA-reactive bile acids present in the human body. The unconjugated and conjugated forms of cholic (CA), chenodeoxycholic (CDCA), deoxycholic (DCA) and lithocholic acid (LCA) were added to calf thymus DNA followed by 1 h of incubation at 37 degrees C. After the incubation the mixture was analyzed by the nuclease P1 modification of 32P-postlabeling. Among the 12 bile acids tested, our results showed that unconjugated CDCA and LCA and the glycine and taurine conjugates of LCA (g-LCA, t-LCA) were able to bind covalently with naked DNA in vitro without intervention of any catalyst. It was also shown that bile acid alone did not give any spot on TLC. These binding reactions depended on the bile acid concentration in a linear manner. The data on the extent of binding at a concentration of 0.1 mg/ml showed values of 28.5 (t-LCA), 23.7 (g-LCA), 3.47 (LCA) and 1.32 (CDCA) adducts per 10(8) nucleotides. These reactive bile acids were also incubated with COLO 205 human colon carcinoma cells and Hep G2 human hepatocellular carcinoma cells for 24 h, but no specific DNA adduct was formed. Further, when LCA or CDCA was administered into male Fischer 344 rats by gavage at a dose of 10 mg/rat every 8 h for 3 days, no specific DNA adduct was detected in their liver or colon. Covalent DNA adducts are believed to cause alteration of the primary structure of genes, which is potentially linked to carcinogenesis. Though our preliminary data failed to detect any bile acid-related DNA adducts in the cultured cells or in rats, the results may provide a basis for assuming some of these bile acids to be potential initiators of colon cancer.
在进行32P后标记分析后开展体外实验，以确定人体中存在的具有DNA反应性的胆汁酸。将胆酸（CA）、鹅去氧胆酸（CDCA）、脱氧胆酸（DCA）和石胆酸（LCA）的未结合形式及结合形式添加到小牛胸腺DNA中，然后在37℃孵育1小时。孵育后，通过32P后标记的核酸酶P1修饰分析混合物。在所测试的12种胆汁酸中，我们的结果表明，未结合的CDCA和LCA以及LCA的甘氨酸和牛磺酸结合物（g-LCA、t-LCA）能够在体外无需任何催化剂干预的情况下与裸露DNA共价结合。还表明单独的胆汁酸在薄层层析上未出现任何斑点。这些结合反应以线性方式依赖于胆汁酸浓度。在浓度为0.1mg/ml时的结合程度数据显示，每10(8)个核苷酸的加合物值分别为28.5（t-LCA）、23.7（g-LCA）、3.47（LCA）和1.32（CDCA）。这些具有反应性的胆汁酸还与COLO 205人结肠癌细胞和Hep G2人肝癌细胞孵育24小时，但未形成特异性DNA加合物。此外，当以每只大鼠10mg的剂量、每8小时一次通过灌胃给雄性Fischer 344大鼠施用LCA或CDCA，持续3天时，在其肝脏或结肠中未检测到特异性DNA加合物。共价DNA加合物被认为会导致基因一级结构的改变，这可能与致癌作用相关。尽管我们的初步数据未能在培养细胞或大鼠中检测到任何与胆汁酸相关的DNA加合物，但这些结果可能为假定其中一些胆汁酸是结肠癌的潜在引发剂提供依据。
