Department of Pathophysiological Science, Faculty of Pharmaceutical Sciences, Hokuriku University, Kanazawa, 920-1181, Japan.
J Gastroenterol. 2010 Jun;45(6):646-55. doi: 10.1007/s00535-009-0193-9. Epub 2010 Jan 19.
Nonalcoholic steatohepatitis (NASH) is a feature of metabolic syndrome. Advanced glycation end-products (AGEs) are formed by the Maillard reaction, which contributes to aging and to certain pathological complications of diabetes. A recent study has suggested that glyceraldehyde-derived AGEs (Glycer-AGEs) are elevated in the sera of patients with NASH. Furthermore, immunohistochemistry of Glycer-AGEs showed intense staining in the livers of patients with NASH. The present study aimed to examine the effect of intracellular Glycer-AGEs on hepatocellular carcinoma (Hep3B) cells.
Cell viability was determined by the WST-1 assay. The slot blot and Western blot were used to detect intracellular Glycer-AGEs, and their localization was analyzed by confocal microscopy. Real-time reverse transcription-polymerase chain reaction was used to quantify the mRNA for the acute phase reactant C-reactive protein (CRP).
Glyceraldehyde (GA), which is the precursor of Glycer-AGEs, induced a concentration- and time-dependent increase in cell death, which was associated with an increase in intracellular Glycer-AGEs formation. Aminoguanidine (AG), which prevents AGEs formation, inhibited the formation of intracellular Glycer-AGEs and prevented cell death. Among the intracellular Glycer-AGEs that were formed, heat shock cognate 70 (Hsc70) was identified as a GA-modified protein, and its modification reduced the activity of Hsc70. Furthermore, intracellular Glycer-AGEs increased the CRP mRNA concentration.
These results suggest that intracellular Glycer-AGEs play important roles in promoting inflammation and hepatocellular death.
非酒精性脂肪性肝炎（NASH）是代谢综合征的一个特征。糖基化终产物（AGEs）是美拉德反应的产物，该反应导致衰老和某些糖尿病的病理并发症。最近的一项研究表明，甘油醛衍生的 AGEs（Glycer-AGEs）在 NASH 患者的血清中升高。此外，Glycer-AGEs 的免疫组化染色在 NASH 患者的肝脏中显示出强烈的染色。本研究旨在研究细胞内 Glycer-AGEs 对肝癌（Hep3B）细胞的影响。
通过 WST-1 测定法测定细胞活力。Slot blot 和 Western blot 用于检测细胞内 Glycer-AGEs，并通过共聚焦显微镜分析其定位。实时逆转录聚合酶链反应用于定量急性反应反应物 C 反应蛋白（CRP）的 mRNA。
Glycer-AGEs 的前体甘油醛（GA）诱导细胞死亡呈浓度和时间依赖性增加，这与细胞内 Glycer-AGEs 形成增加有关。可防止 AGEs 形成的氨基胍（AG）抑制细胞内 Glycer-AGEs 的形成并防止细胞死亡。在形成的细胞内 Glycer-AGEs 中，热休克同源物 70（Hsc70）被鉴定为 GA 修饰的蛋白质，其修饰降低了 Hsc70 的活性。此外，细胞内 Glycer-AGEs 增加了 CRP mRNA 浓度。
这些结果表明，细胞内 Glycer-AGEs 在促进炎症和肝细胞死亡方面发挥重要作用。
