Department of Gastroenterology, The Second Affiliated Hospital of Xuzhou Medical College (The General Hospital of Xuzhou Coal Mineral Group), Xuzhou, Jiangsu, China.
J Dig Dis. 2009 Nov;10(4):310-4. doi: 10.1111/j.1751-2980.2009.00401.x.
To investigate the cholesterol metabolism and mRNA expression of the relevant genes in cholesterol synthesis of the cultured steatotic hepatocytes model.
A steatotic model of hepatocytes was constructed by adding palmitic acid to the growing L-02 cells. These cells were collected at day 3 and 6, respectively. Cells with the culture solution without palmitic acid added served as the control. The contents of intracellular triglyceride (TG) and total cholesterol (TC) were detected by the analysis kit. The expression of sterol-regulatory element binding protein-2 (SREBP-2) and its target gene hydroxymethylglutaryl CoA reductase (HMGCR) and low-density lipoprotein receptor (LDLR) were measured by RT-PCR.
Hepatocyte steatosis was observed at day 3 and became more intense at day 6. The contents of intracellular TG and TC were increased and the expression of the SREBP-2, HMGCR and LDLR mRNA were upregulated in a time-dependent manner in the model group. Compared with the control group, the content of intracellular TG was higher at both day 3 and 6 (P < 0.05), while the content of intracellular TC was significantly increased only at day 6; The expression of HMGCR and LDLR mRNA was upregulated in steatotic hepatocytes at both day 3 and 6 (P < 0.05), whereas the SREBP-2 mRNA was increased only at day 6 (P < 0.05).
Cholesterol accumulation is probably due to the upregulated expression of the relevant genes in the cholesterol synthesis of the steatotic hepatocytes.
研究培养的脂肪变性肝细胞模型中胆固醇代谢及相关基因在胆固醇合成中的 mRNA 表达。
在生长中的 L-02 细胞中加入软脂酸构建脂肪变性肝细胞模型。分别于第 3 天和第 6 天收集这些细胞。以不加软脂酸的培养液培养的细胞作为对照。采用分析试剂盒检测细胞内甘油三酯（TG）和总胆固醇（TC）的含量。通过 RT-PCR 检测固醇调节元件结合蛋白-2（SREBP-2）及其靶基因羟甲基戊二酰辅酶 A 还原酶（HMGCR）和低密度脂蛋白受体（LDLR）的表达。
第 3 天观察到肝细胞脂肪变性，第 6 天变得更加明显。模型组细胞内 TG 和 TC 含量增加，SREBP-2、HMGCR 和 LDLR mRNA 的表达呈时间依赖性上调。与对照组相比，模型组第 3 天和第 6 天细胞内 TG 含量均升高（P < 0.05），而细胞内 TC 含量仅在第 6 天显著增加；第 3 天和第 6 天脂肪变性肝细胞中 HMGCR 和 LDLR mRNA 的表达上调（P < 0.05），而 SREBP-2 mRNA 仅在第 6 天增加（P < 0.05）。
胆固醇的积累可能是由于脂肪变性肝细胞中胆固醇合成相关基因的表达上调所致。
