Objective: To observe whether liraglutide protects HepG2 cells from lipotoxicity by affecting mitogen-activated protein kinase (MAPKs) pathway. Methods: HepG2 cells were induced with 400μmol/L palmitic acid, and cells were treated with a final concentration of 100 nmol/L liraglutide. In addition, JNK inhibitor (SP600125) and p38 MAPK inhibitor (SB203580) were added in advance, respectively. Apoptosis rate, malondialdehyde (MDA) content, and caspase3 activity were detected. Western blot was used to detect p38 mitogen-activated protein kinase (p38 MAPK), c-jun amino terminal kinase (JNK), cytochrome oxidase P450 2E1 (CYP2E1), glucose regulatory protein 78 (GRP78), activated caspase 3, B cell lymphoma associated Protein X (Bax), B cell lymphoma 2 (Bcl-2), and expression of C/EBP homologous protein (CHOP) protein. LSD or Dunnett's T3 test were used to compare the mean of multiple samples. Results: Palmitic acid increased the phosphorylation of p38 MAPK and JNK in HepG2 cells (P< 0.05). Furthermore, it increased the expression of GRP78, CHOP, CYP2E1, MDA, Bax, caspase3 and apoptosis rate, but inhibited the expression of Bcl-2 (Pvalue < 0.05). SP600125 and SB203580 had inhibited oxidative stress and apoptosis induced by palmitic acid (including CYP2E1, MDA, Bax, Bcl-2, caspase3, CHOP) (P< 0.05). The phosphorylation level of p38 MAPK and JNK was reduced with liraglutide and the expression of apoptosis-related proteins (Bax, Bcl-2, caspase3, CHOP) (P< 0.05) was regulated. There was no significant difference in the effect of liraglutide on apoptotic proteins (Bax, Bcl-2, caspase-3, CHOP) (P> 0.05) after pretreatment with those two inhibitors. Conclusion: Palmitic acid has strong lipotoxicity to HepG2 cells and induces apoptosis. Glucagon-like peptide-1 analogue, liraglutide may improve lipotoxicity of palmitic acid by mediating p38 MAPK and JNK pathways.
目的： 观察利拉鲁肽是否通过影响丝裂原活化蛋白激酶(MAPKs)通路起到保护HepG2细胞拮抗脂毒性的作用。 方法： 用400 μmol/L棕榈酸诱导HepG2细胞，同时予终浓度为100 nmol/L利拉鲁肽处理细胞，此外分别提前加入JNK抑制剂（SP600125）和p38 MAPK抑制剂（SB203580），检测细胞凋亡率、丙二醛（MDA）含量、caspase3活性，蛋白质印迹法检测p38丝裂原活化蛋白激酶（p38 MAPK）、c-jun氨基末端激酶(JNK)、细胞色素氧化酶P450 2E1（CYP2E1）、葡萄糖调节蛋白78（GRP78）、活化的半胱氨酸天冬氨酸蛋白酶3（cleaved caspase3）、B细胞淋巴瘤相关蛋白X（Bax）、B细胞淋巴瘤2（Bcl-2）蛋白、C/EBP同源蛋白（CHOP）的表达。多个样本均数比较采用LSD检验或DunnettT3检验。 结果： 棕榈酸能够增加HepG2细胞p38 MAPK、JNK磷酸化水平（P < 0.05），升高GRP78、CHOP、CYP2E1、MDA、Bax、caspase3的表达和细胞凋亡率，抑制Bcl-2的表达(P值均< 0.05)。SP600125和SB203580能够抑制棕榈酸引起的氧化应激和凋亡（包括CYP2E1、MDA、Bax、Bcl-2、caspase3、CHOP）(P值均< 0.05)；利拉鲁肽能够降低p38 MAPK、JNK磷酸化水平，能够调控细胞凋亡相关蛋白（Bax、Bcl-2、caspase3、CHOP）的表达（P值均< 0.05)；而在两种抑制剂预处理后，利拉鲁肽对细胞凋亡蛋白（Bax、Bcl-2、caspase3、CHOP）的影响无统计学意义(P值均> 0.05)。 结论： 棕榈酸对HepG2细胞具有较强的脂毒性，诱导细胞发生凋亡，胰高血糖素样肽-1类似物利拉鲁肽可能通过介导p38 MAPK和JNK途径改善棕榈酸对细胞的脂毒性损伤。.
Keywords: Apoptosis; Endoplasmic reticulum stress; Liraglutide; Oxidative stress.