Phosphatase and Tensin Homolog Deleted on Chromosome 10 (PTEN) Inhibition by 4-Hydroxynonenal Leads to Increased Akt Activation in Hepatocytes
Phosphatase and Tensin Homolog Deleted on Chromosome 10 (PTEN) Inhibition by 4-Hydroxynonenal Leads to Increased Akt Activation in Hepatocytes
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DOI：
10.1124/mol.110.069534
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发表时间：
2011-06-01
影响因子：
3.6
通讯作者：
Petersen, Dennis R.
中科院分区：
文献类型：
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作者：
Shearn, Colin T.;Smathers, Rebecca L.;Petersen, Dennis R.
反应性醛如4-羟基壬烯醛（4-HNE）的产生被认为是酒精性肝病病因学中的重要因素。为了了解4-HNE对肝细胞稳态信号通路的影响，评价了由人肝细胞癌细胞系（HepG 2）和原代大鼠肝细胞组成的细胞模型。用亚细胞毒性浓度的4-HNE处理HepG 2细胞和原代肝细胞导致Akt在30分钟内活化，如通过残基Ser 473和Thr 308的磷酸化增加所证明的。磷脂酰肌醇-3，4，5-三磷酸[PtdIns（3，4，5）P-3[rsqb（]）]的定量和随后的免疫细胞化学导致总PtdIns（3，4，5）P-3增加6倍，并在4-HNE处理后增加质膜的免疫染色。用磷脂酰肌醇3激酶（PI 3 K）抑制剂2-（4-morpholinyl）-8-phenyl-4 H-1-benzopyran-4-one（Ly 294002）或蛋白磷酸酶2A（PP 2A）抑制剂冈田酸（okadaic acid）与4-HNE共处理HepG 2细胞，发现Akt的激活机制是PI 3 K依赖的，而PP 2A不依赖。使用生物素酰肼检测，确定HepG 2细胞与4-HNE的孵育导致称为“10号染色体上缺失的磷酸酶和张力蛋白同源物”（PTEN）的脂质磷酸酶的羰基化增加，所述脂质磷酸酶是Akt活化的关键调节剂。HepG 2细胞和重组PTEN中的活性测定均显示，应用4-HNE后，PTEN脂质磷酸酶活性降低。4-HNE处理的重组PTEN的质谱分析检测到单个4-HNE加合物。随后对HepG 2细胞中4-HNE的Akt依赖性生理后果的分析揭示了中性脂质积累的显著增加。这些结果提供了Akt激活的潜在机制和4-HNE在肝细胞中的细胞后果。
The production of reactive aldehydes such as 4-hydroxynonenal (4-HNE) is proposed to be an important factor in the etiology of alcoholic liver disease. To understand the effects of 4-HNE on homeostatic signaling pathways in hepatocytes, cellular models consisting of the human hepatocellular carcinoma cell line (HepG2) and primary rat hepatocytes were evaluated. Treatment of both HepG2 cells and primary hepatocytes with subcytotoxic concentrations of 4-HNE resulted in the activation of Akt within 30 min as demonstrated by increased phosphorylation of residues Ser473 and Thr308. Quantification and subsequent immunocytochemistry of phosphatidylinositol-3,4,5-trisphosphate [PtdIns(3,4,5)P-3[rsqb(]) resulted in a 6-fold increase in total PtdIns(3,4,5)P-3 and increased immunostaining at the plasma membrane after 4-HNE treatment. Cotreatment of HepG2 cells with 4-HNE and the phosphatidylinositol 3-kinase (PI3K) inhibitor 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (Ly294002) or the protein phosphatase 2A (PP2A) inhibitor okadaic acid revealed that the mechanism of activation of Akt is PI3K-dependent and PP2A-independent. Using biotin hydrazide detection, it was established that the incubation of HepG2 cells with 4-HNE resulted in increased carbonylation of the lipid phosphatase known as "phosphatase and tensin homolog deleted on chromosome 10" (PTEN), a key regulator of Akt activation. Activity assays both in HepG2 cells and recombinant PTEN revealed a decrease in PTEN lipid phosphatase activity after 4-HNE application. Mass spectral analysis of 4-HNE-treated recombinant PTEN detected a single 4-HNE adduct. Subsequent analysis of Akt dependent physiological consequences of 4-HNE in HepG2 cells revealed significant increases in the accumulation of neutral lipids. These results provide a potential mechanism of Akt activation and cellular consequences of 4-HNE in hepatocytes.