Maintenance of bad phosphorylation prevents apoptosis of rat hepatic sinusoidal endothelial cells in vitro and in vivo
Maintenance of bad phosphorylation prevents apoptosis of rat hepatic sinusoidal endothelial cells in vitro and in vivo
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DOI：
10.2353/ajpath.2006.050462
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发表时间：
2006-04-01
影响因子：
6
通讯作者：
Enomoto, K
中科院分区：
文献类型：
--
作者：
Ohi, N;Nishikawa, Y;Enomoto, K
探讨肝窦内皮细胞凋亡的机制。细胞（SEC），我们检查了在培养中的细胞凋亡和缺血再灌注损伤后Bad及其上游信号分子的磷酸化状态。免疫磁珠法分离大鼠SEC，培养2天后，大多数SEC发生凋亡，这与细胞蛋白酪氨酸磷酸化水平降低有关。加入蛋白酪氨酸磷酸酶抑制剂原钒酸盐（OV），可维持细胞蛋白磷酸化，并强烈抑制细胞凋亡。在凋亡过程中，Bad在Ser-112和Ser-136处脱磷酸化，但在OV存在下，Bad的磷酸化状态得以维持。OV激活Akt、细胞外信号调节蛋白激酶和p38促分裂原活化蛋白激酶途径，这些途径参与Bad磷酸化。在不存在OV的情况下，通过RNA干扰耗尽Bad赋予了对凋亡的抗性。OV治疗可减轻缺血再灌注后的肝损伤，并显著抑制SEC凋亡。SEC凋亡在体内与Bad，Akt和细胞外信号调节蛋白激酶的去磷酸化有关，这被OV处理所阻断。我们的数据表明，Bad磷酸化的维持在SEC细胞凋亡的预防中是重要的，OV的抗细胞凋亡特性可能具有治疗效用。
To elucidate the mechanism of apoptosis of liver sinusoidal endothelial. cells (SECs), we examined the phosphorylation status of Bad and its upstream signaling molecules during apoptosis in culture and after ischemia-reperfusion injury. Rat SECs were isolated by the immunomagnetic method, and 2 days after culture, most SECs underwent apoptosis, which was associated with decreased tyrosine phosphorylation of cellular proteins. Addition of orthovanadate (OV), a protein tyrosine phosphatase inhibitor, sustained cellular protein phosphorylation and strongly inhibited apoptosis. Bad was dephosphorylated at Ser-112 and Ser-136 during apoptosis, but the phosphorylation status of Bad was maintained in the presence of OV. OV activated the Akt, extracellular signal-regulated protein kinase, and p38 mitogen-activated protein kinase pathways, which are involved in Bad phosphorylation. in the absence of OV, depletion of Bad by RNA interference conferred resistance to apoptosis. Hepatic injury after ischemia-reperfusion was alleviated by OV treatment, with significant inhibition of SEC apoptosis. SEC apoptosis in vivo was associated with dephosphorylation of Bad, Akt, and extracellular signal-regulated protein kinase, which was blocked by OV treatment. Our data suggest that maintenance of Bad phosphorylation is important in the prevention of SEC apoptosis and that the anti-apoptotic property of OV might have therapeutic utility.