Vrenken Titia E, Buist-Homan Manon, Kalsbeek Allard Jan, Faber Klaas Nico, Moshage Han
Department of Gastroenterology and Hepatology, University Medical Center Groningen, University of Groningen, 9700 RB Groningen, The Netherlands.
J Hepatol. 2008 Nov;49(5):799-809. doi: 10.1016/j.jhep.2008.07.019. Epub 2008 Aug 22.
BACKGROUND/AIMS: Leflunomide is used in the treatment of autoimmune diseases as an anti-inflammatory agent. Leflunomide and its active metabolite A77 1726 modulate mitogen-activated protein kinases (MAPK), Src kinases, the phosphoinositide-3 kinase (PI3K)/Akt-pathway and nuclear factor (NF)-kappaB activation. Both cell protective and cytotoxic effects of leflunomide have been described. Since leflunomide affects pathways involved in hepatocyte cell survival, we examined the effects of A77 1726 on hepatocyte cell death.
Primary rat hepatocytes were exposed to the bile acid glycochenodeoxycholic acid (GCDCA), the superoxide anion donor menadione, or tumor necrosis factor (TNF) alpha in combination with actinomycin D. Activation of MAP-kinases was determined by Western blot analysis. Apoptosis and necrosis were analyzed by acridine orange staining and caspase activity and Sytox Green staining, respectively.
A77 1726 dose-dependently reduces GCDCA-induced apoptosis and necrosis, but not menadione- or TNFalpha/ActD-induced apoptosis. The hepatoprotective effect of A77 1726 does not involve ERK1/2, p38 or PI3K/Akt activation. A77 1726 does not inhibit NF-kappaB activation in hepatocytes.
Since A77 1726 inhibits bile acid-induced apoptosis and does not sensitize hepatocytes to TNFalpha, our results suggest that A77 1726 could be considered for the treatment of chronic liver diseases accompanied by elevated bile acid levels and inflammation.
背景/目的：来氟米特作为一种抗炎药物用于治疗自身免疫性疾病。来氟米特及其活性代谢产物A77 1726可调节丝裂原活化蛋白激酶（MAPK）、Src激酶、磷酸肌醇-3激酶（PI3K）/Akt信号通路以及核因子（NF）-κB的激活。来氟米特的细胞保护作用和细胞毒性作用均有报道。由于来氟米特影响肝细胞存活相关的信号通路，我们研究了A77 1726对肝细胞死亡的影响。
原代大鼠肝细胞分别暴露于胆汁酸甘氨鹅去氧胆酸（GCDCA）、超氧阴离子供体甲萘醌或肿瘤坏死因子（TNF）α与放线菌素D的组合中。通过蛋白质免疫印迹分析确定MAP激酶的激活情况。分别通过吖啶橙染色、半胱天冬酶活性分析和Sytox Green染色分析细胞凋亡和坏死情况。
A77 1726剂量依赖性地减少GCDCA诱导的细胞凋亡和坏死，但不减少甲萘醌或TNFα/放线菌素D诱导的细胞凋亡。A77 1726的肝保护作用不涉及ERK1/2、p38或PI3K/Akt的激活。A77 1726不抑制肝细胞中NF-κB的激活。
由于A77 1726抑制胆汁酸诱导的细胞凋亡且不使肝细胞对TNFα敏感，我们的结果表明A77 1726可考虑用于治疗伴有胆汁酸水平升高和炎症的慢性肝病。
