Shen Jian-gang, Zhang Xiao-lan, Huo Xiao-xia
Department of Gastroenterology, Second Hospital of Hebei Medical University, Shijiazhuang, 050000, China.
Zhonghua Gan Zang Bing Za Zhi. 2008 Nov;16(11):849-53.
To investigate the effects of FAK-related non-kinase (FRNK) on the apoptosis of hepatic stellate cells (HSC) in vitro and on the extracellular signal-regulated kinase (ERK) signal transduction pathway.
HSC were stimulated by fibronectin (FN), and then they were transfected with FAK-related non-kinase (FRNK) plasmids mediated by cationic liposome. The apoptosis of FRNK-induced HSC was examined by annexin-V/propidium iodide double-labeled flow cytometry (FCM), gel electrophoresis and transmission electron microscopy. Levels of FRNK, FAK, p-FAK (Tyr397), ERK1 and p-ERK in HSC were assayed by Western blot on the protein level, and by RT-PCR on the mRNA level.
The expression of FRNK was enhanced after FRNK plasmids were transiently transfected into the HSC. The apoptotic rate of the HSC exposed to FRNK plasmids for 48 h was higher than that in the non-FRNK plasmid group (25.37%+/-1.92% vs 9.28%+/-1.05%, P less than 0.01), and was accompanied by a significantly higher activity of caspase-3 both in the protein and in the mRNA levels [(264.17+/-12.60 vs 185.82+/-9.69), P less than 0.01; (4.19+/-0.48 vs 1.07+/-0.27), P less than 0.01]. After exposure of HSC to FRNK plasmids, compared with the non-FRNK plasmid group, the expressions of p-FAK, ERK1 and p-ERK in protein and mRNA levels were lower; on the contrary, compared with the control group, the expressions of p-FAK, ERK1 and p-ERK in the FN group were higher.
The expression of FRNK was enhanced and the phosphorylation of FAK was inhibited after FRNK was transiently transfected into HSC in vitro. FRNK induces apoptosis of HSC. FAK-ERK signal transduction pathway perhaps is involved in the process.
研究黏着斑激酶相关非激酶（FRNK）对体外肝星状细胞（HSC）凋亡及细胞外信号调节激酶（ERK）信号转导通路的影响。
用纤连蛋白（FN）刺激HSC，然后用阳离子脂质体介导转染FRNK质粒。采用膜联蛋白V/碘化丙啶双标记流式细胞术（FCM）、凝胶电泳和透射电镜检测FRNK诱导的HSC凋亡情况。采用蛋白质免疫印迹法在蛋白质水平以及逆转录聚合酶链反应（RT-PCR）在mRNA水平检测HSC中FRNK、黏着斑激酶（FAK）、磷酸化FAK（Tyr397）、ERK1和磷酸化ERK的水平。
将FRNK质粒瞬时转染入HSC后，FRNK的表达增强。暴露于FRNK质粒48小时的HSC凋亡率高于未转染FRNK质粒组（25.37%±1.92%对9.28%±1.05%，P<0.01），且半胱天冬酶-3在蛋白质和mRNA水平的活性均显著更高[（264.17±12.60对185.82±9.69），P<0.01；（4.19±0.48对1.07±0.27），P<0.01]。HSC暴露于FRNK质粒后，与未转染FRNK质粒组相比，p-FAK、ERK1和磷酸化ERK在蛋白质和mRNA水平的表达较低；相反，与对照组相比，FN组中p-FAK、ERK1和磷酸化ERK的表达较高。
体外将FRNK瞬时转染入HSC后，FRNK表达增强，FAK磷酸化受到抑制。FRNK诱导HSC凋亡。FAK-ERK信号转导通路可能参与了这一过程。
