黏着斑相关非激酶对肝星状细胞迁移的抑制作用与机制研究
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摘要
肝纤维化实质上是机体对肝损伤的一种修复过程,组织适度修复则肝脏结构恢复正常,若修复过度则组织中沉积大量细胞外间质(extracellular matrix, ECM)成分,形成组织疤痕或肝纤维化。肝星状细胞(hepatic stellate cell, HSC)的活化、增殖、黏附、迁移以及ECM 分泌增加是肝纤维化形成的中心环节。
整合素在肝纤维化的形成过程中表达逐渐增加或被诱导表达,在HSC与ECM 的相互作用中发挥重要桥梁作用,黏着斑激酶(focal adhesion kinase, FAK)是整合素信号途径中连接整合素与下游信号分子的媒介,是胞内多个信号途径的交汇点。以FAK 为中心的整合素信号转导通路除了Ras-MAPK 途径外,还有磷脂酰肌醇-3 激酶(phosphatidylinositol 3-kinase, PI3K), 这些信号分子被激活可以调节细胞骨架组装和某些基因表达,引起细胞表型发生相应反应,包括细胞形态、铺展、迁移、增殖、分化和存活等变化。我们以往研究发现FAK、细胞外信号调节激酶(extracellular signal-regulated kinase, ERK)在肝纤维化形成过程中有动态改变。
黏着斑相关非激酶(focal adhesion related non-kinase , FRNK)是FAK 磷酸化的内源性抑制剂。近年来人们对整合素FAK 信号途径及其在多种细胞黏附与迁移中的意义进行了系列研究,但FAK 信号分子是否参与FN介导的HSC 迁移过程尚缺乏证据。
为此,我们应用体外细胞培养技术,从HSC 和ECM 之间的相互作用着手,应用表达质粒转染HSC,研究了选择性阻断FAK 磷酸化对FN诱导的HSC 黏附、迁移的影响,以及该过程中某些信号转导分子的作用。
实验内容主要包括以下3 部分:
第一部分:大鼠肝纤维化组织FAK、PI3K、ERK 与AP-1 的动态表达
目的:探讨FAK、PI3K、ERK 和转录因子AP-1(c-fos、c-jun)特别是其活化形式磷酸化分子在大鼠肝纤维化形成过程中的作用。
方法:本研究应用大鼠肝纤维化动物模型,了解肝纤维化形成情况;采用Western blot 与RT-PCR 共扩增技术研究肌动蛋白、FAK、p-FAK(Tyr~(397))、
Actually, liver fibrosis is a repairing process of the body towards the injury. The moderate repair may reconstruct the liver normal structure, however, if not limited, the excess deposition of extracellular matrix (ECM) finally leads to a scar formation or liver fibrogenesis. The activation, proliferation, adhesion, migration of HSC and the production of ECM are the central events in the liver fibrogenesis.
During liver fibrogenesis, the expression of integrin increases or is induced to express, just like a bridge connecting hepatic stellate cell (HSC) and ECM. Focal adhesion kinase (FAK) is the medium that connects integrin and the downstream signal molecules in integrin-signal transduction pathway, and is the convergence of many signal pathways. In the integrin-signal transduction pathway which FAK is central, except for the Ras-MAPK pathway, there is also phosphatidylinositol-3-kinase (PI3K) signal molecule. Through self-phosphorylation, FAK can recruit and activate the downstream signal molecules, thus modulate the cell framework and the expression of some genes,at last regulate cellular processes such as cell spreading, migration, proliferation, differentiation and cell survival etc. In our past study, FAK and extracellular signal-regulated kinase (ERK) have been found to change dynamically in liver fibrogenesis.
Focal adhesion related non-kinase (FRNK) acts as an endogenous inhibitor of FAK phosphorylation. In recent years, study has focused on the integrin-FAK signal pathway and the role of it in cell adhesion and migration, but whether FAK takes part in the HSC migration process hasn’t been proved.
So with the help of cell culture in vitro, we set about our research from the interaction of HSC and ECM. We transfected HSC using FRNK expressing plasmid and studied the effects on the adhesion and migration of
HSC after selectively breaking the phosphorylation of FAK and the role of some signal molecules in this process. The experiments contained three parts as below: Part 1: The Dynamic Expression of FAK、PI3K、ERK and AP-1 during Hepatic Fibrogenesis in Rats Objective:To investigate the role of FAK、PI3K、ERK and transcription factor AP-1(c-fos, c-jun), especially their phosphorylated forms in the hepatic fibrogenesis Methods:A rat model of common bile duct ligation (BDL)-induced hepatic fibrosis was used to assess the formation of liver fibrosis. Dynamic expressions of FAK、p-FAK (Tyr~(397))、PI3K、ERK、p-ERK and transcription factor AP-1 (c-fos、c-jun) were evaluated by RT-PCR at mRNA level and by Western blot at protein level, respectively. Results: ①Building of liver fibrosis model: The results of Hematoxylin and eosin staining and Masson's trichrome methods showed that in sham-operated group, the structure of liver lobule was integrin and the array of the liver flank was in order. But in model groups, the connective tissue hyperplasiaed broadly and the collagen fiber deposited around the central vein. The structure of the lobule was disturbed even the pseudo-lobules were formed. ②Western blot analysis showed that in the sham-operated group, the actin was expressed a little, but with the development of liver fibrosis, the expression of it increased in model group. And the expression peak happened in the fourth week, 1.82 times of that of 1 wk and 14.53 times of that of sham-operated group; FAK, the molecular weight is 125 kD, expressed in normal rats, but with the period of making model prolonged, the expression increased 14.9、25.8、29.9 and 31.7 times than that of the sham-operated group. Phosphorylation of Tyr~(397) results in the activation of FAK. There was a little expression of p-FAK (Tyr~(397)) in the sham-operated group, but with the development of liver fibrosis, the expression of p-FAK (Tyr~(397)) increased and reached the peak value at the fourth week, which was 1.68 times of that of the first week and 6.16 times of that of sham-operated group, P<0.01; By RT-PCR analysis, FAK mRNA
appeared in the liver of normal rat, and was upregulated on the 2nd day after bile duct ligation, and the upregulation continued during the liver fibrosis, the expression increased 4.7 times in the fourth week. ③The PI3K, molecular weight 85 kD, was expressed in all groups. Compared with the sham-operated group, PI3K increased 1.20、1.54、1.90 and 7.41 times, and the difference was significant, P<0.05;④With the development of liver fibrosis, the expression of ERK1/2 increased, and in the fourth week, ERK1/2 reached 1.89 times of that of sham-operated group. So that was p-ERK: compared with the sham-operated group, the p-ERK expression increased 1.01, 1.11, 1.18 and 1.46 times, respectively. According to our study, with the fibrosis developing, ERK mRNA increased simultaneously. ⑤Using the RT-PCR assay, c-fos and c-jun mRNA were expressed in sham-operated group. The levels of them were highest in the fourth week and 14.33 and 12.38 times of that of the sham-operated group. Conclusions: In liver fibrogenesis, the expression of FAK、PI3K、ERK and transcription factor activator protein-1(c-fos、c-jun) increased obviously. So the increases of them play a pivotal role in the formation and development of liver fibrosis. Part 2: FRNK inhibits the HSC adhesion and migration Objective: FRNK plasmid was used to transfect HSCs, so as to evaluate the effects of breaking the phosphorylation of FAK on the HSC adhesion and migration. Methods: The adhesive inhibition of FRNK on HSCs was examined by toluidine blue colorimetric assay,and the inhibition of migration of FRNK on HSCs was evaluated by improved Boyden chamber. And the levels of FAK in HSCs were assayed by Western blot on protein level and RT-PCR on mRNA level. Results: ①Western blot analysis showed: the FAK expression increased obviously after cultivated with FN, compared with the control group, the expression increased 92.73%,P<0.01;But there was no significant difference among FN group, FN+liposome group and FN+liposome+empty plamid group,
P>0.05. After transfection by FRNK gene for 48 h, the FAK expression of HSCs reduced significantly. Compared with the empty plasmid group, the expression of FAK(45448.7±3388.3)reduced 92.5%, there was statistic difference between them, P<0.01; The expression of p-FAK(Tyr397)protein (3213.6±1090.1) reduced after being transfected by FRNK, P<0.01;After HSCs transfected by plasmid FRNK for 8 h, FAK mRNA expression showed that there was no difference among FN group, FN+liposome group and FN+liposome+empty plamid group, but in the group transfected by FRNK, FAK mRNA reduced 27.4% than that in the empty-plasmid group.②The inhibition of proliferation by FRNK in HSCs: after being transfected by FRNK plasmid, the proliferation of HSCs was inhibited. The inhibition rates were 72.02%、82.04%、89.14% at 12 h、24 h、48 h, respectively. So FRNK plasmid could dramatically inhibit the proliferation of HSCs in time-dependent manner.③The inhibition of FRNK on HSC adhesion: compared with the control group, the adhesion rate increased in FN group, FN+ liposome group, FN+liposome+empty plamid group and non-FRNK group, but there was no statistic difference among them, P=0.767. After transfected by FRNK for 24 h、48 h the adhesion rate of HSCs reduced 37.56% and 38.85% than that of empty-plasmid group, there was significant difference (P=0.000) and was time-dependent. ④The inhibition of FRNK on HSCs migration: Compared with the control group, the cell number of migrating to the bottom chamber increased in FN group, FN+liposome group, FN+liposome+empty plamid group and non-FRNK empty plasmid group, but there was no statistic difference among them. After being transfected by FRNK, the number of crossing membrane of HSCs reduced 47.8% and 56.4% than that of empty-plasmid group, there was statistic difference and in time-dependent manner. Conclusions: The expression and phosphorylation of FAK in HSCs were inhibited after being transfected by FRNK. And FRNK can inhibit FAK both at protein and at mRNA levels, and inhibit the HSC proliferation, cell adhesion and migration in the same time course and in time-dependent
manner. Part 3:The molecular mechanism of the inhibiting migration of FRNK on HSCs Objective: To explore the molecular mechanism of FRNK’s inhibiting HSCs migration induced by fibronectin. Methods: HSCs were cultured in vitro, and the expression of FAK, PI3K, ERK and transcription factor AP-1(c-fos、c-jun)were assessed using Western blot and RT-PCR assay. Results: ①Western blot showed: FAK expression increased obviously after cultivated with FN, compared with the control group, up to 92.73%,P<0.01;But there was no significant difference among FN group, FN+liposome group and FN+liposome +empty plamid group, P>0.05. After transfected by FRNK gene for 48 h, the expression of FAK reduced obviously in HSCs. Compared with the empty plasmid group, the expression of FAK (45448.7±3388.3) reduced 92.5%, there was statistic difference between them, P<0.01; The expression of p-FAK(Tyr~(397)) protein (3213.6±1090.1) reduced after being transfected by FRNK, P<0.01;After transfected by FRNK for 8 h, we examined the level of FAK mRNA by RT-PCR. The results showed that there was no difference among FN group, FN+liposome group and FN+ liposome+empty plamid group, but in the group transfected by FRNK, FAK mRNA reduced 27.4% than that in the empty-plasmid group. ②Western blot showed: Induced by FN, the expression of PI3K(189.9±8.0)increased much more than that in the control group(165.4±13.0). The increasing rate was 14.8%,P=0.013; But there was no significant difference among FN group, FN+liposome group and FN+liposome+empty plamid group, P>0.05. after being transfected by FRNK for 48 h, the expression of PI3K(86.7±6.4)of HSCs reduced obviously, compared with the empty plasmid group (189.4±11.3), it reduced 54.25%. there was statistic difference between them, P<0.01; The expression of PI3K protein reduced after being transfected by FRNK, P=0.000;After transfected by FRNK for 8 h, we examined the level of PI3K p85 mRNA by RT-PCR. The results showed that there is no difference
整合素在肝纤维化的形成过程中表达逐渐增加或被诱导表达,在HSC与ECM 的相互作用中发挥重要桥梁作用,黏着斑激酶(focal adhesion kinase, FAK)是整合素信号途径中连接整合素与下游信号分子的媒介,是胞内多个信号途径的交汇点。以FAK 为中心的整合素信号转导通路除了Ras-MAPK 途径外,还有磷脂酰肌醇-3 激酶(phosphatidylinositol 3-kinase, PI3K), 这些信号分子被激活可以调节细胞骨架组装和某些基因表达,引起细胞表型发生相应反应,包括细胞形态、铺展、迁移、增殖、分化和存活等变化。我们以往研究发现FAK、细胞外信号调节激酶(extracellular signal-regulated kinase, ERK)在肝纤维化形成过程中有动态改变。
黏着斑相关非激酶(focal adhesion related non-kinase , FRNK)是FAK 磷酸化的内源性抑制剂。近年来人们对整合素FAK 信号途径及其在多种细胞黏附与迁移中的意义进行了系列研究,但FAK 信号分子是否参与FN介导的HSC 迁移过程尚缺乏证据。
为此,我们应用体外细胞培养技术,从HSC 和ECM 之间的相互作用着手,应用表达质粒转染HSC,研究了选择性阻断FAK 磷酸化对FN诱导的HSC 黏附、迁移的影响,以及该过程中某些信号转导分子的作用。
实验内容主要包括以下3 部分:
第一部分:大鼠肝纤维化组织FAK、PI3K、ERK 与AP-1 的动态表达
目的:探讨FAK、PI3K、ERK 和转录因子AP-1(c-fos、c-jun)特别是其活化形式磷酸化分子在大鼠肝纤维化形成过程中的作用。
方法:本研究应用大鼠肝纤维化动物模型,了解肝纤维化形成情况;采用Western blot 与RT-PCR 共扩增技术研究肌动蛋白、FAK、p-FAK(Tyr~(397))、
Actually, liver fibrosis is a repairing process of the body towards the injury. The moderate repair may reconstruct the liver normal structure, however, if not limited, the excess deposition of extracellular matrix (ECM) finally leads to a scar formation or liver fibrogenesis. The activation, proliferation, adhesion, migration of HSC and the production of ECM are the central events in the liver fibrogenesis.
During liver fibrogenesis, the expression of integrin increases or is induced to express, just like a bridge connecting hepatic stellate cell (HSC) and ECM. Focal adhesion kinase (FAK) is the medium that connects integrin and the downstream signal molecules in integrin-signal transduction pathway, and is the convergence of many signal pathways. In the integrin-signal transduction pathway which FAK is central, except for the Ras-MAPK pathway, there is also phosphatidylinositol-3-kinase (PI3K) signal molecule. Through self-phosphorylation, FAK can recruit and activate the downstream signal molecules, thus modulate the cell framework and the expression of some genes,at last regulate cellular processes such as cell spreading, migration, proliferation, differentiation and cell survival etc. In our past study, FAK and extracellular signal-regulated kinase (ERK) have been found to change dynamically in liver fibrogenesis.
Focal adhesion related non-kinase (FRNK) acts as an endogenous inhibitor of FAK phosphorylation. In recent years, study has focused on the integrin-FAK signal pathway and the role of it in cell adhesion and migration, but whether FAK takes part in the HSC migration process hasn’t been proved.
So with the help of cell culture in vitro, we set about our research from the interaction of HSC and ECM. We transfected HSC using FRNK expressing plasmid and studied the effects on the adhesion and migration of
HSC after selectively breaking the phosphorylation of FAK and the role of some signal molecules in this process. The experiments contained three parts as below: Part 1: The Dynamic Expression of FAK、PI3K、ERK and AP-1 during Hepatic Fibrogenesis in Rats Objective:To investigate the role of FAK、PI3K、ERK and transcription factor AP-1(c-fos, c-jun), especially their phosphorylated forms in the hepatic fibrogenesis Methods:A rat model of common bile duct ligation (BDL)-induced hepatic fibrosis was used to assess the formation of liver fibrosis. Dynamic expressions of FAK、p-FAK (Tyr~(397))、PI3K、ERK、p-ERK and transcription factor AP-1 (c-fos、c-jun) were evaluated by RT-PCR at mRNA level and by Western blot at protein level, respectively. Results: ①Building of liver fibrosis model: The results of Hematoxylin and eosin staining and Masson's trichrome methods showed that in sham-operated group, the structure of liver lobule was integrin and the array of the liver flank was in order. But in model groups, the connective tissue hyperplasiaed broadly and the collagen fiber deposited around the central vein. The structure of the lobule was disturbed even the pseudo-lobules were formed. ②Western blot analysis showed that in the sham-operated group, the actin was expressed a little, but with the development of liver fibrosis, the expression of it increased in model group. And the expression peak happened in the fourth week, 1.82 times of that of 1 wk and 14.53 times of that of sham-operated group; FAK, the molecular weight is 125 kD, expressed in normal rats, but with the period of making model prolonged, the expression increased 14.9、25.8、29.9 and 31.7 times than that of the sham-operated group. Phosphorylation of Tyr~(397) results in the activation of FAK. There was a little expression of p-FAK (Tyr~(397)) in the sham-operated group, but with the development of liver fibrosis, the expression of p-FAK (Tyr~(397)) increased and reached the peak value at the fourth week, which was 1.68 times of that of the first week and 6.16 times of that of sham-operated group, P<0.01; By RT-PCR analysis, FAK mRNA
appeared in the liver of normal rat, and was upregulated on the 2nd day after bile duct ligation, and the upregulation continued during the liver fibrosis, the expression increased 4.7 times in the fourth week. ③The PI3K, molecular weight 85 kD, was expressed in all groups. Compared with the sham-operated group, PI3K increased 1.20、1.54、1.90 and 7.41 times, and the difference was significant, P<0.05;④With the development of liver fibrosis, the expression of ERK1/2 increased, and in the fourth week, ERK1/2 reached 1.89 times of that of sham-operated group. So that was p-ERK: compared with the sham-operated group, the p-ERK expression increased 1.01, 1.11, 1.18 and 1.46 times, respectively. According to our study, with the fibrosis developing, ERK mRNA increased simultaneously. ⑤Using the RT-PCR assay, c-fos and c-jun mRNA were expressed in sham-operated group. The levels of them were highest in the fourth week and 14.33 and 12.38 times of that of the sham-operated group. Conclusions: In liver fibrogenesis, the expression of FAK、PI3K、ERK and transcription factor activator protein-1(c-fos、c-jun) increased obviously. So the increases of them play a pivotal role in the formation and development of liver fibrosis. Part 2: FRNK inhibits the HSC adhesion and migration Objective: FRNK plasmid was used to transfect HSCs, so as to evaluate the effects of breaking the phosphorylation of FAK on the HSC adhesion and migration. Methods: The adhesive inhibition of FRNK on HSCs was examined by toluidine blue colorimetric assay,and the inhibition of migration of FRNK on HSCs was evaluated by improved Boyden chamber. And the levels of FAK in HSCs were assayed by Western blot on protein level and RT-PCR on mRNA level. Results: ①Western blot analysis showed: the FAK expression increased obviously after cultivated with FN, compared with the control group, the expression increased 92.73%,P<0.01;But there was no significant difference among FN group, FN+liposome group and FN+liposome+empty plamid group,
P>0.05. After transfection by FRNK gene for 48 h, the FAK expression of HSCs reduced significantly. Compared with the empty plasmid group, the expression of FAK(45448.7±3388.3)reduced 92.5%, there was statistic difference between them, P<0.01; The expression of p-FAK(Tyr397)protein (3213.6±1090.1) reduced after being transfected by FRNK, P<0.01;After HSCs transfected by plasmid FRNK for 8 h, FAK mRNA expression showed that there was no difference among FN group, FN+liposome group and FN+liposome+empty plamid group, but in the group transfected by FRNK, FAK mRNA reduced 27.4% than that in the empty-plasmid group.②The inhibition of proliferation by FRNK in HSCs: after being transfected by FRNK plasmid, the proliferation of HSCs was inhibited. The inhibition rates were 72.02%、82.04%、89.14% at 12 h、24 h、48 h, respectively. So FRNK plasmid could dramatically inhibit the proliferation of HSCs in time-dependent manner.③The inhibition of FRNK on HSC adhesion: compared with the control group, the adhesion rate increased in FN group, FN+ liposome group, FN+liposome+empty plamid group and non-FRNK group, but there was no statistic difference among them, P=0.767. After transfected by FRNK for 24 h、48 h the adhesion rate of HSCs reduced 37.56% and 38.85% than that of empty-plasmid group, there was significant difference (P=0.000) and was time-dependent. ④The inhibition of FRNK on HSCs migration: Compared with the control group, the cell number of migrating to the bottom chamber increased in FN group, FN+liposome group, FN+liposome+empty plamid group and non-FRNK empty plasmid group, but there was no statistic difference among them. After being transfected by FRNK, the number of crossing membrane of HSCs reduced 47.8% and 56.4% than that of empty-plasmid group, there was statistic difference and in time-dependent manner. Conclusions: The expression and phosphorylation of FAK in HSCs were inhibited after being transfected by FRNK. And FRNK can inhibit FAK both at protein and at mRNA levels, and inhibit the HSC proliferation, cell adhesion and migration in the same time course and in time-dependent
manner. Part 3:The molecular mechanism of the inhibiting migration of FRNK on HSCs Objective: To explore the molecular mechanism of FRNK’s inhibiting HSCs migration induced by fibronectin. Methods: HSCs were cultured in vitro, and the expression of FAK, PI3K, ERK and transcription factor AP-1(c-fos、c-jun)were assessed using Western blot and RT-PCR assay. Results: ①Western blot showed: FAK expression increased obviously after cultivated with FN, compared with the control group, up to 92.73%,P<0.01;But there was no significant difference among FN group, FN+liposome group and FN+liposome +empty plamid group, P>0.05. After transfected by FRNK gene for 48 h, the expression of FAK reduced obviously in HSCs. Compared with the empty plasmid group, the expression of FAK (45448.7±3388.3) reduced 92.5%, there was statistic difference between them, P<0.01; The expression of p-FAK(Tyr~(397)) protein (3213.6±1090.1) reduced after being transfected by FRNK, P<0.01;After transfected by FRNK for 8 h, we examined the level of FAK mRNA by RT-PCR. The results showed that there was no difference among FN group, FN+liposome group and FN+ liposome+empty plamid group, but in the group transfected by FRNK, FAK mRNA reduced 27.4% than that in the empty-plasmid group. ②Western blot showed: Induced by FN, the expression of PI3K(189.9±8.0)increased much more than that in the control group(165.4±13.0). The increasing rate was 14.8%,P=0.013; But there was no significant difference among FN group, FN+liposome group and FN+liposome+empty plamid group, P>0.05. after being transfected by FRNK for 48 h, the expression of PI3K(86.7±6.4)of HSCs reduced obviously, compared with the empty plasmid group (189.4±11.3), it reduced 54.25%. there was statistic difference between them, P<0.01; The expression of PI3K protein reduced after being transfected by FRNK, P=0.000;After transfected by FRNK for 8 h, we examined the level of PI3K p85 mRNA by RT-PCR. The results showed that there is no difference
引文
1 姜慧卿, 王占魁, 张晓岚, 等. 丹参单体IH764-3 对胆总管结扎大鼠胶原降解的作用及其机制. 胃肠病学和肝病学杂志,2003,12:336-339
2 苏雅娴. 细胞骨架与细胞运动. 张迺蘅主编. 医学分子生物学. 第1 版, 北京, 北京医科大学出版社, 1999:260-281
3 Sundberg LJ, Galante LM, Bill HM, et al. An endogenous inhibitor of focal adhesion kinase blocks Rac1/JNK but not Ras/ERK-dependent signaling in vascular smooth muscle cells. J Biol Chem, 2003, 278: 29783-29791
4 王波, 王天才, 梁扩寰. 细胞因子Rho A在实验性大鼠肝纤维化形成过程中的作用机制. 中华消化杂志, 2004, 24:414-417
5 Guan JL. Role of focal adhesion kinase in integrin signaling. Int J Biochem Cell Biol, 1997, 29:1085-1096
6 Parsons JT. Focal adhesion kinase: the first ten years. J Cell Sci, 2003, 116: 1409-1416
7 姜慧卿, 张晓岚, 王占魁. 等. 大鼠肝纤维化过程中肝组织黏着斑激酶表达增加. 基础医学与临床, 2005, 25:49-53
8 Marra F, Arrighi MC, Fazi M, et al. Extracellular signal-regulated kinase activation differentially regulates platelet-derived growth factor’s actions in hepatic stellate cells, and is induced by in vivo liver injury in the rat. Hepatology. 1999, 30:951-958
9 张晓岚, 孙泽明, 宋梅, 等. 细胞外信号调节激酶1 在大鼠胆汁性肝纤维化形成过程中的含量变化. 胃肠病学和肝病学杂志, 2003,12:440-443
2 苏雅娴. 细胞骨架与细胞运动. 张迺蘅主编. 医学分子生物学. 第1 版, 北京, 北京医科大学出版社, 1999:260-281
3 Sundberg LJ, Galante LM, Bill HM, et al. An endogenous inhibitor of focal adhesion kinase blocks Rac1/JNK but not Ras/ERK-dependent signaling in vascular smooth muscle cells. J Biol Chem, 2003, 278: 29783-29791
4 王波, 王天才, 梁扩寰. 细胞因子Rho A在实验性大鼠肝纤维化形成过程中的作用机制. 中华消化杂志, 2004, 24:414-417
5 Guan JL. Role of focal adhesion kinase in integrin signaling. Int J Biochem Cell Biol, 1997, 29:1085-1096
6 Parsons JT. Focal adhesion kinase: the first ten years. J Cell Sci, 2003, 116: 1409-1416
7 姜慧卿, 张晓岚, 王占魁. 等. 大鼠肝纤维化过程中肝组织黏着斑激酶表达增加. 基础医学与临床, 2005, 25:49-53
8 Marra F, Arrighi MC, Fazi M, et al. Extracellular signal-regulated kinase activation differentially regulates platelet-derived growth factor’s actions in hepatic stellate cells, and is induced by in vivo liver injury in the rat. Hepatology. 1999, 30:951-958
9 张晓岚, 孙泽明, 宋梅, 等. 细胞外信号调节激酶1 在大鼠胆汁性肝纤维化形成过程中的含量变化. 胃肠病学和肝病学杂志, 2003,12:440-443