扶正化瘀大鼠含药血清对HSC-T6细胞mRNA及miRM表达谱调控研究
|
摘要
背景：
肝纤维化是多种慢性肝病向肝硬化发展的必经过程。肝星状细胞扮演了这一过程的核心角色。近30年来,中医药在肝纤维化防控领域发挥了重要作用。众多中药及复方曾在肝星状细胞特定基因表达、胞内信号通路、细胞增殖及凋亡等领域进行过研究。目前认为中医药抗纤维化具有多靶点,多层次的特点,但常规研究很难反映这一特点。全基因组基因芯片及高通量测序技术是基因研究的最新技术,可以对特定时间的mRNA及miRNA基因表达谱进行全面反映,生物信息学的发展则提供了对大量数据进行系统分析的工具。上述技术的出现为中医药研究提供了有力的工具和空前的发展机遇。
扶正化瘀方(胶囊)是临床常用、疗效稳定的抗肝纤维化药物。该复方也是目前研究最系统的抗肝纤维化中成药之一。利用全基因组芯片及高通量测序技术对该药进行研究有助于揭示中医药抗纤维化的重要机制。
目的：
本研究拟探索扶正化瘀含药血清对HSC-T6细胞mRNA及miRNA表达谱的干预特征,阐明该药物在基因层面抗纤维化的作用机制。
方法：
1参照血清药理学原则,按照文献报道方法为大鼠灌服扶正化瘀溶液及对照溶液,采集大鼠血清制备扶正化瘀含药血清及对照血清。对大鼠HSC-T6细胞系进行体外培养24小时,然后同步使用扶正化瘀大鼠含药/对照血清进行干预24小时,提取细胞总RNA,并使用realtime PCR技术,对Ⅰ、Ⅲ型胶原、α-SMA、TGF-β1的mRNA表达水平进行检测,通过上述效应基因的实际表达水平验证实验系统的稳定性。
2使用Affymetrix公司Rat 2302.0对提取的总RNA进行mRNA表达谱的基因芯片检测,获取mRNA的基因芯片表达谱。利用DAVID网站整合数据库对2倍法筛选的差异基因进行基因本体学(GO)、KEGG信号通路(KEGG pathway)分析,对差异基因的功能富集情况和信号通路富集情况进行描述。
3以上述总RNA为材料,使用Solexa miRNA高通量测序法建立miRNA文库,获取miRNA的表达谱。利用targetscan网站提供的预测数据库,对2倍法筛选的miRNA差异基因进行靶基因预测。对预测获得的靶基因利用DAVID网站进行G0及KEGG信号通路分析,对预测靶基因的功能富集情况和信号通路富集情况进行描述。将分析结果与mRNA表达谱的分析结果进行比对,筛选验证研究方向。
4对筛选出的MAPK信号通路,通过western-blot方法,对其核心蛋白(ERK, JNK,p38)及其磷酸化形式进行研究,明确药物对该通路的调控特点。
5结合mRNA表达谱和miRNA巴基因谱的GO分析结果,MAPK通路调控特点、细胞培养中发现的细胞凋亡现象,进一步确定了细胞凋亡为验证方向。使用annexin V/7aad双染色,流式细胞仪分析平台,对HSC-T6细胞凋亡情况进行研究。
结果：
1 mRNA表达谱检测中,差异基因共有637条,其中334条为已知基因。GO分析发现,这些基因在细胞发育、结构发育、细胞过程调节、细胞粘附、细胞组分机化调控方面具有重要作用。KEGG信号通路分析发现具有显著性差异的为RIG-I-like受体信号通路和MAPK信号通路,其中MAPK信号通路与肝纤维化密切相关,其中Prkcg、TNF、Faslg、Rasa2、Gna12、Mapk8、Illr2、Fgf23、Fgf12、Map3k3、Rasgrp2等11条基因对ERK、JNK、p38三条通路有重要调控作用。
2 miRNA表达谱中共有差异基因75个,通过targetscan网站获得预测靶基因1962个,通过DAVID网站分析发现,预测靶基因在发育过程,生物调节,细胞过程,细胞组分机化,生长,生物粘附等方面有富集；KEGG通路分析提示,基因富集具有显著性差异的通路总计29条,去除与肿瘤相关的通路后共剩余16条通路。其中大部分在肝纤维化领域均有研究,而MAPK通路富集最为显著。通过与mRNA表达谱研究结果的比对,确定了MAPK通路为进一步验证方向。
3通过realtime PCR技术,我们发现4项指标的mRNA表达水平均有下降。其中Ⅰ型胶原mRNA表达下降27%,Ⅲ型胶原mRNA表达下降36%,α-SMA mRNA表达下降43%,TGF-β1 mRNA表达下降23%。研究结果提示在扶正化瘀大鼠血清的干预下,维持HSC-T6细胞持续活化的TGF-β1表达下降,HSC-T6细胞的活化标志物表达受到抑制,Ⅰ、Ⅲ型胶原表达发生下调。
4通过对MAPK下三条通路核心蛋白及其磷酸化形式的研究发现扶正化瘀大鼠血清确实对该通路发生了重要的调控：ERK通路上调(P<0.05),JNK通路下调,p38通路上调(P<0.05)。结合HSC-T6细胞本身为活化细胞的背景,我们预测该通路的变化可能会促进细胞凋亡。
5通过对HSC-T6细胞的annexinV、7aad双染色流式细胞仪研究发现,扶正化瘀大鼠含药血清与对照组血清比较,具有明显促进细胞凋亡的生物学效应,无论早期凋亡(P<0.05)、晚期凋亡(P<0.05)、总凋亡(P<0.05),均具有显著性差异。
结论：
1通过上述系列研究,我们证实扶正化瘀大鼠血清对HSC-T6细胞的mRNA及miRNA表达谱产生了广泛的调控作用。
2对两个表达谱的GO分析提示,差异基因在细胞粘附、发育、凋亡等方面存在富集；
3 KEGG通路分析提示,MAPK通路是重要调控通路。
4研究结果提示niRNA的负性调控作用在其中发挥了重要作用。
5扶正化瘀含药血清可促进HSC-T6细胞凋亡。
6 MAPK通路关键蛋白表达水平提示：该药可上调p38通路,下调JNK通路,这可能是扶正化瘀大鼠血清促进HSC-T6的凋亡的主要机制。
7对肝纤维化关键基因的RealtimePCR检测提示：该药可通过抑制TGF-β1表达,降低Ⅰ、Ⅲ型胶原的产生,抑制细胞本身的持续活化。
BACKGROUND
Liver fibrosis, an early pathologic stage of cirrhosis, is the outcome of many chronic liver diseases. Hepatic stellate cell (HSC) plays a central role in this process. In the recent 3 decades, Chinese medicine was committed lot of researches in this field and considered effective on the regulation of gene expression, intracellular signaling pathways, cell proliferation and cell apopatosis. The regulation feature of Chinese medicine is multi-target and multi-level, which is difficult to be reflected by the regular research.
The genome-wide gene chip and high-throughput sequencing technology are the lastest technologies in the gene research field, which can give a global view of the total mRNA and miRNA gene expression profiles at a special time point. The develepment of bioinfomatics provides the powerful tools for the analysis of huge amounts of data. The above progresses make an unprecedented good chance for the development of Chinese medicine research.
FuZheng HuaYu recipe (capsule) is a commonly used and effective anti-hepatic fibrosis drug. On the while, it is one of the drugs with systematic studies. This study is designed to explore the anti-hepatic fibrosis mechanism of Chines medicine by reseaching the mechanism of FuZheng HuaYu recipe with the technologies of the genome-wide gene chip and high-throughput sequencing.
AIM:
This study is designed to explore the mRNA and miRNA expression profiles features of HSC-T6 cells treated by FuZheng HuaYu (FZHY) rat drug serum and elucidate the anti-hepatic fibrosis mechanism in the level of gene.
METHOD:
1. The FZHY rat drug serum and its control serum were prepared from the rats fed with FZHY solution by gavage, according to the serum pharmacological method. The HSC-T6 cells were cultured in the normal culture solution containing 90% DMEM high glucose medium and 10% fetal bovine serum for the first 24 hour and then treated by the FZHY culture solution (90% DMEM high glucose medium and 10% FZHY rat serum) or the control culture solution (90% DMEM high glucose medium and 10% control rat serum) for the next 24 hours. Then the total RNA of the treated HSC-T6 cells was abstracted to commit the real-time PCR experiment. We detected the mRNA levels ofα-SMA, TGF-β1, collagen I and collagen III, which relating to the liver fibrosis closely, to confirm the stability of our research system.
2. The total RNA was detected by the Affymetrix Rat 2302.0 gene chip to achieve the mRNA gene expression profile. Then the differentially expressed genes with more than 2-fold changing were screened out and annotated by the gene ontology and KEGG signaling pathway with the tools and databases integrated by the website DAVID. By the above analysis, the main functions and signaling pathways enriched with differentially expressed genes were found out.
3. The total RNA was detected by the method of Solexa miRNA High-throughput sequencing to achieve the mRNA gene expression profile. The differentially expressed miRNA with more than 2-fold changing were screened out and used to achieve the target genes based on the prediction database of website TARGETSCAN. Then the target genes were annotated by the gene ontology and KEGG signaling pathway with the tools and databases integrated by the website DAVID. The analysis results of mRNA and target genes were compared to find the clues for the follow-up research.
4. The MAPK signaling pathway screened out by the comparison of the annotation results of both mRNA and miRNA gene expression profiles was further confirmed by the western-blot experiment on the key proteins and their Phosphorylated types including ERK, JNK and p38 to elucidate the regulating details.
5. The apoptosis was strongly suggested by the gene ontology analysis of mRNA gene expression profiles and miRNA target gene profile, the regulation features of MAPK pathway and HSC-T6 culture. We made a hypothesis that the FZHY rat serum can induce the apoptosis of HSC-T6 and made a further observation by the annexin V/7aad double staining with the flow cytometry on the apoptosis.
RESULT:
1. In the study of mRNA gene expression profiles, we found 637 differentially expressed genes with more than 2-fold changing and 334 genes within them researched before. The gene ontology analysis suggested that these genes enriched in the process of developmental process, biological regulation, cellular process and biological adhesion. The KEGG pathway analysis suggested the RIG-I-like receptor pathway and MAPK pathway were significantly regulated (P<0.05). The MAPK pathway had a closely relationship with the liver fibrosis and there were 11 genes regulated in this signaling pathway including Prkcg, TNF, Faslg, Rasa2, Gna12, Mapk8, Illr2, Fgf23, Fgf12, Map3k3 and Rasgrp2. These genes had key role in the ERK, JNK and p38 pathway.
2. In the study of miRNA gene expression profiles, we found 75 differentially expressed genes with more than 2-fold changing. Based on the database of website TARGETSCAN, we achieved 1962 target genes. In the following GO analysis, we found these genes enriched in the biological processes:developmental process, biological regulation, cellular process, cellular component organization, growth and et al. In the KEGG signaling pathway analysis, we found that 29 pathway were regulated. Excepting the pathway relating to tumor, there were 16 signaling pathway left, most of which were proved to relating to liver fibrosis. In the 16 pathways, the MAPK signaling pathway was significantly regulated by the miRNA. After comparing the result with mRNA gene expression profiles, we confirm the MAPK signaling pathway for the following up research.
3. In the experiment of real-time PCR, we found that all the 4 kinds of gene expression levels had been down regulated. The mRNA of collagen I, collagen III,α-SMA and TGF-β1 decreased 27%,36%,43% and 23% separately.
4. In the western-blot study about the MAPK key proteins and its Phosphorylated types, we found that the treatment of FZHY rat serum had an important regulation on the MAPK pathway. The ERK pathway was unregulated (P<0.05), the JNK pathway was downregulated (P>0.05) and p38 pathway was upregulated (P<0.05). Considering that the HSC-T6 is the spontaneously activated cell strain, we suggested the above changing may induce the apoptosis of the cells.
5. In the study of apoptosis by annexinV/7aad double staining with flow cytometry, we found the treatment of FZHY rat serum could induce the cell apoptosis comparing to the control serum. Whether the early stage apoptosis or the later stage apoptosis and the total apoptosis, the differences were significant.
CONCLUSION:
By the above 5 experiments, we proved that:
1. FZHY rat serum had a broad regulation on the mRNA and miRNA expression profiles of HSC-T6 cells.
2. The Gene Ontology analysis on the two profiles suggested that the differentially expressed genes mainly enriched in the cellular adhesion, development and apoptosis.
3. The KEGG pathway analysis suggested that the MAPK pathway was the most important regulation target.
4. The negative regulation of miRNA played an important role in the effection of the drug.
5. The FZHY rat serum could induce the apoptosis of HSC-T6 cells.
6. The regulation on the MAPK pathway embodied on the upregulation of p38 and downregulation of JNK, which was probably the most important way to induce the apoptosis of HSC-T6 cells.
7. FZHY rat serum had a stable effect on decreasing the mRNA expression of TGF-β1, which is the mainly mechanism to inhibit the persistent activation of HSC and decrease the collagen I and collagen III
肝纤维化是多种慢性肝病向肝硬化发展的必经过程。肝星状细胞扮演了这一过程的核心角色。近30年来,中医药在肝纤维化防控领域发挥了重要作用。众多中药及复方曾在肝星状细胞特定基因表达、胞内信号通路、细胞增殖及凋亡等领域进行过研究。目前认为中医药抗纤维化具有多靶点,多层次的特点,但常规研究很难反映这一特点。全基因组基因芯片及高通量测序技术是基因研究的最新技术,可以对特定时间的mRNA及miRNA基因表达谱进行全面反映,生物信息学的发展则提供了对大量数据进行系统分析的工具。上述技术的出现为中医药研究提供了有力的工具和空前的发展机遇。
扶正化瘀方(胶囊)是临床常用、疗效稳定的抗肝纤维化药物。该复方也是目前研究最系统的抗肝纤维化中成药之一。利用全基因组芯片及高通量测序技术对该药进行研究有助于揭示中医药抗纤维化的重要机制。
目的：
本研究拟探索扶正化瘀含药血清对HSC-T6细胞mRNA及miRNA表达谱的干预特征,阐明该药物在基因层面抗纤维化的作用机制。
方法：
1参照血清药理学原则,按照文献报道方法为大鼠灌服扶正化瘀溶液及对照溶液,采集大鼠血清制备扶正化瘀含药血清及对照血清。对大鼠HSC-T6细胞系进行体外培养24小时,然后同步使用扶正化瘀大鼠含药/对照血清进行干预24小时,提取细胞总RNA,并使用realtime PCR技术,对Ⅰ、Ⅲ型胶原、α-SMA、TGF-β1的mRNA表达水平进行检测,通过上述效应基因的实际表达水平验证实验系统的稳定性。
2使用Affymetrix公司Rat 2302.0对提取的总RNA进行mRNA表达谱的基因芯片检测,获取mRNA的基因芯片表达谱。利用DAVID网站整合数据库对2倍法筛选的差异基因进行基因本体学(GO)、KEGG信号通路(KEGG pathway)分析,对差异基因的功能富集情况和信号通路富集情况进行描述。
3以上述总RNA为材料,使用Solexa miRNA高通量测序法建立miRNA文库,获取miRNA的表达谱。利用targetscan网站提供的预测数据库,对2倍法筛选的miRNA差异基因进行靶基因预测。对预测获得的靶基因利用DAVID网站进行G0及KEGG信号通路分析,对预测靶基因的功能富集情况和信号通路富集情况进行描述。将分析结果与mRNA表达谱的分析结果进行比对,筛选验证研究方向。
4对筛选出的MAPK信号通路,通过western-blot方法,对其核心蛋白(ERK, JNK,p38)及其磷酸化形式进行研究,明确药物对该通路的调控特点。
5结合mRNA表达谱和miRNA巴基因谱的GO分析结果,MAPK通路调控特点、细胞培养中发现的细胞凋亡现象,进一步确定了细胞凋亡为验证方向。使用annexin V/7aad双染色,流式细胞仪分析平台,对HSC-T6细胞凋亡情况进行研究。
结果：
1 mRNA表达谱检测中,差异基因共有637条,其中334条为已知基因。GO分析发现,这些基因在细胞发育、结构发育、细胞过程调节、细胞粘附、细胞组分机化调控方面具有重要作用。KEGG信号通路分析发现具有显著性差异的为RIG-I-like受体信号通路和MAPK信号通路,其中MAPK信号通路与肝纤维化密切相关,其中Prkcg、TNF、Faslg、Rasa2、Gna12、Mapk8、Illr2、Fgf23、Fgf12、Map3k3、Rasgrp2等11条基因对ERK、JNK、p38三条通路有重要调控作用。
2 miRNA表达谱中共有差异基因75个,通过targetscan网站获得预测靶基因1962个,通过DAVID网站分析发现,预测靶基因在发育过程,生物调节,细胞过程,细胞组分机化,生长,生物粘附等方面有富集；KEGG通路分析提示,基因富集具有显著性差异的通路总计29条,去除与肿瘤相关的通路后共剩余16条通路。其中大部分在肝纤维化领域均有研究,而MAPK通路富集最为显著。通过与mRNA表达谱研究结果的比对,确定了MAPK通路为进一步验证方向。
3通过realtime PCR技术,我们发现4项指标的mRNA表达水平均有下降。其中Ⅰ型胶原mRNA表达下降27%,Ⅲ型胶原mRNA表达下降36%,α-SMA mRNA表达下降43%,TGF-β1 mRNA表达下降23%。研究结果提示在扶正化瘀大鼠血清的干预下,维持HSC-T6细胞持续活化的TGF-β1表达下降,HSC-T6细胞的活化标志物表达受到抑制,Ⅰ、Ⅲ型胶原表达发生下调。
4通过对MAPK下三条通路核心蛋白及其磷酸化形式的研究发现扶正化瘀大鼠血清确实对该通路发生了重要的调控：ERK通路上调(P<0.05),JNK通路下调,p38通路上调(P<0.05)。结合HSC-T6细胞本身为活化细胞的背景,我们预测该通路的变化可能会促进细胞凋亡。
5通过对HSC-T6细胞的annexinV、7aad双染色流式细胞仪研究发现,扶正化瘀大鼠含药血清与对照组血清比较,具有明显促进细胞凋亡的生物学效应,无论早期凋亡(P<0.05)、晚期凋亡(P<0.05)、总凋亡(P<0.05),均具有显著性差异。
结论：
1通过上述系列研究,我们证实扶正化瘀大鼠血清对HSC-T6细胞的mRNA及miRNA表达谱产生了广泛的调控作用。
2对两个表达谱的GO分析提示,差异基因在细胞粘附、发育、凋亡等方面存在富集；
3 KEGG通路分析提示,MAPK通路是重要调控通路。
4研究结果提示niRNA的负性调控作用在其中发挥了重要作用。
5扶正化瘀含药血清可促进HSC-T6细胞凋亡。
6 MAPK通路关键蛋白表达水平提示：该药可上调p38通路,下调JNK通路,这可能是扶正化瘀大鼠血清促进HSC-T6的凋亡的主要机制。
7对肝纤维化关键基因的RealtimePCR检测提示：该药可通过抑制TGF-β1表达,降低Ⅰ、Ⅲ型胶原的产生,抑制细胞本身的持续活化。
BACKGROUND
Liver fibrosis, an early pathologic stage of cirrhosis, is the outcome of many chronic liver diseases. Hepatic stellate cell (HSC) plays a central role in this process. In the recent 3 decades, Chinese medicine was committed lot of researches in this field and considered effective on the regulation of gene expression, intracellular signaling pathways, cell proliferation and cell apopatosis. The regulation feature of Chinese medicine is multi-target and multi-level, which is difficult to be reflected by the regular research.
The genome-wide gene chip and high-throughput sequencing technology are the lastest technologies in the gene research field, which can give a global view of the total mRNA and miRNA gene expression profiles at a special time point. The develepment of bioinfomatics provides the powerful tools for the analysis of huge amounts of data. The above progresses make an unprecedented good chance for the development of Chinese medicine research.
FuZheng HuaYu recipe (capsule) is a commonly used and effective anti-hepatic fibrosis drug. On the while, it is one of the drugs with systematic studies. This study is designed to explore the anti-hepatic fibrosis mechanism of Chines medicine by reseaching the mechanism of FuZheng HuaYu recipe with the technologies of the genome-wide gene chip and high-throughput sequencing.
AIM:
This study is designed to explore the mRNA and miRNA expression profiles features of HSC-T6 cells treated by FuZheng HuaYu (FZHY) rat drug serum and elucidate the anti-hepatic fibrosis mechanism in the level of gene.
METHOD:
1. The FZHY rat drug serum and its control serum were prepared from the rats fed with FZHY solution by gavage, according to the serum pharmacological method. The HSC-T6 cells were cultured in the normal culture solution containing 90% DMEM high glucose medium and 10% fetal bovine serum for the first 24 hour and then treated by the FZHY culture solution (90% DMEM high glucose medium and 10% FZHY rat serum) or the control culture solution (90% DMEM high glucose medium and 10% control rat serum) for the next 24 hours. Then the total RNA of the treated HSC-T6 cells was abstracted to commit the real-time PCR experiment. We detected the mRNA levels ofα-SMA, TGF-β1, collagen I and collagen III, which relating to the liver fibrosis closely, to confirm the stability of our research system.
2. The total RNA was detected by the Affymetrix Rat 2302.0 gene chip to achieve the mRNA gene expression profile. Then the differentially expressed genes with more than 2-fold changing were screened out and annotated by the gene ontology and KEGG signaling pathway with the tools and databases integrated by the website DAVID. By the above analysis, the main functions and signaling pathways enriched with differentially expressed genes were found out.
3. The total RNA was detected by the method of Solexa miRNA High-throughput sequencing to achieve the mRNA gene expression profile. The differentially expressed miRNA with more than 2-fold changing were screened out and used to achieve the target genes based on the prediction database of website TARGETSCAN. Then the target genes were annotated by the gene ontology and KEGG signaling pathway with the tools and databases integrated by the website DAVID. The analysis results of mRNA and target genes were compared to find the clues for the follow-up research.
4. The MAPK signaling pathway screened out by the comparison of the annotation results of both mRNA and miRNA gene expression profiles was further confirmed by the western-blot experiment on the key proteins and their Phosphorylated types including ERK, JNK and p38 to elucidate the regulating details.
5. The apoptosis was strongly suggested by the gene ontology analysis of mRNA gene expression profiles and miRNA target gene profile, the regulation features of MAPK pathway and HSC-T6 culture. We made a hypothesis that the FZHY rat serum can induce the apoptosis of HSC-T6 and made a further observation by the annexin V/7aad double staining with the flow cytometry on the apoptosis.
RESULT:
1. In the study of mRNA gene expression profiles, we found 637 differentially expressed genes with more than 2-fold changing and 334 genes within them researched before. The gene ontology analysis suggested that these genes enriched in the process of developmental process, biological regulation, cellular process and biological adhesion. The KEGG pathway analysis suggested the RIG-I-like receptor pathway and MAPK pathway were significantly regulated (P<0.05). The MAPK pathway had a closely relationship with the liver fibrosis and there were 11 genes regulated in this signaling pathway including Prkcg, TNF, Faslg, Rasa2, Gna12, Mapk8, Illr2, Fgf23, Fgf12, Map3k3 and Rasgrp2. These genes had key role in the ERK, JNK and p38 pathway.
2. In the study of miRNA gene expression profiles, we found 75 differentially expressed genes with more than 2-fold changing. Based on the database of website TARGETSCAN, we achieved 1962 target genes. In the following GO analysis, we found these genes enriched in the biological processes:developmental process, biological regulation, cellular process, cellular component organization, growth and et al. In the KEGG signaling pathway analysis, we found that 29 pathway were regulated. Excepting the pathway relating to tumor, there were 16 signaling pathway left, most of which were proved to relating to liver fibrosis. In the 16 pathways, the MAPK signaling pathway was significantly regulated by the miRNA. After comparing the result with mRNA gene expression profiles, we confirm the MAPK signaling pathway for the following up research.
3. In the experiment of real-time PCR, we found that all the 4 kinds of gene expression levels had been down regulated. The mRNA of collagen I, collagen III,α-SMA and TGF-β1 decreased 27%,36%,43% and 23% separately.
4. In the western-blot study about the MAPK key proteins and its Phosphorylated types, we found that the treatment of FZHY rat serum had an important regulation on the MAPK pathway. The ERK pathway was unregulated (P<0.05), the JNK pathway was downregulated (P>0.05) and p38 pathway was upregulated (P<0.05). Considering that the HSC-T6 is the spontaneously activated cell strain, we suggested the above changing may induce the apoptosis of the cells.
5. In the study of apoptosis by annexinV/7aad double staining with flow cytometry, we found the treatment of FZHY rat serum could induce the cell apoptosis comparing to the control serum. Whether the early stage apoptosis or the later stage apoptosis and the total apoptosis, the differences were significant.
CONCLUSION:
By the above 5 experiments, we proved that:
1. FZHY rat serum had a broad regulation on the mRNA and miRNA expression profiles of HSC-T6 cells.
2. The Gene Ontology analysis on the two profiles suggested that the differentially expressed genes mainly enriched in the cellular adhesion, development and apoptosis.
3. The KEGG pathway analysis suggested that the MAPK pathway was the most important regulation target.
4. The negative regulation of miRNA played an important role in the effection of the drug.
5. The FZHY rat serum could induce the apoptosis of HSC-T6 cells.
6. The regulation on the MAPK pathway embodied on the upregulation of p38 and downregulation of JNK, which was probably the most important way to induce the apoptosis of HSC-T6 cells.
7. FZHY rat serum had a stable effect on decreasing the mRNA expression of TGF-β1, which is the mainly mechanism to inhibit the persistent activation of HSC and decrease the collagen I and collagen III
引文
1中国中西医结合学会肝病专业委员会.肝纤维化中西医结合诊疗指南.[J]中华肝脏病杂志2006,14(11)：866-870
2姚乃礼.中医药抗肝纤维化研究现状与对策.[J]陕西中医.2004.20(6).54-56
3唐智敏,茹清静,张振鄂,等.肝血瘀阻与肝纤维化关系的临床研究[J].中国中西医结合杂志,1997,17(2)：81.
4霍清萍,张延昭.瘀热互结与肝纤维化关系的临床研究[J].新中医,1999,37(4)：36.
5薛博瑜,顾学兰.肝纤维化的病机认识和辨证论治[J].南京中医药大学学报(自然科学版),2000,17(2)：76.
6贲长恩.中药抗肝纤维化研究的现状和展望[J]北京中医药大学学报.1996,19(5)：2-8.
7钱海青.辨证治疗肝炎后肝纤维化[J].浙江中医杂志,1999,15(8)：558.
8李筠.中医证型与肝炎患者血清肝纤维化指标及肝脏病理关系的研究[J].新中医,1999,31(2)：36-37.
9薛东英.丹参酚酸B对大鼠肝星状细胞中丝裂原激活的蛋白激酶通路的抑制作用.中华肝脏病杂志.2004,12(8)：471-474
10 Liu P, Liu CH, Wang HN, et al. Effect of salvianol ic acid B on collagen production and mitogen—activated protein kinase activity in rat hepatic stellate cellS[J]. Acta Pharmacol Sin,2002,23(8):733-738.
11 Wasser S, Ho JM, Ang HK, et al. Salvia miltiorrhiza reduces experimentally induced hepatic fibrosis in rats[J]. J Hepatol 1998,29(5):760-771.
12颜吉丽,李华,范钰,等.虫草多糖对大鼠肝星状细胞核因子—KB活性和肿瘤坏死因子～Ⅱ表达的影响[J].复旦学报(医学版),2003,30(1)：27-29.
13刘玉侃,沈薇,张霞.虫草菌丝对实验性肝纤维化的防治作用及其机制研究[J].中国新药与临床杂志,2004,23(3)：139-143.
14 Muriel P。Moreno MG. Effects of silymarin and vitamins E and C on liver damage induced by prolonged biliary obstruction in the rat[J]. Basic Clin Pharmacol Toxicol,2004, 94(2):99-104.
15 Lieber CS。Leo MA, Cao Q, et al. Silymarin retards the progression of alcohol— induced hepatic fibrosis in baboons. ilymarin retards the progression of alcohol—induced hepatic fibrosiS in baboons[J]. J Clin Gastroenterol,2003,37(4):336-339.
16王红,陈在忠.川芎嗪对大鼠肝纤维化脂质过氧化的影响[J1.中华肝脏病杂志,2000,8(2)：98.
17吴建红,陕光,张端莲,等.川芎嗪治疗后肝纤维化组织中Ⅰ型胶原平均光密度表 达的比较[J].数理医药学杂志,2006,19(2)：134.
18姬成伟,刘晋敏,李亮成,等.川芎嗪阻抑大鼠肝纤维作用实验研究[J].山西医科大学学报,2003,34(4)：293.
19马红.复方861对大鼠肝星状细胞增殖及胶原合成的影响.中国临床药理学与治疗学杂志.1993,3(3)：172-175.
20申凤俊.DMN肝纤维化模型肝组织Ⅰ、Ⅳ型胶原和TIMP-1 mRNA表达及复方861的干预作用.临床和实验医学杂志.2002,1(3)：150-153
21阴赤贞宏.胆管阻塞性大鼠肝纤维化模型肝组织MMP2mRNA的表达及复方861抑制作用的研究[J].临床肝胆病杂志.2002,18(4)：248-251
22丁惠国.复方861对活化肝星状细胞超微结构及细胞骨架蛋白的影响[J].中华肝脏病杂志.2005.13(7)：553-556
23尹姗姗.复方861治疗慢性乙型肝炎肝纤维化与早期肝硬化的临床研究[J].中华肝脏病杂志.2004.12(8)：467-469
24姜春萌.扶正化癖方对大鼠CCL4损伤肝Kupffer细胞功能的影响.中西医结合肝病杂志.2000,10(5)：26-29
25陶艳艳.肝纤维化中TGF-β1信号转导变化特点与中药干预作用[D].上海中医药大学.2005
26刘成海.基于FN/整合素信号途径探讨扶正化瘀方抑制肝星状细胞活化的作用机制[J].上海中医药.2008,42(1)：3-6
27谭春雨.扶正化瘀方对肝纤维化大鼠肝脏明胶酶、TIMP-2和星状细胞凋亡的影响[J].上海中医药大学学报.2003,17(4)：40-44
28刘成海.二甲基亚硝胺致大鼠脂质过氧化变化与药物干预作用.2001,9(z1)：18-21
29闫秀川.扶正化瘀方影响肝细胞凋亡的抗肝纤维化作用机制与部分有效成分[D].上海中医药大学.2010
30谭春雨.扶正化瘀方对DMN肝纤维化模型鼠肝细胞凋亡的影响[J].上海中医药.2005,39(1)：47-49
31陈倩.基于肝组织微血管病变探讨扶正化瘀方药抗肝纤维化的作用机制[D].上海中医药大学.2009.
32崔红燕.扶正化淤方对四氯化碳肝纤维化大鼠基膜型基质金属蛋白酶活性的影响[J].时珍国医国药.2009,20(8)：2045-2047
33顾杰.扶正化瘀胶囊对肝硬化患者门脉血流动力学的影响.上海中医药杂志.2005,39(11)：31-32
1 Fodor SPA, Read JL, Pirning MC,et al. Light-directed, spetially addressable paraliel chemical synthesis. Science,1991,251:767-773
2 Bataller R,Brenner D A. Liver fibrosis[J]. J Clinl Invest,2005,115:209-218.
3 Rat Genome Sequencing Project Consortium for (Participants are arranged under area of contribution, and then by institution.), Richard A. Gibbs. Et al.Genome sequence of the Brown Norway rat yields insights into mammalian evolution. Nature 428,493-521.
4.Inhibition of Proprotein Convertase SKI-1 Blocks Transcription of Key Extracellular Matr ix Genes Regulating Osteoblastic Mineralization Gorski, J. P. et al. Journal of Biological Chemistry 286(3),1836-49,
5.Multiple cytochrome P-450 genes are concomitantly regulated by vitamin A under steady-state conditions and by retinoic acid during hepatic first-pass metabolism Ross, A. C. et al. Physiological Genomics 43(1),57-67,
6强晖.肝纤维化相关基因的筛选及其功能初步研究[D].第二军医大学.2003.
7邵凤娟.肝再生增强因子对肝脏星状细胞基因表达谱的影响[D].哈尔滨医科大学.2006.
8高艳红.肝星状细胞活化抑制基因筛选及其表观调控和功能[D].上海交通大学.2007.
9 Woo SW, Hwang KI, Chung MW, et al. Gene expression profiles during the activation of rat hepatic stellate cells evaluated by cDNA microarray. Arch Pharm Res.2007 Nov;30(11):1410-8
10 De Minicis S, Seki E, Uchinami H, et al. Gene expression profiles during hepatic stellate cell activation in culture and in vivo. Gastroenterology.2007 May; 132(5):1937-46.
11吴诗品,刘真真,周俊,等.DNA微阵矩技术研究肝纤维化差异基因表达谱变化[J].临床肝胆病杂志,2007,23(4)：258-260
12肖琳,成军,郭江,等.筛选转化生长因子β1刺激肝星状细胞差异表达基因的研究[J].中华肝脏病杂.2008,16(10)：752-756.
13陈新美.基于基因表达谱的肝纤维化治疗药物筛选及相关实验研究[D].南方医科大学.2008
14 Shin HW, Park SY, Lee KB, et al. Transcriptional profiling and Wnt signaling activation in proliferation of human hepatic stellate cells induced by PDGF-BB. Korean J Hepatol. 2009 Dec; 15(4):486-95
15 Xia Y, Chen R, Song Z, et al. Gene expression profiles during activation of cultured rat hepatic stellate cells by tumoral hepatocytes and fetal bovine serum. J Cancer Res Clin Oncol.2010 Feb;136(2):309-21.
16 Xu C, chen x, chang c, et al. Analysis of gene expression profiles of liver stellate cells during liver regeneration in rats. Mol Cells.2011 Jan;31(1):17-23. Epub 2010 Nov 26.
17 ZHONG Li-hua, CHENG Jun, ZHU Li-ying. Differentially expressed genes identified by microarray analysis following leptin treatment of hepatic stellate cells[J]. Chinese Medical Journal,2010, Vol.123 No.6:726-729
18江远.基因芯片技术研究莪术不同成分对HSC-T6细胞基因表达的影响[D].湖北中医学院.2004
19董玲,孙剑勇,杜施霖,等.基因芯片研究甘草酸对大鼠肝星状细胞转化生长因子-p信号通路的影响.中华消化杂志[J].2006,26(9)：594-597
20肖凡,魏红山,李国力,等.应用基因表达谱芯片技术筛选GABA作用下肝星状细胞差异表达基因.世界华人消化杂志[J].2007.15(6)：629-632
21幕永平,刘平,王磊,等.一贯煎影响CCL4大鼠肝硬化形成期肝组织基因表达谱的影响.世界科学技术-中医药现代化.2007,9(3)：43-54
22王晓玲,刘平,胡旭东,等.丹酚酸B抑制CC14诱导大鼠肝纤维化形成的基因表达谱分析[J].上海中医药大学学报.2007,21(2)：50-54
23李国力,魏红山,邱爽,等.注射用血栓通对肝星状细胞基因表达的影响[J].世界华人消化杂志.2008,16(4)：417-421.
24 Shin HW, Park SY, Lee KB, et al. Down-regulation of Wnt signaling during apoptosis of human hepatic stellate cells. Hepatogastroenterology.2009 Jan-Feb;56(89):208-12.
1 Gaca MD, zhou X, IsSaR, Kiriella K, Iredale JP, Benyon PC Base-mentmenbrane-like matrix inhibits proliferation and collagen synthesis by actived rat hepatic cells:evidence for matrix dependent deactivation of stellate cells. Matrix Biology.2003.22:229
2 Iredale JP,Benyon RC, Pickering J, et a 1. Mechanimns of spotaneous resolution of rat liver fibrosis. J Clin Invest,1998,102:538-49
3 Friedman SL. The cellular basis of hepatic fibrosis, Mechanism and treatment strategies. N Engl J Med 1993,328-329
4 Saile B, KnitelT Matthes N, et al. CD95/CD95L-mediated apoptosis of the hepatic stellate cell. A mechanism terminating uncontrolled hepatic cell proliferation during hepatic tissue repair. Am J Pathol,1997,151:1265-72
5 WenRong Gong, Adali Pecci, Sylke Roth,et al.Transformation-dependent susceptibility of rat hepatic stellate cells to apoptosis induced by soluble fas ligand. Hepatology.1998. 28.2.492-502
6 Saile B, Matthes N, Knittel T, et al, Transforming growth factor beta and tumor necrosis factor alpha inhiit both apoptosis and proliferation of activated rat hepatic stellate cells. Hepatology,1999,30:196-202
7 Malhi H, Gores G J, Lemasters J J. Apoptosis and necrosis in the liver:a tale of two deaths? [J]. Hepatology,2006,43(2 Suppl):31-44.
8 Gfeen DR Reed JC. Mitochondria and apoptosis. Science 1998; 281:1309-1312.
9 Grondin M, Marion M, Denizeau F et al. Tributyltin induces apoptotic signaling in hepatocytes through pathways involving the endoplasmic reticulum and mitochondria. Toxicol Appl harmacol 2007; 222:57-68.
10 Fiona Oakley, Nathan Trim, Christothea M et al.Hepatocytes Express Nerve Growth Factor during Liver Injury Evidence for Paracrine Regulation of Hepatic Stellate Cell Apoptosis Am J Pathol.2003 November; 163(5):1849-1858.
11 Guo Y T, Leng X s, Li T. et al. Effect of ligand of peroxisome prolifemtor-activated receptor gamma on the biological characters of hepatic stellate cells[J]. World J Gastroenterol,2005,11(30):4735-9.
12 Myung S J, Yoon J H, Gwak G Y, et al. Wnt signaling enhances the activation and survival of human hepatic stollate cells [J].FEBS Lett.2007,581(16):2954-8.
13 Cheng JH, SheH, Han Y P, et al. Wnt antagonism inhibits hepatic stellate cell activation and liver fibrosis [J]. Am J Physiol Gastrointest Liver Physiol,2008,294(1):G39-49.
14 Ikeda H, Nag&shima K, Yanase M, et al. Involvement of Rho/Rho kinase pathway in regulation of apoptosis in rat hepatic stellate cells[J]. Am J Physiol Gastrointest Liver Physiol,2003,285(5):G880-6.
15 Murphy FR, Issa R, Zhou X, et al. Inhibition of apoptosis of activated hepatic stellate cells by tissue inhibitior of metalloproteinasel is mediated via effects on matrix metalloproteinase inhibition:implications for reversibility of liver fibrosis. J Biol Chem, 2002,277(13):11069-11076.
16 Wang XZ, Zhang SJ, Chen YX, et al. Effects of platelet-derived growth factor and interleukin-10 on Fas/Fas-ligand and Bcl-2/Bax mRNA expression in rat hepatic stellate cells in vitro. World J Gastroenterol.2004 Sep 15;10(18):2706-10.
17 Saile B, Eisenbach C, Dudas J,et al.Interferon-gamma acts proapoptotic on hepatic stellate cells (HSC) and abrogates the antiapoptotic effect of interferon-alpha by an HSP70-dependant pathway.Eur J Cell Biol.2004 Sep;83(9):469-76.
18 Cande C, Cecconi F, Dessen P, Kroemer G. (2002). Apoptosis-inducing factor (AIF): Key to the conserved caspase-independent pathways of cell death. Journal of Cell Science. 115(24),4727-4734.
19. van Gurp M, Festjens N, van Loo G, Saelens X, Vandenabeele P. (2003). Mitochondrial intermembrane proteins in cell death. Biochem Biophys Res Commun.304(3),487-497.
20 Tsujimoto Y. (2003). Cell death regulation by the Bcl-2 protein family in the mitochondria. J Cell Physiol.195(2),158-167.
21 Servida F, Lecis D, Scavullo C et al. Novel second mitochondria-derived activator of caspases (Smac) mimetic compounds sensitize human leukemic cell lines to conventional chemotherapeutic drug-induced and death receptor-mediated apoptosis. Invest New Drugs. 2011 Dec;29(6):1264-75. Epub 2010 Jul 8.
22 SiHF, Li J, nI X, et al. Suppressive effects of letlunomide on leptin-induced collagen I production involved in hepatic stellate cell proliferation[J]. ExpBioI Med,2007,232(3): 427-36.
23 Lij, si hf, La X, et al. Suppressive effects of leflunomide on leptin—induced TIMP-1 production involves on hepatic stellate cell proliferation and apoptosis[J]. EurJPharmacol, 2008,580(1-2):63-9.
24 Kumada M, Kihara S, Ouchi N, et al. Adiponectin specifically increased tissue inhibitor of metalloproteinase-1 through interteukin-10 expression in human macmphages[J]. Circulation,2004,109(17):2046-9.
25彭安邦,张秀梅,丁由.Srivivin在活化的肝星状细胞中的表达及反义寡核苷酸对其凋亡的影响.世界华人消化杂志,2010,18：3742：3731.
26 http://www.cellsignal.com/pathways/apoptosis-signaling.jsp
27尤红,王宝恩,马雪梅.中药复方861抑制感形状细胞NF-κB活性的体外研究[J]中华肝脏病杂志2001,9(2)：73-74.
28谭春雨,谭善忠,蒋健,等.扶正化瘀方对DMN肝纤维化模型鼠肝细胞凋亡的影响[J].上海中医药杂志,2005,39(1)：47-49
29赵英恒,何兴祥.复方利肝冲剂对肝星状细胞NF-κB活性的体外研究.[J]世界华人消化杂志,2002,10(1)：95-96.
30刘晓燕.保肝宁对核转录因子-KB的影响及对肝纤维化的调节作用研究.[D].第一军医大学.2004
31 Xiaoxia Geng.Effect of Danshao Huaxian Capsule(丹芍化纤胶囊)on the Expression of Collagen Ⅰ,Ⅲ,and Caspase-3 in Hepatic Fibrosis Rats.[J]CHINESE JOURNAL OF INTEGRATIVE MEDICINE 2004,10(4):211-213
32耿晓霞,吴亚云,程明亮.丹芍化纤胶囊对体外活化HSCs增殖、凋亡及Fas/FasL途径的影响.[J]中华中医药杂志,2005,20(7)：402-406
33杨勤.丹芍化纤胶囊治疗大鼠肝纤维化后肝星状细胞凋亡及细胞周期的变化.[J]中国病理生理杂志.2005,21(5)：975-977
34江远,张玲,何金洋,等.复方肝毒清对铁超载肝星状细胞活化和凋亡的影响.[J]广州中医药大学学报.2010,27(1)：31-32
35胡爱荣,丁一生,程明亮.汉丹肝乐对免疫性肝纤维化大鼠HSCs增殖、活化及凋亡的影响.[A]第十二次全国中西医结合肝病学术会议论文汇编,2003年
36 Lin HJ, Tseng CP, Lin CF,et al.A Chinese Herbal Decoction, Modified Yi Guan Jian, Induces Apoptosis in Hepatic Stellate Cells through an ROS-Mediated Mitochondrial/Caspase Pathway.Evid Based Complement Alternat Med.2011;2011:459531. Epub 2010 Sep 26.
37 Ji G, Wang L, Liu T.Apoptosis of hepatic stellate cell and hepatocyte and regulatory effects of Qinggan Huoxue Recipe on it in alcoholic liver fibrosis rat.Zhongguo Zhong Xi Yi Jie He Za Zhi.2007,27(5):421-5.
38罗云.丹参诱导大鼠肝星状细胞凋亡及其机理的初步研究.[D]重庆医科大学,2000.
39 Chor SY, Hui AY, To KF,et al.Anti-proliferative and pro-apoptotic effects of herbal medicine on hepatic stellate cell.J Ethnopharmacol.2005 Aug 22;100(1-2):180-6.
40李成浩.草苁蓉乙醇提取物对肝星状细胞增殖及胶原合成的影响.[D]延边大学,2004
41 zu J, Fu Y, chen A. Activation of peroxsome proliferator activated receptor-y contributes to the inhibitory effects of curcuminon rat hepatic stellate cell growth[J]. Am J Physiol Gast rointest Liver Physiol,2003,285:G202-G230
42胡爱荣,丁一生,程明亮.汉丹肝乐对免疫性肝纤维化大鼠HSCs增殖、活化及凋亡的影响.[A]第十二次全国中西医结合肝病学术会议论文汇编,2003年
43韩玲.柴胡总皂苷预防大鼠肝纤维化效果的观察.江西中医学院学报.2008,20(6)：70-71
44赵东硕.丹参单体IH764-3与TNF-α对肝星状细胞增殖与凋亡的影响.[D]河北医科大学,2001
45张晓岚,姜慧卿,刘丽,等.丹参单体IH764-3诱导肝星状细胞凋亡.[J]中华内科杂志.2002,41(9)：616-617
46华海婴,郭旭光,杨波等.丹参酮ⅡA磺酸钠注射液对人肝星状细胞的影响.[J]中药药理与临床2007,23(6)：12-14
47钟显飞,蒋明德,马洪德,等.红景天苷对乙醛刺激的大鼠肝星状细胞凋亡的影响.[J]中药新药与临床药理,2004,15(3)：161-163
48汤丽霞,杨光,谭家驹.积雪草酸诱导大鼠肝星状细胞凋亡[J].中草药,2009,40(增刊)230-232
49 Lin YL, Lin CY, Chi CW,etal.Study on antifibrotic effects of curcumin in rat hepatic stellate cells.Phytother Res.2009 Jul;23(7):927-32.
50 Ikeda H, Nagashima K, Yanase M,et al.The herbal medicine inchin-ko-to (TJ-135) induces apoptosis in cultured rat hepatic stellate cells.Life Sci.2006 Apr 4;78(19):2226-33.
51 Pan TL, Wang PW, Leu YL,et al.Inhibitory effects of Scutellaria baicalensis extract on hepatic stellate cells through inducing G2/M cell cycle arrest and activating ERK-dependent apoptosis via Bax and caspase pathway.J Ethnopharmacol.2012 Feb 15;139(3):829-37.
1 K.F.Cheung, D.W.Ye, Z.F.Yang. et.al. Therapeutic efficacy of Traditional Cinese Medicine 319 recipe on hepatic fibrosis induced by carbon tetrachloride in rats[J]. Journal of Ethnopharmacology 124(2009):142-150.
2刘平,胡义扬,刘成,等.扶正化瘀胶囊干预慢性乙型肝炎肝纤维化作用的多中心临床研究[J].中西医结合学报.2003,1(2)：88-102
3 Ping Liu, Yi-Yang Hu, Cheng Liu, et al. Multicenter clinical study on Fuzhenghuayu capsule against liver fibrosis due to chronic hepatitis B[J].World J Gastroenterol 2005,11(19):2892-2899
4 Lewis BP, Burge CB, Bartel DP. Conserved seed pairing, often flanked by adenosines, indicates that thousands of human genes are micro RNA targets[J]. Cell 2005; 120:15-20
5 Calin GA, Sevignani C, Dumitru CD, Hyslop T,Noch E, Yendamuri S, Shimizu M, Rattan S, Bullrich F, Negrini M, Croce CM. Human microRNA genes are frequently located at fragile sites and genomic regions involved in cancers[J]. Proc Natl Acad Sci U S A 2004; 101:2999-3004
6Jin Hou,LI lin, weiping zhou,et al. Identification of miRNomes in Human Liver and Hepatocellular Carcinoma Reveals miR-199a/b-3p as Therapeutic Target for Hepatocellular Carcinoma[J]. Cancer Cell 2011(19),232-243,
7张锦生Micro-RNAs在肝纤维化发生中的作用[J].世界华人消化杂志2009,17(36)：3671-3674
8 Friedman SL,Lalazar A,Crong L,et al. HSC-T6 cells, an immoralized rat hepatic stellate cell line [J]. Hepatology,1997,26 (4Pt):338A
9 Yoshiki Murakami, Hidenori Toyoda, Masami Tanaka, et al. The Progression of Liver Fibrosis Is Related with Overexpression of the miR-199 and 200 Families. PLoS One.2011; 6(1):e16081.
10 Mody N, Leitch J, Amstrong C, et al.Effects of MAP kinase cascade inhibitors on the MKK5/ERK5 pathway J.FEBS Lett 9 2001 S502 (12):21-24.
11 WU LM, WU XX, SUN Y, et a 1. A novel synthetic oleanolic acid derivative (CPU-112) attenuates liver fibrosis in mice through regulating the function of hepatic stellate cells[J]. J Biomed Sci.2008(2).15:251-259.
12任桂芳,叶立红,黄燕,等.乙型肝炎患者肝组织粘着斑激酶、α-平滑肌肌动蛋白的表达及意义.临床肝胆病杂志,2006(6)：410-412.
13扈彩霞.ROCK-I在大鼠肝纤维化形成和肝星状细胞迁移过程中作用机制的研究[D].河北医科大学.2007.
14牛丽文,曹琦,李俊,等JAK/STAT途径调节瘦素诱导的肝星状细胞Ⅰ型胶原基因的表达[J].中国药理学通报.2007,23(10)：4-45
15 Kang Q, Chen A. Curcumin eliminates oxidized LDL roles in activating hepatic stellate cells by suppressing gene expression of lectin-like oxidized LDL receptor-1. Lab Invest 2009; 89:1275-1290
16 Li W, Zhu C, Chen X, Li Y, Gao R, Wu Q. Pokeweed antiviral protein down-regulates Wnt/β-catenin signalling to attenuate liver fibrogenesis in vitro and in vivo. Dig Liver Dis 2011; 43:559-566
17李文庭,陈西柳,李宜,朱传龙,徐瑗瑗,沈强,高人焘.Wnt抑制因子-1对肝星状细胞活化的影响.肝脏2010；15：338-341
1刘成,蒋春萌,刘平.扶正化瘀方对大鼠肝星状细胞旁分泌活化途径的抑制作用[J].中华消化杂志.2001,21(6)：368-369
2 K.F.Cheung, D.W.Ye, Z.F.Yang, et.al. Therapeutic efficacy of Traditional Cinese Medicine 319 recipe on hepatic fibrosis induced by carbon tetrachloride in rats[J]. Journal of Ethnopharmacology 124(2009):142-150.
3刘成海,宣红萍,陶艳艳,等.基于FN/整合素信号途径探讨扶正化瘀方抑制肝星状细胞活化的作用机制[J].上海中医药杂志.2008,42(1)：3-7
4 Gressner AM.Cytokines and cellular Crosstalk involved in the activation of fat storing cells[J]. J-Hepatol.1995,22(2 suppl):28-36.
5 K.F.Cheung, D.W.Ye, Z.F.Yang. et.al. Therapeutic efficacy of Traditional Cinese Medicine 319 recipe on hepatic fibrosis induced by carbon tetrachloride in rats[J]. Journal of Ethnopharmacology 124(2009):142-150
6方刚,林永茎,翁山耕。等.丹参对大鼠肝星状细胞转化生长因子betal与Ⅰ型胶原mRNA袁达的影响.中国现代医学杂志2002；12(13)：17
7王晓玲,刘平,刘成海,等.含药血清药理法观察扶正化瘀方抗肝纤维化的配伍作用[J].中国实验方剂学杂志.1999,5(6)：22-24
8陆雄,刘平,刘成海,等.扶正化瘀方促进二甲基亚硝胺肝纤维化大鼠肝窦毛细血管化逆转作用[J].中医杂志,2003,2：15-17
9 Friedman SL, Lalazar A, Crong L, et al. HSC-T6 cells, an immortalized rat hepatic stellate cell line. Hepatology,1997,26(4Pt):338A
1李校天,杨书良,王军民,等。丹参对肝星状细胞丝裂原活化蛋白激酶通路的抑制作用[J].解放军医学杂志.2006.31(1)：22-25
2薛冬英,洪嘉禾,徐列明.丹参酚酸B对大鼠肝星状细胞中丝裂原激活的蛋白激酶通路的抑制作用[J].中华肝脏病杂志,2004,12(8)：471：474
3 Carlon V, Dcfranco RM,Caligiuri A,et al.Cell adhesion regulates platelet-derived growth factor-induced MAP kinase and PI-3 kinase activation in stellate cells[J]. Hepatology,2002, 36(3):582
4Takashima T, Kawada N, Maeda N.et al. Pepstatin A attenuates the inhibitory effect of N acetyl L-cysteine on proliferation of hepatic myofibroblast(stellate cells). Eur J Phamacol, 2002,451(3):265
5 Alvaro D,Onori P, Metalli VD et al. Intracellular pathway mediating estrogen induced cholangiocyte proliferation in the rat. Hepatology,2002,36(2):297
6 Schnabl B, Bradh am CA, Bennett BL, et al. TAK1/JNK and p38 have opposite effects on rat hepatic stellate cells [J]. Hepatology,2001,34(5):953-963
7 Varel a-Re y M, Montiel-Du a r te C, Oses-Prieto JA, et al. p38 MAPK mediates the regula tion of alphal (I) procollagen mRNA levels by TNF-alpha and TGF-beta in a cell line of rat hepatic stellate cells(1) [J]. FEBS Lett,2002,528 (1-3):133-138.
8 Cao Q, Mak KM, Lieber CS. DLPC decreases TGF-beta induced collagen mRNA by inhibiting p38 MAPK in hepatic stellat e cells [J]. Am J Physiol Gastrointest Liver Physiol, 2002,283 (5):G1051-G1061.
9 Reeves HL, Dac k CL, Peak M, et al. Stress-activated protein kinases in the activation of rat hepatic stellate cells in culture [J] J Hepatol,2000,32 (3):465-472.
10叶平,杨波,吴晓玲,蒋明德.P38 MAPK信号通路主要功能、常见研究方法及对肝纤维化的作用.世界华人消化杂志2011；19(32)：3353-3358
11 Chen HH, Zhao S, Song JG. TGF-betal suppresses apoptosis via differential regulation of MAP kinases and ceramide production[J]. Cell Death Differ.2003 May; 10(5):516-27.
12 Lin SJ, Lee IT, Chen YH, et al.Salvianolic acid B attenuates MMP-2 and MMP-9 expression in vivo in apolipoprotein-E-deficient mouse aorta and in vitro in LPS-treated human aortic smooth muscle cells[J]. J Cell Biochem.2007,100(2):372-84.
13张文炜.丹参酚酸B盐对人肝星状细胞内TGF-β1 MAPK信号传导通路中MEF2的影响[D].上海中医药大学.2005.
1刘成海,刘成,刘平,等.扶正化瘀方对二甲基亚硝胺大鼠肝纤维化原代贮脂细胞形态及功能的影响[J].中国中西医结合脾胃杂志,1997,5(1)：32-35
2谭春雨,谭善忠,蒋健,等.扶正化瘀方对DMN肝纤维化模型鼠肝细胞凋亡的影响[J].上海中医药杂志,2005,39(1):47-49
3 Varel a-Re y M, Montiel-Du a r te C, Oses-Prieto JA, et al. p38 MAPK mediates the regula tion of alphal (I) procollagen mRNA levels by TNF-alpha and TGF-beta in a cell line of rat hepatic stellate cells(1) [J]. FEBS Lett,2002,528 (1-3):133-138.
4 Cao Q, Mak KM, Lieber CS. DLPC decreases TGF-beta induced collagen mRNA by inhibiting p38 MAPK in hepatic stellat e cells [J]. Am J Physiol Gastrointest Liver Physiol, 2002,283(5):G1051-G1061.
5 Reeves HL, Dac k CL, Peak M, et al. Stress-activated protein kinases in the activation of rat hepatic stellate cells in culture [J] J Ilepatol,2000,32 (3):465-472.
6叶平,杨波,吴晓玲,蒋明德.P38 MAPK信号通路主要功能、常见研究方法及对肝纤维化的作用.世界华人消化杂志2011；19(32)：3353-3358.
7 Donnahoo KK, Shames BD, Harken AH, et al. the role of tumor necrosis factor in renal ischemia-reperfusion injury [J]. J Urol.1999 Jul;162(1):196-203.
2姚乃礼.中医药抗肝纤维化研究现状与对策.[J]陕西中医.2004.20(6).54-56
3唐智敏,茹清静,张振鄂,等.肝血瘀阻与肝纤维化关系的临床研究[J].中国中西医结合杂志,1997,17(2)：81.
4霍清萍,张延昭.瘀热互结与肝纤维化关系的临床研究[J].新中医,1999,37(4)：36.
5薛博瑜,顾学兰.肝纤维化的病机认识和辨证论治[J].南京中医药大学学报(自然科学版),2000,17(2)：76.
6贲长恩.中药抗肝纤维化研究的现状和展望[J]北京中医药大学学报.1996,19(5)：2-8.
7钱海青.辨证治疗肝炎后肝纤维化[J].浙江中医杂志,1999,15(8)：558.
8李筠.中医证型与肝炎患者血清肝纤维化指标及肝脏病理关系的研究[J].新中医,1999,31(2)：36-37.
9薛东英.丹参酚酸B对大鼠肝星状细胞中丝裂原激活的蛋白激酶通路的抑制作用.中华肝脏病杂志.2004,12(8)：471-474
10 Liu P, Liu CH, Wang HN, et al. Effect of salvianol ic acid B on collagen production and mitogen—activated protein kinase activity in rat hepatic stellate cellS[J]. Acta Pharmacol Sin,2002,23(8):733-738.
11 Wasser S, Ho JM, Ang HK, et al. Salvia miltiorrhiza reduces experimentally induced hepatic fibrosis in rats[J]. J Hepatol 1998,29(5):760-771.
12颜吉丽,李华,范钰,等.虫草多糖对大鼠肝星状细胞核因子—KB活性和肿瘤坏死因子～Ⅱ表达的影响[J].复旦学报(医学版),2003,30(1)：27-29.
13刘玉侃,沈薇,张霞.虫草菌丝对实验性肝纤维化的防治作用及其机制研究[J].中国新药与临床杂志,2004,23(3)：139-143.
14 Muriel P。Moreno MG. Effects of silymarin and vitamins E and C on liver damage induced by prolonged biliary obstruction in the rat[J]. Basic Clin Pharmacol Toxicol,2004, 94(2):99-104.
15 Lieber CS。Leo MA, Cao Q, et al. Silymarin retards the progression of alcohol— induced hepatic fibrosis in baboons. ilymarin retards the progression of alcohol—induced hepatic fibrosiS in baboons[J]. J Clin Gastroenterol,2003,37(4):336-339.
16王红,陈在忠.川芎嗪对大鼠肝纤维化脂质过氧化的影响[J1.中华肝脏病杂志,2000,8(2)：98.
17吴建红,陕光,张端莲,等.川芎嗪治疗后肝纤维化组织中Ⅰ型胶原平均光密度表 达的比较[J].数理医药学杂志,2006,19(2)：134.
18姬成伟,刘晋敏,李亮成,等.川芎嗪阻抑大鼠肝纤维作用实验研究[J].山西医科大学学报,2003,34(4)：293.
19马红.复方861对大鼠肝星状细胞增殖及胶原合成的影响.中国临床药理学与治疗学杂志.1993,3(3)：172-175.
20申凤俊.DMN肝纤维化模型肝组织Ⅰ、Ⅳ型胶原和TIMP-1 mRNA表达及复方861的干预作用.临床和实验医学杂志.2002,1(3)：150-153
21阴赤贞宏.胆管阻塞性大鼠肝纤维化模型肝组织MMP2mRNA的表达及复方861抑制作用的研究[J].临床肝胆病杂志.2002,18(4)：248-251
22丁惠国.复方861对活化肝星状细胞超微结构及细胞骨架蛋白的影响[J].中华肝脏病杂志.2005.13(7)：553-556
23尹姗姗.复方861治疗慢性乙型肝炎肝纤维化与早期肝硬化的临床研究[J].中华肝脏病杂志.2004.12(8)：467-469
24姜春萌.扶正化癖方对大鼠CCL4损伤肝Kupffer细胞功能的影响.中西医结合肝病杂志.2000,10(5)：26-29
25陶艳艳.肝纤维化中TGF-β1信号转导变化特点与中药干预作用[D].上海中医药大学.2005
26刘成海.基于FN/整合素信号途径探讨扶正化瘀方抑制肝星状细胞活化的作用机制[J].上海中医药.2008,42(1)：3-6
27谭春雨.扶正化瘀方对肝纤维化大鼠肝脏明胶酶、TIMP-2和星状细胞凋亡的影响[J].上海中医药大学学报.2003,17(4)：40-44
28刘成海.二甲基亚硝胺致大鼠脂质过氧化变化与药物干预作用.2001,9(z1)：18-21
29闫秀川.扶正化瘀方影响肝细胞凋亡的抗肝纤维化作用机制与部分有效成分[D].上海中医药大学.2010
30谭春雨.扶正化瘀方对DMN肝纤维化模型鼠肝细胞凋亡的影响[J].上海中医药.2005,39(1)：47-49
31陈倩.基于肝组织微血管病变探讨扶正化瘀方药抗肝纤维化的作用机制[D].上海中医药大学.2009.
32崔红燕.扶正化淤方对四氯化碳肝纤维化大鼠基膜型基质金属蛋白酶活性的影响[J].时珍国医国药.2009,20(8)：2045-2047
33顾杰.扶正化瘀胶囊对肝硬化患者门脉血流动力学的影响.上海中医药杂志.2005,39(11)：31-32
1 Fodor SPA, Read JL, Pirning MC,et al. Light-directed, spetially addressable paraliel chemical synthesis. Science,1991,251:767-773
2 Bataller R,Brenner D A. Liver fibrosis[J]. J Clinl Invest,2005,115:209-218.
3 Rat Genome Sequencing Project Consortium for (Participants are arranged under area of contribution, and then by institution.), Richard A. Gibbs. Et al.Genome sequence of the Brown Norway rat yields insights into mammalian evolution. Nature 428,493-521.
4.Inhibition of Proprotein Convertase SKI-1 Blocks Transcription of Key Extracellular Matr ix Genes Regulating Osteoblastic Mineralization Gorski, J. P. et al. Journal of Biological Chemistry 286(3),1836-49,
5.Multiple cytochrome P-450 genes are concomitantly regulated by vitamin A under steady-state conditions and by retinoic acid during hepatic first-pass metabolism Ross, A. C. et al. Physiological Genomics 43(1),57-67,
6强晖.肝纤维化相关基因的筛选及其功能初步研究[D].第二军医大学.2003.
7邵凤娟.肝再生增强因子对肝脏星状细胞基因表达谱的影响[D].哈尔滨医科大学.2006.
8高艳红.肝星状细胞活化抑制基因筛选及其表观调控和功能[D].上海交通大学.2007.
9 Woo SW, Hwang KI, Chung MW, et al. Gene expression profiles during the activation of rat hepatic stellate cells evaluated by cDNA microarray. Arch Pharm Res.2007 Nov;30(11):1410-8
10 De Minicis S, Seki E, Uchinami H, et al. Gene expression profiles during hepatic stellate cell activation in culture and in vivo. Gastroenterology.2007 May; 132(5):1937-46.
11吴诗品,刘真真,周俊,等.DNA微阵矩技术研究肝纤维化差异基因表达谱变化[J].临床肝胆病杂志,2007,23(4)：258-260
12肖琳,成军,郭江,等.筛选转化生长因子β1刺激肝星状细胞差异表达基因的研究[J].中华肝脏病杂.2008,16(10)：752-756.
13陈新美.基于基因表达谱的肝纤维化治疗药物筛选及相关实验研究[D].南方医科大学.2008
14 Shin HW, Park SY, Lee KB, et al. Transcriptional profiling and Wnt signaling activation in proliferation of human hepatic stellate cells induced by PDGF-BB. Korean J Hepatol. 2009 Dec; 15(4):486-95
15 Xia Y, Chen R, Song Z, et al. Gene expression profiles during activation of cultured rat hepatic stellate cells by tumoral hepatocytes and fetal bovine serum. J Cancer Res Clin Oncol.2010 Feb;136(2):309-21.
16 Xu C, chen x, chang c, et al. Analysis of gene expression profiles of liver stellate cells during liver regeneration in rats. Mol Cells.2011 Jan;31(1):17-23. Epub 2010 Nov 26.
17 ZHONG Li-hua, CHENG Jun, ZHU Li-ying. Differentially expressed genes identified by microarray analysis following leptin treatment of hepatic stellate cells[J]. Chinese Medical Journal,2010, Vol.123 No.6:726-729
18江远.基因芯片技术研究莪术不同成分对HSC-T6细胞基因表达的影响[D].湖北中医学院.2004
19董玲,孙剑勇,杜施霖,等.基因芯片研究甘草酸对大鼠肝星状细胞转化生长因子-p信号通路的影响.中华消化杂志[J].2006,26(9)：594-597
20肖凡,魏红山,李国力,等.应用基因表达谱芯片技术筛选GABA作用下肝星状细胞差异表达基因.世界华人消化杂志[J].2007.15(6)：629-632
21幕永平,刘平,王磊,等.一贯煎影响CCL4大鼠肝硬化形成期肝组织基因表达谱的影响.世界科学技术-中医药现代化.2007,9(3)：43-54
22王晓玲,刘平,胡旭东,等.丹酚酸B抑制CC14诱导大鼠肝纤维化形成的基因表达谱分析[J].上海中医药大学学报.2007,21(2)：50-54
23李国力,魏红山,邱爽,等.注射用血栓通对肝星状细胞基因表达的影响[J].世界华人消化杂志.2008,16(4)：417-421.
24 Shin HW, Park SY, Lee KB, et al. Down-regulation of Wnt signaling during apoptosis of human hepatic stellate cells. Hepatogastroenterology.2009 Jan-Feb;56(89):208-12.
1 Gaca MD, zhou X, IsSaR, Kiriella K, Iredale JP, Benyon PC Base-mentmenbrane-like matrix inhibits proliferation and collagen synthesis by actived rat hepatic cells:evidence for matrix dependent deactivation of stellate cells. Matrix Biology.2003.22:229
2 Iredale JP,Benyon RC, Pickering J, et a 1. Mechanimns of spotaneous resolution of rat liver fibrosis. J Clin Invest,1998,102:538-49
3 Friedman SL. The cellular basis of hepatic fibrosis, Mechanism and treatment strategies. N Engl J Med 1993,328-329
4 Saile B, KnitelT Matthes N, et al. CD95/CD95L-mediated apoptosis of the hepatic stellate cell. A mechanism terminating uncontrolled hepatic cell proliferation during hepatic tissue repair. Am J Pathol,1997,151:1265-72
5 WenRong Gong, Adali Pecci, Sylke Roth,et al.Transformation-dependent susceptibility of rat hepatic stellate cells to apoptosis induced by soluble fas ligand. Hepatology.1998. 28.2.492-502
6 Saile B, Matthes N, Knittel T, et al, Transforming growth factor beta and tumor necrosis factor alpha inhiit both apoptosis and proliferation of activated rat hepatic stellate cells. Hepatology,1999,30:196-202
7 Malhi H, Gores G J, Lemasters J J. Apoptosis and necrosis in the liver:a tale of two deaths? [J]. Hepatology,2006,43(2 Suppl):31-44.
8 Gfeen DR Reed JC. Mitochondria and apoptosis. Science 1998; 281:1309-1312.
9 Grondin M, Marion M, Denizeau F et al. Tributyltin induces apoptotic signaling in hepatocytes through pathways involving the endoplasmic reticulum and mitochondria. Toxicol Appl harmacol 2007; 222:57-68.
10 Fiona Oakley, Nathan Trim, Christothea M et al.Hepatocytes Express Nerve Growth Factor during Liver Injury Evidence for Paracrine Regulation of Hepatic Stellate Cell Apoptosis Am J Pathol.2003 November; 163(5):1849-1858.
11 Guo Y T, Leng X s, Li T. et al. Effect of ligand of peroxisome prolifemtor-activated receptor gamma on the biological characters of hepatic stellate cells[J]. World J Gastroenterol,2005,11(30):4735-9.
12 Myung S J, Yoon J H, Gwak G Y, et al. Wnt signaling enhances the activation and survival of human hepatic stollate cells [J].FEBS Lett.2007,581(16):2954-8.
13 Cheng JH, SheH, Han Y P, et al. Wnt antagonism inhibits hepatic stellate cell activation and liver fibrosis [J]. Am J Physiol Gastrointest Liver Physiol,2008,294(1):G39-49.
14 Ikeda H, Nag&shima K, Yanase M, et al. Involvement of Rho/Rho kinase pathway in regulation of apoptosis in rat hepatic stellate cells[J]. Am J Physiol Gastrointest Liver Physiol,2003,285(5):G880-6.
15 Murphy FR, Issa R, Zhou X, et al. Inhibition of apoptosis of activated hepatic stellate cells by tissue inhibitior of metalloproteinasel is mediated via effects on matrix metalloproteinase inhibition:implications for reversibility of liver fibrosis. J Biol Chem, 2002,277(13):11069-11076.
16 Wang XZ, Zhang SJ, Chen YX, et al. Effects of platelet-derived growth factor and interleukin-10 on Fas/Fas-ligand and Bcl-2/Bax mRNA expression in rat hepatic stellate cells in vitro. World J Gastroenterol.2004 Sep 15;10(18):2706-10.
17 Saile B, Eisenbach C, Dudas J,et al.Interferon-gamma acts proapoptotic on hepatic stellate cells (HSC) and abrogates the antiapoptotic effect of interferon-alpha by an HSP70-dependant pathway.Eur J Cell Biol.2004 Sep;83(9):469-76.
18 Cande C, Cecconi F, Dessen P, Kroemer G. (2002). Apoptosis-inducing factor (AIF): Key to the conserved caspase-independent pathways of cell death. Journal of Cell Science. 115(24),4727-4734.
19. van Gurp M, Festjens N, van Loo G, Saelens X, Vandenabeele P. (2003). Mitochondrial intermembrane proteins in cell death. Biochem Biophys Res Commun.304(3),487-497.
20 Tsujimoto Y. (2003). Cell death regulation by the Bcl-2 protein family in the mitochondria. J Cell Physiol.195(2),158-167.
21 Servida F, Lecis D, Scavullo C et al. Novel second mitochondria-derived activator of caspases (Smac) mimetic compounds sensitize human leukemic cell lines to conventional chemotherapeutic drug-induced and death receptor-mediated apoptosis. Invest New Drugs. 2011 Dec;29(6):1264-75. Epub 2010 Jul 8.
22 SiHF, Li J, nI X, et al. Suppressive effects of letlunomide on leptin-induced collagen I production involved in hepatic stellate cell proliferation[J]. ExpBioI Med,2007,232(3): 427-36.
23 Lij, si hf, La X, et al. Suppressive effects of leflunomide on leptin—induced TIMP-1 production involves on hepatic stellate cell proliferation and apoptosis[J]. EurJPharmacol, 2008,580(1-2):63-9.
24 Kumada M, Kihara S, Ouchi N, et al. Adiponectin specifically increased tissue inhibitor of metalloproteinase-1 through interteukin-10 expression in human macmphages[J]. Circulation,2004,109(17):2046-9.
25彭安邦,张秀梅,丁由.Srivivin在活化的肝星状细胞中的表达及反义寡核苷酸对其凋亡的影响.世界华人消化杂志,2010,18：3742：3731.
26 http://www.cellsignal.com/pathways/apoptosis-signaling.jsp
27尤红,王宝恩,马雪梅.中药复方861抑制感形状细胞NF-κB活性的体外研究[J]中华肝脏病杂志2001,9(2)：73-74.
28谭春雨,谭善忠,蒋健,等.扶正化瘀方对DMN肝纤维化模型鼠肝细胞凋亡的影响[J].上海中医药杂志,2005,39(1)：47-49
29赵英恒,何兴祥.复方利肝冲剂对肝星状细胞NF-κB活性的体外研究.[J]世界华人消化杂志,2002,10(1)：95-96.
30刘晓燕.保肝宁对核转录因子-KB的影响及对肝纤维化的调节作用研究.[D].第一军医大学.2004
31 Xiaoxia Geng.Effect of Danshao Huaxian Capsule(丹芍化纤胶囊)on the Expression of Collagen Ⅰ,Ⅲ,and Caspase-3 in Hepatic Fibrosis Rats.[J]CHINESE JOURNAL OF INTEGRATIVE MEDICINE 2004,10(4):211-213
32耿晓霞,吴亚云,程明亮.丹芍化纤胶囊对体外活化HSCs增殖、凋亡及Fas/FasL途径的影响.[J]中华中医药杂志,2005,20(7)：402-406
33杨勤.丹芍化纤胶囊治疗大鼠肝纤维化后肝星状细胞凋亡及细胞周期的变化.[J]中国病理生理杂志.2005,21(5)：975-977
34江远,张玲,何金洋,等.复方肝毒清对铁超载肝星状细胞活化和凋亡的影响.[J]广州中医药大学学报.2010,27(1)：31-32
35胡爱荣,丁一生,程明亮.汉丹肝乐对免疫性肝纤维化大鼠HSCs增殖、活化及凋亡的影响.[A]第十二次全国中西医结合肝病学术会议论文汇编,2003年
36 Lin HJ, Tseng CP, Lin CF,et al.A Chinese Herbal Decoction, Modified Yi Guan Jian, Induces Apoptosis in Hepatic Stellate Cells through an ROS-Mediated Mitochondrial/Caspase Pathway.Evid Based Complement Alternat Med.2011;2011:459531. Epub 2010 Sep 26.
37 Ji G, Wang L, Liu T.Apoptosis of hepatic stellate cell and hepatocyte and regulatory effects of Qinggan Huoxue Recipe on it in alcoholic liver fibrosis rat.Zhongguo Zhong Xi Yi Jie He Za Zhi.2007,27(5):421-5.
38罗云.丹参诱导大鼠肝星状细胞凋亡及其机理的初步研究.[D]重庆医科大学,2000.
39 Chor SY, Hui AY, To KF,et al.Anti-proliferative and pro-apoptotic effects of herbal medicine on hepatic stellate cell.J Ethnopharmacol.2005 Aug 22;100(1-2):180-6.
40李成浩.草苁蓉乙醇提取物对肝星状细胞增殖及胶原合成的影响.[D]延边大学,2004
41 zu J, Fu Y, chen A. Activation of peroxsome proliferator activated receptor-y contributes to the inhibitory effects of curcuminon rat hepatic stellate cell growth[J]. Am J Physiol Gast rointest Liver Physiol,2003,285:G202-G230
42胡爱荣,丁一生,程明亮.汉丹肝乐对免疫性肝纤维化大鼠HSCs增殖、活化及凋亡的影响.[A]第十二次全国中西医结合肝病学术会议论文汇编,2003年
43韩玲.柴胡总皂苷预防大鼠肝纤维化效果的观察.江西中医学院学报.2008,20(6)：70-71
44赵东硕.丹参单体IH764-3与TNF-α对肝星状细胞增殖与凋亡的影响.[D]河北医科大学,2001
45张晓岚,姜慧卿,刘丽,等.丹参单体IH764-3诱导肝星状细胞凋亡.[J]中华内科杂志.2002,41(9)：616-617
46华海婴,郭旭光,杨波等.丹参酮ⅡA磺酸钠注射液对人肝星状细胞的影响.[J]中药药理与临床2007,23(6)：12-14
47钟显飞,蒋明德,马洪德,等.红景天苷对乙醛刺激的大鼠肝星状细胞凋亡的影响.[J]中药新药与临床药理,2004,15(3)：161-163
48汤丽霞,杨光,谭家驹.积雪草酸诱导大鼠肝星状细胞凋亡[J].中草药,2009,40(增刊)230-232
49 Lin YL, Lin CY, Chi CW,etal.Study on antifibrotic effects of curcumin in rat hepatic stellate cells.Phytother Res.2009 Jul;23(7):927-32.
50 Ikeda H, Nagashima K, Yanase M,et al.The herbal medicine inchin-ko-to (TJ-135) induces apoptosis in cultured rat hepatic stellate cells.Life Sci.2006 Apr 4;78(19):2226-33.
51 Pan TL, Wang PW, Leu YL,et al.Inhibitory effects of Scutellaria baicalensis extract on hepatic stellate cells through inducing G2/M cell cycle arrest and activating ERK-dependent apoptosis via Bax and caspase pathway.J Ethnopharmacol.2012 Feb 15;139(3):829-37.
1 K.F.Cheung, D.W.Ye, Z.F.Yang. et.al. Therapeutic efficacy of Traditional Cinese Medicine 319 recipe on hepatic fibrosis induced by carbon tetrachloride in rats[J]. Journal of Ethnopharmacology 124(2009):142-150.
2刘平,胡义扬,刘成,等.扶正化瘀胶囊干预慢性乙型肝炎肝纤维化作用的多中心临床研究[J].中西医结合学报.2003,1(2)：88-102
3 Ping Liu, Yi-Yang Hu, Cheng Liu, et al. Multicenter clinical study on Fuzhenghuayu capsule against liver fibrosis due to chronic hepatitis B[J].World J Gastroenterol 2005,11(19):2892-2899
4 Lewis BP, Burge CB, Bartel DP. Conserved seed pairing, often flanked by adenosines, indicates that thousands of human genes are micro RNA targets[J]. Cell 2005; 120:15-20
5 Calin GA, Sevignani C, Dumitru CD, Hyslop T,Noch E, Yendamuri S, Shimizu M, Rattan S, Bullrich F, Negrini M, Croce CM. Human microRNA genes are frequently located at fragile sites and genomic regions involved in cancers[J]. Proc Natl Acad Sci U S A 2004; 101:2999-3004
6Jin Hou,LI lin, weiping zhou,et al. Identification of miRNomes in Human Liver and Hepatocellular Carcinoma Reveals miR-199a/b-3p as Therapeutic Target for Hepatocellular Carcinoma[J]. Cancer Cell 2011(19),232-243,
7张锦生Micro-RNAs在肝纤维化发生中的作用[J].世界华人消化杂志2009,17(36)：3671-3674
8 Friedman SL,Lalazar A,Crong L,et al. HSC-T6 cells, an immoralized rat hepatic stellate cell line [J]. Hepatology,1997,26 (4Pt):338A
9 Yoshiki Murakami, Hidenori Toyoda, Masami Tanaka, et al. The Progression of Liver Fibrosis Is Related with Overexpression of the miR-199 and 200 Families. PLoS One.2011; 6(1):e16081.
10 Mody N, Leitch J, Amstrong C, et al.Effects of MAP kinase cascade inhibitors on the MKK5/ERK5 pathway J.FEBS Lett 9 2001 S502 (12):21-24.
11 WU LM, WU XX, SUN Y, et a 1. A novel synthetic oleanolic acid derivative (CPU-112) attenuates liver fibrosis in mice through regulating the function of hepatic stellate cells[J]. J Biomed Sci.2008(2).15:251-259.
12任桂芳,叶立红,黄燕,等.乙型肝炎患者肝组织粘着斑激酶、α-平滑肌肌动蛋白的表达及意义.临床肝胆病杂志,2006(6)：410-412.
13扈彩霞.ROCK-I在大鼠肝纤维化形成和肝星状细胞迁移过程中作用机制的研究[D].河北医科大学.2007.
14牛丽文,曹琦,李俊,等JAK/STAT途径调节瘦素诱导的肝星状细胞Ⅰ型胶原基因的表达[J].中国药理学通报.2007,23(10)：4-45
15 Kang Q, Chen A. Curcumin eliminates oxidized LDL roles in activating hepatic stellate cells by suppressing gene expression of lectin-like oxidized LDL receptor-1. Lab Invest 2009; 89:1275-1290
16 Li W, Zhu C, Chen X, Li Y, Gao R, Wu Q. Pokeweed antiviral protein down-regulates Wnt/β-catenin signalling to attenuate liver fibrogenesis in vitro and in vivo. Dig Liver Dis 2011; 43:559-566
17李文庭,陈西柳,李宜,朱传龙,徐瑗瑗,沈强,高人焘.Wnt抑制因子-1对肝星状细胞活化的影响.肝脏2010；15：338-341
1刘成,蒋春萌,刘平.扶正化瘀方对大鼠肝星状细胞旁分泌活化途径的抑制作用[J].中华消化杂志.2001,21(6)：368-369
2 K.F.Cheung, D.W.Ye, Z.F.Yang, et.al. Therapeutic efficacy of Traditional Cinese Medicine 319 recipe on hepatic fibrosis induced by carbon tetrachloride in rats[J]. Journal of Ethnopharmacology 124(2009):142-150.
3刘成海,宣红萍,陶艳艳,等.基于FN/整合素信号途径探讨扶正化瘀方抑制肝星状细胞活化的作用机制[J].上海中医药杂志.2008,42(1)：3-7
4 Gressner AM.Cytokines and cellular Crosstalk involved in the activation of fat storing cells[J]. J-Hepatol.1995,22(2 suppl):28-36.
5 K.F.Cheung, D.W.Ye, Z.F.Yang. et.al. Therapeutic efficacy of Traditional Cinese Medicine 319 recipe on hepatic fibrosis induced by carbon tetrachloride in rats[J]. Journal of Ethnopharmacology 124(2009):142-150
6方刚,林永茎,翁山耕。等.丹参对大鼠肝星状细胞转化生长因子betal与Ⅰ型胶原mRNA袁达的影响.中国现代医学杂志2002；12(13)：17
7王晓玲,刘平,刘成海,等.含药血清药理法观察扶正化瘀方抗肝纤维化的配伍作用[J].中国实验方剂学杂志.1999,5(6)：22-24
8陆雄,刘平,刘成海,等.扶正化瘀方促进二甲基亚硝胺肝纤维化大鼠肝窦毛细血管化逆转作用[J].中医杂志,2003,2：15-17
9 Friedman SL, Lalazar A, Crong L, et al. HSC-T6 cells, an immortalized rat hepatic stellate cell line. Hepatology,1997,26(4Pt):338A
1李校天,杨书良,王军民,等。丹参对肝星状细胞丝裂原活化蛋白激酶通路的抑制作用[J].解放军医学杂志.2006.31(1)：22-25
2薛冬英,洪嘉禾,徐列明.丹参酚酸B对大鼠肝星状细胞中丝裂原激活的蛋白激酶通路的抑制作用[J].中华肝脏病杂志,2004,12(8)：471：474
3 Carlon V, Dcfranco RM,Caligiuri A,et al.Cell adhesion regulates platelet-derived growth factor-induced MAP kinase and PI-3 kinase activation in stellate cells[J]. Hepatology,2002, 36(3):582
4Takashima T, Kawada N, Maeda N.et al. Pepstatin A attenuates the inhibitory effect of N acetyl L-cysteine on proliferation of hepatic myofibroblast(stellate cells). Eur J Phamacol, 2002,451(3):265
5 Alvaro D,Onori P, Metalli VD et al. Intracellular pathway mediating estrogen induced cholangiocyte proliferation in the rat. Hepatology,2002,36(2):297
6 Schnabl B, Bradh am CA, Bennett BL, et al. TAK1/JNK and p38 have opposite effects on rat hepatic stellate cells [J]. Hepatology,2001,34(5):953-963
7 Varel a-Re y M, Montiel-Du a r te C, Oses-Prieto JA, et al. p38 MAPK mediates the regula tion of alphal (I) procollagen mRNA levels by TNF-alpha and TGF-beta in a cell line of rat hepatic stellate cells(1) [J]. FEBS Lett,2002,528 (1-3):133-138.
8 Cao Q, Mak KM, Lieber CS. DLPC decreases TGF-beta induced collagen mRNA by inhibiting p38 MAPK in hepatic stellat e cells [J]. Am J Physiol Gastrointest Liver Physiol, 2002,283 (5):G1051-G1061.
9 Reeves HL, Dac k CL, Peak M, et al. Stress-activated protein kinases in the activation of rat hepatic stellate cells in culture [J] J Hepatol,2000,32 (3):465-472.
10叶平,杨波,吴晓玲,蒋明德.P38 MAPK信号通路主要功能、常见研究方法及对肝纤维化的作用.世界华人消化杂志2011；19(32)：3353-3358
11 Chen HH, Zhao S, Song JG. TGF-betal suppresses apoptosis via differential regulation of MAP kinases and ceramide production[J]. Cell Death Differ.2003 May; 10(5):516-27.
12 Lin SJ, Lee IT, Chen YH, et al.Salvianolic acid B attenuates MMP-2 and MMP-9 expression in vivo in apolipoprotein-E-deficient mouse aorta and in vitro in LPS-treated human aortic smooth muscle cells[J]. J Cell Biochem.2007,100(2):372-84.
13张文炜.丹参酚酸B盐对人肝星状细胞内TGF-β1 MAPK信号传导通路中MEF2的影响[D].上海中医药大学.2005.
1刘成海,刘成,刘平,等.扶正化瘀方对二甲基亚硝胺大鼠肝纤维化原代贮脂细胞形态及功能的影响[J].中国中西医结合脾胃杂志,1997,5(1)：32-35
2谭春雨,谭善忠,蒋健,等.扶正化瘀方对DMN肝纤维化模型鼠肝细胞凋亡的影响[J].上海中医药杂志,2005,39(1):47-49
3 Varel a-Re y M, Montiel-Du a r te C, Oses-Prieto JA, et al. p38 MAPK mediates the regula tion of alphal (I) procollagen mRNA levels by TNF-alpha and TGF-beta in a cell line of rat hepatic stellate cells(1) [J]. FEBS Lett,2002,528 (1-3):133-138.
4 Cao Q, Mak KM, Lieber CS. DLPC decreases TGF-beta induced collagen mRNA by inhibiting p38 MAPK in hepatic stellat e cells [J]. Am J Physiol Gastrointest Liver Physiol, 2002,283(5):G1051-G1061.
5 Reeves HL, Dac k CL, Peak M, et al. Stress-activated protein kinases in the activation of rat hepatic stellate cells in culture [J] J Ilepatol,2000,32 (3):465-472.
6叶平,杨波,吴晓玲,蒋明德.P38 MAPK信号通路主要功能、常见研究方法及对肝纤维化的作用.世界华人消化杂志2011；19(32)：3353-3358.
7 Donnahoo KK, Shames BD, Harken AH, et al. the role of tumor necrosis factor in renal ischemia-reperfusion injury [J]. J Urol.1999 Jul;162(1):196-203.