虫草多糖体外诱导大鼠骨髓间充质干细胞分化为肝细胞的研究
|
摘要
我国是病毒性肝炎的高发区,尤其是在农村及经济不发达地区,其发病率高达10-15%,由病毒性肝炎导致的重型肝炎或肝衰竭,其死亡率高达50-70%或更高,由慢性肝炎引起的肝纤维化或肝硬化,严重影响了患者的生存质量。
肝纤维化进一步可发展至肝硬化,甚或终末期肝病阶段,发展至终末期肝病则缺乏广泛可行的有效治疗手段,尽管肝移植可作为治疗肝衰竭和肝硬化失代偿期的有效手段,但肝移植存在费用昂贵,手术复杂,术后可能发生严重的排斥反应,生存率不高等缺点,尤其是供体肝缺乏,使得这一治疗措施难以广泛推广。
骨髓间充质干细胞(MSCs)是一群具有多向分化潜能的干细胞,在特定的诱导条件下,可以分化为多种造血以外的组织,如成骨细胞、神经元细胞、脂肪细胞、成纤维细胞、心肌细胞、肝细胞等。MSCs来源广泛,取材容易,便于自体移植,且在体外长期培养过程中,MSCs始终保持其多向分化的潜能,遗传背景相当稳定,体外基因转染率高,并能稳定高效地表达外源基因,故体内植入反应弱,是一种理想的组织工程种子细胞,具有治疗急慢性肝衰竭的潜能。因此,MSCs向肝系细胞的定向分化已受到学者们的广泛关注。
1理论研究
现代医学对MSCs了诱导分化为肝细胞的研究取得了一定的进展,但是单纯的MSCs进行在体移植,进入体内很容易被扩散或蛋白酶水解,能达到坏死肝细胞的MSCs不多,其肝细胞的分化率不仅很低,实际应用前景有限;而体外诱导分化则是在化学制剂的条件下实现,其肝细胞的分化率也很低,而且其临床安全性和长期有效性还难以实现。因此,如何促进MSCs向肝细胞完全意义上的转化,以便更有效改善病变肝组织的结构和功能将成为未来相关研究的重中之重。
中药对MSCs诱导作用的研究起步较晚,近年来,不少研究机构证明中药对MSCs的定向分化有调控作用,并且有比传统的西医学方法高的诱导率。已有学者成功运用中药有效成分诱导MSCs分化为神经细胞、心肌细胞、脂肪细胞、成骨细胞等,目前国内外还没有关于应用中药单体有效成分诱导MSCs分化为肝细胞的研究报道,我们在众多的中药中,选用临床上较常用的治疗肝病的名贵中药冬虫夏草的有效成分虫草多糖来进行研究。现代中药药理研究表明,虫草多糖对肝细胞的损害及肝纤维化有较好的保护作用。而MSCs具有多种多分化潜能,根据既往利用中药有效成分诱导MSCs定向分化的研究成果,理论上相信虫草多糖在适当的条件下可以诱导MSCs分化为肝细胞,研究探索中药诱导MSCs分化为肝细胞,不仅将为应用中医药治疗急性肝衰竭提供新的理论依据,而且由于中药固有的安全性和长期有效性,可以克服目前现代医学研究中存在的某些困境,因此有可能与细胞移植治疗急性肝衰竭、肝硬化失代偿期的方法结合,带来广阔的应用前景,所以积极开展中药诱导MSCs分化为肝细胞的研究有重要意义。
随着中医药的临床实践的不断深入,人们从中药中发现了可以治疗肝纤维的有效药物及其组分或成分,经过临床及实验得到了验证,显示中医药在抗肝纤维化治疗中的优势及潜力。冬虫夏草是我国传统滋补中药,有“保肺,益肾,补精髓,止血化痰,已劳咳,治膈症皆良”之功效,现代研究发现了其具有多种药理作用,拓展了其应用范围,已有体外实验表明虫草制剂可减轻实验型肝纤维化大鼠模型的肝纤维化指标,保护肝细胞,本研究使用虫草菌丝制剂(百令胶囊)来治疗乙肝后肝纤维化患者,观察其临床疗效,并初步探讨虫草制剂治疗肝纤维化的机制,为临床应用提供更多的科学依据。
2实验研究
2.1实验一虫草多糖体外诱导大鼠MSCs分化为肝细胞的研究
目的:探讨虫草多糖能否体外诱导大鼠MSCs分化为肝细胞及其方法条件。方法:取成年Wister大鼠骨髓,采用贴壁法进行培养,流式细胞仪法检测细胞表面抗原CD34和CD44。对MSCs分组进行诱导,虫草组:用虫草多糖进行诱导,终末浓度为0.15mg/ml;阳性对照组:肝细胞生长因子(HGF)和表皮生长因子(EGF)联合诱导,浓度分别为20ng/ml和10ng/ml;空白对照组仅用基本培养基。倒置显微镜下连续观察细胞形态变化及增殖情况。于培养的7天、14天、21天、28天以免疫组化法检测肝细胞特异性标记AFP、CK18、ALB的表达,PAS法进行糖原染色。结果:原代培养的MSCs 24h小时后部分细胞贴壁,48h时大部分细胞贴壁,一周后细胞体积增大,多见长梭形、多角形细胞。细胞增殖明显增快,9天左右铺满瓶壁,胰酶消化传代后细胞形态多为梭形,较前均匀,部分多角形,增殖更快,成集落式生长,一般7天可铺满瓶壁90%。连续传代,P4代后,细胞形态变化不大。分离纯化的P3代MSCs检测CD34为阴性表达,CD44为阳性。对诱导分化后的细胞行免疫组化鉴定:虫草组及阳性对照组7天时出现AFP表达,14天时表达增强,28天时表达减弱;CK18阳性对照组7天时出现表达,14天时表达增强。虫草组则14天时出现表达,而后持续。ALB 7天时两组表达阴性,14天时出现阳性;对照组的AFP阳性率高于虫草组。糖原染色两组14天时出现阳性结果,28天时表达减弱。空白组结果皆为阴性。结论:虫草多糖可以体外诱导大鼠MSCs分化为类肝细胞。
2.2实验二:百令胶囊治疗肝炎后肝纤维化临床疗效观察目的:探索治疗肝纤维化的有效药物。观察虫草菌丝制剂(百令胶囊)对慢性乙肝、早期肝硬化患者肝纤维化的疗效。方法:选择广州中医药大学及广东省中医院2008年1月-2008年12月份住院及门诊治疗后长期随访的慢性活动性肝炎及早期肝硬化患者36例。随机分为对照组和治疗组,两组各18例,全部组别给予肝泰乐0.2g,3次/日、维生素C2.0,口服,3次/日,甘利欣150mg+GS 250ml静滴2周,后改为甘草酸二胺胶囊,2粒,口服,3次/日等基础护肝药物。治疗组在对照组基础上给予百令胶囊(虫草菌丝体干粉,中美浙江华东制药有限公司生产)0.2g/粒,4粒/次,3次/天,口服。观察患者症状改善及血清生化学指标情况,3个月为一疗程。一个疗程后,评定疗效。结果:从临床疗效看,治疗组显效5例,有效11例,无效2例,总有效率为89%;对照组显效2例,有效10例,无效6例,总有效率为67%,两组总有效率比较有统计学意义(P＜0.05),治疗组的疗效优于对照组。两组治疗后,患者症状皆有改善,但治疗组的改善更为显著,与治疗前对比,差异有显著性(P＜0.05);对照组与治疗前相比差异无统计学意义。在肝功能检测方面两组自身治疗前后比较都有显著性差异(P＜0.05),治疗后两组肝功指标比较差异无统计学意义(P＞0.05)。两组治疗后,肝纤维化指标HA、PCⅢ、LN、Ⅳ-C皆有所改善,组内前后对比有显著差异(P＜0.05);治疗组的效果更为优异,与对照组比较差异有统计学意义(P＜0.05)。结论:百令胶囊(虫草菌丝体干粉)可有效治疗肝纤维化及早期肝硬化,保护肝细胞,改善患者的肝功能及HA、LN、PCⅢ、Ⅳ-C血清学指标。
China is a region of high incidence of viral hepatitis its incidence rate is as high as 10-15%,especially in rural and underdeveloped areas of the economy.Serious hepatitis or liver failure caused by viral hepatitis,its mortality rate as high as 50-70%or higher.Hepatic cirrhosis and hepatic fibrosis caused by chronic hepatitis,seriously affected patients in the quality of life.
Hepatic fibrosis can further develope cirrhosis of liver,even final stage liver disease. Lack of effective therapeutic tool in the stage,although liver transplantation can be used as the effective means of treating liver failure and cirrhosis,but it has several drawbacks, Expensive,complex surgery,may occur severe rejection after surgery and the survival rate is not high.the lack of donor liver is a big problem.Those restrictions make it difficult to promote this therapeutic tool.
Mesenchymal stem cells(MSCs)is a group of multi-differentiation potential of stem cells.It can differentiate into a variety of out the hemopoietic tissue in specific conditions. for example,osteoblast,neuron cell,adipocyte,fibroblast,cardiac muscle cell,hepatocyte etc.
MSCs can be obtained from different organizations of the body,and can be autografted conveniently.In the course of long-term culture in vitro,MSCS has always been to maintain its multi-directional differentiation potential,its genetic background is relatively stable in vitro,it has a high rate of gene transfection and can express foreign genes stable and efficient.Therefore,when MSCs is implanted in the body,the weak response to implan in vivo is one of the features.So MSCS is an ideal seed cells for tissue engineering,wih the potential of treating acute and chronic liver failure.MSCs can directed to differentiate into liver cells,which has been widespread concerned by scholars.
1.Theoretical research
Modern medicine has made some progress in MSCs differentiate into liver cells.But MSCs can easily diffused or hydrolysed by enzymatic for pure MSCs transplantation in vivo,the MSCs that can achieve hepatocyte is not much,and it differentiation rate is low,so its prospects is lmited for practical application.The differentiation in vitro is realized under conditions of chemical praeparatum,the differentiation rate of it is low too.And its clinical safety and long-term effectiveness is also difficult to achieve,therefore it's the most important how to promote the MSCS into hepatocytes fully and more effectively to improve the structure and functions of liver organizations.
The study was started relatively late that make use of chinese medicine induce MSCs differentiate.Many research institutions have proved chinese medicine the differentiation regulation to MSCs in recent years,and wih higher induce rate than Western medicine method.It were reported that scholars applied active ingredients of Chinese medicine had been successful induced MSCs differentiate into nerve cells、adipocyte,cadiocyte, fibroblast and hepatocyte etl.There are no reports that someone apply monomer composition of traditional Chinese medicine to induce MSCs differentiate into liver cells at home and abroad.Chinese caterpillar fungus is a commonly Chinese medicine used to treat liver disease in clinical,We chose its active ingredients Chinese caterpillar fungus polysaccharide to study.Pharmacological studies show that Cordyceps polysaccharide:it has protective effect to liver damage and liver fibrosis.MSCs has a wide range of differentiate potential,According to oriented research results of applying chinese medicine active ingredients to induce differentiation of MSCs,we can believe theoretically that cordyceps polysaccharide can induce MSCs differentiate into liver cells in the right conditions.Research on traditional Chinese medicine induced MSCs to differentiate into liver cells,not only provide a new theoretical basis for applying chinese medicine to treat acute liver failure,but also can overcome some dilemma in modern medical research due to the inherent safety and long-term effectiveness of chinese medicine.It may be related to the methods that treat acute liver failure and cirrhosis by cell transplantation,Bring about broad application prospects.So it is important to carry out chinese medicine induce MSCS to differentiate into liver cells.
Along with the deepening of clinical practice of chinese medicine,people had been found some effective drugs and compositions from Chinese medicine that can treat hepatic fibrosis.Clinical and Experimental have been verificated those effects,this showed advantages and potential of chinese medicine in the treatment of hepatic fibrosis.Chinese caterpillar fungus is a traditional chinese tonic medicine,it can protect the lungs and beneficial to the kidneys,enrich the essence,relieve cough and reduce sputum,treat chronic cough and swallowing disorders.Modern research has found a variety of pharmacological effects of it,expanded its scope of application.Experiments have been showed that cordyceps can reduce indicators of experimental liver fibrosis rat model about liver fibrosis, protect liver cells.In this study,we made use of the preparations of Cordyceps sinensis(Bailing Capsule)to treat liver fibrosis with hepatitis B,observe the clinical efficacy and explore mechanisms for the treatment of liver fibrosis.Provide more scientific basis for the clinical application of it.
2.Experimental Study
ExperimentⅠ:Cordyceps polysaccharide induces differentiation of adult rat MSCs into a hepatocyte lineage in vitro
Objective:To research the possibility of the rat mesenchymal stem cells(MSCs)to differentiate into hepatocyte-like with Cordyceps polysaccharide in vitro.Methods:Bone marrow in the femurs of Wistar rat was collected by flushing under sterile condition.MSCs were separated and cultured according to the direct anchoring method,identified corpuscular surface antigen of CD34 and CD44 by using flow cytometric analys.MSCs were divided into 3 groups to induce differentiate.Cordyceps group:induced with Cordyceps polysaccharide.The final concentration of it was 0.15mg/ml.Masccline Control group:Induced by HGF and EGF,the concentration of each was 20ng/ml and 10ng/ml. Blank group:only cultivated with basis medium.The morphological changes and proliferation of MSCSs were observed by phase-contrast microscope.On day 7,14,21 and 28,AFP,CK18 and ALB of MSCSs of each group were examined,the expressions of them were detected with immunohistochemistry technique,staining for glycogen with PAS method.Results:After 24h in primary culture,MSCs adhered to the plastic surface,most cells adhered surface,cell volume increase after a week.The cells spindle-shaped and polygonal,cell proliferation significantly faster,the cells covered culture flask about 9 days. Subcultured after trypsin digestion the cells morphology are most of the spindle patterns more homogeneous than before,proliferated even faster,the cells into a colony-type growth.7 days can covered with 90%surface of culture flask.Continuous passage.Little change in cell morphology after p4.MSCs collected from the femora of Wister rats expressed antigens of CD44,but not CD34.Immunocytochemical analysis for AFP showed positive staining reaction on day7,in Cordyceps group and Masccline Control group, strengthen on day 14,then weaken on day 28.Positive staining reaction on 14 day.Masceline Control group was higher than Cordyceps group in ratio of masccline for AFP.On 14 day both group show positive staining reaction in staining for glycogen of PAS,then weaken on day 28.The blank group all showed positive staining reaction. Conclusion:MSCs induced by cordyceps polysaccharide in vitro can differentiate into hepatocyte-like cells.
ExperimentⅡ:Bailing capsule to treat hepatic fibrosis after hepatitis clinical observation
Objective:To explore the effective drug of treating hepatic fibrosis.Observe cordyceps sinensis preparation(Bailing Capsule)for hepatic fibrosis and early liver fibrosis clinical efficacy in patients.Methods:Choose chronic active hepatitis and early liver cirrhosis 36 cases of patients,which cases come from the First Affiliated Hospital of Guangzhou University of TCM and the Guangdong Provincial TCM Hospital since January 2008 to December 2008.The 36 cases randomly divided into control group and treatment group.18 cases of each group.Two groups all gave glucurolactone 0.2g,3 times/d,po;Vitamin C 2g, 3 times/d,po;Diammonium glycyrrhizinate 150mg+GS 250ml ivgtt in 2 weeks,1 times/day,replaced by diammonium Glycyrrhizinate capsule after 2 weeks,2 pills,po,3 times/day;Treatment group gave Bailing Capsule.Treatment group gave Bailing Capsule(Cordyceps Mycelium Powder,East China Pharmaceutical Co,Ltd, Zhejiang,China and the United States)in addition than the control group,0.2g / tablets,4 tablets/times,3 times/d,po.Observed symptom improvement and serum biochemical indicators in patients,3 months for a course,after a course of treatment,assess of efficacy.
Results:See from the Clinical Efficacy 5 cases of treatment group markedly,11 cases of effective,2 cases of invalid,the total effective rate was 89%;2 cases of the control group markedly,10 cases of effective,6 cases of invalid,the total effective rate was 67%,The total efficiency of the two groups was statistically significant(P＜0.05),Efficacy of the treatment group was superior to the control group.The patients had improvement in symptoms in two groups after treatment,however,the improvement of the treatment group was more significantly,Compared with before treatment,there was a significant difference (P＜0.05),The control group compared with before treatment there was no significant difference(P＞0.05).Comparison of the two groups own before and after the treatment has its significant difference in liver function testing(P＜0.05).The two groups was no significant difference in indicators of liver function after treatment(P＞0.05).Indicators of hepatic fibrosis HA、PCⅢ、LN、Ⅳ-C all were improved in two groups after treatment, Comparison before and after treatment in group were significantly different(P＜0.05),the effect of the treatment group is more superior,Compared with the control group were statistically significant differences.Conclusions:Bailing Capsule(cordyceps sinensis preparation)can treat liver fibrosis and early cirrhosis,protects liver cells,improve liver function and serum indicators of patients with hepatic fibrosis.
肝纤维化进一步可发展至肝硬化,甚或终末期肝病阶段,发展至终末期肝病则缺乏广泛可行的有效治疗手段,尽管肝移植可作为治疗肝衰竭和肝硬化失代偿期的有效手段,但肝移植存在费用昂贵,手术复杂,术后可能发生严重的排斥反应,生存率不高等缺点,尤其是供体肝缺乏,使得这一治疗措施难以广泛推广。
骨髓间充质干细胞(MSCs)是一群具有多向分化潜能的干细胞,在特定的诱导条件下,可以分化为多种造血以外的组织,如成骨细胞、神经元细胞、脂肪细胞、成纤维细胞、心肌细胞、肝细胞等。MSCs来源广泛,取材容易,便于自体移植,且在体外长期培养过程中,MSCs始终保持其多向分化的潜能,遗传背景相当稳定,体外基因转染率高,并能稳定高效地表达外源基因,故体内植入反应弱,是一种理想的组织工程种子细胞,具有治疗急慢性肝衰竭的潜能。因此,MSCs向肝系细胞的定向分化已受到学者们的广泛关注。
1理论研究
现代医学对MSCs了诱导分化为肝细胞的研究取得了一定的进展,但是单纯的MSCs进行在体移植,进入体内很容易被扩散或蛋白酶水解,能达到坏死肝细胞的MSCs不多,其肝细胞的分化率不仅很低,实际应用前景有限;而体外诱导分化则是在化学制剂的条件下实现,其肝细胞的分化率也很低,而且其临床安全性和长期有效性还难以实现。因此,如何促进MSCs向肝细胞完全意义上的转化,以便更有效改善病变肝组织的结构和功能将成为未来相关研究的重中之重。
中药对MSCs诱导作用的研究起步较晚,近年来,不少研究机构证明中药对MSCs的定向分化有调控作用,并且有比传统的西医学方法高的诱导率。已有学者成功运用中药有效成分诱导MSCs分化为神经细胞、心肌细胞、脂肪细胞、成骨细胞等,目前国内外还没有关于应用中药单体有效成分诱导MSCs分化为肝细胞的研究报道,我们在众多的中药中,选用临床上较常用的治疗肝病的名贵中药冬虫夏草的有效成分虫草多糖来进行研究。现代中药药理研究表明,虫草多糖对肝细胞的损害及肝纤维化有较好的保护作用。而MSCs具有多种多分化潜能,根据既往利用中药有效成分诱导MSCs定向分化的研究成果,理论上相信虫草多糖在适当的条件下可以诱导MSCs分化为肝细胞,研究探索中药诱导MSCs分化为肝细胞,不仅将为应用中医药治疗急性肝衰竭提供新的理论依据,而且由于中药固有的安全性和长期有效性,可以克服目前现代医学研究中存在的某些困境,因此有可能与细胞移植治疗急性肝衰竭、肝硬化失代偿期的方法结合,带来广阔的应用前景,所以积极开展中药诱导MSCs分化为肝细胞的研究有重要意义。
随着中医药的临床实践的不断深入,人们从中药中发现了可以治疗肝纤维的有效药物及其组分或成分,经过临床及实验得到了验证,显示中医药在抗肝纤维化治疗中的优势及潜力。冬虫夏草是我国传统滋补中药,有“保肺,益肾,补精髓,止血化痰,已劳咳,治膈症皆良”之功效,现代研究发现了其具有多种药理作用,拓展了其应用范围,已有体外实验表明虫草制剂可减轻实验型肝纤维化大鼠模型的肝纤维化指标,保护肝细胞,本研究使用虫草菌丝制剂(百令胶囊)来治疗乙肝后肝纤维化患者,观察其临床疗效,并初步探讨虫草制剂治疗肝纤维化的机制,为临床应用提供更多的科学依据。
2实验研究
2.1实验一虫草多糖体外诱导大鼠MSCs分化为肝细胞的研究
目的:探讨虫草多糖能否体外诱导大鼠MSCs分化为肝细胞及其方法条件。方法:取成年Wister大鼠骨髓,采用贴壁法进行培养,流式细胞仪法检测细胞表面抗原CD34和CD44。对MSCs分组进行诱导,虫草组:用虫草多糖进行诱导,终末浓度为0.15mg/ml;阳性对照组:肝细胞生长因子(HGF)和表皮生长因子(EGF)联合诱导,浓度分别为20ng/ml和10ng/ml;空白对照组仅用基本培养基。倒置显微镜下连续观察细胞形态变化及增殖情况。于培养的7天、14天、21天、28天以免疫组化法检测肝细胞特异性标记AFP、CK18、ALB的表达,PAS法进行糖原染色。结果:原代培养的MSCs 24h小时后部分细胞贴壁,48h时大部分细胞贴壁,一周后细胞体积增大,多见长梭形、多角形细胞。细胞增殖明显增快,9天左右铺满瓶壁,胰酶消化传代后细胞形态多为梭形,较前均匀,部分多角形,增殖更快,成集落式生长,一般7天可铺满瓶壁90%。连续传代,P4代后,细胞形态变化不大。分离纯化的P3代MSCs检测CD34为阴性表达,CD44为阳性。对诱导分化后的细胞行免疫组化鉴定:虫草组及阳性对照组7天时出现AFP表达,14天时表达增强,28天时表达减弱;CK18阳性对照组7天时出现表达,14天时表达增强。虫草组则14天时出现表达,而后持续。ALB 7天时两组表达阴性,14天时出现阳性;对照组的AFP阳性率高于虫草组。糖原染色两组14天时出现阳性结果,28天时表达减弱。空白组结果皆为阴性。结论:虫草多糖可以体外诱导大鼠MSCs分化为类肝细胞。
2.2实验二:百令胶囊治疗肝炎后肝纤维化临床疗效观察目的:探索治疗肝纤维化的有效药物。观察虫草菌丝制剂(百令胶囊)对慢性乙肝、早期肝硬化患者肝纤维化的疗效。方法:选择广州中医药大学及广东省中医院2008年1月-2008年12月份住院及门诊治疗后长期随访的慢性活动性肝炎及早期肝硬化患者36例。随机分为对照组和治疗组,两组各18例,全部组别给予肝泰乐0.2g,3次/日、维生素C2.0,口服,3次/日,甘利欣150mg+GS 250ml静滴2周,后改为甘草酸二胺胶囊,2粒,口服,3次/日等基础护肝药物。治疗组在对照组基础上给予百令胶囊(虫草菌丝体干粉,中美浙江华东制药有限公司生产)0.2g/粒,4粒/次,3次/天,口服。观察患者症状改善及血清生化学指标情况,3个月为一疗程。一个疗程后,评定疗效。结果:从临床疗效看,治疗组显效5例,有效11例,无效2例,总有效率为89%;对照组显效2例,有效10例,无效6例,总有效率为67%,两组总有效率比较有统计学意义(P＜0.05),治疗组的疗效优于对照组。两组治疗后,患者症状皆有改善,但治疗组的改善更为显著,与治疗前对比,差异有显著性(P＜0.05);对照组与治疗前相比差异无统计学意义。在肝功能检测方面两组自身治疗前后比较都有显著性差异(P＜0.05),治疗后两组肝功指标比较差异无统计学意义(P＞0.05)。两组治疗后,肝纤维化指标HA、PCⅢ、LN、Ⅳ-C皆有所改善,组内前后对比有显著差异(P＜0.05);治疗组的效果更为优异,与对照组比较差异有统计学意义(P＜0.05)。结论:百令胶囊(虫草菌丝体干粉)可有效治疗肝纤维化及早期肝硬化,保护肝细胞,改善患者的肝功能及HA、LN、PCⅢ、Ⅳ-C血清学指标。
China is a region of high incidence of viral hepatitis its incidence rate is as high as 10-15%,especially in rural and underdeveloped areas of the economy.Serious hepatitis or liver failure caused by viral hepatitis,its mortality rate as high as 50-70%or higher.Hepatic cirrhosis and hepatic fibrosis caused by chronic hepatitis,seriously affected patients in the quality of life.
Hepatic fibrosis can further develope cirrhosis of liver,even final stage liver disease. Lack of effective therapeutic tool in the stage,although liver transplantation can be used as the effective means of treating liver failure and cirrhosis,but it has several drawbacks, Expensive,complex surgery,may occur severe rejection after surgery and the survival rate is not high.the lack of donor liver is a big problem.Those restrictions make it difficult to promote this therapeutic tool.
Mesenchymal stem cells(MSCs)is a group of multi-differentiation potential of stem cells.It can differentiate into a variety of out the hemopoietic tissue in specific conditions. for example,osteoblast,neuron cell,adipocyte,fibroblast,cardiac muscle cell,hepatocyte etc.
MSCs can be obtained from different organizations of the body,and can be autografted conveniently.In the course of long-term culture in vitro,MSCS has always been to maintain its multi-directional differentiation potential,its genetic background is relatively stable in vitro,it has a high rate of gene transfection and can express foreign genes stable and efficient.Therefore,when MSCs is implanted in the body,the weak response to implan in vivo is one of the features.So MSCS is an ideal seed cells for tissue engineering,wih the potential of treating acute and chronic liver failure.MSCs can directed to differentiate into liver cells,which has been widespread concerned by scholars.
1.Theoretical research
Modern medicine has made some progress in MSCs differentiate into liver cells.But MSCs can easily diffused or hydrolysed by enzymatic for pure MSCs transplantation in vivo,the MSCs that can achieve hepatocyte is not much,and it differentiation rate is low,so its prospects is lmited for practical application.The differentiation in vitro is realized under conditions of chemical praeparatum,the differentiation rate of it is low too.And its clinical safety and long-term effectiveness is also difficult to achieve,therefore it's the most important how to promote the MSCS into hepatocytes fully and more effectively to improve the structure and functions of liver organizations.
The study was started relatively late that make use of chinese medicine induce MSCs differentiate.Many research institutions have proved chinese medicine the differentiation regulation to MSCs in recent years,and wih higher induce rate than Western medicine method.It were reported that scholars applied active ingredients of Chinese medicine had been successful induced MSCs differentiate into nerve cells、adipocyte,cadiocyte, fibroblast and hepatocyte etl.There are no reports that someone apply monomer composition of traditional Chinese medicine to induce MSCs differentiate into liver cells at home and abroad.Chinese caterpillar fungus is a commonly Chinese medicine used to treat liver disease in clinical,We chose its active ingredients Chinese caterpillar fungus polysaccharide to study.Pharmacological studies show that Cordyceps polysaccharide:it has protective effect to liver damage and liver fibrosis.MSCs has a wide range of differentiate potential,According to oriented research results of applying chinese medicine active ingredients to induce differentiation of MSCs,we can believe theoretically that cordyceps polysaccharide can induce MSCs differentiate into liver cells in the right conditions.Research on traditional Chinese medicine induced MSCs to differentiate into liver cells,not only provide a new theoretical basis for applying chinese medicine to treat acute liver failure,but also can overcome some dilemma in modern medical research due to the inherent safety and long-term effectiveness of chinese medicine.It may be related to the methods that treat acute liver failure and cirrhosis by cell transplantation,Bring about broad application prospects.So it is important to carry out chinese medicine induce MSCS to differentiate into liver cells.
Along with the deepening of clinical practice of chinese medicine,people had been found some effective drugs and compositions from Chinese medicine that can treat hepatic fibrosis.Clinical and Experimental have been verificated those effects,this showed advantages and potential of chinese medicine in the treatment of hepatic fibrosis.Chinese caterpillar fungus is a traditional chinese tonic medicine,it can protect the lungs and beneficial to the kidneys,enrich the essence,relieve cough and reduce sputum,treat chronic cough and swallowing disorders.Modern research has found a variety of pharmacological effects of it,expanded its scope of application.Experiments have been showed that cordyceps can reduce indicators of experimental liver fibrosis rat model about liver fibrosis, protect liver cells.In this study,we made use of the preparations of Cordyceps sinensis(Bailing Capsule)to treat liver fibrosis with hepatitis B,observe the clinical efficacy and explore mechanisms for the treatment of liver fibrosis.Provide more scientific basis for the clinical application of it.
2.Experimental Study
ExperimentⅠ:Cordyceps polysaccharide induces differentiation of adult rat MSCs into a hepatocyte lineage in vitro
Objective:To research the possibility of the rat mesenchymal stem cells(MSCs)to differentiate into hepatocyte-like with Cordyceps polysaccharide in vitro.Methods:Bone marrow in the femurs of Wistar rat was collected by flushing under sterile condition.MSCs were separated and cultured according to the direct anchoring method,identified corpuscular surface antigen of CD34 and CD44 by using flow cytometric analys.MSCs were divided into 3 groups to induce differentiate.Cordyceps group:induced with Cordyceps polysaccharide.The final concentration of it was 0.15mg/ml.Masccline Control group:Induced by HGF and EGF,the concentration of each was 20ng/ml and 10ng/ml. Blank group:only cultivated with basis medium.The morphological changes and proliferation of MSCSs were observed by phase-contrast microscope.On day 7,14,21 and 28,AFP,CK18 and ALB of MSCSs of each group were examined,the expressions of them were detected with immunohistochemistry technique,staining for glycogen with PAS method.Results:After 24h in primary culture,MSCs adhered to the plastic surface,most cells adhered surface,cell volume increase after a week.The cells spindle-shaped and polygonal,cell proliferation significantly faster,the cells covered culture flask about 9 days. Subcultured after trypsin digestion the cells morphology are most of the spindle patterns more homogeneous than before,proliferated even faster,the cells into a colony-type growth.7 days can covered with 90%surface of culture flask.Continuous passage.Little change in cell morphology after p4.MSCs collected from the femora of Wister rats expressed antigens of CD44,but not CD34.Immunocytochemical analysis for AFP showed positive staining reaction on day7,in Cordyceps group and Masccline Control group, strengthen on day 14,then weaken on day 28.Positive staining reaction on 14 day.Masceline Control group was higher than Cordyceps group in ratio of masccline for AFP.On 14 day both group show positive staining reaction in staining for glycogen of PAS,then weaken on day 28.The blank group all showed positive staining reaction. Conclusion:MSCs induced by cordyceps polysaccharide in vitro can differentiate into hepatocyte-like cells.
ExperimentⅡ:Bailing capsule to treat hepatic fibrosis after hepatitis clinical observation
Objective:To explore the effective drug of treating hepatic fibrosis.Observe cordyceps sinensis preparation(Bailing Capsule)for hepatic fibrosis and early liver fibrosis clinical efficacy in patients.Methods:Choose chronic active hepatitis and early liver cirrhosis 36 cases of patients,which cases come from the First Affiliated Hospital of Guangzhou University of TCM and the Guangdong Provincial TCM Hospital since January 2008 to December 2008.The 36 cases randomly divided into control group and treatment group.18 cases of each group.Two groups all gave glucurolactone 0.2g,3 times/d,po;Vitamin C 2g, 3 times/d,po;Diammonium glycyrrhizinate 150mg+GS 250ml ivgtt in 2 weeks,1 times/day,replaced by diammonium Glycyrrhizinate capsule after 2 weeks,2 pills,po,3 times/day;Treatment group gave Bailing Capsule.Treatment group gave Bailing Capsule(Cordyceps Mycelium Powder,East China Pharmaceutical Co,Ltd, Zhejiang,China and the United States)in addition than the control group,0.2g / tablets,4 tablets/times,3 times/d,po.Observed symptom improvement and serum biochemical indicators in patients,3 months for a course,after a course of treatment,assess of efficacy.
Results:See from the Clinical Efficacy 5 cases of treatment group markedly,11 cases of effective,2 cases of invalid,the total effective rate was 89%;2 cases of the control group markedly,10 cases of effective,6 cases of invalid,the total effective rate was 67%,The total efficiency of the two groups was statistically significant(P＜0.05),Efficacy of the treatment group was superior to the control group.The patients had improvement in symptoms in two groups after treatment,however,the improvement of the treatment group was more significantly,Compared with before treatment,there was a significant difference (P＜0.05),The control group compared with before treatment there was no significant difference(P＞0.05).Comparison of the two groups own before and after the treatment has its significant difference in liver function testing(P＜0.05).The two groups was no significant difference in indicators of liver function after treatment(P＞0.05).Indicators of hepatic fibrosis HA、PCⅢ、LN、Ⅳ-C all were improved in two groups after treatment, Comparison before and after treatment in group were significantly different(P＜0.05),the effect of the treatment group is more superior,Compared with the control group were statistically significant differences.Conclusions:Bailing Capsule(cordyceps sinensis preparation)can treat liver fibrosis and early cirrhosis,protects liver cells,improve liver function and serum indicators of patients with hepatic fibrosis.
引文
[1].Friendenstrein A,Chailakhyan R,Gerasimov U,et al.Cell Tissue Kinet,1987;20:263-272.
[2].Pittenger MF,Mackay AM,Beck SC,et al.Multilinage potential of adult human mesenchymal stem cells.Scienses,1999;284(5411):143-147.
[3].Reyes M,Verfaillie CM.Characterization of multipotent adult progenitor cells,a subpopulation of mesenchymal stem cells.Ann New York Acad Sci,2001;938:231-235.
[4].Guo ZK,Yang JQ,Liu xd,et al.Biological features of mesenchymal stem cells from human bone marrow.Chin Med J,2001;114:950-953.
[5].Weissman IL.Translating stem and progenitor cell biology to the clinic:barriers and opportunites.Science,2000;287:1442-1446
[6].Lin H.Annu Reu Genet,1997;31:455-491.
[7].Cmnpognoli C,Roberts IA,Kumar S,etal.Identification of mesenchymal stem-progenitor cells in human first trimester fetal Deans RJ,Moseley AB.Mesenchymal stem cells:biology and potential clinical uses.Exp Hematol,2002;28(8):875.
[8].Deans RJ,Moseley AB.Mesenchymal stem cells:biology and potential clinical uses.Exp Hematol,2002;28(8):875
[9].傅文玉,路燕蒙,朴英杰.人骨髓间充质干细胞培养及多能性研究中国血液学杂志.2002:23(4):202-204.
[10].Gross TJ and Hunninghake GW.N Engl J Med,2004;345,517-525.
[11].姚青等,大鼠骨髓间充质干细胞体外诱导分化为心脏起搏样细胞的研究.中国心脏起搏与心电生理杂志,2007;21(1):57-58
[12].曹欣欣等,人骨髓间充质干细胞向心肌样细胞分化.基础医学与临床,2007;27(2):159-160
[13].Wang JS,Shum-Tim D,Augelli BJ,et al.Marrow stromal cells for celular cardiomyoplasty:feasibility and potentialclinical advantages.J Thorc Cardiovaso Surg,2000;120(5):999
[14].Wakitani S,Saito T,Caplan AI.Myogenic cels derived from rat bone marrow mesenchymal stem cells exposed to azacytidine.Muscle Nerve,1995;18:417.
[15].丁曷东等,心肌梗死后骨髓间充质干细胞移植对心室肌细胞复极活动的影响. 中华心律失常学杂志,2007;(11)2:136-137.
[16].Orllc D.Kajstura J,Chimenti S,et al.Nature,2001;4:339-351.
[17].王韫芳,裴雪涛.成体干细胞:从基础到临床.中华医学杂志,2004;16(84):1327-1328.
[18].赛因其木格等,大黄素诱导人骨髓问充质干细胞向成骨细胞分化的研究.中国中西医结合外科杂志,2006;12(5):447-448.
[19].Kadiyala s,Young RG,Thlde MA.Culture expanded canine mesenchymal stem cells possess osteochondrogenic potential in vivo and in vitro.Cell Transplant,1997;6:125.
[20].Pittenger MF,Mackay AM,Beck SC,etal.Multi—line.age potential of aduh human mesenchymal stem cells.Science,1999;284(5411):143.
[21].Yoo JIJ,Barthel TS,Nishimum K,eI al.The chondro genic potential ofh.man bone marrow-derived mesenchymal pro-genitor cells.J Bone Joint Surg,1998;80(12):1745.
[22].李林等,人骨髓间充质干细胞的分离、培养和体外定向诱导成类软骨细胞的实验研究.齐齐哈尔医学院报,2007;28(3):275-277.
[23].Balfoth B,Weisser J,Stemkop F。 et al.Transplantation of allagraft chondmcytes embeded inagarosegelin to cartilage defects ofrabbits.Osteoarthritis Cartilage,1998;6:50.
[24].Worster AA,Nixon AJ,Brower-Toland BD.et al.Efect of transforming growth factor betaon chondrogenic diferentiation of cultured equine mesenehymal stem cells.Am J VetRes,2000;61(9):1003.
[25].Yan ZQChen YS.Li WJ el al.Treatment of osteonec rosis of the femoraI head by percutaneous decompression and autologous bone marrow mononuclear cellinfusion.Chin J TraumatoI 2006;9(1):265.
[26].Ji WF,Tong PJ,Zheng W B,et al.ExperimentaI study on treatment of femoral head necrosis with arteriaI perfusion of marrow stem cells.Zhongguo Xiufu Chongjian Waike Zazhi.2004;24(11):999-1002.
[27].Terada M,Namazaki T,Oka M,et,al.Bone marrow cells adopt the phenotype of othercells by spontaneous cell fusion.Nature,2002;416:542-545.
[28].Ying QY,Nichols J,Evans EP,et al.Changing potency by spontaneous fusion.Nature,2002;416:545-574.
[29].Woodbury D,Schwarz EJ,Prockop DJ,et al.Adult rat and human marrow stromal cells differentiate in—to neurons[J].J Neurosci Res,2000;61:364-370.
[30].苏平等,银杏内酯 B 对骨髓间充质干细胞分化为神经样细胞的影响.广东医学,2007;28(1):33-34.
[31].肖庆忠,李浩威,温冠媚等.麝香多肽体外诱导成年大鼠和人诱导骨髓间充质干细胞定向分化为神经元的研究.中国生理病理杂志,2002;18(10):1179-1182.
[32].刘金保,董晓先,董燕湘等.多种中药诱导大鼠骨髓间质干细胞转变为神经元样细胞.中国药物与临床,2003;3(3):234-236
[33].Ohta M,Suzuki Noda et al.Bone marrow stromal cells infused into the cerettospinal flfid promote ftmctioml recovery of the injured rat spinal cord wiht reduced cavity formation.Exp Neurol,2004;187:266-278.
[34].Malmaood Lu D.Qu C,et al.Long-term recovery after bone marrow stromal cell treatmem of tmmatie train injury in rats.J Neurosurg,2006.104:272-277.
[35].MarinaM andBarbaraI.SocNephrol.2004;15:1794-1804.
[36].张中华等,髓间充质干细胞体内分化为肾小管上皮细胞的实验研究.江西医学检验,2007;25(1):15-17.
[37].赵嫒等,大鼠骨髓间充质干细胞分离培养及向脂肪细胞分化的实验研究。四川解剖学杂志,2007;15(1):31-32.
[38].刘弘光等,移植骨髓间充质干细胞治疗大鼠糖尿病的研究.中国组织化学与细胞化学杂志.2007;16(1):104-105.
[39].OH SH,Miyazaki M,Kouch H,et al.Hepatocyte growth factor induces differentiation of adult rat bone marrow cells into a hepatocyte lineage in vitro[J].Bioch Bioph Res commun,2000;279(2):500-504.
[40].Avital I,Inderbitzin D,Aoki T,et al.Isolation,characterization,and transplantation of bone marrow derived hepatocyte stem cells[J].Biochem Biophys Res Commun,2001;288(1):156-164.
[41].蔡云峰,闵军,何劲松等.骨髓源性肝干细胞的确认及定向分化的实验研究[J].中国普通外科杂志,2003;12(4):287-290.
[42].何忠杰,方驰华,马俊勋等.肝细胞生长因子与表皮细胞生长因子联合诱导大鼠骨髓间充质干细胞分化为类肝细胞[J].解放军医学杂志,2006;5(31):446-449.
[43].Wang P P,Wang J H,Yan Z P,et al.Expression of hepatocyte like phenotypes in bone marrow stromal cells after HGF induction[J].Biochem Biophys Res Commun,2004;320(3):712-716.
[44].Lee KD,Kuo TK,Whang-Peng J,Chung YF,Lin CT,Chou SH,Chen JR,Chen YP,Lee OK.In vitro hepatic differentiation of hum an mesenchvm al stem cells[J].Hepatology,2004;40:1275-1284.
[45].Theise ND,Nimmakayalu M,Gardner R,et al.Liver from bone marrow in humans[J].Hepatology,2000;32(1):11-16.
[46].陆海华等.小鼠骨髓间充质干细胞体内定向诱导分化与治疗肝功能损伤的实验研究[J].中华医学杂志,2007;87(4):223-224.
[47].Sato Y,Araki H,Kao J,et al.Human mesenchymal stem cells xenografted directly to rat liver are differentiated into human hepatocytes without fusion.Blood[J],2005;106:756-763.
[48].Oyagi Hirose M,Kojima M,Okuyama M,KawaseM,Nakamura T,Ohgushi H,Yagi K.Therapeutic effect of transplanting HGF-treated bone marrow mesenchymal cells into CCl_4-inlured rats[J].Hepatology 2006;44:742-748.
[49].Hung SC,Lu CY,Shyue SK,Liu H C,H o LL.Lineage differentiation-associated loss of adenoviral susceptibility and Coxsackie-adenovirus receptor expression in human mesenchymal stem cells.Stem Cells 2004;22:1321-1329
[50].TsuiTY,Lau CK,Ma J,Glockzin G Obed A,Schlitt HJ,Fan ST.Adeno-associated virus-mediated heme oxygenase-gene transfer suppresses the progression of micronodular cirrhosis in rats[J].World Gastroenterology,2006;12:2016-2023.
[51].Tcrai S.Sakaida I.Yamamoto N.et al.An in viva model for moni-toring trans-diffcrcntiation of bone marrow cells into functional hepatocytes[J].[J].Biochem,2003;134(4):551-558.
[52].Yamamoto N.Tcrai S.Ohata S.et al.A subpopulation of bone marrow cells depleted by a novel antibody.anti-Liv8 is useful for cell therapy to repair damaged liver[J].Biochem Biophys Res com,2004;313(4):1110-1118.
[53].Miyazakit,Studies on on fungal polysaccharides XX.Galactom annan Of Cordyceps sinesis[J].Chem Pham Bull,1977;25(12):3325-3328.
[54].龚敏,朱勤,王彤等.冬虫夏草多糖的分子结构与免疫活性[J].生物化学杂志,1990;6(6):486-492.
[55].李楠,宋建国,刘金云等.蛹虫草与冬虫夏草化学成分比较[J].吉林农业大学学报.1995;17(增刊):80-83.
[56].白云娥,李青山,王毅等.冬虫夏草多搪的含量测定[J].山西医科大学学报,2000:31(2):129-130.
[57].Ng TB.Tranditional Chinese medicine and the search for new antineoplastic drugs.Life Science[J],1997;60(25):2349.
[58].郭风彩.冬虫夏草多糖的药理学研究进展[J].现代医药卫生,2006;22 (7):999-1000.
[59].刘超,卢珊,姬美蓉等.冬虫夏草对白血病 NK 细胞影响的体外研究[J].中国中西医结合杂志,1992;12(5),267-269.
[60].张椒兰,孙云汉,刘晓平,等.冬虫夏草及人工虫草菌丝抗小鼠 Lewis 肺癌的研究[J].中药通报,1987;12(2):53.
[61].刘湘.从冬虫夏草得到的蛋白多糖0909具有抗肿瘤和免疫促进活性[J].国外医药·植物药分册,1995;10(2):91.
[62].龚敏,朱勤,王彤,等.虫草多糖的分子结构与免疫活性[J].生物化学杂志,1990;6(6):486-491.
[63].刘晓平.冬虫夏草及人工虫草菌丝对 Y 线照射后小鼠血小板和免疫器官的影响[J].中药通报1988;13(4):44.
[64].龚晓健,季晖.人工冬虫夏草对小鼠免疫功能的影响[J].中国药科大学报,2000;31(1):53
[65].程琪琳,邱德凯,张泳.虫草脂质体对小鼠巨噬细胞的作用[J].上海免疫学杂志,1989;9(3):143.
[66].靖大道,邱德凯,萧树东等.虫草多糖对人外周血 IL-2,IL-2R 及 IFN-r 调节作用的研究[J].上海免疫学杂志,1995;15(6):321-323.
[67].张建,韦星呈等.北虫草多糖对人肿瘤坏死因子α、白细胞介素-2及其可溶性受体水平的影响[J].中国实用内科杂志,1997;17(12):727-728.
[68].孙荣玲、李国忠复方虫草多糖脂质体口服液对慢性乙型肝炎细胞免疫的影响[J].中国药师.2005;8(10).848-849.
[69].夏宝森,李晓康.FTY720:一个来源于冬虫夏草的新型免疫抑制剂[J].上海免疫杂志2002;22(2):79-81.
[70].匡彦德,王美英,钱琴芳等.冬虫夏草免疫增强作用机理的研究.上海免疫学杂志1989;9(1):6.
[71].许维祯,李丽芬,石扣兰等.冬虫夏草菌丝体对单氨氧化酶及免疫功能的影响[J].上海中医药杂志1988;(1):48.
[72].陈道明,张淑兰,于志洁等.冬虫夏草及人工培养菌丝对小鼠血小板生成的影响及超微结构观察[J].中药通报,1987;12(1):47.
[73].祝希嫒,史勇,刘晓明等.人工培养冬虫夏草菌粉对细胞免疫的抑制作用[J].中西医结合杂志,1990;10(8):485-486.
[74].夏益平.冬虫夏草口服液治疗慢乙肝36例疗效观察[J].右江医学,2000;28(2):91-92.
[75].季晖等.人工虫草菌丝多糖的分离提取及其降血糖作用研究[J].中国药科大学学报,1993;24(1)39-42.
[76].黄志江,季晖,李萍.人工虫草多糖降血糖作用及其机制研究.中国药科人学学报,2002;33(1):51-54.
[77].Kiho T.Hypoglycemic activity of a cs-F30 from the cultural mycelium of Cordyceps sinensisand its effecton glucosemetabo-lism in mouse liver.Biolpharm.Bul[J],1996;19(2):294.
[78].许建明,丁长海,李连德,等.冬虫夏草多糖保护小鼠免疫性肝损伤的筛选研究[J].安徽医科大学学报,1999;34(3):173-175.
[79].刘玉凤,李彩霞.冬虫夏草多糖脂质体治疗慢性乙型肝炎86例临床观察[J].山西中医,1998;14(1):15-16.
[80].赵立群.复方冬虫夏草多糖脂质体口服液治疗肝炎后肝硬化120例疗效观察[J].中医药学刊.2005;23(7).1300-1300.
[81].马雄,邱德凯,徐军.虫草多糖脂质体抗肝纤维化的实验研究[J].中国中医基础杂志,1999;5(9):28-30.
[82].颜吉丽等.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-α表达的影响[J].复旦学报:医学版.2003;30(1):27-29.
[83].杨俊何,凌耀生.虫草多糖的抗氧化作用[J].广东药学院学报,1997;13(1):35-38.
[84].李泓俊,彭欣辉,吴应莲.虫草多糖脂质体对小鼠免疫性肝纤维化TGF-β_1mRNA 表达的影响[J].临床医学研究,2004;21(5):500-503.
[85].李风华、刘平等,虫草多糖逆转DMN诱导大鼠肝纤维化的作用及机制研究[J].中国中药杂志,2006;31(23):1968-1971.
[86].马雄,邱德凯,徐军.虫草多糖脂质体抗肝纤维化的实验研究[J].中国中医基础杂志,1999;5(9):28-30.
[87].方步武,刘平,刘成,等.虫草多糖抗免疫损伤性大鼠肝纤维化的作用及其机理研究[J].上海中医药杂志,2000;(9):37-40.
[88].方士英,虫草多糖对小鼠化学性肝损伤的保护作用[J].安徽医科大学学报.2004;39(3):201-204.
[89].曾宪可,唐玉,袁盛榕.冬虫夏草和中国拟青霉对小鼠免疫性肝损伤的保护作用[J].中国药学杂志,2001;36(3):161.
[90].张小强.冬虫夏草与人工虫草菌丝体对四氯化碳所致急性化学性肝损伤的保护作用[J].环境与职业医学,2003;20(6):422-426.
[91].夏益平.冬虫夏草口服液治疗慢乙肝36例疗效观察[J].右江医学.2000;28(2).91-92.
[92].Sato M,Suzuki S,Senoo H.Hepatic stellate cells:Unique characteristics in cell biology and phenotype[J].Cel Struct Funct,2003;28(2):105-112.
[93].Gressner AM,Weiskirchen R,Breitkopf K,et al.Roles of TGFβ betain hepatic fibrosis[J].Front Biosci,2002;7:793-807.
[94].王萍.血小板衍生生长因子与肝纤维化[J].国外医学.消化疾病分册,2000;20(1):40-43.
[95].Holmes A,Abraham DJ,Sa Set al.CTGF and SMADs,maintenanee of seleroderma phenotype is independent of SMADsignaling[J].J Biol Chem,2001;276(14):10594-10601.
[96].彭小东,戴立里.结缔组织生长因子与肝纤维化的研究进展[J].胃肠病学与肝病学杂志,2005;14(4):425-27.
[97].Saxena N K,Titus M A,Ding X,et al.Leptin as a novel profibrogenie cytokine in hepatic stellate cells:mitogenesis and inhibition of apoptosis mediated by extracellular regulated kinase(ErK)and Akt phosphory lation[J].FASEB J,2004;8(13):1612-1614.
[98].Cao Q,Mak K M,Lieber C S,et al.Leptin enhances alphal(I) collagen gene expression in LX-2 human hepatic stellate cellsthrough JAK-mediated H-2O_2 dependent MAPK pathways[J].J Cell Biochem,2006;97(1):188-197.
[99].何生松,徐标,韩春荣.环醇对小鼠日本血吸虫病肝纤维化即早基因与 TGF-β1、TIMP1及胶原表达的影响[J].世界华人消化杂志,2007;15(35):3678-3684.
[100].Wu JY,Wu YB,et al.Effects of endothelin-1 On hepatic stellate cell proliferation,collagen synthesis and secretion intracellcular flee calcium concentration[J].World J Gastroenterol,2004;10(18):2697-2700.
[101].Bataller R,Gines P,Nicolas JM,et al.Angiotensin Ⅱ induces contraction and proliferation of human hepatic stem cells[J].Gastroenterology,2000;118(6):1149-56.
[102].Bataller R,Sancho-Bru P,Gines P,et al.Activated human hepaticsteHate cells express the rennin-an giotensin system and synthesize angiotensin [J].Gastroenterology,2003;125(1):117-25.
[103].Balk SK,Jo Hs,Suk KT,etal.Inhibitory efect ofangiotensin Ⅱ receptor antagonist on the contraction and growth of hepatic stellate cells[J].Korean J Gastroenterol,2003;42(2):134-41.
[104].Kurikawa N,Suga M,Kuroda S,et al.An angiotensin Ⅱ type 1 receptor antagonist,oimesartanmedoxomil,improves experimental liver fibrosis by suppression of proliferation and collagen synthesis in activated hepatic steHate cells[J].Br J Pharmacol,2003;139(6):1085-94.
[105].李小刚.细胞因子网络在肝损害中作用的演剧进展[J].国外医学·内科学分 册,1997;24(1):20.
[106].张文胜,王宝恩,王泰龄,等.慢性乙型肝炎纤维化无创性诊断模型的建立[J].中华肝脏病杂志,2006;14:169-173.
[107].Itagaki T,ShimizuI,ChengX,et al.Opposing efect s of oes-tradiol and progesterone On intraeellular pathways and aetivationproeesses in the Oxidative 8 tress induced activation of cultured rat hepatic stellate celJs.Gut.2005;54:1782-1789.
[108].Novitskiy G,Potter J J,Wang L,et al.Influences of reactive oxygen species and nitric oxideon hq-icfibrogenesis.Liverht,2006;26:1248-1257.
[109].BatallerR,BrennerDA.Hepatic stellate cells as a target for the treatment of liver fibrosis[J].S-nin Liver Dis,2006;21:437-451.
[110].Ziol M,Handra2Luca A,Kettaneh A,et al.Noninvasive assessment of livel fibrosis by rrleasuH nent of-ifness in patients with chronic hepatitis [J].Hepatologv,2005:41:48-54
[111].中华医学会肝病学分会脂肪肝和酒精性肝病学组.非酒精性脂肪性肝病诊疗指南[J].中华肝脏病志,2006;14:161-163.
[112].CoiliA,Fraquelli M,Andreoletti M,etal.Severe liverfibrosisor cirrhosis:accuracy of US for detection analysis of 300 cases [J].Radiology,2003;227:89-94.
[113].Zheng RQ,Wang QH,Lu MD,et al.laver fibrosis in chronic viral hepatitis:an ultrasonographic study.World J Gastroenterol,2003;9:2484-2489.
[114].Aube C,Oberti F,Korali N,et al.UItrasonographic diagnosisof hepatic fibrosis of cirrhosis[J].J Hepatol,1999;30:472-478.
[115].Galani S Gramantieri L,Venturoli N,et al.W hat is the criterion for differentiating chronic hepatitis from compensated cirrhosis?A prospective study comparing uItrasonography andpercutaneous liver biopsy[J].J Hepatol,1997;27:979-985.
[116].Bernatik T,Strobel D,Hahn EC,et al.Doppler measurements:a surrogate marker of liver fibrosis[J].Eur J Gastroenterol Hepatol,2002;14:383-387.
[117].Taura T,Nakam ura K,Takashima S,et al.Heterogeneity of hepatic parenchymal enhancement on computed tomography during arterial portography:quantitative analysis of correlation with severity of hepatic fibrosis[J].Hepatol Res,2001;20:192-192.
[118].陈克敏,柴维敏.肝纤维化患者肝脏形态学改变的 CT、MR 评价肝脏[J].2000;5(4):209-210.
[119].van Beers BE,Leconte I,M aterne R,et al.Hepatic perfusion parameters in chronic liver disease:dynamic CT measurements correlated with disease severity[J].Am J Roentgenol,2001;(176):667-673.
[120].Materne R,Annet L,Dechambre S,et al.Dynamic computed tomography with low and high nmolecular mass contrast agents to assess microvascular permeability modifications in a model of liver fibrosis[J].Clin Sci(Lond).2002;1(13):213-216.
[121].Cho SG,Kim MY,Kim J,etal.Chronic hepatitis:invivo proton MR spectroscopic evaluation of the liver and correlation with histopathologic findings[J].Radiology,2001;221(3):1740.
[122].Rouvire O,Yin M,Dresner M A,et al.M R elastography of the liver:preliminary results[J].Radiology,2006;240(2):440.
[123].李晶,张菁.重组α-干扰素治疗慢性乙型肝炎肝纤维化46例疗效观察[J].临床和实验医学杂志.2006;5(10):1546-1547.
[124].张岩,白雪帆,李艳红等.拉米夫定治疗慢性乙型肝炎的疗效及肝组织学改变[J].中国误诊学杂志,2006;5(5):804-806.
[125].Leung NW,Lai CL,Chang TT,et al.Extended lamivudine treatment in patients with chronic hepatitis B enhances hepatitis B antigen seroconversion rates:results after 3 years of therapy[J].Hepatology,2001;33(6):1527-1532.
[126].Bataller R,Brenner DA.Hepatic stellate cells as target for the treatmentn of liver fibrosis[J].Semin Liver Dis,2001;21(3):437-45.
[127].谭海荣,吴谦,潘竞锵,等.水飞蓟宾对抗大鼠酒精性脂肪肝作用及机制[J].中国临床药理学与治疗学,2004;9(7):803-806.
[128].Boigk G,Stroedter L,Herbst H,et al.Silymarin retards collagen accumulationin early and advanced biliary fibrosis secondary to complete bile duct obliteration in rats[J].Hepatology,1997;26(3):643-649.
[129].许倩,许建娟.腺苷蛋氨酸治疗妊娠肝内胆汁淤积症疗效探讨[J].中国优生与遗传杂志,2004:12(1):83-84.
[130].汪余勤,程五风,李宣海.维生素 E 与肝纤维化[J].临床肝胆病杂志,2000;16(1):19.
[131].Dogru Abbasoglu S,Balkan J,Kanbagli O,et al.Aminoguanidineian inducible nitric oxide synthase inhibitor,plus N-acetylcysteine treatment reduce the lipopOlysaccharide-augmented hepatotoxicity in rats with cirrhosis[J].Hum Exp Toxicol,2002;21(7):359-364.
[132].Vendemiale G,Grattagliano I,Caruso ML,et al.Increased oxidativestress in dimethylnitrOsamine-induced liver fibrosis in the rat:effect of N.acetylcysteine and interferon-alpha[J].Toxicol Appl Pharmaeol,2001;175(2):130-139.
[133].Meurer SK,Lahme B,Tihaa L,et al.N-acetyl-L-cysteine suppresses TGFβ a signaling at distinct molecular steps:the biochemical and biological efficacy of a muhfunetional,antifibrotie drug[J].Biochem Pharmacol,2005;70(7):1026-1034.
[134].Arrieta O,Rodriguez JL,Rosaa V,et al.Colchicine delays the development of hepatocellular carcinoma in patients with hepatitis virus-related liver cirrhosis[J].Cancer,2006;107(8):1852-1858.
[135].Poupon RE,Huet PM,Poupon R,et al.A randomized trial corapa-ring colchicine and ursodexyeholie acid combination to ursodeoxycholic acid in primary billiary cirhosis[J].UDCA—PBC Study Group.Hepatology.1996;24(5):1098-1103.
[136].程书权.肝纤维化的现代药物治疗研究[J].中国处方药,2005;3(5):36-40.
[137].阳学风,曾明新,张明亮等.马洛替酯抑制肝纤维化的实验与临床研究[J].世界华人消化杂志,1999;7(3):224-226.
[138].何方平,孙惠蓉,张跃新等.四种药物对大鼠肝纤维化防治作用的实验研究[J].新疆医科大学学报,2003;26(4):338.
[139].LieberCS.New concepts of thepathogenesis of alcoholic liver disease lead to novel treatments[J].Cur Gastroenterol,2004;6(1):60-65.
[140].Wamoto H。 Nakamuta M,Tada S,et al.Platelet-derived growth factorreceptor tyrosine kinase inhibitor AG1295 attenuates rat hepatic stelate cell growth[J].Lab Clin Med,2000;135(5):406.
[141].Cut X,Shimizu 1,Lu G,et al.Inhibitory efect of a soluble transforming growth factor beta type Ⅱ receptor on the activation of rat hepatic stellate cells in primary culture.Hepar,2003;39(5):731.
[142].齐洪军,胡蔓菁.从“痰瘀”角度浅析肝纤维化的病因病机及治疗[J].四川中医,2004;22(6):26-28.
[143].刘为民,姚乃礼.络病理论与肝纤维化临床[J].中医药学报,2003;31(1):2-3.
[144].叶放,薛博瑜,周珉,等.论湿热瘀毒与肝纤维化[J].南京中医药大学学报,2005;21(6):346-349.
[145].唐世利,刘其政,王艳妮.化瘀软肝汤治疗肝纤维化70例[J].陕西中医,2002:23:588-589.
[146].程明亮,付荣泉.中药复方对猪血清免疫性肝纤维化肝中氧化及抗氧化的影响[J].中华肝脏病杂志,2006;14(3):217-218.
[147].赵晓晨,张琳.复方丹参注射液抗肝纤维化的疗效观察[J].山西中医学院学报,2007;7(6):38.
[148].齐洪军,胡曼青,王长松,等.祛瘀化痰汤抗大鼠肝纤维化的作用及其机制[J].江苏中医杂志,2003;24(7):55-57
[149].方谦和,孙维娜.方谦和和解法治疗肝纤维化的经验[J]北京中医,2004:23(3):143-144.
[150].肖会泉,刘妮,杨俭勤,等.肝舒胶囊对慢性乙型肝炎肝纤维化的临床疗效观察[J].山东中医杂志,2006;25(10):663-665.
[151].褚裕义,孙克伟,刘伟士,等.疏肝理脾片抗大鼠免疫性肝纤维化研究[J].中国中西医结合脾胃杂志。1998;6(2):98.
[152].龙爱华,陈瑾,王树槐.抑肝健脾复方抗大鼠肝纤维化的实验研究[J].中西医结合肝病杂志,2002;12(1):23-25.
[153].刘爱丽,魏敏,甑增国,等.小柴胡汤凯西莱联用抗肝纤维化50例[J].陕西中医,2005;26(9):873-874.
[154].郭晓华,王宜芳.复肝片治疗慢性乙型肝炎肝纤维化的临床观察中成药.2006;28(12):1760-1762.
[155].李新华滋阴养肝汤治疗慢性乙型肝炎后肝纤维化的临床观察上海中医药杂志.2006;40(11):14-15
[156].蔡春江,褚志敏,崔书芬,等.柔肝方治疗慢性乙肝肝纤维化的临床研究[J].四川中医,2006;24(4):43-45.
[157].郭晓萍,侯允天,李东良,等.柔肝煎冲剂对实验性肝纤维化大鼠血清透明质酸及Ⅲ型前胶原的影响[J].中医药学报,1996;12(5):42.
[158].叶放,薛博瑜,周眠,等.论湿热瘀毒与肝纤维化[J].南京中医药大学学报,2005;21(6):346-349.
[159].于世瀛,贲长恩,杨美娟,等.清热利湿方药抗肝纤维化的形态学和免疫组织化学定量研究[J].北京中医药大学学报,1997;20(1):28-30.
[160].李日向,常占杰,叶青.清木丹颗粒治疗慢性乙型肝炎湿热留恋瘀毒型肝纤维化的临床研究[J].陕西中医学院学报,2005;28(2):8-9.
[161].朱清静,聂广.剔毒护肝方抗鸭乙型肝炎肝纤维化的作用.中西医结合肝病杂志.1998;8(2):84-87.
[162].李明奎.芪茜化纤汤治疗肝纤维化37例临床观察[J].山西中医学院学报, 2006;7(4):28.
[163].李召忠,王啸,隋在云,等.血隆冲剂抗肝纤维化的临床与实验研究[J].中国中西医结合杂志,2001;21(11):813-815.
[164].唐苾芯,李金华,黄存垣.益肝消癥散抗慢性肝炎肝纤维化临床研究[J].江西中医学院学报.2004;16(2).40-43.
[165].王晓峰,董培红,陈威,等.汉防己甲素对脂多糖刺激的大鼠肝星状细胞增殖的影响[J].现代实用医学,2006;18(7):452-454
[166].Chor SY,Hal AY,To KF,et al.Anti-proliferative and pro-apoptotic effects of herbal medicine on hepatic stellate cell[J].Ethnopharmacol.2005;100(1-2):180-186.
[167].徐永红,边城,李定国,等.汉防己甲素对成纤维细胞增殖及胶原合成的作用[J].天津医药,2004;32(10):633-635.
[168].权启镇,王要军,权源,等.复方汉防己冲剂抗肝纤维化的实验研究,中华肝脏病杂志,2007;15(11):873-87.
[169].周光德,李文淑,赵景民等.复方鳖甲软肝片抗肝纤维化机制的临床病理研究.解放军医学杂志,2004;29(7):563-564.
[170].王晓玲,崔云华,胡旭东等.丹酚酸 B 对大鼠肝星状细胞增殖周期的抑制作用.中华消化杂志,2004;24(1):59.
[171].胡义扬,王润平,赵长青,等,丹酚酸 A 对实验性大鼠肝纤维化Ⅰ型胶原及其基因表达的影响[J].中国中医药科技,1999;6(4):235-237.
[172].胡义扬,王润平,张学军,等.体外四氯化碳损伤肝细胞对肝星状细胞活化的影响及其丹酚酸 A 的干预效果[J].中华肝脏病杂志,2000;8(5):299-301.
[173].王志凌,成军,刘妍,等.甘草甜素抑制肝星状细胞增殖作用的研究.肝脏,2005:10(3):225-226.
[174].贾道全,张正,罗成福,等,甘草甜素逆转肝纤维化及早期肝硬化的作用探讨[J].中华消化杂志,2001;21(12):754-756.
[175].王吉耀,郭津生,刘淑玲,等.甘草甜素对肝硬化动物模型肝脏内 NF-KB 结合活性的抑制作用[J].中华肝脏病杂志,1999;7(1):42-43.
[176].郭津生,王吉耀,宋后燕,等,强力宁对大鼠肝硬化肝脏内肝细胞生长因子表达的影响.中华肝脏病杂志,1999;7(2):112.
[177].崔德广,李素云,王生耀.干扰素 alb 联合苦参碱治疗肝炎肝纤维化40例[J].中国新药杂志,2007;16(4):314-316.
[178].卢清,张清波,张继明,等.氧化苦参碱对大鼠肝星状细胞旁分泌活化途径的抑制作用[J].肝脏,2004;9(1):31-33.
[179].黄瑾,胡晋红,邱磊等.阿魏酸钠抑制肝星状细胞合成胶原的机制.药学学 报,2004;38(8):577-580
[180].吴建红,陕光,张端莲,等.川芎嗪抗实验性肝纤维化组织中 TGF-β1阳性面积率表达的比较[J].数理医药学杂志,2006;19(3):237-238.
[181].谈博,宋健平,张奉学,等.川芎嗪对 HSC-T6细胞 smad 蛋白细胞内转位的影响[J].中药新药与临床药理,2006;17(5):320-322
[182].Lucena MI.Andrade lad.de la Cruz JP.et al.Effects of silymatin MZ-8on oxidative strcss in paticnts with alcoholic cirrhosis,Rcsults of arandomized Doudle-blind,placebo-controlled clinical study.Int Clin Pharmacol Ther,2002;40(1):2-8.
[183].徐列明,刘平,刘成,等.桃仁提取物合并虫草菌丝抗肝纤维化的实验研究[J].中国中医药科技,1995;2(1):18-20.
[184].王文兵,载立里,郑元义等,三七总皂苷对大鼠肝纤维化防治机制[J].世界华人消化病杂志,2004;12(8):1939-1942.
[185].展玉涛,刘宾,李定国,等.大黄素抗肝纤维化的作用机制[J].中华肝脏病杂志,2004:12(4):245-246.
[186].Akamatsu S,Watanabe A,Tamesada M,et al.Hepatoprotective effect of extracts from Lentinusedodes mycelia on dimethylnitrosamine-induced liver injury[J].Biol Pharm Bull,2004:27(12):1957-1960.
[187].Van de Casteele M,Roskams T,Van der Elat I,et al.Halofuginonecan worsen liver fibrosis in bile ductobstructed rats[J].Liver Int,2004;24(5):502-509
[188].张锡流,梁健,杨光业,等.牛磺酸对大鼠肝纤维化组织Ⅲ型胶原的影响[J].中国中西医结合消化杂志,2005;13(4):245-247.
[189].黄琳芸,钟鸣,杨增艳,等.排钱草总生物碱对肝纤维化大鼠血清干扰素和肝脏组织病理学的影响[J].中国中医药科技,2006;13(2):101-102.
[190].Petersen BE.Bone marrow as a potential source of hepatiC oval cellS[J].Science,1999;284:1168-1170.
[191].王要军,孙自勤,权自镇等.冬虫夏草对大鼠实验性肝纤维化的防治作用及其机理研究[J].中国中药杂志,1996;21(3):179-181.
[192].Niklason LE,G ao J,AbbottW M,etal.Functionalarteries grown in vitro.Science 1999;284(5413):489-493.
[193].Campagnnoli C,Roberts lAG,Kumar S,et al.Identification of mesenchymal stem/progenitor cel ls in humans first-trimester fetal blood,liver,and marrow[J].Blood,2001;98:2396-2402.
[194].Martinez C,Hofmann T J,Marino R,et al.Human bone marrow mesenchymal stromal cells express the neural ganglioside GD2:a novel surface marker for the identification of MSCs[J].Blood,2007;Epub ahead of print.
[195].Gang E J,Bosnakovski D,Figueiredo C A,et al.SSEA-4 identifies mesenchymal stem cells from bone marrow[J].Blood,2007;109(4):1743-1751.
[196].Skarpen E,Oksvold MP,Grosvik H,et al.Altered regulation of EGF receptor signaling following a partial hepatectomy[J].J Cell Physiol,2005;202(3):707-716
[197].RE Schwartz,M Reyes,L Koodie,et al.Muhipotent adult progenitor cellsfrom bone marow diferentiate into functional hepatocyte-like cells[J].ClinInvest,2002;109(10):1291-1302.
[198].鄂征·组织培养和分子细胞学技术[M]·第1版·北京:北京出版社.
[199].刘树辉,曹中伟,秦书俭,等.SD 大鼠骨髓间充质干细胞的分离培养方法的研究[J].锦州医学院学报,2006;27(2):35-36.
[200].张刚庆,方驰华,池达智.肝细胞生长因子诱导骨髓间充质干细胞向肝细胞分化的实验研究[J].中华外科杂志,2005;43(11):716-720
[201].Xin-Qin Kang,Wei-Jin Zang,Tu-Sheng Song,et al.Rat bonemarrow mesenchymal stem cells differentiate into hepatocytes in vitro[J].World J Gastroenterol,2005;1:3479-3481.
[202].Terada M,Namazaki T,Oka M,et,al.Bone marrow cells adopt the phenotype of other cells by spontaneous cellfusion[J].Nature,2002;416:542-545.
[203].Zaret KS.H epatocyte differentiation:from the endoderm and beyond[J].Curr Opin Genet Dev,2001;11:568-574.
[204].Jung J,Zheng M,Goldfarb M.et al.Initiation of mammalian liver development from endoderm by fibroblast growth factors[J].Science,1999;284:1998-2003.
[205].Kamiya A,Kinoshita T,Miyajima A.Oncostatin M and hepatocyte growth factor induce hepatic maturation via distinct signaling pathways [J].FEBS Lett,2001;492:90-94
[206].Uehara Y,Mori C,Noda T,et al.Rescue of embryonic lethality in hepatocyte growth factor/scatter factor knockout mice[J].Genesis,2000;27:99-103.
[207].Okumoto K,Saito T,Hattori E,et al.Diferentiation of bone marow cells into cells that express liver-specific genes in vitro:implication of the Notch signals in differentiation[J].Biochem Biophys Res Comman,2003;304(4):691-695.
[208].kumoto K,Saito T,Hattori E,et al.Expression of Notch signalling markers in bone maiTow cells that differentiate into a liver cell lineage in a rat transplant model[J].Hepatol Res,2005;31(1):7-12.
[209].Schwartz RE,Reyes M,Koodie L,et al.Muhipotent adult progenitor cells from bone maiToW differentiate into functional hepatocyte-like cells[J].J Clin Invest,2002;109(10):1291-1302.
[210].Saji Y,Tamura S,Yoshida Y,et al Basic fibroblast growth factor promotes the trans diferentiation of mouse bone marow cells into hepatie lineage cells via multiple liver-enriched transcription factors[J].J Hepatol,2004;41(4):545-550.
[211].Avital I,Inderbitzin D,Aoki T,et al.Isolation,characterization,and transplantation of bone marrow-derived hepatocyte stem cells[J].Biochem Biophys Res Commun,2001;288:156-164.
[212].Suzuki A,lwama A,Miyashita H,et al.Role for growth factors and extracellular matrix in controlling differentiation of prospectively isolated hepatic stem cells Development,2003;130:2513-2524.
[213].黎晖,周健洪,陈东风,等.龟板对大鼠骨髓间充质干细胞向成骨分化的 影响[J].中药新药与临床药理,2005;3(16):159-161.
[214].徐展望,张建新,谭国庆,等.中药骨碎补提取液对兔骨髓基质细胞体外成骨分化的影响[J].中医正骨,2006;6(18):15-16.
[215].陈杰,陈新月.肝纤维化诊断研究进展[J].国际流行病学传染病杂志,2006;3(6):414.
[216].Oberti F.Valseda E.Piletle C,et al.Noineasive diagnosia of hepeticfibrosis or cirrhosis[J].Castroenterology.1997;113(5):1609-1611.
[217].王静艳,高红,王兆荃,等.Ⅲ型前胶原层粘连蛋白在肝纤维化诊断中的意义[J].中国医科大学学报,1997;26(2):17.
[218].Obata K,Twata T,Tnoue K,et al.One step sandwich enzyme immunoassay for human type Ⅳ collagen using monodonalantibodies.Clin Dim Acta[J],1989;181:293.
[219].Okazaki I,Watanabe T,Hozawa S,et al.Molecularmechanism of the reversibility of hepatic fibrosis:with special reference to the role of matrix metalloproteinases[J].J Gastroenterol Hepatol,2000;15(Suppl):D26-32.
[220].黄字琦,高毅.大鼠肝纤维化基质金属蛋白酶及其抑制因子的表达[J].世界华人消化杂志,1999;7:795-796.
[221].潘爱萍,林英辉,潘元平等.肝纤维化血清学标志物与肝脏病理相关性研究[J].南方医科大学学报.2007;27(12).-1935-1936,1938.
[222].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.
[223].王究波,刘平.虫草菌丝提取物干预与治疗二甲基亚硝胺诱导大鼠肝硬化的实验研究[J].中国中西医结合杂志.2008;28(7):617-622.
[224].刘平,王晓玲.扶正与化瘀影响肝星状细胞活化、胶原生成及肝细胞白蛋白生成的配伍作用研究.上海中医药大学学报.1999;13(3):46-50.
[225].王继,陈俊红.活血、益气、软坚不同中医治则逆转大鼠肝纤维化的实验观察[J].中国中医基础医学杂志.2006;12(6):420-422.
[226].孙玉凤,冯志杰.黄芪抗肝纤维化的实验研究[J].河北中医药学报.2008;23(1):9-10.
[2].Pittenger MF,Mackay AM,Beck SC,et al.Multilinage potential of adult human mesenchymal stem cells.Scienses,1999;284(5411):143-147.
[3].Reyes M,Verfaillie CM.Characterization of multipotent adult progenitor cells,a subpopulation of mesenchymal stem cells.Ann New York Acad Sci,2001;938:231-235.
[4].Guo ZK,Yang JQ,Liu xd,et al.Biological features of mesenchymal stem cells from human bone marrow.Chin Med J,2001;114:950-953.
[5].Weissman IL.Translating stem and progenitor cell biology to the clinic:barriers and opportunites.Science,2000;287:1442-1446
[6].Lin H.Annu Reu Genet,1997;31:455-491.
[7].Cmnpognoli C,Roberts IA,Kumar S,etal.Identification of mesenchymal stem-progenitor cells in human first trimester fetal Deans RJ,Moseley AB.Mesenchymal stem cells:biology and potential clinical uses.Exp Hematol,2002;28(8):875.
[8].Deans RJ,Moseley AB.Mesenchymal stem cells:biology and potential clinical uses.Exp Hematol,2002;28(8):875
[9].傅文玉,路燕蒙,朴英杰.人骨髓间充质干细胞培养及多能性研究中国血液学杂志.2002:23(4):202-204.
[10].Gross TJ and Hunninghake GW.N Engl J Med,2004;345,517-525.
[11].姚青等,大鼠骨髓间充质干细胞体外诱导分化为心脏起搏样细胞的研究.中国心脏起搏与心电生理杂志,2007;21(1):57-58
[12].曹欣欣等,人骨髓间充质干细胞向心肌样细胞分化.基础医学与临床,2007;27(2):159-160
[13].Wang JS,Shum-Tim D,Augelli BJ,et al.Marrow stromal cells for celular cardiomyoplasty:feasibility and potentialclinical advantages.J Thorc Cardiovaso Surg,2000;120(5):999
[14].Wakitani S,Saito T,Caplan AI.Myogenic cels derived from rat bone marrow mesenchymal stem cells exposed to azacytidine.Muscle Nerve,1995;18:417.
[15].丁曷东等,心肌梗死后骨髓间充质干细胞移植对心室肌细胞复极活动的影响. 中华心律失常学杂志,2007;(11)2:136-137.
[16].Orllc D.Kajstura J,Chimenti S,et al.Nature,2001;4:339-351.
[17].王韫芳,裴雪涛.成体干细胞:从基础到临床.中华医学杂志,2004;16(84):1327-1328.
[18].赛因其木格等,大黄素诱导人骨髓问充质干细胞向成骨细胞分化的研究.中国中西医结合外科杂志,2006;12(5):447-448.
[19].Kadiyala s,Young RG,Thlde MA.Culture expanded canine mesenchymal stem cells possess osteochondrogenic potential in vivo and in vitro.Cell Transplant,1997;6:125.
[20].Pittenger MF,Mackay AM,Beck SC,etal.Multi—line.age potential of aduh human mesenchymal stem cells.Science,1999;284(5411):143.
[21].Yoo JIJ,Barthel TS,Nishimum K,eI al.The chondro genic potential ofh.man bone marrow-derived mesenchymal pro-genitor cells.J Bone Joint Surg,1998;80(12):1745.
[22].李林等,人骨髓间充质干细胞的分离、培养和体外定向诱导成类软骨细胞的实验研究.齐齐哈尔医学院报,2007;28(3):275-277.
[23].Balfoth B,Weisser J,Stemkop F。 et al.Transplantation of allagraft chondmcytes embeded inagarosegelin to cartilage defects ofrabbits.Osteoarthritis Cartilage,1998;6:50.
[24].Worster AA,Nixon AJ,Brower-Toland BD.et al.Efect of transforming growth factor betaon chondrogenic diferentiation of cultured equine mesenehymal stem cells.Am J VetRes,2000;61(9):1003.
[25].Yan ZQChen YS.Li WJ el al.Treatment of osteonec rosis of the femoraI head by percutaneous decompression and autologous bone marrow mononuclear cellinfusion.Chin J TraumatoI 2006;9(1):265.
[26].Ji WF,Tong PJ,Zheng W B,et al.ExperimentaI study on treatment of femoral head necrosis with arteriaI perfusion of marrow stem cells.Zhongguo Xiufu Chongjian Waike Zazhi.2004;24(11):999-1002.
[27].Terada M,Namazaki T,Oka M,et,al.Bone marrow cells adopt the phenotype of othercells by spontaneous cell fusion.Nature,2002;416:542-545.
[28].Ying QY,Nichols J,Evans EP,et al.Changing potency by spontaneous fusion.Nature,2002;416:545-574.
[29].Woodbury D,Schwarz EJ,Prockop DJ,et al.Adult rat and human marrow stromal cells differentiate in—to neurons[J].J Neurosci Res,2000;61:364-370.
[30].苏平等,银杏内酯 B 对骨髓间充质干细胞分化为神经样细胞的影响.广东医学,2007;28(1):33-34.
[31].肖庆忠,李浩威,温冠媚等.麝香多肽体外诱导成年大鼠和人诱导骨髓间充质干细胞定向分化为神经元的研究.中国生理病理杂志,2002;18(10):1179-1182.
[32].刘金保,董晓先,董燕湘等.多种中药诱导大鼠骨髓间质干细胞转变为神经元样细胞.中国药物与临床,2003;3(3):234-236
[33].Ohta M,Suzuki Noda et al.Bone marrow stromal cells infused into the cerettospinal flfid promote ftmctioml recovery of the injured rat spinal cord wiht reduced cavity formation.Exp Neurol,2004;187:266-278.
[34].Malmaood Lu D.Qu C,et al.Long-term recovery after bone marrow stromal cell treatmem of tmmatie train injury in rats.J Neurosurg,2006.104:272-277.
[35].MarinaM andBarbaraI.SocNephrol.2004;15:1794-1804.
[36].张中华等,髓间充质干细胞体内分化为肾小管上皮细胞的实验研究.江西医学检验,2007;25(1):15-17.
[37].赵嫒等,大鼠骨髓间充质干细胞分离培养及向脂肪细胞分化的实验研究。四川解剖学杂志,2007;15(1):31-32.
[38].刘弘光等,移植骨髓间充质干细胞治疗大鼠糖尿病的研究.中国组织化学与细胞化学杂志.2007;16(1):104-105.
[39].OH SH,Miyazaki M,Kouch H,et al.Hepatocyte growth factor induces differentiation of adult rat bone marrow cells into a hepatocyte lineage in vitro[J].Bioch Bioph Res commun,2000;279(2):500-504.
[40].Avital I,Inderbitzin D,Aoki T,et al.Isolation,characterization,and transplantation of bone marrow derived hepatocyte stem cells[J].Biochem Biophys Res Commun,2001;288(1):156-164.
[41].蔡云峰,闵军,何劲松等.骨髓源性肝干细胞的确认及定向分化的实验研究[J].中国普通外科杂志,2003;12(4):287-290.
[42].何忠杰,方驰华,马俊勋等.肝细胞生长因子与表皮细胞生长因子联合诱导大鼠骨髓间充质干细胞分化为类肝细胞[J].解放军医学杂志,2006;5(31):446-449.
[43].Wang P P,Wang J H,Yan Z P,et al.Expression of hepatocyte like phenotypes in bone marrow stromal cells after HGF induction[J].Biochem Biophys Res Commun,2004;320(3):712-716.
[44].Lee KD,Kuo TK,Whang-Peng J,Chung YF,Lin CT,Chou SH,Chen JR,Chen YP,Lee OK.In vitro hepatic differentiation of hum an mesenchvm al stem cells[J].Hepatology,2004;40:1275-1284.
[45].Theise ND,Nimmakayalu M,Gardner R,et al.Liver from bone marrow in humans[J].Hepatology,2000;32(1):11-16.
[46].陆海华等.小鼠骨髓间充质干细胞体内定向诱导分化与治疗肝功能损伤的实验研究[J].中华医学杂志,2007;87(4):223-224.
[47].Sato Y,Araki H,Kao J,et al.Human mesenchymal stem cells xenografted directly to rat liver are differentiated into human hepatocytes without fusion.Blood[J],2005;106:756-763.
[48].Oyagi Hirose M,Kojima M,Okuyama M,KawaseM,Nakamura T,Ohgushi H,Yagi K.Therapeutic effect of transplanting HGF-treated bone marrow mesenchymal cells into CCl_4-inlured rats[J].Hepatology 2006;44:742-748.
[49].Hung SC,Lu CY,Shyue SK,Liu H C,H o LL.Lineage differentiation-associated loss of adenoviral susceptibility and Coxsackie-adenovirus receptor expression in human mesenchymal stem cells.Stem Cells 2004;22:1321-1329
[50].TsuiTY,Lau CK,Ma J,Glockzin G Obed A,Schlitt HJ,Fan ST.Adeno-associated virus-mediated heme oxygenase-gene transfer suppresses the progression of micronodular cirrhosis in rats[J].World Gastroenterology,2006;12:2016-2023.
[51].Tcrai S.Sakaida I.Yamamoto N.et al.An in viva model for moni-toring trans-diffcrcntiation of bone marrow cells into functional hepatocytes[J].[J].Biochem,2003;134(4):551-558.
[52].Yamamoto N.Tcrai S.Ohata S.et al.A subpopulation of bone marrow cells depleted by a novel antibody.anti-Liv8 is useful for cell therapy to repair damaged liver[J].Biochem Biophys Res com,2004;313(4):1110-1118.
[53].Miyazakit,Studies on on fungal polysaccharides XX.Galactom annan Of Cordyceps sinesis[J].Chem Pham Bull,1977;25(12):3325-3328.
[54].龚敏,朱勤,王彤等.冬虫夏草多糖的分子结构与免疫活性[J].生物化学杂志,1990;6(6):486-492.
[55].李楠,宋建国,刘金云等.蛹虫草与冬虫夏草化学成分比较[J].吉林农业大学学报.1995;17(增刊):80-83.
[56].白云娥,李青山,王毅等.冬虫夏草多搪的含量测定[J].山西医科大学学报,2000:31(2):129-130.
[57].Ng TB.Tranditional Chinese medicine and the search for new antineoplastic drugs.Life Science[J],1997;60(25):2349.
[58].郭风彩.冬虫夏草多糖的药理学研究进展[J].现代医药卫生,2006;22 (7):999-1000.
[59].刘超,卢珊,姬美蓉等.冬虫夏草对白血病 NK 细胞影响的体外研究[J].中国中西医结合杂志,1992;12(5),267-269.
[60].张椒兰,孙云汉,刘晓平,等.冬虫夏草及人工虫草菌丝抗小鼠 Lewis 肺癌的研究[J].中药通报,1987;12(2):53.
[61].刘湘.从冬虫夏草得到的蛋白多糖0909具有抗肿瘤和免疫促进活性[J].国外医药·植物药分册,1995;10(2):91.
[62].龚敏,朱勤,王彤,等.虫草多糖的分子结构与免疫活性[J].生物化学杂志,1990;6(6):486-491.
[63].刘晓平.冬虫夏草及人工虫草菌丝对 Y 线照射后小鼠血小板和免疫器官的影响[J].中药通报1988;13(4):44.
[64].龚晓健,季晖.人工冬虫夏草对小鼠免疫功能的影响[J].中国药科大学报,2000;31(1):53
[65].程琪琳,邱德凯,张泳.虫草脂质体对小鼠巨噬细胞的作用[J].上海免疫学杂志,1989;9(3):143.
[66].靖大道,邱德凯,萧树东等.虫草多糖对人外周血 IL-2,IL-2R 及 IFN-r 调节作用的研究[J].上海免疫学杂志,1995;15(6):321-323.
[67].张建,韦星呈等.北虫草多糖对人肿瘤坏死因子α、白细胞介素-2及其可溶性受体水平的影响[J].中国实用内科杂志,1997;17(12):727-728.
[68].孙荣玲、李国忠复方虫草多糖脂质体口服液对慢性乙型肝炎细胞免疫的影响[J].中国药师.2005;8(10).848-849.
[69].夏宝森,李晓康.FTY720:一个来源于冬虫夏草的新型免疫抑制剂[J].上海免疫杂志2002;22(2):79-81.
[70].匡彦德,王美英,钱琴芳等.冬虫夏草免疫增强作用机理的研究.上海免疫学杂志1989;9(1):6.
[71].许维祯,李丽芬,石扣兰等.冬虫夏草菌丝体对单氨氧化酶及免疫功能的影响[J].上海中医药杂志1988;(1):48.
[72].陈道明,张淑兰,于志洁等.冬虫夏草及人工培养菌丝对小鼠血小板生成的影响及超微结构观察[J].中药通报,1987;12(1):47.
[73].祝希嫒,史勇,刘晓明等.人工培养冬虫夏草菌粉对细胞免疫的抑制作用[J].中西医结合杂志,1990;10(8):485-486.
[74].夏益平.冬虫夏草口服液治疗慢乙肝36例疗效观察[J].右江医学,2000;28(2):91-92.
[75].季晖等.人工虫草菌丝多糖的分离提取及其降血糖作用研究[J].中国药科大学学报,1993;24(1)39-42.
[76].黄志江,季晖,李萍.人工虫草多糖降血糖作用及其机制研究.中国药科人学学报,2002;33(1):51-54.
[77].Kiho T.Hypoglycemic activity of a cs-F30 from the cultural mycelium of Cordyceps sinensisand its effecton glucosemetabo-lism in mouse liver.Biolpharm.Bul[J],1996;19(2):294.
[78].许建明,丁长海,李连德,等.冬虫夏草多糖保护小鼠免疫性肝损伤的筛选研究[J].安徽医科大学学报,1999;34(3):173-175.
[79].刘玉凤,李彩霞.冬虫夏草多糖脂质体治疗慢性乙型肝炎86例临床观察[J].山西中医,1998;14(1):15-16.
[80].赵立群.复方冬虫夏草多糖脂质体口服液治疗肝炎后肝硬化120例疗效观察[J].中医药学刊.2005;23(7).1300-1300.
[81].马雄,邱德凯,徐军.虫草多糖脂质体抗肝纤维化的实验研究[J].中国中医基础杂志,1999;5(9):28-30.
[82].颜吉丽等.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-α表达的影响[J].复旦学报:医学版.2003;30(1):27-29.
[83].杨俊何,凌耀生.虫草多糖的抗氧化作用[J].广东药学院学报,1997;13(1):35-38.
[84].李泓俊,彭欣辉,吴应莲.虫草多糖脂质体对小鼠免疫性肝纤维化TGF-β_1mRNA 表达的影响[J].临床医学研究,2004;21(5):500-503.
[85].李风华、刘平等,虫草多糖逆转DMN诱导大鼠肝纤维化的作用及机制研究[J].中国中药杂志,2006;31(23):1968-1971.
[86].马雄,邱德凯,徐军.虫草多糖脂质体抗肝纤维化的实验研究[J].中国中医基础杂志,1999;5(9):28-30.
[87].方步武,刘平,刘成,等.虫草多糖抗免疫损伤性大鼠肝纤维化的作用及其机理研究[J].上海中医药杂志,2000;(9):37-40.
[88].方士英,虫草多糖对小鼠化学性肝损伤的保护作用[J].安徽医科大学学报.2004;39(3):201-204.
[89].曾宪可,唐玉,袁盛榕.冬虫夏草和中国拟青霉对小鼠免疫性肝损伤的保护作用[J].中国药学杂志,2001;36(3):161.
[90].张小强.冬虫夏草与人工虫草菌丝体对四氯化碳所致急性化学性肝损伤的保护作用[J].环境与职业医学,2003;20(6):422-426.
[91].夏益平.冬虫夏草口服液治疗慢乙肝36例疗效观察[J].右江医学.2000;28(2).91-92.
[92].Sato M,Suzuki S,Senoo H.Hepatic stellate cells:Unique characteristics in cell biology and phenotype[J].Cel Struct Funct,2003;28(2):105-112.
[93].Gressner AM,Weiskirchen R,Breitkopf K,et al.Roles of TGFβ betain hepatic fibrosis[J].Front Biosci,2002;7:793-807.
[94].王萍.血小板衍生生长因子与肝纤维化[J].国外医学.消化疾病分册,2000;20(1):40-43.
[95].Holmes A,Abraham DJ,Sa Set al.CTGF and SMADs,maintenanee of seleroderma phenotype is independent of SMADsignaling[J].J Biol Chem,2001;276(14):10594-10601.
[96].彭小东,戴立里.结缔组织生长因子与肝纤维化的研究进展[J].胃肠病学与肝病学杂志,2005;14(4):425-27.
[97].Saxena N K,Titus M A,Ding X,et al.Leptin as a novel profibrogenie cytokine in hepatic stellate cells:mitogenesis and inhibition of apoptosis mediated by extracellular regulated kinase(ErK)and Akt phosphory lation[J].FASEB J,2004;8(13):1612-1614.
[98].Cao Q,Mak K M,Lieber C S,et al.Leptin enhances alphal(I) collagen gene expression in LX-2 human hepatic stellate cellsthrough JAK-mediated H-2O_2 dependent MAPK pathways[J].J Cell Biochem,2006;97(1):188-197.
[99].何生松,徐标,韩春荣.环醇对小鼠日本血吸虫病肝纤维化即早基因与 TGF-β1、TIMP1及胶原表达的影响[J].世界华人消化杂志,2007;15(35):3678-3684.
[100].Wu JY,Wu YB,et al.Effects of endothelin-1 On hepatic stellate cell proliferation,collagen synthesis and secretion intracellcular flee calcium concentration[J].World J Gastroenterol,2004;10(18):2697-2700.
[101].Bataller R,Gines P,Nicolas JM,et al.Angiotensin Ⅱ induces contraction and proliferation of human hepatic stem cells[J].Gastroenterology,2000;118(6):1149-56.
[102].Bataller R,Sancho-Bru P,Gines P,et al.Activated human hepaticsteHate cells express the rennin-an giotensin system and synthesize angiotensin [J].Gastroenterology,2003;125(1):117-25.
[103].Balk SK,Jo Hs,Suk KT,etal.Inhibitory efect ofangiotensin Ⅱ receptor antagonist on the contraction and growth of hepatic stellate cells[J].Korean J Gastroenterol,2003;42(2):134-41.
[104].Kurikawa N,Suga M,Kuroda S,et al.An angiotensin Ⅱ type 1 receptor antagonist,oimesartanmedoxomil,improves experimental liver fibrosis by suppression of proliferation and collagen synthesis in activated hepatic steHate cells[J].Br J Pharmacol,2003;139(6):1085-94.
[105].李小刚.细胞因子网络在肝损害中作用的演剧进展[J].国外医学·内科学分 册,1997;24(1):20.
[106].张文胜,王宝恩,王泰龄,等.慢性乙型肝炎纤维化无创性诊断模型的建立[J].中华肝脏病杂志,2006;14:169-173.
[107].Itagaki T,ShimizuI,ChengX,et al.Opposing efect s of oes-tradiol and progesterone On intraeellular pathways and aetivationproeesses in the Oxidative 8 tress induced activation of cultured rat hepatic stellate celJs.Gut.2005;54:1782-1789.
[108].Novitskiy G,Potter J J,Wang L,et al.Influences of reactive oxygen species and nitric oxideon hq-icfibrogenesis.Liverht,2006;26:1248-1257.
[109].BatallerR,BrennerDA.Hepatic stellate cells as a target for the treatment of liver fibrosis[J].S-nin Liver Dis,2006;21:437-451.
[110].Ziol M,Handra2Luca A,Kettaneh A,et al.Noninvasive assessment of livel fibrosis by rrleasuH nent of-ifness in patients with chronic hepatitis [J].Hepatologv,2005:41:48-54
[111].中华医学会肝病学分会脂肪肝和酒精性肝病学组.非酒精性脂肪性肝病诊疗指南[J].中华肝脏病志,2006;14:161-163.
[112].CoiliA,Fraquelli M,Andreoletti M,etal.Severe liverfibrosisor cirrhosis:accuracy of US for detection analysis of 300 cases [J].Radiology,2003;227:89-94.
[113].Zheng RQ,Wang QH,Lu MD,et al.laver fibrosis in chronic viral hepatitis:an ultrasonographic study.World J Gastroenterol,2003;9:2484-2489.
[114].Aube C,Oberti F,Korali N,et al.UItrasonographic diagnosisof hepatic fibrosis of cirrhosis[J].J Hepatol,1999;30:472-478.
[115].Galani S Gramantieri L,Venturoli N,et al.W hat is the criterion for differentiating chronic hepatitis from compensated cirrhosis?A prospective study comparing uItrasonography andpercutaneous liver biopsy[J].J Hepatol,1997;27:979-985.
[116].Bernatik T,Strobel D,Hahn EC,et al.Doppler measurements:a surrogate marker of liver fibrosis[J].Eur J Gastroenterol Hepatol,2002;14:383-387.
[117].Taura T,Nakam ura K,Takashima S,et al.Heterogeneity of hepatic parenchymal enhancement on computed tomography during arterial portography:quantitative analysis of correlation with severity of hepatic fibrosis[J].Hepatol Res,2001;20:192-192.
[118].陈克敏,柴维敏.肝纤维化患者肝脏形态学改变的 CT、MR 评价肝脏[J].2000;5(4):209-210.
[119].van Beers BE,Leconte I,M aterne R,et al.Hepatic perfusion parameters in chronic liver disease:dynamic CT measurements correlated with disease severity[J].Am J Roentgenol,2001;(176):667-673.
[120].Materne R,Annet L,Dechambre S,et al.Dynamic computed tomography with low and high nmolecular mass contrast agents to assess microvascular permeability modifications in a model of liver fibrosis[J].Clin Sci(Lond).2002;1(13):213-216.
[121].Cho SG,Kim MY,Kim J,etal.Chronic hepatitis:invivo proton MR spectroscopic evaluation of the liver and correlation with histopathologic findings[J].Radiology,2001;221(3):1740.
[122].Rouvire O,Yin M,Dresner M A,et al.M R elastography of the liver:preliminary results[J].Radiology,2006;240(2):440.
[123].李晶,张菁.重组α-干扰素治疗慢性乙型肝炎肝纤维化46例疗效观察[J].临床和实验医学杂志.2006;5(10):1546-1547.
[124].张岩,白雪帆,李艳红等.拉米夫定治疗慢性乙型肝炎的疗效及肝组织学改变[J].中国误诊学杂志,2006;5(5):804-806.
[125].Leung NW,Lai CL,Chang TT,et al.Extended lamivudine treatment in patients with chronic hepatitis B enhances hepatitis B antigen seroconversion rates:results after 3 years of therapy[J].Hepatology,2001;33(6):1527-1532.
[126].Bataller R,Brenner DA.Hepatic stellate cells as target for the treatmentn of liver fibrosis[J].Semin Liver Dis,2001;21(3):437-45.
[127].谭海荣,吴谦,潘竞锵,等.水飞蓟宾对抗大鼠酒精性脂肪肝作用及机制[J].中国临床药理学与治疗学,2004;9(7):803-806.
[128].Boigk G,Stroedter L,Herbst H,et al.Silymarin retards collagen accumulationin early and advanced biliary fibrosis secondary to complete bile duct obliteration in rats[J].Hepatology,1997;26(3):643-649.
[129].许倩,许建娟.腺苷蛋氨酸治疗妊娠肝内胆汁淤积症疗效探讨[J].中国优生与遗传杂志,2004:12(1):83-84.
[130].汪余勤,程五风,李宣海.维生素 E 与肝纤维化[J].临床肝胆病杂志,2000;16(1):19.
[131].Dogru Abbasoglu S,Balkan J,Kanbagli O,et al.Aminoguanidineian inducible nitric oxide synthase inhibitor,plus N-acetylcysteine treatment reduce the lipopOlysaccharide-augmented hepatotoxicity in rats with cirrhosis[J].Hum Exp Toxicol,2002;21(7):359-364.
[132].Vendemiale G,Grattagliano I,Caruso ML,et al.Increased oxidativestress in dimethylnitrOsamine-induced liver fibrosis in the rat:effect of N.acetylcysteine and interferon-alpha[J].Toxicol Appl Pharmaeol,2001;175(2):130-139.
[133].Meurer SK,Lahme B,Tihaa L,et al.N-acetyl-L-cysteine suppresses TGFβ a signaling at distinct molecular steps:the biochemical and biological efficacy of a muhfunetional,antifibrotie drug[J].Biochem Pharmacol,2005;70(7):1026-1034.
[134].Arrieta O,Rodriguez JL,Rosaa V,et al.Colchicine delays the development of hepatocellular carcinoma in patients with hepatitis virus-related liver cirrhosis[J].Cancer,2006;107(8):1852-1858.
[135].Poupon RE,Huet PM,Poupon R,et al.A randomized trial corapa-ring colchicine and ursodexyeholie acid combination to ursodeoxycholic acid in primary billiary cirhosis[J].UDCA—PBC Study Group.Hepatology.1996;24(5):1098-1103.
[136].程书权.肝纤维化的现代药物治疗研究[J].中国处方药,2005;3(5):36-40.
[137].阳学风,曾明新,张明亮等.马洛替酯抑制肝纤维化的实验与临床研究[J].世界华人消化杂志,1999;7(3):224-226.
[138].何方平,孙惠蓉,张跃新等.四种药物对大鼠肝纤维化防治作用的实验研究[J].新疆医科大学学报,2003;26(4):338.
[139].LieberCS.New concepts of thepathogenesis of alcoholic liver disease lead to novel treatments[J].Cur Gastroenterol,2004;6(1):60-65.
[140].Wamoto H。 Nakamuta M,Tada S,et al.Platelet-derived growth factorreceptor tyrosine kinase inhibitor AG1295 attenuates rat hepatic stelate cell growth[J].Lab Clin Med,2000;135(5):406.
[141].Cut X,Shimizu 1,Lu G,et al.Inhibitory efect of a soluble transforming growth factor beta type Ⅱ receptor on the activation of rat hepatic stellate cells in primary culture.Hepar,2003;39(5):731.
[142].齐洪军,胡蔓菁.从“痰瘀”角度浅析肝纤维化的病因病机及治疗[J].四川中医,2004;22(6):26-28.
[143].刘为民,姚乃礼.络病理论与肝纤维化临床[J].中医药学报,2003;31(1):2-3.
[144].叶放,薛博瑜,周珉,等.论湿热瘀毒与肝纤维化[J].南京中医药大学学报,2005;21(6):346-349.
[145].唐世利,刘其政,王艳妮.化瘀软肝汤治疗肝纤维化70例[J].陕西中医,2002:23:588-589.
[146].程明亮,付荣泉.中药复方对猪血清免疫性肝纤维化肝中氧化及抗氧化的影响[J].中华肝脏病杂志,2006;14(3):217-218.
[147].赵晓晨,张琳.复方丹参注射液抗肝纤维化的疗效观察[J].山西中医学院学报,2007;7(6):38.
[148].齐洪军,胡曼青,王长松,等.祛瘀化痰汤抗大鼠肝纤维化的作用及其机制[J].江苏中医杂志,2003;24(7):55-57
[149].方谦和,孙维娜.方谦和和解法治疗肝纤维化的经验[J]北京中医,2004:23(3):143-144.
[150].肖会泉,刘妮,杨俭勤,等.肝舒胶囊对慢性乙型肝炎肝纤维化的临床疗效观察[J].山东中医杂志,2006;25(10):663-665.
[151].褚裕义,孙克伟,刘伟士,等.疏肝理脾片抗大鼠免疫性肝纤维化研究[J].中国中西医结合脾胃杂志。1998;6(2):98.
[152].龙爱华,陈瑾,王树槐.抑肝健脾复方抗大鼠肝纤维化的实验研究[J].中西医结合肝病杂志,2002;12(1):23-25.
[153].刘爱丽,魏敏,甑增国,等.小柴胡汤凯西莱联用抗肝纤维化50例[J].陕西中医,2005;26(9):873-874.
[154].郭晓华,王宜芳.复肝片治疗慢性乙型肝炎肝纤维化的临床观察中成药.2006;28(12):1760-1762.
[155].李新华滋阴养肝汤治疗慢性乙型肝炎后肝纤维化的临床观察上海中医药杂志.2006;40(11):14-15
[156].蔡春江,褚志敏,崔书芬,等.柔肝方治疗慢性乙肝肝纤维化的临床研究[J].四川中医,2006;24(4):43-45.
[157].郭晓萍,侯允天,李东良,等.柔肝煎冲剂对实验性肝纤维化大鼠血清透明质酸及Ⅲ型前胶原的影响[J].中医药学报,1996;12(5):42.
[158].叶放,薛博瑜,周眠,等.论湿热瘀毒与肝纤维化[J].南京中医药大学学报,2005;21(6):346-349.
[159].于世瀛,贲长恩,杨美娟,等.清热利湿方药抗肝纤维化的形态学和免疫组织化学定量研究[J].北京中医药大学学报,1997;20(1):28-30.
[160].李日向,常占杰,叶青.清木丹颗粒治疗慢性乙型肝炎湿热留恋瘀毒型肝纤维化的临床研究[J].陕西中医学院学报,2005;28(2):8-9.
[161].朱清静,聂广.剔毒护肝方抗鸭乙型肝炎肝纤维化的作用.中西医结合肝病杂志.1998;8(2):84-87.
[162].李明奎.芪茜化纤汤治疗肝纤维化37例临床观察[J].山西中医学院学报, 2006;7(4):28.
[163].李召忠,王啸,隋在云,等.血隆冲剂抗肝纤维化的临床与实验研究[J].中国中西医结合杂志,2001;21(11):813-815.
[164].唐苾芯,李金华,黄存垣.益肝消癥散抗慢性肝炎肝纤维化临床研究[J].江西中医学院学报.2004;16(2).40-43.
[165].王晓峰,董培红,陈威,等.汉防己甲素对脂多糖刺激的大鼠肝星状细胞增殖的影响[J].现代实用医学,2006;18(7):452-454
[166].Chor SY,Hal AY,To KF,et al.Anti-proliferative and pro-apoptotic effects of herbal medicine on hepatic stellate cell[J].Ethnopharmacol.2005;100(1-2):180-186.
[167].徐永红,边城,李定国,等.汉防己甲素对成纤维细胞增殖及胶原合成的作用[J].天津医药,2004;32(10):633-635.
[168].权启镇,王要军,权源,等.复方汉防己冲剂抗肝纤维化的实验研究,中华肝脏病杂志,2007;15(11):873-87.
[169].周光德,李文淑,赵景民等.复方鳖甲软肝片抗肝纤维化机制的临床病理研究.解放军医学杂志,2004;29(7):563-564.
[170].王晓玲,崔云华,胡旭东等.丹酚酸 B 对大鼠肝星状细胞增殖周期的抑制作用.中华消化杂志,2004;24(1):59.
[171].胡义扬,王润平,赵长青,等,丹酚酸 A 对实验性大鼠肝纤维化Ⅰ型胶原及其基因表达的影响[J].中国中医药科技,1999;6(4):235-237.
[172].胡义扬,王润平,张学军,等.体外四氯化碳损伤肝细胞对肝星状细胞活化的影响及其丹酚酸 A 的干预效果[J].中华肝脏病杂志,2000;8(5):299-301.
[173].王志凌,成军,刘妍,等.甘草甜素抑制肝星状细胞增殖作用的研究.肝脏,2005:10(3):225-226.
[174].贾道全,张正,罗成福,等,甘草甜素逆转肝纤维化及早期肝硬化的作用探讨[J].中华消化杂志,2001;21(12):754-756.
[175].王吉耀,郭津生,刘淑玲,等.甘草甜素对肝硬化动物模型肝脏内 NF-KB 结合活性的抑制作用[J].中华肝脏病杂志,1999;7(1):42-43.
[176].郭津生,王吉耀,宋后燕,等,强力宁对大鼠肝硬化肝脏内肝细胞生长因子表达的影响.中华肝脏病杂志,1999;7(2):112.
[177].崔德广,李素云,王生耀.干扰素 alb 联合苦参碱治疗肝炎肝纤维化40例[J].中国新药杂志,2007;16(4):314-316.
[178].卢清,张清波,张继明,等.氧化苦参碱对大鼠肝星状细胞旁分泌活化途径的抑制作用[J].肝脏,2004;9(1):31-33.
[179].黄瑾,胡晋红,邱磊等.阿魏酸钠抑制肝星状细胞合成胶原的机制.药学学 报,2004;38(8):577-580
[180].吴建红,陕光,张端莲,等.川芎嗪抗实验性肝纤维化组织中 TGF-β1阳性面积率表达的比较[J].数理医药学杂志,2006;19(3):237-238.
[181].谈博,宋健平,张奉学,等.川芎嗪对 HSC-T6细胞 smad 蛋白细胞内转位的影响[J].中药新药与临床药理,2006;17(5):320-322
[182].Lucena MI.Andrade lad.de la Cruz JP.et al.Effects of silymatin MZ-8on oxidative strcss in paticnts with alcoholic cirrhosis,Rcsults of arandomized Doudle-blind,placebo-controlled clinical study.Int Clin Pharmacol Ther,2002;40(1):2-8.
[183].徐列明,刘平,刘成,等.桃仁提取物合并虫草菌丝抗肝纤维化的实验研究[J].中国中医药科技,1995;2(1):18-20.
[184].王文兵,载立里,郑元义等,三七总皂苷对大鼠肝纤维化防治机制[J].世界华人消化病杂志,2004;12(8):1939-1942.
[185].展玉涛,刘宾,李定国,等.大黄素抗肝纤维化的作用机制[J].中华肝脏病杂志,2004:12(4):245-246.
[186].Akamatsu S,Watanabe A,Tamesada M,et al.Hepatoprotective effect of extracts from Lentinusedodes mycelia on dimethylnitrosamine-induced liver injury[J].Biol Pharm Bull,2004:27(12):1957-1960.
[187].Van de Casteele M,Roskams T,Van der Elat I,et al.Halofuginonecan worsen liver fibrosis in bile ductobstructed rats[J].Liver Int,2004;24(5):502-509
[188].张锡流,梁健,杨光业,等.牛磺酸对大鼠肝纤维化组织Ⅲ型胶原的影响[J].中国中西医结合消化杂志,2005;13(4):245-247.
[189].黄琳芸,钟鸣,杨增艳,等.排钱草总生物碱对肝纤维化大鼠血清干扰素和肝脏组织病理学的影响[J].中国中医药科技,2006;13(2):101-102.
[190].Petersen BE.Bone marrow as a potential source of hepatiC oval cellS[J].Science,1999;284:1168-1170.
[191].王要军,孙自勤,权自镇等.冬虫夏草对大鼠实验性肝纤维化的防治作用及其机理研究[J].中国中药杂志,1996;21(3):179-181.
[192].Niklason LE,G ao J,AbbottW M,etal.Functionalarteries grown in vitro.Science 1999;284(5413):489-493.
[193].Campagnnoli C,Roberts lAG,Kumar S,et al.Identification of mesenchymal stem/progenitor cel ls in humans first-trimester fetal blood,liver,and marrow[J].Blood,2001;98:2396-2402.
[194].Martinez C,Hofmann T J,Marino R,et al.Human bone marrow mesenchymal stromal cells express the neural ganglioside GD2:a novel surface marker for the identification of MSCs[J].Blood,2007;Epub ahead of print.
[195].Gang E J,Bosnakovski D,Figueiredo C A,et al.SSEA-4 identifies mesenchymal stem cells from bone marrow[J].Blood,2007;109(4):1743-1751.
[196].Skarpen E,Oksvold MP,Grosvik H,et al.Altered regulation of EGF receptor signaling following a partial hepatectomy[J].J Cell Physiol,2005;202(3):707-716
[197].RE Schwartz,M Reyes,L Koodie,et al.Muhipotent adult progenitor cellsfrom bone marow diferentiate into functional hepatocyte-like cells[J].ClinInvest,2002;109(10):1291-1302.
[198].鄂征·组织培养和分子细胞学技术[M]·第1版·北京:北京出版社.
[199].刘树辉,曹中伟,秦书俭,等.SD 大鼠骨髓间充质干细胞的分离培养方法的研究[J].锦州医学院学报,2006;27(2):35-36.
[200].张刚庆,方驰华,池达智.肝细胞生长因子诱导骨髓间充质干细胞向肝细胞分化的实验研究[J].中华外科杂志,2005;43(11):716-720
[201].Xin-Qin Kang,Wei-Jin Zang,Tu-Sheng Song,et al.Rat bonemarrow mesenchymal stem cells differentiate into hepatocytes in vitro[J].World J Gastroenterol,2005;1:3479-3481.
[202].Terada M,Namazaki T,Oka M,et,al.Bone marrow cells adopt the phenotype of other cells by spontaneous cellfusion[J].Nature,2002;416:542-545.
[203].Zaret KS.H epatocyte differentiation:from the endoderm and beyond[J].Curr Opin Genet Dev,2001;11:568-574.
[204].Jung J,Zheng M,Goldfarb M.et al.Initiation of mammalian liver development from endoderm by fibroblast growth factors[J].Science,1999;284:1998-2003.
[205].Kamiya A,Kinoshita T,Miyajima A.Oncostatin M and hepatocyte growth factor induce hepatic maturation via distinct signaling pathways [J].FEBS Lett,2001;492:90-94
[206].Uehara Y,Mori C,Noda T,et al.Rescue of embryonic lethality in hepatocyte growth factor/scatter factor knockout mice[J].Genesis,2000;27:99-103.
[207].Okumoto K,Saito T,Hattori E,et al.Diferentiation of bone marow cells into cells that express liver-specific genes in vitro:implication of the Notch signals in differentiation[J].Biochem Biophys Res Comman,2003;304(4):691-695.
[208].kumoto K,Saito T,Hattori E,et al.Expression of Notch signalling markers in bone maiTow cells that differentiate into a liver cell lineage in a rat transplant model[J].Hepatol Res,2005;31(1):7-12.
[209].Schwartz RE,Reyes M,Koodie L,et al.Muhipotent adult progenitor cells from bone maiToW differentiate into functional hepatocyte-like cells[J].J Clin Invest,2002;109(10):1291-1302.
[210].Saji Y,Tamura S,Yoshida Y,et al Basic fibroblast growth factor promotes the trans diferentiation of mouse bone marow cells into hepatie lineage cells via multiple liver-enriched transcription factors[J].J Hepatol,2004;41(4):545-550.
[211].Avital I,Inderbitzin D,Aoki T,et al.Isolation,characterization,and transplantation of bone marrow-derived hepatocyte stem cells[J].Biochem Biophys Res Commun,2001;288:156-164.
[212].Suzuki A,lwama A,Miyashita H,et al.Role for growth factors and extracellular matrix in controlling differentiation of prospectively isolated hepatic stem cells Development,2003;130:2513-2524.
[213].黎晖,周健洪,陈东风,等.龟板对大鼠骨髓间充质干细胞向成骨分化的 影响[J].中药新药与临床药理,2005;3(16):159-161.
[214].徐展望,张建新,谭国庆,等.中药骨碎补提取液对兔骨髓基质细胞体外成骨分化的影响[J].中医正骨,2006;6(18):15-16.
[215].陈杰,陈新月.肝纤维化诊断研究进展[J].国际流行病学传染病杂志,2006;3(6):414.
[216].Oberti F.Valseda E.Piletle C,et al.Noineasive diagnosia of hepeticfibrosis or cirrhosis[J].Castroenterology.1997;113(5):1609-1611.
[217].王静艳,高红,王兆荃,等.Ⅲ型前胶原层粘连蛋白在肝纤维化诊断中的意义[J].中国医科大学学报,1997;26(2):17.
[218].Obata K,Twata T,Tnoue K,et al.One step sandwich enzyme immunoassay for human type Ⅳ collagen using monodonalantibodies.Clin Dim Acta[J],1989;181:293.
[219].Okazaki I,Watanabe T,Hozawa S,et al.Molecularmechanism of the reversibility of hepatic fibrosis:with special reference to the role of matrix metalloproteinases[J].J Gastroenterol Hepatol,2000;15(Suppl):D26-32.
[220].黄字琦,高毅.大鼠肝纤维化基质金属蛋白酶及其抑制因子的表达[J].世界华人消化杂志,1999;7:795-796.
[221].潘爱萍,林英辉,潘元平等.肝纤维化血清学标志物与肝脏病理相关性研究[J].南方医科大学学报.2007;27(12).-1935-1936,1938.
[222].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.
[223].王究波,刘平.虫草菌丝提取物干预与治疗二甲基亚硝胺诱导大鼠肝硬化的实验研究[J].中国中西医结合杂志.2008;28(7):617-622.
[224].刘平,王晓玲.扶正与化瘀影响肝星状细胞活化、胶原生成及肝细胞白蛋白生成的配伍作用研究.上海中医药大学学报.1999;13(3):46-50.
[225].王继,陈俊红.活血、益气、软坚不同中医治则逆转大鼠肝纤维化的实验观察[J].中国中医基础医学杂志.2006;12(6):420-422.
[226].孙玉凤,冯志杰.黄芪抗肝纤维化的实验研究[J].河北中医药学报.2008;23(1):9-10.