肝卵圆细胞、肝脏局部微环境改变及Wnt5a表达与肝癌形成、转移的初步研究
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摘要
第一部分肝癌起源于肝卵圆细胞免疫组化研究及临床意义
目的有关原发性肝癌的细胞起源存在两种学说,即成熟肝细胞去分化学说和肝干细胞分化受阻学说,这两种学说目前尚有争议。本实验通过测定肝硬化、肝癌和癌旁组织肝干细胞标志的表达,观察肝干细胞标志的表达阳性细胞的细胞学形态,结合肝癌患者的临床病理特征,探讨肝脏卵圆细胞在肝癌形成中所扮演的角色及其与临床病理因素的相互关系。
方法应用免疫组化的方法,测定63例肝细胞肝癌及其癌旁组织、15例肝炎后肝硬化组织标本及10例正常肝组织标本C-KIT+、AFP、CK8/18、CK7和CK19的表达,用光学显微镜观察上述组织标本干细胞标记标染的细胞形态、分布,回顾性总结63例HCC患者的临床病理资料,分析HCC标本C-KIT+、AFP、CK8/18、CK7和CK19的表达与性别、血清HBsAg状态、HBcAb状态、血清AFP、肿瘤大小、肝内病灶数目、是否有包膜侵犯、肝癌细胞病理分化分级、肝功能评级、肝硬化程度、TNM分期及术后2年生存期等13个临床病理参数的关系。
结果63例HCC及其癌旁组织、15例肝炎后肝硬化组织标本及10例正常肝组织标本免疫组织化学染色显示,CK8/18标染阳性率分别是95.2%、96.8%,100%和100%,四组间CK8/18阳性表达率和表达强度评分差异均无显著性(x~2=1.274,P＞0.05;F=1.465,P＞0.05);CK7标染阳性率分别是46.0%、58.7%,26.7%和0,四组间CK7阳性表达率差异有显著性(x~2=14.86,P＜0.01),肝癌组织、癌旁组织CK7表达率均分别高于肝硬化、正常肝组织(x~2=6.887,P＜0.01;x~2=13.133,P＜0.01);CK7标染表达强度评分比较,肝癌组织、癌旁组织CK7阳性表达强度评分高于肝硬化组织及正常组织(F=21.26,P＜0.01);四种组织实质肝细胞和肿瘤细胞均未见CK19阳性表达;四组间C-KIT阳性表达率差异有显著性(x~2=25.089,P＜0.01),HCC、癌旁及肝硬化组织C-KIT阳性表达率显著高于正常组织,癌旁组织C-KIT阳性病例表达强度评分显著高于HCC、肝硬化及正常组织(F=36.19,P＜0.01);HCC组织AFP阳性率及表达强度评分均显著高于癌旁组织、肝硬化及正常组织(x~2=5.567,P＜0.05;F=15.76,P＜0.01)。HCC实质组织中可见符合肝卵圆细胞形态特点CK7、C-KIT及AFP阳性细胞,呈点状、片状或弥漫性分布,以癌结节边缘多见。HCC癌组织中,CK7阳性表达与肝癌病人的性别、血清HBsAg状态、HBcAb状态、血清AFP、肿瘤大小、肝内病灶数目、是否有包膜侵犯、肝癌细胞病理分化分级、肝功能评级、肝硬化程度、TNM分期及术后2年生存期等病理参数均无关(P＞0.05),;C-KIT阳性表达与血清AFP、肿瘤转移、肝癌细胞病理分级及肿瘤TNM分期有关(P＜0.01),和其余病理因素无关(P＞0.05);组织中AFP阳性表达与血清AFP、肿瘤转移、肝癌细胞病理分化分级、肿瘤TNM分期及术后2年生存期等参数有关(P＜0.01),和其余病理因素无关(P＞0.05)。
结论HCC及癌旁组织中存在表达肝干细胞标记表型并具有卵圆细胞形态的细胞,提示HCC的发生与肝脏干细胞/卵圆细胞密切相关,表达CK7、C-KIT和AFP的肝卵圆细胞参与了HCC形成;肝卵圆细胞表型标记CK7、C-KIT和AFP在HCC组织和癌旁组织有不同的表达规律,它们可能代表不同的干细胞群,在肝炎慢性损伤修复和癌变过程中具有不同作用;HCC及其癌旁组织干细胞表型标记表达具有多种临床意义;癌旁CK7和C-KIT表达与肝卵圆细胞异质性、增殖和组织修复有关,癌组织C-KIT和AFP表达预示着卵圆细胞由癌旁组织中的增殖修复功能向癌组织的恶性转化。
第二部分肝癌局部微环境改变与肝癌浸润、转移
目的研究细胞外基质tenascin(TN)在HCC(hepatocellular carcinoma,HCC)中的表达及HCC癌、癌旁组织微血管密度(MVD-CD105)分布,探讨TN对HCC肿瘤微血管生成、浸润、转移及临床预后的影响,MVD-CD105在HCC癌、癌旁组织分布规律、临床意义及CD105标染HCC组织微血管的特异性。
方法采用免疫组织化学方法检测63例HCC癌组织、癌旁组织及15例肝炎后肝硬化组织、10例正常肝组织中TN表达,同时检测63例HCC癌组织、癌旁组织微血管密度(MVD-CD105),分析TN表达与HCC病理学特点及肿瘤微血管生成的关系,比较HCC癌组织和癌旁组织MVD-CD105分布特点及其与临床参数的关系。
结果TN阳性率HCC癌组织65.1%,癌旁组织69.8%,肝硬化组织20.0%,正常肝组织无阳性表达;HCC癌组织及癌旁组织TN表达阳性率及表达强度评分显著高于肝硬化组织及正常肝组织(x~2=27.67,P＜0.01;F=12.82,P＜0.01);癌组织TN表达阳性率与癌旁组织比较差异无统计学意义(x~2=0.325,P＞0.05);TN表达与肝癌的组织学分级,病灶数量,有否包膜浸润、肿瘤转移及肿瘤微血管生成密切相关(P＜0.01)。63例HCC及其癌旁组织血管CD105阳性表达率为100%,癌旁组织表达强度高于癌组织(x~2=9.184,P＜0.01);HCC组织MVD-CD105为(58.37±21.45),癌旁组织为(81.62±19.86),HCC癌旁组织MVD-CD105分布显著高于癌组织(t=2.438,P＜0.05);HCC组织中的MVD-CD105阳性表达分布与病人的性别、年龄、HBsAg状态、甲胎蛋白水平、肿瘤大小、病灶个数、包膜浸润、肝癌细胞组织学分级、肝功能评级、是否合并肝硬化及2年存活时间无明显关系(P＞0.05),与肿瘤转移及TNM分期有关(P＜0.01);癌旁组织的MVD-CD105阳性表达分布与病人的性别、年龄、HBsAg状态、甲胎蛋白水平、肿瘤大小、病灶个数、包膜浸润、肝癌细胞组织学分级、肝功能评级、是否合并肝硬化及肿瘤有无转移无关(P＞0.05),而与肿瘤TNM分期及2年存活时间有关(P＜0.05)。
结论HCC癌组织TN的表达上调促进了肿瘤的新生血管形成,导致了术后肿瘤的早期复发和转移;肝癌组织TN检测可有效反映肝癌的恶性程度、浸润、转移及临床预后;CD105标染HCC组织MVD较CD34未体现出肿瘤微血管标染特异性方面的优越性;肝癌组织MVD-CD105高表达、分布意味着肿瘤的局部转移和进展;CD105分子不能作为抗肝癌肿瘤血管生成靶向治疗的靶标,但肝癌MVD-CD105的研究对于后续综合治疗有指导作用。
第三部分Wnt5a表达与HCC形成、侵袭和转移
目的探讨Wnt5a基因与蛋白在HCC的表达及对HCC的发生、转移的影响,为肝癌发病及侵袭转移的分子机制提供新的理论依据,为肝癌治疗开辟新途径。
方法应用实时荧光定量RT-PCR检测28例HCC癌组织及癌旁组织中Wnt5amRNA的表达,同时采用免疫组织化学染色方法检测Wnt5a蛋白的表达和MVD-CD105分布,统计分析它们与临床病理特征之间的关系。
结果28例HCC及其癌旁组织、10例正常肝组织均可见Wnt5a mRNA转录表达,HCC为(0.78±0.15),癌旁组织(0.81±0.17),正常组织为(0.41±0.11),HCC及其癌旁组织表达水平均高于正常肝组织(F=6.958,P＜0.001);HCC组织和癌旁组织转录表达水平未见差异(t=1.16,P=0.227);28例HCC及其癌旁组织、10例正常肝组织Wnt5a蛋白阳性表达率分别为32.1%、89.3%和30.0%,癌旁组织Wnt5a蛋白表达阳性率显著高于HCC和正常肝组织(x~2=21.808,P＜0.001);HCCWnt5a mRNA转录表达水平与性别、年龄、HBsAg状态、肿瘤大小、肿瘤病灶数目、包膜有无侵犯、肿瘤有无转移、肝功能Child评级、肝硬化及肿瘤TNM分期无关(P＞0.05),与血清AFP浓度、HCC病理分化分级及肝卵圆细胞增殖表达有关(P＜0.05);HCCWnt5a蛋白表达水平与性别、年龄、HBsAg状态、肿瘤大小、肿瘤病灶数目、包膜有无侵犯、肿瘤有无转移、肝功能Child评级、肝硬化及C-KIT蛋白表达无关(P＞0.05),但与血清AFP浓度水平、HCC病理分化分级和肿瘤TNM分期有关(P＜0.05);HCC组织中Wnt5a蛋白表达阳性组MVD-CD105计数为(46.51±15.38),阴性组为(38.24±8.32),差异未见显著性(t=1.16,P=0.227)。
结论HCC组织Wnt5a mRNA高转录表达与Wnt5a蛋白低翻译表达并不矛盾;Wnt5a mRNA高转录表达对细胞的生长、迁移、分化及转化具有双向调节功能,发挥抑制肿瘤浸润、转移作用,是机体修复机制的一种体现;HCC中Wnt5a蛋白表达减弱或缺失是肝癌发生的晚期事件,是肿瘤进展、浸润和转移的标志;HCC组织中Wnt5a蛋白表达与HCC组织MVD-CD105分布是与肿瘤TNM分期有关的两个相互独立的病理因素。
PartⅠStudy on the potentiality of hepatocellular carcinoma originating from hepatic oval cells by immunohistochemisty and its clinical significance
Objetive There are two theories on the origin of hepatocellular carcinoma and the two theories are still controversial.This experiment is to explore whether hepatocellular carcinoma(HCC)originates from hepatic oval cells and whether there is any relationship beween origin of HCC and clinicopathologic factors of HCC.
Methods Specimens were obtained from 63 cases of hepatocellular carcinoma and its adjacent non-tumorous tissues,15 cases of liver cirrhotic tissues and 10 normal liver tissues,so they were divided into four groups.Cytokeratin(CK)8/18,CK7,CK19, C-KIT and AFP were used as surface marker of oval cells.The expressions of surface marker of oval cells were semiquantitiatively assessed by immunohistochemisty.Cell morphologic features and distributions labled by CK8/18,CK7 CK19,C-KIT and AFP were observed with optical microscope.Clinical and pathological data including gender,serum HBsAg status,HBcAb status,serum AFP,tumor size,number of intrahepatic lesions and so on were summuried.
Results The positive rates labled by CK8/18 were 95.2%in HCC,96.8%in adjacent non-tumorous tissues,100%in cirrhotic tissues and 100%in normal tissues, respectively.There were no significant difference of the positive rates and intensity scores labled by CK8/18 among four groups(x~2=1.274,P＞0.05;F=1.465,P＞0.05). The positive rates labled by CK7 were 46.0%in HCC、58.7%in adjacent non-tumorous tissues of HCC,26.7%in cirrhotic tissues and negative in normal tissues,respectively.There were significant difference of the positive rates and intensity scores labled by CK7 among four groups(x~2=14.86,P＜0.01;F=14.73, P＜0.01).The positive rates and intensity scores labled by CK7 were significantly higher in HCC than those in cirrhotic and normal tissues(x~2=6.887,P＜0.01; F=21.26,P＜0.01),so were in adjacent non-tumorous tissues(x~2=13.133,P＜0.01; F=21.26,P＜0.01).The hepatic cells and tumor cells were no labled by CK19 in all specimens.There were significant difference of the positive rates and intensity scores labled by C-KIT among four groups(x~2=25.089,P＜0.01).The positive rates and intensity scores labled by C-KIT were significantly higher in HCC,adjacent non-tumorous tissues and cirrhotic tissues than those in normal tissues.The intensity scores labled by C-KIT were significantly higher in adjacent non-tumorous tissues than those in HCC,cirrhotic and normal tissues(F=36.19,P＜0.01).The positive rates and intensity scores labled by AFP were significantly higher in HCC than those in adjacent non-tumorous tissues,cirrhotic and normal tissues(x~2=5.567,P＜0.05;F= 15.76,P＜0.01).Hepatic oval cells labled by CK7,C-KIT and AFP were seen among normal or tumor cells,especially on the edge of tumor nodules.There is no correlation between CK7 positive expression in HCC and all involed clinicopathologic factors(P＞0.05).C-KIT positive expression was associated with the following clinicopathologic factors,such as serum AFP,tumor metastasis,cancer cells ES grading and TNM staging(P＜0.01),but not with the others(P＞0.05).AFP positive expression in HCC was associated with the serum AFP,tumor metastasis,cancer cells ES grading,TNM staging and postoperative survival periods of 2 years(P＜0.01),but not with the others(P＞0.05).
Conclusions The oval cells labled by CK7,C-KIT and AFP take part in the occurrence of HCC and may be one of the original cells for HCC.The oval cells labled by CK7,C-KIT and AFP represent various stem subpopulations and play different roles for repairment of chronic hepatitic injury and carcinogenesis.Expressions of stem cell phenotypic markers in HCC and paracancerous tissue represent a variety of clinical significance.Expressions of CK7 and C-KIT in adjacent non-tumorous tissues is related to the heterogeneity of hepatic oval cells,proliferation and tissue repairment.Expressions of C-KIT and AFP in HCC indicate malignant transformation of oval cells.
PartⅡChanges of Tenascin and MVD-CD105 in hepatocellular cancer tissues and their correlations with metastasis
Objective To study the expression of tenascin in hepatocellular carcinoma(HCC) and to investigate the roles of tenascin expression in microvessel angiogenesis,invasion, metastasis and prognosis of HCC.To evaluate the expression and distribution of CD105 in HCC and adjacent non-tumorous tissues,and then to discuss the specificity of CD 105 labling microvessel density(MVD) and its clinical significance.
Methods Formalin fixed,paraffin wax embedded specimens from 63 patients with HCC were stained with anti-tenascin and anti-CD105 monoclonal antibody,So were done by adjacent non-tumorous specimens from the same patients.The correlation was analysed between expression of tenascin and clinicopathological features as well as microvessel angiogenesis.The correlation was analysed between the expression and distribution of MVD-CD 105 in HCC and clinicopathological features,so was also analysed in adjacent non-tumorous tissues.
Results Tenascin immunoreactivity was seen in 65.1%specimens of HCC,69.8%of adjacent non-tumorous,20.0%of liver cirrhosis and riegative of normal liver specimens.Positivity and intensity of tenascin expressions in specimens of HCC and adjacent non-tumorous tissues were higher than those in specimens of cirrhotic and normal liver(x~2=27.67,P＜0.01;F=12.82,P＜0.01).Tenascin expression was associated with E-S histological grading,tumour numbers,capsule invasion,tumour metastasis and microvessel angiogenesis.CD105 immunoreactivity was seen in all specimens of HCC and adjacent non-tumorous tissues.Immunoreactivity intensity was higher in adjacent non-tumorous tissues than that in HCC(x~2=9.184,P＜0.01). The mean scores of MVD-CD105 were higher in adjacent non-tumorous tissues than that in HCC.(81.62±19.86 vs 58.37±21.45;t=2.438,P＜0.05).The levels of expression and distribution of MVD-CD105 in HCC were associated with tumor metastasis and TNM staging(P＜0.01),but not with gendar,age,HBsAg status,AFP level,tumor size,tumor number,capsule infiltration,E-S histological grading,liver function,cirrhosis and survival periods of 2 years(P＞0.05).The levels of expression and distribution of MVD-CD105 in adjacent non-tumorous tissues were associated with TNM staging and survival periods of 2 years(P＜0.01),but not with gendar,age, HBsAg status,AFP level,tumor size,tumor number,capsule infiltration,E-S histological grading,liver function,cirrhosis and tumor metastasis(P＞0.05).
Conclusions The up-regulation of tenascin expression may play an important role in invasion and metastasis of HCC and lead to poor prognosis.The superiority of MVD labled with CD 105 is not reflected compared with MVD labled with CD34,The latter is pan-endothelial cell maker that reacts not only with prolifering vessles but also with established vessles in the tumor.The higher level of expression and distribution of MVD-CD105 in HCC means that the local tumor metastasis and progress.CD105 can not be serve as an appropriate targeting for antiangenesis therapy in HCC,but the study on MVD-CD105 in HCC may guide the comprehensive treatment postoperatively.
PartⅢThe roles of Wnt5a in hepatocellular carcinoma development, invasion and metastasis
Objective To detect the expression of Wnt5a at the levels of transcription and translation and to evaluate the roles of Wnt5a expression in microvessel angiogenesis, invasion,metastasis and prognosis of HCC.
Methods The level of Wnt5a mRNA expression in HCC and adjacent non-tumorous tissues of 28 cases was detected by real-time fluorescence quantitative RT-PCR.The level of Wnt5a protein expression was detected with the same specimens by immunohistochemistry,so was done the level of expression and distribution of MVD-CD105.The correlation was analysed between the expression level of Wnt5a mRNA,Wnt5a protein and MVD-CD105 in HCC and clinicopathological features.
Results Wnt5a mRNA transcription was seen in all specimens harvested from cancerous and adjacent non-tumorous tissues of 28 cases with HCC and normal liver tissues of 10 cases.The mean expression level is 0.78±0.15 in HCC,0.81±0.17 in adjacent non-tumorous tissues and 0.41±0.11 in normal liver tissues.The expression levels were significantly higher in HCC and adjacent non-tumorous tissues than that in normal liver tissues(F=6.958,P＜0.001).There was no significant difference of Wnt5a mRNA transcription between cancerous and adjacent non-tumorous tissues (t=1.16,P=0.227).Wnt5a immunoreactivity was 32.1%in specimens of HCC, 89.3%in the adjacent non-tumor and 30.0%in normal liver.Wnt5a immunoreactivity in adjacent non-tumorous tissues were significantly higher than that in HCC and normal liver tissues(x~2=21.808,P＜0.001).There were no significant differences of the mean scores of MVD-CD105 between Wnt5a protein expression higher and lower specimens.(46.51±15.38 vs 38.24±8.32;t=1.132,P＞0.05).The level of Wnt5a mRNA transcription in HCC was associated with serum AFP level,E-S histological grading and proliferation of hepatic oval cells(P＜0.05),but not with gendar,age,HBsAg status,tumor size,tumor numbers,capsule infiltration,tumor metastasis,liver function,cirrhosis and TNM staging(P＞0.05).The level of Wnt5a protein in HCC was associated with serum AFP level,E-S histological grading and TNM staging(P＜0.05),but not with gendar,age,HBsAg status,tumor size,tumor numbers,capsule infiltration,tumor metastasis,liver function,cirrhosis and immunoreactivity of C-KIT(P＞0.05).
Conclusions It is not incompatible that Wnt5a mRNA is increased transcription expression and Wnt5a protein is reduced expression in HCC.Increased transcription expression of Wnt5a mRNA can regulate cell growth,migration,differentiation and transformation bidirectionally and play a tumor suppressor role,which embodys one kinds of repairing mechanisms of preserving cells normal morphology and function. The reduced or lost expression of Wnt5a protein in HCC should be a late event during development of HCC,which implicates tumor invasion and metastasis.The expression of Wnt5a protein and the distribution of MVD-CD105 in HCC are two independent cliniclpathological factors associated with TNM staging.
目的有关原发性肝癌的细胞起源存在两种学说,即成熟肝细胞去分化学说和肝干细胞分化受阻学说,这两种学说目前尚有争议。本实验通过测定肝硬化、肝癌和癌旁组织肝干细胞标志的表达,观察肝干细胞标志的表达阳性细胞的细胞学形态,结合肝癌患者的临床病理特征,探讨肝脏卵圆细胞在肝癌形成中所扮演的角色及其与临床病理因素的相互关系。
方法应用免疫组化的方法,测定63例肝细胞肝癌及其癌旁组织、15例肝炎后肝硬化组织标本及10例正常肝组织标本C-KIT+、AFP、CK8/18、CK7和CK19的表达,用光学显微镜观察上述组织标本干细胞标记标染的细胞形态、分布,回顾性总结63例HCC患者的临床病理资料,分析HCC标本C-KIT+、AFP、CK8/18、CK7和CK19的表达与性别、血清HBsAg状态、HBcAb状态、血清AFP、肿瘤大小、肝内病灶数目、是否有包膜侵犯、肝癌细胞病理分化分级、肝功能评级、肝硬化程度、TNM分期及术后2年生存期等13个临床病理参数的关系。
结果63例HCC及其癌旁组织、15例肝炎后肝硬化组织标本及10例正常肝组织标本免疫组织化学染色显示,CK8/18标染阳性率分别是95.2%、96.8%,100%和100%,四组间CK8/18阳性表达率和表达强度评分差异均无显著性(x~2=1.274,P＞0.05;F=1.465,P＞0.05);CK7标染阳性率分别是46.0%、58.7%,26.7%和0,四组间CK7阳性表达率差异有显著性(x~2=14.86,P＜0.01),肝癌组织、癌旁组织CK7表达率均分别高于肝硬化、正常肝组织(x~2=6.887,P＜0.01;x~2=13.133,P＜0.01);CK7标染表达强度评分比较,肝癌组织、癌旁组织CK7阳性表达强度评分高于肝硬化组织及正常组织(F=21.26,P＜0.01);四种组织实质肝细胞和肿瘤细胞均未见CK19阳性表达;四组间C-KIT阳性表达率差异有显著性(x~2=25.089,P＜0.01),HCC、癌旁及肝硬化组织C-KIT阳性表达率显著高于正常组织,癌旁组织C-KIT阳性病例表达强度评分显著高于HCC、肝硬化及正常组织(F=36.19,P＜0.01);HCC组织AFP阳性率及表达强度评分均显著高于癌旁组织、肝硬化及正常组织(x~2=5.567,P＜0.05;F=15.76,P＜0.01)。HCC实质组织中可见符合肝卵圆细胞形态特点CK7、C-KIT及AFP阳性细胞,呈点状、片状或弥漫性分布,以癌结节边缘多见。HCC癌组织中,CK7阳性表达与肝癌病人的性别、血清HBsAg状态、HBcAb状态、血清AFP、肿瘤大小、肝内病灶数目、是否有包膜侵犯、肝癌细胞病理分化分级、肝功能评级、肝硬化程度、TNM分期及术后2年生存期等病理参数均无关(P＞0.05),;C-KIT阳性表达与血清AFP、肿瘤转移、肝癌细胞病理分级及肿瘤TNM分期有关(P＜0.01),和其余病理因素无关(P＞0.05);组织中AFP阳性表达与血清AFP、肿瘤转移、肝癌细胞病理分化分级、肿瘤TNM分期及术后2年生存期等参数有关(P＜0.01),和其余病理因素无关(P＞0.05)。
结论HCC及癌旁组织中存在表达肝干细胞标记表型并具有卵圆细胞形态的细胞,提示HCC的发生与肝脏干细胞/卵圆细胞密切相关,表达CK7、C-KIT和AFP的肝卵圆细胞参与了HCC形成;肝卵圆细胞表型标记CK7、C-KIT和AFP在HCC组织和癌旁组织有不同的表达规律,它们可能代表不同的干细胞群,在肝炎慢性损伤修复和癌变过程中具有不同作用;HCC及其癌旁组织干细胞表型标记表达具有多种临床意义;癌旁CK7和C-KIT表达与肝卵圆细胞异质性、增殖和组织修复有关,癌组织C-KIT和AFP表达预示着卵圆细胞由癌旁组织中的增殖修复功能向癌组织的恶性转化。
第二部分肝癌局部微环境改变与肝癌浸润、转移
目的研究细胞外基质tenascin(TN)在HCC(hepatocellular carcinoma,HCC)中的表达及HCC癌、癌旁组织微血管密度(MVD-CD105)分布,探讨TN对HCC肿瘤微血管生成、浸润、转移及临床预后的影响,MVD-CD105在HCC癌、癌旁组织分布规律、临床意义及CD105标染HCC组织微血管的特异性。
方法采用免疫组织化学方法检测63例HCC癌组织、癌旁组织及15例肝炎后肝硬化组织、10例正常肝组织中TN表达,同时检测63例HCC癌组织、癌旁组织微血管密度(MVD-CD105),分析TN表达与HCC病理学特点及肿瘤微血管生成的关系,比较HCC癌组织和癌旁组织MVD-CD105分布特点及其与临床参数的关系。
结果TN阳性率HCC癌组织65.1%,癌旁组织69.8%,肝硬化组织20.0%,正常肝组织无阳性表达;HCC癌组织及癌旁组织TN表达阳性率及表达强度评分显著高于肝硬化组织及正常肝组织(x~2=27.67,P＜0.01;F=12.82,P＜0.01);癌组织TN表达阳性率与癌旁组织比较差异无统计学意义(x~2=0.325,P＞0.05);TN表达与肝癌的组织学分级,病灶数量,有否包膜浸润、肿瘤转移及肿瘤微血管生成密切相关(P＜0.01)。63例HCC及其癌旁组织血管CD105阳性表达率为100%,癌旁组织表达强度高于癌组织(x~2=9.184,P＜0.01);HCC组织MVD-CD105为(58.37±21.45),癌旁组织为(81.62±19.86),HCC癌旁组织MVD-CD105分布显著高于癌组织(t=2.438,P＜0.05);HCC组织中的MVD-CD105阳性表达分布与病人的性别、年龄、HBsAg状态、甲胎蛋白水平、肿瘤大小、病灶个数、包膜浸润、肝癌细胞组织学分级、肝功能评级、是否合并肝硬化及2年存活时间无明显关系(P＞0.05),与肿瘤转移及TNM分期有关(P＜0.01);癌旁组织的MVD-CD105阳性表达分布与病人的性别、年龄、HBsAg状态、甲胎蛋白水平、肿瘤大小、病灶个数、包膜浸润、肝癌细胞组织学分级、肝功能评级、是否合并肝硬化及肿瘤有无转移无关(P＞0.05),而与肿瘤TNM分期及2年存活时间有关(P＜0.05)。
结论HCC癌组织TN的表达上调促进了肿瘤的新生血管形成,导致了术后肿瘤的早期复发和转移;肝癌组织TN检测可有效反映肝癌的恶性程度、浸润、转移及临床预后;CD105标染HCC组织MVD较CD34未体现出肿瘤微血管标染特异性方面的优越性;肝癌组织MVD-CD105高表达、分布意味着肿瘤的局部转移和进展;CD105分子不能作为抗肝癌肿瘤血管生成靶向治疗的靶标,但肝癌MVD-CD105的研究对于后续综合治疗有指导作用。
第三部分Wnt5a表达与HCC形成、侵袭和转移
目的探讨Wnt5a基因与蛋白在HCC的表达及对HCC的发生、转移的影响,为肝癌发病及侵袭转移的分子机制提供新的理论依据,为肝癌治疗开辟新途径。
方法应用实时荧光定量RT-PCR检测28例HCC癌组织及癌旁组织中Wnt5amRNA的表达,同时采用免疫组织化学染色方法检测Wnt5a蛋白的表达和MVD-CD105分布,统计分析它们与临床病理特征之间的关系。
结果28例HCC及其癌旁组织、10例正常肝组织均可见Wnt5a mRNA转录表达,HCC为(0.78±0.15),癌旁组织(0.81±0.17),正常组织为(0.41±0.11),HCC及其癌旁组织表达水平均高于正常肝组织(F=6.958,P＜0.001);HCC组织和癌旁组织转录表达水平未见差异(t=1.16,P=0.227);28例HCC及其癌旁组织、10例正常肝组织Wnt5a蛋白阳性表达率分别为32.1%、89.3%和30.0%,癌旁组织Wnt5a蛋白表达阳性率显著高于HCC和正常肝组织(x~2=21.808,P＜0.001);HCCWnt5a mRNA转录表达水平与性别、年龄、HBsAg状态、肿瘤大小、肿瘤病灶数目、包膜有无侵犯、肿瘤有无转移、肝功能Child评级、肝硬化及肿瘤TNM分期无关(P＞0.05),与血清AFP浓度、HCC病理分化分级及肝卵圆细胞增殖表达有关(P＜0.05);HCCWnt5a蛋白表达水平与性别、年龄、HBsAg状态、肿瘤大小、肿瘤病灶数目、包膜有无侵犯、肿瘤有无转移、肝功能Child评级、肝硬化及C-KIT蛋白表达无关(P＞0.05),但与血清AFP浓度水平、HCC病理分化分级和肿瘤TNM分期有关(P＜0.05);HCC组织中Wnt5a蛋白表达阳性组MVD-CD105计数为(46.51±15.38),阴性组为(38.24±8.32),差异未见显著性(t=1.16,P=0.227)。
结论HCC组织Wnt5a mRNA高转录表达与Wnt5a蛋白低翻译表达并不矛盾;Wnt5a mRNA高转录表达对细胞的生长、迁移、分化及转化具有双向调节功能,发挥抑制肿瘤浸润、转移作用,是机体修复机制的一种体现;HCC中Wnt5a蛋白表达减弱或缺失是肝癌发生的晚期事件,是肿瘤进展、浸润和转移的标志;HCC组织中Wnt5a蛋白表达与HCC组织MVD-CD105分布是与肿瘤TNM分期有关的两个相互独立的病理因素。
PartⅠStudy on the potentiality of hepatocellular carcinoma originating from hepatic oval cells by immunohistochemisty and its clinical significance
Objetive There are two theories on the origin of hepatocellular carcinoma and the two theories are still controversial.This experiment is to explore whether hepatocellular carcinoma(HCC)originates from hepatic oval cells and whether there is any relationship beween origin of HCC and clinicopathologic factors of HCC.
Methods Specimens were obtained from 63 cases of hepatocellular carcinoma and its adjacent non-tumorous tissues,15 cases of liver cirrhotic tissues and 10 normal liver tissues,so they were divided into four groups.Cytokeratin(CK)8/18,CK7,CK19, C-KIT and AFP were used as surface marker of oval cells.The expressions of surface marker of oval cells were semiquantitiatively assessed by immunohistochemisty.Cell morphologic features and distributions labled by CK8/18,CK7 CK19,C-KIT and AFP were observed with optical microscope.Clinical and pathological data including gender,serum HBsAg status,HBcAb status,serum AFP,tumor size,number of intrahepatic lesions and so on were summuried.
Results The positive rates labled by CK8/18 were 95.2%in HCC,96.8%in adjacent non-tumorous tissues,100%in cirrhotic tissues and 100%in normal tissues, respectively.There were no significant difference of the positive rates and intensity scores labled by CK8/18 among four groups(x~2=1.274,P＞0.05;F=1.465,P＞0.05). The positive rates labled by CK7 were 46.0%in HCC、58.7%in adjacent non-tumorous tissues of HCC,26.7%in cirrhotic tissues and negative in normal tissues,respectively.There were significant difference of the positive rates and intensity scores labled by CK7 among four groups(x~2=14.86,P＜0.01;F=14.73, P＜0.01).The positive rates and intensity scores labled by CK7 were significantly higher in HCC than those in cirrhotic and normal tissues(x~2=6.887,P＜0.01; F=21.26,P＜0.01),so were in adjacent non-tumorous tissues(x~2=13.133,P＜0.01; F=21.26,P＜0.01).The hepatic cells and tumor cells were no labled by CK19 in all specimens.There were significant difference of the positive rates and intensity scores labled by C-KIT among four groups(x~2=25.089,P＜0.01).The positive rates and intensity scores labled by C-KIT were significantly higher in HCC,adjacent non-tumorous tissues and cirrhotic tissues than those in normal tissues.The intensity scores labled by C-KIT were significantly higher in adjacent non-tumorous tissues than those in HCC,cirrhotic and normal tissues(F=36.19,P＜0.01).The positive rates and intensity scores labled by AFP were significantly higher in HCC than those in adjacent non-tumorous tissues,cirrhotic and normal tissues(x~2=5.567,P＜0.05;F= 15.76,P＜0.01).Hepatic oval cells labled by CK7,C-KIT and AFP were seen among normal or tumor cells,especially on the edge of tumor nodules.There is no correlation between CK7 positive expression in HCC and all involed clinicopathologic factors(P＞0.05).C-KIT positive expression was associated with the following clinicopathologic factors,such as serum AFP,tumor metastasis,cancer cells ES grading and TNM staging(P＜0.01),but not with the others(P＞0.05).AFP positive expression in HCC was associated with the serum AFP,tumor metastasis,cancer cells ES grading,TNM staging and postoperative survival periods of 2 years(P＜0.01),but not with the others(P＞0.05).
Conclusions The oval cells labled by CK7,C-KIT and AFP take part in the occurrence of HCC and may be one of the original cells for HCC.The oval cells labled by CK7,C-KIT and AFP represent various stem subpopulations and play different roles for repairment of chronic hepatitic injury and carcinogenesis.Expressions of stem cell phenotypic markers in HCC and paracancerous tissue represent a variety of clinical significance.Expressions of CK7 and C-KIT in adjacent non-tumorous tissues is related to the heterogeneity of hepatic oval cells,proliferation and tissue repairment.Expressions of C-KIT and AFP in HCC indicate malignant transformation of oval cells.
PartⅡChanges of Tenascin and MVD-CD105 in hepatocellular cancer tissues and their correlations with metastasis
Objective To study the expression of tenascin in hepatocellular carcinoma(HCC) and to investigate the roles of tenascin expression in microvessel angiogenesis,invasion, metastasis and prognosis of HCC.To evaluate the expression and distribution of CD105 in HCC and adjacent non-tumorous tissues,and then to discuss the specificity of CD 105 labling microvessel density(MVD) and its clinical significance.
Methods Formalin fixed,paraffin wax embedded specimens from 63 patients with HCC were stained with anti-tenascin and anti-CD105 monoclonal antibody,So were done by adjacent non-tumorous specimens from the same patients.The correlation was analysed between expression of tenascin and clinicopathological features as well as microvessel angiogenesis.The correlation was analysed between the expression and distribution of MVD-CD 105 in HCC and clinicopathological features,so was also analysed in adjacent non-tumorous tissues.
Results Tenascin immunoreactivity was seen in 65.1%specimens of HCC,69.8%of adjacent non-tumorous,20.0%of liver cirrhosis and riegative of normal liver specimens.Positivity and intensity of tenascin expressions in specimens of HCC and adjacent non-tumorous tissues were higher than those in specimens of cirrhotic and normal liver(x~2=27.67,P＜0.01;F=12.82,P＜0.01).Tenascin expression was associated with E-S histological grading,tumour numbers,capsule invasion,tumour metastasis and microvessel angiogenesis.CD105 immunoreactivity was seen in all specimens of HCC and adjacent non-tumorous tissues.Immunoreactivity intensity was higher in adjacent non-tumorous tissues than that in HCC(x~2=9.184,P＜0.01). The mean scores of MVD-CD105 were higher in adjacent non-tumorous tissues than that in HCC.(81.62±19.86 vs 58.37±21.45;t=2.438,P＜0.05).The levels of expression and distribution of MVD-CD105 in HCC were associated with tumor metastasis and TNM staging(P＜0.01),but not with gendar,age,HBsAg status,AFP level,tumor size,tumor number,capsule infiltration,E-S histological grading,liver function,cirrhosis and survival periods of 2 years(P＞0.05).The levels of expression and distribution of MVD-CD105 in adjacent non-tumorous tissues were associated with TNM staging and survival periods of 2 years(P＜0.01),but not with gendar,age, HBsAg status,AFP level,tumor size,tumor number,capsule infiltration,E-S histological grading,liver function,cirrhosis and tumor metastasis(P＞0.05).
Conclusions The up-regulation of tenascin expression may play an important role in invasion and metastasis of HCC and lead to poor prognosis.The superiority of MVD labled with CD 105 is not reflected compared with MVD labled with CD34,The latter is pan-endothelial cell maker that reacts not only with prolifering vessles but also with established vessles in the tumor.The higher level of expression and distribution of MVD-CD105 in HCC means that the local tumor metastasis and progress.CD105 can not be serve as an appropriate targeting for antiangenesis therapy in HCC,but the study on MVD-CD105 in HCC may guide the comprehensive treatment postoperatively.
PartⅢThe roles of Wnt5a in hepatocellular carcinoma development, invasion and metastasis
Objective To detect the expression of Wnt5a at the levels of transcription and translation and to evaluate the roles of Wnt5a expression in microvessel angiogenesis, invasion,metastasis and prognosis of HCC.
Methods The level of Wnt5a mRNA expression in HCC and adjacent non-tumorous tissues of 28 cases was detected by real-time fluorescence quantitative RT-PCR.The level of Wnt5a protein expression was detected with the same specimens by immunohistochemistry,so was done the level of expression and distribution of MVD-CD105.The correlation was analysed between the expression level of Wnt5a mRNA,Wnt5a protein and MVD-CD105 in HCC and clinicopathological features.
Results Wnt5a mRNA transcription was seen in all specimens harvested from cancerous and adjacent non-tumorous tissues of 28 cases with HCC and normal liver tissues of 10 cases.The mean expression level is 0.78±0.15 in HCC,0.81±0.17 in adjacent non-tumorous tissues and 0.41±0.11 in normal liver tissues.The expression levels were significantly higher in HCC and adjacent non-tumorous tissues than that in normal liver tissues(F=6.958,P＜0.001).There was no significant difference of Wnt5a mRNA transcription between cancerous and adjacent non-tumorous tissues (t=1.16,P=0.227).Wnt5a immunoreactivity was 32.1%in specimens of HCC, 89.3%in the adjacent non-tumor and 30.0%in normal liver.Wnt5a immunoreactivity in adjacent non-tumorous tissues were significantly higher than that in HCC and normal liver tissues(x~2=21.808,P＜0.001).There were no significant differences of the mean scores of MVD-CD105 between Wnt5a protein expression higher and lower specimens.(46.51±15.38 vs 38.24±8.32;t=1.132,P＞0.05).The level of Wnt5a mRNA transcription in HCC was associated with serum AFP level,E-S histological grading and proliferation of hepatic oval cells(P＜0.05),but not with gendar,age,HBsAg status,tumor size,tumor numbers,capsule infiltration,tumor metastasis,liver function,cirrhosis and TNM staging(P＞0.05).The level of Wnt5a protein in HCC was associated with serum AFP level,E-S histological grading and TNM staging(P＜0.05),but not with gendar,age,HBsAg status,tumor size,tumor numbers,capsule infiltration,tumor metastasis,liver function,cirrhosis and immunoreactivity of C-KIT(P＞0.05).
Conclusions It is not incompatible that Wnt5a mRNA is increased transcription expression and Wnt5a protein is reduced expression in HCC.Increased transcription expression of Wnt5a mRNA can regulate cell growth,migration,differentiation and transformation bidirectionally and play a tumor suppressor role,which embodys one kinds of repairing mechanisms of preserving cells normal morphology and function. The reduced or lost expression of Wnt5a protein in HCC should be a late event during development of HCC,which implicates tumor invasion and metastasis.The expression of Wnt5a protein and the distribution of MVD-CD105 in HCC are two independent cliniclpathological factors associated with TNM staging.
引文
1.Strick-Marchand H,Morosan S,Charneau P,et al.Bipotential mouse embryonic liver stem cell lines contribute to liver regeneration and differentiate as bile ducts and hepatocytes.Proc Natl Acad Sci USA,2004,101(22):8360-8365.
2.金世龙,刘宝华,刘宏鸣,等.大鼠肝卵圆细胞的分离培养及干细胞标志分析.中国普外基础与临床杂志,2007,14(1):68-71.
3.Zheng YW,Taniguchi H.Diversity of hepatic stem cells in the fetal and adult liver.Semin Liver Dis,2003,23(4):337-348.
4.Paku S,Schnur J,Nagy P,et al.Origin and structural evolution of the early proliferating oval cells in rat liver.Am J Pathol,2001,158(4):1313-1321
5.Lemmer ER,Shepard EG,Blakolmer K,Kirsch RE,Robson SC:Isolation from human fetal liver of cells co-expressing CD34 haematopoietic stem cell and CAM 5.2 pancytokera-tin markers.J Hepatol,1998,29:450-454.
6.Libbrecht L,Desmet V,Van DB,et al.The immunohistochemical phenotype of dysplastic foci in human liver:correlation with putative progenitor cells.J Hepatol,2000,33:76-84.
7.Van EP,Desmet VJ.Cytokeratin and the liver.Liver,1993,13:113-122.
8.Sobin L H,Wittekind C,eds.TNM classification of malignant rumors[M].6th ed.New York:Wiley—Liss,2002:81-83.
9.Edmondson HA,Steiner PE.Primary carcinoma of the liver:a study of 100 cases among 48,900 necropsies.Cancer,1954,7:462-503.
10.Hsia CC,Evarts RP,Nakatsukasa H,et al.Occurrence of oval type cells in hepatitis B virus associated human hepatocacinogenesis.Hepatology,1992,16(6):1327-1333.
11.Mattern J,Koom(a|¨)gi R,Volm M.Association of vascular endothelial growth factor expression with intratumoral microvessel density and tumour cell proliferation in human epidermoid lung carcinoma.Br J Cancer,1996,73(7):931-934.
12.Klein WM,Molmenti EP,Colombani PM,et al.Primary liver carcinoma arising in people younger than 30 years.Am J Clin Pathol,2005,124(4):512-518.
13.Kara B,Doran F,Kara IO,et al.Expression of c-kit protooncogen in hepatitis B virus-induced chronic hepatitis,cirrhosis and hepatocellular carcinoma:has it a diagnostic role? Int J Clin Pract, 2008, 62(8): 1206-1211.
14. Tsuji M, Kashihara T, Terada N, et al. An immunohistochemical study of hepatic atypical adenomatous hyperplasia, hepatocellular carcinoma, and cholangiocarcinoma with alpha-fetoprotein, carcinoembryonic antigen, CA19-9, epithelial membrane antigen, and cytokeratins 18 and 19. Pathol Int, 1999, 49 (4): 310-317.
15. Desmet VJ. Organizational principles; in Arias IM, Boyer JL, Chisari FV (eds): The Liver, Biology and Pathology. Philadelphia, Lippincott Williams & Wilkins, 2001, pp3-15.
16. Libbrecht L, Roskams T. Hepatic progenitor cells in human liver diseases. Semin Cell Dev Biol, 2002,13: 389-396.
17. Roskams TA, Theise ND, Balabaud C, et al. Nomenclature of the finer branches of the biliary tree: canals, ductules, and ductular reactions in human livers. Hepatology, 2004, 39(6): 1739-1745.
18. De Vos R, Desmet V. Ultrastructural characteristics of novel epithelial cell types indentified in human pathologic liver specimens with chronic ductular reaction. Am J Pathol, 1992,140, (6): 1441-1450.
19. Thorgeirsson SS. Hepatic stem cells. Am J Pathol, 1993, 142: 1331-1333.
20. Strick-Marchand H, Morosan S, Charneau P, et al. Bipotential mouse embryonic liver stem cell lines contribute to liver regeneration and differentiate as bile ducts and hepatocytes . Proc Natl Acad Sci USA, 2004, 101(22) : 8360-8365.
21. Thorgeirsson SS. Hepatic stem cells in liver regeneration. FASEB J, 1996, 10: 1249-1256.
22. Runge D, Runge DM, Jager D, et al. Serum-free, long-term cultures of human hepatocytes: maintenance of cell morphology, transcription factors, and liver specific functions. Biochem Biophys Res Commun, 2000, 269(1): 46-53.
23. Braun KM, Thompson AW, Sandgren EP. Hepatic microenvironment affects oval cell localization in albumin urokinase type plasminogen activator transgenic mice. Am J Pathol, 2003, 162(1):195-202.
24. Lowes KN, Brennan BA, Yeoh GC, et al. Oval cell numbers in human chronic liver diseases are directly related to disease severity. Am J Pathol, 1999, 154: 537-541.
25. Libbrecht L, Desmet V, Van Damme B, et al. Deep intralobular extension of human hepatic'progenitor cells'correlates with parenchymal inflammation in chronic viral hepatitis: can 'progenitor cells' migrate? J Pathol, 2000, 192: 373-378.
26. Eleazar JA, Memeo L, Jhang JS, et al. Progenitor cell expansion: an important source of hepatocyte regeneration in chronic hepatitis. J Hepatol, 2004, 41: 983-991.
27. Moore KA, Lemischka IR. Stem cells and their niches. Science, 2006, 311(5769): 1880-1885.
28. Sell S. Stem cell origin of cancer and differentiation therapy. Crit Rev Oncol Hematol, 2004, 51:1-28.
29. Dontu M, Al-Hajj, Abdallah WM, et al. Stem cell in normal breast development and breast cancer. Cell Prolif, 2003, 36(Suppll): 59-72.
30. Singh SK, Hawkins C, Clarke ID, et al. Identification of human brain tumour initiating cells. Nature, 2004,432: 396-401.
31. Burger PE, Xiong X, Coetzee S, et al. Sca-1 expression identifies stem cell in the proximal region of prostatic ducts with high capacity to reconstitute prostatic tissue. Proc Natl Acad USA, 2005,102: 7180-7185.
32. Sell S. Cellular origin of cancer: dedifferentiation or stem cell maturation arrest? Environ Health Perspect, 1993,101 Suppl 5: 15-26.
33. Sell S, Pierce G B. Maturation arrest of stem cell differentiation is a common pathway for the cellular origin of teratocarcinomas and epithelial cancers. Lab Invest, 1994, 70(1): 6-22.
34. Sell S. Heterogeneity and plasticity of hepatocyte lineage cells. Hepatology, 2001, 33(3) : 738-750.
35. Tanaka S, Yamamoto T, Tanaka H, et al. Potentiality of combined hepatocellular and intrahepatic cholangiocellular carcinoma originating from a hepatic precursor cell: Immunohistochemical evidence. Hepatol Res, 2005, 32(1) .' 52-57.
36. Ruck P, Xiao JC, Kaiserling E. Small epithelial cells and the histogenesis of hepatoblastoma. Electron microscopic, immunoelectron microscopic, and immunohistochemical findings.Am J Pathol,1996,148(1):321-329.
37.Robrechts C,De-Vos R,Van-den-Heuvel M,et al.Primary liver tumor of intermediate hepatocyte-bile duct cell,phenotype:a progenitor cell tumour? Liver,1998,18:288-293.
38.Sell S,Xu KL,Huff WE,et al.Aflatoxin exposure produces serum alphafetoprotein elevations and marked oval cell proliferation in young male Pekin ducklings.Pathology,1998,30(1):34-39.
39.Tsao MS,Grisham JW.Hepatocarcinomas,cholangiocarcinomas,and hepatoblastomas produced by chemically transformed cultured rat liver epithelial cells-a light and electron microscopic analysis.Am J Pathol,1985,127:168-181.
40.陈琼荣,向锦,廖冰,等.卵圆细胞在实验性肝癌发生过程中的演变特征.癌症,2007,26(7):719-723.
41.Yoon DS,Jeong J,Park YN,et al.Expression of biliary antigen and its clinical significance in hepatocellular carcinoma.Yonsei Med J,1999,40(5):472-477.
42.Van Eyken P,Sciot R,Paterson A,et al.Cytokeratin expression in hepatocellular carcinoma:an immunohistochemical study.Hum Pathol,1988,19(5):562-568.
43.Uenishi T,Kubo S,Yamamoto T,et al.Cytokeratin 19 expression in hepatocellular carcinoma predicts early postoperative recurrence.Cancer Sci,2003,94(10):851-857.
44.Fausto N.Liver regeneration and repair:hepatocytes,Progenitor cells,and stem cells.Hepatology,2004,39:1477-1487.
45.Seeger C,Mason WS.Hepatitis B virus biology.Microbiol Mol Biol Rev,2000,64:51-68.
46.Blakolmer K,Jaskiewicz K,Dunsford HA,et al.Hematopoietic stem cell markers are expressed by ductal plate and bile duct cells in developing human liver.Hepatology,1995,21(6):1510-1516.
47.Kiiasov AP,Gumerova AA,Bilalov MM,et al.Cytokeratin expression in the pre-and postnatal ontogeny of the rat liver.Ontogenez,1997,28(5):389-93.
48.Fu Y,Zhang W,Liu Y F,et al.Abnormal expression of cytokeratins in hepatocellular carcinoma cell lines and its mechanism.J Fourth Mil Med Univ,2001,22(7):598-603.
49.Van Eyken P,Sciot R,Desmet VJ.A cytokeratin immunohistochemical study of cholestatic liver disease:evidence that hepatocytes can express bile duct type cytokeratins.Histopathol,1989,15(2):125-135.
50.Balaton AJ,Gotheil N-SC,Callard P,et al.Distribution between hepatocellular carcinoma,cholangiocarcionma,and metastatic carcinoma based on immunohistochemical staining for carcinoembryonic antigen and for CK19 on paraffin sections.J Pathol,1988,156:305-310.
51.Uenishi T,Kubo S,Hirohashi K,et al.Expression of bile duct-type cytokeratin in hepatocellular carcinoma in patients with hepatitis C virus and prior hepatitis B virus infection.Cancer Lett,2002,178(1):107-112.
52.肖家诚,王天翔,Ruck P等.肝硬化组织中卵圆细胞的光镜、电镜与免疫电镜观察.临床与实验病理学杂志,2002,18(1):57-60.
53.Goodman ZD,Ishak KG,Langloss JM,et al.Combined hepatocellular-cholangiocarcinoma.A histologic and immunohistochemical study.Cancer,1985,55:124-135.
54.Tickoo SK,Zee SY,Obiekwe S,et al.Combined hepatocellular-cholangiocarcinoma:a histopathologic,immunohistochemical,and in situ hybridization study.Am J Surg Pathol,2002,26:989-997.
55.Kim H,Park C,Han KH,et al.Primary liver carcinoma of intermediate (hepatocyte-cholangiocyte) phenotype.J Hepatol,2004,40:298-304.
56.Liaw YF,Tai DI,Chu CM,et al.Early detection of hepatocellular carcinoma in patients with chronic type B hepatitis A prospective study.Gastroenterology,1986,90:263-267.
57.Crosby HA,Kelly DA,Strain AJ.Human hepatic stem-like cells isolated using c-kit or CD34 can differentiate into biliary epithelium.Gastroenterology,2001,120:534-544.
58.Matsusaka S,Tsujimura T,Toyosaka A,et al.Role of c-kit receptor tyrosine kinase in development of oval cells in the rat 2-acetylaminoffuorene/partial hepatectomy model.Hepatology,1999,29:670-676.
59.Blakolmer K,Jaskiewicz K,Dunsford HA,et al.Hematopoietic stem cell markers are expressed by ductal plate and bile duct cells in developing human liver. Hepatology, 1995,21:1510-1516.
60. Yamamoto T, Uenishi T, Ogawa M, et al. Immunohistologic Attempt to Find Carcinogenesis from Hepatic Progenitor Cell in Hepatocellular Carcinoma. Digestive Surgery, 2005, 22(5):364-370.
1.吴孟超.原发性肝癌的诊断和治疗进展.中华外科杂志,1998,36(9):515-518.
2.王悦华,冯玉泉,刘永雄,等.原发性肝癌的外科治疗.中华普通外科杂志,1997,12(6):325-328.
3.Poon RT,Fan ST,Ng IO,et al.Different risk factors and prognosis for late intrahepatic recurrence after resection of hepatocarcinoma.Cancer,2000,89(3):5O0-507.
4.Folkman J.What is the evidence that tumors are angiogenesis dependent?.J Natl Cancer Inst,1990,82(1):4-6.
5.齐庆安,别平.原发性肝细胞肝癌血小板源性生长因子的表达和肿瘤血管形成关系的相关研究.消化外科,2005,4(2):119-123.
6.穆四清,张峰.原发性肝癌复发转移的分子机理.中国普外基础与临床杂志,2007,14(1):49-51.
7.Chiquet-Ehrismann R.Tenascins,a growing family of extracellular matrix proteins.Experientia,1995,51(9-10):853-862.
8.Midwood KS,Schwarzbauer JE.Tenascin-C modulates matrix contraction via focal adhesion kinase-and Rho-mediated signaling pathways.Mol Biol Cell,2002,13(10):3601-3613.
9.Chiquet-Ehrismann R.Anti-adhesive molecules of the extracellular matrix.Curr Opin Cell Biol,1991,3(5):800-804.
10.Jones PL,Cowan KN,Rabinovitch M.Tenascin-C,proliferation and subendothelial fibronectin in progressive pulmonary vascular disease.Am J Pathol,1997,150(4):1349-1360.
11.Erickson HP,Bourdon MA.Tenascin:an extracellular matrix protein prominent in specialized embryonic tissues and tumors.Annu Rev Cell Biol,1989,4(5):71-92.
12.Koukoulis GK,Gould VE,Bhattacharyya A,et al.Tenascin in normal,reactive,hyperplastic,and neoplastic tissues:biologic and pathologic implications.Hum Pathol,1991,22(7):636-643.
13.Boxer GM,Tsiompanou E,Levine T,et al.Immunohistochemical expression of vascular endothelial growth factor and microvessel counting as prognostic indicators in node-negative colorectal cancer. Tumour Biol, 2005, 26(1):1-8.
14. Vermeulen PB, Gasparinig G, Fox SB, et al. Quantification of angiogenesis in solid human tumours: an international consensus on the methodology and criteria of evaluation. Eur J Cancer, 1996, 32A(14): 2474-2484.
15. Wang JM, Kumar S, Pye D, et al. Breast carcinoma: comparative study of tumor vasculature using endothelial cell markers. J Nat Cancer Inst, 1994, 86:386-388.
16. Miller DW, Graulich W, Karges B, et al. Elwvated expression of endogl in, a component of the TGF-O-receptor complex, correlates with proliferation of tumor endothelial cells. Int J Cancer, 1999, 81:568-572.
17. Brewar CA, Setterdahl JJ, Li MJ, et al. Endoglin expression as a mesure of microvessel density in cervical cancer. Obstet Gynecol, 2000, 96:224-228.
18. Tanaka F, Otake Y, Yanagihara K, et al. Evaluation in angiogenesis in non-small cell lung cancer: comparison between anti-CD34 antibody and anti-CD 105 antibody. Clin. Cancer Res, 2001,7:3410-3415.
19. Mattern J, Koomagi R, Volm M. Association of vascular endothelial growth factor expression with intratumoral microvessel density and tumour cell proliferation in human epidermoid lung carcinoma. Br J Cancer, 1996, 73(7): 931-934.
20. Weidner N. Current pathologic methods for measuring intratumoral microvessel density within breast carcinoma and other solid tumors. Breast Cancer Res Treat, 1995,36(2):169-180.
21.Donjacour AA, Cunha GR. Stromal regulation of epithelial function. Cancer Treat Res, 1991,53:335-364.
22. Lee YJ, Streuli, C H. Extracellular matrix selectively modulates the response of mammary epithelial cells to different soluble signalling ligands. J Cell Biol, 1999, 274: 22401-22408.
23. Jones JL, Critchley DR, Walker RA. Alteration of stromal protein and integrin expression in breast-a marker of premalignant change? J Pathol, 1992, 167: 399-406.
24. Ishihara A, Yoshida T, Tamaki H, et al. Tenascin expression in cancer cells and stroma of human breast cancer and its prognostic significance. Clin Cancer Res,1995,1 (9) :1035-1041.
25. Jahkola T, Toivonen T, Virtanen I, et al. Tenascin-C expression in invasion border of early breast cancer: a predictor of local and distant reccurence. Br J Cancer, 1998,78 (11) :1507-1513.
26. Zagzag D, Friedlander DR, Dosik J, et al. Tenascin-C expression by angiogenic vessels in human astrocytomas and by human brain endothelial cells in vitro. Cancer Res, 1996,56 (1) :182-189.
27. Yoshida T, Ishihara A, Hirokawa Y, et al. Tenascin in breast cancer development-is epithelial tenascin a marker for poor prognosis? Cancer Lett, 1995,90 (1) :65-73.
28. Folkman J. Tumor angiogenesis. Adv Cancer Res, 1985,43: 175-203.
29. Liotta LA, Steeg PS, Stetler-Stevenson WG. Cancer metastasis and angiogenesis: an imbalance of positive and negative regulation. Cell, 1991, 64: 327-336.
30. Srivastava A, Laidler P, Davies RP, et al. The prognostic significance of tumor vascularity in intermediate thickness (0.76-4.0mm thick) skin melanoma. A quantitative histologic study. Am J Pathol, 1988,133: 419- 423.
31. Macchiarini P, Fontani G, Hardin MJ, et al. Relation of neovascularization to metastasis of non-small cell lung cancer. Lancet, 1992, 340:145-146.
32. Weidner N, Semple JP, Welch WR, et al. Tumor angiogenesis and metastasis: Correlation in invasive breast carcinoma. N Engl J Med, 1991, 324:1-8.
33. Dickinson AJ, Fox SB, Persad RA, et al. Quantification of angiogenesis as an independent predictor of prognosis in invasive bladder carcinoma. Br J Urol, 1994, 74: 762-766.
34. Weidner N, Carroll PR, Flax J, et al. Tumor angiogenesis correlates with metastasis in invasive prostate carcinoma. Am J Pathol, 1993, 143:401-409.
35. Wiggins DL, Granai CO, Steinhoff MM, et al. Tumor angiogenesis as a prognostic factor in cervical carcinoma. Gynecol Oncol, 1995, 56:353-356.
36. El-Assal ON, Yamanoi A, Soda Y, et al. Clinical significance of microvessel density and vascular endothelial growth factor expression in hepatocellular carcinoma and surrounding liver: possible involvement of vascular endothelial growth factor in the angiogenesis of cirrhotic liver. Hepatology, 1998, 27: 1554-1562.
37. Tanigawa N, Lu C, Mitsui T, et al.Quantitation of sinusoid-like vessels in hepatocellular carcinoma: its clinical and prognostic significance. Hepatology, 1997,26: 1216-1223.
38. Poon RT, Ng IO, Lau C, et al.Tumor microvessel density as a predictor of recurrence after resection of hepatocellular carcinoma: a prospective study. J Clin Oncol, 2002, 20: 1775-1785.
39. Wang JM, Kumar S, Pye D, et al. Breast carcinomaxomparative study of tumor vasculature using two endothelial cell markers. J Natl Cancer Inst, 1994, 86:386-388.
40. Burrows FJ, Derbyshire EJ, Tazzari PL, et al. Up-regulation of endoglin on vascular endothelial cells in human solid tumors implications for diagnosis and therapy. Clin Cancer Res, 1995,1:1623-1634.
41. Matsuno F, Haruta Y, Kondo M, et al.Induction of lasting complete regression of preformed distinct solid rumors by targeting the tumor vasculature using two new anti-endoglin monoclonal antibodies. Clin Cancer Res, 1999, 5: 371-382.
42. Tanaka F, Otake Y, Yanagihara K, et al. Evaluation of angiogenesis in non-small cell lung cancer: comparison between anti-CD34 antibody and anti-CD105 antibody. Clin Cancer Res, 2001, 7: 3410-3415.
43. Yu DC, Zhuang LY, Sun XT, et al. Particular distribution and expression pattern of endoglin (CD105) in the liver of patients with hepatocellular carcinoma. BMC Cancer, 2007, 7:122-132.
1. Cadigan KM, Liu YI. Wnt signaling: complexity at the surface. J Cell Sci, 2006, 119(Pt 3): 395-402.
2. Jamora C, Fuchs E. Intercellular adhesion, signaling and the cytoskeleton. Nat Cell Biol, 2002,4(4):E101-108.
3. Muller T, Bain G, Wang X, et al.Regulation of epithelial cell migration and tumor formation by β-catenin signaling. Exp Cell Res, 2002,280(1): 119-133.
4. Kurayoshi M, Oue N, Yamamoto H, et al. Expression of Wnt-5a is correlated with aggressiveness of gastric cancer by stimulating cell migration and invasion.Cancer Res, 2006, 66(21): 10439-10448.
5. Dejmek J, Dejmek A, Safholm A, et al. Wnt-5a protein expression in primary dukes B colon cancers identifies a subgroup of patients with good prognosis. Cancer Res, 2005, 65(20): 9142-9146.
6. Jonsson M, Dejmek J, Bendahl PO, et al. Loss of Wnt-5a protein is associated with early relapse in invasive ductal breast carcinomas. Cancer Res, 2002, 62(2): 409-416.
7. Sobin L H, Wittekind C, eds. TNM classification of malignant tumors [M]. 6th ed. New York: Wiley—Liss, 2002: 81-83.
8. Edmondson HA, Steiner PE. Primary carcinoma of the liver: a study of 100 cases among 48,900 necropsies. Cancer, 1954, 7: 462-503.
9. Mattern J, Koomagi R, Volm M. Association of vascular endothelial growth factor expression with intratumoral microvessel density and tumour cell proliferation in human epidermoid lung carcinoma. Br J Cancer, 1996, 73(7): 931-934.
10. Weidner N. Current pathologic methods for measuring intratumoral microvessel density within breast carcinoma and other solid tumors. Breast Cancer Res Treat, 1995,36(2):169-180.
11. Nusse R, Varmus HE. Many tumors induced by the mouse mammary tumor virus contain a provirus intergration in the same region of the host genome. Cell, 1982, 31:99-109.
12. Masaru K. WNT and FGF gene clusters. International journal of oncology, 2002, 21: 1269-1273.
13. Du SJ, Purcell SM, Christian JL, et al. Identification of distinct classes and functional domains of Wnts through expression of wild-type and chimeric proteins in Xenopus embryos. Mol Cell Biol, 1995, 15:2625-2634.
14. Shimizu H, Julius MA, Giarre M, et al. Transformation by Wnt family proteins correlates with regulation of beta-catenin.Cell Growth Differ, 1997, 8: 1349-1358.
15. Giles RH, van Es JH, Clevers H. Caught up in a Wnt storm: Wnt signaling in cancer. Biochim Biophys Acta, 2003, 1653:1-24.
16. Torres MA, Yang-Snyder JA, Purcell SM, et al. Activities of the Wnt-1 class of secreted signaling factors are antagonized by the Wnt-5A class and by a dominant negative cadherin in early Xenopus development. J Cell Biol, 1996, 133: 1123-1137.
17. Clark CC, Cohen I, Eichstetter I, et al. Molecular cloning of the human proto-oncogene Wnt-5A andmapping of the gene (WNT-5A) to chromosome 3p14-p21. Genomics, 1993,18 (2):249-260.
18. Mikels A J,Nusse R. Purified Wnt5a protein activates or inhibits beta-catenin-TCF signaling depending on receptor context. PLoS Biol, 2006,4(4):e115.
19. Clark CC, Cohen IR, Iozzo RV, et al. Molecular cloning of the human proto-oncogene Wnt-5A and mapping of the gene(WNT5A)to chromosome 3p14-p21. Genomics, 1993,18(2):249-260.
20. Wang HY, Malbon CC. Wnt signaling, Ca2+, and cyclic GMP: visualizing Frizzled functions. Science, 2003, 300(5625): 1529-1530.
21. Varma RR, Hector SM, Clark K. et al.Gene expression profiling of a clonal isolate of oxaliplatin-resistant ovarian carcinoma cell line A2780/C10. Oncol Rep, 2005, 14(4): 925-932.
22. Bachmann IM, Straume O, Puntervoll HE. et al. Importance of P-cadherin, beta-catenin, and Wnt5a/frizzled for progression of melanocytic tumors and prognosis in cutaneous melanoma. Clin Cancer Res, 2005, 11(24 Pt l):8606-8614.
23. Ying J, Li H, Yu J, et al. WNT5A exhibits tumor-suppressive activity through antagonizing the Wnt/beta-catenin signaling, and is frequently methylated in colorectal cancer. Clin Cancer Res, 2008,14(1):55-61.
24. Blanc E,Roux GL,Benard J, et al.Low expression of Wnt-5a gene is associated with high-risk neuroblastoma. Oncogene, 2005,24(7): 1277-1283.
25. Dejmek J, Leandersson K, Manjer J, et al.Expression and signaling activity of Wnt-5a/discoidin domain receptor-1 and Syk plays distinct but decisive roles in breast cancer patient survival. Clin Cancer Res, 2005 ,11(2 Pt l):520-528.
26. Liang H, Chen Q, Coles AH, et al. Wnt5a inhibits B cell proliferation and functions as a tumor suppressor in hematopoietic tissue. Cancer Cell, 2003, 4(5): 349-360.
27. Roman-Gomez J, Jimenez-Velasco A, Cordeu L, et al. WNT5A, a putative tumour suppressor of lymphoid malignancies, is inactivated by aberrant methylation in acute lymphoblastic leukaemia. Eur J Cancer, 2007 ,43(18):2736-2746.
28. Ying J, Li H, Chen YW, et al. WNT5A is epigenetically silenced in hematologic malignancies and inhibits leukemia cell growth as a tumor suppressor. Blood, 2007,110(12):4130-4132.
29. Kremenevskaja N, von Wasielewski R, Rao AS, et al. Wnt-5a has tumor suppressor activity in thyroid carcinoma. Oncogene, 2005,24(13):2144-2154.
30. Iozzo RV, Eichstetter I, Danielson KG. Aberrant expression of the growth factor Wnt-5a in human malignancy. Cancer Res, 1995, 55(16):3495-3499.
31. Huang CL, Liu D, Nakano J, et al.Wnt5a expression is associated with the tumor proliferation and the stromal vascular endothelial growth factor--an expression in non-small-cell lung cancer. J Clin Oncol, 2005,23(34):8765-8773.
32. Lejeune S, Huguet EL, Hamby A, et al. Wnt5a cloning,expression,and up-regulation in human primary breast cancers. Clin Cancer Res, 1995, 1(2): 215-222.
33. Liu XH, Pan MH, Lu ZF, et al. Expression of Wnt-5a and its clinicopathological significance in hepatocellular carcinoma. Dig Liver Dis, 2008, 40(7):560-567.
34. Huguet EL, Smith K, Bicknell R, et al. Regulation of Wnt-5a mRNA expression in human mammary epithelial cells by cell shape, confluence, and hepatocyte growth factor. J Biol Chem, 1995,270:12851-12856.
35. Olson DJ, Oshimura M, Otte AP. Ectopic Expression of wnt-5a in Human Renal Cell Carcinoma Cells Suppresses in vitro Growth and Telomerase Activity. Tumor Biol, 1998,19:244-252.
36. Harley CB, Villeponteau B. Telomere and telomerase in aging and cancer. Curr Opin Genet Dev, 1995, 5:249-255.
37. Nathan J. Palpant, So-ichiro, et al. Non-canonical Wnt Signaling Enhances differentiation of Scal+/c-kit+ Adipose-derived Murine Stromal Vascular Cells into Spontaneously Beating Cardiac Myocytes. J Mol Cell Cardiol, 2007, 43(3): 362-370.
38. Mishra L, Derynck R, Mishra B. Transforming growth factor-beta sig-naling in stem cells and cancer. Science, 2005, 310:68-71.
39. Boxer GM, Tsiompanou E, Levine T, et al. Immunohistochemical expression of vascular endothelial growth factor and microvessel counting as prognostic indicators in node-negative colorectal cancer. Tumour Biol, 2005, 26(1): 1-8.
40. Srivastava A, Laidler P, Davies RP, et al. The prognostic significance of tumor vascularity in intermediate thickness (0.76-4.0mm thick) skin melanoma. A quantitative histologic study. Am J Pathol, 1988, 133: 419-423.
41. Macchiarini P, Fontani G, Hardin MJ, et al. Relation of neovascularization to metastasis of non-small cell lung cancer. Lancet, 1992, 340:145-146.
42. Weidner N, Semple JP, Welch WR, et al. Tumor angiogenesis and metastasis: Correlation in invasive breast carcinoma. N Engl J Med, 1991, 324:1-8.
43. Dickinson AJ, Fox SB, Persad RA, et al. Quantification of angiogenesis as an independent predictor of prognosis in invasive bladder carcinoma. Br J Urol, 1994, 74: 762-766.
44. Weidner N, Carroll PR, Flax J, et al. Tumor angiogenesis correlates with metastasis in invasive prostate carcinoma. Am J Pathol, 1993, 143:401-409.
45. Wiggins DL, Granai CO, Steinhoff MM, et al. Tumor angiogenesis as a prognostic factor in cervical carcinoma. Gynecol Oncol, 1995, 56:353-356.
46. Masckauchan TN, Agalliu D, Vorontchikhina M, et al. Wnt5a Signaling Induces Proliferation and Survival of Endothelial Cells In Vitro and Expression of MMP-land Tie-2. Mol Biol Cell, 2006,17(12):5163-5172.
47. Cheng CW, Yeh JC, Fan TP, et al. Wnt5a-mediated non-canonical Wnt signalling regulates human endothelial cell proliferation and migration. Biochem Biophys Res Commun. 2008, 365(2):285-290.
48. Goodwin AM, Kitajewski J, D'Amore PA. Wntl and Wnt5a affect endothelial proliferation and capillary length; Wnt2 does not. Growth Factors, 2007, 25(1): 25-32.
2.金世龙,刘宝华,刘宏鸣,等.大鼠肝卵圆细胞的分离培养及干细胞标志分析.中国普外基础与临床杂志,2007,14(1):68-71.
3.Zheng YW,Taniguchi H.Diversity of hepatic stem cells in the fetal and adult liver.Semin Liver Dis,2003,23(4):337-348.
4.Paku S,Schnur J,Nagy P,et al.Origin and structural evolution of the early proliferating oval cells in rat liver.Am J Pathol,2001,158(4):1313-1321
5.Lemmer ER,Shepard EG,Blakolmer K,Kirsch RE,Robson SC:Isolation from human fetal liver of cells co-expressing CD34 haematopoietic stem cell and CAM 5.2 pancytokera-tin markers.J Hepatol,1998,29:450-454.
6.Libbrecht L,Desmet V,Van DB,et al.The immunohistochemical phenotype of dysplastic foci in human liver:correlation with putative progenitor cells.J Hepatol,2000,33:76-84.
7.Van EP,Desmet VJ.Cytokeratin and the liver.Liver,1993,13:113-122.
8.Sobin L H,Wittekind C,eds.TNM classification of malignant rumors[M].6th ed.New York:Wiley—Liss,2002:81-83.
9.Edmondson HA,Steiner PE.Primary carcinoma of the liver:a study of 100 cases among 48,900 necropsies.Cancer,1954,7:462-503.
10.Hsia CC,Evarts RP,Nakatsukasa H,et al.Occurrence of oval type cells in hepatitis B virus associated human hepatocacinogenesis.Hepatology,1992,16(6):1327-1333.
11.Mattern J,Koom(a|¨)gi R,Volm M.Association of vascular endothelial growth factor expression with intratumoral microvessel density and tumour cell proliferation in human epidermoid lung carcinoma.Br J Cancer,1996,73(7):931-934.
12.Klein WM,Molmenti EP,Colombani PM,et al.Primary liver carcinoma arising in people younger than 30 years.Am J Clin Pathol,2005,124(4):512-518.
13.Kara B,Doran F,Kara IO,et al.Expression of c-kit protooncogen in hepatitis B virus-induced chronic hepatitis,cirrhosis and hepatocellular carcinoma:has it a diagnostic role? Int J Clin Pract, 2008, 62(8): 1206-1211.
14. Tsuji M, Kashihara T, Terada N, et al. An immunohistochemical study of hepatic atypical adenomatous hyperplasia, hepatocellular carcinoma, and cholangiocarcinoma with alpha-fetoprotein, carcinoembryonic antigen, CA19-9, epithelial membrane antigen, and cytokeratins 18 and 19. Pathol Int, 1999, 49 (4): 310-317.
15. Desmet VJ. Organizational principles; in Arias IM, Boyer JL, Chisari FV (eds): The Liver, Biology and Pathology. Philadelphia, Lippincott Williams & Wilkins, 2001, pp3-15.
16. Libbrecht L, Roskams T. Hepatic progenitor cells in human liver diseases. Semin Cell Dev Biol, 2002,13: 389-396.
17. Roskams TA, Theise ND, Balabaud C, et al. Nomenclature of the finer branches of the biliary tree: canals, ductules, and ductular reactions in human livers. Hepatology, 2004, 39(6): 1739-1745.
18. De Vos R, Desmet V. Ultrastructural characteristics of novel epithelial cell types indentified in human pathologic liver specimens with chronic ductular reaction. Am J Pathol, 1992,140, (6): 1441-1450.
19. Thorgeirsson SS. Hepatic stem cells. Am J Pathol, 1993, 142: 1331-1333.
20. Strick-Marchand H, Morosan S, Charneau P, et al. Bipotential mouse embryonic liver stem cell lines contribute to liver regeneration and differentiate as bile ducts and hepatocytes . Proc Natl Acad Sci USA, 2004, 101(22) : 8360-8365.
21. Thorgeirsson SS. Hepatic stem cells in liver regeneration. FASEB J, 1996, 10: 1249-1256.
22. Runge D, Runge DM, Jager D, et al. Serum-free, long-term cultures of human hepatocytes: maintenance of cell morphology, transcription factors, and liver specific functions. Biochem Biophys Res Commun, 2000, 269(1): 46-53.
23. Braun KM, Thompson AW, Sandgren EP. Hepatic microenvironment affects oval cell localization in albumin urokinase type plasminogen activator transgenic mice. Am J Pathol, 2003, 162(1):195-202.
24. Lowes KN, Brennan BA, Yeoh GC, et al. Oval cell numbers in human chronic liver diseases are directly related to disease severity. Am J Pathol, 1999, 154: 537-541.
25. Libbrecht L, Desmet V, Van Damme B, et al. Deep intralobular extension of human hepatic'progenitor cells'correlates with parenchymal inflammation in chronic viral hepatitis: can 'progenitor cells' migrate? J Pathol, 2000, 192: 373-378.
26. Eleazar JA, Memeo L, Jhang JS, et al. Progenitor cell expansion: an important source of hepatocyte regeneration in chronic hepatitis. J Hepatol, 2004, 41: 983-991.
27. Moore KA, Lemischka IR. Stem cells and their niches. Science, 2006, 311(5769): 1880-1885.
28. Sell S. Stem cell origin of cancer and differentiation therapy. Crit Rev Oncol Hematol, 2004, 51:1-28.
29. Dontu M, Al-Hajj, Abdallah WM, et al. Stem cell in normal breast development and breast cancer. Cell Prolif, 2003, 36(Suppll): 59-72.
30. Singh SK, Hawkins C, Clarke ID, et al. Identification of human brain tumour initiating cells. Nature, 2004,432: 396-401.
31. Burger PE, Xiong X, Coetzee S, et al. Sca-1 expression identifies stem cell in the proximal region of prostatic ducts with high capacity to reconstitute prostatic tissue. Proc Natl Acad USA, 2005,102: 7180-7185.
32. Sell S. Cellular origin of cancer: dedifferentiation or stem cell maturation arrest? Environ Health Perspect, 1993,101 Suppl 5: 15-26.
33. Sell S, Pierce G B. Maturation arrest of stem cell differentiation is a common pathway for the cellular origin of teratocarcinomas and epithelial cancers. Lab Invest, 1994, 70(1): 6-22.
34. Sell S. Heterogeneity and plasticity of hepatocyte lineage cells. Hepatology, 2001, 33(3) : 738-750.
35. Tanaka S, Yamamoto T, Tanaka H, et al. Potentiality of combined hepatocellular and intrahepatic cholangiocellular carcinoma originating from a hepatic precursor cell: Immunohistochemical evidence. Hepatol Res, 2005, 32(1) .' 52-57.
36. Ruck P, Xiao JC, Kaiserling E. Small epithelial cells and the histogenesis of hepatoblastoma. Electron microscopic, immunoelectron microscopic, and immunohistochemical findings.Am J Pathol,1996,148(1):321-329.
37.Robrechts C,De-Vos R,Van-den-Heuvel M,et al.Primary liver tumor of intermediate hepatocyte-bile duct cell,phenotype:a progenitor cell tumour? Liver,1998,18:288-293.
38.Sell S,Xu KL,Huff WE,et al.Aflatoxin exposure produces serum alphafetoprotein elevations and marked oval cell proliferation in young male Pekin ducklings.Pathology,1998,30(1):34-39.
39.Tsao MS,Grisham JW.Hepatocarcinomas,cholangiocarcinomas,and hepatoblastomas produced by chemically transformed cultured rat liver epithelial cells-a light and electron microscopic analysis.Am J Pathol,1985,127:168-181.
40.陈琼荣,向锦,廖冰,等.卵圆细胞在实验性肝癌发生过程中的演变特征.癌症,2007,26(7):719-723.
41.Yoon DS,Jeong J,Park YN,et al.Expression of biliary antigen and its clinical significance in hepatocellular carcinoma.Yonsei Med J,1999,40(5):472-477.
42.Van Eyken P,Sciot R,Paterson A,et al.Cytokeratin expression in hepatocellular carcinoma:an immunohistochemical study.Hum Pathol,1988,19(5):562-568.
43.Uenishi T,Kubo S,Yamamoto T,et al.Cytokeratin 19 expression in hepatocellular carcinoma predicts early postoperative recurrence.Cancer Sci,2003,94(10):851-857.
44.Fausto N.Liver regeneration and repair:hepatocytes,Progenitor cells,and stem cells.Hepatology,2004,39:1477-1487.
45.Seeger C,Mason WS.Hepatitis B virus biology.Microbiol Mol Biol Rev,2000,64:51-68.
46.Blakolmer K,Jaskiewicz K,Dunsford HA,et al.Hematopoietic stem cell markers are expressed by ductal plate and bile duct cells in developing human liver.Hepatology,1995,21(6):1510-1516.
47.Kiiasov AP,Gumerova AA,Bilalov MM,et al.Cytokeratin expression in the pre-and postnatal ontogeny of the rat liver.Ontogenez,1997,28(5):389-93.
48.Fu Y,Zhang W,Liu Y F,et al.Abnormal expression of cytokeratins in hepatocellular carcinoma cell lines and its mechanism.J Fourth Mil Med Univ,2001,22(7):598-603.
49.Van Eyken P,Sciot R,Desmet VJ.A cytokeratin immunohistochemical study of cholestatic liver disease:evidence that hepatocytes can express bile duct type cytokeratins.Histopathol,1989,15(2):125-135.
50.Balaton AJ,Gotheil N-SC,Callard P,et al.Distribution between hepatocellular carcinoma,cholangiocarcionma,and metastatic carcinoma based on immunohistochemical staining for carcinoembryonic antigen and for CK19 on paraffin sections.J Pathol,1988,156:305-310.
51.Uenishi T,Kubo S,Hirohashi K,et al.Expression of bile duct-type cytokeratin in hepatocellular carcinoma in patients with hepatitis C virus and prior hepatitis B virus infection.Cancer Lett,2002,178(1):107-112.
52.肖家诚,王天翔,Ruck P等.肝硬化组织中卵圆细胞的光镜、电镜与免疫电镜观察.临床与实验病理学杂志,2002,18(1):57-60.
53.Goodman ZD,Ishak KG,Langloss JM,et al.Combined hepatocellular-cholangiocarcinoma.A histologic and immunohistochemical study.Cancer,1985,55:124-135.
54.Tickoo SK,Zee SY,Obiekwe S,et al.Combined hepatocellular-cholangiocarcinoma:a histopathologic,immunohistochemical,and in situ hybridization study.Am J Surg Pathol,2002,26:989-997.
55.Kim H,Park C,Han KH,et al.Primary liver carcinoma of intermediate (hepatocyte-cholangiocyte) phenotype.J Hepatol,2004,40:298-304.
56.Liaw YF,Tai DI,Chu CM,et al.Early detection of hepatocellular carcinoma in patients with chronic type B hepatitis A prospective study.Gastroenterology,1986,90:263-267.
57.Crosby HA,Kelly DA,Strain AJ.Human hepatic stem-like cells isolated using c-kit or CD34 can differentiate into biliary epithelium.Gastroenterology,2001,120:534-544.
58.Matsusaka S,Tsujimura T,Toyosaka A,et al.Role of c-kit receptor tyrosine kinase in development of oval cells in the rat 2-acetylaminoffuorene/partial hepatectomy model.Hepatology,1999,29:670-676.
59.Blakolmer K,Jaskiewicz K,Dunsford HA,et al.Hematopoietic stem cell markers are expressed by ductal plate and bile duct cells in developing human liver. Hepatology, 1995,21:1510-1516.
60. Yamamoto T, Uenishi T, Ogawa M, et al. Immunohistologic Attempt to Find Carcinogenesis from Hepatic Progenitor Cell in Hepatocellular Carcinoma. Digestive Surgery, 2005, 22(5):364-370.
1.吴孟超.原发性肝癌的诊断和治疗进展.中华外科杂志,1998,36(9):515-518.
2.王悦华,冯玉泉,刘永雄,等.原发性肝癌的外科治疗.中华普通外科杂志,1997,12(6):325-328.
3.Poon RT,Fan ST,Ng IO,et al.Different risk factors and prognosis for late intrahepatic recurrence after resection of hepatocarcinoma.Cancer,2000,89(3):5O0-507.
4.Folkman J.What is the evidence that tumors are angiogenesis dependent?.J Natl Cancer Inst,1990,82(1):4-6.
5.齐庆安,别平.原发性肝细胞肝癌血小板源性生长因子的表达和肿瘤血管形成关系的相关研究.消化外科,2005,4(2):119-123.
6.穆四清,张峰.原发性肝癌复发转移的分子机理.中国普外基础与临床杂志,2007,14(1):49-51.
7.Chiquet-Ehrismann R.Tenascins,a growing family of extracellular matrix proteins.Experientia,1995,51(9-10):853-862.
8.Midwood KS,Schwarzbauer JE.Tenascin-C modulates matrix contraction via focal adhesion kinase-and Rho-mediated signaling pathways.Mol Biol Cell,2002,13(10):3601-3613.
9.Chiquet-Ehrismann R.Anti-adhesive molecules of the extracellular matrix.Curr Opin Cell Biol,1991,3(5):800-804.
10.Jones PL,Cowan KN,Rabinovitch M.Tenascin-C,proliferation and subendothelial fibronectin in progressive pulmonary vascular disease.Am J Pathol,1997,150(4):1349-1360.
11.Erickson HP,Bourdon MA.Tenascin:an extracellular matrix protein prominent in specialized embryonic tissues and tumors.Annu Rev Cell Biol,1989,4(5):71-92.
12.Koukoulis GK,Gould VE,Bhattacharyya A,et al.Tenascin in normal,reactive,hyperplastic,and neoplastic tissues:biologic and pathologic implications.Hum Pathol,1991,22(7):636-643.
13.Boxer GM,Tsiompanou E,Levine T,et al.Immunohistochemical expression of vascular endothelial growth factor and microvessel counting as prognostic indicators in node-negative colorectal cancer. Tumour Biol, 2005, 26(1):1-8.
14. Vermeulen PB, Gasparinig G, Fox SB, et al. Quantification of angiogenesis in solid human tumours: an international consensus on the methodology and criteria of evaluation. Eur J Cancer, 1996, 32A(14): 2474-2484.
15. Wang JM, Kumar S, Pye D, et al. Breast carcinoma: comparative study of tumor vasculature using endothelial cell markers. J Nat Cancer Inst, 1994, 86:386-388.
16. Miller DW, Graulich W, Karges B, et al. Elwvated expression of endogl in, a component of the TGF-O-receptor complex, correlates with proliferation of tumor endothelial cells. Int J Cancer, 1999, 81:568-572.
17. Brewar CA, Setterdahl JJ, Li MJ, et al. Endoglin expression as a mesure of microvessel density in cervical cancer. Obstet Gynecol, 2000, 96:224-228.
18. Tanaka F, Otake Y, Yanagihara K, et al. Evaluation in angiogenesis in non-small cell lung cancer: comparison between anti-CD34 antibody and anti-CD 105 antibody. Clin. Cancer Res, 2001,7:3410-3415.
19. Mattern J, Koomagi R, Volm M. Association of vascular endothelial growth factor expression with intratumoral microvessel density and tumour cell proliferation in human epidermoid lung carcinoma. Br J Cancer, 1996, 73(7): 931-934.
20. Weidner N. Current pathologic methods for measuring intratumoral microvessel density within breast carcinoma and other solid tumors. Breast Cancer Res Treat, 1995,36(2):169-180.
21.Donjacour AA, Cunha GR. Stromal regulation of epithelial function. Cancer Treat Res, 1991,53:335-364.
22. Lee YJ, Streuli, C H. Extracellular matrix selectively modulates the response of mammary epithelial cells to different soluble signalling ligands. J Cell Biol, 1999, 274: 22401-22408.
23. Jones JL, Critchley DR, Walker RA. Alteration of stromal protein and integrin expression in breast-a marker of premalignant change? J Pathol, 1992, 167: 399-406.
24. Ishihara A, Yoshida T, Tamaki H, et al. Tenascin expression in cancer cells and stroma of human breast cancer and its prognostic significance. Clin Cancer Res,1995,1 (9) :1035-1041.
25. Jahkola T, Toivonen T, Virtanen I, et al. Tenascin-C expression in invasion border of early breast cancer: a predictor of local and distant reccurence. Br J Cancer, 1998,78 (11) :1507-1513.
26. Zagzag D, Friedlander DR, Dosik J, et al. Tenascin-C expression by angiogenic vessels in human astrocytomas and by human brain endothelial cells in vitro. Cancer Res, 1996,56 (1) :182-189.
27. Yoshida T, Ishihara A, Hirokawa Y, et al. Tenascin in breast cancer development-is epithelial tenascin a marker for poor prognosis? Cancer Lett, 1995,90 (1) :65-73.
28. Folkman J. Tumor angiogenesis. Adv Cancer Res, 1985,43: 175-203.
29. Liotta LA, Steeg PS, Stetler-Stevenson WG. Cancer metastasis and angiogenesis: an imbalance of positive and negative regulation. Cell, 1991, 64: 327-336.
30. Srivastava A, Laidler P, Davies RP, et al. The prognostic significance of tumor vascularity in intermediate thickness (0.76-4.0mm thick) skin melanoma. A quantitative histologic study. Am J Pathol, 1988,133: 419- 423.
31. Macchiarini P, Fontani G, Hardin MJ, et al. Relation of neovascularization to metastasis of non-small cell lung cancer. Lancet, 1992, 340:145-146.
32. Weidner N, Semple JP, Welch WR, et al. Tumor angiogenesis and metastasis: Correlation in invasive breast carcinoma. N Engl J Med, 1991, 324:1-8.
33. Dickinson AJ, Fox SB, Persad RA, et al. Quantification of angiogenesis as an independent predictor of prognosis in invasive bladder carcinoma. Br J Urol, 1994, 74: 762-766.
34. Weidner N, Carroll PR, Flax J, et al. Tumor angiogenesis correlates with metastasis in invasive prostate carcinoma. Am J Pathol, 1993, 143:401-409.
35. Wiggins DL, Granai CO, Steinhoff MM, et al. Tumor angiogenesis as a prognostic factor in cervical carcinoma. Gynecol Oncol, 1995, 56:353-356.
36. El-Assal ON, Yamanoi A, Soda Y, et al. Clinical significance of microvessel density and vascular endothelial growth factor expression in hepatocellular carcinoma and surrounding liver: possible involvement of vascular endothelial growth factor in the angiogenesis of cirrhotic liver. Hepatology, 1998, 27: 1554-1562.
37. Tanigawa N, Lu C, Mitsui T, et al.Quantitation of sinusoid-like vessels in hepatocellular carcinoma: its clinical and prognostic significance. Hepatology, 1997,26: 1216-1223.
38. Poon RT, Ng IO, Lau C, et al.Tumor microvessel density as a predictor of recurrence after resection of hepatocellular carcinoma: a prospective study. J Clin Oncol, 2002, 20: 1775-1785.
39. Wang JM, Kumar S, Pye D, et al. Breast carcinomaxomparative study of tumor vasculature using two endothelial cell markers. J Natl Cancer Inst, 1994, 86:386-388.
40. Burrows FJ, Derbyshire EJ, Tazzari PL, et al. Up-regulation of endoglin on vascular endothelial cells in human solid tumors implications for diagnosis and therapy. Clin Cancer Res, 1995,1:1623-1634.
41. Matsuno F, Haruta Y, Kondo M, et al.Induction of lasting complete regression of preformed distinct solid rumors by targeting the tumor vasculature using two new anti-endoglin monoclonal antibodies. Clin Cancer Res, 1999, 5: 371-382.
42. Tanaka F, Otake Y, Yanagihara K, et al. Evaluation of angiogenesis in non-small cell lung cancer: comparison between anti-CD34 antibody and anti-CD105 antibody. Clin Cancer Res, 2001, 7: 3410-3415.
43. Yu DC, Zhuang LY, Sun XT, et al. Particular distribution and expression pattern of endoglin (CD105) in the liver of patients with hepatocellular carcinoma. BMC Cancer, 2007, 7:122-132.
1. Cadigan KM, Liu YI. Wnt signaling: complexity at the surface. J Cell Sci, 2006, 119(Pt 3): 395-402.
2. Jamora C, Fuchs E. Intercellular adhesion, signaling and the cytoskeleton. Nat Cell Biol, 2002,4(4):E101-108.
3. Muller T, Bain G, Wang X, et al.Regulation of epithelial cell migration and tumor formation by β-catenin signaling. Exp Cell Res, 2002,280(1): 119-133.
4. Kurayoshi M, Oue N, Yamamoto H, et al. Expression of Wnt-5a is correlated with aggressiveness of gastric cancer by stimulating cell migration and invasion.Cancer Res, 2006, 66(21): 10439-10448.
5. Dejmek J, Dejmek A, Safholm A, et al. Wnt-5a protein expression in primary dukes B colon cancers identifies a subgroup of patients with good prognosis. Cancer Res, 2005, 65(20): 9142-9146.
6. Jonsson M, Dejmek J, Bendahl PO, et al. Loss of Wnt-5a protein is associated with early relapse in invasive ductal breast carcinomas. Cancer Res, 2002, 62(2): 409-416.
7. Sobin L H, Wittekind C, eds. TNM classification of malignant tumors [M]. 6th ed. New York: Wiley—Liss, 2002: 81-83.
8. Edmondson HA, Steiner PE. Primary carcinoma of the liver: a study of 100 cases among 48,900 necropsies. Cancer, 1954, 7: 462-503.
9. Mattern J, Koomagi R, Volm M. Association of vascular endothelial growth factor expression with intratumoral microvessel density and tumour cell proliferation in human epidermoid lung carcinoma. Br J Cancer, 1996, 73(7): 931-934.
10. Weidner N. Current pathologic methods for measuring intratumoral microvessel density within breast carcinoma and other solid tumors. Breast Cancer Res Treat, 1995,36(2):169-180.
11. Nusse R, Varmus HE. Many tumors induced by the mouse mammary tumor virus contain a provirus intergration in the same region of the host genome. Cell, 1982, 31:99-109.
12. Masaru K. WNT and FGF gene clusters. International journal of oncology, 2002, 21: 1269-1273.
13. Du SJ, Purcell SM, Christian JL, et al. Identification of distinct classes and functional domains of Wnts through expression of wild-type and chimeric proteins in Xenopus embryos. Mol Cell Biol, 1995, 15:2625-2634.
14. Shimizu H, Julius MA, Giarre M, et al. Transformation by Wnt family proteins correlates with regulation of beta-catenin.Cell Growth Differ, 1997, 8: 1349-1358.
15. Giles RH, van Es JH, Clevers H. Caught up in a Wnt storm: Wnt signaling in cancer. Biochim Biophys Acta, 2003, 1653:1-24.
16. Torres MA, Yang-Snyder JA, Purcell SM, et al. Activities of the Wnt-1 class of secreted signaling factors are antagonized by the Wnt-5A class and by a dominant negative cadherin in early Xenopus development. J Cell Biol, 1996, 133: 1123-1137.
17. Clark CC, Cohen I, Eichstetter I, et al. Molecular cloning of the human proto-oncogene Wnt-5A andmapping of the gene (WNT-5A) to chromosome 3p14-p21. Genomics, 1993,18 (2):249-260.
18. Mikels A J,Nusse R. Purified Wnt5a protein activates or inhibits beta-catenin-TCF signaling depending on receptor context. PLoS Biol, 2006,4(4):e115.
19. Clark CC, Cohen IR, Iozzo RV, et al. Molecular cloning of the human proto-oncogene Wnt-5A and mapping of the gene(WNT5A)to chromosome 3p14-p21. Genomics, 1993,18(2):249-260.
20. Wang HY, Malbon CC. Wnt signaling, Ca2+, and cyclic GMP: visualizing Frizzled functions. Science, 2003, 300(5625): 1529-1530.
21. Varma RR, Hector SM, Clark K. et al.Gene expression profiling of a clonal isolate of oxaliplatin-resistant ovarian carcinoma cell line A2780/C10. Oncol Rep, 2005, 14(4): 925-932.
22. Bachmann IM, Straume O, Puntervoll HE. et al. Importance of P-cadherin, beta-catenin, and Wnt5a/frizzled for progression of melanocytic tumors and prognosis in cutaneous melanoma. Clin Cancer Res, 2005, 11(24 Pt l):8606-8614.
23. Ying J, Li H, Yu J, et al. WNT5A exhibits tumor-suppressive activity through antagonizing the Wnt/beta-catenin signaling, and is frequently methylated in colorectal cancer. Clin Cancer Res, 2008,14(1):55-61.
24. Blanc E,Roux GL,Benard J, et al.Low expression of Wnt-5a gene is associated with high-risk neuroblastoma. Oncogene, 2005,24(7): 1277-1283.
25. Dejmek J, Leandersson K, Manjer J, et al.Expression and signaling activity of Wnt-5a/discoidin domain receptor-1 and Syk plays distinct but decisive roles in breast cancer patient survival. Clin Cancer Res, 2005 ,11(2 Pt l):520-528.
26. Liang H, Chen Q, Coles AH, et al. Wnt5a inhibits B cell proliferation and functions as a tumor suppressor in hematopoietic tissue. Cancer Cell, 2003, 4(5): 349-360.
27. Roman-Gomez J, Jimenez-Velasco A, Cordeu L, et al. WNT5A, a putative tumour suppressor of lymphoid malignancies, is inactivated by aberrant methylation in acute lymphoblastic leukaemia. Eur J Cancer, 2007 ,43(18):2736-2746.
28. Ying J, Li H, Chen YW, et al. WNT5A is epigenetically silenced in hematologic malignancies and inhibits leukemia cell growth as a tumor suppressor. Blood, 2007,110(12):4130-4132.
29. Kremenevskaja N, von Wasielewski R, Rao AS, et al. Wnt-5a has tumor suppressor activity in thyroid carcinoma. Oncogene, 2005,24(13):2144-2154.
30. Iozzo RV, Eichstetter I, Danielson KG. Aberrant expression of the growth factor Wnt-5a in human malignancy. Cancer Res, 1995, 55(16):3495-3499.
31. Huang CL, Liu D, Nakano J, et al.Wnt5a expression is associated with the tumor proliferation and the stromal vascular endothelial growth factor--an expression in non-small-cell lung cancer. J Clin Oncol, 2005,23(34):8765-8773.
32. Lejeune S, Huguet EL, Hamby A, et al. Wnt5a cloning,expression,and up-regulation in human primary breast cancers. Clin Cancer Res, 1995, 1(2): 215-222.
33. Liu XH, Pan MH, Lu ZF, et al. Expression of Wnt-5a and its clinicopathological significance in hepatocellular carcinoma. Dig Liver Dis, 2008, 40(7):560-567.
34. Huguet EL, Smith K, Bicknell R, et al. Regulation of Wnt-5a mRNA expression in human mammary epithelial cells by cell shape, confluence, and hepatocyte growth factor. J Biol Chem, 1995,270:12851-12856.
35. Olson DJ, Oshimura M, Otte AP. Ectopic Expression of wnt-5a in Human Renal Cell Carcinoma Cells Suppresses in vitro Growth and Telomerase Activity. Tumor Biol, 1998,19:244-252.
36. Harley CB, Villeponteau B. Telomere and telomerase in aging and cancer. Curr Opin Genet Dev, 1995, 5:249-255.
37. Nathan J. Palpant, So-ichiro, et al. Non-canonical Wnt Signaling Enhances differentiation of Scal+/c-kit+ Adipose-derived Murine Stromal Vascular Cells into Spontaneously Beating Cardiac Myocytes. J Mol Cell Cardiol, 2007, 43(3): 362-370.
38. Mishra L, Derynck R, Mishra B. Transforming growth factor-beta sig-naling in stem cells and cancer. Science, 2005, 310:68-71.
39. Boxer GM, Tsiompanou E, Levine T, et al. Immunohistochemical expression of vascular endothelial growth factor and microvessel counting as prognostic indicators in node-negative colorectal cancer. Tumour Biol, 2005, 26(1): 1-8.
40. Srivastava A, Laidler P, Davies RP, et al. The prognostic significance of tumor vascularity in intermediate thickness (0.76-4.0mm thick) skin melanoma. A quantitative histologic study. Am J Pathol, 1988, 133: 419-423.
41. Macchiarini P, Fontani G, Hardin MJ, et al. Relation of neovascularization to metastasis of non-small cell lung cancer. Lancet, 1992, 340:145-146.
42. Weidner N, Semple JP, Welch WR, et al. Tumor angiogenesis and metastasis: Correlation in invasive breast carcinoma. N Engl J Med, 1991, 324:1-8.
43. Dickinson AJ, Fox SB, Persad RA, et al. Quantification of angiogenesis as an independent predictor of prognosis in invasive bladder carcinoma. Br J Urol, 1994, 74: 762-766.
44. Weidner N, Carroll PR, Flax J, et al. Tumor angiogenesis correlates with metastasis in invasive prostate carcinoma. Am J Pathol, 1993, 143:401-409.
45. Wiggins DL, Granai CO, Steinhoff MM, et al. Tumor angiogenesis as a prognostic factor in cervical carcinoma. Gynecol Oncol, 1995, 56:353-356.
46. Masckauchan TN, Agalliu D, Vorontchikhina M, et al. Wnt5a Signaling Induces Proliferation and Survival of Endothelial Cells In Vitro and Expression of MMP-land Tie-2. Mol Biol Cell, 2006,17(12):5163-5172.
47. Cheng CW, Yeh JC, Fan TP, et al. Wnt5a-mediated non-canonical Wnt signalling regulates human endothelial cell proliferation and migration. Biochem Biophys Res Commun. 2008, 365(2):285-290.
48. Goodwin AM, Kitajewski J, D'Amore PA. Wntl and Wnt5a affect endothelial proliferation and capillary length; Wnt2 does not. Growth Factors, 2007, 25(1): 25-32.