肝细胞癌组织中Glypican-3基因表达及其调控机制

目的：探讨肝细胞癌中Glypican-3（GPC3）基因表达及调控机制。方法:采用RT-PCR和聚合酶链反应-单链构象多态性(PCR-SSCP)方法分别检测48例肝细胞癌的癌组织、39例癌旁组织和31例正常肝组织中GPC3 mRNA的表达和基因突变；免疫组化S-P法检测GPC3、P53和PCNA蛋白的表达。结果:GPC3 mRNA在肝细胞癌的癌组织中的阳性表达率为77.1%，但其在癌旁和正常肝组织中不表达；肝细胞癌的癌组织中未发现GPC3基因突变；GPC3与P53蛋白表达无相关性（r＝-0.12574，P＞0.05），肝细胞癌的癌组织中GPC3 表达阳性和阴性的增殖细胞核抗原(PCNA)平均指数分别为（46.32±27.54）%、（39.83±21.47）%,P＞0.05。结论:肝细胞癌组织中GPC3 mRNA高表达与基因突变无关，没有直接影响P53和PCNA蛋白表达。     

肝细胞核因子-1α和肝细胞核因子-4α在人肝细胞癌中的表达及意义

目的 检测肝细胞癌（HCC）标本中肝细胞核因子-1α(HNF-1α)和肝细胞核因子-4α(HNF-4α)的表达,探讨其在肝癌发生、发展中的作用。 方法 采用反转录聚合酶链反应（RT-PCR）和免疫组织化学方法检测HNF-1α、HNF-4α mRNA和蛋白质在肝癌组织和癌旁肝组织中的表达。 结果 HNF-1α mRNA在癌旁肝组织中的相对表达量0.818±0.371明显高于肝细胞癌组织0.383±0.102,差异有显著性（P＜0.05）。HNF-4α mRNA在癌旁肝组织中的相对表达量0.846±0.384明显高于肝细胞癌组织0.397±0.105,差异有显著性（P＜0.05）。HNF-1α和HNF-4α mRNA 的表达与肿瘤分化程度有关（P＜0.05）。HNF-1α 蛋白在癌旁肝组织中的阳性率（92.3%）明显高于肝细胞癌组织（42.3%）,差异有显著性（P＜0.05）。HNF-4α 蛋白在癌旁肝组织中的阳性率（96.2%）明显高于肝细胞癌组织（50.0%）,差异有显著性（P＜0.05）。HNF-1α和HNF-4α 蛋白的表达与肿瘤分化程度有关（P＜0.05）。在肝细胞癌组织中HNF-1α与HNF-4α mRNA表达水平呈负相关关系。 结论 HNF-1α和HNF-4α在肝细胞癌中表达下调,这可能与肝癌的发生、发展相关。     

TPD52基因与原发性肝细胞癌患者预后相关性分析

目的探索TPD52基因在原发性肝细胞癌中的表达及与患者预后的关系。方法荧光定量RT-PCR检测33对肝癌组织及其对应的癌旁组织中TPD52 mRNA表达水平;免疫组化检测173例肝癌石蜡标本中TPD52蛋白表达情况。结果肝癌组织中TPD52 mRNA表达水平较癌旁组织低(P0.001);肝癌组织TPD52蛋白表达水平低于癌旁组织,且其表达水平与TNM分期存在负相关(P=0.023);TPD52高表达的患者较低表达的患者预后好(P=0.003,P=0.013),且能明显延长肿瘤小于5 cm、肿瘤单发或TNM分期为Ⅲ期的患者的总生存期(P=0.015,P=0.025,P=0.018)和无病进展生存期(P=0.023,P=0.047,P=0.034)。结论 TPD52低表达与原发性肝癌的发生发展有高度相关性,是肝癌的独立预后因素。     

MKK-4、MMP-9基因的表达与原发性肝癌肿瘤侵袭转移的关系

目的:检测m itogen activated prote in k inase k inase 4(MKK-4)、MMP-9基因在原发性肝癌中的表达,探讨原发性肝癌MKK-4与MMP-9 mRNA表达水平间的相互关系,及两者对原发性肝癌侵袭转移的影响。方法:应用逆转录-聚合酶链反应(RT-PCR)检测34例原发性肝癌癌组织和相应癌旁组织及12例正常肝组织中MKK-4 mRNA、MMP-9 mRNA的表达。结果:癌旁与正常肝组织的MMP-9 mRNA表达差异无显著统计学意义(P>0.05),癌中的MMP-9 mRNA表达增高,且转移癌与未转移癌组间有差异(P<0.05),转移癌组、未转移癌组分别与正常组及癌旁组织比较有差异(P<0.01);MKK-4 mRNA在正常及癌旁组织中表达无差异(P>0.05),转移癌组、未转移癌组分别与正常组及癌旁组比较有差异(P<0.01),转移癌中较未转移癌中的表达低(P<0.05);MKK-4或MMP-9的mRNA表达均与肿瘤的大小、分化无关(P>0.05);MKK-4 mRNA与MMP-9 mRNA的表达有一定的相关性(r=-0.925,P<0.01)。结论:提示MKK-4 mRNA与MMP-9 mRNA的表达改变与原发性肝癌侵袭转移的发生发展有一定的关系。     

Hedgehog通路下游转录因子Gli1在肝细胞癌中的表达及其临床意义

目的:探讨hedgehog信号通路下游转录因子Gli1在肝细胞癌中的表达及临床意义。方法:收集63例肝细胞癌组织及相对应的癌旁组织构建成组织芯片,应用免疫组化、RT-PCR法检测Gli1在肝细胞癌及癌旁组织中的表达情况。结果:Gli1蛋白在肝癌组织与癌旁组织中表达具有显著性差异;Gli1mRNA含量在肝癌组织及癌旁组织中具有显著性差异;肝细胞癌中Gli1蛋白表达和mRNA含量与病理分级、门脉癌栓有无、淋巴结转移与否、原发肿瘤分级、TNM分期密切相关。结论:Gli1的上调参与了肝细胞癌的发生发展,与肿瘤转移侵润有关。Gli1可能成为肝细胞癌重要生物学标志和生物治疗靶点。     

肝细胞癌中EMS1蛋白的表达

目的 探讨EMS1蛋白表达与肝癌发生、发展及临床病理特征的关系.方法 采用免疫组化SP法,对78例肝癌组织、其中68例附癌旁肝组织和8例正常肝组织中EMS1蛋白表达进行检测.结果 EMS1蛋白表达定位于细胞的胞质,着色呈黄色至棕黄色.肝癌中阳性表达率89.69%,高于癌旁及正常肝组织30.26%(P＜0.001);无肝硬化肝癌阳性表达率73.91%,低于伴有肝硬化95.56% (P＜0.05);伴静脉内癌栓组94.12%与无静脉内癌栓组70.59%差异有显著性 (P＜0.05);随着组织分级的增高(分化程度的降低),阳性表达率明显增高,其差异有显著性 (P＜0.05);而与性别、年龄、肿瘤大小无关.结论 EMS1蛋白高表达与肝癌发生、肝硬化、静脉内癌栓及组织学分级相关.     

NY-ESO-1和LAGE-1癌症-睾丸抗原在肝细胞癌的表达

目的　检测NY- ESO -1和LAGE- 1癌症睾丸抗原在肝细胞癌中的表达,探讨其作为肝细胞癌免疫治疗靶标的可行性及其与肝癌生物学行为的关系。方法　逆转录聚合酶链反应(RT -PCR)和免疫组织化学EnVision二步法检测30例肝细胞癌新鲜标本NY ESO -1和LAGE -1的表达;另将191例肝癌石蜡组织制成芯片观察NY- ESO- 1蛋白在肝癌中的分布和表达。结果NY- ESO -1和LAGE -1基因mRNA在肝癌中的阳性表达率分别是33. 3% (10 /30)和16. 7% (5 /30),至少表达1种基因mRNA者为36 7% (11 /30);NY- ESO -1蛋白主要分布在肝癌细胞胞质,有效标本中NY ESO 1表达13 8% (24 /174),小肝癌、中晚期肝癌、发生转移肝癌中的阳性表达率逐渐升高,分别为6 8% (3 /44)、16 2% (21 /130)、23 1% (12 /52),不发生转移的肝癌仅为9 8% ( 12 /122 ),其中转移组与无转移组之间比较差异有统计学意义(P<0 .05),全部病例癌组织与癌旁组织比较差异有统计学意义(P<0 .01)。NY- ESO-1蛋白的阳性表达与肿瘤大小无关,所有癌旁肝组织均未见NY -ESO -1和LAGE -1的mRNA和蛋白的表达。结论　NY -ESO- 1 /LAGE- 1在肝细胞癌组织中的特异性表达提示其可作为肝细胞癌特异性免疫治疗潜在的靶标;NY -ESO -1在肝癌早期出现,随病情进展表达率逐步增高,转移患者最高,提示N     

H2-Calponin蛋白在4种癌组织的表达及临床意义

 目的 探索肿瘤相关抗原H2-Calponin mRNA及其蛋白在肺癌、胃癌、鼻咽癌和肝癌中表达及其临床意义。方法 用免疫组织化学和原位RT-PCR检测癌组织中H2-Calponin蛋白及其mRNA的表达分布和定位情况。结果 H2-Calponin蛋白主要分布在胞质;在肺癌、胃癌、鼻咽癌和肝癌中阳性率分别为30.0%、27.5%、45.0%和51.8%,肝癌组织阳性率最高。H2-Calponin mRNA 定位主要在细胞核周;H2-Calponin mRNA在肺癌、胃癌、鼻咽癌和肝癌中表达阳性率分别为17.5%、10.0%、20.0%和50.0%,在肝癌组织阳性率最高(P<0.05)。H2-Calponin在伴有转移的肝癌组织中的阳性表达率高于无转移的肝癌组织中的阳性表达率(P<0. 05),但该蛋白在4种不同癌组织中的表达与患者的性别、年龄、及癌组织的病理分级等因素无关。结论 肝癌组织高表达H2-Calponin蛋白和mRNA,可能具有肝癌特异性且与肝癌是否伴有转移相关。     

癌症高表达蛋白在肝细胞肝癌中的表达及意义

目的:研究癌症高表达蛋白(Hec1)在原发性肝细胞肝癌(HCC)中的表达和意义。方法:采用免疫组化SP法检测27例正常肝组织、30例癌旁组织及62例HCC组织中Hec1的表达;用Westernblot检测上述组织中Hec1蛋白的表达。结果:在正常肝、癌旁组织及HCC组织中,Hec1蛋白的阳性表达率分别为0%、20.8%、62.9%,各组织中Hec1蛋白的表达差异均有统计学意义(P<0.05)。Hec1蛋白的表达与HCC的Edmondson分级、侵袭转移显著相关。Westernblot结果表明,在正常肝、癌旁组织及HCC组织中,Hec1蛋白的平均表达量的分别为0、0.12±0.03、0.89±0.16,差异有统计学意义(P<0.05)。结论:Hec1蛋白的过度表达可能在HCC的发生和发展中起重要作用。     

Nuclear translocation of survivin in hepatocellular carcinoma: a key to cancer cell growth?

Survivin is a recently described anti-apoptosis protein and regulator of cell division. Its expression and localization in hepatocellular carcinoma (HCC) and in normal liver tissue has not been fully elucidated. We examined the expression of survivin, Fas, proliferating cell nuclear antigen (PCNA), and apoptosis in 47 specimens of hepatocellular carcinoma (HCC) and surrounding nonmalignant hepatic tissues. To further determine the relationship between survivin expression and cell proliferation and apoptosis, we performed double immunostaining for survivin and PCNA TUNEL staining in the same HCC specimens. Positive immunostaining for survivin was present in 35 of 47 (74%) HCCs. Twenty-two of 35 survivin-positive HCCs (63%) showed punctate nuclear staining in HCC cells, and the remaining 13 showed predominant cytoplasmic staining. In contrast, nonmalignant hepatocytes showed only cytoplasmic staining. HCC cells had significantly higher PCNA-labeling and apoptotic indices compared with the case of nonmalignant hepatic tissue (P<0.001). Furthermore, nucleus-positive HCC specimens for survivin showed the highest PCNA labeling index. The nuclear localization of survivin in HCC cells correlated with tumor cell de-differentiation with the exception of the HepG2 cell line. Survivin expression was inversely associated with apoptosis and was strongly associated with Fas expression (P=0.01). All 4 HCC cell lines examined showed survivin expression and punctate nuclear localization. Our results indicate that survivin is localized to the cytoplasm in quiescent nonmalignant liver cells to suppress apoptosis and translocates into the nucleus in HCC cells. In conclusion, translocation of survivin from the cytoplasm to the nucleus may constitute an important regulatory mechanism for cell proliferation and differentiation in HCC.     

MDM2和P53蛋白表达与原发性肝细胞肝癌发生的关系

目的:研究MDM2和突变型P53蛋白表达与原发性肝细胞肝癌(HCC)发生的关系。方法: 用免疫组织化学SP法,检测55例HCC癌组织、23例癌旁组织、10例正常肝组织MDM2和突变型P53蛋白表达情况。结果: 55例HCC中MDM2蛋白阳性表达17例(30.9%),突变型P53蛋白阳性表达23例(41.8%),二者均阳性表达11例(20%),MDM2和突变型P53蛋白表达有相关性(r=0.310, P＜0.05)。23例癌旁组织MDM2蛋白阳性1例,突变型P53蛋白阳性表达2例,肝癌组织和癌旁组织MDM2和突变型P53蛋白表达有差异(P＜0.05)。10例正常肝组织无MDM2和突变型P53蛋白表达。结论: 肝细胞癌组织有MDM2和突变型P53过量表达,MDM2蛋白和/或 p53 基因突变使 p53 基因功能失活在HCC发生中可能有重要作用。     

RB tumor suppressor gene expression in hepatocellular carcinomas from patients infected with the hepatitis B virus

Hepatitis B virus (HBV) infection is closely associated with the development of hepatocellular carcinoma (HCC), but definite mechanisms by which it could play an etiologic role have not yet been identified. Modifications of the function of the RB tumor suppressor gene, which regulates the cell cycle, could provide such a mechanism. In the present study, the expression of the protein product of RB, pRB, was evaluated by immunohistochemical staining in HCC tissues from 25 patients from China and the United States, adjacent nontumorous liver from 19 of those patients, five human HCC cell lines, three human hepatoblastoma cell lines, and five specimens of normal human liver. Representative samples were also evaluated by western blot. Altered expression of RB was detected in eight HCC tissues (pRB undetectable in five HCCs and detected in <1% of nuclei of HCC cells in three others); all eight had detectable hepatitis B surface or core antigen in the adjacent nontumorous liver, indicating active HBV infection. pRB was detected in 10--95% of nuclei (normal expression) in the remaining 17 HCCs, and in many nuclei in all 19 nontumorous livers, and in the 5 normal livers. No pRB staining was detected in the nuclei of three HCC cell lines, but pRB was detected in > 90% of nuclei of the other HCC and hepatoblastoma cell lines. The relationship of pRB expression to mutations of the p53 tumor suppressor gene was also examined. The absence of detectable nuclear pRB by immunohistochemical staining was associated with the presence of presumed mutant p53 detected by immunohistochemical staining in four out of five HCC cases. In addition, all three HCC cell lines lacking detectable pRB also had a p53 mutation or a p53 deletion. HCCs with altered pRB expression included more grade III and IV tumors (8/8,100%) than did HCCs with normal pRB expression (7/17, 41%) (P < 0.02), suggesting that abnormal pRB expression may be associated with more advanced histologic grades of HCC. These data indicate that interference with the normal function of the tumor suppressor gene RB or its product pRB, often with concomitant p53 mutation, may be one of several mechanisms that contribute to the development or progression of HCC in humans infected with HBV. © 1994Wiley-Liss, Inc.     

人肝癌组织中CD34和血管内皮生长因子的表达及微血管密度的 …

目的 探讨ＣＤ３４和血管内皮生长因子（ＶＥＧＦ）在人肝癌组织中的表达及微血管密度（ＭＶＤ）的病理意义。方法 对３０例人肝细胞肝癌９ＨＣＣ）及相应癌旁组织,１０例肝硬化,５例轻度慢性肝炎和４例正常肝组织,进行了ＣＤ３４、ＶＥＧＦ免疫组织化学ＳＰ法检测,对ＣＤ３４阳性血管进行ＭＶＤ计数,对ＶＥＧＦ进行半定量计数,并结合肝癌的病理特征进行分析。结果 ＨＣＣ组织中ＣＤ３４呈广泛,窦隙状阳性表达,而在正常及     

High-throughput tissue microarray analysis of c-myc activation in chronic liver diseases and hepatocellular carcinoma

Amplification of 8q23-qter is common in human hepatocellular carcinoma (HCC). c-myc, an oncogene located on 8q24, may be important in hepatocarcinogenesis. The present study aimed to evaluate c-myc activation in hepatocarcinogenesis and its clinicopathological significance. High-throughput analysis of c-myc gene amplification and expression using dual-color fluorescence in situ hybridization and immunohistochemistry was performed on tissue microarrays consisting of 458 liver samples comprising HCCs, nontumorous livers and normal livers. HCCs demonstrated frequent c-myc amplification (30% when corrected for chromosome 8 aneusomy). In contrast, the noncancerous livers, which were mostly chronic hepatitis and cirrhosis, exhibited no c-myc amplification. Despite c-myc amplification, the HCCs exhibited less nuclear c-myc expression than the livers with chronic liver diseases and normal livers (P <0.001 and 0.004, respectively). The HCCs also had less cytoplasmic c-myc staining than the livers with chronic liver diseases (P = 0.002). Despite their absence of c-myc amplification, however, the livers with chronic disease had significantly increased expression of both nuclear and cytoplasmic c-myc protein compared with normal livers (P = 0.015 and 0.009, respectively). Clinicopathologically, the reduction in nuclear c-myc was more marked in HCCs with venous permeation and absence of tumor encapsulation (P = 0.013 and 0.021, respectively), whereas HCCs with cytoplasmic c-myc were positively associated with larger tumor size (P = 0.027). There was no significant association between c-myc amplification and protein expression levels in HCC. Our results suggest that overexpression of c-myc in chronic liver diseases may play an important role in the predisposition to hepatocarcinogenesis. Although c-myc was amplified in HCC, there appears to be a tight regulation by independent pathways of c-myc activation in hepatocarcinogenesis.     

Value of glypican 3 immunostaining in the diagnosis of hepatocellular carcinoma on needle biopsy

The diagnostic value of glypican 3 (GPC3) immunostaining on needle biopsy specimens has not been well assessed. In this study, 120 liver needle biopsy specimens, including 46 from cirrhotic livers and 74 hepatocellular carcinomas (HCCs), were immunohistochemically examined for expression of GPC3. The results showed strong cytoplasmic and membranous staining in 36 HCCs (49%), among which 20 cases (56%) showed diffuse immunoreactivity. None of the 46 cirrhotic livers exhibited positive GPC3 immunostaining. The nonneoplastic liver tissues (cirrhotic or noncirrhotic) that were present in the majority of the HCC cases were also completely negative for GPC3 expression. These data demonstrate that GPC3 is a reliable immunohistochemical marker for the diagnosis of HCC on needle biopsy specimens when positive. However, the detection rate in our series seems lower than that reported in studies using resection specimens as the study materials. Our findings emphasize that GPC3 immunoreactivity can be focal and that negative staining should not be viewed as evidence to exclude the diagnosis of HCC in challenging needle biopsy specimens.     

黄曲霉毒素高污染区肝细胞癌p（53）蛋白与乙肝病毒感染的相关性研究

应用免疫组化ＳＰ法，检查桂西南黄曲霉毒素（ＡＦＢ１）高污染区１０９例肝细胞癌（ＨＣＣ）组织中突变型ｐ５３蛋白的表达，并与相应病例中乙型肝炎病毒表面抗原（ＨＢｓＡｇ）的表达进行相关性研究。结果发现：ＨＣＣ中ｐ５３蛋白阳性率高达６８．８％（７５／１０９），ｐ５３蛋白表达与病人性别、肿瘤大小及有无乙型肝炎病毒（ＨＢＶ）感染无关（Ｐ＞０．０５）。说明在ＡＦＢ１高污染区，ｐ５３基因突变是ＨＣＣ中非常普遍的现象。在ＨＣＣ病例中，ＨＢｓＡｇ阳性率为８４．２％（８５／１０１），其中同一病例ｐ５３蛋白及ＨＢｓＡｇ均为阳性者占５７．０％（５８／１０１），而ＨＢｓＡｇ阳性、突变型ｐ５３蛋白阴性者为２６．０％（２７／１０１）。表明ＨＢＶ仍然是肝细胞癌发生的主要原因，但ＨＢＶ的致癌机制具有多样性，并非均引起ｐ５３基因的改变。     

Overexpression of X-linked inhibitor of apoptosis in human hepatocellular carcinoma

The X-linked inhibitor of apoptosis (XIAP) is a member of a novel family of inhibitors of apoptosis. Since suppression of apoptosis is fundamentally important for carcinogenesis and tumor growth, we investigated the expression and function of XIAP in human hepatocellular carcinomas (HCCs). XIAP was expressed constitutively in HCC cell lines. Fourteen out of 20 (70%) HCC tissues demonstrated moderate or strong cytoplasmic staining for XIAP, whereas non-tumor parts showed negative or weak staining for XIAP by immunohistochemistry. In addition, XIAP expression was inversely correlated with apoptosis, but not with proliferation in HCC tissues. These results indicated that XIAP is a principal inhibitor of apoptosis overexpressed in human HCCs and that XIAP may be a potential target for gene therapy of human HCCs.     

Glypican-3 as a potential differential diagnosis marker for hepatocellular carcinoma: a tissue microarray-based study

The differential diagnosis between hepatocellular carcinoma (HCC) and benign hepatic lesions is still difficult and new biochemical markers for HCC are required. The aim of this study was to assess the differential diagnostic value of glypican-3 (GPC3) immunostaining in HCC patients. 147 cases of surgically excised HCC tissues, 94 cases from needle biopsies, and tissue microarrays were used for this study. The tissue microarrays contained 449 specimens including: 115 HCC, 25 intrahepatic cholangiocellular carcinoma, 29 lung adenocarcinoma, 23 squamous cell lung carcinoma, 53 ovary adenocarcinoma, 44 renal cell carcinoma, 30 prostate acinar adenocarcinoma, 42 breast carcinoma, 41 gastric carcinoma and 47 colorectal carcinoma. The immunolocalization of GPC3 was measured using immunohistochemical staining. Among 147 surgically excised HCC samples, 87.1% (128/147) were GPC3 positive. No GPC3 expression, however, was observed in paracarcinomatous and cirrhotic tissues. In needle biopsy tissues, GPC3 was positively expressed in 81.9% (77/94). Among tissue microassays, HCCs showed positive GPC3 expression in 55.7% (64/115), while 9.6% (5/52) of lung carcinoma and 5.7% (3/53) of ovary adenocarcinoma also were positively stained. The other tumor types showed negative GPC3 expression. In conclusion, our results show that GPC3 is specifically overexpressed in HCC tissue and may be regarded as a potential marker for differential diagnostic hepatocellular carcinoma.