Prognostic significance of co‐expression of nm23 and p57 protein in hepatocellular carcinoma

Aim: To investigate the unbalance of proliferation and apoptosis and the functions of cell‐cycle proteins and apoptotic factor in metastasis of hepatocellular carcinoma (HCC) and their effect in prognosis. Methods: Proliferation index and apoptosis index, as well as seven relatively molecular markers, namely p15, p34, p53, p57, p73, survivin and nm23, were evaluated by immunohistochemistry and TUNEL in HCC tissues and compared to adjacent non‐cancerous tissues and normal liver tissues. Furthermore, the prognostic significance by follow‐up and mutual relationships for each clinicopathologic factor and molecular marker were analysed. Results: The dysregulation between proliferation and apoptosis and the abnormal expression of seven molecular markers were observed in HCC tissues. The unbalance of proliferation and apoptosis and abnormal expressions of p15, p34, p57 and nm23 were correlated with TNM stage and extrahepatic metastasis. In particular, the abnormal co‐expression of nm23/p57 correlated with advanced TNM stage and bigger tumor size and was an independent prognostic factor of HCC. Conclusion: The unbalance of proliferation and apoptosis and abnormal expression of cell‐cycle proteins promote metastasis of HCC. Moreover, the abnormal co‐expression of nm23/p57 may be a useful molecular marker for metastasis and unfavourable prognosis for HCC.     

p16INK4A gene alterations are not a prognostic indicator for survival in patients with hepatocellular carcinoma undergoing curative hepatectomy

BACKGROUND AND AIM: Hepatocellular carcinoma (HCC) is a common malignancy worldwide that is highly associated with chronic hepatitis B or C infection and cirrhosis. The tumor suppressor gene p16INK4A is an important component of the cell cycle and inactivation of the gene has been found in a variety of human cancers. The present study was performed to determine genetic and epigenetic alterations in the p16INK4A tumor suppressor gene and the effect of these on HCC progression. METHODS: The status of p16INK4A was evaluated in 117 HCC tumoral nodules and 110 corresponding peritumoral tissues by loss of heterozigosity (LOH) at the 9p21-22 region, homozygous deletions, single-strand conformation polymorphism-polymerase chain reaction (PCR) mutational analysis and methylation specific PCR. RESULTS: The most frequent inactivation mechanism was hypermethylation of the promoter region, which was found in 63.2% of the tumor samples and in 28.2% of the peritumoral samples. Loss of heterozygosity at the 9p21 region was detected in 27.3% and 10% of tumor and peritumoral tissues, respectively. Homozygous deletions and mutations were less common events in hepatocarcinogenesis. The authors found 5.9% of the tumor cases with exon 2 homozygous deletions and 8.6% with mutations. Two polymorphisms were detected, one at codon 148 (GCG --> ACG, Ala --> Thr) in three cases and the other in exon 3 at 540 bp (34.2% of the samples). No association was found between inactivation of p16INK4A and clinicopathological characteristics or prognosis. CONCLUSION: p16INK4A is altered frequently and early in HCC, being the predominant mechanism of inactivation promoter hypermethylation. The present results suggest that the p16INK4A gene plays an important role in the pathogenesis of HCC.     

The role of lncRNAs in hepatocellular carcinoma: opportunities as novel targets for pharmacological intervention

Long non-coding RNA (lncRNA) is commonly defined as an RNA with a length of greater than 200 nucleotides, frequently up to 100 kb. Numerous studies have shown that dysregulation of lncRNAs may directly relate to a number of human diseases, particularly in oncology where lncRNAs appear to play an important role. LncRNAs may also play a potentially novel and critical role in the development and progression of hepatocellular carcinoma (HCC). This article discusses lncRNAs as a new possibility for diagnostic and therapeutic approaches for HCC. The authors introduce the relationship between some lncRNAs and HCC, including carcinogenesis, development, metastasis and prognosis. In addition, the authors suggest that the discovery of lncRNAs may encourage the discovery and development of new therapeutic modalities for HCC and that their regulation may be a promising potential treatment for HCC. Clinical studies are required to determine the therapeutic effect of regulating lncRNA in humans with HCC.     

Apoptosis and proliferation of human hepatocellular carcinoma

Abstract: To investigate the contribution of apoptosis, a major mechanism of cell death, in the growth of hepatocellular carcinoma, we analyzed both apoptosis and cell proliferation in human hepatocellular carcinoma. We used the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling method and proliferative cell nuclear antigen staining, respectively. Among 21 hepatocellular carcinoma specimens examined, four were well, ten were moderately, and seven were poorly differentiated hepatocellular carcinoma. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling-positive cells in hepatocellular carcinoma were scattered individually or were sometimes clustered in the tumors. The terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling indices were 0.35±0.09, 0.81±0.29, and 1.9±0.94 in well, moderately, and poorly differentiated hepatocellular carcinoma, respectively. The proliferative cell nuclear antigen labeling indices were 6.6±0.9, 13.1±3.5, and 26.7±6.3 in hepatocellular carcinoma in the same respective order of differentiation. The differences in both terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling indices and proliferative cell nuclear antigen labeling indices (p>0.05) were significant between well, moderately and poorly differentiated hepatocellular carcinoma. There was a positive correlation between the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling and proliferative cell nuclear antigen labeling indices in hepatocellular carcinoma (r=0.84, p>0.001). This study showed that the proliferation rate and the incidence of apoptosis increased as the differentiation grade of hepatocellular carcinoma was lowered, suggesting a rapid turnover of cancer cells in the lower differentiation grades. Apoptosis may thus play an important role in the growth of hepatocellular carcinoma.     

Rab23 is a potential biological target for treating hepatocellular carcinoma

AIMTo elucidate the role of Rab23 in hepatocellular carcinoma(HCC)by assessing the expression of Rab23 in HCC tissue and in HCC cell lines.METHODSPrimary tumors(n = 100)were stained with Rab23 antibodies using immunohistochemistry and in situ hybridization in tissue microarrays.Relationships between gene expression and pathology parameters were analysed.The biological significance of Rab23 in Hep-3B cells was examined by knocking down Rab23 gene expression.We designed a pair of doublestranded RNAs against human rab23 and transfected siRNA into Hep-3B cells.Rab23 expression in these cells was examined using RT-PCR and Western blots.We investigated cell growth by MTT assays and fluorescenceactivated cell sorting.RESULTSHigh cytoplasmic and nuclear expression of Rab23 was found in 38 of 71(53.5%)and in 49 of 68 HCC patients(72%)respectively,which correlated with tumor size.HCC cell lines expressed Rab23.In Hep3B cells,siRNA for Rab23 decreased Rab23 mRNA by 4.5-fold and protein expression by 2-fold.Survival rates at 24 and 48 h for Hep-3B cells transfected with siRNA were lower and about 30% Hep-3B cells were apoptotic.Knocking down rab23 suppressed Hep3B cell growth,suggesting that rab23 could play an important role in Hep3B cell growth.CONCLUSIONRab23 is overexpressed and/or activated in HCC.Rab23 may be both a HCC predictor and a target for treating HCC.     

肝癌组织中p27蛋白表达与肿瘤增殖凋亡活性的研究

研究肝细胞癌组织中p27表达对细胞增殖与凋亡活性的影响.采用Elivision法检测47例肝癌及相应癌旁肝组织中p27及PCNA的表达,TUNEL法检测原位细胞凋亡.结果显示肝癌组织中细胞增殖与凋亡指数明显高于癌旁肝组织.肝癌组织中p27染色指数明显低于癌旁肝组织与正常肝组织.低分化肝癌组织中p27表达和凋亡指数均明显降低.存在肝外转移的肝癌组织中p27表达降低而增殖指数明显增高.进一步研究显示,p27高表达组肝癌组织中凋亡指数明显高于低表达组,而增殖指数却显著降低.提示p27蛋白表达降低与肝癌的发生和进展有着密切关系,并可能是导致肝癌细胞增殖凋亡失衡的因素之一.     

Nm23-H1 expression in intrahepatic or extrahepatic metastases of hepatocellular carcinoma

Abstract: Aims/Background: Decreased expression of nm23, a putative metastasis suppressor gene, has been reported to be related to either intrahepatic metastasis or a poor prognosis in hepatocellular carcinoma (HCC). The aim of this study was to elucidate the true role of nm23-H1 expression in both intrahepatic and distant metastases of HCC. Methods: Thirteen patients with single-nodule HCC, seven patients with HCC having satellite nodules and seven patients with HCCs having extrahepatic metastases were included in this study. The expression of nm23-H1 protein was immunohistochemically examined in both primary and metastatic nodules. Results: Ten of 13 single-nodule HCCs were found to overexpress nm23-H1 protein. All main tumors, having satellite nodules, were found to overexpress nm23-H1 protein, except for two HCCs, which only partially expressed nm23-H1 protein. Regarding the nm23-H1 expression in intrahepatic metastases, most nodules overexpressed the protein. The expression of nm23-H1 was found to be low in only one intrahepatic metastasis specimen, while its primary tumor was also found to show a low expression of nm23-H1 protein. Microscopic portal vein invasion was found in three of the five patients studied, and all cancer cells in portal invasion overexpressed nm23-H1 protein. Nm23-H1 protein was expressed in all distant metastatic tumors and the staining intensity of most metastatic nodules was similar to that of the primary tumors. Conclusions: Our study demonstrated that nm23-H1 expression did not always decrease but instead tended to increase at both intrahepatic and extrahepatic metastatic sites. Based on these findings, nm23-H1 expression is not considered to be a reliable indicator of either intrahepatic or distant metastasis in HCC.     

Overexpression of extracellular signal-regulated protein kinase and its correlation with proliferation in human hepatocellular carcinoma.

BACKGROUND: Dysregulation of cell proliferation is one of the most important features of human cancers including hepatocellular carcinoma (HCC). However, the molecular basis underlying the proliferation of HCC has not been fully clarified. Because a previous study reported that overexpression of extracellular signal-regulated protein kinase (ERK), which transduces extracellular growth stimuli to the nuclei, was frequently observed in several human cancers, this study was performed to analyze the expression of ERK in human HCC and its correlation with HCC proliferation. METHODS: Twenty-four paired samples of primary HCCs and corresponding noncancerous liver tissues, and six samples of histologically normal liver tissue were obtained from surgically resected materials. The expression of ERK was examined by immunoblotting and immunohistochemical analysis. Proliferative activity of each HCC was examined by immunohistochemical demonstration of proliferative cell nuclear antigen (PCNA). RESULTS: The ERK1 and ERK2 expression in HCCs was significantly higher than that in noncancerous liver tissues, and the ERK1 expression in noncancerous liver tissues from patients with HCC was higher than that in tissue from normal liver. Immunohistochemical examination revealed enhanced accumulation of ERK1 in the nuclei of HCC cells. Regression analysis revealed a significant correlation between ERK expression and PCNA labeling index in HCC. CONCLUSIONS: Our findings suggest that ERK overexpression contributes to the proliferation of HCC.     

肝细胞癌p53及nm23-H1 mRNA表达的意义

目的探讨ｐ５３,ｎｍ２３Ｈ１与原发性肝细胞癌（ＨＣＣ）发生发展的关系．方法运用原位分子杂交技术对４９例ＨＣＣ中ｐ５３和ｎｍ２３Ｈ１基因ｍＲＮＡ进行检测,并结合临床病理特征进行分析．结果ｐ５３ｍＲＮＡ杂交阳性２３例,占４６９％;ｐ５３ｍＲＮＡ过表达与肿瘤的肝内转移．包膜侵犯及Ｅｄｍｏｎｄｓｏｎ分级相关（Ｐ＜００５）;ｎｍ２３Ｈ１ｍＲＮＡ阳性表达２７例,占５５１％;ｎｍ２３Ｈ１ｍＲＮＡ表达与肿瘤肝内转移及ＴＮＭ分期呈负相关（Ｐ＜００５）;同时发现ｐ５３ｍＲＮＡ过表达和ｎｍ２３Ｈ１ｍＲＮＡ低表达在ＨＣＣ肝内转移中具有协同作用．结论ｐ５３和ｎｍ２３Ｈ１参与ＨＣＣ的发生发展,ｐ５３过表达及ｎｍ２３Ｈ１低表达提示ＨＣＣ肝内转移．     

原发性肝癌中p15、p16基因缺失和STK15基因过表达的研究

目的 探讨肿瘤组织p15、p16基因缺失和STK15基因过表达与原发性肝癌的关系。 方法 术中取原发性肝癌组织和相应癌旁正常组织标本30例,术前均未经化学疗法及放射疗法治疗。提取DNA并应用聚合酶链反应(PCR)技术检测p15第二外显子(p15E2)和p16第二外显子(p16E2)纯台缺失。提取RNA,逆转录合成cDNA,应用PCR技术检测STK15基因的表达。以β-肌动蛋白基因作为内对照。测定癌组织和癌旁正常组织中STK15基因和β-肌动蛋白基因的平均密度值(ADV)。 结果 在癌组织中p15E2缺失率为13.3％(4／30),p16E2缺失率为16.7％(5／30),p15E2和p16E2共缺失率为6.7％(2／30)。在30例癌组织中有19例(63.3％)STK15基因表达高于邻近正常组织。STK15基因ADV／β-肌动蛋白基因ADV比值,癌组织为1.53±0.31,癌旁正常组织为0.91±0.25,t=2.86,P<0.01。 结论 p15E2、p16E2纯合缺失和STK15过表达可能在原发性肝癌的发生发展中发挥一定作用。     

Hypoxia and hepatocellular carcinoma: The therapeutic target for hepatocellular carcinoma

Hypoxia enhances proliferation, angiogenesis, metastasis, chemoresistance, and radioresistance of hepatocellular carcinoma (HCC); suppresses differentiation and apoptosis of HCC; and consequently leads to resistance of transarterial embolization (with or without chemotherapy). Because transarterial embolization contributes to angiogenesis via inducing hypoxia, therapy combined with transarterial embolization and antiangiogenic therapy provides a new strategy for the treatment of HCC. Unfortunately, hypoxia leads to the escape of HCC cells from transarterial embolization and antiangiogenic therapy. Thus combined therapy that induces and targets hypoxia may be of benefit to HCC patients. Because angiogenesis plays an important role in recurrence of HCC after resection, antiangiogenic therapy is beneficial to HCC patients following surgical resection of the tumor.     

血吸虫病患者肝癌细胞p53、p16蛋白表达的相关性研究

目的:探讨血吸虫病合并原发性肝细胞癌(hepatocellular carcinoma,HCC)与p53、p16蛋白的基因之间的相关性及其作用机制。方法:58例HCC患者分为两组:1组(HCC合并血吸虫病组)23例和2组(HCC不合并血吸虫病组)35例。采用免疫组织化学方法检测所有患者的p53、p16蛋白表达。结果:p53蛋白阳性表达率在1组中和2组中分别为73.9%(17/23)和31.4%(11/35),两组比较差异有显著性意义(P<0.01);而p16蛋白阳性率在1组和2组中分别为34.8%(8/23)和28.6%(10/35),两组比较差异无显著性意义(χ2=0.25,P>0.05)。p53蛋白及p16蛋白同时阳性仅7例,阳性率为12.1%(7/28);其中1组为13.0%(3/23),2组为11.4%(4/35),两组比较差异无显著性意义(χ2=0.052,P>0.05)。结论:患有日本血吸虫的肝细胞癌患者的癌组织中有较高的p53基因突变率,说明血吸虫感染对肝细胞癌组织细胞中p53突变蛋白的过量表达有促进作用;同时,血吸虫患者肝细胞癌患者肝组织细胞中p16肿瘤抑制基因-p16基因的丢失普遍。p16蛋白表达缺失,其与p53突变蛋白的过量表达同时存在,两者共同促进肿瘤的增长,可能是HCC恶性增殖的原因之一。     

Novel therapeutic target for cancer stem cells in hepatocellular carcinoma

Cancer is a disease of genetic and epigenetic alterations, which are emphasized as the central mechanisms of tumor progression in the multi-stepwise model. Discovery of rare subpopulations of cancer stem cells (CSCs) has created a new focus in cancer research. The heterogeneity of tumors can be explained with the help of CSCs supported by anti-apoptotic signaling. CSCs mimic normal adult stem cells by demonstrating unique characteristics of self-renewal and pluripotency, and the critical role for tumor growth and resistance to anti-cancer therapy. We found that CD13 was a surface marker for CSCs in human liver cancer cell lines and clinical samples, and that CD13+ CSCs were associated with a hypoxic marker in clinical hepatocellular carcinoma (HCC) sample, suggesting that CD13+ CSCs have the critical role in tumor growth and resistance to anti-cancer therapy in liver cancers. In this review article, we update recent findings regarding the involvement of CSCs, especially in HCC.     

姜黄素对肝癌Hep1细胞增殖及凋亡的影响

目的进一步探讨姜黄素的抗肿瘤作用机制。方法CCK-8法检测不同浓度姜黄素作用不同时间肝癌Hepl细胞的增殖抑制率;流式细胞术检测Hepl细胞凋亡率以及各周期时相变化。结果姜黄素作用后Hepl细胞增殖抑制率明显升高,呈时间剂量效应关系;15μmol／L作用后,C0＋G1期细胞显著增多,而G2＋M期细胞显著减少,细胞凋亡率增高（P均〈0．05）。结论姜黄素可有效抑制Hepl细胞的生长和增殖,且具有明显的量效关系;其机制可能为干扰细胞周期进程,促进细胞凋亡。