Adjacent Normal Hepatic Tissues Research Articles

LINC01023 Promotes the Hepatoblastoma Tumorigenesis via miR-378a-5p/WNT3 Axis.

Hepatoblastoma is the most common type of hepatic tumors occurring in children between 0 and 5years. And the exact pathophysiology of the disease is still mysterious. Accumulating studies on LncRNA have shown its pivotal role in the development and progression of distinct human cancers. However, the role of LINC01023 in hepatoblastoma is unknown. The relative expression of LINC01023, miR-378a-5p, and Wnt3 on hepatoblastoma tissue and cell lines was determined by quantitative polymerase chain reaction (qRT-PCR). The effect of LINC01023 downregulation and upregulation on cell proliferation, colony formation and apoptosis activities in HUH6 and HepG2 Cells was assessed by CKK8, clonogenic and flow cytometry analysis, respectively. Dual luciferase, RNA immunoprecipitation (RIP), and RNA pull-down were performed to confirm the interaction between LINC01023 and miR-378a-5p. Similarly, Dual luciferase assay was performed to confirmed the interaction between Wnt3 and miR-378a-5p. The xenograft tumorgenicity test was performed to elucidate the tumorgenicity potential of LINC01023. LINC01023 was significantly upregulated in hepatoblastoma tissue and cell lines rather than in adjacent normal hepatic tissue and QSG7701 cell lines. LINC01023 silencing attenuated cell proliferation, colony formation and increased cell apoptosis. Conversely, LINC01023 upregulation results in significant increase in cell proliferation, and colony formation activities however, a significant reduction in apoptosis activity was reported. Interaction between the LINC01023 and WNT3 was confirmed by dual luciferase assay. Xenograft animal tumorgenicity test confirmed the in-vivo tumorigenesis potential of LINC01203. To the best of our knowledge, this study is the first study demonstrating the role of LINC01023 in hepatoblastoma tumorigenesis through the LINC01023/miR-378a-5p/Wnt3 axis. It could be a potential therapeutic target and a prognostic biomarker in hepatoblastoma.

Abstract 2785: 13C-MRI detects increased lactate flux in HCC following hepatic thermal A\ablation and correlates with PFKFB3 Expression

Abstract Thermal ablation (TA) is an accepted therapy for early HCC and hepatic metastases from colorectal and breast cancers. However, suboptimal TA is a risk factor for early HCC recurrence and has been associated with a reduction in overall survival. This effect has been attributed to hypoxic response in periablative tissue mediated by HIF-1. Malignant transformation leads to increased anaerobic glycolysis even under normoxic conditions. This process is detectable by hyperpolarized 13C MRI (h13C MRI), which enables real-time high-resolution imaging of the conversion from pyruvate to lactate and may be used as a potential predictor for therapeutic response in HCC. We have identified two potential metabolic regulators and therapeutic targets with a direct link to lactate flux which may be monitored with h13C MRI. PFKFB3 has been identified as a crucial metabolic factor in several cancer types, encoding for inducible 6-phosphofructo-2-kinase and enhancing production of F2,6P2, consequently activating PK1 and glycolysis. Expression levels of PFKFB3 have been found to be higher in malignancies than in normal tissues, attributed to loss of glycolytic control and accelerating neoplastic activity. Hexokinase 3 (HK3), a key catalyst in the first committed steps in glucose metabolism, is also a key promoter involved in epithelial-mesenchymal transition. When we exposed an HCC rat cell line, N1S1, to heat stress, simulating the marginal zone of TA therapy, 20%-90% of cells survived at 40°C-50°C and there was no survival by 55°C. 5-8 days following this treatment, cell proliferation and mRNA expression of glycolysis-related genes were analyzed. Cellular proliferation rate was significantly higher in the heat-treated groups. mRNA expression of PFKFB3 was increased after thermal treatment, while HK3 expression was significantly decreased. We then developed an in vivo orthotopic rat model of a solitary N1S1 HCC and used h13C MRI to measure in vivo tumor lactate flux in response to radiofrequency ablation (RFA), a type of thermal ablation, of adjacent normal hepatic tissue, simulating local treatment of a distant lesion. h13C MRI demonstrated significantly increased tumor 13C lactate flux in the RFA arm relative to the control group (change in lactate-to-pyruvate ratio of 0.236±0.278 vs -0.035±0.107, p=0.0465, n=6), while background liver lactate flux was unchanged. We found that this correlated with mRNA expression of PFKFB3 and HK3, which was statistically significantly greater within tumors from the RFA group compared to control tumors (PFKFB3, 1.052±0.091 vs 0.655±0.294, p=0.0004, n=6; HK3, 0.751±0.130 vs 0.501±0.163, p=0.0036, n=6). The PFKFB3 expression patterns were concordant between in vitro and in vivo experiments. Our data shows that PFKFB3 may be an attractive treatment target for HCC, and h13C MRI may be a noninvasive technique well-suited to monitor treatment response. Citation Format: Qianhui Dou, Aaron K. Grant, Cody Callahan, David Mwin, Muneeb Ahmed, Leo L. Tsai. 13C-MRI detects increased lactate flux in HCC following hepatic thermal A\ablation and correlates with PFKFB3 Expression [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2785.

MTA1 Promotes Hepatocellular Carcinoma Progression by Downregulation of DNA-PK-Mediated H1.2T146 Phosphorylation.

Global incidence and mortality associated with hepatocellular carcinoma (HCC) is steadily increasing. Metastasis-associated 1 (MTA1) can induce tumorigenesis and metastatic progression in HCC. However, the mechanistic details of MTA1-mediated regulation of HCC has not been completely defined. Epigenetic histone modification is closely related to tumor development. Histone cluster 1 H1 family member c (H1.2) is important for epigenetic histone modification and chromatin remodeling; however, whether it has a role in HCC tumorigenesis is not known. In the current study, we confirmed that MTA1 promoted HCC cell growth and migration. Our results further show that MTA1 inhibited the phosphorylation of histone cluster 1 H1 family member c (H1.2) at threonine-146 residue (T146) (H1.2T146ph). MTA1 inhibited H1.2T146ph by mediating proteasomal degradation of the DNA protein kinase (DNA-PK). Pharmacological inhibition of proteasomal degradation of DNA-PK or genetic ablation of E3 ligase mouse double minute 2 (MDM2) rescued expression of DNA-PK, and subsequent phosphorylation of H1.2. MTA1's role in HCC was inhibited by ectopic expression of H1.2T146ph in HCC cell lines. Our results showed that H1.2T146ph can bind to MTA1 target genes. Collectively, our study confirms that MTA1 functions as an oncogene and promotes HCC progression. The epigenetic histone modifier H1.2T146ph exerts critical role in the regulation of MTA1-induced tumorigenesis. MTA1 regulates posttranslational activation of H1.2 by regulating the cognate kinase, DNA-PK, via the ubiquitin proteasome system. MTA1 expression was inversely correlated to both DNA-PK and phosphorylated H1.2 in HCC tissue specimens compared to tumor adjacent normal hepatic tissue, revealing that the MTA1/MDM2/DNA-PK/H1.2 is an important therapeutic axis in HCC.

Functional analysis of miR-767-5p during the progression of hepatocellular carcinoma and the clinical relevance of its dysregulation.

Aberrant microRNA (miRNA) expression is a central hallmark of hepatocellular carcinoma (HCC) and identification of the mechanisms underlying the miRNA actions should provide invaluable resource for revealing the molecular basis of different malignant behaviors in HCC. Previous high-throughput analysis has identified miR-767-5p as a unique miRNA signature of HCC, but the biological relevance and corresponding molecular basis of miR-767-5p in HCC is still in its infancy. The current study was, therefore, designed to elucidate whether changes in miR-767-5p expression levels affect HCC pathogenesis, and to further identify the putative targets. miR-767-5p expression was observed to be upregulated by ~ 3.7-fold in surgical HCC specimens as compared to that in adjacent normal hepatic tissues, and this up-regulation trend correlated well to disease progression and predicted a poor prognosis in HCC patients. Functionally, miR-767-5p-overexpressing cells had a significantly higher proliferative, migratory, and invasive potential, and exhibited an enhanced anchorage-dependent clonogenesis and a tumor formation potential in vivo. Mechanistically, PMP22, a core component of integral membrane glycoprotein of peripheral nervous system myelin, was further identified as a direct down-stream target of miR-767-5p in HCC cells. Conversely, stable ectopic expression of PMP22 abrogated the promoting effects of miR-767-5p on HCC aggressive phenotype. Collectively, the available data suggest that as a potent oncomiR, miR-767-5p actions along HCC progression are in part mediated by its function as a posttranscriptional repressor of PMP22 signaling.

A novel miR-0308-3p revealed by miRNA-seq of HBV-positive hepatocellular carcinoma suppresses cell proliferation and promotes G1/S arrest by targeting double CDK6/Cyclin D1 genes

BackgroundPersistent infection with hepatitis B virus (HBV) accounts for the majority of hepatocellular carcinoma (HCC), but the molecular mechanisms underlying liver carcinogenesis are still not completely understood. Increasing evidence demonstrates that microRNAs (miRNAs) play significant functional roles in virus–host interactions. The aim of this study was to explore differentially expressed miRNA profiles and investigate the molecular mechanism of miR-0308-3p in HBV-positive HCC carcinogenesis.MethodsHigh-throughput sequencing was used to detect novel miRNAs in three samples of HBV-positive HCC tissue compared to matched HBV-negative HCC tissue. The Cancer Genome Atlas (TCGA) database was used to mine miRNAs related to HBV-positive HCC. Bioinformatics analyses were conducted to predict the miRNAs’ possible biological and pathway regulatory functions. Quantitative polymerase chain reaction (qPCR) was then applied to evaluate the expression levels of randomly selected miRNAs. CCK-8 was used to measure cell proliferation and cell cycles were analyzed using flow cytometry. A dual luciferase reporter gene assay was used to confirm the downstream targets of miR-0308-3p.ResultsIn total, there were 34 overlapping miRNAs in both our miRNA-seq data and the TCGA database. We found two overlapping miRNAs in both the HBV-positive HCC samples and the TCGA database, and 205 novel pre-miRNA sequences were predicted. miR-522 and miR-523 were markedly overexpressed in HBV-positive HCC and were associated with a significantly poorer long-term prognosis (miR-522, HR 2.19, 95% CI 1.33–3.6, p = 0.0015; miR-523HR 1.5, 95% CI 1–2.44, p = 0.0047). Of note, we found that the novel miR-0308-3p was markedly downregulated in HBV-positive HCC samples and HCC cancer cell lines compared with HBV-negative HCC samples and adjacent normal hepatic tissue. Moreover, elevated expression of miR-0308-3p was found to inhibit proliferation of cancer cells by promoting G1/S cell cycle arrest but did not influence the apoptosis of cancer cells. A dual luciferase reporter activity assay identified that miR-0308-3p acted directly on the target sequence of the CDK6 and Cyclin D1 mRNA 3ʹUTR to suppress CDK6 and Cyclin D1 expression.ConclusionsMiR-0308-3p upregulation dramatically suppressed HCC cell proliferation and induced G1/S cell cycle arrest by directly targeting CDK6/Cyclin D1. These findings reveal a novel molecular mechanism for activation of G1/S arrest in HCC and may prove clinically useful for developing new therapeutic targets.

Overexpression of Cullin7 is associated with hepatocellular carcinoma progression and pathogenesis

BackgroundOverexpression of Cullin7 is associated with some types of malignancies. However, the part of Cullin7 in hepatocellular carcinoma remains unclear. The aim of this study was to investigate the role of Cullin7 in pathogenesis and the progression of hepatocellular carcinoma.MethodsIn the present study, the expression of Cullin7 in hepatocellular carcinoma cell lines and five surgical hepatocellular carcinoma specimens was detected with quantitative reverse transcription PCR and western blotting. In addition, the protein expression of Cullin7 was examined in 162 cases of archived hepatocellular carcinoma using immunohistochemistry.ResultsWe found elevated expression of both mRNA and protein levels of Cullin7 in hepatocellular carcinoma cell lines, and Cullin7 protein was significantly upregulated in hepatocellular carcinoma compared with paired normal hepatic tissues. The immunohistochemistry analysis revealed that overexpression of Cullin7 occurred in 69.1% of hepatocellular carcinoma samples, which was a significantly higher rate than that in adjacent normal hepatic tissue (P < 0.01). Statistical analysis found that overexpression of Cullin7 was significantly associated with lymph node metastasis, tumor thrombus of the portal vein and advanced clinical stage (P < 0.05). Furthermore, by overexpressing Cullin7 in hepatocellular carcinoma HepG2 cells, we revealed that Cullin7 could significantly enhance cell proliferation, growth, migration and invasion. Conversely, knocking down Cullin7 expression with short hairpin RNAi in hepatocellular carcinoma HepG2 cells inhibited cell proliferation, growth, migration and invasion.ConclusionOur studies provide evidence that overexpression of Cullin7 plays an important role in the pathogenesis and progression of hepatocellular carcinoma and may be a valuable marker for hepatocellular carcinoma management.

Overexpression of Rabl3 and Cullin7 is associated with pathogenesis and poor prognosis in hepatocellular carcinoma

The expression of Rabl3 and Cullin7 is relevant to the carcinogenesis of certain cancers. However, the relationship of this expression with hepatocellular carcinoma remains unclear. To study the protein expression of Rabl3 and Cullin7 and to evaluate their role in hepatocarcinogenesis, in 162 cases of hepatocellular carcinoma, we used immunohistochemistry to investigate the expression of Rabl3 and Cullin7 in both the cancer tissues and the normal hepatic tissues around the hepatocellular carcinoma. The results demonstrated that the rates of positive Rabl3 and Cullin7 expression were 80.2% and 69.1%, respectively, in hepatocellular carcinoma tissues. However, the rates of positive Rabl3 and Cullin7 expression were 31.5% and 29.0%, respectively, in adjacent normal hepatic tissues. Rabl3 and Cullin7 were expressed at significantly higher rates in hepatocellular carcinoma compared with adjacent normal hepatic tissues (P<.01). The rates of positive Rabl3 and Cullin7 expression were higher in the hepatocellular carcinoma tissues of patients with lymph node metastasis, tumor thrombi in the portal vein and an advanced clinical stage (P<.05). A positive correlation between the expression of Rabl3 and the expression of Cullin7 (r=0.27, P<.001) was also observed in our hepatocellular carcinoma cohort. Moreover, patients with positive expression for both Rabl3 and Cullin7 had a remarkably shorter survival time compared with patients with negative expression for both proteins (P<.05). Therefore, the expression of the Rabl3 and Cullin7 proteins may play an important role in the pathogenesis and progression of hepatocellular carcinoma and could be used as a prognostic indicator in patients with hepatocellular carcinoma.

Abstract 4726: TonEBP promotes hepatocellular carcinoma and is associated with poor prognosis

Abstract Carcinogenesis is initiated by genetic changes due to aberrant processing of genetic information. Tumor microenvironment, however, should not be overlooked as cancer risk because they influence the development and progression. Indeed, the role of inflammation has received a lot of attention lately. Specifically, hepatocellular carcinoma (HCC) has been considered as an inflammation-associated tumor like colorectal cancer and lung cancer. In addition, high rate of recurrence is the major feature of HCC, and there is a need to identify novel biomarkers for post-operative prognosis of HCC patients Tonicity-responsive enhancer binding protein (TonEBP), also known as nuclear factor of activated T cell 5 (NFAT5), is a key transcriptional cofactor for the expression of pro-inflammatory genes such as TNF-α, IL-1β, and cyclooxygenase 2 (COX-2). Since chronic hepatic inflammation is essential for hepatocellular carcinogenesis, we asked whether TonEBP plays a role in HCC. To investigate the relevance of TonEBP in HCC development, TonEBP expression in hepatic tumors and their adjacent normal tissues were analyzed from of HCC patients. TonEBP expression was higher in tumors than adjacent normal hepatic tissues in 94% of HCC patients (n = 104). In a mouse model of diethylnitrosamine (DEN)-induced HCC, TonEBP expression was also higher in 100% of the tumors. Downregulation of miR-223 was responsible for the elevated TonEBP expression. miR-223 targeted the 3’-UTR of TonEBP gene and suppressed TonEBP translation. Interestingly, univariate analyses revealed that TonEBP expression in normal tissues adjacent to HCC was significantly associated with tumor size, tumor grade, vascular invasion, viral DNA replication, α-fetoprotein, PIVKA-II, recurrence, metastasis, and death. In addition, Kaplan-Meier estimation and multivariate analyses showed that the TonEBP expression was an independent prognostic marker for recurrence and death in HCC patients. In mice, TonEBP haplo-deficiency reduced both DEN- and DEN/high-fat diet (HFD)-induced HCC in association with lower COX-2 transcription and milder hepatic inflammation. Acute hepatic inflammation induced by DEN or lipopolysaccharide was also reduced in hepatocyte- or myeloid-specific TonEBP knockout mice. TonEBP-mediated activation of hepatic COX-2 promoter was dependent on the transcription factor Yin Yang 1 (YY1). TonEBP interacted with YY1 through the Rel-homology domain of TonEBP and spacer domain of YY1. In aggregate, our data demonstrate that TonEBP-mediated hepatic inflammation is a critical regulator of hepatocarcinogenesis. The miR-223-YY1/TonEBP-COX-2 pathway drives hepatic inflammation and commits the stressed hepatocytes to malignant fate. Thus, TonEBP promotes HCC via COX-2 expression and is a strong post-operative prognosticator of poor outcome in HCC patients. Citation Format: Jun Ho Lee, Jae hee Suh, Soo Youn Choi, Hyun Je Kang, Gap Ryol Lee, Whaseon Lee-Kwon, Neung Hwa Park, Hyug Moo Kwon. TonEBP promotes hepatocellular carcinoma and is associated with poor prognosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4726. doi:10.1158/1538-7445.AM2017-4726

The overexpression of Rabl3 is associated with pathogenesis and clinicopathologic variables in hepatocellular carcinoma.

Overexpression of Rabl3 is associated with some malignancies. However, their relationship with hepatocellular carcinoma remains unclear. In this study, the expression of Rabl3 in hepatocellular carcinoma cell lines, and four pairs of matched hepatocellular carcinoma tissues and their adjacent normal hepatic tissues were detected by quantitative reverse transcription polymerase chain reaction and western blot. In addition, the protein expression of Rabl3 was examined in 162 cases of hepatocellular carcinoma by immunohistochemistry. Rabl3 in hepatocellular carcinoma cell lines was elevated at both messenger RNA and protein levels, and the Rabl3 protein was significantly upregulated by upto 3.3-fold in hepatocellular carcinoma compared with the paired normal hepatic tissues. Immunohistochemical analysis revealed that overexpressions of Rabl3 were 80.2% in hepatocellular carcinoma. Rabl3 is expressed at significantly higher rates in hepatocellular carcinoma compared with adjacent normal hepatic tissue (p < 0.01). Statistical analysis suggested the upregulation of Rabl3 was significantly associated with lymph node metastasis, tumor thrombus of the portal vein, and advanced clinical stage (p < 0.05). Furthermore, we found that overexpression of Rabl3 in hepatocellular carcinoma cells could significantly enhance cell proliferation and growth ability. Conversely, silencing Rabl3 by small hairpin RNA interference caused an inhibition of cell proliferation and growth. Our studies suggest that the Rabl3 is a valuable marker of hepatocellular carcinoma progression and that the overexpression of Rabl3 plays an important role in the development and pathogenesis of hepatocellular carcinoma.

Effect of CD16a, the surface receptor of Kupffer cells, on the growth of hepatocellular carcinoma cells.

FcγRIIIa (CD16) is a low-affinity Fc receptor of IgG. As the idio-binding receptor of IgG Fc, it plays an important role in the antibody-dependent cellular cytotoxicity of natural killer cells. The aim of the present study was to investigate the distribution of Kupffer cells (KCs) and the expression of their surface receptor FcγRIIIa in hepatocellular carcinoma. Furthermore, we also aimed to observe the functional mechanism of FcγRIIIa. Immunohistochemical analysis was employed to study KCs and FcγRIIIa. In order to explore the role of FcγRIIIa in the growth of cancer cells, KCs and H22 tumor cells were co-cultured in different serum. The mRNA expression levels of tumor necrosis factor (TNF)-α and FcγRIIIa were analyzed by RT-qPCR; the TNF-α and FcγRIIIa protein expression levels were examined by enzyme-linked immunosorbent assay and western blot analysis, respectively. Our results showed that the number of Kuppfer cells in cancerous tissues (21.6±7.8) was lower than those in para-cancerous (68.8±9.1) tissues and adjacent normal hepatic tissues (62.0±1.9) (P<0.01); this decreased with the reduction in the differentiation degree of cancer (P<0.05). FcγRIIIa-positive cells were similar in morphology to KCs, and their distributive tendency was coincident (P<0.05). The increase in CD16a mRNA levels in the group treated with immune serum was 3.9-, 4.9- and 3.9-fold greater than that in the ordinary serum group at different time points, and CD16a protein expression also markedly increased (P<0.05). However, these effects were inhibited by the addition of anti-IgG Fc serum (P<0.05). The results of the present study suggested that FcγRIIIa resided in KCs, and it contributed to the inhibition of the growth of liver tumor cells.

In vitro and in vivo magnetic resonance imaging with chlorotoxin-conjugated superparamagnetic nanoprobes for targeting hepatocarcinoma.

The present study aimed to assess the in vitro and in vivo magnetic resonance imaging (MRI) features of chlorotoxin (CTX)-conjugated superparamagnetic iron oxide (SPIO) nanoprobes. CTX-conjugated nanoprobes were composed of SPIO coated with polyethylene glycol (PEG) and conjugated with CTX. The nanoprobes were termed SPIO-PEG-CTX. MRI of the SPIO and SPIO-PEG-CTX solutions at a different concentration was performed with a 3.0-T MRI scanner (Philips Achieva 3.0T X Series; Phillips Healthcare, The Netherlands). Rabbit VX2 hepatocarcinoma was established by a traditional laparotomy method (injection of the tumor particles into the liver using a 15G syringe needle) following approval by the institutional animal care and use committee. Contrast-enhanced MRI of VX2 rabbits (n=8) was performed using the same MRI scanner with SPIO‑PEG-CTX solutions as the contrast agent. Data were analyzed with calibration curve and a paired t-test. The SPIO-PEG-CTX nanoparticles were successfully prepared. With increasing concentrations of the solutions, the MRI signal intensity was increased at T1WI, but decreased at T2WI, which were the same as that for SPIO. Rabbit VX2 carcinoma appeared as a low MRI signal at T1WI, and high at T2WI. After injection of the contrast agent, the MRI signal of carcinoma was decreased relative to that before injection at T2WI (1,161±331.5 vs. 1,346±300.5; P=0.004<0.05), while the signal of the adjacent normal hepatic tissues was unchanged (480.6±165.1 vs. 563.4±67.8; P=0.202>0.05). The SPIO-PEG-CTX nanoparticles showed MRI negative enhancement at T2WI and a targeting effect in liver cancer, which provides the theoretical basis for further study of the early diagnosis of hepatocellular carcinoma.

AhR expression is increased in hepatocellular carcinoma

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor implicated in multiple cellular processes and its expression has been shown to play a critical role in tumorigenesis. However, the role of AhR in tumorigenesis of hepatocellular carcinoma remains unclear. In the current study, we investigated the role of AhR in hepatocellular carcinoma tumorigenesis and progression by (a) measuring the expression levels of AhR in liver lesions and (b) assessing the correlation between AhR expression and clinicopathologic parameters. The tissue microarray used in this study contained hepatocellular carcinoma tissues (n = 94), cancer adjacent normal hepatic tissues (n = 5) and normal hepatic tissues (n = 5), which were immunohistochemically assessed for AhR expression. Significantly stronger AhR staining was observed for hepatocellular carcinoma tissues than for cancer adjacent normal hepatic tissues (P = 0.003) and normal hepatic tissues (P = 0.004). In addition, AhR expression was associated with T stage (P = 0.03). The results from this study suggest that an increase in AhR expression is associated with hepatocellular carcinoma progression and may have a potential role in the treatment of hepatocellular carcinoma.

Expression of HIWI in Human Hepatocellular Carcinoma

In this study, the investigation of the expression of HIWI and its protein in hepatocellular carcinoma (HCC) was performed, and the relationships between HIWI expression and the location of HCC metastases were analyzed. Sets of fresh HCC and matched adjacent normal hepatic tissue and paraffin-embedded tissue slides were provided by the hospital hepatology and pathology departments. RT-PCR, Western blot, and immunohistochemistry were performed to detect HIWI mRNA and protein. Correlations between HIWI expression and patient's age, sex, type of tumor, and metastasis location were recorded. HIWI mRNA and protein levels were significantly higher in HCC tissues than in adjacent normal hepatic tissue (P<0.05). Immunohistochemistry showed positive staining for HIWI in cell cytoplasm; however, the number of HIWI-positive cells in HCC tissue (65.2%; 60/92) was significantly higher than in adjacent normal hepatic tissue (27.2%; 25/92) (P<0.05). HIWI expression was not correlated with patients' age, gender, tumors' size, and location but correlated with metastasis involving lymph nodes and other remote organs (P<0.05). HIWI expression is significantly higher in HCC tissue than in adjacent normal hepatic tissue. The results of this study suggest that HIWI may have a crucial role in HCC carcinogenesis and could serve as a potential biomarker or treatment target for HCC.

Up-regulation of Tiam1 and Rac1 Correlates with Poor Prognosis in Hepatocellular Carcinoma

T-cell lymphoma invasion and metastasis 1 (Tiam1) specifically activates Rho-like GTPases (e.g. Rac1) and Tiam1-Rac1 pathway affects the migration and invasion of many tumors, such as nasopharyngeal carcinoma, breast cancer and retinoblastoma. However, no studies have yet comprehensively examined the involvement of Tiam1-Rac1 pathway in hepatocellular carcinoma. In this study, we examined the relationship of the up-regulation of Tiam1 and Rac1 with clinicopathological features in patients with hepatocellular carcinoma. Expression of Tiam1 and Rac1 was assessed in 242 hepatocellular carcinoma tissues and their adjacent normal hepatic tissues by performing immunohistochemistry and was gauged regarding stage, grade and survival. Immunohistochemistry showed that patients with a high clinical stage hepatocellular carcinoma (III-IV) and α-fetoprotein levels had a higher tendency to express Tiam1 and Rac1 on tumor cells than the patients with low pathologic grade hepatocellular carcinoma (I-II) (P = 0.008 and 0.01, respectively) and low α-fetoprotein levels (P = 0.006 and 0.002, respectively). In addition, Tiam1 and Rac1 up-regulation was also significantly associated with vascular invasion status (both P = 0.02), intrahepatic metastasis status (P = 0.009 and 0.01, respectively) and histological differentiation (P = 0.008 and 0.009, respectively) of patients with hepatocellular carcinoma. Moreover, post-operative survival analysis indicated that hepatocellular carcinoma patients with strong Tiam1 (P = 0.01) and Rac1 (P = 0.02) expression had shorter disease-specific survival than those with weak expression. Multivariate analysis also showed that Tiam1 and Rac1 overexpression could be two predictors of poor prognosis (P = 0.02 and 0.03, respectively). The current study demonstrated for the first time that the Tiam1-Rac1 pathway may play a critical role in tumor progression of hepatocellular carcinoma. The expression of Tiam1 and Rac1 can be considered as the two useful indicators for predicting the prognosis of hepatocellular carcinoma.

A study of experimental hepatic amoebiasis in the rabbit.

Progressive acute and chronic hepatic amoebiasis was readily produced in rabbits by subcapsular intrahepatic inoculation of approximately 2 × 104 trophozoites of Entamoeba histolytica (along with their bacterial associates), at 0·05 ml per animal. Infant rabbits, one to four weeks old, were more susceptible to infection than those aged 8·12 weeks. Rabbits with hepatic lesions showed anorexia, reduced activity and significant loss in weight and death.The necrotic lesion appeared about two days post-infection; it was yellowish-pink and later became softer than the adjacent normal hepatic tissue from which it was sharply demarcated. Histological examination of the amoebic lesions showed extensive necrosis with polymorphonuclear and mononuclear cellular infiltration, with the presence of amoebae. In advanced cases, a fibrotic reaction was observed at the edge of the lesions.