牦牛肝蛋白分离纯化及其抗氧化活性研究(英文)

新鲜牦牛肝匀浆、磷酸缓冲液抽提、硫酸铵分级沉淀,并采用响应面法对提取条件进行优化。在最佳提取条件下提取的粗蛋白经DEAE-52离子交换层析和Sephadex G-200凝胶柱洗脱得到7种不同的蛋白。用SDS-聚丙烯酰胺凝胶电泳对各蛋白的分析结果,各蛋白均成单一条带表明纯化效果较好,其分子量分布在97.2 KD-116 KD之间。采用化学发光法对其在体外抗氧化活性研究,主要包括超氧阴离子自由基、羟自由基、1,1-二苯基-2-三硝基苯肼自由基的清除实验,结果表明几种蛋白均具有抗氧化作用,尤其蛋白5和蛋白1作用较为突出,在250 mg/L浓度时,对超氧阴离子的清除率为82.91%和72.92%。上述结果为牦牛肝作为抗氧化保健食品或功能食品的研究和开发提供了依据。

丁二磺酸腺苷蛋氨酸在治疗妊娠肝内胆汁淤积症中的应用

目的:观察丁二磺酸腺苷蛋氨酸对妊娠肝内胆汁淤积症(ICP)患者的治疗作用。方法:治疗组15例采用丁二磺酸腺苷蛋氨酸静脉滴注,对照组使用熊去氧胆酸口服。对比观察治疗前后血清、胆红素、胆汁酸及肝功能的变化。结果:丁二磺酸腺苷蛋氨酸治疗后,多数患者的症状及肝功能均可以明显好转。两者比较后者复发率较高,且有一定的不良反应。结论:丁二磺酸腺苷蛋氨酸作为对ICP的主要用药,效果明显,具有重要的临床实用价值。

复方鸽蛋益肝化瘀丸治疗肝硬化66例

目的观察复方鸽蛋益肝化瘀丸治疗肝硬化的疗效。方法将患者随机分为治疗组和对照组。治疗组用复方鸽蛋益肝化瘀丸,每次5粒,每日3次。对照组服用肝肽乐,每次2片,每日3次,益肝灵每次2片,每日3次,治疗3个月后观察。结果治疗组有效率92.5%,对照组有效率65%,两组疗效相比,差异有显著性,P<0.01结论中药复方鸽蛋益肝化瘀丸可有效治疗早期肝硬化。

复方鸽蛋益肝化瘀丸治疗肝硬化120例临床疗效分析

复方鸽蛋益肝化瘀丸,主要有鸽蛋、柴胡、赤芍、鳖甲、山甲、人参、当归、黄芪、茯苓、苍术、阿胶等药组成。具有疏肝理气,活血化瘀、益气软坚、健脾补肾的作用。在临床上单纯用中药复方鸽蛋益肝化瘀丸治疗肝硬化120例,疗效非常显著,无明显副作用,总有效率达93.3%。

Expression of the Glypican-3 Gene in α-fetoprotein-negative Human Hepatocellular Carcinoma

Objective： To investigate the expression of Glypican-3 gene in （α-fetoprotein （AFP）-negative hepatocellular carcinoma （HCC） and clarify whether Glypican-3 expression is a useful parameter for the diagnosis of hepatocelhllar carcinoma （HCC）, especially for AFP-negative ones. Methods： Forty-one specimens of AFP-negative hepatocellular carcinoma and para-carcimoma tissue were studied for the expression of Glypican-3 by Northern blot. The expression of Glypican-3 protein was detected immunohistochemically with specific polyclonal antibody. Results： Northern blot analysis indicated that the expression of Glypican-3 mRNA was intensively detected in 30 of 41 AFP-negative HCC （73.17%）. In contrast, Glypican-3 mRNA was only weakly detected in 4 of the surrounding non-tumor liver tissues. There was significant difference in the Glypican-3 mRNA expression in large tumors （〉5 cm） （79.31%） and in small tumors （〈5 cm） （41.67%） （P〈0.01）. Gypican-3 mRNA was more frequently overexpressed in poorly differentiated HCC than in well differentiated ones （76.47% vs. 42.86%, P〈0.05）. The Glypican-3 expression was not correlated with age. sex. ttbsAg seropositivity, fibroeapsule, portal venous embolus and intrahepatic metastasis. The overexpression of Glypican-3 protein in HCC was targeted in tumor cells, not in bile duct cells and other interstitial cells. Conclusion： Glypican-3 was specially overexpressed in AFP-negative HCC, and its expression was closely correlated with the tumor size and tumor grade. Glypican-3 gene may play important roles in hepatocareinogenesis, and can be used as a new biochemical marker of HCC, especially for AFP-negative HCC.

Regulation of Bcl-2 Family in TIP30-Induced Apoptosis in Hepatoblastoma Cells

Objective: To investigate the role of TIP30 in the apoptotic signal pathwayin HepG2, and Hep3B and Hu-7 hepatoblastoma cell lines. Methods: In order to confirm whether TIP30conducted Bcl-2 family was involved in apoptosis signal pathway, MTT assay, in situ 3' end labellingof DNA assay and Western blot were carried out to detect the diverse apoptotic function of TIP30and the regulation of Bcl-2 family. Results: TIP30 induced apoptosis as evidenced by morphologicalchanges in hepatoblastoma cells, which was accompanied by up-regulating Bax and Bad proteins andstimulating them from cytoplasm to mitochondria, and down-regulating Bcl-xl, while it had no effecton the level of Bak protein. Conclusion: TIP30 induced apoptosis partly by modulating the proteinlevels of members of Bcl-2 family in hepatoblastoma cells. Elucidating the mechanism by which TIP30induces cell death might establish it as an anticancer factor.

Expression of Lung Resistance Protein （LRP） Gene in Hepatocellular Carcinoma and Its Significance

Objective: To study the multidrug resistance (MDR) mechanism of lung resistance protein (LRP) gene in hepatocellular carcinoma (HCC), and the relations among the expression of the LRP gene and clinicopathologic features, the influence of α-fetoprotein (AFP), and prognosis of patients who received adjuvant chemotherapy after resection of HCC. Methods: The expression of the LRP gene encoding LRP and mRNA LRP was detected in tissues from 54 untreated patients with HCC, adjacent tissues from 24 patients with HCC and archival paraffin-embedded tissues from 12 patients with posthepatitic cirrhosis. The relationship between the LRP gene expression and the change of AFP level was analyzed in the 24 postoperative HCC patients whose AFP was measured after 2 weeks. All of the HCC patients were followed up. Results: The percentage of positive expression of LRP and mRNA LRP in the 3 tissues was 61.1%, 33.3%, 16.7%, and 75.9%, 37.5%, 33.3% respectively. There was significant difference between the untreated HCC tissue and other tissues (P<0.05). No difference existed between the LRP gene expression and clinicopathologic findings, age, sex, and tumor size (P>0.05), but the expression was related to the degree of differentiation of HCC (P<0.05). The effective rate of AFP in the LRP gene positive expression group or in postoperative chemotherapeutic patients was very lower than that in the negative group (P<0.05). Although the mean survival time of postoperative HCC patients in negative LRP gene expression group was longer than that of positive group, there was no difference between them (P<0.05). Conclusion: LRP gene expression is related to MDR of HCC and initiates the intrinsic MDR. Detection of LRP gene expression is of great guiding significance in accessing chemotherapeutic resistance of HCC. As an index to chemotherapy of HCC, detection of LRP expression provides evidence for making individual chemotherapeutic treatment,and reversing MDR in HCC. Although LRP gene expression correlates with the tumor differential degree (P<0.05), it perhaps does not relate with the prognosis of HCC patients.

Overexpression and mutations of tumor suppressor gene p53 in hepatocellular carcinoma

AIMS To examine the prevalance of p53 mutations in hepatocellular carcinoma (HCC) from Chongqing area and the relationship between the p53 mutations and clinicopathological features of HCC,as well as the risk factors. METHODS The overexpression and point mutations of tumor suppressor gene p53 in 38 cases of HCC were detected by a sensitive antigen retrieval fluid (ARF) immunohistochemical method and polymerase chain re- action(PCR)-restriction fragment length polymorphism (RFLP),and single strand conformation polymorphism (SSCP)-silver staining analysis. RESULTS The results showed that 16 of 38 HCCs had positive p53 protein (42.1%),7 HCCs had p53 mutation at 249 (18.4 % ) and 2 HCCS had point muta- tion within exon 7 other than 249. Among 9 cases of HCC with mutations,8 cases demonstrated positive p53 protein,its coincidental rate was 88.9%. The overexpression and mutations of p53 were significantly related to the differentiation and metastasis of HCCs. The frequency of p53 mutations was consistent with high prevalence of HBV and a moderate aflatoxin B1 (AFB1) exposure in our area. CONCLUSIONS The results suggest that AFB1 acts synergistically with HBV in the generation of p53 mutations. Furthermore,dietary exposure to AFB1 may mainly contribute to the tumor specific mutation at codon 249,while HBV may account for other scattered mutations in HCC.

Specifically targeted antiviral therapy for hepatitis C virus

Hepatitis C virus （HCV） infection affects 180 million people worldwide with the predominant prevalence being infection with genotype 1, followed by genotypes 2 and 3. Standard anti-HCV therapy currently aims to enhance natural immune responses to the virus, whereas new therapeutic concepts directly target HCV RNA and viral enzymes or influence host-virus interactions. Novel treatment options now in development are focused on inhibitors of HCV- specific enzymes, NS3 protease and NS5B polymerase. These agents acting in concert represent the concept of specifically targeted antiviral therapy for HCV （STAT-C）. STAT-C is an attractive strategy in which the main goal is to increase the effectiveness of antiviral responses across all genotypes, with shorter treatment duration and better tolerability. However, the emergence of resistant mutations that limit the use of these compounds in monotherapy complicates the regimens. Thus, a predictable scenario for HCV treatment in the future will be combinations of drugs with distinct mechanisms of action. For now, it seems that interferon will remain a fundamental component of any new anti-HCV therapeutic regimens in the near future; therefore, there is pressure to develop forms of interferon that are more effective, less toxic, and more convenient than pegylated interferon.

Hepatitis C Virus non-structural 5A abrogates signal transducer and activator of transcription-1 nuclear translocation induced by IFN-α through dephosphorylation

AIM： To study the effect of Hepatitis C virus nonstructural 5A （HCV NSSA） on IFNα induced signal transducer and activator of transcription-1 （STAT1） phosphorylation and nuclear translocation.METHODS： Expression of STAT1 Tyr701 phosphorylation at different time points was confirmed by Western blot, and the time point when p-STAT1 expressed most, was taken as the IFN induction time for further studies. Immunocytochemistry was used to confirm the successful transient transfection of NS5A expression plasmid. Immunofluorescene was performed to observe if there was any difference in IFNα-induced STAT1 phosphorylation and nuclear translocation between HCV NSSA-expressed and non-HCV NSSA-expressed cells. Western blot was used to compare the phosphorylated STAT1 protein of the cells.RESULTS： Expression of HCV NS5A was found in the cytoplasm of pCNS5A-transfected Huh7 cells, but not in the PRC/ CMV transfected or non-transfected cells, STAT1 Tyr701 phosphorylation was found strongest in 30 min of IFN induction, STAT1 phosphorylation and nuclear import were much less in the presence of HCV NS5A protein in contrast to pRC/CMV-transfected and non-transfected cells under fluorescent microscopy, which was further confirmed by Western blot.CONCLUSION： HCV NSSA expression plasmid is successfully transfected into Huh7 cells and HCV NS5A protein is expressed in the cytoplasm of the cells. IFN-α is able to induce STAT1 phosphrylation and nuclear translocation, and this effect is inhibited by HCV NS5A protein, which might be another possible resistance mechanism to interferon alpha therapy.

Globulin-platelet model predicts minimal fibrosis and cirrhosis in chronic hepatitis B virus infected patients

AIM： To establish a simple model consisting of the rou- tine laboratory variables to predict both minimal fibrosis and cirrhosis in chronic hepatitis B virus （HBV）-infected patients. METHODS： We retrospectively investigated 114 chron- ic HBV-infected patients who underwent liver biopsy in two different hospitals. Thirteen parameters were analyzed by step-wise regression analysis and correla- tion analysis. A new fibrosis index [globulin/platelet （GP） model] was developed, including globulin （GLOB） and platelet count （PLT）. GP model = GLOB （g/mL） x 100/PLT （x 109/L）. We evaluated the receiver operating characteristics analysis used to predict minimal fibrosis and compared six other available models. RESULTS： Thirteen clinical biochemical and hemato- logical variables [sex, age, PLT, alanine aminotransfer- ase, aspartate aminotransferase （AST）, albumin, GLOB, total bilirubin （T.bil）, direct bilirubin （D.bil）, glutamyl-transferase, alkaline phosphatase, HBV DNA and pro- thrombin time （PT）] were analyzed according to three stages of liver fibrosis （F0-F1, F2-F3 and F4）. Bivariate Spearman＇s rank correlation analysis showed that six variables, including age, PLT, T.bil, D.bil, GLOB and PT, were correlated with the three fibrosis stages （FS）. Cor- relation coefficients were 0.23, -0.412, 0.208, 0.220, 0.314 and 0.212; and P value was 0.014, 〈 0.001, 0.026, 0.018, 0.001 and 0.024, respectively. Univariate analysis revealed that only PLT and GLOB were signifi- cantly different in the three FS （PLT： F = 11.772, P 〈 0.001; GLOB： F = 6.612, P = 0.002）. Step-wise multiple regression analysis showed that PLT and GLOB were also independently correlated with FS （R2 = 0.237）. By Spearman＇s rank correlation analysis, GP model was significantly correlated with the three FS （r = 0.466, P 〈 0.001）. The median values in F0-F1, F2-F3 and F4 were 1.461, 1.720 and 2.634. Compared with the six available models （fibrosis index, AST-platelet ratio, FIB-4, fibrosis-cirrhosis index and age-AST model and age-PLT ratio）, GP model showed a highest correlation coefficient. The sensitivity and positive predictive value at a cutoff value 〈 1.68 for predicting minimal fibrosis F0-F1 were 72.4% and 71.2%, respectively. The speci- ficity and negative predictive value at a cutoff value 〈 2.53 for the prediction of cirrhosis were 84.5% and 96.7%. The area under the curve （AUC） of GP model for predicting minimal fibrosis and cirrhosis was 0.762 [95% confidence interval （CI）： 0.676-0.848] and 0.781 （95% CI： 0.638-0.924）. Although the differences were not statistically significant between GP model and the other models （P all 〉 0.05）, the AUC of GP model was the largest among the seven models. CONCLUSION： By establishing a simple model using available laboratory variables, chronic HBV-infected patients with minimal fibrosis and cirrhosis can be di- agnosed accurately, and the clinical application of this model may reduce the need for liver biopsy in HBV- infected patients.

Clinicopathological significance of expression of paxillin, syndecan-1 and EMMPRIN in hepatocellular carcinoma

AIM: To evaluate the relationship of expression of paxillin,syndecan-1 and EMMPRIN proteins with clinicopathological features in hepatocellular carcinoma (HCC).METHODS: Fifty-one patients who underwent HCC resection were recruited in the study. Paxillin, syndecan1 and EMMPRIN proteins in HCC tissues were detected with immunohistochemical staining.RESULTS: Of 51 cases of HCC, 23 (45%) exhibited paxillin protein positive expression. Of 42 cases of adjacent nontumor liver tissues, 24 (57%) exhibited positive expression.Positive paxillin protein expression was associated with low differentiation (r= 0.406, P= 0.004), with the presence of portal vein thrombosis (r = 0.325, P = 0.021), with extra-hepatic metastasis (r = 0.346, P = 0.014). Of 51cases of HCC, 28 (55%) exhibited syndecan-1 protein positive expression. Of 42 cases of adjacent non-tumor liver tissues, 23 (55%) exhibited positive expression.Positive snydecan-1 protein expression was associated with well differentiation (r = 0.491, P = 0.001), with no extra-hepatic metastasis (r = 0.346, P = 0.014). Of 51cases of HCC, 28 (55%) exhibited EMMPRIN protein positive expression. Of 42 cases of adjacent non-tumor liver tissues, 21 (50%) exhibited positive expression.Expression of EMMPRIN protein was not associated with serum AFP level, HBsAg status, presence of microsatellite nodule, tumor size, presence of cirrhosis and necrosis,differentiation, presence of portal vein thrombosis, extrahepatic metastasis, disease-free survival and overall survival (P＞0.05). Expression of paxillin protein was correlated conversely with the expression of syndecan-1protein in HCC (r = -0.366, P = 0.010).CONCLUSION: Expression of paxillin and syndecan-1proteins in HCC may affect its invasive and metastatic ability of the tumor. There may be a converse correlation between the expression of paxillin and syndecan-1 protein in HCC. Expression of EMMPRIN protein may be detected in HCC, but it may play little role in the invasion and metastasis of HCC.

Bystin-like protein is upregulated in hepatocellular carcinoma and required for nucleologenesis in cancer cell proliferation

The bystin-like （BYSL） gene was previously characterized to encode an accessory protein for cell adhesion that participates in early embryo implantation. It is also involved in 40S ribosomal subunit biogenesis and is found to be expressed in rapidly growing embryo and cancer cell lines. In order to explore the role of BYSL in cancer cell proliferation and growth, we used hepatocellular carcinoma （HCC） as a model. Here, we report that BYSL is crucial for HCC cell growth both in vitro and in vivo. Expression levels of BYSL mRNA and protein in human HCC specimens were markedly increased compared with those seen in adjacent non-cancerous tissue. In vitro, inhibition of BYSL by short hairpin RNA decreased HCC cell proliferation, induced apoptosis and partially arrested the cell cycle in the G2/M phase. In vivo, HCC cells treated with BYSL siRNA failed to form tumors in nude mice after subcutaneous implantation. To determine the cellular basis for BYSL RNAi-induced cell growth arrest, BYSL subcellular localization in mitotic and interphase HepG2 cells was examined. BYSL was present at multiple stages during nucleologenesis, including in nucleolus-derived foci （NDF）, perichromosomal regions and the prenucleolar body （PNB） during mitosis. BYSL depletion remarkably suppressed NDF and PNB formation, and disrupted nucleoli assembly after mitosis, resulting in increased apoptosis and reduced tolerance of HCC cells to serum starvation. Taken together, our studies indicate that upregulated BYSL expression plays a role in hepatocarcinogenesis.

Metabolomic changes in fatty liver can be modified by dietary protein and calcium during energy restriction

AIM： To characterise the effect of energy restriction （ER） on liver lipid and primary metabolite profile by using metabolomic approach. We also investigated whether the effect of energy restriction can be further enhanced by modification of dietary protein source and calcium. METHODS： Liver metabolomic profile of lean and obese C57BI/6J mice （n = 10/group） were compared with two groups of weight-reduced mice. ER was performed on control diet and whey protein-based high-calcium diet （whey ＋ Ca）. The metabolomic analyses were performed using the UPLC/MS based lipidomic platform and the HPLC/MS/MS based primary metabolite platform.RESULTS： ER on both diets significantly reduced hepatic lipid accumulation and lipid droplet size, while only whey ＋ Ca diet significantly decreased blood glucose （P 〈 0.001） and serum insulin （P 〈 0.01）. In hepatic lipid species the biggest reduction was in the level of triacylglycerols and cerarnides while the level of cholesterol esters was significantly increased during ER. Interestingly, diacylglycerol to phospholipid ratio, an indicator of relative amount of diabetogenic diglyceride species, was increased in the control ER group, but decreased in the whey ＋ Ca ER group （P 〈 0.001, vs obese）. ER on whey ＋ Ca diet also totally reversed the obesity induced increase in the relative level of lipotoxic cerarnides （P 〈 0.001, vs obese; P 〉 0.05, vs lean）. These changes were accompanied with up-regulated TCA cycle and pentose phosphate pathway rnetabolites. CONCLUSION： ER-induced changes on hepatic rnetabolornic profile can be significantly affected by dietary protein source. The therapeutic potential of whey protein and calcium should be further studied.

Construction of a fusion protein expression vector MK-EGFP and its subcellular localization in different carcinoma cell lines

AIM： To construct an expression plasmid encoding human wild-type midkine （MK） and enhanced green fluorescence protein （EGFP） fusion protein （MK-EGFP）, and to analyze the subcellular localization of MK in different cardnoma cell lines. METHODS： Two kinds of MK coding sequences with or without signal peptide were cloned into plasmid pEGFP-N2, and the recombinant plasmids constructed were introduced into HepG2, MCF7 and DU145 cells, respectively, by transfection. With the help of laser scanning confocal microscopy, the expression and subcellular localization of MK-GFP fusion protein could be detected. RESULTS： Compared with the GFP control, in which fluorescence was detected diffusely over the entire cell body except in the nucleolus, both kinds of fusion protein MK-GFP were localized exclusively to the nucleus and accumulated in the nucleolus in the three kinds of cancer cell lines. CONCLUSION： This study reveals the specific nucleolar translocation independent of signal peptide, which may be involved in the mechanism that MK works. It provides valuable evidence for further study on the functions of MK in nucleus and its possible mechanisms, in which ribosomal RNA transcription and ribosome assembly are involved.

Expression of β-catenin in hepatocellular carcinoma

AIM: The beta-catenin has been recognized as a critical member of the Wnt signaling pathway and plays an important role in the generation/differentiation of many tissues. Inappropriate activation of this pathway has been implicated in carcinogenesis. The mechanism underlying the development as well as its prognosis of hepatocellular carcinoma (HCC) has remained unclear. The purpose of this study is to analyze the expression of beta-catenin in HCC in relation to histological grades and viral hepatitis backgrounds. METHODS: Thirty-two sections were selected at random from autopsy and surgical cases of HCC. Immuohistologically, the location and positivity of beta-catenin expression in HCC was examined. RESULTS: Normal hepatocytes did not express beta-catenin. In 78% of HCC beta-catenin was expressed at the membrane of the cells, with or without cytoplasmic and/or nuclear expression. The tumor cells with well-and moderately-differentiated grades expressed frequently at the membrane and cytoplasm compared with poorly-differentiated type. Nuclear expression of beta-catenin was prone to occur in the tumor cells of poorly-differentiated grade. There were 15% of hepatitis C virus (HCV) backgrounds with nuclear expression. In contrast, there was 38% with nuclear expression in hepatitis B virus (HBV) backgrounds. In nonB-nonC hepatitis, no case expressed nuclear beta-catenin. CONCLUSION: The beta-catenin expression in HCC cells was heterogenous among types of hepatitis viral infection. Wnt signaling pathway might be deeply involved in less-differentiated HCC and HBV background. (C) 2005 The WJG Press and Elsevier Inc. All rights reserved.

Effect of Daxx on cholesterol accumulation in hepatic cells

AIM： To study the effect of Daxx on cholesterol accumulation in hepatic cells. METHODS： Sprague Dawley （SD） rats were fed a normal or high fat diet for 6 wk, and serum lipids and Daxx expression of hepatic tissues were measured by immunoblot assays. HepG2 cells were transfected with the pEGFP-C1/Daxx or pEGFP-C1 plasmid. Cells stably transfected with Daxx were identified by RTPCR analysis. Total cholesterol levels were determined by high performance liquid chromatography. Activated- SREBP and caveolin-1 were assayed by western blotting. RESULTS： Hepatic Daxx protein was higher in normal rats than in high fat diet-fed rats. Noticeable negative correlations were seen between Daxx and LDL-C （γ=-7.56, ρ=0.018）, and between Daxx and TC （γ=-9.07, ρ= 0.01）, respectively. The total cholesterol of HepG2/GFP-Daxx cells was lower than that of control cells or HepG2/GFP cells （9.28±0.19 vs 14.36± 4.45 or 13.94±2.62, both P 〈 0.05）. Furthermore, in HepG2/GFP cells, the expression of activated SREBP was lower than that of control cells, whereas caveolin-1 expression was higher. CONCLUSION： Overexpression of Daxx in HepG2 cells decreased intracellular cholesterol accumulation, which might be associated with inhibition of SREBP activity and an increase in caveolin-1 expression.

Silencing SMYD3 in hepatoma demethylates RIZI promoter induces apoptosis and inhibits cell proliferation and migration

AIM： To investigate the role of SMYD3 in hepatocellular carcinoma （HCC） development and progression and to verify whether its regulation activity was through RIZ1 inactivation. METHODS： Expression of SMYD3 in HCC cell lines and tissues were measured; silencing of SMYD3 by RNA interference （RNAi） was effectuated, hepatoma cell proliferation, migration and apoptosis were tested, with RIZl CpG promoter methylation, and corresponding mRNA expression were investigated. RESULTS： SMYD3 over-expression in HCC was associated with RIZl hypermethylation and mRNA down-expression. Suppression of SMYD3 expression de- methylated RIZl CpG promoter （P 〈 0.01） and increased RIZl mRNA expression （P 〈 0.01）. Consequently, SMYD3 down-expression with RIZl de-methylation strongly inhibited hepatoma cell growth （MTT inhibitory rates： Pgenesil-1-s1 60.95%± 7.97%, Pgenesil-1-s2 72.14% ± 9.68% vs Pgenesil-1-hk 6.89% ± 4.12%, P 〈 0.01） and migration （Pgenesil-1-s1 4.24% ± 1.58%, Pgenesil- 1-s1 4.87% ± 0.73% vs Pgenesil-1 19.03% ± 4.63%, Pgenesil-1-hk 19.95% ±5.21%, P 〈 0.01） and induced apoptosis （FCM subG1 phase Pgenesil-1-s1 19.07% ＋ 1.78%, Pgenesil-1-s2 17.68% ± 2.36% vs Pgenesil-1 0.47% ± 0.12%, Pgenesil-1-hk 1.46% ± 0.28%, P 〈 0.01. TUNEL-positive cells： Pgenesil-1-s1 40.24%± 5.18%, Pgenesil-1-s2 38.48% ± 4.65% vs Pgenesil-1 2.1B% - 1.34%, Pgenesil-1-hk 2.84%± 1.22%, P 〈 0.01） in HepG2 cells. CONCLUSION： These results demonstrate that SMYD3plays a critical role in the carcinogenesis and progression of HCC, The proliferation, migration induction and apoptosis inhibition activities of SMYD3 may be mediated through RIZl CpG promoter hypermethylation.

Removal selectivity of Prometheus:A new extracorporeal liver support device

AIM： To evaluate whether treatment with the Prometheus system significantly affects cytokines, coagulation factors and other plasma proteins. METHODS： We studied nine patients with acute-onchronic liver failure and accompanying renal failure. Prometheus therapy was performed on 2 consecutive days for up to 6 h in all patients. Several biochemical parameters and blood counts were assessed at regular time points during Prometheus treatment. RESULTS： We observed a significant decrease of both protein-bound （e.g. bile acids） and water-soluble （e.g. ammonia） substances after Prometheus therapy. Even though leukocytes increased during treatment （P〈 0.01）, we found no significant changes of C-reactive protein, interleukin-6, and tumor necrosis factor-o plasma levels （all P 〉 0.5）. Further, antithrombin 3, factor II and factor V plasma levels did not decrease during Prometheus therapy （all P 〉0.5）, and the INR remained unchanged （P = 0.4）. Plasma levels of total protein, albumin, and fibrinogen were also not altered during Prometheus treatment （all P 〉 0.5）. Finally, platelet count did not change significantly during therapy （P= 0.6）. CONCLUSION： Despite significant removal of protein- bound and water-soluble substances, Prometheus therapy did not affect the level of cytokines, coagulation factors or other plasma proteins. Thus, the filters and adsorbers used in the system are highly effective and specific for water-soluble substances and toxins bound to the albumin fraction.

Liver expression of steroid hormones and Apolipoprotein D receptors in hepatocellular carcinoma

AIM: To evaluate the tissular expression of Androgen (A), Estrogen (E) and Progesterone (Pg) receptors, and Apolipoprotein D (ApoD), in liver tumors from resected hepatocellular carcinoma (HCC) cases in order to assess their possible relationship to prognosis. METHODS: We performed an immunohistochemical study using tissue microarrays (containing more than 260 cancer specimens, from 31 HCC patients and controls) to determine the presence of specif ic antibodies against AR, ER, PgR and ApoD, correlating their findings with several clinico-pathological and biological variables. The staining results were categorized using a semi-quantitive score based on their intensity, and the percentage of immunostained cells was measured. RESULTS: A total of 21 liver tumors (67.7%) were positive for AR; 16 (51.6%) for ER; 26 (83.9%) for PgR and 12 (38.7%) stained for ApoD. We have found a wide variability in the immunostaining score values for each protein, with a median (range) of 11.5 (11.5-229.5) for AR; 11.1 (8.5-65) for ER; 14.2 (4-61) for PgR; and 37.7 (13.8-81.1) for ApoD. A history of heavy ethanol consumption, correlated positively with AR and PgR and negatively with ER status. HCV chronic infection also correlated positively with AR and PgR status. However, the presence of ApoD immunostaining did not correlate with any of these variables. Tumors with a positive immuno-staining for PgR showed a better prognosis. CONCLUSION: Our results indicate a moderate clinical value of the steroid receptor status in HCC, emphasizing the need to perform further studies in order to evaluate the possible role of new hormonal-based therapies.