中药对肝基因表达的影响

1 单味药及有效成分对肝基因表达的影响 1.1 丹参及其有效成分 姜传仓等采用斑点印迹杂交和Northen印迹杂文技术,发现丹参能升高大鼠肝及培养的人成纤维细胞低密度脂蛋白(LDL)受体mRNA水平,而丹参主要有效成分丹参素的作用较丹参注射液明显,且有量效依赖关系。但降血清总胆固醇(TC)及LDL-胆固醇(LDL-c)作用不明显,一旦停止高脂喂饲,则引起TC及LDL-c水平降低。

人胚肝脂代谢相关基因的表达谱

运用cDNA微阵列结合荧光半定量聚合酶链反应技术 ,首次大规模分析人胚发育过程中肝脏脂代谢相关基因的表达情况 ,比较孕早、晚期人胚肝脂代谢相关基因的表达谱。抽提孕早、晚期胎儿肝脏组织总RNA ,取等量mRNA逆转录标记 33P ,制成cDNA探针后与人类全基因cDNA微阵列芯片杂交 ,用相应软件分析杂交信号扫描结果。有差异表达的基因再用实时荧光半定量PCR方法进一步确认。在进入研究的可信基因中 ,与脂代谢相关的基因共有 135条 ,其中 2 8条有差异表达 (2倍以上 ) ,随胎龄增长 ,脂肪酸分解的相关基因呈现上调趋势 ,脂肪酸和胆固醇合成的相关基因呈现下调趋势 ,差异范围从 0 .4 3～ 8.3倍。检测 15例胚肝组织的脂质含量 ,发现胆固醇和甘油三酯水平随胎龄增长而逐渐下降。人类胚胎肝脏的胆固醇和甘油三酯的合成率及含量可能随着发育成熟呈现下降趋势。

肝组织特异表达人核受体nr5a2转基因小鼠的构建(英文)

人乙肝病毒增强子ⅡB1结合因子(hB1F)系FtzF1(NR5A)亚家族的新成员。经基因重组法将人hb1fcDNA置于小鼠白蛋白增强子/启动子序列下游构建成肝特异重组载体,通过原核显微注射将该载体导入小鼠受精卵原核,经注射且状态良好的卵回输至假孕母鼠输卵管。产下仔鼠经PCR和Southernblotting鉴定,同时RTPCR和Westernblotting分析转基因的表达。阳性Founder鼠与正常C57鼠交配以建立转基因纯系小鼠,F1代以PCR法鉴定。结果共获得4只PCR鉴定转基因阳性Founder鼠,其中一只同时经Southernblotting鉴定为阳性。RTPCR和Westernblotting结果显示,外源基因在转基因小鼠的肝组织成功表达。遗传学分析表明,转基因已整合入小鼠基因组并可稳定遗传。

Survivin基因在肝门部胆管癌组织中的表达及其与nm23和PTEN相关性的研究

目的:探讨Survivin基因在肝门部胆管癌组织中的表达情况以及与nm23和PTEN基因表达的相关性。方法:应用免疫组织化学方法测定肝门部胆管癌及相应癌旁正常组织中Survivin、nm23和PTEN基因的表达,分析Survivin基因表达与肿瘤组织病理分级、淋巴结转移以及nm23和PTEN基因的相关性。结果:44例肝门部胆管癌组织中,Survivin基因阳性表达率63.6%,15例癌旁组织中Sur-vivin基因阳性表达率为6.67%,二者比较差异有统计学意义(P<0.05);Survivin基因表达与病理分级无关(P>0.05),与淋巴结转移无关(P>0.05);Survivin基因表达与PTEN及nm23的表达呈负相关(P<0.05)。结论:Survivin基因的高表达在肝门部胆管癌的发生、进展和转移过程中可能起重要作用;联合检测Survivin与PTEN及nm23基因表达可能有助于临床诊断及判断预后。

大鼠马钱子碱中毒肝细胞Bcl-2和Caspase-3蛋白表达的研究

目的:研究马钱子碱中毒后不同时间段内肝细胞Bcl-2和Caspase-3表达的病理变化。方法:用20只Wistar大鼠为正常对照组,40只Wistar大鼠复制马钱子中毒模型,用免疫组织化学技术对中毒后大鼠肝细胞Bcl-2和Caspase-3的蛋白表达进行检测,并用图像分析对检测结果进行分析。结果:大鼠在马钱子中毒低、中剂量后的1～7d不同时间段内,肝细胞Bcl-2和Caspase-3蛋白阳性表达数均明显高于正常对照组,中毒组与对照组相比,差异有显著性。结论:Bcl-2和Caspase-3参与了马钱子碱中毒的病理生理过程。

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.

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.

Down-regulated Expression of Cytochrome P450 Related Genes Involved in Hepatocellular Carcinoma

Objective： To analyze the differentially expressed genes （DEGs） among hepatocellular carcinoma （HCC）, para-cancerous tissue （PCT） and normal liver tissue （NLT） by using hgilent microarray and explore the target genes related to the development and progression of HCC. Methods： Total RNA of matched HCC, PCT and NLT of HCC patients was isolated using one step Trizol method and qualified using Lab-onchip method, cRNAs were synthesized, fluoresce labeled and purified after total RNAs purified. The RNAs of HCC and NLT, HCC and PCT were hybridized to Agilent microarray （21 074 probes） respectively. The fluorescence intensities were detected by Agilent scanner and quantified by Feature Extraction software. The DEGs which both differentially expressed in HCC vs NLT and HCC vs PCT were selected as candidate genes and confirmed by SYBR Green I stained real time RT-PCR. Results： （1） The total RNAs, reverse transcription products and fluoresce labeled cRNAs were of high quality; （2） There were 420 up-regulated genes and 552 down-regulated genes in 2-fold DEGs, including 23 genes related to cytochrome P450; （3） CYP2C8 was selected as a candidate gene of 10-fold down-regulated genes. The results of real time RTPCR, using β-actin as an internal control, revealed that the 2^-ΔCt values of CYP2C8 in HCC, PCT and NLT were 0.009383, 0.316812 and 0.607182, respectively. Conclusion： （1） The high throughput and effective Agilent oligomicroarray can screen novel therapy target genes by analyzing the DGEs in development and progression of HCC; （2） The development and progression of HCC is a complicated process involving multigenes and multiprocedures; （3） Down-regulated expression of cytochrome P450 related genes maybe involved in the development and progression of HCC due to the increased activity of certain carcinogens and maybe the exact mechanism need further study.

Liver-specific gene expression in mesenchymal stem cells is induced by liver cells

AIM： The origin of putative liver cells from distinct bone marrow stem cells, e.g. hematopoietic stem cells or multipotent adult progenitor cells was found in recent in vitro studies. Cell culture experiments revealed a key role of growth factors for the induction of liver-specific genes in stern cell cultures. We investigated the potential of rat mesenchymal stem cells （MSC） from bone marrow to differentiate into hepatocytic cells in vitro. Furthermore, we assessed the influence of cocultured liver cells on induction of liver-specific gene expression. METHODS： Mesenchymal stem cells were marked with green fluorescent protein （GFP） by retroviral gene transduction. Clonal marked MSC were either cultured under liver stimulating conditions using fibronectin-coated culture dishes and medium supplemented with SCF, HGF, EGF, and FGF-4 alone, or in presence of freshly isolated rat liver cells. Cells in cocultures were harvested and GFP＋ or GFP- cells were separated using fluorescence activated cell sorting. RT-PCR analysis for the stem cell marker Thyl and the hepatocytic markers CK-18, albumin, CK-19, and AFP was performed in the different cell populations. RESULTS： Under the specified culture conditions, rat MSC cocultured with liver cells expressed albumin-, CK-18, CK-19, and AFP-RNA over 3 weeks, whereas MSC cultured alone did not show liver specific gene expression, CONCLUSION： The results indicate that （1） rat MSC from bone marrow can differentiate towards hepatocytic lineage in vitro, and （2） that the microenvironment plays a decisive role for the induction of hepatic differentiation of rMSC.

Chemokine expression in hepatocellular carcinoma versus colorectal liver metastases

AIM： To evaluate and compare the expression profiles of CXCL12 （SDF-1）, CCL19 （MIP-3β）, CCL20 （MIP-3a） and CCL21 （6Ckine, Exodus2） and their receptors on RNA and protein levels in hepatocellular carcinoma （HCC） versus colorectal liver metastases （CRLM） and to elucidate their impact on the carcinogenesis and progression of malignant liver diseases. METHODS： Chemokine expression was analyzed by RT-PCR and ELISA in 11 cases of HCC specimens and in 23 cases of CRLM and corresponding adjacent nontumorous liver tissues, respectively. Expressions of their receptors CXCR4, CCR6 and CCR7 were analyzed by RT- PCR and Western blot analysis in the same cases of HCC and CRLM. RESULTS： Significant up-regulation for CCL20/CCR6 was detected in both cancer types. Moreover, CCL20 demonstrated significant overexpression in CRLM in relation to the HCC tissues. Being significantly up-regulated only in CRLM, CXCR4 displayed an aberrant expression pattern with respect to the HCC tissues. CONCLUSION： Correlation of CXCR4 expression with CRLM suggests CXCR4 as a potential predictive factor for CRLM. High level expression of CCL20 and its receptor CCR6 in HCC and CRLM with marked up-regulation of CCL20 in CRLM in relation to HCC tissues indicates involvement of the CCL20/CCR6 ligand-receptor pair in the carcinogenesis and progression of hepatic malignancies.

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.

Inhibitory effect of adeno-associated virus-mediated gene transfer of human endostatin on hepatocellular carcinoma

AIM: To investigate the effect of adeno-associated virusmediated gene transfer of human endostatin on the growth of hepatocellular carcinoma (HCC).METHODS: HCC cell line Hep3B was infected with recombinantadeno-associated virus containing human endostatin gene (rAAV2-hEndo). The results of transfection were detected by RT-PCR and SDS-PAGE assay. MTT assay was used to observe the effects of supernatant of transfected cells on ECV304 cell proliferation. An animal model of HCC was established by injecting Hep3B cells subcutaneously into the back of nude mice. Intratumoral injection of rAAV2hEndo, empty virus and phosphate-buffered saline were given sequentially. Serum endostatin was determined byELISA, the inhibitory effect of endostatin on the growth of xenograft was assessed in 3 wk.RESULTS: The results of RT-PCR and SDS-PAGE assay confirmed that rAAV2-hEndo successfully transfected Hep3B cells, and endostatin was secreted from Hep3B cells to medium. The supernatant of transfected cells markedly inhibited the proliferation of ECV304 cells (P＜0.01). Intratumoral injection of rAAV2-hEndo (2×1010v.g.) led to a sustained serum endostatin level ofapproximately (86.71±5.19) ng/mL. The tumor volumeand microvessel density were less in rAAV2-hEndo group than in control groups (P＜0.01).CONCLUSION: Human endostatin can be stably expressed by adeno-associated virus-mediated gene transfer and effectively inhibit the growth of HCC.

Expression and significance of heparanase and nm23-Hl in hepatocellular carcinoma

AIM: To explore the relation between heparanase (HPA) and nm23-H1 in hepatocellular carcinoma (HCC), and whether they could be used as valuable markers in predicting post-operative metastasis and recurrence of HCC. METHODS: Reverse transcription-polymerase chain reaction and immunohistochemistry (S-P method) were used to measure the expressions of HPA mRNA and nm23-H1 protein in primary tumor tissue and paracancerous tissue of 33 cases of HCC. Paracancerous tissues of 9 cases of benign liver tumor were used as normal controls. The results were analyzed in combination with the results of clinicopathological examination and follow-up. RESULTS: The positive expression of HPA gene was significantly higher in primary tumor tissues of HCC (48.5%, 16/33) as compared to the paracancerous tissues of HCC and normal controls (3.03%, 1/33) (P<0.01). HPA expression was not related with the size of tumor, envelope formation, AFP level, HBsAg state and cirrhosis of liver. The positive rates of HPA mRNA in the group with high tendency to metastasis or recurrence and in the group with metastasis or recurrence during the follow-up were significantly higher than those in the group with low tendency to metastasis or recurrence (62.5% vs 37.5%, P<0.05) and in the group without metastasis or recurrence (78.6% vs 21.4%, P<0.01). The poorly differentiated tumor and tumor of TNM stages Ⅲ-Ⅳ had a higher positive rate of HPA gene expression than the well differentiated tumor and tumor of TNM stages Ⅰ-Ⅱ (66.7% vs 33.3%, P<0.05). The positive expression rate of nm23-H1 protein in HCC tissue was significantly lower than that in corresponding non-cancerous or normal liver tissue (45.5, 72.7, 88.9%, P<0.05). nm23-H1 expression was not related with the size of tumor, envelope formation, AFP level, HBsAg state, cirrhosis of liver, Edmondson grade, and TNM stage (P>0.05). The positive rates of nm23-H1 in the group with high tendency to metastasis and recurrence and in patients with metastasis or recurrence during the follow-up were obviously higher than those in the group with low tendency to metastasis and recurrence (P= 0.018) and in the patients without metastasis and recurrence (P = 0.024); but no significant difference was found between HPA positive and negative groups (P = 0.082). According to the results of follow-up, the rate of accuracy in predicting metastasis of positive HPA, negative nm23-H1 and combination of positive HPA with negative nm23-H1 was 78.6% (11/14), 68.8% (11/16) and 88.9% (8/9), respectively. CONCLUSION: Expression of HPA and/or nm23-H1 is related with metastasis and recurrence of HCC. Detection of the expression rate of HPA and nm23-H1 may help increase the accuracy in predicting post-operative metastasis and recurrence of HCC.

Overexpression of decoy receptor 3 in hepatocellular carcinoma and its association with resistance to Fas ligand-mediated apoptosis

AIM： To characterize the expression and genomic amplification of decoy receptor 3 （DcR3） in hepatocellular carcinoma （HCC） and to evaluate the role of DcR3 in apoptosis.METHODS： We examined 48 cases of HCC for DcR3 expression by RT-PCR and DcR3 gene amplification by quantitative genomic PCR. DcR3 protein was detected by immunohistochemistry. Terminal deoxynucleotidyl transferase-mediated dUTP digoxigenin nick and labeling （TUNEL） was used to identify the apoptosis cells in tissues. Primary hepatoma cell culture and MTT test were used to evaluate the protection against FasL- and chemicalinduced apoptosis by DcR3 expression. RESULTS： DcR3 mRNA overexpression was detected in 60% HCC （29/48） patients. The occurrence of HCC was not associated with amplification of the gene. One sample base substitution was found in three sites as a sequence in Genbank. The expression of DcR3 in HCC was associated with the apoptotic index （0.067±0.04 vs 0.209±0.12, P〈0. 01）, size of mass, stage, and infiltration or metastasis （41.2% vs71.0%, 40% vs75%, 51.8% vs84.6%, P〈0. 05）. DcR3 expression could protect hepatoma cells against apoptosis induced by FasL, but not by chemicals. CONCLUSION： These data suggest that in addition to gene amplification there may be another mechanism underlying DcR3 overexpression. The effect of overexpression of DcR3 on the apoptosis of cancer cells may have direct therapeutic implications for the management of HCC.

Genomic-wide analysis of lymphatic metastasis-associated genes in human hepatocellular carcinoma

AIM： To identify the genes related to lymph node metastasis in human hepatocellular carcinoma （HCC）, 32 HCC patients with or without lymph node metastasis were investigated by high-throughput microarray comprising 886 genes. METHODS： The samples of cancerous and noncancerous paired tissue were taken from 32 patients with HCC who underwent hepatectomy with lymph node dissection. Total RNA was extracted from the cells obtained by means of laser microdissection （LCM） and was amplified by the T7-based amplification system. Then, the amplified samples were applied in the cDNA microarray comprising of 886 genes. RESULTS： The results demonstrated that 25 upregulated genes such as cell membrane receptor, intraceliular signaling and cell adhesion related genes, and 48 down-regulated genes such as intracellular signaling and cell cycle regulator-related genes, were correlated with lymph node metastasis in HCC. Amongst them were included some interesting genes, such as MET, EPHA2, CCND1, MMP2, MMP13, CASP3, CDH1, and PTPN2. Expression of 16 genes （MET, CCND1, CCIVD2, VEGF, KRT18, RFC4, BIRC5, CDC6, MMP2, BCL2A1, CDH1, VIM, PDGFRA, PTPN2, SLC25A5 and DSP） were further confirmed by real-time quantitative reverse transcriptional polymerase chain reaction （RT-PCR）. CONCLUSION： Tumor metastasis is an important biological characteristic, which involves multiple genetic changes and cumulation. This genome-wide information contributes to an improved understanding of molecular alterations during lymph node metastasis in HCC. It may help clinicians to predict metastasis of lymph nodes and assist researchers in identifying novel therapeutic targets for metastatic HCC patients.

Genes transactivated by hepatitis C virus core protein, a microarray assay

AIM: To explore the new target genes transactivated by hepatitis C virus (HCV) core protein and to elucidate the pathogenesis of HCV infection. METHODS: Reverse transcribed cDNA was subjected to microarray assay. The coding gene transactivated by HCV core protein was cloned and analyzed with bioinformatics methods. RESULTS: The expressive vector of pcDNA3.1(-)-core was constructed and confirmed by restriction enzyme digestion and DNA sequencing and approved correct. mRNA was purified from HepGZ and HepG2 cells transfected with pcDNA3.1(-)-core, respectively. The cDNA derived was subjected to microarray assay. A new gene named HCTP4 was cloned with molecular biological method in combination with bioinformatics method. CONCLUSION: HCV core is a potential transactivator. Microarray is an efficient and convenient method for analysis of differentially expressed genes.

Possible mechanism for hepatitis B virus X gene to induce apoptosis of hepatocytes

AIM： To investigate the possible mechanism for HBV X gene to induce apoptosis of hepatocyte HL-7702 cells.METHODS： HBV X gene eukaryon expression vector pcDNA3-X was established and transfected into HL-7702 cells by lipid-mediated transfection, including transient and stable transfection. Positive clones were screened by incubating in the selective medium with 600 μg/mL G418 and named HL-7702/HBV-encoded X protein （HBx） cells. The expressions of Fas/FasL, Bax/Bcl-2, and c-myc mRNA were measured by semi-quantitative RT-PCR in HL-7702/HBx and control group, respectively.RESULTS： RT-PCR analysis confirmed that HBV X gene was transfected into HL-7702 cells successfully. By semiquantitative RT-PCR analysis, Bax and c-myc mRNA levels in HL-7702/HBx cells of transient transfection were significantly higher than those in control, FasL and c-myc mRNA levels in HL-7702/HBx cells of stable transfection were significantly higher than those in control, whereas the Bcl-2 mRNA levels in HL-7702/HBx cells of transient and stable transfection were significantly lower than thosein control.CONCLUSION： HBV X gene may promote the apoptosis of hepatocytes by regulating the expressions of Fas/FasL, Bax/Bcl-2, and c-myc gene in a dose-dependent manner.

Increased DNA binding activity of NF-κB,STAT-3,SMAD3 and AP-1 in acutely damaged liver

AIM: To investigate the role of genes and kinetics of specific transcription factors in liver regeneration, and to analyze the gene expression and the activity of some molecules crucially involved in hepatic regeneration. METHODS: USING gel-shift assay and RT-PCR, transcription factors, such as NF-κB, STAT-3, SMAD3 and AP-1, and gene expression of inducible nitric oxide synthase (iNOS), hepatocyte growth factor (HGF) and c-met were analyzed in an animal model of chemically induced hepatectomy. RESULTS: Gene expression of HGF and its receptor c-met peaked at 3 h and 24 h after acute CCl4 intoxi- cation. iNOS expression was only observed from 6 to 48 h. Transcriptional factor NF-κB had an early activation at 30 min after acute liver damage. STAT-3 peaked 3 h post- intoxication, while AP-1 displayed a peak of activation at 48 h. SMAD3 showed a high activity at all analyzed times. CONCLUSION: TNF-α and IL-6 play a central role in hepatic regeneration. These two molecules are responsible for triggering the cascade of events and switch-on of genes involved in cell proliferation, such as growth factors, kinases and cyclins which are direct participants of cell proliferation.

Effects of Wei Chang An on expression of multiple genes in human gastric cancer grafted onto nude mice

AIM： To investigate the expression of multiple genes in Chinese jianpi herbal recipe Wei Chang An （WCA） in human gastric cancer cell line SGC-7901. METHODS： A human gastric adenocarcinoma cell line SGC-7902 grafted onto nude mice was used as the animal model. The mice were randomly divided into 3 groups, one control and the two representing experimental conditions. Animals in the two experimental groups received either WCA over a 34-d period or 5-fluorouracil （5-FU） over 6-d period starting at 8th d after grafting. Control animals received saline on an identical schedule. Animals were killed 41 d after being grafted. The expression profiles in paired WCA treated gastric cancer samples and the N.S. control samples were studied by using a cDNA array representing 14181 cDNA clusters. The alterations in gene expression levels were confirmed by Real-time Quantitative polymerase chain reaction （qPCR）. RESULTS： When compared with controls, the average tumor inhibitory rate in WCA group was 44.32% ± 5.67% and 5-FU 47.04% ± 22.33% （P 〈 0.01, respectively）. The average labeling index （LI） for PCNA in WCA group and 5-FU group was significantly decreased compared with the control group. Apoptotic index （AI） was significantly increased to 9.72% ± 4.52% using the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate fluorescence nick end labeling （TUNEL） method in WCA group compared with the controls 2.45% ± 2.37%. 5-FU group was also found to have a significantly increased AI compared with the controls. The expression of cleaved Caspase-3 in WCA group and 5-FU group was significantly increased compared with the control group respectively. There were 45 different expressed sequence tags （ESTs） among the control sample pool and WCA sample pool. There were 24 ESTs up-regulated in WCA samples and 21 ESTs down-regulated. By using qPCR, the expression level of Stat3, rap2 interacting protein x （RIPX）, regulator of differentiation 1 （ROD1） and Bcl-2 was lower in WCA group than that in control group respectively. By using SP immunohistochemical method the expression of Phospho-Stat3 （Tyr705） and Bcl-2 in WCA group and 5-FU group was significantly decreased compared with the control group respectively. CONCLUSION： WCA could inhibit gastric cancer cell SGC-7901 growth in vivo. WCA could induce gastric cancer cell apoptosis and suppress proliferation. Its mechanisms might be involved in the down-regulation of Star3, RIPX, ROD1 and Bcl-2 gene.

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.