Expression and its clinicopathological features of tumor rejective antigen 1 in human tissues of hepatocellular carcinoma and liver cirrhosis / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the expression of tumor rejective antigen 1 in hepatocellular carcinoma (HCC) and liver cirrhosis(LC) tissues, and the relationship between clinicopathological feature and HCC.</p><p><b>METHODS</b>The expressions of TRA1 mRNA and its protein were detected by reverse transcription polymerase chain reaction (RT-PCR) and Western blot respectively. Immunohistochemical staining was used to further examine the expression of TRA1 protein in LC, HCC and control tissues. The relationship between clinicopathological feature and HCC was analyzed. Data of RT-PCR and Western blot were analyzed by One-way ANOVA; results of immunohistochemical staining were analyzed by Fisher's exact test and correlation analysis using Spearman rank correlation.</p><p><b>RESULTS</b>RT-PCR data showed that the expression of TRA1 mRNA was higher in HCC and LC tissues than that in the normal liver tissues (F values were 20.821 and 12.311 respectively, P is less than 0.05). The expression of TRA1 protein in HCC and LC tissues was signifIcantly higher than that in control by Western blot (F values were 21.231 and 20.125 respectively, P < 0.05). The immunohistochemical data showed the expression of TRA1 protein was gradually increased in HCC group than that in the LC group and control group, and the positive expression rate of TRA1 was 57.14%, 78.95% and 93.75% respectively. The expression of TRA1 protein was negatively correlated with HCC differentiation (r = -0.4655, P = 0.0073) and positively correlated with HCC TNM staging (r = 0.5157, P = 0.0025).</p><p><b>CONCLUSION</b>The over-expression of TRA1 in hepatocirrhosis and HCC is correlated with the formation and development of HCC. It may be a prognostic marker for the diagnosis of HCC and be associated with the degree of differentiation and HBV infection. It can be used as a marker for prognostic prediction of HCC.</p>

Effects of antisense RNA of connective tissue growth factor expressing plasmid on rat liver fibrosis / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To observe the effects of antisense RNA of connective tissue growth factor (CTGF) on rat liver fibrosis.</p><p><b>METHODS</b>Gene recombinant techniques were used to construct a rat antisense RNA of CTGF recombinant plasmid which could be expressed in eukaryotic cells. The recombinant plasmids were encapsulated with lipofectamine and then transducted into a carbon tetrachloride (CCl4) induced rat liver fibrosis model. Expression of CTGF was assessed by RT-PCR, Western blot and immunohistochemistry. Immunohistochemistry was used to identify type I and III collagens. HE stained liver slides were used for pathological study.</p><p><b>RESULTS</b>The mRNA and protein expression of CTGF in the fibrotic liver transfected with antisense-CTGF were significantly decreased compared with those of the controls (P<0.01). The depositions of type I and type III collagens were also decreased (P<0.05). Antisense-CTGF also minimized the pathological fibrosis in the rat livers (P<0.01).</p><p><b>CONCLUSION</b>The results demonstrate that the antisense RNA of CTGF recombinant plasmid has certain effects in preventing liver fibrosis and makes it a possible candidate for use in future gene therapy.</p>

The expression of platelet-derived growth factor (PDGF) receptor-beta and its correlation with extracellular matrix in hepatic tissue in hepatic fibrosis rats / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the expression of PDGF receptor-beta and its correlation with extracellular matrix in hepatic tissue during hepatic fibrosis.</p><p><b>METHODS</b>The model of hepatic fibrosis in rats was induced by carbon tetrachloride. PDGF receptor-beta subunit, collagen I, collagen III and a-SMA in hepatic tissues of these rats were examined using immunohistochemistry. The correlation between PDGF receptor-beta subunit and collagen I, III was analyzed using SAS software after the results of immunohistochemistry were semi-quantified.</p><p><b>RESULTS</b>PDGF receptor-beta subunit and a-SMA were not detected in normal controls. Collagen I and III were distributed in the portal tracts and beneath the endothelia of the central veins and of the Disse spaces. Two weeks after CCl4 injection, the PDGF receptor-beta and a-SMA were detected, and the expression of collagen I and III increased. At the end of 4 and 6 weeks, the above four proteins were further increased. Two weeks after CCl4 injection, PDGF receptor-beta had no apparent correlation with collagen I and III. However, PDGF receptor-beta had a significant correlation with collagen I and III 2 weeks later, and the correlation coefficient was 0.74 and 0.60 respectively at 4 weeks, and 0.83 and 0.67 respectively at 6 weeks. PDGF receptor-beta had a significant correlation with a-SMA during the whole process of hepatic fibrosis and the correlation coefficient was 0.62, 0.69 and 0.81, respectively at the time of 2, 4 and 6 weeks after CCl4 injection.</p><p><b>CONCLUSION</b>The PDGF receptor-beta was overexpressed during the process of hepatic fibrosis development, and it significantly correlated with collagen I and collagen III.</p>

Preparation and purification of siRNA targeting a proliferation-inducing ligand of pancreatic cancer cell line / 第二军医大学学报

Objective:To prepare and purify siRNA targeting a proliferation-inducing ligand targeted(APRIL-siRNA),so as to provxde a basis for studying the role of APRIL in human pancreatic cancer.Methods:pET-22b-APRIL was constructed to express APRIL dsRNA of human pancreatic cancer cell line CFPAC-1 in E.coli and the product was purified by chromatography using CF-11 column.APRIL dsRNA was digested by RNaseⅢto prepare APRIL siRNA,then the reaction mixture was loaded onto a DEAE ion exchange chromatography to remove RNaseⅢfrom oligonucleotides,and size exclusion chromatography was used to purify 21 bp siRNA.The purified APRIL siRNA was used to transfect Chinese hamster ovary(CHO)cells and the expression of APRIL in CHO cells was observed under fluorescence microscope Results:APRIL dsRNA was successfully expressed in E.coli after IPTG induction and was purified by CF-11 column.dsRNA was hydrolyzed with RNaseⅢand was purified by DEAE ion exchange chromatography and size exclusion chromatography.15% nondenaturing PAGE and 12% SDS- PAGE confirmed that RNaseⅢwas removed from oligonucleotides and 21 bp siRNA was purified with size exclusion chromatography.It was also found that APRIL siRNA obviously depressed APRIL expression in CHO cells.Conclusion:We have successfully constructed APRIL siRNA targeting APRIL gene of CFPAC-1 cells with in vitro transcription,which provides a basis for knock-down of APRIL gene in CFPAC-1 cells.