Dynamic alteration of telomerase expression and its diagnostic significance in liver or peripheral blood for hepatocellular carcinoma

AIM: To investigate the dynamic alteration of telomerase expression during development of hepatocellular carcinoma (HCC) and its diagnostic implications in liver tissues or peripheral blood mononuclear cells for HCC. METHODS: Dynamic expressions of liver telomerase during malignant transformation of hepatocytes were observed in Sprague-Dawly (SD) rats fed with 0.05% of 2-fluoenyacetamide (2-FAA). Total RNA and telomerase were extracted from rat or human liver tissues. The telomerase activities in livers and in circulating blood were detected by a telomeric repeat amplification protocol-enzyme-linked immunosorbent assay (TRAP-ELISA), and its diagnostic value was investigated in patients with benign or malignant liver diseases. RESULTS: The hepatoma model displayed the dynamic expression of hepatic telomerase during HCC development. The telomerase activities were consistent with liver total RNA levels (r = 0.83, P < 0.01) at the stages of degeneration, precancerosis, and cancerization of hepatocytes. In HCC patients, the telomerase levels in HCC tissues were significantly higher than in their adjacent non-cancerous tissues, but liver total RNA levels were lower in the former than in the latter. Although the circulating telomerase of HCC patients was abnormally expressed among patients with chronic liver diseases, the telomerase activity was a non-specific marker for HCC diagnosis, because the incidence was 15.7% in normal control, 25% in chronic hepatitis, 45.9% in liver cirrhosis, and 85.2% in HCC, respectively when absorbance value of telomerase activity was more than 0.2. If the value was over 0.6, the incidence was 60% in HCC group and 0% in any of the others (P < 0.01) except in two cases with liver cirrhosis. However, the combination of circulating telomerase with serum alpha-fetoprotein level could increase the positive rate and the accuracy (92.6%, 125 of 135) of HCC diagnosis. CONCLUSION: The overexpression of telomerase is associated with HCC development, and its abnormality in liver tissues or in peripheral blood could be a useful marker for diagnosis and prognosis of HCC.     

Abnormal expression of HSP gp96 associated with HBV replication in human hepatocellular carcinoma

BACKGROUND: Heat shock protein (HSP) gp96 is a member of the HSP90 family and presumably overexpresses as a result of stimulation by mutated or abnormal proteins. Its abnormal expression correlates with carcinogenesis, progression and prognosis of hepatocellular carcinoma (HCC). In this study, we investigated the pathological characteristics of liver gp96 expression and its relationship with hepatitis B virus (HBV) replication in HCC patients. METHODS: Tumor specimens were prospectively collected from 30 HCC patients undergoing liver resection. Total RNAs were extracted from HCC or their noncancerous tissues. The distribution of gp96 expression in hepatocytes was investigated by streptavidin peroxidase (S-P) immunohistochemistry and tissue HBV-DNA was detected by the in situ molecular hybridization technique. The association of gp96 expression with HBV replication, and the histopathological characteristics of HCC were analyzed. RESULTS: The gp96 was strongly expressed in HCC (73.3%, 22 of 30) and weakly (46.7%, 14 of 30) in non-cancerous tissues. The gp96 expression in HCC tissues was correlated with degree of tumor differentiation and tumor size, but not with tumor number (P>0.05). Immunohistochemical analysis showed that 17 of 19 HCC patients with HBVDNA -positive were strongly expressed for gp96, whereas only 5 of 11 patients with HBV-DNA-negative were positive for gp96. A significant difference was found between the two groups (89.5% vs. 45.5%, P<0.05). CONCLUSIONS: The abnormal expressions of HSP gp96 in HCC tissues are associated with HBV replication. This finding indicates that HBV infection plays an important role in the development of HCC.     

Expression of focal adhesion kinase and α5andβ1integrins in carcinonas and its clinical significance

AIM：To detect the expression pattern of FAK(focal adhesion kinase)and integrinα5andβ1 subunits in different kinds of cancerous tissues and to study their correlation with clinicopathological data including tumor type,grade and lymph node status.METHODS:Using an immunohistochemical technique,we examined the expression of FAK and integrin and subunits in cancerous and noncancerous tissues obtained from75patients with gastric carcinomas,21colorectal carcinomas,16 hepatocellular carcinomas,20uterocervical carcinomas,and20breast carcinomas.RESULTS:The staining of FAK was stronger in cancerous than in noncancerous areas,Enhanced expression of FAKwas detected in poor-differentiated carcinoma of the stomach and colorectum.Tumors with lymph node metastases had more FAKprotein than those without metastases.In addition.the deeper the extent of tumor infiltration,the higher the FAKexpression.The expression of integrinα5andβ1subunits was lower in cancerous areas than in noncancerous areas,but it was higher in well-differentiated cancerous tissues than in poor differentiated tissues.The relationship between the expression of integrinα5andβ1subunits and infiltration or metastasis was not significant.Cancerous tissues with stronger FAK expression(++or+++)also had a higher expression of integrinα5andβ1subunits in the tumor and its unaffected margins.CONCLUSION:FAKis a better marker for carcinogenesis and the progression of cancer than integrinα5orβ1subunit,and it may be not only a transformation-linked enzyme but also a progression-linked enzyme.     

Expression of gap junction genes connexin 32, connexin 43 and their proteins in hepatocellular carcinoma and normal liver tissues

AIM To investigate the significance and mechanism of cx32 mRNA, cx43 mRNA and their proteins in hepatocarcinogenesis.METHODS Sixty-one cases of HCC and 14 cases of normal liver tissues were detected by immunohistochemical and in situ hybridization (ISH) methods.RESULTS In HCC grades Ⅰ,Ⅱ,Ⅲ and normal liver tissues, the positive rates of Cx32 protein were 55.6%, 42.1%, 18.2% and 92.9%,respectively. The detection rates of Cx43 protein were 44.4%, 26.3%, 12.1% and 78.6%,respectively. There was significant difference in Cx32 and Cx43 protein between HCC and normal liver tissues (P＜0.01). ISH the positive rates of cx32 mRNA shown by ISH in HCC grades Ⅰ,Ⅱ,Ⅲ and normal liver tissues were 88.9%, 84.2%,87.9% and 92.9%, respectively. Those of cx43 mRNA were 77.8%, 78.6%, 78.8% and 85.7%,respectively. There was no statistical difference in the positive rates of cx32 mRNA and cx43 mRNA between HCC and normal liver tissue (P＞0.05).CONCLUSION The aberrant location of Cx32 and Cx43 proteins could be responsible for progression of hepatocarcinogenesis, and the defect of cx genes in post-translational processing might be the possible mechanism.     

Preventive effect of regional radiotherapy with phosphorus-32 glass microspheres in hepatocellular carcinoma recurrence after hepatectomy

AIM： To evaluate the preventive effects of phosph- orus-32 glass microspheres （P32-GMS） in the recurrence of massive hepatocellular carcinomas （HCCs） after tumor resection. METHODS： Twenty-nine patients with massive HCCs received local P^32-GMS implantation after liver tumors were removed, while the other 38 patients with massive HCCs were not treated with P^32-GMS after hepatectomies. The radioactivity of the blood, urine and liver were examined. The complications, HCC recurrence and overall survival rates in the patients were analyzed. RESULTS： P^32-GMS implanted in the liver did not cause systemic absorption of p^32. There were no significant differences of postoperative complications between the patients with and without P^32-GMS treatment. The shortterm （six months and 1 year） and long-term （2, 3 and over 3 years） recurrence rates in patients who received P^32-GMS radiotherapy were significantly decreased, and the overall survival rates in this group were significantly improved. CONCLUSION： P^32-GMS implantation in the liver can significantly decrease the postoperative recurrence and improve the overall survival in HCCs patients after hepatectomy. This therapy may provide an innovative method in prevention of HCC recurrence after operation.     

Correlation between expression of cyclooxygenase-2 and the presence of inflammatory cells in human primary hepatocellular carcinoma： Possible role in tumor promotion and angiogenesis

AIM: To investigate the association of cyclooxygenase-2 (COX-2) expression with angiogenesis and the number and type of inflammatory cells (macrophages/Kupffer cells; mast cells) within primary hepatocellular carcinoma (HCC) tissues and adjacent non-tumorous (NT) tissues. METHODS: Immunohistochemistry for COX-2, CD34, CD68 and mast cell tryptase (MCT) was performed on 14 well-characterized series of liver-cirrhosis-associated HCC patients. COX-2 expression and the number of inflammatory cells in tumor lesions and surrounding liver tissues of each specimen were compared. Moreover, COX-2, CD34 staining and the number of inflammatory cells in areas with different histological degrees within each tumor sample were comparatively analyzed. RESULTS: The percentage of COX-2 positive cells was significantly higher in NT tissues than in tumors. COX-2 expression was higher in well-differentiated HCC than in poorly-differentiated tissues. Few mast cells were observed within the tumor mass, whereas a higher number was observed in the surrounding tissue, especially in peri-portal spaces of NT tissues. Abundant macrophages/ Kupffer cells were observed in NT tissues, whereas the number of cells was significantly lower in the tumor mass. However, a higher cell number was observed in the well-differentiated tumor and progressively decreased in relation to the differentiation grade. Within the tumor, a positive correlation was found between COX-2 expression and the number of macrophages/Kupffer cells and mast cells. Moreover, there was a positive correlation between CD34 and COX-2 expression in tumor tissues. Comparison between well- and poorly-differentiated HCC showed that the number of CD34-positive cells decreased with dedifferentiation. However, COX-2 was the only independent variable showing a positive correlation with CD34 in a multivariate analysis. CONCLUSION: The presence of inflammatory cells and COX-2 expression in liver tumor suggests a possible relationship with tumor angiogenesis. COX-2 expressing cells and the number of macrophages/Kupffer cells and mast cells decrease with progression of the disease.