| ObjectiveTo analysis the expression of Wnt-5a, Ror2 mRNA and protein in hepatoc-ellular carcinoma, to research wether Ror2 is the downstream genen of Wnt-5a; to analysis the relationship among Wnt-5a, Ror2 and Wnt/β-Catenin classic path,to analysis the role of wnt classic and inclassic pathway in the carcinogenesis and development of hepatocellular carcinoma according to the expression of P-Catenin.And research the relationship among Wnt-5a,Ror2 and several clinical pathology agent and survival rate of the patient after operation, to provide theory basis for prevention and cure criterion prognosis of hepatocellular carcinoma.Method1. In our study, we collectde 19 cases of the fresh hepatocellular carcin-oma samples from exairesis (which to get out of the way of necrosis), real time fluorescent quantitation polymerasechain reaction (FQ-PCR) was used to detect the expression of Wnt-5a,Ror2 mRNA in hepatocellular carcinoma and the corresponding peri-tumor tissue(beyond the tumor edge two centimeters).2.SP immunohistochemical method was used to detect the expression of Wnt-5a,Ror2 andβ-catenin protein in 62 cases of hepatocellular carcinoma and the corresponding peri-tumor tissues,10 cases cirrhosis tissues,6 cases normal hepatocellular tissues from exairesis the hepatocellular tissues were operated due to Bile duct stone calculus in liver or liver break caused by trauma.3. We investigated the prognosis of every patient suffer from HCC, to research the value of Wnt-5a,Ror2 in HCC prognosis; we did multiplicity by Cox proportion risk model to evaluate the existence index of patients'prognosis.Results一. RT-PCR results 1. The level of Wnt-5a mRNA in hepatocellular carcinoma and peri-tumor were 0.102030±0.158560,0.020113±0.022084 respectively,with a significant difference(t=2.22, P=0.039), Wnt-5a mRNA was up-regulation in73.68% (14/19) hepato-Cellular carcinoma.2. The level of Ror2 mRNA in hepatocellular carcinoma and peri-tumor were 0.001665±0.002585,0.006504±0.012512,with no significant difference (t=-1.934,P=0.069), Ror2 mRNA was down-regulation in 73.68%(14/19) hepatocellular carcinoma.二. Immunohistochemical results1. We found that Wnt-5a expression was weakly positive or negative in kytoplasm of all normal hepatocellular tissues, it was asytematic and medium intensity positive in 90%(9/10) cirrhosis tissues and in 82.26%(51/62) peri-tumor tissues, the weakly positive or negative rate of Wnt-5a in hepato-cellular carcinoma was 85.49%(53/62). Compare with the cirrhosis tissues and peri-tumor tissues Wnt-5a expresseion was dowm-regulation obviously or negative in hepatocellular carcinoma,with a signifinace difference (P<0.001). Wnt-5a expression was dowm-regulation obviously or negative in hepatocellular carcinoma, was significantly associated with high tumor stages(P=0.004), high serum AFP level (P=0.013) and high Ki-67 index (P=0.006), but was not correlated with sex of patients,age,size of tumor, HBsAg, Hepatic cirrhosis and the differentiation level of tumor(P>0.05).2. Ror2 expression was weakly positive or negative in cytoplasm of normal hepatocellular tissues, it was asytematic and medium intensity positive in 80% cirrhosis tissues (8/10) and in 85.48%(53/62) peri-tumor tissues, and the weakly positive or negative rate of Wnt-5a in hepatocellular carcinoma was 80.65% (50/62).Compare with the peri-tumor tissues and cirrhosis tissues Ror2 expresseion was dowm-regulation obviously or negative in hepatocellular carcinoma,with a signifinace difference (P<0.001). Ror2 expression was dowm-regulation obviously or negative in hepatocellular carcinoma, was significantly associated with high tumor stages (P=0.021), high serum AFP level (P=0.004), but was not correlated with sex of patients, age,size of tumor,hepatic cirrhosis, HBsAg and the differentiation level of tumor (P>0.05).3. The cell membranous expression rate ofβ-catenin in normal liver tissues was 100%, it presents brown and yellow. The membranous expression ofβ-catenin was reduced and it was observed in cytoplasm and/or cellular nucleus,in hepatocellular carcinoma tissues. There was no nucelus expression in hepatic cirrhosis and peri-HCC.The expression ofβ-catenin was down regulation in membranous and abnormal cytoplasm and nucelus expression in67.74%(42/62) HCC, and there was statistics difference with the normal membranous expression (P<0.001).The statistics result show that abnormal expression ofβ-catenin was associated with tumor lowly differentiation level (P=0.001),but was not correl-ated with sex of patients, age, size of tumor, stages of tumor,AFP level, HbsAg, hepatic cirrhosis (P>0.05)。三. The correlation between gene and protein1. There was negative correlation between Wnt-5a mRNA and Ror2 mRNA expression in hepatocellular carcinoma (r=-0.571, P=0.011)2. There was no correlation between Wnt-5a mRNA and Wnt-5a protein expression in hepatocellular carcinoma (r=-0.367, P=0.123)3. There was positive correlation between Ror2 mRNA and Ror2 protein expression in hepatocellular carcinoma (r=0.457, P=0.049)4. There was positive correlation between Wnt-5a and Ror2 protein down-regulation expression in hepatocellular carcinoma(r=0.340, P<0.01), and there was negative correlation between Wnt-5a, Ror2 down-regulation expression andβ-catenin protein abnormal expression in hepatocellular carcinoma (r=-0.389,P=0.002;r=-0.300,P=0.018)四. The survival analysis after HCCKaplan-Meier survival analysis show that:The negative expression of Wnt-5a and Ror2 in HCC has correlation with the survival time and rate of patient (P<0.05).Proportional hazard regression model indicate that:the patients'survival time after operation has correlation with poor tumor stage (P=0.005) tumor lowly differentiation level (P=0.004), and hepatic cirrhosis (P=0.011) but was not correlated with sex of patients, age, size of tumor, AFP level,HbsAg(P>0.05)。Conclusion1. The down-regulation of Wnt-5a protein expression is a frequent event in the occurrence and deveploment of HCC, Wnt-5a may play the role of anti-nocogene in the occurrence and deveploment of HCC, it correlation with the suivival rate and prognosis of patient, prompt us it may be a diagnosis and bad prognosis maker of HCC.2. Down-regulation of Ror2 protein expression is a frequent and early period event in the occurrence and development of HCC, it ma play the role of anti-nocogene in the occurrence and deveploment of HCC,it correlation with the suivival rate and prognosis of patient, abnormal expression of Ror2 may was associated with the occurrence and development of HCC.3.β-catenin ectopy expression may be associated with the occurrence and development of HCC;β-catenin plays the role of proto-oncogene in the process.4. There was positive correlation between Wnt-5a and Ror2 protein expression in HCC, prompt us that Ror2 may be the downstream gene of Wnt-5a, both of them mediate Wnt inclassic pathway. |
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