Relationship Between Genetic Instability Of Nm23H1 Gene And Clinical Pathological Characteristic In Original Hepatocellular-Gallbladder Tumor

BackgroundThe malignant tumors have threatened human health and lives severely. One of the most important reasons is that the microinvasion or micrometastasis exists at the time of surgery and can not be controlled. Therefore it is very important to clarify the mechanism of invasion and metastasis, and quite significant to establish some definite molecular marker and target for the therapy.It has been investigated that the expression of anti-oncoprotein becomes low with the occurrence and metastasis of tumor, then the growth and metastasis of tumor cell are out of control. The main reason that the expression of anti-oncoprotein reduces is anti-oncogenes inactivity. To investigate the mechanism of genetic inactivity, scientists did lots of research on many anti-oncogenes such as P53, P16, FHIT et al, and found that the genetic instability, the representative of microsatellite instability and loss of heterozygosity, might be one important factor that leads to genetic mutation, the anti-oncogenes function maladjustment, and the tumor occurrence.Microsatellite instability (MSI) and loss of heterozygosity (LOH), the alteration in length and strength of short tandem repeat sequences are important molecular characteristics of many human tumors. Many researches indicated that MSI or LOH of anti-oncogenes plays an important role in occurrence of sporadic tumor. MSI was firstfound in tumors of the hereditary non-polyposis colon carcinoma (HNPCC) syndrome, and then it is found in many other kinds of tumors, such as colon cancer, gastric cancer, carcinoma of endometrium, breast cancer, prostatic cancer and pancreatic cancer, et al. Many studies suggested that the occurrence of MSI increase the frequence of mutation, and moreover, it leads to mutation of genes which has great relationship with tumor which may be an important mechanism of tumor occurrence. According to Knudson's two-hit hypothesis of TSG (tumor suppressor gene) inactivation, the common chromosomal region of LOH is a potential site harbouring TSG, thus LOH is regarded as a valuable molecular genetic marker to find TSGnm23H1 gene is an important anti-oncogene which locates at 17th chromosome (17q21). Steeg et al found that the nm23H1 protein/NDPK, the expression product of nm23H1 gene, could suppress tumor metastasis effectively. However, the studies were mostly on the immunochemistry research of the nm23H1 protein expression, the genetic research of nm23H1 gene on tumor metastasis was rarely reported.ObjectiveTo investigate MSI and LOH of locus D17S396 in original hepatocellular-gallbladder tumor, and their effect on the expression of nm23H1, which would reveal the function mechanism of anti-oncogene and provide experimental basis for metastasis mechanism of cancer.Methods(1) The technology of extraction DNA from paraffin-embedded materials. (2) Polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and displaying band by ordinary silver stain. (3) Expression of nm23H1 was detected by Envision immunohistochemistry. (4) Leica-Qwin computer imaging techniques. (5) SPSSstatistic software analyzed the experimental results.Results(1) Study on genetic instability of nm23H1 gene in gallbladder tumorâ‘ In our experiment, the frequency of heredity instability of malignant gallbladder tumors is 42.55%, which is higher than that of gallbladder adenomas, while there were no heredity instability occured in chronic cholecystitis tissue. The frequency of LOH seemed higher with the deteriorism of gallbladder tumor. Among 47 gallbladder carcinomas, the frequency of LOH is different between different differentiation cases (P <0.05), and the frequency of LOH in liver and lymph node metastasis cases was significantly higher than those without metastasis (P<0.01). Moreover, the frequency of LOH was higher in stage Nevin IV and V when compared with stage â… , â…¡ and â…¢. However, the frequency of MSI showed contrary correlation with some clinicopathologic characteristics compared with LOH. â‘¡The expression of nm23H1 in gallbladder carcinoma, gallbladder adenoma and chronic cholecystitis tissue were 46.81% , 26.09% and 10% (P<0.05). Moreover, the case with lymph node metastasis showed significantly lower nm23H1 expression than those without lymph node metastasis (P<0.01 ). Nevin stage â…£ and V also exhibited lower nm23H1 expression levels compared with stage â… , â…¡ and â…¢. Furthermore, there was no difference in nm23H1 protein expression intensity analyzed by computer imaging techniques. â‘¢ In gallbladder carcinomas, the positive frequency of nm23H1 protein in LOH positive group was 11.11%, which was lower than that of LOH negative group (P <0.05).(2) Study on genetic instability of nm23H1 gene in hepatocellular carcinoma(HCC)â‘ the frequency of heredity instability of HCCs are 35.42%. The frequency of LOH in the cases with lymph node or distant organs metastasis or not and with intrahepatic metastasis or embolus of portal vein or not are significantly different (P<0.01), it was higher in stage TNM â…¢ than in stage â…  , â…¡. Moreover, it was higher in high tendency to invasion ormetastasis cases than those in the low tendency cases (P<0.01). â‘¡The expression of nm23H1 are 56.25%. It was significantly different in Edmondson grade,TNM stage and in lymph node or distant organ metastasis cases (P<0.01). the cases with higher tendency of invasion or metastasis exhibited lower nm23H1 expression compared with low tendency cases (P<0.01). â‘¢the positive rate of nm23H1 protein in LOH positive group was lower than that of LOH negative group (P<0.05).ConclusionsThe results indicated that the heredity instability of nm23H1 gene may be implicated in pathogenesis and progression of hepatocelluar-gallbladder tumor. The occurrence of LOH may be molecule marker for the deteriorism of hepatocelluar-gallbladder tissue. Both MSI and LOH of nm23H1 gene controlled the development of hepatocelluar-gallbladder tumor independently in different paths. LOH could inhibit the expression of nm23H1 in local tissue of hepatocelluar-gallbladder carcinoma, which endowed it with high aggressive and poor prognosis. Increasing the amount of nm23H1 protein expression could effectively restrain hepatocelluar-gallbladder carcinoma metastasis and improve prognosis of patients.