A Case-control Study On The Association Between Genetic Polymorphisms Of DNA Rapair And Hepatic Cell Cancer Susceptibility

Hepatocellular carcinoma is one of the malignant tumor that threaten our healthy and life obviously.Although hepatocelllar carcinoma has been proved to be related to the infection of hepatitis B,aflatoxin and so on,only a few part of population that exposed to the risk factors suffered hepatocellular carcinoma.The population or individual susceptibility may be relation to the cause,that is different population or individuals had different liability to the carcinogen.The main factors that decide susceptibility are genetic differences.DNA repair gene and metabolic enzymatic gene are chief components of tumor-related gene.These gene possess single nucleotide polymorphisms whose product usually has different biologic activity and therefore different susceptibilities to the same carcinogen have been expressed.DNA repair system is a chief defense barrier in vivo,taking part in repairing DNA injury caused by internal and outside environmental factors.The absence or decrease of DNA repair ability could increase the risk of genetic mutation and cell canceration. Therefore,individuals that have lower DNA repair ability would be susceptible to tumor.XPD and XRCC1 are important DNA repair gene,participating the repair of incision of DNA sequence and nucleotide.Many sites on XPD and XRCC1 exist SNP phenomenon,moreover,they are related to various tumor susceptibility,such as lung cancer,gastric carcinoma,breast cancer and so on.However,very few research on the relationship of XPD and XRCC1 with primary hepatocellular carcinoma had been reported. Our research explored the relationship of common DNA repair genetic polymorphisms such as XPD Lys751Gln,XRCC1 Arg194Trp,XRCC1 Arg280His, XCCR Arg399Gln and susceptibility to primary hepatocellular carcinoma by case control study by polymerase chain reaction restriction fragment length polymorphism(CR-RFLP).The research emphasized the relationship of genetic site SNP and hereditary susceptibility of primary hepatocellular carcinoma.The major aims of our research were the relationship between genetic polymorphisms and susceptibility to hepatocellular carcinoma and the interaction of genetic polymorphisms and hepatitis infection in susceptibility to hepatocellular carcinoma.The research provided evidence on confirming the high risk population of hepatocellular carcinoma.Material and MethodThe experimental subjects composed of 100 cases with primary hepatocellular carcinoma and 111 healthy controls in Liaoning Province.All the cases were patients hospitalized in intervention department of the first affiliated hospital of China Medical University.,with the diagnosis of primary hepatocellular carcinoma according to the clinical diagnostic standard of primary hepatocellular carcinoma approved on the 8th Chinese hepatocelluar carcinoma academic conference in September,2001.Every case had pathologic evidence or increased value of AFP,and all of them had never received radiochemotherapy.Participants in the control group came from population undertaking healthy physical examination in the physical examination center in the same hospital without hereditary history or history of tumor.The normal control were matched by 1:1 according to age(5 years),sex and case frequency.The demography material and related risk factor of each subject were collected while collecting blood samples.The polymorphism genetic type of Lys751Gln of XPD gene and Arg194Trp, Arg280His,Arg399Gls of XRCC1 gene were analyzed by PCR-RFLP.DNA were distilled from anticoagulated blood by potassium iodide.PCR amplified polymorphisms of XPD-751,XRCC1-194,XRCC1-280 and XRCC1-399.The designed primer of each gene was as follows,XPD 751: 5'-GCCCGCTCTGGATTATACG-3′and 5′-CTATCATCTCCTGGCCCCC-3′; XRCC1-194:5' GCC AGG GCC CCT CCT TCA A 3' and 5' TAC CCT CAG ACC CAC GAG T 3';XRCC1-399:5' TTG TGC TTT CTC TGT GTC CA3' and 5' TCC TCC AGC CTT TTC TGA TA 3';XRCC1-280:5'CCA GTG GTG CTA ACC TAA TC3' and 5'CAC TCA GCA CCA CTA CCA CA3'.The PCR reaction condition of XRCC1-194 and XRCC1-399 were 2min predenaturation at 94℃,then 30s at 94℃, 30s at 57℃,45s at 72℃with 30 circulations as above and 7min extension at last.The condition of XRCC1-280 was the same except for changing 57℃to 59℃in the 30 circulations.Electrophoresis analyzed products of PCR.The products were incised by enzyme.The incision enzyme were PstⅠfor XPD-751,RsaⅠfor XRCC1-280,MspⅠfor XRCC1-399 and PvuⅡfor XRCC1-194.5μl PCR product was digested with corresponding restrictive endoneuclease over night.The enzymatic incised products were analyzed by 1.5%agarose gel electrophoresis.SPSS12.0 was used for Statistic analysis.The distribution differences of polymorphisms of XPD751,XRCC1-194,XRCC1-280,XCRR1-399 and related risk between the case group and control group were analyzed by chi-square test.The relationship of polymorphism genotype with risk of hepatocellular carcinoma and interaction with exposed risk factor were analyzed by Logistic regression.Results1.The frequence of polymorphism genotype of Lys/Lys,Lys/Gln,Gln/Gln of XPD751 gene in patients with hepatocellular carcinoma were 76(76.00%),23(23.00%)和1(1.00%)respectively,which in the control group were 88.28%,9.91%and 1.8%. The difference were highly significant between the groups(p=0.033).2.It was observed by Logistic regression analysis that the risk of suffering hepatocellular carcinoma increased significantly in individuals carrying at least one allele of 751Gln(Lys/Gln和Gln/Gln genotype)(OR=2.38,95%CI was 1.14-4.98, p=0.019).3.Interaction existed between Lys/Gln,Gln/Gln genotype of XPD751 gene and hepatitis B infection.The risk of suffering hepatocellular carcinoma in individuals with hepatitis B infection carrying at least a allele of 751Gln was 123.75 fold increased than that without both hepatitis B infection and Gln gene.(95%CI was15.56-984.00).4.The frequence of polymorphism genotype Arg/Arg,Trp/Trp of XRCC1-194 in patients with hepatocellular carcinoma were 46.00%,50.00%and 4.00%respectively, while that in the healthy group were 51.35%,,38.74%and 9.91%respectively.The comparison had no significant difference(p=0.110).5.The risk of suffering hepatocellular carcinoma did not have significant increase between individuals carrying Arg/Trp and Trp/Trp genotype by Logistic regression analysis(OR=1.24;95%CIwas 0.71-2.13,p=0.437).6.The frequence of polymorphism genotype Arg/Arg,Trp/His of XRCC1-280 in patients with hepatocellular carcinoma were 79.00%,20.00%and 1.00%respectively, while that in the healthy group were78.38%,18.92%and 2.7%respectively.The comparison had not significant difference(p=0.912).7.The risk of suffering hepatocellular carcinoma did not have significant increase between individuals carrying Arg/His and His/His genotype by Logistic regression ananlysis(OR=0.096;95%CI was 0.50-1.86,p=0.912).8.The frequence of polymorphism genotype Arg/Arg,Trp/Gln,Gln/Gln of XRCC1-399 in patients with hepatocellular carcinoma were 40.00%,53.00%and 7.00%respectively,while that in the healthy group were62.16%,27.93%and 9.91% respectively.The comparison had significant difference(p=0.001).9.Interaction existed between Arg/Gln,Gln/Gln genotype of XPD399 gene and hepatitis B infection.The risk of suffering hepatocellular carcinoma in individuals with hepatitis B infection carrying at least a allele of 399Gln was 170.76 fold increased than that without both hepatitis B infection and Gln gene.(95%CI was 40.02-717.03).Conclusion(1)XPD gene Polymorphism of Lys751Gln was a hereditary susceptible factor of hepatocellular carcinoma,and 751Gln gene was a risk gene for hepatocellular carcinoma.(2)XPD gene Interaction existed in hepatocellular carcinoma occurrence between polymorphism of Lys751Gln and exposed risk factor of hepatocellular carcinoma.(3)XRCC1 gene Polymorphism of Arg399Gln was a hereditary susceptible factor of hepatocellular carcinoma,and 399Gln gene was a risk gene for hepatocellular carcinoma.(4)XRCC1 gene Interaction existed in hepatocellular carcinoma occurrence between polymorphism of Arg399Gln and exposed risk factor of hepatocellular carcinoma.(5)XRCC1 gene Polymorphism of Arg194/Trp,Arg280/His gene had no relationship with hereditary susceptibility of hepatocellular carcinoma.