Association Between DNA Double Strand Break Repair Gene Polymorphisms And Risk Of Glioma In A Chinese Population

Background and Objectives:Brain tumor is one of serious tumor which threat to human health and life, the incidence of brain tumor shows a rising trend in recent years. Gliomas also called neuroepithelial tumor, neuroectodermal tumor, which originated in the neural ectoderm. In the countries with high incidence of gliomas, nervous system tumor mortality rate can be as high as 7/10 million. As WHO reported in 1998andmonitoring results of the cause of death in some countries, mortality rate of gliomas ranking the first in total mortality of children under 10 years old, ranking the second in tumor death of people under 34 years old, ranking the third in total mortality of people 35-54 years old. The prevalence rate of gliomas was different between people with different race,gender,age and occupation. Previous studies found that the risk factor of gliomas including behavior and life style, physical and chemical factors,biological and virus factors,and genetic factors. Excessive consumption of cured foods contain N-nitroso compounds and its precursors could increase the risk of gliomaS,consumption of fresh fruits and vegetables could reduce the risk of glioma;exposure of chemical composition of environment is a certain risk of glioma,in particular, people exposure to chemicals,whose children shows significantly increase risk of gliomas; X-ray exposure by large dose and many times is a risk factor for glioma pathogenesis, but diagnostic dose of X-ray exposure showed no increased risk; Simian virus 40 (SV40)andpolyomavirus(JCV)is correlated with glioma; otherwise, immune state, infection disease history, family history of gliomas are also associated with glioma. Although people exposed to the same risk factors, only a few suffering from gliomas. Above facts suggested that Whether the individual suffering from gliomas is determined by a genetic susceptibility to environmental factors. The differences of susceptible of disease is derived from the differences of DNA sequence. So the research object on susceptible of glioma gradually turned to the research of single nucleotide polymorphism(SNPs). Single nucleotide polymorphism is the phenomenon that occur in the same kind of organisms but different individuals, of whom, the same DNA allele shows different sequence of single nucleotide. Some functional SNPs could influence the gene expression,thus presents the heredity difference. There are several ways of DNA repair in mammals, including base excision repair, nucleotide excision repair, mismatch repair, DNA double strand break repair, direct repair, etc. The influence of genetic factors in the repair system will cause a corresponding change in repair ability,cause DNA damage, gene mutation and cell canceration. DNA double strand break damage is one of the most serious form of DNA damage, which was difficult to repair, caused chromosome breakage and cell death, was the basis of chromosome abnormality and tumor susceptibility. Double strand breakrepair pathwayincluding non homologous end joining(NHEJ) and homologous recombination (HR), which could repair DNA damage collaboratively. NHEJ repair pathway does not require intact DNA template in the repair process, which connected the broken ends of fracture of DNA directly. NHEJ repair pathway is the main repair pathway in mammalian cells for DNA double strand breaks. HR repair pathway needs homologous complete DNA as template to repair, which plays an important role in the double strand break repair as its accuracy of repair. There were some reports concerning the polymorphism of DSBR gene and cancer susceptibility, but researches concerning relationship between polymorphism of DSBR gene and glioma were less. The main purpose of this study is to understand the demographic characteristics, family history, environmental exposure, life style and other factors on the impact of glioma pathogenesis,to found out the differences in the distribution of alleles and genotypes of LIG4 rs3093737, LIG4 rs3093739; XRCC4 rs131616625 XRCC4 rs2731858, XRCC4 rs7715771; XRCC5 rs828704, XRCC5 rs3770502, XRCC5 rs9288516;XRCC6 rs6519265, XRCC6 rs132793;XRCC3 rs3212092, XRCC3 rs861530, RAD54L rs1048771, NBS1 rs1805794, XRCC7 rs10109984 between cases and control group,to analyse the influence of LIG4 rs3093737, LIG4 rs3093739;XRCC4 rs13161662, XRCC4 rs2731858, XRCC4 rs7715771; XRCC5 rs828704, XRCC5 rs3770502, XRCC5 rs9288516;XRCC6 rs6519265, XRCC6 rs132793;XRCC3 rs3212092, XRCC3 rs861530, RAD54L rs1048771, NBS1 rs1805794XRCC7 rs10109984 on gliomas susceptibility after demographic characteristics, family history, environmental exposure factors were adjusted,to provide valuable reference for glioma screening or early diagnosis and treatment.Materials and MethodsThis study was conducted by frequency matched case-control study.224 gliomas patients who have been histologically diagnosed were included as case group from Novembre 2010 to December 2013.controls were selected from 200 people who received health examination with no history of cancer and neurological disease,had not received chemotherapy,had no blood relationship with case group. Controls were frequency matched by gender and age with case group. Self-designed questionnaire was used to collect the general demographic characteristics of patients, including name, gender, age, occupation, marital status, place of residence,economiccondition, medical security, family history, smoking, cured products consumption, X ray exposure history and other information.5 ml venous blood wes collected by EDTA-Na2 anticoagulant blood sampling from all the subjects, which were kept in the refrigerator at-20℃. Blood genome were extracted from blood samples according to blood genome extraction kit(DP319)instructions. Detection of gene polymorphism was conducted by Taqman probe method. SNP site were selected on the basis of other reference which were consider with highly tumor susceptibility or controversial.25 L PCR reaction system was used for DNA amplify in real time fluorescent quantitative PCR instrument, survey database was established by Epidata3.0 software, and statistical analysis was performedbySPSS17.0. Descriptive statistics analysis was used to difference of demographic characteristics, family history of cancer, behavioral risk factors, environmentrisk factors exposure between case group and controls. Hardy-Weinberg balance test was conducted to realize the frequency distribution of SNP locus in the control group. The difference of distribution of target alleles and genotypes were clarified between case group and controls. Single factor non-conditional logistic regression and multivariate non-conditional logistic regression analysis were conducted respectively to find the association between those candidate SNP sites and gliomas susceptible.Resultspart 1:(1)The degree of education of case group were mainly primary school and below(96 people, account for 42.86%),in the next place were junior middle school(66 people, account for 29.02%) and senior middle school or secondary specialized school(41 people, account for 18.30%),22 people were University or college degree or above,accounted for 9.82%.113 people live in city,accounting for 50.89%, 110people live in the rural,accounte for 49.11%.33 people were single of marital status,accounted for 14.73%,married or cohabiting people with a higher proportion of 73.66%(165 people),26 people were divorced or widowed accounted for 11.61%.The occupation of case group were mainly farmers(92 people, account for 41.07%),workers were accounted for 36.61%(82 people),21 people were cadre(9.38%),29 people engaged in other occupation (12.95%). In controls, the degree of education were mainly primary school and below(74 people, account for 37.00%),in the next place were junior middle school(70 people, account for 35.00%) and senior middle school or secondary specialized school(31 people, account for 15.50%),25 people were Universityorcollege degree or above,accounted for 12.50%. 110 people live in city,accounting for 55.00%,90 people live in the rural,accounte for 45.00%.25 people were single of marital status,accounted for 12.50%,married or cohabiting people with a higher proportion of 79.00%(158 people),17 people were divorced or widowed accounted for 8.50%.The occupation of controls were mainly workers(70 people, account for 35.00%),farmers were accounted for 39.50%(79people),29 people were cadre(14.50%),22 people engaged in other occupation (11.00%). There were no differences between case group and control group in those demographic characteristics(P>0.05).(2)The case group with family history of cancer was 23.66%, which was 14.00% in control group, there was statistical differences between case and control(χ2=6.381,P<0.05). The case group of drinkers accounted for 45.98%, smokers accounted for 29.91%, people who like eating cured meat products accounted for 61.16%, occupational x-ray expose accounted for 4.91%, occupational chemical contact accounted for 4.91%.The control group of drinkers accounted for 39.00%, smokers accounted for 28.50%, people who like eating cured meat products accounted for 59.50%, occupational x-ray expose accounted for 4.00%, occupational chemical contact accounted for 5.00%,there were no statistical differences between case and control of those factors.(3) Hardy-Weinberg balance test results shows that the candidate gene SNPs of NHEJ pathway were conforms to Hardy-Weinberg law of genetic equilibrium(p value was 0.633、0.924、0.732、0.057、0.245、0.431、0.095、0.916、0.876、0.993% 0.900 respectively).The distribution of genotype frequency in this study could represent the region Chinese Han population.(4)The allele frequency distribution of rs3093739 of LIG4 gene, rs13161662 of XRCC4 gene, rs2731858 of XRCC4 gene and rs9288516 of XRCC5 gene were different between case and control gruop (P<0.05). The allele frequency distribution of other selected gene loci of LIG4 gene, XRCC4 gene, XRCC5 gene and all the selected gene loci of XRCC6, XRCC7 were no statistical difference(P>0.05).(5)The genotype frequency distribution of rs3093739 of LIG4 gene, rs13161662 of XRCC4 gene, rs2731858 of XRCC4 gene, rs828704 and rs9288516 of XRCC5 gene were different between case and control gruop (P<0.05). The genotype frequency distribution of other selected gene loci of LIG4 gene, XRCC4 gene, XRCC5 gene and all the selected gene loci of XRCC6, XRCC7 were no statistical difference(P>0.05)(6)Single factor uncondition logistic regression was used to anlyse the influence of candidate gene loci of NHEJ pathway on glioma susceptibility,the result shows that,compared with TT gene type, TC and CC gene type of LIG4 rs3093739 may increase risk of illness (OR=1.544, OR95%C.I.=1.070-2.229);compared with AA gene type, AG and GG gene type of XRCC4 rs13161662 may reduce risk of illness (OR=0.536,OR95%C.I.= 0.377-0.762);compared with GGgene type, GA and AA gene type of XRCC4 rs2731858 may increase risk of illness (OR=1.365,OR95%C.I.= 1.004-1.856);compared with TT gene type, TC and CC gene type of XRCC5 rs9288516 may increase risk of illness (OR=1.450,OR95%C.I= 1.106-1.903).(7) Multi factor uncondition logistic regression was used to anlyse the influence of candidate gene loci of NHEJ pathway on glioma susceptibility, demographic characteristics, family history, unhealthy life style, and environmental exposure were adjusted as the confounding factors.The result shows that,compared with TT gene type, TC and CC gene type of LIG4 rs3093739 may increase risk of illness (OR=1.556,OR95%C.I.= 1.050-2.305);compared with AA gene type, AG and GG gene type of XRCC4 rs13161662 may reduce risk of illness (OR=0.505,OR95%C.I.= 0.346-0.736);compared with AA gene type, AC and CC gene type of XRCC5 rs828704 may reduce risk of illness (OR=0.698,OR95%C.I.= 0.478-0.920);compared with TT gene type, TC and CC gene type of XRCC5 rs9288516 may increase risk of illness (OR=1.413,OR95%C.I.= 1.071-1.913).part 2:(1) Hardy-Weinberg balance test results shows that the candidate gene SNPs of HR pathway were conforms to Hardy-Weinberg law of genetic equilibrium(p value was 0.214,0.268,0.651,0.057 respectively).The distribution of genotype frequency in this study could represent the region Chinese Han population.(2)The allele frequency distribution of rs3212092 of XRCC3 gene, rs861530 of XRCC3 gene, rs1048771 of RAD54L gene were different between case and control gruop (P<0.05). The allele frequency distribution of all the selected gene loci of NBS1 were no statistical difference(P>0.05).(3)The genotype frequency distribution of rs3212092 of XRCC3 gene, rs861530 of XRCC3 gene, rs 1048771 of RAD54L gene were different between case and control gruop (P< 0.05). The genotype frequency distribution of all the selected gene loci of NBS1 were no statistical difference(P>0.05).(4)Single factor uncondition logistic regression was used to anlyse the influence of candidate gene loci of HR pathway on glioma susceptibility,the result shows that,compared with CC gene type, CT and TT gene type of XRCC3 rs3212092 may increase risk of illness (OR=1.774, OR95%C.I.=1.106-2.847);compared with AA gene type, AG and GG gene type of XRCC3 rs861530 may increase risk of illness (OR=1.420,OR95%C.I.= 1.083-1.862);compared with CC gene type, CT and TT gene type of RAD54L rslO48771 may increase risk of illness (OR=1.495,OR95%C.I.= 1.144-1.954).(5) Multi factor uncondition logistic regression was used to anlyse the influence of candidate gene loci of HR pathway on glioma susceptibility, demographic characteristics, family history, unhealthy life style, and environmental exposure were adjusted as the confounding factors.The result shows that,compared with CC gene type, CT and TT gene type of XRCC3 rs3212092 may increase risk of illness (OR=1.790, OR95%C.I.= 1.095-2.928);compared with AA gene type, AG and GG gene type of XRCC3 rs861530 may increase risk of illness (OR=1.401,OR95%C.I.= 1.053-1.864).Conclusions:(1)The candidate gene SNPs of this study were conforms to Hardy-Weinberg law of genetic equilibrium, the distribution of genotype frequency in this study could represent the region Chinese Han population.(2) There weresignificant statistical differences between case and control on family history of cancer and no statistical differences between case and control on other factors as drinking,smoking,eating cured meat, occupational x-ray expose and occupational chemical contact.(3) The allele and genotype frequency distribution of rs3093739 of LIG4 gene, rs13161662 of XRCC4 gene, rs2731858 of XRCC4 gene and rs9288516 of XRCC5 gene were different between case and control group in NHEJ pathway. The allele and genotype frequency distribution of rs3212092 of XRCC3 gene, rs861530 of XRCC3 gene, rs 1048771 of RAD54L gene were different between case and control gruop (P< 0.05)in HR pathway.(4) Single factor uncondition logistic regression analysis shows that there was relevance between LIG4 rs3093739、XRCC4 rs13161662、XRCC4 rs2731858、 XRCC5 rs9288516 and gliomas in NHEJ pathway,and there was relevance between XRCC3 rs3212092、XRCC3 rs861530、RAD54L rs1048771 and gliomas in HR pathway.(5) Demographic characteristics, family history, unhealthy life style, and environmental exposure were adjusted as the confounding factors,multi factoruncondition logistic regression shows that LIG4 rs3093739(OR=1.556,OR95% C.I.= 1.050-2.305),XRCC4 rs13161662(OR=0.505,OR95% C.I.= 0.346-0.736), XRCC5 rs828704(OR=0.698,OR95%C.I.= 0.478-0.920), XRCC5 rs9288516(OR=1.413,OR95%C.I.= 1.071-1.913) in NHEJ pathway and XRCC3 rs3212092(OR=1.790,OR95%C.I.= 1.095-2.928), XRCC3 rs861530(OR=1.401,OR95%C.I.= 1.053-1.864)in HR pathway has relationship with gliomas.