| Hepatocellular carcinoma (HCC) is a common one of highly malignant tumor. It is estimated that the number of new cases worldwide is 60 million and deaths from liver cancer to 58 million each year. China is one of the areas of high incidence of liver cancer, 55% of the world occurred in China. Guangxi is the high incidence area of HCC in China, which accounts for the county Fusui of all malignant as high as 61.51%, topping its tumor death cause list. HCC features as high malignant, long dormancy, lurk onset, rapid progress, short course, and high mortality rate, and almost 100% case fatality rate, posing an overwhelmingly threat to human health. As far as the current medical treatment feasibility is concerned, few can definitely changes the course and the end of HCC. So the investigation into the etiology and susceptibility is of great significance.The occurrence of HCC is the synergetic results of both heredity and environment elements. Previous studies indicate that the main environment risk factors of HCC include chronic HBV or HCV infection, AFB1 in food, smoking and drinking. Of chronic HBV infection is an important cause of HCC in China and more than 90% of patients with HCC associated with HBV infection. This suggests that the body's scavenging ability of HBV are individual differences, which means that hosts are infected with HBV disease progression and the outcome after large extent determined by their antiviral immunity (or inflammation) related gene polymorphism. With the increase understanding of pathogenesis of host immune response against HBV, the present greater attention to the immune response of cytokine genes, polymorphisms may be a major impact of their course and outcome of HBV infection.Guangxi is the high incidence area of HCC in China, but also the largest gathering of Zhuang people . The incidence of liver cancer racial differences, need to further clarify the genetic level. In some pathogenic gene polymorphism studies, first of all to understand their distribution in the normal population is characterized by correct analysis of the relationship between gene polymorphisms and disease prerequisite. Because of geography, population and other limitations, the current Guangxi Zhuang and Han populations in HCC-related cytokine gene polymorphisms are very few.A large number of studies show that TNF-α, interleukin-1B, IL-12B, transforming growth factorβ1 is closely related to the development of HCC. In order to verify the above assumptions, we have launched a hospital-based molecular epidemiological study.There are two parts of the study. Firstly, to obtain the Guangxi Zhuang and Han populations corresponding population genetics data, we analysis gene polymorphism of TNF-α, interleukin-1B, IL-12B, transforming growth factorβ1,which associated with HCC, with 200 cases of Zhuang and 232 cases of Han. Secondly, aiming at approaching their relations with HCC, we carried through a study on based-hospital molecular Epidemiology and selected 772 cases of HCC patients as case group, 852 cases of non-cancer patients as control group.Objectives To approach the distribution frequency and characteristics in Guangxi Zhuang and Han populations , we analysis gene polymorphism of TNF-α, interleukin-1B, IL-12B, transforming growth factorβ1 ,which associated with HCC.Methods The research was designed as A hospital-based epidemiological survey methods. From June 2007 to June 2010, we randomly selected hospital trauma treatment ,whose parents and grandparents were living in the Guangxi, 200 of Zhuang , 232 of Han. All of the cases from the First Affiliated Hospital of Guangxi Medical University and the First Affiliated Hospital of Guangxi Traditional Chinese Medicine University, 303 Hospital of Guangxi, first People's Hospital of Liuzhou, and no blood relationship between them. All subjects are rule out liver, kidney, endocrine, and cardiovascular and cerebrovascular diseases, no history and clinical features of the tumor, without genetic diseases and infectious diseases. All subjects were surveyed face to face by the specifically trained investigators. The questionaire features include basic datum, disease history, personal history, family history, smoking history, and drinking history etc. 2ml fasting venous blood were collected at the second day morning, and genome DNA were extracted at the same day. Genotypes of TNF-αrs1799964,TNF-αrs1800629,TNF-αrs1800630,IL-1B rs1143623,IL-1B rs1143627,IL-1B rs16944,IL-12B rs3212227,TGFβ1 rs1800469 were detected using Applied Biosystems TaqMan genotyping platform according to the manufacture's recommendations. All data were input and analyzed statistically in SPSS 13.0 for Windows. Student's t-test was used to compare mean value of the samples. While categorical data were compared with the use of the chi-square test or the Fisher's exact propability. Odd ratio and 95% confidence interval indicate relative risk scale, and were analyzed with the non-conditional logistic regress model. Shesis software was applied to test Hardy-Weinberg genetic balance, haplotype frequency and linkage disequilibrium.Results (1) Demographic characteristics of the object: There were no statistically differences in the distributions of age and sex between Zhuang and Han groups (P>0.05), respectively.(2) Hardy-Weinberg equilibrium tests: SHEsis software was applied to perform Hardy-Weinberg equilibrium tests, the results showed that all genotypes of the observations fit well with the theoretical value (P> 0.05), showed that samples of this experiment with a group representative.(3) Cytokine gene polymorphisms of the genotypes and allele frequency: The genotype analysis showed that allele mutation frequency of gene sites of TNF-αrs1799964,TNF-αrs1800629,TNF-αrs1800630,IL-1B rs1143623,IL-1B rs1143627,IL-1B rs16944,IL-12B rs3212227 in the Zhuang group were 20.20%,9.80%,20.50%,45.80%,49.00%,50.20%,47.20%, respectively, with no statistically significant difference with the Han group (21.60%,9.10%,22.40%,45.90%,49.10%,48.30%,47.40%)(all P>0.05). Also their genotype distribution showed no statistic difference. The allele of TGFβ1 rs1800469 sites of C and T were 67.20% and 32.80% in the Zhuang group, as compared with 59.30% and 40.70% in the Han; the distribution frequency of TGFβ1 rs1800469 sites of CC, CT,TT were 44.00%,46.50%,9.50%,in the Zhuang group, respectively, as compared with 36.21%,46.12%,17.67% in the Han, all with a statistical significance (P<0.05).(4) TNF-αand IL-1B polymorphisms in linkage disequilibrium test: SHEsis software was applied to perform linkage disequilibrium test between TNF-α3 SNP sites and between IL-1B 3 SNP sites,the results showed these sites exist in strong linkage disequilibrium in Zhuang group and Han group , respectively.(5) TNF-αand IL-1B haplotypes analysis: SHEsis software was applied to perform haplotyping analysis. The results showed that CAG,TCA and TCG were the most common haplotypes in TNF-αrs1799964,TNF-αrs1800630,TNF-αrs1800629 among Zhuang and Han, and CCT,GCT and GTC were the most common haplotypes in IL-1B rs1143623,IL-1B rs16944,IL-1B rs1143627 among the two groups, but with no statistical difference in the rates between the two groups(P<0.05).Conclusion (1) There were significant differences in the frequency distribution of TGFβ1 rs1800469 locus C and T allele in the Zhuang and Han groups. And the same result exited the frequency distribution of TGFβ1 rs1800469 (P <0.05). Subjects carrying TGFβ1 rs1800469 loci homozygous wild genotype CC is more in Zhuang group, and carry rs1800469 TT homozygous mutation genotype more in Han group.(2) There were strong linkage disequilibrium between TNF-αrs1799964,TNF-αrs1800630 and TNF-αrs1800629. The CAG,TCA and TCG were the most common haplotypes in both Zhuang and Han groups. And there were also strong linkage disequilibrium between IL-1B rs1143623,IL-1B rs16944 and IL-1B rs1143627. The CCT,GCT and GTC were the most common haplotypes in two groups, but with no statistical difference in the rates between the two groups. Objectives To study the susceptibility of cytokines TNF-α, interleukin-1B, IL-12B, transforming growth factorβ1 gene polymorphism with primary hepatocellular carcinoma in Guangxi population.Methods The research was designed as a case-control study based on hospital. From June 2007 to June 2010, 772 newly histopathologically diagnosed HCC patients from First Affiliated Hospital of Guangxi Medical University and the First Affiliated Hospital of Guangxi Traditional Chinese Medicine University, 303 Hospital of Guangxi, first People's Hospital of Liuzhou were enrolled as the case group who had never received chemotherapy and radiotherapy and had signed the study consensus protocol, with 852 cases of non-cancer patients in the Spinal Orthopedic Department,Traumatic Department,Aesthetic Plastic Surgery and Ophthalmology as the control, with the case and the control matched in terms of ages, sex, and habitation. All subjects were surveyed face to face by the specifically trained investigators. The questionaire features include basic datum, disease history, personal history, family history, smoking history, and drinking history etc. 2ml fasting venous blood were collected at the second day morning, and genome DNA were extracted at the same day. 8 SNP loci of TNF-αrs1799964,TNF-αrs1800629,TNF-αrs1800630,IL-1B rs1143623,IL-1B rs1143627,IL-1B rs16944,IL-12B rs3212227, respectively, were measured with high flux TaqMan MGB, a fluorescent quantitative real time PCR method. All data were input and analyzed statistically in SPSS 13.0 for Windows. Student's t-test was used to compare mean value of the samples. While categorical data were compared with the use of the chi-square test or the Fisher's exact propability. Odd ratio and 95% confidence interval indicate relative risk scale, and were analyzed with the non-conditional logistic regress model. Shesis software was applied to test Hardy-Weinberg genetic balance, haplotype frequency and linkage disequilibrium.Results (1) Demographic characteristics of the object: There were no statistically differences in the distributions of age, sex, and nationality between cases and controls (P>0.05), respectively.(2) The clinical data of subjects and dietary behavior: The results showed that there were no significant difference in HCVAb distributed among the case and control group (P> 0.05).And there were statistically significant differences between cases and controls in the distribution of frequencies with hepatitis B history, consumption of fish health, liver cancer family history, smoking history, drinking history, HBsAg, HBsAb, HBeAg, HBeAb, HBcAb, etc(P <0.05).(3) Hardy-Weinberg equilibrium tests: The results showed that all genotypes of the observations fit well with the theoretical value (P> 0.05), showed that samples of this experiment with a group representative.(3) Cytokine gene polymorphisms of the genotypes and allele frequency: The genotype analysis showed that allele mutation frequency of gene sites of TNF-αrs1799964,TNF-αrs1800629,TNF-αrs1800630,IL-1B rs1143623,IL-1B rs1143627,IL-1B rs16944,IL-12B rs3212227,TGFβ1 rs1800469 in the case group were 21.50%,10.43%,20.60%,45.01%,49.09%,49.16%,45.79% and 37.44%,respectively, with no statistically significant difference with the control group (20.25%,10.45%,18.19%,45.83%,49.00%,49.47%,47.83%,36.38%)(all P>0.05). Also their genotype distribution showed no statistic difference. The distribution frequency of TNF-αrs1800630 sites of CC,CA,AA were 65.67%,27.46% and 6.87% in the case group, respectively, as compared with67.02%,29.58% and 3.40% in the control, all with a statistical significance (P<0.05). HCC risk in patients with AA genotype was 2.058 times that of CC, with a significant difference(P<0.05), indicating genotype of rs1800630 AA might be correlated statistically with the onset of HCC.(4) Stratification analysis showed that after stratified with drinking, in subgroup of without drinking, HCC risk of group of TNF-αrs1800630 A allele were 1.839 times that of TNF-αrs1800630 C (0.010);After stratified with history of hepatitis, smoking and HBsAg positive, no risk increasing genotype was found.(5)TNF-αand IL-1B polymorphisms in linkage disequilibrium test: SHEsis software was applied to perform linkage disequilibrium test between TNF-α3 SNP sites and between IL-1B 3 SNP sites,the results showed these sites exist in strong linkage disequilibrium in control group .(6) TNF-αand IL-1B haplotypes analysis: SHEsis software was applied to perform haplotyping analysis. The results showed that CAG,CCG,TCA and TCG,were the most common haplotypes in TNF-αrs1799964,TNF-αrs1800630,TNF-αrs1800629 among study and control groups, but with no statistical difference in the rates between the two groups(P<0.05). And CCT,CTT,GCC,GCT and GTC were the most common haplotypes in IL-1B rs1143623,IL-1B rs16944,IL-1B rs1143627 among the two groups. The aplotypes of GCC and CTT were the anti-risk haplotypes from HCC, the distribution of case group was 3.00% and 1.90%, as compared with 6.50% and 5.50% in the case group, all with a statistical significance (P = 3.73 e-006 and 8.02e-008). HCC risk in patients with CTT and GCC haplotypes were 0.446 times and 0.332 times that of CCT, GCT and GTC haplotypes. The GTC is the high risk haplotype, its frequency distribution of case group was 46.70%,and 43.10% in the control group, there are statistically significant differences in the distribution among the case and control groups (P = 0.034), HCC risk in patients with GTC haplotype were 1.162 times that of others.Conclusion (1) TNF-αrs1800630 AA genotype is the risk allele of HCC in Guangxi. In subgroup of drinking, HCC risk of group of TNF-αrs1800630 A genotype were 1.839 times that of TNF-αrs1800630 C.(2) Family history of HCC, history of hepatitis, raw fish eating, HBsAg positive, drinking, smoking and TNF-αrs1800630 AA genotype carrier were HCC risk factors in Guangxi.(3) Linkage disequilibrium exists in polymorphism sites of TNF-αrs1799964,TNF-αrs1800630,TNF-αrs1800629, with CAG,CCG,TCA and TCG as the most common haplotype, but no significant difference in all estimated haplotype frequency distribution between the case and the control. CCT,CTT,GCC,GCT and GTC were the most common haplotypes in IL-1B rs1143623,IL-1B rs16944,IL-1B rs1143627 among the two groups. The aplotypes of GCC and CTT were the anti-risk haplotypes from HCC, HCC risk in patients with CTT and GCC haplotypes were 0.446 times and 0.332 times that of CCT, GCT and GTC haplotypes. The GTC is the high risk haplotype, HCC risk in patients with GTC haplotype were 1.162 times that of others. |
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