| CHAPTER ⅠAssociation between Glycoprotein CD44 Gene Polymorphism and Risk of Susceptibility to Hepatocellular CarcinomaObjective: CD44 is a widely distributed and with a high degree of heterogeneity single-chain transmembrane glycoprotein adhesion molecules, playing an important role in tumor cell proliferation, differentiation, invasion and metastasis. The study detected genotype and allele frequencies distribution of CD44 gene rs8193, rs10836347 and rs13347 to explore the association between CD44 gene rs8193, rs10836347 and rs13347 polymorphism and the risk of HCC.Method: A total of 204 HCC and 210 controls were included in this study, restriction fragment length polymorphism and DNA sequencing were used to detect the polymorphism of CD44 gene rs8193, rs10836347 and rs13347. The χ2 test was used to analyze whether the distribution of the SNP genotype frequencies in HCC group and the healthy controls group consistent with Hardy-Weinberg equilibrium(HWE). Binary logstic regression adjusted for sex, age, smoking and drinking four factors were used to calculate the odds ratio(OR) and 95% confidence intervals(CI) between the genetic polymorphisms and the risk of HCC.Results:1. The genotype frequencies in the HCC group and controls group of the three SNPs of CD44 gene were found to be in HWE(all P>0.05), indicating that the sample selected had group representation.2. The study found that the rs8193 had CC, CT and TT three genotypes. Using the CC genotype as a reference genotype, the CT and TT genotype were not associated with HCC risk(all P>0.05). Using the C allele as a reference genotype, T allele was not associated with HCC risk(P>0.05). Both in a dominant model and recessive model analysis, not genotype was found to be associated with the risk of HCC(all P>0.05).3. The study found that rs10836347 had CC and CT genotypes. Using the CC genotype as a reference genotype, TT genotype were not associated with HCC risk(P=0.253). Using the C allele as a reference genotype, T allele was not associated with HCC risk(P=0.271).4. The study found that rs13347 had CC, CT and TT three genotypes. Using the CC genotype as a reference genotype, patients with CT genotype showed 1.626-fold(95% CI: 1.057-2.500, P=0.027) higher risks of HCC. Patients carrying TT genotype showed 1.965-fold(95% CI: 1.043-3.702, P=0.037) higher risks of HCC. Using C allele as reference genotype, T allele showed 1.461-fold(95% CI: 1.091-1.956, P=0.011) higher risks of HCC. In the dominant model analysis, CT+TT compared to CC genotype showed 1.697-fold(95% CI: 1.13-2.549, P=0.011) higher risks of HCC. However, in the recessive model analysis, it has found no correlation between the polymorphisms and the risk of HCC(P=0.549).5. Stratified analysis according to the risk factors of age, sex, smoking and alcohol consumption, in women subgroup, patients with CD44 rs13347 CT genotype exhibited 2.675-fold(95% CI: 1.004-7.132, P=0.049) and TT genotype exhibited 5.922-fold(95% CI: 1.083-33.164, P=0.040) higher risks of HCC than those of patients with the CC genotype. In age≥50 years subgroup, patients with CD44 rs13347 CT genotype exhibited 2.157 fold(95% CI: 1.113-4.181, P=0.023) and TT exhibited 3.405-fold(95% CI: 1.263-9.178, P=0.015) higher risks of HCC than those of patients with the CC genotype. In drinking subgroup, for the rs10836347 site, CT genotype significantly increased the risk of HCC to 4.552-fold(95% CI: 1.205-17.192, P=0.025). For the rs13347 site, using the CC genotype as reference, CT genotype was no associated with the risk of HCC(P=0.179), patients with CD44 rs13347 TT genotype significantly increased the risk of HCC to 4.549-fold(95% CI: 1.244-16.635, P=0.022);6. The haplotype distributions of CD44 rs8193, rs10836347 and rs13347 were evaluated. The TTT haplotype significantly increased the risk of HCC(OR=207.16, P=0.014), indicating that TTT haplotype maybe a risk for HCC.Conclusion: The polymorphism of CD44 gene rs8193, rs10836347 was not associated with the risk of HCC. T mutant allele of rs13347 may increase the risk of HCC.CHAPTER Ⅱ Comparative serum glycoprotein of Hepatocellular Carcinoma with different sexObjective: Glycosylation is one of the most common protein posttranslational modification. The same protein with different sugar chain structure may present different function. The incidence of HCC in male is higher than that in female, while survival in female is better than that in male. In this part, we use lectin microarray technology to detect the serum glycoprotein spectrum between male and female in the process of HCC. The aim of this study is to find its characteristic changes of glycoprotein spectrum and the different sugar spectrum signs between male and female in the process of HCC, providing new ideas and experimental basis for the study of difference in the incidence of HCC between male and female.Method: Experiment process was as followed(1) low-abundance serum proteins were collected: Mixing the equal volume serum of four groups(control group of male, HCC group of male, control group of female and HCC group of female), low-abundance protein was collected using the high-abundance protein removal kits.(2) After desalting and ultrafiltration, detect the concentration of the low abundance proteins.(3) The target proteins were labeled with Cy5 using Lightning-Link kits.(4) Prepare the lectin microarray and hybridization.(5)Lectin microarray was scaned and data was collected.(6) Data was analysis by the SPSS 13.0, two independent samples t test or non-parametric test was used.(7) Lectin immuno-bloting.Results: Between the control group and HCC group of male, 12 different lectin affinity signals were found, including AAL, Con A, DSL, ECL, LCA, MNA-G, MNA-M, PHA-E, PSA, SNA,EBL, SNA-I, WFA,WFL. In the process of HCC, terminal α Fuc, α-man biantennary, Fucα1-6Glc NAc, Gal, oligomannosyl residues, bisecting Glc NAc and biantennary N-glycans increased, while galactosyl(β-1,4) N-acetylglucosamine and Gal NAcα/β-1,3/6Gal structure decreased. Between the control group and HCC group of female, 13 different lectin affinity signals were found, including AAL, CALSEPA, Con A, DSL, LCA, LTL, MNA-M, PHA-E, RCA I, RCA120, SNA,EBL, SNA-I, WFA, WFL, WGA. In the process of HCC, terminal α Fuc, Fucα1-6Glc NAc, bisecting Glc NAc increased, while high mannose, biantennary and Gal NAcα/β-1,3/6Gal decreased. Between the HCC group of male and HCC group of female, 7 differenct lectin affinity signals were found, indicated that in the HCC group of male, biantennary, N-Acetyl-D-Galactosamine, Gal, oligomannosyl residues increased, while bisecting and Gal NAcα/β-1,3/6Gal decreased compare with female.Conclution: Different sex of HCC patients had different serum protein, sugar chain structure changes associated with the higher incidence and higher mortality in male with HCCCHAPTER Ⅲ Diagnostic Value of DKK1 in Hepatocellular Carcinoma: A Meta-analysisObjective: To evaluate the diagnostic value of serum DKK1 in patients with hepatocellular carcinoma.Method: Pub Med, EMBASE, Science Direct, Elsevier, Springer, EBSCO, Cochrane library, WF, VIP, CMCC and CNKI were searched for relevant reports publish before January 2016. Eligibility studies were identified and determined according to the criteria of inclusion and exclusion. The characteristics of the included articles were appraised and extracted. Meta-Di Sc 1.4 was used to statistic the data. Three methods were used to exam the heterogeneity between the included articles. According to the heterogeneity results, the proper effect model was selected to calculate pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio and diagnostic odds ratio(DOR). Then the summary receiver operating characteristic(SROC) curve was performed and the area under the curve(AUC) was calculated.Results: A total of 452 articles were searched and 11 articles were included in this meta-analysis, providing 13 results about diagnosis value of DKK1 in HCC. The meta-analysis showed that when using DKK1 diagnosis HCC the pooled sensitivity was 72%, the pool specificity was 88%, the DOR was 25.95 and the AUC was 0.872. Using DKK1 combine AFP diagnosis HCC,the pooled sensitivity was 81%, the pool specificity was 85%, the DOR was 31.28 and the AUC was 0.913.Conclusion: The sensitivity of serum DKK1 in diagnosing HCC is high. Using DKK1 combine AFP to diagnosis HCC can significantly improve the diagnostic performance. |
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