| Objective:Esophageal cancer is regarded as the most common malignancy worldwide related with human health. Compared with other nations, there has higher incidence in China. The mortality rate of esophageal cancer is 23.53% of malignant tumor mortality in our country. In spite of major advancement in cancer treatment, prognosis is still poor.The matrix metalloproteinases(MMPs) are a family of proteolytic enzymes which play critical roles in cancer development and aggression. At present, there have 5 categories and 23 species MMPs in human. MMP-9, a member of the MMPs family also known as gelatinase B, can degrade the different components of extracellular matrix and IV collagenanse and the integrity of the basement membrane(BM). Therefore, MMP-9 is involved in the occurrence of tumor cells, various stages of development and metastasis.The single nucleotide polymorphisms(SNPs) are the most common type in genetic polymorphism. SNPs almost appear in human genome, the frequency of SNPs exceeds 1% in human. The alteration of genetic structure may affect the transcription and protein products. Therefore, it is very importment to investigate the SNPs. There are C-1562T, R279Q, P574R, R668Q SNPs in MMP-9 gene, and those SNPs might change the development and metastasis of gastric cancer, breast cancer, colorectal cancer by influencing transcription and expression of MMP-9. However, whether the association of the MMP-9 polymorphism with susceptibility to esophageal squamous cell carcinoma(ESCC) is not yet to be determined.Our study is designed to investigate the association of the C-1562T, R279Q, P574R polymorphism of MMP-9 gene with susceptibility to ESCC, and to provide experimental basis for deeper understanding and knowledge of the related genetic background about development and transfer of esophageal squamous cell cancer in southwest China, which might provide a theoretical basis for early detection, early diagnosis of ESCC and a new method for treatment of ESCC.Methods:1. Subjects. All subjects were unrelated Han nationality and were resident in southwest China, including a experimental group of 235 ESCC patients and a control group of 300 healthy volunteers.2. To analyze the SNP of MMP-9 gene.(1) Genome DNA of blood samples were extracted by using Wizard Genomic DNA Purification kit.(2) MMP-9 C-1562T, R279Q, P574R polymorphism were analyzed by using the polymerase chain reaction-restriction fragment length polymorphisms(PCR-RFLPs) technique.3. Statistical analyses: Statistical analyses were performed by using SPSS13.0 software package (SPSS Company, Chicago, Illinois, USA).(1) Hardy-Weinberg analysis was performed via comparing the observed and expected genotype frequencies by using Chi-square test.(2) The distributions of genotype and alleltype were analyzed by using Chi-square test.(3) The odds ratio (OR) and 95% confidence interval (CI) were calculated by using an unconditional logistic regression model.(4) Pair-wised linkage disequilibrium analysis were conducted online by SHEsis (http://analysis.bio-x.cn).Results:1. The association analysis between the C-1562T SNP of MMP-9 and ESCC.The observed genotype frequencies of MMP-9 C-1562T SNP in controls did not deviate significantly from those expected from Hardy-Weinberg equilibrium(P>0.05). The allelotype and genotype distribution of the MMP-9 C-1562T SNP in the overall ESCC patients was no significantly difference from that in healthy controls (P>0.05). Compared with individuals with the C/C genotype, individuals with T allele (C/T or T/T genotype) did not change the risk of developing ESCC (age, gender, smoking and drinking status adjusted OR=0.73, 95%CI=0.49~1.09). When stratified by gender, compared with individuals with the C/T and T/T genotype, individuals with C/T genotype in male group had lower risk of occurrence ESCC (age, smoking and drinking status adjusted OR=0.54, 95%CI =0.33~0.87). When stratified by age, compared with individuals with the C/T and T/T genotype, individuals with C/T genotype in the group who were belowed 50 years had a tendency of reducing the risk of occurrence ESCC (gender, smoking and drinking status adjusted OR=0.25, 95%CI =0.09~0.72). When stratified by smoking status, compared with individuals with the C/T and T/T genotype, individuals with C/T genotype in smoker group had lower risk of occurrence ESCC (age, gender, drinking status adjusted OR=0.46, 95%CI =0.24~0.86). When stratified by drinking status, compared with individuals with the C/T and T/T genotype, individuals with C/T genotype in drinker group had lower risk of occurrence ESCC (age, gender, smoking status adjusted OR=0.36, 95%CI =0.18~0.72).2. The association analysis between the R279Q SNP of MMP-9 and ESCC.The observed genotype frequencies of MMP-9 R279Q SNP in controls did not deviate significantly from those expected from Hardy-Weinberg equilibrium(P>0.05). The allelotype and genotype distribution of the MMP-9 R279Q SNP in the overall ESCC patients was significantly different from that in healthy controls (P<0.05). When stratified by gender, compared with individuals with the Q/Q genotype, individuals with R/R genotype in female group significantly increased risk of occurrence ESCC (age, smoking and drinking status adjusted OR=3.39, 95%CI =1.37~8.04). When stratified by age, compared with individuals with the Q/Q genotype, individuals with R/R genotype in the group who were belowed 50 years had a tendency of increasing the risk of occurrence ESCC (gender, smoking and drinking status adjusted OR=3.39, 95%CI =1.23~9.29). When stratified by smoking status, compared with individuals with the Q/Q genotype, individuals with R/R genotype in smoker group had higher risk of occurrence ESCC (age, gender, drinking status adjusted OR=1.78, 95%CI =0.96~3.31). When stratified by drinking status, compared with individuals with the Q/Q genotype, individuals with R/R genotype in drinker group had higher risk of occurrence ESCC (age, gender, smoking status adjusted OR=2.40, 95%CI =1.13~5.11).3. The association analysis between the P574R SNP of MMP-9 and ESCC.The observed genotype frequencies of MMP-9 P574R SNP in controls did not deviate significantly from those expected from Hardy-Weinberg equilibrium(P>0.05). The allelotype and genotype distribution of the MMP-9 P574R SNP in the overall ESCC patients was significantly different from that in healthy controls (P<0.05). When stratified by gender, age, smoking and drinking status, respectively, compared with individuals with the R/R genotype, individuals with P/R genotype significantly reduce the risk of occurrence ESCC, but individuals with P/P genotype did not change the risk of occurrence ESCC.4. The association analysis between the C-1562T, R279Q, P574R SNP of MMP-9 and ESCC patients with lymphatic metastasis.(1) The frequency of the MMP-9 1562C/C, C/T+T/T genotypes was 82.4%, 17.6% in patients with lymphatic metastasis, and 71.7%, 28.3% in patients without lymphatic metastasis, respectively. The difference was not statistical significance.(2) The frequof the MMP-9 279R/R, R/Q, Q/Q genotypes was 39.2%, 43.1%, 17.7% in patients with lymphatic metastasis, and 40.7%, 31.5%, 27.8% in patients without lymphatic metastasis, respectively. The difference was not statistical significance.(3) The frequency of the MMP-9 574P/P, P/R, R/R genotypes in patients with lymphatic metastasis was 62.7%, 23.5%, 13.7%, and in patients without lymphatic metastasis was 70.7%, 14.7%, 14.7%, respectively. The difference was not statistical significance.5. There were no linkage disequilibrium between C-1562T and R279Q, R279Q and P574R, C-1562T and P574R.Conclusions:1. Individuals with MMP-9 574P/R genotype and 1562C/C genotype had a tendency of reducing the risk of ESCC occurrence. MMP-9 C-1562T, P574R SNPs were the protective factors of ESCC occurrence in southwest China.2. Individuals with MMP-9 279R/R genotype had a tendency of increasing the risk of ESCC occurrence. MMP-9 R279Q SNP was one of the risk factors of ESCC occurrence in southwest China.3. There had no significant association between the C-1562T, R279Q, P574R SNP of MMP-9 and ESCC patients with lymphatic metastasis in southwest China.4. In our study, there were no linkage disequilibrium between C-1562T and R279Q, R279Q and P574R, C-1562T and P574R.
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