| Backgrounds: CRC is caused by multi-factors and multi-stages. In this process, DNA damage accumulates in cells over time as a result of exposure to exogenous and endogenous toxic agents. DNA damage, if unrepaired, has the potential to generate genome instability, which can further cause the loss of control in cell apoptosis,proliferation and differentiation, and initiated carcinomatous changes at last. To prevent such deleterious effects and safeguard the integrity of the genome, cells possess multiple mechanisms to repair DNA damage. One such system is the critical pathway known as DNA mismatch repair (MMR). However, the MMR genes have inheritance polymorphisms. The different susceptibility to CRC may be induced by different repair capacity of MMR proteins which caused by Different allele combinations.Objective: To explore the frequency distributions of genetic polymorphisms of MMR genes, and their interaction with environmental factors in colorectal carcinogenesis, a population-based case-control study will be conducted.Methods: In this study, we recruited CRC cases in 3 counties (Jintan, Taixing and Huaian) of Jiangsu province, China. A population study based on cancer registration was performed with association of the Centers of Disease Control in the 3 counties. The patients with onset CRC from January 1 to December 31 of 2004 were investigated. One hundred eighty blood samples collected from healthy individuals in the same geographic origin were taken as the control group.Blood samples were taken at the time of interview. Genomic DNA was extracted from peripheral blood using QIAamp DNA blood mini kit. IVS10+12G>A and IVS12-6T>C in hMSH2 gene was analyzed using a PCR-based DHPLC, the existence of them were verified by DNA sequencing. Bioinformatics software was used to analysis the etiological mechanism of these two SNPs. Basic variables of controls and cases were compared byχ2 tests. Odds ratio (OR) and its 95% confidence interval (95%CI) were estimated by unconditional logistic regression. All analyses were performed using STATA7.0.Results:1. MMR gene polymorphisms and CRC1.1 IVS10+12G>A and CRC: The genotypic frequencies of the IVS10+12G>A were 63.9% in cases, significantly higher than 51.7% of the controls(p<0.05). Compared to the GG genotype, the GA+AA genotypes could increase the risk of CRC, with the OR of 1.65(95%CI:1.01～2.71). when stratified according family history, this polymorphism substantially increased the risk of CRC for population from CRC genealogy(p<0.05), with the OR of 2.68(95%CI:1.16～6.67).1.2 IVS12-6T>C and CRC: The genotypic frequencies of the IVS12-6T>C were 48.1% in cases, not substantially different from 37.8% of the controls(p>0.05). when stratified according family history, this polymorphism substantially increased the risk of CRC for population from CRC genealogy(p<0.05), with the OR of 2.68(95%CI:1.16～6.67) .2. The interaction between environmental factors and MMR gene polymorphism.2.1 Environmental factors and CRC: Smoking,drinking,high monthly consumption of fried food and high monthly consumption of sauerkraut were all risk factors of CRC(p<0.05). The OR were 1.96(95%CI:1.17～3.27),2.20(95%CI:1.30～3.73),2.29(95%CI:1.37～3.83),1.85(95%CI:1.10～3.11)respectively.2.2 Environmental factors and IVS10+12G>A: No statistical significant association was shown between smoking/drinking and IVS10+12G>A. The interaction of IVS10+12G>A and high monthly consumption of fried food or sauerkraut all significantly increased the risk of CRC(p<0.05), the OR value were 2.90(95%CI:1.41～6.11) and 2.98(95%CI:1.48～6.20) respectively. Besides, this polymorphism significantly increased the risk of CRC for population younger than 50 years(p<0.05), with the OR of 3.42(95%CI:1.36～9.43).2.3 Environmental factors and IVS12-6T>C: No statistical significant association was shown between smoking/drinking and IVS12-6T>C. The interaction of IVS12-6T>C and high monthly consumption of fried food or sauerkraut all significantly increased the risk of CRC(p<0.05), the OR were 2.14(95%CI:1.01～4.57) and 2.25(95%CI: 1.13～4.52) respectively. Besides, this polymorphism significantly increased the risk of CRC for population younger than 50 years and with a male gende(rp<0.05), with the OR of 4.17(95%CI:1.70～10.43) and 2.02(95%CI:1.09～3.76) respectively.3. Gene-gene interaction and CRC: In combinative analysis, an interaction was shown existed between IVS10+12G>A and IVS12-6T>C, and individuals with these two SNPs simultaneously had an significantly higher risk of CRC(p<0.05), with the OR of 2.43(95%CI:1.17～5.09).4. Results of Bioinformatics software analysis Four Bioinformatics softwares were used to analysis the relations of these two SNPs with pre-mRNA splicing. Three of them had shown IVS12-6T>C and two of them had shown IVS10+12G>A may be deleterious variations for pre-mRNA splicing. IVS12-6T>C may destroy the combination motifs of SR protein and interfere the intronic splicing enhancers (ISEs). And IVS10+12G>A may produce new ISEs and cryptic splicing sites. All of them possibly produce new sites for splicing regulation.Conclusions: IVS10+12G>A other than IVS12-6T>C may be associated with genetic susceptibility to CRC. However, IVS10+12G>A and IVS12-6T>C all were significantly associated with mobility of familial patients and patients with onset ages<50 years, and IVS12-6T>C may play an etiological role in male patients yet. With the cross-over study, we found the genotype of IVS10+12G>A and IVS12-6T>C all were interacted with high monthly consumption of fried food and sauerkraut. Besides, IVS10+12G>A and IVS12-6T>C had a interaction in increasing risk of CRC. Bioinformatics softwares analysis had shown that these two SNPs may be deleterious variations for pre-mRNA splicing.
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