| Objective:To explore the correlation of the COL1A1 gene polymorphism with the ACL injuries by detecting the SNP of the -1997G/T locus and+1245G/T locus used real-time fluorescence quantitative PCR; to explore the genetic mechanism of ACL injuries by detecting the content and proportion of collagen types Ⅰ and Ⅲ in ACL tissue with immunohistochemistry.Methods:A total of 40 adult patients who diagnosed with ACL injuries under arthroscopy were chosen as experimental group, and there were 20 cases in non-contact ACL injuries group and 20 cases in contact ACL injuries group. In addition,20 cases of normal adult patients from traumatic amputation were chosen as control group, which without any history of ACL injuries. The specimen were collected from injury ACL residual and normal ACL from traumatic amputation. Some specimens were saved in -80℃ refrigerator, the others were fixed in 10% formaldehyde immediately. The SNP of the-1997G/T locus and+1245G/T locus were detected real-time fluorescence quantitative PCR, chi-square test was applied to analyse HWE and genotype frequency distribution of each group, and the logistic regression was applied to assess odds ratios(OR)and 95% confidence interval(95%CI), that the correlation of the COL1A1 gene polymorphism and ACL injuries was analysed;The expression of type Ⅰ and type Ⅲ collagen were detected by Biosens Digital Imaging System v6.0 software after immunohistochemistry, which were indicated by the expression of type Ⅰ and type Ⅲ collagen; The data were analysed with SPSS20.0.Results:1. The law of Hardy-Weinberg was applied to test the genotype frequencies in the population of each group, the P>0.05, indicated that the population was representative.2. The-1997G/T locus of COL1A1 gene, the genotype frequency of GG, TT and GT were 15%,55%,30% in the normal ACL group,while GG, TT and GT genotype frequency were 35%,25%,40% in the non-contact ACL injuries group and 25%,55%,20% in the contact ACL injuries group, In pair-wise comparison among non-contact ACL injuries group,contact ACL injuries group and the normal ACL group, the genotype frequency of T/T was lower in the non-contact ACL injuries group,there was statistical significance (P<0.05).Binary logistic analysis suggested that lower frequency of the COL1A1TT (-1997G/T polymorphisms) genotype was significantly correlation with increased the susceptibility of non-contact ACL injuries(OR95%CI=0.21 (0.05-0.8)); The COL1A1GG, TT, GT (-1997G/T polymorphisms) genotype were not correlation with the susceptibility of contact ACL injuries (OR95%CI=1.42(0.28-2.41), OR95%CI=3.21(0.57-2.61),OR95%CI=1.25 (0.35-3.63)).3. The -1997G/T locus of COL1A1 gene, the allele frequency of G and T were30%,70%in the normal ACL group,while G and T allele frequency were55%,45% in the non-contact ACL injuries group and35%,65% in the contact ACL injuries group, In pair-wise comparison among non-contact ACL injuries group,contact ACL injuries group and the normal ACL group, there was no statistical significance (P>0.05).Binary logistic analysis suggested that the COL1A1G, T (-1997G/T polymorphisms) allele were not correlation with the susceptibility of ACL injuries (OR95%CI=1.42 (0.29-2.93),OR95%CI=1.25 (0.39-4.73), OR95%CI=1.57 (0.33-2.56),OR95%CI=1.98 (0.35-3.63)).4. The+1245G/T locus of COL1A1 gene, the genotype frequency of GG, TT and GT were 20%,55%,25% in the normal ACL group,while GG, TT and GT genotype frequency were 35%,20%,45% in the non-contact ACL injuries group and 25%,50%,25% in the contact ACL injuries group, In pair-wise comparison among non-contact ACL injuries group, contact ACL injuries group and the normal ACL group, the genotype frequency of T/T was lower in the non-contact ACL injuries group, there was statistical significance (P<0.05). Binary logistic analysis suggested that lower frequency of the COL1A1TT(+1245G/T polymorphisms) genotype was significantly correlation with increased the susceptibility of non-contact ACL injuries (OR95%CI=4.00 (1.21-16.27));The COL1A1GG, TT, GT(+1245G/T polymorphisms) genotype were not correlation with the susceptibility of contact ACL injuries (OR95%CI=1.19 (0.57-3.35),OR95%CI=2.99 (0.41-2.23),OR95%CI=1.21 (0.31-2.91)).5. The+1245G/T locus of COL1A1 gene, the allele frequency of G and T were45%,55% in the normal ACL group,while G and T allele frequency were 57.5%,2.5% in the non-contact ACL injuries group and 37.5%,62.5% in the contact ACL injuries group, In pair-wise comparison among non-contact ACL injuries group, contact ACL injuries group and the normal ACL group, there was no statistical significance (P>0.05).Binary logistic analysis suggested that the COL1A1G, T (+1245G/T polymorphisms) allele were not correlation with the susceptibility of ACL injuries(OR95%CI=0.99 (0.46-2.98),OR95%CI=1.31 (0.65-3.59), OR95%CI=1.78 (0.58-2.99),OR95%CI=1.41 (0.75-4.68)).6. The expression of type Ⅰ collagen was 5194.97±619.31 in the normal ACL group, while the expression of type Ⅰ collagen was 1654.25±492.93 in the non-contact ACL injuries and 4858.30±985.69 in the contact ACL injuries, In pair-wise comparison among non-contact ACL injuries group, contact ACL injuries group and the normal ACL group, The expression of type Ⅰ collagen was lower in the non-contact ACL injuries group,there was statistical significance (P<0.05), however,the expression of type Ⅰ collagen was no statistical significance between contact ACL injuries and normal ACL group.7. The expression of type Ⅲ collagen was 2599.25±564.03 in the normal ACL group, while the expression of type Ⅲ collagen was 4257.15±800.82 in the non-contact ACL injuries and 2586.90±624.88 in the contact ACL injuries, In pair-wise comparison among non-contact ACL injuries group, contact ACL injuries group and the normal ACL group, The expression of type Ⅲ collagen was higher in the non-contact ACL injuries group,there was statistical significance (P<0.05), however, the expression of type Ⅲ collagen was no statistical significance between contact ACL injuries and normal ACL group.8. The ratio of type Ⅰ to type Ⅲ collagen was 2.04±0.62 in the normal ACL group,while the ratio of type Ⅰ to type Ⅲ collagen was 0.40±0.14 in the non-contact ACL injuries and 2.07±0.95 in the contact ACL injuries, In pair-wise comparison among non-contact ACL injuries group, contact ACL injuries group and the normal ACL group, The ratio of type Ⅰ to type Ⅲ collagen was higher in the non-contact ACL injuries group,there was statistical significance (P<0.05), however, the ratio of type Ⅰ to type Ⅲ collagen was no statistical significance between contact ACL injuries and normal ACL group.Conclusions:1.The TT genotype of COL1A1(-1997G/Tlocus,+1245G/Tlocus) gene polymorphism was correlation with the susceptibility of ACL injuries, and the higher frequency of the COL1A1TTgenotype was significantly correlation with reduced the risk of ACL injuries compared with the GG andGT genotype.2. It had no significant correlation between the allele of COL1A1 (-1997G/T locus,+1245G/T locus)gene polymorphism and the ACL injuries.3.It was reduced the risk of ACL injuries by increasing the IOD of type Ⅰ, ratio of type Ⅰ to type Ⅲ collagen and reducing the IOD of type Ⅲ by converting of G'T in the COL1A1 gene of COL1A1 (-1997G/T locus,+1245G/T locus). |
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