Association Between Homocysteine Level And MTHFR Polymorphism In Patients With Thoracic Aortic Dissection

Objective: Thoracic aortic dissection(TAD)is a macrovascular disease with high mortality and intractable treatment,and its incidence tends to be young.At present,there is still no reliable biomarkers that can be used to provide early warning.Hypertension is an important risk factor of aortic dissection.More than 75% patients with hypertension in China are accompanied by elevated homocysteine(Hcy).As the limiting enzyme of Hcy metabolism,methylene tetrahydrofolate reductase(MTHFR)plays a key role in the metabolism of Hcy in vivo.Single nucleotide polymorphism(SNP)could lead to dysfunction of genes,thus contributed to the occurrence and development of diseases.However,the relationship between MTHFR SNP rs1801133 and TAD has not been reported.Particularly,whether this SNP combined with Hcy metabolism can be used as an early warning marker for TAD still remains uncertain.In order to investigate the proportion of increased Hcy in patients with TAD,and analyze whether the increase of Hcy level is related to the MTHFR gene polymorphism,and whether Hcy level combined with the polymorphism of MTHFR gene can be used as an early warning screening tool for TAD?Methods: Firstly,detection of Hcy levels in blood samples of 117 patients with TAD and 43 healthy people by enzymatic cycling assay;Secondly,the MTHFR protein structure was simulated using the bioinformatics tool Swiss-model software to observe the difference between normal and mutated protein structures and to predict the biological functions of SNP rs1801133 through the 3DSNP database;Finally,genomic DNA was extracted and combined with MTHFR rs1801133 primers for PCR,and the amplified products were rapidly genotyped using T7 Endonuclease Ⅰ(T7EI).Then,we combined with the results of gene sequencing to verify the relationship between rs1801133 and Hcy in TAD patients.Results: Among 117 patients with TAD,the Hcy concentration was(35.53 ± 15.77)μM,and the percentage of Hcy levels above 15 μM was 89.75%;among 43 healthy people,the Hcy concentration was(10.54 ± 2.79)μM,and the percentage of Hcy over15 μM was 6.98%.It was found that the level of Hcy in TAD group was significantly higher than that in normal control group(P <0.0001).Compared with the normal protein,the structure of the mutant MTHFR protein is loose and the folding direction is obviously changed.rs1801133 is closely related to the MTHFR gene,is highly conserved among different species,and binds more to cell regulators that shape vascular structure.The results of gene sequencing showed that the primers were highly specific.After genotyping of all blood samples,it was found that the detection rate of MTHFR rs1801133 homozygous mutation was 82.8% in 105 TAD patients with elevated Hcy levels.It showed that the level of Hcy in patients with aortic dissection was significantly increased,and it was positive related to rs1801133 genotype.Then PCR/T7 EI method was established to identify the genotype of TAD patients.Conclusion: The level of Hcy in patients with TAD was significantly increased.TAD patients with elevated Hcy have an extremely high rate of detection of SNP rs1801133 homozygous mutations.The established PCR/T7 EI method of rs1801133 provided a rapid and effective screening tool for those with high risk for TAD.