Expression Of NOX4 Before The Deep Vein Thrombosis Formation Experimental And Clinical Study

ObjectOn Acute traumatic thrombosis of deep vein thrombosis model process, the former for thrombosis, the formation, not the formation of the state, testing rat vein blood NOX4 gene expression trends. Both deep vein thrombosis in clinical blood samples, focusing on the blood of the expression of NOX4. Analized the function which is promotion of thrombosis for NOX4 in deep vein thrombosis, Determine the transcription levels of human blood cells in the early diagnosis and NOX4 DVT as the early diagnosis of deep venous thrombosis candidate molecular markers.Method1. Summarized and analyzed rat femoral veins Gene chip data and real-time PCR for rat blood cell①In the establishment of acute traumatic deep vein thrombosis model, gene chip detection for rat femoral vein tissue,and Summarized for microarray data, Combined with Fold Change analysis, to screen differential expression genes from the Affymetrix 230A gene chips detection data of each group and to perform GO classification.(Signal Log2 ratio:comparison gene increases:Logo Ratio≥1, gene discreases:Log2 Ratio≤-1).②The genes of NOX family in the data, by adjusting the relevant parameters to determine the significantly differentially expressed genes, Parameter is set to Single Log2 Ratio≥3 or Single Log2 Ratio≤-3, analysis of NOX4 in the process of deep venous thrombosis tendency in rats;③By BLAST homology with human comparison, a clear homology with the human genome;④In the establishment of acute traumatic deep vein thrombosis model, based on observation time and thrombosis will be 100 SD rats were divided into normal control group (A group), early thrombosis group (B group), the peak of thrombosis group (C) and no thrombosis group (D group), extraction of total RNA of blood cells in each group, and quality testing:⑤Applicate Oligo6.69 primer design software to design PCR primers of rat NOX4,then reverse transcript total RNA to cDNA, using real-time PCR detection of each group blood cell of the rat NOX4 gene expression before and after thrombosis, initially identified differentially expressed genes.2. RT-PCR detection Blood samples of clinical DVT①Develop clinical standards for blood sample collection Clinical samples based on the observation time and grouped as follows thrombosis:Normal control group (Al group), Trauma control group(A2 group) the peak of thrombosis group (B group) and no thrombosis group (C group), extraction of total RNA in each group and the quality of blood testing.②Applicate Oligo6.69 primer design software to design PCR primers of human NOX4, then reverse transcript total RNA to cDNA, and with GAPDH as an internal reference, using fluorescence RT-PCR detect NOX4 gene expression before and after thrombosis from each group blood cell of clinical DVT blood samples.③Analyzed and summarized the experimental data and compared with expression change of vascular tissue, combine with physiological roles and participation in the development of other disease mechanism, Analysis the status and role in the deep venous thrombosis. Early diagnosis candidate markers of rats DVT in transcription levels maybe selected.Result1. Through animal experiments Analysis of the data, we found that the gene NOX4 instantly from trauma to the peak of thrombosis in both vascular endothelial cells in any significant increase in the blood test results also showed significant upward trend (BvsA:1.186, CvsA:4.716. DvsA:6.443). It is speculated that the gene may play an important role in the formation, and thrombosis are closely related.2. The NOX4 gene of rat and human is highly homologous to the clinical blood cells NOX4 by semi-quantitative PCR, showed that the peak in the form of deep vein hrombosis of the group and do not form a group to form was expressed, but the difference was not significant, which preliminary experiments may be related to insufficient number of samples included and the inclusion criteria lack of imperfections.Conclusion1, Since traumatic immediate time to the peak of venous thrombosis, NOX4 was significantly up-regulated in vascular tissue and blood cells of rat DVT model, which could play an important role in deep vein thrombosis.2. NOX4 in the rat experiments before the formation of deep vein thrombosis were high expression of deep vein thrombosis as an early diagnostic marker of the candidate molecule identified further research needed. Part of clinical trials, with the growing number of samples, the inclusion criteria of sound and continue to develop, NOX4 in human blood cells DVT cases may appear similar results with rats.