Experimental Study Of Cathepsin Gene Family And Early Diagnosis Of Deep Venous Thrombosis

ObjectiveIn transcription level, to reasach the expression changes of cathepsin gene family in blood of rats model of traumatic deep vein thrombosis before/ater, and non-formation; Analyze the roles of them involve in vascular endothelial cell extracellular matrix degradation, endothelial cell apoptosis and inhibit fibrinolytic activity. To screening molecular markers can be sensitive diagnosis deep venous thrombosis early.Methods1.To see Dr. Wang Bing's thesis "Establishment of a rat Acute traumatic deep vein thrombosis model and the femoral vein wall gene expression study" in rats modeling and grouping method, Analysis the chip data and summarize the expression regular pattern of cathepsin gene family in the femoral vein endothelial cells.2 Search ncbi GenBank database and cnki database to determine rat cathepsin gene family members and homology comparison with the person by BLAST. Then Access to their physiological functions involved.3 Re-establish 100 rat models of acute traumatic deep vein thrombosis as follow groups:normal control group (A), thrombosis in the initial phase (B), thrombosis peak (C) and the formation of thrombus group (D). Four groups total RNA was extracted separately and quality testing them.4 Applicate Oligo6.69 primer design software to design RT-PCR primers of all cathepsin family members,then reverse transcript total RNA to cDNA.5 With GAPDH as an internal reference, using semi-quantitative PCR detection of all members of the cathepsin gene family expression before and after thrombosis, initially identified differentially expressed genes, detected expression of rats cathepsin genes have been locked in each group by fluorescence real-time quantitativePCR.6 Analyzed and summarized The experimental data and compared with expression change of vascular endothelial cells, early diagnosis candidate markers of rats DVT in transcription levels can be selected.Through they expression changes regular pattern of deep-vein thrombosis, physiological roles and participation in the development of other disease mechanism, Analysis the status and role in the deep venous thrombosis.Results1 An analysis of pre-chip data, we found, some cathepsin genes are closely related with the formation of DVT. Ten members are highly expressed in vein endothelial cells since the injury immediately to the peak of thrombosis.Considering the family members are similar higher in gene and protein sequence, and thus its function is similar, suggested that family members may play similar or DVT formation of synergies.2 Eighteen rat cathepsin gene family members are:cathepsin 7,cathepsin 8,cathepsin A,cathepsin B,cathepsin C,cathepsin D, cathepsin E,cathepsin F,cathepsin G,cathepsin H,cathepsin K,cathepsin L1, cathepsin M, Cathepsin Q,cathepsin Q like 2,cathepsin R,cathepsin S,cathepsin W, Rat and human cathepsin genes are highly homologous, the family mainly player antigen presentation, apoptosis, matrix degradation and other physiological functions, also involve lysosomal pathway and of systemic lupus erythematosus pathway.3 Rat model was successful Re-established, Twice the rate of thrombosis was no significant difference ntegrity and purity of each group RNA extracted fromBlood cells is Satisfied with the requirement of real time quantitative PCR experiment4 All the members of cathepsin gene family by semi-quantitative PCR, the results show that cathepsin A, B, C, D, E, F, G, H, K, L1, W has a difference in expression before and after thrombosis, Them real-time PCR test showed cathepsinB, cathepsinC, cathepsinF, cathepsinH are significantly raised.in the peak of thrombosis group compared with the initial thrombosis group,and cathepsinB, cathepsinC up is extremely significant.5 The results contrast with previous microarray results, In the process of thrombosis,cathepsin B, cathepsin C express in the femoral vein endothelial cells and blood cells showed upward trend. The first time we propose that the two can be as deep vein thrombosis in rats sensitive candidate molecular markers of early diagnosis.6 After Endothelial cell injuryed, cathepsin B to promote the degradation of extracellular matrix, endothelial cell apoptosis, endothelial cell dysfunction, and thus enlarge the procoagulant activity of endothelial cells, then promoting The formation and development of DVT. Conclusion1 Cathepsin family is closely related with rats traumatic deep vein thrombosis.2 Cathepsin family through play roles of degradation of extracellular matrixand,endothelial cell apoptosis and inhibit fibrinolytic activity to promote the development of thrombosis.3 Cathepsin B, cathepsin C can be as candidate molecular markers of early diagnosis deep venous thrombosis.