Real-time Fluorescent Quantitation PCR Detects Tissue Factor Expression Level In Acute Pulmonary Thromboembolism Of Rats At Different Time

ObjectivePulmonary thromboembolism (PTE) gradually draws clinicians'attention because of the unexpected and sudden onset, its high lethality and mutilation rate. Although there are still not explicit large-scale epidemic investigations in China, various reported datas indicated that the incidence rate is not that low. The fatality rate of the untreated pulmonary embolism is 20-30%, while reasonable treatment can reduce the rate to2- 8%. As recognized in the traditional theory, that tissue factor is the initial factor to trigger exogenous blood coagulation and the strongest support for blood vessel injuries as well. New researches also indicated that the coordination between tissue factor and the vascular endothelial growth factor plays the vital role in recanalization and the formation of new blood vessels. Among the relative fewer reports about tissue factor, no clear conclusion has been drawn at present about the dynamic changes of tissue factor during different phases and whether it is an essential factor in coagulation process. Therefore, the identification of the pathogenesis of pulmonary thromboembolism, especially the parts tissue factor takes in the process, could help us to better understand the theoretical basis, treatment and clinical methods for this disease. This article focused on the difference of TF expression in lung tissue during different time and introduced the new quota technology simultaneously for the novel detected methods for thromboembolism diseases.Method1 Adopt partial thrombus induction methods to duplicate acute pulmonary thromboembolism model of rats during different phases. 2 Employ SYBRGreen I real-time fluorescent quantitation PCRtechnology to establish tissue factor (TF) standard product and its standard curve.3 Detect the alternations of tissue factor (TF) in lung tissue in 1st,7th,14th after embolism formation with the standard products as established previously.ResultThe animal model of pulmonary thromboembolism is successfully established. Thrombus formation were confirmed in pulmonary artery by pathology. The standard products and standard curve were constructed by real-time fluorescent quantitation PCR. Using cDNA as the standard product, which is the products of reverse tanscription of total RNA, we got better amplification curve, standard curve, and melt curve. The standard curve demonstrates well linear relationship (house-keeping gene GAPDH: R2=0.998, goal gene TF: R2=0.998, R2> 0.98) and accurate quota can be achieved in a broad scope. The amplification rate is satisfactory. (house-keeping gene GAPDH: E=95%, goal gene TF: E=89.4%,0.8